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2011 - 20175
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Subject: Veterinary Physiology × End date: 2019 × Start date: 2011 ×
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Now showing 1 - 5 of 5
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    ThesisItemOpen Access
    LEPTIN RECEPTOR EXPRESSION IN CULTURED OVARIAN FOLLICLES OF SHEEP
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2017-05) SRAVANI PRAGNA, K; Siva Kumar, A.V.N(MAJOR); Rambabu Naik, B; Punya Kumari, B
    ABSTRACT : The expression of Leptin receptor (LepR) mRNA and protein was studied in sheep using Real time quantitative PCR and Immunohistochemistry in the cumulus cells and oocytes from : (i) In vivo grown preantral, early antral, antral, large antral follicles and Cumulus Oocyte Complexes (COCs) obtained from large antral follicles and subjected to 24h of In Vitro Maturation ( IVM) and (ii) Preantral follicles (PFs’) exposed to TCM199B, TCM199BL (10ng/mL) and TCM199BHGL (10ng/mL) media for 3min, cultured in vitro for two, four or six days and subsequently matured in vitro for 24h in respective cultures separately. LepR was observed at all stages of in vivo grown and in vitro cultured ovarian follicles in both cumulus cells and oocytes. Expression levels and patterns of LepR mRNA in PFs’ cultured in TCM199BHGL was similar to in vivo in all the stages except in cumulus cells from COCs after in vitro maturation for 24h, where the expression was significantly higher which was a positive effect. The expression of LepR in cumulus cells among the three different in vitro culturing conditions was significantly higher in PFs’ cultured in TCM199BHGL medium. In the oocytes, the Leptin receptor expression was similar or significantly higher in PFs’ cultured in TCM199BHGL medium than in vivo grown follicles except in PFs’ exposed to 3min stage which could suggest the synergistic action of growth factors and hormones with Leptin. Immunohistochemistry studies revealed highest intensity of Leptin receptor protein expression in the oocytes. Mild to moderate staining was observed in cumulus cells with good intensity in large antral stages. This protein expression coincided with gene expression. From our studies it is concluded that Leptin along with other growth factors and hormones supplementation stimulated the expression of Leptin receptor mRNA and protein on par with that of in vivo /even higher than that of in vivo condition.
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    ThesisItemOpen Access
    INFLUENCE OF LEPTIN ON THE EXPRESSION OF p53 and BAK GENES IN CULTURED OVARIAN FOLLICLES IN SHEEP
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2015-11) PRABHAVATHI, T; RAO, V.H(MAJOR); SIVA KUMAR, A.V.N; VEERABRAHMAIAH, K
    ABSTRACT : The present study was conducted to know the influence of Leptin on the expression of p53 and BAK (apoptotic) genes in the sheep preantral follicles (PFs’) cultured in vitro. Ovaries of sheep were collected from the local slaughter house. Intact PFs’ (250-400μm) were isolated and placed individually in 20μl droplets of standard medium (TCM 199 B + 2 μg/ml FSH, 0.1μg/ml T4 + 10 ng/ml IGF-I + 1 mIU/ml of GH) supplemented with Leptin (10ng/ml) for 2, 4 or 6 days. Quantitative expression of Test genes (p53 and BAK) and Reference genes (18s RNA, RPLPO, HPRT1) were studied in the cumulus cells and oocytes isolated at different developmental stages (preantral, early antral, antral and large antral follicles) of the in vivo and in vitro grown follicles. The entire experiment was repeated twice. Duplicate samples of cDNA from each replicate of the experiment were subjected to relative-RT-qPCR. The pattern of expression of p53 gene in the cumulus cells from ovarian follicles grown in vivo and cultured in Leptin was similar from preantral to large antral follicle stage. In the oocytes, however, such similarity was restricted up to the antral follicle stage only. It is concluded that Leptin in culture medium mimiced the pattern of p53 expression in in vitro as in in vivo to some extent. The BAK gene expression was undetected in the cumulus cells and oocytes isolated from all the stages of both in vivo and in vitro developed ovarian follicles. Accordingly it is concluded that (i) Leptin has no influence on the expression of BAK gene in cultured ovarian follicles in sheep (ii) BAK may not be an important regulator of apoptosis in ovarian follicles and (iii) p53 plays a relatively more significant role than BAK in the regulation of apoptosis in ovarian follicles.
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    ThesisItemOpen Access
    AMELIORATION OF HEAT STRESS INDUCED OXIDATIVE DISTURBANCES BY HERBAL ANTIOXIDANTS IN BROILER CHICKEN
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2011-04) SWATHI, B; Gupta, P.S.P(MAJOR); Nagalaxmi, D; Rajasekhar Reddy, A; RaJu, M.V.L.N
    ABSTRACT: A study was petformed to evaluate the antioxidative potential of herbals Tulsi (Ocimum sanctum) and Turmeric (Curcuma longa) in combating heat stress with two batches of broiler chicks in hot summer months. Each batch consisted of a total of 108 one day OM Vencobb broiler chicks which were grouped and offered 9 types of dietary regimens wtth variable concentrations and combinations of antioxidants. A group of 12 chicks was raised separately in stress free environment on basal diet without any antioxidant supplementation as control. Body weights and feed intake were measured at weekly intentab. The blood samples were colleded at 4" and 6" wk of age and haernatological estimations (RBC, WBC, Hb. & PCV) were carried out within 2 hrs of collection. While serum samples were preserved at -20°C until utilized for estimation of protein profiles (TP, albumen, globulin concentrations and A:G ratios), serum cortisol concentrations, humoral and cell mediated immune responses. Plasma samples were prepared by centrifugation, stored and utilized for analysis of enzymatic and non enzymatic antioxidant concentration. At the end of trail, breast and meat samples were procured by excision of sacrificed birds for studying organoleptic properties and lipid peroxidation. Histopathological studies of liver, bursa, brain and spleen were conducted to observe the tissue changes at the end of the experiment. The performance parameters (Body weights and feed efficiency), haernatologoical parameters (RBC, Hb. PCV and WBC), enzymatic (GSH-PX, Catalase and SOD), non enzymatic (Reducedglutathione) antioxidant concentrations, immunological indices (HI titers and CMI responses) were signifiintly low (PsO.01) in heat stressed birds compared to control and antioxidant supplemented groups. However, WBC count, serum cortisol concentrations, serum ALP and ALT activities were significantly (PS0.01) higher in heat stressed birds compared to control group. Histopathological study evidenced degenerative changes and pathological lesions in tissues collected from heat 8tmssed birds, While no lesions of pathological importance were observed with the tissues d control group. Sensory chamderistics of breast meat samples from heat stressed birds scored low on 5 point hedonic scale (3.42 for colour, 3.12 for texture, 3.42 for juiciness, 3.22 for tlavour and 3.14 for overall acceptability) compared to higher scores recorded for meat samples of control group. Dietary supplementation with herbals Tulsi and Turmeric was proved beneficial in terms of broiler performance, haematological, serological values. immunological indims and antioxidant status. Their inclusion also contributed to low levels of serum cortisol and improved sensory characteristics and reduced lipoperoxidative damage of broiler meat. Of the two herbals employed for the study. Turmeric had contributed to hgher body weights (1629.17 g and 1627.50 g with 0.2% and 0.4% level of inclusions, respectively), which were comparable with the weights achieved by supplementing the diet either with vitamin E alone (1637.52 g) and its combination with Selenium (1656.67 g) end control group (1684.17 g) at the end of 6'" week, Feed efficiency though low in the initial stages, an accelerated trend was noticed from 4m wk onwards wrth Turmeric supplementation. And even sensory attributes showed higher sensory scores with Turmeric supplementation at either of the dose. While, Tulsi, contributed to significantly (Ps0.01) higher concentration of enzymatic and non enzymatic antioxidants, blood and serum variables, and immunological indices at either of the dose supplemented. Herbals at higher concentration (Ocimum at 0.5% and Turmeric at 0.4%) had shown improved antioxidant -8. However, the two herbals when given in combination at different doses could not yield any additive effect. The benefdal results obtained with the supplementation of herbals at different doses were however remained low when compared to the values obtained with the diets inclusive of vitamin E alone, or its combination with Se and control group. But, considering the cost of these synthetic antioxidants vitamin E and Se and maintaining the birds in stress free environment, the herbals employed Tulsi and Turmeric in this study may be suggested as natural and economical alternatives in amelioration of heat stress and achieving higher performance
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    ThesisItemOpen Access
    IN VITRO PRODUCTION OF MEIOTICALLY COMPETENT OOCYTES FROM GOAT PREANTRAL FOLLICLES
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2013-09) SHANKARAIAH, PAKALA; SWATHI, B(MAJOR); SRINIVASA PRASAD, Ch; ARUNA KUMARI, G
    ABSTRACT : A series of experiments were carried out with different combinations of various growth factors and hormones like Growth Hormone (GH), Insulin like Growth Factor-I (IGF-I), Epidermal Growth Factor (EGF), Thyroxine (T4) and Follicle Stimulating Hormone (FSH) in culture medium to support growth and development of goat preantral follicles in vitro for a period of eight days and then the oocytes retrieved from cultured preantral follicles were kept for in vitro maturation in four different protocols of IVM media. The effect of Growth Hormone (GH), Insulin like Growth Factor-I (IGF-I), Epidermal Growth Factor (EGF), Thyroxine (T4) and Follicle Stimulating Hormone (FSH) in different combinations was investigated on in vitro development of goat preantral follicles (PFs). Tissue culture medium 199 (TCM 199M) supplemented with Sodium Pyruvate, L-Glutamine, Ascorbic acid, Cysteine and Gentamicin sulfate was used as control medium. Goat PFs (diameter ranging from 150-400μ) were cultured for a period of eight days in different combinations of above mentioned hormones and growth factors. The development of the cultured PFs was assessed by the proportions of follicles exhibiting growth, increase in follicular diameter, antrum formation and extrusion of oocytes in vitro. After eight days of culture period, oocytes were retrieved from different treatments of preantral follicles (PFs) and kept for in vitro maturation in incubator at 38.5oC, 5% CO2 in different IVM media. After 27 hours of in vitro maturation period, oocytes were denuded and evaluated for nuclear status by staining with propidium iodide. In Experiment–1: Based on the previous studies in this laboratory the best concentrations were taken for different growth factors and hormones and evaluated the effect in different combinations of those growth factors and hormones on in vitro development of preantral follicles (PFs). PFs were cultured in various combinations in total of five treatments (T1- T5) (including control) at a concentration of GH (2 mIU/ml), IGF-I (20 ng/ml), EGF (50 ng/ml), Thyroxine (1μg/ml) and FSH (2 ng/ml) supplemented to control media. The proportion of PFs exhibiting growth was highest (98.00±0.87) when cultured in T4+FSH+GH+EGF (T3) supplemented medium but this was not statistically significant to those cultured in other treatments. However, the proportion of PFs exhibiting antrum formation (89.33±1.86) and extrusion of oocyte (15.55±2.09) were also highest when the medium was supplemented with T4+FSH+GH+EGF (T3) with a significant difference with follicles cultured in other treatments. The proportion of PFs showing increase in diameter (42.66±2.06 μ) was highest in T4+FSH+GH+EGF (T3) supplemented medium and this was significantly different to those cultured in treatment groupsT1 (T4+FSH), T2 (T4+FSH+EGF) and T5 (control). However, there was no significant difference with T4+FSH+EGF+IGF (T4) Experiment–2: Based on the results from experiment -1, experiment -II was designed to investigate the maturation status of the in vitro mature oocytes in different IVM media. After culture period of 8 days, oocytes were retrieved from cultured preantral follicles (T1-T5), were kept for in vitro maturation for a period of 27hrs in different IVM media (IVM-I, IVM-II, IVM-III and IVM-IV) and antral oocytes were kept for IVM as control-II in different IVM media. Among in vitro maturation protocols of our study the proportion of oocytes exhibiting M-II stage were maximum in IVM-III. And among treatments T3 (T4+FSH+GH+EGF) had a highest percentage of M II stage in all IVM media and these results were statistically different (P ≤ 0.05) with oocytes of other treatments.
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    ThesisItemOpen Access
    INFLUENCE OF LEPTIN ON IN VITRO DEVELOPMENT OF PREANTRAL FOLLICLES OF SHEEP
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2014-11) Siva Kumar, A.V.N.; Rao, V.H(MAJOR); Punya Kumari, B
    ABSTRACT : The role of human and ovine leptin on in vitro development of preantral follicles (PFs’) isolated from sheep ovaries was studied. Experiment I, was conducted to standardize the optimum concentration of human leptin for the growth of PFs’. The parameters set for the evaluation of the growth of PFs’ were increase in diameter, proportion of PFs’ exhibiting growth, antrum formation and maturation of oocytes to M II stage. All these parameters were higher in the leptin (10ng/ml) supplemented medium. Experiment II was conducted with the optimum level of leptin (10ng/ml) derived in experiment I to evaluate the effect of human or ovine leptin @10ng/ml in combination with various growth factors viz., TCM 199 B with FSH, T4, IGF-I and GH. Two controls viz., TCM 199 B and standard medium i.e., TCM 199 B supplemented with FSH (2ug/ml), T4 (1μg/ml), IGF-I (10ng/ml) and GH (1mIU/ml) were used. In the standard medium supplemented with human leptin, the proportion of PFs’ exhibiting growth (66.14±3.9), average increase in the diameter (11.1±0.87 μ), antrum formation (51.5±5.33) and maturation of oocytes to M II stage (24.1±3.7) were higher compared to the standard medium and standard medium supplemented with ovine leptin (10ng/ml). But there was no significant difference between the standard medium and the standard medium supplemented with ovine/human leptin. It is concluded that (i) the optimum dose of leptin for the growth of sheep PFs’ in vitro was 10ng/ml and (ii) human or ovine leptin supported similar growth of PFs’ cultured in the standard medium.