Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241101EnglishN2018January10General SciencesStratosphere-mesosphere Coupling Through Vertically Propagating Gravity Waves During Mesospheric Temperature Inversion (MTI): An Evidence
English0106G. Venkata ChalapathiEnglish S. EswaraiahEnglish M. Venkat RatnamEnglish K. Niranjan KumarEnglish P. Vishnu PrasanthEnglish Jaewook LeeEnglish Yong Ha KimEnglish S.V.B. RaoEnglishObjective: It is theoretically observed that atmospheric gravity waves play a key role in vertical coupling during the Mesosphere Temperature Inversion (MTI). Therefore, the present paper describes the observational evidence for vertical coupling between the stratosphere and mesosphere through the short-period gravity waves (GWs), during the Mesosphere Temperature Inversion (MTI) over a tropical region, Gadanki (13.5oN, 79.2oE), India.
Method: The combined observations of Mesosphere-Stratosphere-Troposphere (MST) Radar and Rayleigh LIDAR located at Gadanki is utilized to study the vertical coupling. We used a unique experimental design from the two ground-based instruments that scan the lower and middle atmosphere simultaneously during the observational campaign. This kind of combined instruments are very sparsely located on the same site to make the observations unique to understand the vertical coupling processes of GWs.
Result: The vertical flux of the horizontal momentum of GWs of periods in the range 20 min. to 2h is investigated in the mesosphere using the MST Radar winds. The emphasis is made on the variability of zonal and meridional momentum fluxes in the mesosphere and possible reasons for the variability of fluxes during MTI. It is observed that raise in momentum fluxes of ~7 m2/ s2 in the eastward flux and ~10 m2/s2 in southward flux at mesospheric altitudes during the MTI.
Conclusion: The gravity wave (GW) analysis using the LIDAR temperature profiles indicate the connection between GW breaking at mesosphere altitudes and temperature inversion and thus the turbulence caused mesospheric echoes. The study suggests the prospect of coupling between stratosphere and mesosphere during the MTI.
EnglishGravity wave coupling, Mesospheric Temperature Inversion (MTI), MST Radar, Rayleigh LIDARIntroduction
The role of atmospheric gravity waves (GWs) in modulating the energy budget of the Mesosphere-Lower thermosphere (MLT) has been well recognized (Fritts and Alexander, 2003). The short-period GWs are playing a key role in vertical coupling of the different regions of the atmosphere (Sato, 1993;Fritts and Alexander, 2003). The tropical region is found to be a fountain of generating short-period GWs (Fritts and Alexander, 2003), and their generation and propagation studies are well established in the lower atmosphere over the Indian tropical region (Dhaka et al., 2002; Debashis Nath et al.,2009 and references therein). However, its effects on the MLT is sparsely studied over this region.
Earlier studies highlighted the importance of GWs in changing the MLT thermal structure (Fritts and Alexander, 2003). It was understood that vertically propagating GWs begins to break at the level where there is a sudden change in the temperature lapse rate or at the critical level(Meriwetherand Gerrard, 2004), and deposit large amount of energy and momentum (Fritts and Nastrom, 1992; Gardner and Yang, 1998; Ramesh and Sridharan, 2012). The studies on GW effects on the mesosphere temperature and the Mesosphere Temperature Inversion (MTI) are well existed (Garcia and Solomon, 1985; Hauchecorne et al., 1987; Venkat Ratnam et al., 2003; Ramesh and Sridharan, 2012). The occurrence mechanism of MTI or “mesospheric inversion layer” (MIL) is clearly discussed by Meriwether and Gerrard (2004). The occurrence of MIL and its characteristics are well studied using LIDAR’s, Radars, Rocket Sonde, and Satellites over different geographic locations (Ramesh et al., 2014 and references therein). Thus the earlier studies described the relation between MTI and GW breaking, however, the relation between the mesospheric radar echoes and MTI and GWs breaking studies sparsely exist over the tropical region. Further, the simultaneous ground-based observations both in the stratosphere and mesosphere and the quantification of GW breaking at mesosphere altitudes is still incomplete. In the present study, we made an attempt to unclear this issue and presented the vertical coupling using the unique data set over a tropical region Gadanki.
To this end, up to our knowledge concerned, this is the first report on the mesospheric echo phenomenon and their relation to GW breaking and thus the MTI. The details of the data used and methodology followed for the current study is briefly given in section 2, the results and discussions are provided in section 3, comprehensive summary, and key findingsare emphasized in section 4.
Data
In the present study, the unique data set of Mesosphere-Stratosphere-Troposphere (MST) Radar and Rayleigh LIDAR located in a tropical region, Gadanki(13.5oN, 79.2oE), India is utilized. Short-period (20 min.-2 h) GW momentum fluxes are estimated in the mesosphere using the MST Radar wind data, and LIDAR temperature profiles are used to represent the MTI and GW characteristics both at the stratosphere and mesosphere altitudes.
Methodology
MST radar
The Gadanki MST Radar is high power VHF radar with a peak transmitter power of2.5 MW and operates at 53 MHz. The MST Radar provides winds from surface to 22 km and 65-85 km. More details of MST Radar can be found in Rao et al. (1995). As mesospheric echoes from the MST Radar are mainly due to ?uctuations in electron density gradients and irregularities, they are con?ned to day-time and hence the availability of data is restricted to ∼10:00 to 17:00h Indian Standard Time (IST) (IST=UT+05:30 h).The structure and characteristics of MST Radar mesospheric echoes and mesospheric wind estimation are discussed in Kumar et al. (2007) and Eswaraiah et al. (2011). For the present study, a clear analysis of radar radial velocity profiles of individual days and defined percentage occurrence (PO) of echoes are performed for reliable wind estimation; the detailed procedure is given in Eswaraiah et al (2013). The reliable estimates of GW momentum flux can be obtained using a threshold of 20% of PO. Thus, the echoes in the range bins with PO less than 20% are omitted. For the flux estimation, the symmetric beam radar method of Vincent and Reid (1983) is utilized and the procedure has been discussed in Eswaraiah et al. (2013).
The momentum fluxes of horizontal winds have been estimated using the following equations
Momentum flux for the E-W beam:
moreover, for the N-S beam:
Where is zonal momentum flux, is meridional momentum flux and are square of radial wind perturbation in east, west, north and south directions, respectively, and θ is the anglebetween the radar beam to the zenith. Two consecutive radar beams make an angle θ, each beam being at zenith. In the present case, it is 10o.
Rayleigh Lidar
The Gadanki Rayleigh LIDAR data is used to observe the MTI and GWs in the stratosphere and mesosphere, but not for GW momentum flux estimation. The LIDAR employs the second harmonic of Nd: YAG pulsed laser at 532 nm with a pulse energy of ~ 550 mJ and Rayleigh receiver is used, which employs a Newtonian telescope. The LIDAR provides photon count profiles with an altitude resolution of 300m and time resolution of 250s (5000 laser shots were integrated for one profile). Although the temperature can be derived up to 85 km, due to low SNR at higher altitudes, the altitude range considered for the present study is from 35 to 75 km. The basic method of deriving the temperature profile from the measured photon count is similar to the procedure given by Hauchecorne and Chanin (1980). Usually, the random errors occur while deriving the temperature and they vary from 2 K at 30 km to 4 K at 75 km (Parameswaran et al., 2000). The total vertical flux of horizontal momentum using LIDAR temperature profiles is discussed in Eswaraiah et al., (2013).More details of this instrument and method of analysis can be found from Bhavani Kumar et al. (2001), Siva Kumar et al. (2001) and Ratnam et al. (2002).
For the present study, MST Radar data on 18 November 1998 and 11 January 1999 has been used. Similarly, the Rayleigh LIDAR night time temperature measurements on 18 November 1998 and 11 January 1999 have been used. For 18/19 November 1998, the temperature measurements were made from 03:20 h IST to 05:40 h IST and for 11/12 January 1999 the observations were made between 21:29 h IST to 04:10 h IST on the next day. The simultaneous MST radar observations in the mesosphere and Lidar observations are very rarely will match, since the radar mesospheric echoes are highly sporadic and they will not every day. Though the observational data is old, the scope of the present study is sparse and it is new.
Discussions
Figures 1(a) and 1(b) depicts the radar reflectivity in terms of SNR (Signal to Noise Ratio) observed on 18 November 1998 and 11 January 1999, respectively. Corresponding late night time mean temperature profiles observed by Rayleigh LIDAR are shown in Figure 1(c) and 1(d). Temperature inversion in the mesosphere (Fig.1(c)) is seen on 18 November 1998 but not on 11 January 1999. On the temperature inversion day active radar (MST) echoes at mesospheric altitudes just below the temperature inversion layer (below 75 km) are observed, whereas on non-inversion day no radar echoes are seen in the mesosphere, indicating a close association between mesospheric radar echo occurrence and temperature inversion. Past studies on mesosphere thermal structure (Ratnam et al., 2002; Kishore Kumar et al., 2008) explained the reasons behind this phenomenon. It is observed that whenever the strong mesospheric radar echoes are noticed in the day-time, then mesospheric temperature inversion is evident in the associated night-time LIDAR temperature profiles. It is also seen that whenever there is no temperature inversion, there are no strong echoes observed in the MST Radar observations (Ratnam et al., 2002).The temperature inversion at mesospheric altitudes and turbulence generated to form mesospheric echoes can be understood by verifying the short-period GW features in both the stratosphere and mesosphere and their breaking at mesosphere altitudes. To find the existence of short-period GWs and their propagation from the lower atmosphere to mesosphere, the periodicities and propagation of GW shave been tested. The GW features and their propagation has been verified using the simultaneous observations of MST Radar and LIDAR from the troposphere to mesosphere altitudes by performing spectral analysis. Wavelet analysis has been applied for the de-trended radial velocities of radar, both at the troposphere (18 km) and mesosphere (70.8 km) altitudes and for the temperature perturbations of LIDAR at the stratosphere (45.1 km) and mesosphere (77.2 km) altitudes on 18 November 1998 and are shown in Figure 2.The magnitude of wavelet power is shown in logarithmic values. From the figure, it can be noticed that in the mesosphere at 70.8 km (Figs.2(a) and 2(b)),30 min. to 1 h and 1-2 h period waves are present between 11:30 h-12:30 h IST in east beam and again between 14:30 h-15:30 h IST in the north beam of MST Radar observations. Since the MST Radar will not give background wind information in the gap region (20-60 km), the LIDAR temperature perturbations for the wave information is taken into consideration. At 45 km (Fig.2(c)) in the stratosphere, 30 min. to 1 h period and at 77.2 km (Fig.2(d)) 30 min. to 2 h period, waves are evidenced. The similar analysis has been performed at 18 km (Figs. (2e) and (2f)) i.e., in the troposphere, and it shows that the waves with periodicities up to 1 h are significant both in east and north beams of radar radial velocities. The analysis resembles that at the troposphere altitudes waves with a wide spectrum of periodicities exist and with altitude the waves with low periods propagate to the mesosphere altitude and break when there exists temperature inversion in the background, resulting in turbulence at the mesosphere altitudes below the inversion layer (Fig.1(a)). Further, to show the difference in wave propagation on inversion and non-inversion days, the 30 min. to 1 h and 1-2 h periodicities are filtered from LIDAR temperature perturbations and shown along with daily mean temperature profiles in Figure 3. Figs.3(a), and 3(b) shows the upward propagation of 30 min. to 1h and 1-2 h period waves respectively, on 18/19 November 1998. Fig.3(c) shows the mean temperature profile on 18/19 November 1998. Figs.3(d) and 3(e) and 3(f) are same as (a,b,c), but on 11/12 January 1999. Usually, the waves with low periods (or high frequency) can propagate to the mesosphere (above 60 km) and others get filtered as their phase speed match with the background wind speeds. On 11 January 1999 (Lower panel), the 30 min. to 1h and 1-2 h period waves (Figs.3(d) and 3(e)) continuously existed in the mesosphere altitudes without breaking and hence no inversions in resulting mean temperature. In contrast, on 18 November 1998 (Upper panel), 30 min. to 1h and 1-2 h period waves (Figs.3(a) and 3(b)) are continuously observed above 75 km altitude and below that they are not frequent, which tells that the waves are breaking at ~ 75 km throughout the day and hence at 75 km large inversion appeared in the mean temperature (Fig.3(c)). So far it is unresolved that whether due to inversion waves will break or due to the breaking of the waves inversion occurs. Further, to elucidate the GW breaking at mesosphere altitudes, the characteristics of short-period GWs have been estimated and displayed in Figure 4.
Fig.4(a) shows the vertical wavelength of short-period GWs during the observational period, Fig.4(b) depicts the GW amplitude with height, Fig.4(c) presents the momentum flux deposition by short-period GWs, and Fig.4(d) exhibits the variation of Brunt Väisala Frequency (BVF) of short-period GWs. From the figure, it is evident that the significant 10-20 km vertical wavelength GWs exist during the span of observational period, and the amplitude is increasing with height and peak amplitude is reaching ~ 75 km and results in the wave breaking by producing the large zonal momentum flux at 75 km (Fig.4(c)). Sudden variation in BVF can be seen when the wave breaking and depositing large flux into the background. The analysis further supports the vertical propagation of short-period GWs and their breaking at ~ 75 km.
Using the procedure given in section 2, the time variation of zonal and meridional momentum flux of 20 min. to 2 h GWs during the inversion day(18 Nov.1998) are estimated and presented in Figure 5.The fluxes are evaluated using the procedure shown above and averaged over about 15 min. Even though a high degree of temporal variability in zonal momentum fluxes are seen from Figure 5, a significant momentum flux determination can only be obtained by averaging it over reasonably long periods (Kudeki and Franke, 1998). In the mesosphere, at times of (Figure 2) dominant wave presence, sudden enhancement in flux is observed (Figs.5(a) and 5(b)). The zonal flux is quite low after ~ 14:00 h IST (Fig.5(a)) whereas the meridional flux (Fig.(5b)) continues to be strong and fluctuating. As shown in Figure 5 at 70.8 km in the forenoon hours the dominant flux is eastward, and later it is southward. Momentum fluxes are mainly eastward (~7 m2/s2) between 11:30-12:30 h IST and later observed in southward (~ 10 m2/s2) direction between 14:30-15:30 h IST. In the troposphere, no significant changes are noticed in the fluxes which could be attributed to small wave amplitudes due to high density.
Conclusions
By Utilizing the combined observations of MST Radar, and Rayleigh LIDAR, at a tropical station, Gadanki (13.5oN, 79.2oE), India, the existence and propagation of the short-period (~of periods 20 min. to 2 h) gravity waves during the mesospheric temperature inversion (MTI) has presented. Further, the coupling between the Stratosphere and mesosphere has been investigated during MTI through LIDAR temperature measurements and estimating the momentum fluxes in the mesosphere.
The main outcomes or conclusions of the present study are summarized below:
The relation between MST Radar mesospheric echoes and mesospheric temperature inversion (MTI) obtained with LIDAR daily mean temperatures have been noticed and it is consistent with earlier reports.
Using LIDAR temperature profiles, the propagation of short-period GWs during the MTI and non-MTI cases and their breaking at 75 km leads to mesospheric turbulence and the formation of MTI has been observed.
Upward propagation of GWs and their breaking has been tested by studying the characteristics of the short-period GWs using the LIDAR temperature profiles.
The present case study during MTI showed the rise in momentum fluxes with ~7 m2/s2 in the eastward flux and ~10 m2/s2 in southward flux at mesospheric altitudes with no significant variation in the lower atmosphere. This could be due to large wave breaking at mesosphere altitudes.
The present study suggests that during MTI, the Stratosphere and the mesosphere are coupled through the propagation of short-period GWs, and hence the waves are transporting energy and momentum from lower atmosphere to the mesosphere. The role of GWs in causing the turbulence to form the mesospheric radar echoes is highly substantial.
Acknowledgements: We deeply appreciate the National Atmospheric Research Laboratory (NARL) for providing the data used in the present study. SE acknowledges for financial support by the Korea Polar Research Institute (PE17020), Korea and Chungnam National University, Daejeon, Korea. We alsoacknowledge the technical staff who helped to retrieve the data from the MST radar and Rayleigh Lidar.
Authors acknowledge the immense help received from the scholars whose articles are cited and included in references of this manuscript. The authors are also grateful to authors / editors / publishers of all those articles, journals and books from where the literature for this article has been reviewed and discussed.
Figure 1. Time-Altitude plot of signal to noise ratio (SNR) observed on (a) 18 November 1998 and (b) 11 January 1999. (c) and (d) are nightly mean temperature profile observed on respective nights along with standard deviation.
Figure 2.Wavelet analysis applied to the radial velocities obtained using MST radar on 18 November 1998 at 70.8 km and 18 km (a,b,e,f) and for Lidar temperature at 45.1 km and 77.2 km (c,d). The black line represents the cone of influence.
Figure 3. (a) and (b) show 30 min. to 1 h and 1 to 2 h period oscillations along with mean temperature (c) on the Inversion day (18 November 1998) (top panel). (d), (e) and (f) are same as Figs.3(a), (b) and (c) but during Non-Inversion day (11 January 1999) (bottom panel).
Figure 4. (a)Vertical wavelength of 20 min.-2 h GWs and vertical profiles of (b) Amplitude, (c) Zonal momentum, and (d) Brunt-Vaisala frequency on Inversion day (18 November 1998).
Figure 5. Zonal (left panels) and meridional (right panels) momentum fluxes at different altitudes observed on 18 November 1998 in the troposphere (bottom panels) and mesosphere (top panels).
Englishhttp://ijcrr.com/abstract.php?article_id=2411http://ijcrr.com/article_html.php?did=2411
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Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241101EnglishN2018January10HealthcareSecondary Ocular Involvement in Acute Myeloid Leukemia– M0: A Rare Case Report
English1012Deepika JoshiEnglish Ashok Kumar BairwaEnglish Sourav ShristiEnglishWe report a case of 45 year old female patient who presented with ocular pain and blurring of vision in both eyes (BCVA in right eye 0.1 logMAR and in left eye 0.1 logMAR). An ophthalmological evaluation revealed bilateral pale conjunctiva, vitreous degeneration, dilated and tortuous retinal vessels with multiple flame shaped and dot-blot haemorrhages, cotton wool spots and roth spots in both eyes. IOP in both eyes was 10 mm Hg. Haematological evaluation revealed the presence of Acute Myeloid Leukemia-M0 (undifferentiated) with Hb 3.2 gm%. General physical examination showed severe pallor. On examination bilateral
basal crepts and sternal tenderness found. Other causes of severe anaemia have been ruled out. This case documents the occurence of leukemic retinopathy in rare type of leukemia i.e Acute Myeloid Leukemia-M0 (3-5% of Acute Myeloid Leukemia).
Differential diagnosis: Thrombocytopenia, Severe anaemia, Hyperviscocity.
EnglishAcute Myeloid Leukemia– M0, Roth spots, Basal crepts, Sternal tenderness, Leukemic retinopathyIntroduction
Acute Myeloid Leukemia is subdivided into M0 – M7 . Acute Myeloid Leukemia- M0 is rare ( 3 – 5 % of Acute Myeloid Leukemia).3Ocular involvement in leukemia is classified as (1) Direct or primary leukemic infiltration (2) Indirect or secondary ocular involvement . The direct leukemic infiltration show three patterns: anterior segment uveal infiltration , orbital infiltration and neuro-ophthalmic signs of C.N.S. The secondary changes are result of haematological abnormalities of leukemia such as anaemia, thrombocytopenia, hyperviscocity . These can manifest as retinal or vitreous haemorrhage and vascular occlusions .1 The retina is involved more often than any other ocular tissue. It is estimated that 69 % of all patients with leukemia show fundus changes at some point of course of their disease.4
Case report
A 45 year old female patient referred to our ophthalmology department with complaint of ocular pain and blurring of vision in both eyes . She also had shortness of breath and easy fatigability. There was no history of any systemic afflictions such as Diabetes mellitus , HTN , any vasculitis, blood dyscrasias, blood transfusion . On general examination patient was calm, cooperative and conscious. Severe pallor was present, afebrile, P.R – 114/min , B.P – 130/90, R.R – 18/min . Systemic examination revealed bilateral basal crepitations and sternal tenderness.
On ophthalmological examination : Best corrected visual acuity was 0.1 logMAR (OD) and 0.1 logMAR (OS) . Conjunctiva was pale . Slit lamp examination revealed bilateral vitreous degeneration. I.O.P was 10 mm Hg (OD) and 10 mm Hg (OS) by NCT. Dilated fundus examination showed clear media with normal sized optic disc. The vessels were dilated and tortuous with multiple flame shaped and dot blot haemorrhages in all four quadrants of eye . Multiple cotton wool spots and Roth spots were present in both eyes .
In R.N.T Medical College all routine investigations were performed in which Hb = 3.2gm/dl ; RBC = 1.14 m/cu mm; TLC = 14,400( raised ) ; DLC = Lymphocytes = 58.5% ( High) , Granulocytes = 24 .6 % ( Low) ; MCHC = 30.70 (low) ; Reticulocyte count = 2.00 ; ESR = 49 ; SGOT =546 (raised) ; SGPT (raised) ; HCV antibodies = NR ; MP QBC = negative ; VDRL = NR ; HBsAg = negative. Peripheral blood smear - RBC – moderate anisopoikilocytosis, moderate hypochromic, pencil cells seen
WBC – 18000 cells / cu mm (Raised count)
DLC – Blast cells: 50% (raised)
Lymphocytes: 10%
Polyneutrophils: 25 %
Band form: 7%
Myelocytes: 8%
PLATELETS – adequate in number.
HAEMOPARASITES – not seen
Based on these ophthalmological, systemic and haematological findings diagnosis of Acute Myeloid Leukemia– M0 with CHF was made.
TREATMENT RECEIVED : The patient received 2 units of blood transfusion(Packed Cell Volume) , Vit B 12 and folate supplements and Injection Furosemide 2 amp I.V stat in our hospital and after she was diagnosed with Acute Myeloid Leukemia– M0 patient was referred to Haemato – Oncologist for further management .
Discussion
Leukemias are malignant neoplasms of haematopoetic stem cells, characterized by diffuse replacement of bone marrow by neoplastic cells . Ophthalmic involvement can be classified into two major categories (1)Primary or Direct leukemic infiltration (2.) Secondary or Indirect involvement. The direct leukemic infiltration show three patterns: anterior segment uveal infiltration, orbital infiltration and neuro-ophthalmic signs of C.N.S leukemia that include optic nerve infiltration, cranial nerve palsies and papilloedema. The secondary changes are the result of haematological abnormalities of leukemia such as anaemia, thrombocytopenia, hyperviscocity and immunosuppresion. These can manifest as Retinal or Vitreous haemorrhage, infections and as vascular occlusions. The ophthalmic manifestations of leukemia tend to involve more of posterior than anterior segment structures of eye and resulted more from secondary haematological complications rather than primary leukemic infiltration .1
In the era before effective antileukemic therapy, retinopathy was believed to be of no prognostic significance in acute leukemia. However recent reports have demonstrated that the presence of ocular involvement is associated with poor prognosis in acute childhood leukemias. Therefore it is important to consider an ophthalmic evaluation at the time of diagnosis of acute leukemia in adults and children2.
The acute leukemias are characterized by replacement of bone marrow with very immature cells called BLASTS. Under normal conditions, blasts forms constitute fewer than 5% of nucleated cells of bone marrow and are seen in peripheral blood except during periods of profound over-production of blood cells. Blast cells are primitive precursors lacking many of features of differentiation. Lymphoid blasts are differentiated from myeloid blasts on the basis of standard morphologic and cytochemical differences, based on these acute leukemias are subdivided . Within French – American – British (FAB) classification, ALL has been subdivided into three types : L1 , L2 , L3 , while subdivisions of Acute Myeloid Leukemia are called M0 – M7. Acute Myeloid Leukemia-M0 is rarest (3-5% of leukemias).3
The retina is involved in leukemia more often than any other ocular tissue. It is estimated that 69 % of all patients with leukemia show fundus changes at some point of course of their disease.4
The early manifestations (because of haematological disturbances) are venous dilation and tortousity .5
Haemorrhages may occur in all levels of retina, usually in posterior pole and may extend into vitreous . They may be round or flame shaped and often has white component. The white area consists of leukemic cells and debris, platelet fibrin aggregates or septic emboli. A similar clinical picture can be seen in severe anaemia , thrombocytopenia and hyperviscocity.6
Cotton wool spots can be seen and are probably due to ischaemia from anaemia, hyperviscocity or leukemic infiltrate7
In this case patients complete systemic , ophthalmological and haematological examination clearly demonstrates Acute Myeloid Leukemia– M0kind of Acute Myeloid Leukemia(Acute Myeloid Leukemia) .
Conclusion:
This is rare case entity as very few cases of ACUTE MYELOID LEUKEMIA– M0 are reported in literature . Although the ophthalmologist has a secondary role in treatment of leukemia , a prompt recognition of the ocular manifestations and their importance as a sign of possible extramedullary disease is crucial if appropriate therapy is to be initiated . We suggest that full collaboration among Physicians , Oncologists , Haematologists and Ophthalmologists is needed with prompt ophthalmic assessment of patients suspected to have eye manifestations .
Englishhttp://ijcrr.com/abstract.php?article_id=2413http://ijcrr.com/article_html.php?did=24131 . Reddy SC, Menon BS. A prospective study of ocularmanifestations in childhood acute leukaemia. ActaOphthalmol Scand 1998; 76: 700–703
2. Ohkoshi K, Tsiaras WG. Prognostic importance ofophthalmic manifestations in childhood leukaemia. Br JOphthalmol 1992; 76: 651–655
3. Willman CL. Acute leukemias: a paradigm for the integration of new technologies in diagnosis andclassification. Mod Pathol 1999; 12: 218–228
4.Alemayehu W, Shamebo M, Bedri A, Mengistu Z. Ocular manifestations of leukaemia in Ethiopians. Ethiop Med J 1996; 34: 217–224
5.Ballantyne AJ, Michaelson IC. Textbook of the Fundus of theEye. Williams and Wilkins: Baltimore, 1970; 290–292
6.Allen RA, Straatsma BR. Ocular involvement in leukaemia and allied disorders. Arch Ophthalmol 1961; 66: 490–508
7. Sharma T, Grewal J, Gupta S, Murray PI. Ophthalmic manifestations of acute leukaemia: The ophthalmologist’s role. Eye 2004;18:66
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241101EnglishN2018January10HealthcareA Comparative Study of Isolation of Obligate Anaerobes and Facultative Anaerobes in Microbial Flora of Non- Vital Teeth in Diabetic (Type II) and Non-Diabetic Patients
English1317Pradeep ShettyEnglish Monika ChawlaEnglish Sanjyot MulayEnglish Karan BhargavaEnglish Tanaya KumarEnglish Vasudevan VidhyaEnglishIntroduction: The aim of the study was to evaluate and compare the anaerobic microflora (obligate and facultative) of the root canals of non vital teeth with periapical lesions, in diabetic (Type II) and non-diabetic patients.
Methods: Access cavity preparation and collection of necrotic pulp tissue samples was done aseptically with the help of absorbent paper points placed in the canal for 60 seconds. Samples were processed anaerobically and were subjected to microbiological analysis using culture techniques for the evaluation of microbial flora.
Results: There was a statistically significant difference between mean and SD values of obligate and facultative anaerobes in diabetic (Type II) and non-diabetic groups when all parameters are compared (pEnglishNecrotic pulp, Diabetic type II, AnaerobesIntroduction:
Around 61 million people in India are diabetic. International statistics estimate that 366 million people had diabetes mellitus in 2011; and that by 2030, this number will rise to 552 million. Around 983,000 deaths in India were caused due to diabetes in the last year.1
A group of complex multisystem metabolic disorders because of deficient insulin secretion due to dysfunction of pancreatic β-cell and/or insulin resistance in muscle as well as liver is termed as Diabetes Mellitus.2 According to findings; diabetes mellitus aggravates the risk of caries, gingivitis and periodontal disease. Increased local inflammation causes intensive increase in the level and effects of diabetes with a sharp shoot up in blood glucose level, leading to uncontrolled hyperglycemia of the patient. Thus all the causative infections concerning those of the dental pulp should be mandatorily removed.3
Literature has very well documented the complex relationship between diabetes mellitus and periodontal disease. However, the association between diabetes mellitus and dental pulp and the advancement and repair of different endodontic infections has not been satisfactorily studied.3
There is an established positive, besides insignificant alliance between diabetes and presence of Porphyromonas, Prevotella, Eubacteria, Fusobacterium and Enterococcus species.5 Foregoing studies have demonstrated that in retaliation to pernicious root canal microflora particularly bacteria, the diabetic host may mature with more consequential infections.4 When the diabetic and non-diabetic population is compared, patients with diabetes mellitus and periapical lesion have shown notably diminished healing succeeding endodontic therapy.6
Hence, this study aims to isolate obligate anaerobes and facultative anaerobes from root canal of non-vital teeth in diabetic (Type II) and non-diabetic patients.
Material and Methodology:
Patient Selection
Patients presenting to the Department of Endodontics and Conservative Dentistry, Dr. D Y Patil Dental College were screened for the study. Comprehensive medical and dental histories were obtained from each patient. Inclusion criteria for the study were teeth with pulpal disease and periapical lesions which could be treated with endodontic therapy, patients with a history of controlled diabetes (Type II), patients either taking oral medication or insulin for the disease, patients in the age range of 45-65 years were selected. Written consent was obtained from all participating patients. Patients who met the inclusion criteria were subjected to electric pulp testing and pre-operative radiograph to investigate the presence of periapical lesion.
Group 1: 30 patients with type II diabetes mellitus were subjected to HbA1C test to determine the degree of their glycemic control. [≥7mmol(≥126mg/dl)]
Group 2: 30 patients with no history of diabetes were selected. HbA1C test was carried out on patients with no history of diabetes to confirm their non-diabetic status. [Englishhttp://ijcrr.com/abstract.php?article_id=2414http://ijcrr.com/article_html.php?did=2414
http://articles.timesofindia.indiatimes.com/2011-12-14/india/30515422 1 diabetic- patients- high- blood- sugar-level (14th December 2011).
Juan J Segura-Egea 1, Lizett Castellanos-Cosano 1, Guillermo Machuca et al. Diabetes mellitus, periapical inflammation and endodontic treatment outcome. Med Oral Patol Oral Cir Bucal. 2012 Mar 1;17 (2):356-61.
IB. Bender and A. B. Bender. Diabetes Mellitus and the Dental Pulp. J Endod2003;29(6):383–9.
K V Ramana. Clinical Microbiology: Reemphasizing the Role of Anaerobic Bacteria in Human Infections. J Med Microb Diagn 2012, 1:4.
Ashraf F. Fouad. Diabetes Mellitus as a Modulating Factor of Endodontic Infections. J Dent Educ 2003;67(4):459-467.
Ashraf F. Fouad, Jody Barry, Melissa Caimano et al. PCR-Based Identification of Bacteria Associated with Endodontic Infections.J. Clin. Microbiol 2002;40(9):3223-31.
Bissada NF, Sharawy AM. Histologic study of gingival and pulpal vascular change in human diabetics. Egypt Dent J 1970;16:283-96.
Bennett C.M, Guo M, Dharmage S.C. HbA1c as a screening tool for detection of type 2 diabetes: a systematic review. Diab Med 2007; 24: 333–343
Pishipati Vinayak Kalyan Chakravarthy. Diabetes mellitus: An endodontic perspective. Eur J Dent 2013;2(3).
Falk H, Hugoson A, Thorstensson H, Number of teeth, prevalence of caries and periapical lesions in insulin dependent diabetes. Scand J Den Res 1989; 97: 198-206
Lamster IB, Borgnakke WS, Taylor GW. The relationship between oral health and Diabetes mellitus. JADA 2008; 139: 19s-24s
Nayak Moksha, et al. Diabetes mellitus and apical periodontitis. Endodontology (2013): 103-108.
Möller AJ, Fabricius L, Dahlen G. Influence on periapical tissues of indigenous oral bacteria and necrotic pulp tissue in monkeys. Scand J Dent Res. 1981;89(6):475-84.
Alencar AH, Pimenta FC, Ito IY, Bruno KF, Leonardo MR. Determinação dos microrganismos no canal radicular, antes do preparobiomecânico e após a utilização da medicaçãointracanal, emdentes com necrosepulpar e reaçãoperiapicalcrônica. ArqOdontol.2005;41(2):105-92.
Brown Jr. LR, Rudolph Jr. CE. Isolation and identification of microorganisms from unexposed canals of pulp-involved teeth. OralSurg. Oral Surg. 1957;10(10):1094-9
Socranscky SS, Mcdonald JB, Sawyer S. The cultivation of Treponemamicrodentium as surface colonies. Arch Oral Biol. 1959;1(2):171-2
Winkler KC, Van Amerongen J. Bacteriologic results from 4.000 root canal cultures. Oral Surg Oral Med Oral Pathol. 1959;12(7):857-75
Drucker, D. B. and Natsiou, I. Microbial ecology of the dental root canal. Microb Ecol Health Dis 2000 12, 160–169
Fabricius L, Dahlén G, Holm SE, Möller AJR. Influence of combinations of oral bacteria on periapical tissue of monkeys. Scand J Dent. 1982;90(3):200-6.
Gomes BP, Lilley JD, Drucker DB. Associations of endodontic symptoms and signs with particular combinations of specific bacteria. Int Endod J. 1996;29(2):69-75.
Assed S, Ito IY, Leonardo MR, Silva LAB, Lopatin D. Anaerobic microorganisms in root canals of human teeth with chronic apical periodontitis detected by indirect immunofluorescence. Endo Dent Traumatol. 1996;12(2):66-9
Morse DR. Endodontic microbiology in the 1970s. Int Endod J. 1981;14(2):69-79.
Ferreira CM, Bonifácio KC, Fröner IC, Ito IY. Evaluation of the antimicrobial activity of three irrigating solutions in teeth with pulpal necrosis. Braz Dent J. 1999;10(1):15-21
Haapasalo M. Bacteroidesspp in dental root canal infections. Endod Dent Traumatol. 1989;5(1):1-10.
de Lima Guimarães, Soares NL, Otoch, Machado H, de Andrade, Cavalcante L. Microbiological evaluation of infected root canals and their correlation with pain. Rev Sul-Bras de Odontol 2012;9:31
Van Winkelhoff AJ, Carlee AW, de Graaff J. Bacteroidesendodontalis and other black-pigmented Bacteroides species in odontogenic abscesses. Infect Immun 1985;49:494–8.
Sundqvist G, Johansson E, Sjo¨gren U. Prevalence of black-pigmented Bacteroides species in root canal infections. J Endodon 1989;15:13–9.
Sundqvist GK, Eckerbom MI, Larsson AP, Sjo¨gren UF. Capacity of anaerobic bacteria from necrotic dental pulps to induce purulent infections. Infect Immun 1979;25:685–93.
Siqueira JF Jr, Rôças IN. Exploiting molecular methods to explore endodontic infections: Part 1—current molecular technologies for microbiological diagnosis. J Endod. 2005 Jun;31(6):411-23.
Ferrari PHP, Cai S, Bombana AC. Effectof Effect of endodontic procedures on enterococci, enteric bacteria and yeasts in primary endodontic infections. IntEndodJ.2005;38(6):372-80 Int Endod J. 2005;38(6):372-80.
Dougherty WJ, Bae KS, Watkins BJ, Baumgartner JC. Black-pigmented bacteria in coronal and apical segments of infected root canals. J Endod. 1998;24(5):356-8.
Drucker DB, Gomes BPFA, Lilley JD. Role of anaerobic species in endodontic infection. Clin Infect Dis. 1997;25(2):220-1.
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241101EnglishN2018January10HealthcareHydroprocedures- The Miracle Tool of Phacoemulsification: Visual Outcome and Complications of Fifteen Cases of Phacoemulsification Performed in a Regional Institute of Ophthalmology
English1821Garima AgrawalEnglishIntroduction: It has been our observation that in phacoemulsification many layers of hydrodelineation help in naturally cleaving the natural crystalline lens layer by layer. Subsequent emulsification of each layer is easy. The remaining epinuclear and cortical plate is easy to remove by repeating hydrodissection or hydrodelineation as required.
Aim: The aim of this study has been to document the visual outcome and complications of fifteen cases of phacoemulsification of patients with nuclear sclerosis one and two using multiple layers of hydrodelineation performed at our Regional Institute of Ophthalmology.
Methodology: Fifteen patients having visually significant cataract with nuclear sclerosis one and two were enrolled in our study. Phacoemulsification was performed in all fifteen eyes via the temporal approach. We created multiple waves of hydrodelineation at deeper nuclear levels as identified on the slit lamp examination as natural layers of the endonucleus at approximately one mm depth intervals. Thus we created as small a central nuclear chunk as possible. This was then held by the phaco probe and directly chopped and emulsified. The remaining nuclear, epinuclear and cortical layers were removed layer by layer via gripping
with phaco probe at edge of shell and subsequently emulsifying the layer. The other steps were as per protocol .
Results: All the fifteen patients had good postoperative visual recovery with no major intraoperative or postoperative complications for the reported period of six weeks.
Discussion: We report successful phacoemulsification using multiple layers of hydroprocedures to “divide” the nucleus along its natural cleavage planes followed subsequently by emulsification.
Conclusion: We report successful phacoemulsification using multiple layers of hydroprocedures to “divide” the nucleus along its natural cleavage planes followed subsequently by emulsification.
EnglishHydroprocedures, Phacoemulsification, Layer by layerIntroduction
It has been our observation that in phacoemulsification many layers of hydrodelineation help in naturally cleaving the natural crystalline lens layer by layer. Subsequent emulsification of each layer is easy. The remaining epinuclear and cortical plate is easy to remove by repeating hydrodissection or hydrodelineation as required. This increases safety levels as there is always a plate of epinucleus to protect the posterior capsule. We have found this to be a safe method for early cataracts as nuclear sclerosis one and two grade as well as posterior polar cataract. This method has been documented for posterior polar cataracts but not for nuclear sclerosis type of cataracts.1
We designed a study to document the visual outcome and complications of fifty cases of phacoemulsification using many layers of hyrdodelineation followed by emulsification. The cases were performed at our Regional Institute of ophthalmology by a single experienced surgeon.
The crystalline lens is a highly ordered structure with a close orderly packing of lens fibre cells. The lens has an elliptical shape and occupies the space between the iris and the vitreous. It is held in a fairly fixed position by the zonular fibres that run from the lens to the ciliary body and by its close approximation to the vitreous on its posterior and equatorial sides. The lens fibres with their highly organized concentric shells make up the bulk of the lens.
In infants the cortical lens cells surrounding the fetal nucleus are nucleated. As new cells form the older ones are displaced deeper into the cortex and lose their nuclei. The deeper cells lose their nuclei and their outlines become less defined. By thirty years of age a considerable number of older cells have accumulated to form the lens nucleus. The cells comprising the lens cortex surround the nucleus and many of them retain their nuclei. The cells of the lens cortex and nucleus have come to be known as lens fibres because of their length. Cells from the equator migrate towards the anterior and posterior poles.
We have observed that the colour of the lens including the capsule, cortex, epinucleus and nucleus deepens or darkens progressively as we move from anterior to posterior. Authors have observed and correlated the hardness of the lens with its colour. It is mentioned that as there is synthesis of secondary lens fibres, cells from the equator migrate towards anterior and posterior poles, pushing the older cells deeper .Thus the lens is formed in concentric layers.
Hydroprocedures are a documented step of phacoemulsification. Fine’s cortical cleaving hydrodissection is in the plane between the cortex and the capsule. There is a plane between cortex and nucleus seen as a dark zone in early nuclear sclerosis grade one and two cataracts. Definite mention has not been made of this zone in texts. A plane of fluid wave has to be passed in this zone. There is another dark zone between the epinucleus and the endonucleus. A fluid plane is also created in this zone. Within the endonucleus too there are concentric arrangement of lens fibres which may be cleaved by planes of fluid.
The term hydrodelineation was introduced by Anis to describe the act of separating an outer epinuclear shell or multiple shells from the compact inner nuclear material, the endonucleus by forceful irrigation of fluid into the inner nuclear mass.
It has been our observation that although the cortex, epinucleus and endonucleus are concentric still the posterior cortex, epinucleus and endonucleus are browner than the anterior counterparts as if we could draw a vertical line from the superior equator to the inferior equator to divide the lens into two zones, an anterior softer zone and a posterior harder zone. We also observed that not only the nucleus colour but also the compactness of the lens matter observed on slit lamp biomicroscopy may contribute to its hardness.
In phacoemulsification after capsulorrhexis we remove the anterior cortex ,epinucleus and superficial endonucleus by aspiration alone and it comes easily. The posterior endonucleus, epinucleus and cortex are left. The posterior endonucleus ( the core lens matter ) is held with the phaco probe and alternately vertically chopped and emulsified. The softer lens layers are the gripped and emulsified/aspirated layer by layer. The remaining cortex is removed by irrigation- aspiration.
Aim
The aim of this study has been to document the visual outcome and complications of fifteen cases of phacoemulsification of patients with nuclear sclerosis one and two using multiple layers of hydrodelineation performed at our Regional Institute of Ophthalmology.
Methodology
The study was carried out at our Regional Institute of Ophthalmology from January 2017 to April 2017. Fifteen patients with visually significant cataract with nuclear sclerosis one and two were enrolled in our study. All patients were subjected to a thorough anterior and posterior segment examination, intraocular pressure measurement and intraocular lens power calculation.
Phacoemulsification was performed in all fifteen eyes via the temporal approach. A side port was made with microvitreoretinal blade on the right hand side of the surgeon .1.4% sodium hyaluronate was then injected into the anterior chamber. Capsulorrhexis was performed using a 26 and half guage needle. Another side port was then made at 180 degrees to the first port with a microvitreoretinal blade. Main port was then made using a 3.2 mm keratome at the temporal side. Fine’s cortical cleaving hydrodissection was then carried out using a 27 guage hydrodissection cannula filled with normal saline mounted on a 5 ml syringe. The anterior capsule edge was lifted and freed from the underlying cortex for one mm. The edge was lifted and fluid was injected .The end point of hydrodissection was determined by completion of the fluid wave and lifting up of the nucleus. The nucleus was gently pushed posteriorly. The cannula was then inserted into the cortex and the epinucleus and fluid was injected at the first point of resistance. The end point was noted as completion of the fluid wave and appearance of a golden rim.
Subsequently we created multiple waves of hydrodelineation at deeper nuclear levels as identified on the slit lamp examination as natural layers of the endonucleus at approximately one mm depth intervals. Thus we created as small a central nuclear chunk as possible. This was then held by the phaco probe and directly chopped and emulsified.
Thereafter the remaining nuclear, epinuclear and cortical layers were removed layer by layer via gripping with phaco probe at the edge of the shell and subsequently emulsifying the layer.
Phacoemulsification was completed thus in all the fifteen eyes . Foldable intraocular lens was then implanted under the cover of 1.4% sodium hyaluronate in the capsular bag. Viscoelastic wash was carried out followed by wound closure.
All patients were examined on first postoperative day. Best corrected visual acuity was documented along with intraocular pressure. Thorough anterior segment examination was carried out using slit-lamp biomicroscopy. The patients were followed up on day four, 2 weeks, four weeks and 6 weeks. Final best corrected visual acuity was documented at six weeks. Thorough anterior segment and dilated posterior segment examination was done to account for best corrected visual acuity less than 6/6.
Results
All the fifteen patients had good postoperative visual recovery with clear cornea and intraocular lens in place and a round, regular reacting pupil on first postoperative day. The best corrected visual acuity at 6 weeks postoperative was 6/6 in 12 patients. Two patients had best corrected visual acuity of 6/18 due to dry age related macular degeneration changes. One patient had a best corrected visual acuity of 6/36 due to macular dystrophy.
There were no major intraoperative or postoperative complications for the reported period of six weeks.
Discussion
We report successful phacoemulsification using multiple layers of hydroprocedures to “divide” the nucleus along its natural cleavage planes for grade one and two nuclear sclerosis cataracts followed subsequently by emulsification.
Vajpayee RB et al first reported ‘ Layer by layer ’ phacoemulsification in posterior polar cataract with pre-existing posterior capsular rent. They described the technique as a single ring of hydrodelineation to separate the inner nucleus from the superficial epinucleus and cortex .1 The inner firm nucleus was phacoemulsified. Each layer of epinucleus and cortex was sequentially aspirated in layers using an automated bimanual irrigation and aspiration cannula. They reported successful phacoemulsification followed by implantation of a foldable intraocular lens in the bag.
We document this same procedure for nuclear sclerosis grade one and two cataracts. The procedure reduces the amount of phaco energy used as we are emulsifying a small nucleus. The softer layers can be taken by aspiration alone. The procedure keeps the posterior capsule protected by its layers of softer lens material till nucleus management is done.
The procedure is a safe and effective addition to the current armamentarium of phacoemulsification procedures and complements them.
Conclusion
We report successful phacoemulsification of nuclear sclerosis grade one and two cataracts using multiple layers of hydroprocedures to “divide” the nucleus along its natural cleavage planes followed subsequently by emulsification. The procedure has been previously reported for posterior polar cataracts.
We document this procedure in early nuclear sclerosis cataracts. This procedure represents a safe and effective addition to the current armamentarium of phacoemulsification methods.
Acknowledgement
Authors acknowledge the immense help received from the scholars whose articles are cited and included in references of this manuscript. The authors are also grateful to authors/editors/publishers of all those articles journals and books from where the literature for this article has been reviewed and discussed.
Ethical Clearance: Taken
Informed Consent: Taken
Source of funding : none
Conflict of Interest: none
Englishhttp://ijcrr.com/abstract.php?article_id=2415http://ijcrr.com/article_html.php?did=2415
Jaffe NS, Jaffe MS, Jaffe GF. Surgical Anatomy. In: Jaffe NS, Jaffe MS, Jaffe GF. Cataract Surgery and its Complications. 6th Ed. Mosby;1997. p 48-51.
Schell J, Boulton ME. Basic Science of the Lens. In: Yanoff M, Duker JS, editors. Ophthalmology.4th Ed. China: Elsevier Saunders;2009. p 329.
Jaffe NS, Jaffe MS, Jaffe GF. Surgical Technique. Hydrodissection and Hydrodelineation. In: Jaffe NS, Jaffe MS, Jaffe GF. Cataract Surgery and its Complications.6th Ed.Mosby;1997.p72.
Packer M. Hydrodissection and Hydrdelineation .Small Incision and Femtosecond Laser Cataract Surgery. In: Yanoff M, Duker JS, editors. Ophthalmology.4th Ed. China: Elsevier Saunders;2009. p 372.
Packer M. Nucleofractis Techniques. Small Incision and Femtosecond Laser Cataract Surgery. In: Yanoff M, Duker JS, editors. Ophthalmology.4th Ed. China: Elsevier Saunders;2009. p 372-373.
Vajpayee RB, Sinha R, Sanghvi A, Sharma N, Titiyal JS, Tandon R. ‘Layer by layer ‘ phacoemulsification in posterior polar cataract with pre-existing posterior capsular rent. Eye 2008;22:1008-1010.
Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241101EnglishN2018January10HealthcareStudies on In-Vitro Interaction of Artemether-Lumefantrine with Food Components
English2226Awofisayo Sunday O.English Arhewoh Matthew I.English Okhamafe Augustine O.EnglishThis work evaluates in vitro interaction of food components with artemether–lumefantrine (AL) in tablet. Pelletized food components (i.e., starch, albumin, sunflower oil and carbonated drink) with AL tablet were observed using Fourier transform infrared (FTIR) spectroscopy while spectra obtained were compared using essential FTIR (eFTIR) software. The influence (i.e., instantaneous change in pH (IPH) and acid buffering capacity (ABC)) of food components on Fasted State Simulated Gastric Fluid (FaSSGF) and Fed State Simulated Intestinal Fluid (FeSSIF) were assessed alongside dissolution kinetics of actives in AL tablets. Artemether and lumefantrine produced identical spectra features with reference infra red (IR) spectra. No change in spectra features of AL on carbohydrate, vitamin A and oil treatment but noted with albumin and the carbonated drinks. IPH values for albumin revealed statistical higher value (p < 0.05) than other food components in FaSSGF. ABC for albumin treatment was statistically lower (p < 0.05) than for other food components in FaSSGF. Artemether overall release was significantly reduced by starch, lactose and carbonated drink (p < 0.05) while that of lumefantrine was significantly increased by sunflower oil (p < 0.05). There was interaction between AL and albumin, or carbonated drinks which can be of importance in drug treatment outcome.
EnglishArtemether-lumefantrine; Food component; In vitro interaction; Dissolution kinetics; Instantaneous pH change, Acid buffering capacityINTRODUCTION
The oral route is the most convenient for drug administration when treating uncomplicated Plasmodium falciparum malaria in the tropics. Consequently, a majority of World Health Organization (WHO) recommended drugs are presented as solid formulations for oral administration (Katzung, 2009). Artemether-lumefantrine (AL) is a co-formulated artemisinin combination therapy (ACT) available for paediatric and adult use and widely prescribed across sub-Saharan Africa (Katzung, 2009).
Factor that are critical to the kinetics of dissolution as presented by the Nernst Brunner and Levieh modifications of the Noyes-Whitney model include physiochemical properties of the dissolving compound (i.e., pKa, solubility, crystalline energy, specific surface area) and certain prevailing conditions of the gastrointestinal (GI) tract) (Kostewicz et al, 2002). Physiological parameters that play an important role include pH, surface tension and volume of the GI luminal fluid, solubilization and buffer capacity of substances present in the GI tract (Dressman et al, 2001). These parameters change following ingestion of food (Schmidt and Dalhoff, 2002). Amphiphilic bile components (i.e., bile salts and lecithin) may increase in concentration following a meal (Martinez and Amidon, 2002). Biliary secretion has been demonstrated to increase the in vivo dissolution rate for some poorly soluble compounds. In order to develop a predictive in vitro model to forecast the in vivo performance of drugs, the Fasted State Simulated Gastric Fluid (FaSSGF), Fed State Simulated Gastric Fluid (FeSSGF), Fasted State Simulated Intestinal Fluid (FeSSIF) and Fed State Simulated Intestinal Fluid (FeSSIF) were proposed and are now widely employed (Vertzoni et al, 2005). FeSSIF-V2 (version 2) was further advanced following its close compositional resemblance to intestinal fluid (Jantratid et al, 2008).
In the present study, the aim was to assess in vitro interaction of food components with AL tablet.
MATERIALS AND METHODS
Materials
AL tablet (Coartem®) was purchased in Uyo, Southern Nigeria. The food components were similarly purchased in Lagos, Nigeria. FaSSIF/FeSSIF/FaSSGF and FeSSIF-V2 powder were products of biorelevant.com, UK. Details of the drug and food components are presented in Table 1. Acetonitrile, tetrahydrofuran (THF) and potassium dihydrogen phosphate were of HPLC grade, products of Sigma Aldrich, Germany. All other reagents were of analytical grade.
Methods
Preparation of standard solution of internal standard, artemether and lumefantrine
Ten milligram of halofantrine was transferred into a 10 mL volumetric flask containing 6.0 mL of methanol to dissolve. The powder was dissolved and thereafter made up to mark to produce stock concentration of 1000 µg/mL. Reference standard powder of artemether 20 mg and lumefantrine 120 mg were weighed accurately into different flasks. Artemether and lumefantrine powder were dissolved to produce stock solutions of 2 mg/mL and 12 mg/mL, respectively.
Preparation of mixed standard solutions of AL and (IS)
Equal volumes (2 mL) of artemether and lumefantrine stock solutions were dispensed into 5 mL sample bottles to produce 1 and 6 mg/mL of the standards, respectively. Serial dilutions of the mixed standard and reference standard solutions were made to obtain graded concentrations of AL ranging from 0.01/10 to 2/10 mg/mL. Mixed standard solutions were prepared using acetonitrile and THF (50:50, %) as diluents. The resulting solutions were spiked with IS solution to give 5 mg/mL.
Preparation of hydrochloric acid/sodium chloride solution pH 1.6
A weight of 1.0 g of sodium chloride was dissolved in 0.450 L of distilled water. The pH was adjusted to 1.6 with HCl. Either HCl or NaOH were spiked as applicable to give the exact pH 1.6, thereafter made up to 0.5 L with distilled water.
Preparation of sodium hydroxide/glacial acetic acid/sodium chloride–Buffer solution pH 5
A weight of 2.020, 4.325, and 5.937 g of sodium hydroxide pellet, glacial acetic acid and sodium chloride, respectively, were dissolved in 0.450 L of distilled water. The pH was adjusted with either of 1N HCl or NaOH solution.
Preparation of FaSSGF, FeSSIF and FeSSIF–V2
A weight of 1.120 g of FaSSIF/FeSSIF powder was dissolved in 0.25 L of acid buffer (pH 1.6). This was stirred until the powder was completely dissolved. The solution was made to 0.5 L mark. A weight of 5.60 g of FaSSIF/FeSSIF/FaSSGF powder was dissolved in 0.25 L of buffer (pH 5.0). The solutions were made up to volume with the buffer and allowed to stand for 2 h before use. FeSSIF-V2 was prepared by dissolving 5.0 g of FeSSIF-V2 powder in 5.0 L of distilled water.
Spectroscopic determination
Infra red (IR) studies were performed using Fourier transform infra-red (FTIR) spectrophotometer, FTIR-84005 (Schimadzu, Japan). One milligram of AL powder and 200 mg of dried KBr powder were mixed and compressed into a tablet shaped disk and scanned at a speed of 2 mm/s over a wavenumber region from 4000 to 500 cm-1. Essential FTIR (eFTIR) software was employed for spectral analysis.
Dissolution kinetics of AL in the presence of FeSSIF-V2
A total weight of 2 g of each solid food components or 1.7 mL of sunflower oil or 35 cL of carbonated drinks was dispersed in 500 mL FeSSIF–V2 in a flask. One tablet of the AL product was subsequently placed in the dissolution apparatus; USP type 2.The dissolution profile was determined by sampling 5 mL of the medium at 0, 5, 15, 30, 45, 60 and 90 min. Samples were analyzed using HPLC system.
Statistical analysis
The data obtained were analyzed using Minitab for Windows statistical package (Minitab, Inc., USA). Post hoc comparison was performed with Hsu’s test.
RESULT
Albumin had a pH change of 1.51 units in FaSSGF (Table 2). Similarly, it increased the pH in FaSSGF from 1.62 to 3.13 units and revealed an increase of 0.17 units in FeSSIF. Albumin also demonstrated significantly lower (p < 0.05) ABC compared to the carbohydrates in FaSSGF (Table 2).The ABC of Cocacola® was significantly higher (p < 0.05) than Fanta®. Carbonated drinks treatment presented the highest ABC. The extent of artemether release in the presence of fructose was statistically higher than for starch (pEnglishhttp://ijcrr.com/abstract.php?article_id=2416http://ijcrr.com/article_html.php?did=2416
Katzung BG. Basic and Chemical Pharmacology 11th Ed McGraw – Hill; 2009.
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Kostewicz ES, Branus U, Becker R, Dressman JB. 2002. Forecasting the oral absorption behaviour of poorly soluble weak bases using solubility and dissolution studies in biorelevant media. Pharm Res, 29:345 – 50.
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Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241101EnglishN2018January10HealthcareTotal Thyroidectomy for Large Goitres: Case series
English2730Nagarajan RaghupathyEnglish Suresh K. SinhaEnglishAim: To present the experience of performing Total thyroidectomy for Large Multinodular goitres.
Case Report: Series of nine patients with large multinodular goitres treated in our institute between April 2016 and March 2017 were reported here. All of them underwent Total thyroidectomy. The clinical evaluation, intra operative management and complications
were analysed retrospectively.
Discussion: In this series of nine patients,seven were Females and two were Males, age ranged from 26 to 68 years, seven of those patients had thyroid volume of more than 150 mL by neck ultrasonography and the other two had large goitres with retrosternal extension. Tracheal deviation was found in all nine patients and two patients had tracheal narrowing as well. Thyroid function test revealed three of the patients were hyperthyroid, one hypothyroid, and the rest euthyroid. Technetium(Tc99) scintigraphy showed cold nodule in only one patient out of the three hyperthyroid cases. Fine needle aspiration (FNAC) showed benign
pathology (Bethesda II) in six patients, one patient had follicular neoplasm (Bethesda IV). FNAC was not performed in two toxic goitres that did not have cold nodule. Total thyroidectomy was performed successfully in all nine patients.
Conclusion: Total Thyroidectomy can be safely performed even in large goitres; no incidence of tracheomalacia was observed; none had permanent hypoparathyroidism or recurrent laryngeal nerve injury.
EnglishLarge multinodular goitre, Total thyroidectomy, TracheomalaciaIntroduction
Benign multinodular goitres can be treated with thyroxine suppression therapy, radioactive iodine, or surgery (1). Total thyroidectomy for benign multinodular goitre circumvents reoperations and provides immediate relief from toxic symptoms inthe cases of hyperthyroidism(2). However, performing total thyroidectomy in large goitres is highly challenging due to the distorted anatomy.
Case Reports
Series of nine patients with large multinodular goitres treated in our institute between April 2016 and March 2017 were reported here, (seven Females and two Males), age ranged from 26 to 68 years. Clinical and ultra-sonographic examination of the neck was done in all nine patients. All of them had Grade 2goitreaccording to world health organization’s classification of goitres (3). Three patients were toxic without eye signs. Clinically lower border of the goitre was not palpable in two patients but Pemberton sign was negative.
Ultrasonography of the neck performed with 7.5 MHz probe by the same radiologist. It did not reveal any features of malignancy in all nine patients. Maximum dimensions (length, width, and thickness) were measured for each lobe. Each lobe volume was calculated by using the formula: length× width×thickness×0.479 (4).Addition of both lobe volumes gave the total volume of the gland. Volume of the thyroid gland was greater than 150 mL in seven patients and the other two patients had retrosternal extension. Volume of the gland could not be accurately measured by ultrasonography in patients with retrosternal extension. So, MRI or CT scan should be used for that purpose(5). As the gland was massive, CT scan was done in all patients to study the position of trachea, oesophagus, vasculature, and retrosternal extension. CT scan of neck revealed tracheal shift in all nine patients and two of them had tracheal narrowing as well (Figure 1 and 2). CT scan of chest was done along with neck in two patients, which showed retrosternal extension of multinodular goitre. Lower border of the goitre reached above D4 level (Figure 1 and 2).
Figure 1. CT scan of neck (axil view) showing large multinodular goitre with tracheal shift to right.
Figure 2. CT scan of neck and mediastinum (coronal view) showing large goitre with tracheal shift to right, narrowing of tracheal lumen and retrosternal extension.
Thyroid function tests revealed three (33%) patients to be hyperthyroid, one hypothyroid, and the others(55.5%)to be euthyroid (Table 1). Technetium(Tc99) scintigraphy was done in three patients whose TSH were supressed. Those three patients had high uptake and only one patient had a single cold nodule. Fine needle aspiration (FNAC) showed benign pathology (Bethesda II) in six patients and one patient had follicular neoplasm (Bethesda IV). FNAC was not performed in two toxic goitres without cold nodule. Mean serum calcium level was 9.03mg% (range:8.4–9.6 mg%) (Table 1).
Table 1. Preoperative summary of nine patients with large multinodular goitres.
Management and operative procedure
Toxic multinodular goitre patients were given a course of Carbimazole to achieve euthyroid status before surgery. One hypothyroid patient was started on Thyroxine and was made euthyroid before surgery. Video laryngoscopy wasperformed in all patients and it showed mobile vocal cords in all nine patients. Risks and complications of total thyroidectomy were explained to the patients and informed consent was taken. Adequate blood was reserved before surgery.
Fibreoptic scope was used for intubation. Nasogastric tube was put in all patients. It was useful in dissecting the mass without injuring the oesophagus. Skin incision was made from anterior border of one sternomastoid to the other and it was even longer in a two cases. Subplatysmal ?aps were raised and anterior jugular vein was ligated if required. Deep cervical fascia was opened in the midline. Both sternohyoid and sternothyroid were divided to get adequate exposure. Ligasure was used for dissection and vessel sealing. Middle thyroid vein was secured with energy device.Superior pedicle was dissected and ligated safely. Branches of inferior thyroid artery were coagulated and divided. All parathyroid glands were identified and preserved. In three patients, single parathyroid gland was auto transplanted in sternomastoid muscle due to injured vasculature(6). Recurrent laryngeal nerve was identi?ed in both sides in all patients and preserved. Ligasure minimised the blood loss (7,8)(Figure 3). Mean blood loss in our series was 106.6 mL. Retrosternal portion of goitres were gently dissected with finger while avoiding traction. Both retrosternal goitres were removed through cervical incision(9,10).Perfect haemostasis was secured. Tracheomalacia was ruled out and then the neck wound was closed with a suction drain. All nine patients were safely extubated at the end of surgery. Blood transfusion was not required in any case.
Figure 3. Intra operative picture of Thyroidectomy.
Postoperative complications
Transient hypoparathyroidism occurred in two out of nine cases (22.2%)and one patient had transient hoarseness (Table 2). Postoperative haematoma or infection did not happen in any of the cases. All of them discharged three days after the surgery. All nine specimens were benign by histopathology (Table 2).
Table 2. Intra and postoperative parameters in patients with large multinodular goitre.
Discussion
Multinodular goitre, enlarged to moderate to massive proportions, is very common, especially in patients hailing from rural India. Clinical, functional, ultrasonography with 7.5-10 MHz probe constitute the initial evaluation(1).CT scan is recommended in large goitres to assess the tracheal shift, adequacy of tracheal lumen, and retrosternal extension (11).Total thyroidectomy for benign multinodular goitre avoids reoperations and gives immediate relief from toxic symptoms in cases of hyperthyroidism. Complication rate is also low with meticulous dissection(2).
High dose radioiodine therapy has been triedin large goitres with volume more than 150 mL in both euthyroid and toxic thyroid patients. One third of goitre size reduction was achieved only after one year with high doses of I131 (26.7–124.9 mCi) therapy. Relief from toxic symptoms and compression werealso not immediate (5).
Intubation in massive goitres is a challenge to anaesthetist. Awake intubation was done using fibreoptic scopein allof our patients. Meticulous surgical techniquesare essential topreserve parathyroid and recurrent laryngeal nerve. Parathyroid gland has to be autotransplantedin cases of injury to its blood supply. Injured parathyroid gland has to be minced into thin slices and put into a pouch created in sternomastoid muscle. This measure circumventspermanent hypoparathyroidism (6). Ligasure energy device saves operating time and blood loss(7,8). Retrosternal goitres reaching above aortic arch (D4 level) can be removed through cervical route (9,10).
Long standing extrinsic compression over the trachea due to large goitres may soften the tracheal cartilages. It has been commonly suggested that after the removal of such large goitres, airway maycollapse in excess of 50% of diameter and warrant an emergency tracheostomy(12). After the removal of large goitre, tracheomalacia has to be ruled out. It can be done by the following steps; Oropharynx has to be cleared with suction. Endotracheal tube cuff needs to be de?ated. Tracheomalacia would cause tracheal collapse over the tube and prevent peri-tubal air leak. Therefore, the presenceof peri-tubal air leak after the cuff deflation rulesout Tracheomalacia (13). Trachea can also be palpated and the firm consistency of the tracheal cartilage excludes tracheomalacia. None of our patients developed tracheomalacia and all of them extubated safely at the end of surgery. No incidence of tracheomalacia was reported even in high risk patients with tracheal compression(14).However, it is safe to rule out tracheomalacia before closure.
Transient hypoparathyroidism occurred in two of our patients and one had transient hoarseness of voice. Our study included only nine cases but other studies with large number of cases also con?rmed that total thyroidectomy for large benign multinodular goitres is a safe surgical procedure(15,16,17).
Conclusion
Total thyroidectomy can be safely performed even for large multinodular goitres with minimal complications.
Acknowledgement
Authors acknowledge the immense help received from the scholars whose articles are cited and included in the references of this manuscript. The authors are also grateful to authors/ editors/ publishers of all those articles, journals and books from where the literature for this article has been reviewed and discussed.
Source of funding: No funding received. Conflict of interest: Nil.
Englishhttp://ijcrr.com/abstract.php?article_id=2417http://ijcrr.com/article_html.php?did=2417
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Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241101EnglishN2018January10HealthcareComparison of Cardiovascular Risk Between Diabetic and Non - Diabetic End Stage Renal Disease (ESRD) Patients Undergoing Haemodialysis
English3137Priya K.English Sangeeta K.EnglishBackground: This study proposes to see whether end stage renal disease (ESRD) patients undergoing haemodialysis with additional diabetes mellitus exhibit increased cardiovascular risk markers in comparison to end stage renal disease (ESRD) patients undergoing haemodialysis without diabetes mellitus.
Objectives: In this study, an attempt has been made to compare the Lipid profile and fibrinogen as inflammatory markers for assessing cardiovascular risk between diabetic and non-diabetic end stage renal disease (ESRD) patients undergoing maintenance haemodialysis.
Material and Methods: We enrolled 32 patients of both sex suffering from diabetes mellitus end stage renal disease undergoing haemodialysis and 32 non- diabetic end stage renal disease undergoing haemodialysis for this study. Results: We found that there is significant increase in the parameters like plasma fibrinogen, plasma glucose concentration, atherogenic index (TC/HDLc), serum lipid profile (TC, LDLc, TG, VLDLc) and decrease in serum HDLc in diabetic ESRD patients in comparison to non diabetic ESRD patients undergoing maintenance haemodialysis. Conclusion: We conclude that elevation in plasma fibrinogen level, serum lipid profile and atherogenic index (TC/HDLc) in diabetic ESRD patients undergoing maintenance haemodialysis which indicates higher risk of adverse cardiac events and atherosclerosis as a major cause of coronary heart disease in diabetic ESRD patients.
EnglishPlasma Fibrinogen, Haemodialysis, DiabetesINTRODUCTION
Diabetes mellitus is a group of metabolic diseases characterized by hyperglycaemia resulting from defects in insulin secretion, insulin action, or both. It is a chronic disease that causes serious health complications including renal (kidney) failure, heart disease, stroke and blindness. Diabetes mellitus is associated with a number of changes in thrombotic and fibrinolytic coagulation factor level or activity, which collectively increases the risk of thrombus formation. Hyperglycaemia and insulin resistance induce qualitative and quantitative changes in clotting factors, resulting in dense and compact clot structure and resistance to fibrinolysis [1]. Fibrinogen is the major coagulation protein in blood by mass, the precursor of fibrin and an important determinant of blood viscosity and platelet aggregation [2]. As a clotting factor, fibrinogen is an essential component of the blood coagulation system, being the precursor of fibrin. Fibrinogen has been identified as a major independent risk factor for cardiovascular disease. There are several mechanisms by which fibrinogen may increase cardiovascular risk. First, it binds specifically to activate platelets via glycoprotein iib/iiia, contributing to platelet aggregation. Second, increased fibrinogen levels promote fibrin formation. Third, it is a major contributor to plasma viscosity. Finally, it is an acute phase reactant that is increased in cardiovascular diseases [6]. High fibrinogen levels are associated with more compact clot structure, whereas elevated plasminogen activator inhibitor – I (PAI-I) levels impairs the fibrinolytic process. In addition to the quantitative changes, qualitative changes in clotting factors can affect the structure of the clots. High serum glucose has been shown to increase glycation of fibrinogen and clots formed from glycated fibrinogen have a more compact structure and increased resistance to lysis. A by-product of protein glycation that is glycoaldehyde induces post-translational modifications in fibrinogen, which impairs the fibrinolytic process, other post-translational modifications in fibrinogen such as oxidation, a known pathogenic process in diabetes mellitus, can also modify clot structure [1]. Diabetes mellitus is associated with an increase in low density lipoprotein cholesterol, and triglycerides, and low high density lipoprotein cholesterol levels. The alterations in the level and properties of LDL and HDL together contribute to the increased risk for Coronary artery disease (CAD) in diabetics [7]. Cardiovascular morbidity and mortality is markedly increased in diabetes mellitus with end stage renal disease (ESRD) patients undergoing haemodialysis (HD). As atherogenesis is mediated by inflammation of vessel walls and as evidence evolves that atherosclerosis and diabetes mellitus share a common inflammatory basis. Lipid profile and fibrinogen level has been compared as inflammatory markers for assessing cardiovascular risk between diabetic and nondiabetic end stage renal disease (ESRD) patients undergoing maintenance haemodialysis in this study. Atherogenic index (TC/HDLc) has also been determined and compared between control group and diabetes mellitus patients.
Hence, this study proposes to see whether end stage renal disease (ESRD) patients undergoing haemodialysis with additional diabetes mellitus exhibit increased cardiovascular risk markers in comparison to end stage renal (ESRD) patients undergoing haemodialysis without diabetes mellitus.
EXPERIMENTAL SECTION
Study Design and Subjects: This study was conducted in the Department of Biochemistry Teerthanker Mahaveer Medical College and Research Centre and Dialysis unit of Teerthanker Mahaveer Hospital, Moradabad, U.P, and India. The investigation was conducted on 32 patients of both sex suffering from diabetes mellitus end stage renal disease undergoing haemodialysis and 32 non-diabetic end stage renal disease undergoing haemodialysis. Inclusion criteria: Patients undergoing maintenance HD for atleast one year. Exclusion criteria: Patients with any infective diseases such as AIDS, Hepatitis B and Hepatitis C, and treatment with cholesterol lowering drugs, tissue plasminogen activator (tPA), anticoagulant therapy , pregnant women, and postmenopausal state conditions (which can alter the plasma fibrinogen and serum lipid profile level ) were excluded from the study. Sample collection: Blood samples were collected from an anticubital vein between 8 and 9 AM after an overnight fast from all subjects and dispensed into following vials for various biochemical tests:
1. Fluoride oxalate vial for fasting plasma glucose estimation.
2. Plain vial for Total cholesterol, Total triglycerides and High density lipoprotein cholesterol.
3. Vial containing 3.8% tri sodium citrate for fibrinogen estimation.
Analysis of Sample:
• Estimation of Plasma fibrinogen level by KjeldahlNesslerization method.
• Estimation of fasting blood sugar (FBS) level by GOD-POD method (Glucose oxidase and peroxidase).
• Estimation of serum Cholesterol by end point CHODPAP method.
• Estimation of serum triglycerides by end point GPOTRINDER method.
• Estimation of serum HDLc by end point TRINDER reaction.
Calculation
• VLDLc = [TG/5]
• LDLc = [Total Cholesterol] - [HDLc] - [(TG/5)] (Friedewald equation)
• Atherogenic index = [TC/HDLc]
Statistical analysis
Mean ± SD were calculated for all the parameters analyzed and were compared by Student’s t-test (2 tailed) using SPSS. P-values considered significant were as follows:- P Englishhttp://ijcrr.com/abstract.php?article_id=2418http://ijcrr.com/article_html.php?did=2418
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Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241101EnglishN2018January10HealthcareCharacterization and Invitro Cytotoxicity of T-2 Toxin Isolated from Corn
English3846P. RachithaEnglish K. KrupashreeEnglish Fouzia AmreenEnglish Farhath KhanumEnglishAim: Main objective of the study is to characterize and evaluate invitro cytotoxicity of T-2 toxin isolated from corn.
Methodology: The fungal strain was isolated by standard blotter method; morphological identification was done using scanning electron microscope (SEM). The fungal strain was identified as F. sporotrichioides based on its cultural and morphological characteristics and internal transcribed spacer (ITS/ITS4) primers. The toxin was characterized using thin layer chromatography (TLC), HPLC and Fourier Transform Infrared Spectroscopy (FTIR)
Result: 100ml of culture filtrate yielded 10.126μg of toxin. The functional groups of toxin were identified by (FTIR) method. Invitro cytotoxicity of T-2 toxin was evaluated on human keratinocytes, hepatoma and neuron cell lines. HaCaT and HepG2 cells were
found to be more sensitive than SHSY5Y to T-2 toxin exposure at 50-100ng and 100-200ng.
Discussion and conclusion: T-2 toxin was extracted and characterized from corn samples, by analytical and molecular methods in addition FTIR analysis verifies the functional groups of T-2 toxin. Further invitro cytotoxicity was evaluated using different human cell lines. Comparably skin and liver cell lines found more sensitive to T-2 toxin exposure. Sensitivity of the cell lines to the toxin at ng may be due to the purity of the toxinKey Words: Plasma Fibrinogen, Haemodialysis, Diabetes.
EnglishINTRODUCTION
T-2 mycotoxin, a low molecular weight (250-500 Daltons) non-volatile compound produced by fungi of the genera Fusarium, Myrothecium, and Stachybotrys1 . Predominantly F. sporotrichioides is responsible for the production of T-2 toxin and HT-2 toxin. The most important factors that influence T-2 toxin production are weather conditions and moisture content. T-2 toxin is produced under a wide temperature range (0 to 32 °C), with maximum production at temperatures below 15 °C 2, 3. T-2 toxin is tremendously heat stable and is the only mycotoxin known to have been used as a bio threat weapon delivered via food or water sources, as well as, by means of droplets, aerosols, or smoke from various scattering systems and exploding munitions 4, 5. Chemically, T-2 toxin is a tetra cyclic sesquiterpenoid with 12, 13- epoxytrichothec-9-ene ring system6 with hydroxyl group at the C-3 position, acetyloxy groups at C-4 and C-15 positions, atom of hydrogen at C-7 position and an esterlinked isovaleryl group at the C-8 position 7 . The toxicity can be reduced by the cleavage of esters 8, 9. T-2 toxin is a well known inhibitor of protein, DNA and RNA syntheses and also it is known to interfere with the metabolism of membrane phospholipids and increase the level of liver lipid peroxides 10, 11. The T-2 toxin is well-known to cause acute and chronic toxicity, as well as alimentary toxic aleukia (ATA) and KashinBeck Disease (KBD) in humans and animals. It has been shown to cause alteration of blood–brain barrier (BBB)12- 15. T-2 toxin toxicological effects have been summarized in Food and Agricultural Organization (FAO), World Health Organization (WHO), Council for Agricultural Science and Technology and the Expert Committee on Food Additives 16, 17. European Union states; T-2 toxin as a common contaminate to cereals and cereal based products and there are several reports from different regions of the world associating damage of agricultural field by T-2 toxin and its producing organisms and it extends to humans and animals 18,19. Sensitive and precise analytical methods have been developed for measuring mycotoxins in cereals and cereal based products in order to gauge the risk of animal /human exposure 1-3. T-2 toxin toxicity is the most severe compare to other trichothecenes. Major target organs include skin, liver, nerves etc. In the present study, the fungal species growing on corn was isolated and identified as T-2 Toxin producing organism by molecular characterization and T-2 Toxin identified by FTIR analysis and toxicity of the toxin produced was evaluated in human keratinocytes, hepatoma, and neuron cells in vitro.
MATERIALS AND METHOD
Potato dextrose agar, Potato dextrose broth, glutaraldehyde(Himedia Bangalore), methanol, ethyl acetate, ethanol, silica gel, (SRL, Bangalore), T-2 toxin (Chempure Mumbai), ethidium bromide (Sigma Banglore) were used.
Isolation of Fusarium species from corn sample
Isolation of Fusariumwas carried out by standard blotter method. Corn samples were collected from different localities. Corn seeds were incubated for 3-5 days under moist condition on petriplates. As the spore of fungi developed into mycelia, it was transferred aseptically to sub-culture on potato dextrose agar to develop pure colonies, and the colony morphology was examined under microscope.
Scanning electron microscopy
After observation under microscope the sample subjected to scaning electron microscopy (SEM). The mycelia were fixed in glutaraldehyde in 0.1M PBS for 30 min., washed with 0.1M PBS and dehydrated by immersing in ice cold ethanol for 10min. The dehydrated samples were smeared on silver stub like a thin film, and were coated by cathodic spraying (Polaron gold). The SEM observations were made using a ZEISS Instrument (EHT=15.00 kv, signal A=VPSE G3).
Fusarium species DNA Extraction
7 days old mycelium mat was collected and frozen at -800 C and lyophilized. Lyophilized sample were homogenizedinlysis buffer and extraction was carried by phenol chloroform method. DNA was quantified in nanodrop spectrophotometer, PCR was performed to amplify DNA 20.
Polymerase Chain Reaction
The PCR of fusarium was performed using universal primers (ITS/ITS4 primers). Standard protocol was followed to amplify the DNA, after successful amplication products were inspected by agarosegel electrophoresis containing (0.01%) ethidium bromide and observed under UV(Ultra violet) light. Gels were documented using Gbox (GE Health Care, Mumbai).
Fusarium culture for toxin production
Culture preparation F. sporotrichioides MTCC 1894 Venkataramana et al. 21was used as the test organism and was obtained from Department of Microbiology, Defence Food Research Laboratory, Mysuru. The fungus was purified and harvested at room temperature on potato dextrose broth (PDB).
Toxin extraction and purification
The F. sporotrichioides was inoculated to PDB broth and incubated at for 3 weeks incubation method was followed according to Busman et al., 22 with slight modification. After incubation the broth was filtered and the filtrate was collected and mixed with equal volume of ethanol and incubated for 24 hrs, then it was concentrated using flash evaporator. The extracted toxin was stored in amber vial for further characterization.
Thin Layer Chromatography (TLC)
T-2 toxin was extracted and detected by thin layer chromatographic method23.The plates were prepared by adding 50 ml of distilled water to 25g silica gel powder and mixed uniformly to get homogenous mixture (slurry) and then drawn on glass plates of 20 x 20cm using TLC applicator with thickness of 2mm and allowed to dry. The coated plates were activated by drying in forced draft hot air oven at 1100 C for one to two hours. The activated plates were then cooled to room temperature. Extracted toxin (crude) was spotted on TLC plate with capillary tube and allowed to run using toluene: ethyl acetate: formic acid (6:3:1) as mobile phase. After that the plate were dried, viewed under UV light (366nm) and calculated Rf value.
High performance liquid chromatography (HPLC)
Reverse phase HPLC with C-18 column was used. The extracted toxin dissolved in methanol was passed through membrane filters, Methanol: water (70:30) mixture was used as mobile phase. 20 µl of the toxin was injected to HPLC column and made to run through the column for 20 min. The flow rate of the column was adjusted to 0.7 ml and wavelength to 228nm. Toxin was detected in UV detector system. Standard toxin was also injected to determine the appropriate peak and concentration of toxin extracted23.
Fourier transformed infrared spectroscopy
The functional groups of extracted toxin were anlyzed using FTIR ( Nicolet 380 Thermo) based on the peak values in the region of IR radiation. The toxin was passed into the FTIR, the functional groups of the compounds were separated based on their peaks ratio. The FTIR spectra were recorded in the absorption range between 4000 and 500cm‾1.
Cell Culture Studies
The Human skin cell line HaCaT, human liver cancer cell line HepG2 and human neuroblastoma cell line SHSY5Y was procured from National Center for Cell Sciences, Pune, India. The SHSY5Y and HaCaT cell lines were cultured in DMEM/F-12 mixture and HepG2 cell line were cultured in MEM/F-12 mixture supplemented with 10% FBS, antimycotic solution (Sigma, St.Louis, MO, USA). Cells were maintained and incubated at 37°C in a humidified atmosphere and 5% CO2 atmosphere 95% air at 370 C. The three cell lines were treated with different concentration of T-2 toxin (ng) for 24 hr.
Cell viability assay
To asses the changes in the metabolic activity of the cell MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] was used. The Tetrazolium dye used to measure the cell proliferation and cell viability. The HaCaT, HepG2 and SHSY5Y cell were seeded in different 96 well plates and treated with T-2 toxin at different concentration. After treatment incubated with MTT (0.5 mg/ml) at 370 C for 4hrs and the formed formazan crystals dissolved in DMSO. The absorbance measured at 540nm (VERSA max Hidex plate chameleon TM V (Finland)). The cell viability expressed as percent of control.
Lactate dehydrogenase (LDH) leakage assay
Lactate dehydrogenase (LDH), leakage a biomarker of cellular cytotoxicity and cytolysis. Cell injury was assessed by measuring the LDH efflux to the media after cells has been treated for 24 hr. The protocol following the manufacturers’ instructions (LDH-estimation kit (Agappe-11407002, Mysore, India)). The HaCaT, HepG2 and SHSY5Y cell were seeded in different 24 well plates and cell count set as 1X104 cell/well. The cells were subjected to different concentration of T-2 Toxin. After the treatment the cells were centrifugedat 2500Xg for 5min at 40 C and supernatant was collected and measured the LDH activity estimated.
Propidium iodide staining
The HaCaT, HepG2 and SHSY5Y cells were seeded at 1X104 cell/well into a six well plate. At 90% confluence, the cells were treated with T-2 toxin and incubated for 24 hrs. After incubation the cells were washed with PBS and fixed with 100% ethanol. The fixed cells stained with 100ng/ml propidium iodide for 20 min and observed under fluorescent microscope.
Statistical analysis
The data were represented as the mean ± SD. Experiments were analyzed using one-way ANOVA followed by Tukey post hoc test. Statistical significance is indicated *p ≤ 0.05, **p ≤ 0.01, and ***p ≤ 0.001.
RESULTS
Morphological observation of isolated sample
Morphological identifications of the fungal isolates were made using the criteria of 24-26. The isolated sample on PDA formed cottony colony brownish, thread like filaments the hyphae and each branching contains aerial mycelia, clusters of hyphae, called sporodochia. These species have irregular shaped globular microconidia, that are measured 5-7 μm in diameter (Figure 1).
Microscopic observation of isolated hyphae
The microscopic observation was done in inverted microscope and scanning electron microscope (40 x magnification).The isolated hyphae were in tubular shape and conidia was identified with branches. The mycelia were measured in inverted microscope found to be 3.5µm and in scanning electron microscopy measured around 1-5 µm (Figure 2).
Molecular Identification
The morphological identifications were further confirmed by molecular method (PCR) (Figure 3). A single band of 500bp was obtained from the isolate and that was compared with standard culture of Fusarium sporotrichioides and amplified with ITS/ITS4 primers.
Thin Layer Chromatography (TLC)
The samples showed sky blue colour bands under UV transilluminator at 365nm. Rf valueof T-2 toxin on silica gel found to be 0.48mm, comparable to standard T-2 toxin (Figure 4).
High performance liquid chromatography (HPLC)
High performance liquid chromatography was used to quantify the toxin in the extract with UV detector. The amount of toxin was found to be 10.126 µg from 100ml broth culture (Figure 5).
Fourier transformed infrared spectroscopy
FTIR is qualitative and quantitative analytical technique for identification of functional groups of organic, inorganic samples and chemical compounds. The extracted sample revealed the presence of alkanes, alkenes, esters, alcohols and carboxylic acid, compared with standard (Figure 6 and Table 1 and 2).
Cell viability assay Cell viability was determined by Tetrazolium dye for T-2 toxin with increasing concentration on three different cell lines: HaCaT, HepG2 and SHSY5Y. The results showed that, after 24 hr exposure T-2 toxin decreased cell viability in a dose dependent manner. 50-75ng T-2 toxin was sufficient to reduce viability of HaCaT cells is 50-70% for HepG2 cells 100-150ng of toxin was required and for SHSY5Y 200-250ng was required (Figure 7). Compare to HaCaT and HepG2 cells SHSY5Y showed increased cell viability. The HaCaT cell line more sensitive to T-2 toxin. Lactate dehydrogenase (LDH) leakage assay The cytotoxicity of T-2 toxin further carried out by assuring LDH activity in the culture media. The LDH a cytosolic enzyme leakes into the surrounding medium if the plasma membrane gets damaged and is used to asses membrane damage with T-2 Toxin challenge of HaCaT, HepG2 and SHSY5Y cells in dose dependent manner. As cell viability showed that HaCaT is more sensitive to T-2 Toxin. The highest LDH release was found in HaCat cells 50-75ng, followed by HepG2 (100-150ng) and SHSY5Y (200-250). As shown in Figure 7(C), The shape of T-2 Toxin treated cells become irregular and destruction of monolayer was observed and compared with control cells. Propidium iodide staining To confirm the cytotoxic effects of T-2 toxin on cell lines, nuclear condensation was studied by Propidium iodide staining. The Propidium iodide does not pass through viable cell membrane however reach the nucleus and it can pass through the damaged membranes. In Control cells had minimal propidium iodide stained cells, while treated groups had increased number of propidium iodide stained cells (Figure 8).
DISCUSSION
T-2 toxin is a type A trichothecene mycotoxin produced by different fusarium species, including F. sporotrichioides, F. poae and F. acuminatum27-29. These species grow on a variety of cereals, grains, particularly in oats, barley, rye,wheat, maize, rice, beans, soybean and in some cereal-based products especially in cold climate region or wet storage condition 30. Several surveys have revealed F. sporotrichioides causes root, stem and ear rot, with severe reductions in crop yield. In addition F. sporotrichioidesis capable of producing T-2 toxin which can infect pre harvest plants or stored grains31,32. There are an array of studies on fusarium isolation, its characterization and inhibition of toxin producing species. The present study focused on molecular and analytical characterization of T-2 toxin and T-2 toxin producing organism. FTIR characterization was unaccustomed method for exploration of functional groups in microbiology. Fusarium mycotoxin can be produced on every part of the plant, hence the need for accurate identification at the species level. Fusarium isolation was carried out by standard blotter method. However identification was based on morphological features and molecular method. In the bright field microscope, hyphae and conidia measured 5-7 μm in diameter. Identification of fusarium and other species followed through the SEM for morphological investigation, microconidia were with a smooth cell wall and maintained membrane structure, diameters measured around 1-5μm33,34. Many studies have been discovered ITS/ITS4 primers established recognition of fusarium35-38. Taking into consideration all these available datawe aimed to confirm the rDNA region by providing ITS/ITS4 target sequence for molecular detection of F. sporotrichioides. PCR method with above primers successfully tested against isolates which exhibited greatest homology in the target. T-2 toxin a metabolite of F. sporotrichioides was extracted in ethanol and characterized using TLC, HPLC and FTIR.T-2 toxin visualized as bluish spot on the TLC plates and compared with standard toxin Rf value found to be 0.48 mm. In our study quantification of T-2 toxin was done as per the method of 19with slight modification using UV detector investigated 10.126 µg toxin.
According to previous reports T-2 toxin is more toxic than other mycotoxin. In the present study, we tested T-2 toxin cytotoxicity by MTT and LDH leakage assays. The data showed human skin cell line and liver cell line are the more sensitive of all the cell lines tested, whereas neuronal cell is the least cytotoxic to T-2 toxin. Albarenque et al. 39showed T-2 toxin decreased the cell viability at 25 μg in rat keratinocyte primary cultures. In the present study HaCaT cell viability decreased at 50-75ng. According to Bouaziz et al.40 T-2 toxin induced maximum cell death in HepG2 compare to zearalenone and ochratoxin A at 60nM with increasing concentration, this report proves that cell viability of Hepg2 with T-2 toxin causes toxicity (100-150ng) comparably same concentration. Therefore, these results suggest T-2 toxin are potent inducer of hepatotoxicity. Weidner et al 41 worked on T-2 toxin induced neurotoxicity with human astrocytes primary culture and shown that T-2 toxin disturb the blood-brain barrier (BBB) in vitro. They reported cytotoxicity at 10 nM and disturbance of barrier at 75nM. In our study using SHSY5Y cell line to the toxicity was noticeable at 200 -250mg.In the present study we could grow the T-2 toxin producing fusarium species on corn, isolate and characterize the species upto molecular level. T-2 Toxin was extracted and characterized. And toxicity of the toxin at cellular level was carried out using three different cell lines, A liver cell line (HepG2), a skin cell line (HaCaT) and neuroblastoma (SHSY5Y) cell line. The results obtained are qualitatively similar to the once reported by others. Sensitivity of the cell lines to the toxin at ng may be due to the purity of the toxin.
CONCLUSION
F. sporotrichioides was isolated from corn sample and T-2 Toxin was extracted, characterization of species done by molecular method and T-2 toxin was quantified using analytical techniques. Further different dosage of T-2 toxin in different cell lines evaluated cytotoxicity
ACKNOWLEDGMENTS
The authors are grateful to Ex Director Dr. HV Batra and Present Director R.K Sharma, Defence Food Research Laboratory, Mysuru, for providing all the necessary facilities, constant guidance and encouragement during this investigation. Authors acknowledge the immense help received from the scholars whose articles are cited and included in references of this manuscript. The authors are also grateful to authors/ editors/ publishers of all those articles, journal and books from where this article has been reviewed and discussed.
Conflict of interest: None
???????Source of funding: None
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Radiance Research AcademyInternational Journal of Current Research and Review2231-21960975-5241101EnglishN2018January10HealthcareA Descriptive Study of Amblyopia in Children: A Hospital Based Study
English0709Imtiyaz A. LoneEnglish Natasha KoulEnglish Reyaz A. UntooEnglishIntroduction: Amblyopia is the most common cause of visual impairment in children. The prevalence of amblyopia in children has been estimated at between 1% and 4%. Most cases are associated with strabismus, anisometropia or a combination of strabismus and anisometropia.
Aims and objectives: To assess the profile of patients with visual disability due to amblyopia in Kashmir and to suggest measures for restricting visual disability due to Amblyopia.
Methods: This prospective study was conducted in the Postgraduate Department of Ophthalmology Sher-i-Kashmir institute of Medical Sciences Medical College , Srinagar during a period of 12 months from 1st March 2015 to 29th February 2016. All the patients between 30 months to 19 years of age with amblyopia who attended the Ophthalmology OPD were included in the study. Visual acuity measurements were done using Snellens chart for older patients and for 2-4 year old children by using Cardiff acuity cards, Kay picture test or Teller test.
Results: The mean age of the 103 children was 12.25 years (±2.89) ranging from 4-19 years with 58 males and 45 females. 64 children were amblyopic in right eye, 37 in left eye and 2 had bilateral amblyopia .The cause of the amblyopia was strabismus in 32 children (31%), anisometropia in 60 (58%), and both strabismus and anisometropia in 5 (5%). 5 children had amblyopia due to congenital cataract with 1 of them having bilateral amblyopia. One child had amblyopia due to severe congenital ptosis.
Conclusions: There were significantly greater number of anisometropic amblyopia patients than strabismic amblyopia (pEnglishAmblyopia, Anisometropia, strabismusINTRODUCTION
The term ‘amblyopia’ comes from the Greek word ‘amblupos’ which means ‘dim-sighted’ (1). Amblyopia occurs when there is suboptimal vision in one or both eyes despite best-corrected spectacle correction, and when there are no other anatomical ocular or cerebral visual pathway abnormalities to explain this visual impairment (2). It occurs as a result of disrupted or incomplete visual development during early childhood (3, 4).
Normal visual development commences at childbirth when the child opens his/her eyes for the first time. It improves very rapidly in the first 6 months of life and then more gradually, reaching adult levels when the child is aged 4-6 years .This is accompanied by differential development of the retina, foveal region, increased synaptic density within the primary visual cortex, and pruning of extraneous neuronal receptive fields; all of which result in improved spatial resolution and contrast sensitivity (i.e. vision). This process is a competitive one, with neurons from each eye competing for space within the cortex (5,6).
Well-known ocular risk factors to amblyopia include high hypermetropia (>4.00D), high myopia (>8.00D), astigmatism (>2.00D), lid ptosis, childhood cataract and strabismus (2, 7).
Amblyopia can be classified as either being unilateral or bilateral. Unilateral amblyopia occurs when the visual image in one eye is compromised or blurred so that, that eye is selectively disadvantaged. In contrast, bilateral amblyopia can occur when there are similar levels of obstruction/blur in both eyes. This is often the result of high uncorrected refractive error (e.g. hypermetropia, myopia or astigmatism) or equal obstruction in both eyes. Amblyopia can also be classified according to etiology (i.e., refractive, strabismic or deprivational) (2). In general, refractive amblyopia is more common than strabismic amblyopia. Clinically, amblyopia is defined by two or more lines difference in visual acuity between the eyes, (8) however other monocular visual functions are also affected, including grating acuity, vernier acuity and contrast sensitivity. Another classification based on visual acuity in amblyopic eye is:
Mild : V.A > 6/ 24
Moderate : V.A 6/24 -6/60
Severe : V.A < 6/6
AIMS AND OBJECTIVES
The present study was done to assess the profile of patients with visual disability due to amblyopia in Kashmir and to suggest measures for restricting visual disability due to amblyopia.
MATERIALS AND METHODS
The present study was conducted in the post-graduate department of Ophthalmology Sher-i-Kashmir institute of Medical Sciences Medical College Hospital, Srinagar. Children between 30 months and 19 years of age with two or more lines difference in visual acuity of two eyes or ≤ 6/12 best corrected visual acuity (BCVA) in bilateral cases and having anisometropia, strabismus, congenital cataract or ptosis were included in the study. Patients having some other disease in the eye responsible for low vision like any macular disorders, pathological myopia, history of trauma and those not falling within the age group taken for the study were excluded from the study.
The patients were registered on a pre-structured proforma which included patients demographic details, visual acuity, pin hole vision testing, refraction, retinoscopy, anterior segment examination and fundus examination.
Testing and examination protocol included:
Visual acuity measurement using Snellens chart for older patients and for 2-4 year old children by using Cardiff acuity cards, or Teller test.
Ocular motility.
Squint evaluation.
Refraction with and without cycloplegia.
Detailed anterior segment examination with slit-lamp microscope.
Fundus examination (with direct and indirect ophthalmoscope).
RESULTS
A total of 32528 patients attended Ophthalmology OPD during the study period out of whom 8245 were in the 2-19 year age group. 103 patients were registered for the purpose of the study based on inclusion and exclusion criteria. This roughly puts the prevalence of amblyopia in this age group at 1.25%.
The mean age of the children was 12.25 years (±2.89) ranging from 4-19 years. There were 58 males and 45 females. 64 children (62.14%) were amblyopic in the right eye, 37(35.92%) in the left eye and 2 (1.94%) had bilateral amblyopia. Thus there were a total of 105 amblyopic eyes
The cause of the amblyopia was strabismus in 32 children (31%), anisometropia in 60 (58%), and both strabismus and anisometropia in 5 (5%). 5 children had amblyopia due to congenital cataract. One child had amblyopia due to severe congenital ptosis.
Among those with strabismic amblyopia (pure or combined anisometropic and strabismic), 32 patients (86.5%) had esotropia and 5 (13.5%) had exotropia.
65 patients had anisometropic amblyopia (pure or combined). Out of these 43 patients (66.2%) had hypermetropic anisometropia, 17 (26.1%) had myopic anisometropia and 5(7.7%) had astigmatic anisometropia.
Among 43 patients with hypermetropic anisometropia, the degree of anisometropia ranged from +1D to +6.5 D. Of the total of 17 patients with myopic anisometropia, the degree of anisometropia ranged from -2 D to -4.5 D. 5 patients had astigmatic anisometropia which ranged from -1 D to -2.5 D.
DISCUSSION
Amblyopia is one of the most common causes of visual impairment in both children and adults with a prevalence varying between 0.2% and 12% depending on the subsets of the population studied (9,10) . Lack of adequate understanding or knowledge about this preventable and easily treatable condition, provided compliant treatment is started early, is often the reason why very few patients are referred to eye hospitals or specialists for the amelioration of the same especially in a developing country like India.(11)
Amblyopia and associated strabismus can have devastating psychosocial and economic fall-outs. Failure to develop binocular vision and unilateral or bilateral visual impairment may prevent the individual from pursuing certain occupations. Severe amblyopia is also considered a significant risk factor for blindness in case an individual loses sight in the fellow eye.
Though the present study suffers from a selection bias, as the data is hospital based, this analysis may form the basis of future population-based studies. However, one advantage of a study in a hospital referral practice with a very wide base of patients coming from all parts of the state, is the immense variety and numbers that can be seen, which can be utilized to elucidate the clinical profile of that condition under as ideal a circumstance as possible.
Another important factor that this study reveals is the relatively older ages of presentation to a specialty clinic irrespective of the sub-type of amblyopia present. Seven to eight years is the critical time after which therapeutic measures for the treatment of amblyopia become less effective and unfortunately the average age of presentation of most patients to the hospital in our study was more than that (12.25 years). This reiterates that efforts to screen patients for amblyopia and educate personnel at every level to suspect, diagnose, treat or refer the patient as soon as possible. Prevalence of amblyopia in our study was found to be 1.25% in children aged between 30 months - 19 years. Amblyopia was more prevalent in males than females, though the difference was not statistically significant (p=0.20). The most common cause of the amblyopia in our study was anisometropia (58%) followed by strabismus (31%), and both strabismus and anisometropia (5%). It is precisely the most common form of amblyopia that is often overlooked, as unlike strabismus pure anisometropia is not usually noticeable to those around the child concerned.
CONCLUSION:
Amblyopia is one of the causes of visual impairment in children with anisometropia and strabismus being the main reason. However the corrective measures if taken at the earliest prevent the devastating psychosocial and economic fall-outs.
Source of Funding : Sher-i-Kashmir institute of Medical Sciences Medical College Srinagar being a Government run Hospital , all the funding during the study period were provided by the State only after getting an ethical clearance from the Hospital ethical committee .
Conflicts of Interest : Nil
Englishhttp://ijcrr.com/abstract.php?article_id=2548http://ijcrr.com/article_html.php?did=25481 . http://www.merriam-webster.com/dictionary/amblyopia, 4 Oct 2012.
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