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ThesisItem Open Access APPLICATION AND EVALUATION OF MESENCHYMAL STEM CELLS ON WOUND HEALING IN RAT(West Bengal University of Animal & Fishery Sciences, Kolkata, 2023) MAHESH A, KUMARA DATTHA; Pradhan, S.; Mondal, S.; Das, K.; Bag, Sadhan; Dey, SamirThe primary goal of this study is to assess the wound healing effect of mesenchymal stem cells and condition media in xenogeneic transplantation on rat skin full thickness excisional wounds. MSCs derived from bovine umbilical cord were isolated and cultured using explant and enzymatic methods. The MSC isolation technique was standardised using enzymatic digestion, as was the explant method. Because of their migratory properties, MSCs emerged from explants by day 4 and reached 70-90% confluence by day 10. MSCs were digested with collagenase (1mg/ml) for 4 hours at 370C before being plated and cultured, reaching 90% confluence after 72 hours. MSCs exhibited strong plastic adherence and fibroblast-like morphology. MSCs were cultured in vitro and differentiated into osteocytes, chondrocytes, and adipocytes according to International Society for Cell Therapy and Mesenchymal Stem Cell Committee guidelines. Xenogeneic bovine UC-MSC and bovine UC-MSC derived condition media were transplanted into two treatment groups and compared to positive, negative (vehicle), and wound healing ointment groups. A full thickness (10mm) skin punch biopsy was performed on both sides of the shoulder region, and MSC and CM were injected at the wound's periphery. Wound healing was evaluated using wound contraction (up to day 14), gross morphological evaluation (up to day 7), cytological examination of the wound bed area (up to day 7), and histo-morphological evaluation of skin samples collected on days 7, 14, 21, and 28. Daily photographs were taken from the same height, and wound closure was accelerated in the MSC and CM treated groups. Gross morphological parameters were scored, and it was discovered that all wounds in all experimental groups were grossly healthy until day 7, with enhanced granulation tissue noted in MSC and CM treated groups. Cytological analysis revealed a significant increase in mononuclear infiltration after 72-96 hours in the CM and MSC groups. Overall, histomorphological observations revealed that MSC and CM treated groups had significantly accelerated wound closure, advanced re-epithelisation and neovascularisation, mature extra-cellular matrix deposition with better collagen arrangement, and reduced scarring. Every parameter was scored and statistically evaluated.ThesisItem Open Access ROLE OF CURCUMINOID AND SESAMOID OIL IN COLON CANCER DEVLOPMENT AND PROGRESSION(West Bengal University of Animal & Fishery Sciences, Kolkata, 2023) SENAPATI, SANJIB; Hansda, R.N.; Mukhopadhayay, S.K.; Pradhan, S.; Mondal, Samiran; Dey, SamirThesisItem Open Access EFFECT OF CHRONIC STRESS ON REPRODUCTIVE CYCLE AND EXPRESSION PATTERN OF RFRP-3 GENE IN ADULT FEMALE RATS(West Bengal University of Animal & Fishery Sciences, Kolkata, 2023) TZUDIR, AKUMMENLA; Banerjee, D; Bag, S.; Das, K.; Mukherjee, J.; Mondal, S.; Polley, S.The present study was conducted to analyze the effect of chronic stress on the reproductive cycle and expression pattern of the RFRP-3 gene in adult female rats. The experiments were performed on adult female Wistar rats which were divided into two groups viz. Control and the Stressed groups, and were housed individually in polypropylene cages. The rats of the Stressed group were subjected to 3 hrs. of restraint or immobilization stress daily in a modified rat restrainer for 28 days. The reproductive cycle of the female rats was monitored and the different stages of the oestrous cycle (proestrus, estrus, metestrus and diestrus) were determined through the visual assessment of the vagina of the rats and the vaginal exfoliated cell cytology. All rats from both the Control and the Stressed groups were tested for behavioural traits after the completion of the trial period. Different tests viz. open field locomotion test, Porsolt’s forced swim test, Light/Dark Transition test and the Elevated plus maze test were performed to observe the anxiety/depressive behaviour of the rats.The experimental animals of both groups were sacrificed at the end of the trial period. Plasma was separated from the collected blood samples and was estimated for haematological parameters (Hb, PCV, TEC, TLC and DLC), biochemical parameters (Blood glucose, Total antioxidant, T3, T4 and Cortisol) and reproductive hormones (LH, FSH, Progesterone and Estradiol). The tissue samples collected for histological studies were ovary and for the study of the expression of the RFRP-3 gene were brain (hypothalamus). During the visual assessment of the vagina, the vaginal opening of the rat appeared to be wide and moist and the tissues appeared pink during proestrus. During estrus, the vagina appeared similar to proestrus but was less pink and moist. In metestrus, a pale and dry vaginal opening was seen and during the proestrus, the vaginal opening was very moist, small and closed. The result of the vaginal cell cytology showed that a mixture of epithelial cells was seen in the proestrus. The estrus stage was characterized mainly by the presence of anucleated cornified (keratinized) cells. Metestrus shows the presence of leucocytes and some keratinized cells. Polymorphonuclear leucocytes were seen in diestrus. The behavioural analysis of the rats showed that the rats of the stressed group showed more depressive/anxiety-like behaviour when compared to the control group. The PCV and Hb of the stressed rats were highly significantly(P<0.01)lower than the control rats. The TLC and the TEC between the two groups were found to be not statistically significant (P>0.05). A significant (P<0.05) variation was observed in the blood glucose, T3, T4 and Cortisol between the two groups. The variation in the total antioxidant was found to be non-significant (P>0.05). A significant (P<0.05) variation was observed in the LH of the two groups. The variation in the mean with standard error of FSH, Estradiol (E2) and Progesterone of the two groups viz. the Control and the stressed group was found to be not statistically significant (p>0.05). A significant (P<0.05) increase in the expression of the RFRP-3 gene was seen in the hypothalamus of the stressed rat. The results lead us to conclude that chronic stress causes significant changes in the normal blood physiology of the rat. Chronic stress also alters the concentration of the reproductive hormones which confirms the relationship between stress-induced female infertility and the expression of RFRP-3.ThesisItem Open Access EFFECT OF DIETARY SUPPLEMENTATION OF VITAMIN E AND SELENIUM ON THE GROWTH, HEMATOBIOCHEMICAL, ENDOCRINE AND IMMUNE STATUS OF GROWER GHUNGROO PIGS(West Bengal University of Animal & Fishery Sciences, Kolkata, 2022) Hussain, Rubayat; Mukherjee, J.; Das, P.K.; Banerjee, Dipak; Batabyal, Subhasis; Bag, SadanThe focus of the present investigation was to highlight the findings of effectiveness of dietary supplementation of vitamin E and selenium in grower Ghungroo pigs on growth, hematobiochemical, endocrine, immunity and antioxidant status. Eighteen healthy female grower Ghungroo pigs (Susscroffa domestica) of 2-3 months were selected randomly for the experiment and divided into three groups. The control group (n=6) fed only the basal diet, animals of treatment-1 (T1) group (n=6) were supplied with vitamin E (in form of α tocopherol acetate) @11IU/Kg feed and animals of treatment-2 (T2) group of animals (n=6) were supplemented with selenium (as sodium selenite) @0.15mg/Kg feed daily as per NRC recommendations. The body weight, morphometric parameters of growth and physiological responses were recorded at the interval of 15 days. Blood samples were collected from all the experimental animals on day 15, day 30, day 45 and day 60 of supplementation. Vitamin E and selenium were estimated in the plasma. Hematological parameters were evaluated as per standard procedures. Blood biochemical, endocrine (cortisol, T3 and T4) and total antioxidant status were evaluated by commercially available kits as per manufacturer’s protocol. In vitro phagocytic activity and lymphocyte proliferation response were evaluated by colorimetric NBT and MTT assay respectively. The supplementation of vitamin E and selenium significantly (P<0.01) increased plasma vitamin E and selenium concentration in grower Ghungroo pigs from 15th day of supplementation and persisted till the end of experiment. The supplementation of vitamin E and selenium did not have any significant effect on the growth related morphometric parameters in Ghungroo pigs. The supplementation of vitamin E increased haemoglobin and MCH concentration whereas, selenium supplemented pigs showed higher PCV, MCV, TEC and TLC. The supplementation of vitamin E significantly increased plasma glucose and decreased AST and ALT whereas, selenium supplementation resulted higher total protein in experimental pigs. Vitamin E-treated group had a considerably greater T3 and T4 level and lower cortisol level than the control and selenium-treated groups. Vitamin E supplementation improved the immune status of the growing Ghungroo pigs in terms of in vitro lymphocyte proliferation response, phagocytic activity of neutrophils along with increased total antioxidant status. In conclusions, the present investigation depicted that dietary supplementation of vitamin E had beneficial effect on the immunity and antioxidant status in grower Ghungroo pigs without affecting normal growth and physio-biochemical functions.ThesisItem Open Access Effects of dietary turmeric and lemon essential oil on weaning and post-weaning performances of Ghoongroo piglets(West Bengal University of Animal & Fishery Sciences, Kolkata, 2022) DAS, ANIL KUMAR; Das, P.K.; Sanyal, Sagar; Mukherjee, J.; Batabyal, Subhasis; Patra, A.; Naskar, S.ThesisItem Open Access MULTIPLE DOSE EXPOSURE STUDY OF Psidium guajava Linn. ON EXPERIMENTALLY INDUCED INFLAMMATORY BOWEL DISEASE IN ANIMAL MODEL(West Bengal University of Animal & Fishery Sciences, Kolkata, 2023) MEHRA, SANDEEP; Hansda, R.N.; Kumar, Deepak; Mukhapadhayay, S.K.; Mondal, Samiran; Pradhan, Saktipada; Batabyal, KunalThe current study attempted to screen Psidium guajava leaves for their influence on experimentally produced inflammatory bowel illness (IBD). Inflammatory bowel disease (IBD) is a prevalent chronic inflammatory gastrointestinal condition. Crohn's disease (CD) and ulcerative colitis are the two most common kinds of IBD (UC). The anti inflammatory and antioxidant properties of Psidium guajava leaves were investigated in DSS-induced IBD. The experiment was performed using Dextran sulphate Sodium (DSS) 2 % DSS is added to the drinking water for 7 consecutive days except in negative controle. Aquous extract of P. guajava will be given for 28 days [from day 1 to day 28 with DSS(7days )] to know the antiinflammatory and antioxidant properties the rats will be divided into following namely group I: negative control, Group II: positive control, Group IlI: P. guajava leaf extract 30mg/kg, Group IV: P. guajava leaf extract 60 mg/kg and Group V: P guajava leaf extract 120mg/kg. Haemoglobin,TEC, TLC, DLC, Total Platelet Count, and other common haematological parameters and serum biochemistry such as ALT, AST, Kidney Function Test (crt.and urea) total protein and serum electrolyte sodium were assessed and found maximum of test shows significance difference. In cytology-based analysis, anal cytology for representative group of positive control(dss 2%) were taken every day for 7 consecutive days with using swab method in anal rectal region. In histomorphological of small intestine found that mucosal ulcer ,erosion,necrosis amd infiltration of inflammatory cell.In kidney they have been normal architecture with focal. In liver no histopthological changes except glycogen accumulation in liver. In gross pathology inflammatory nodule is seen on 1st and 2nd week of experiment. Histopathological changes of liver, kidney and intestine were significantly reduced by aqueous extract of Psidium guajava in last week of experiment. Therefore psidium guajava has a good protective potential in inflammatory bowel disease.ThesisItem Open Access EFFECT OF YAPAVA TRIPLINERVIS IN LABEO ROHITA AGAINST BACTERIAL INFECTION WITH SPECIAL EMPHASIS ON ULCER CAUSED BY AEROMONAS HYDROPHILA(West Bengal University of Animal & Fishery Sciences, Kolkata, 2022) SAHA, ANKITA; Dash, Gadadhar; Pramanik, Koushik; Abraham, T.J.; Sau, S.K.; Karmakar, S.Motile Aeromonas Septicemia, caused mainly by Aeromonas sp. is an economically significant problem in freshwater water fish farming industry. The goal of the study is to determine the inhibitory effect of Ayapana triplinervis against Aeromonas hydrophila and determining the efficacy of A. triplinervis against ulcer disease of fish caused by A. hydrophila. In bacterial susceptibility test 2.5 μl of Ayapana triplinervis leaf chloroform extract gave zone of inhibition through disc diffusion assay and the minimum inhibitory concentration was determined through agar dilution method. The minimum inhibitory concebtration was 6% Ayapana triplinervis leaf powder (ALP) against A. hydrophila present in 0.1 ml of 108 CFU/ml through agar dilution method in petri plate. The efficacy of Ayapana triplinervis was evaluated by treating the Aeromonas hydrophila infected Labeo rohita with Ayapana triplinervis leaf powder (ALP) admixed with feed at different doses. The Ayapana triplinervis feed treatment at doses of 10 gm/kg feed and 100gm/kg feed gave better results and were effective in reducing mortalities in diseased Labeo rohita. Histopathological findings suggested marked improvement in the gill, spleen, skin, kidney and liver tissues of Labeo rohita infected with Aeromonas hydrophila upon Ayapana triplinervis treatment at higher doses for 14 consecutive days. Ayapana triplinervis also restored serum biomarkers like glucose, creatinine, calcium, ALT and AST to normal range after 14 days treatment with Ayapana triplinervis leaf powder at the doses of 10gm/kg feed and 100gm/kg feed. The hematological properties like total erythrocyte count, haemoglobin concentration, packed cell volume and total leucocyte count were also normal in Labeo rohita challenged with alpha haemolytic and antigenic Aeromonas hydrophila after 14 days Ayapana treatment at the doses of 10gm ALP/kg feed and 100 gm ALP/feed. This study has been suggested to treat ulcer disease in Labeo rohita caused by Aeromonas hydrophila by Ayapana triplinervis leaf (in powder form) as an effective herbal therapeutic agent.ThesisItem Open Access Evaluation of biosafety of oral oxolinic acid administration to Nile tilapia Oreochromis niloticus (L.) juveniles(West Bengal University of Animal & Fishery Sciences, Kolkata, 2022) PATEL, JAYKUMAR BHAGUBHA; Abraham, T.J.; Hoque, F.; Dash, G.; Nagesh, T.S.; Sau, S.K.Oxolinic acid (OA) is one of the broad-spectrum quinolones that is a critically important medicine for humans and is used as a second-line treatment in aquaculture at 12 mg/kg biomass/day for 7 consecutive days. The present study was executed to evaluate the biosafety of OA and to determine its efficacy against Aeromonas hydrophila infection in Nile tilapia Oreochromis niloticus juveniles. The study also evaluated the biochemical, histopathological and haematological alterations in O. niloticus when fed the OA feed. The biosafety experiments were carried out at 0-10 times the therapeutic dose (12 mg) for 21 days. A significant dose-dependent reduction in feed intake and biomass and an increase in mortalities was recorded. Dietary OA administration caused a dose-dependent effect on fish erythrocyte morphology and haematological parameters. A significant increase in plasma glucose, creatinine, alkaline phosphatase, alanine transaminase and aspartate transaminase was documented during the dosing period. On the other hand, the plasma calcium and chloride levels decreased significantly. The gills, kidneys, liver, spleen, and intestine showed mild to marked histopathological changes, indicating the possible toxic effects of OA on different organs of fish. The majority of the alterations, however, recovered upon cessation of OA-dosing. The edible tissue residues, as analyzed by LC-MS/MS, peaked on day 21 of OA-dosing and decreased upon cessation of OA-dosing in all the dosing groups. The residue levels of OA in the muscle tissue of the therapeutic dose group were well within the maximum residue limit (MRL) set by the European Medicines Evaluation Agency (EMEA) and Food Safety and Standards Authority of India (FSSAI). Further, the efficacy of OA was evaluated by challenging O. niloticus juveniles with A. hydrophila intramuscularly and subsequently treated with medicated OA feed for 7 consecutive days at 12 mg/kg biomass/day. The OA treatment gave better results in terms of reducing the clinical biochemical parameters and recovery against A. hydrophila infection compared to the untreated group. The rate of wound healing was also faster in the OA-treated group compared to the untreated group, wherein the wounds healed completely within 12 days of post-injection. Therefore, the efficacy of OA against A. hydrophila infections in fish was established. Although the current study hinted at the safety and tolerability of OA in O. niloticus juveniles in tropical Indian conditions, care must be exercised for its aquacultural application because of its listing as a critically important medicine for humans.ThesisItem Open Access Comparative evaluation of growth and physio-metabolic responses of Osteobrama belangeri (Valenciennes, 1844) in freshwater v/s saline water(West Bengal University of Animal & Fishery Sciences, Kolkata, 2022) BHUNIA, SAYAN; Ghosh, T.K.; Biswas, Gouranga; Das, S.K.; Sau, S.K.; Chowdhury, S.; Saha, B.Salinity is one of the stressful environmental factors which affects life, metabolism, and distribution of aquatic animals. The present study on ՙՙComparative evaluation of growth and physio-metabolic responses of Osteobrama belangeri (Valenciennes, 1844) in freshwater v/s saline water ՚՚ was carried out to find the suitable salinity for growth performance and survival of pengba (Osteobrama belangeri) in inland saline water and to find the effects of different salinity concentrations on haematological, serum biochemical, and proximal parameters of this fish. In first phase of the experiment, 60 fingerlings of O.belangeri (11.78 ±0.057 g)were stocked in control and treatments (each having triplicate containing 10 sp each per replicate ) for 60 days to run a growth study by feeding with formulated (32 % protein ) feed @4% BW . Then in second phase, 18 days experimental study was run considering freshwater as control ( C ) and saline water as treatment (T) with gradually increasing salinities (2,4,6,8,10 and 12 ppt) at 72 hours interval in each salinity concentration. The result indicated that O. belangeri can tolerate the salinities up to 12 ppt. in inland saline water where as 100% mortality was observed in 12 ppt . MLS96h was calculated and LC 50 was determined using mortality data. Almost similar rate of growth and survival was found up to 8 ppt compared to control. Salinity more than 8 ppt appeared to be unsuitable for O. belangeri due to increased mortality and impaired growth. A few important physiological parameters i.e proximal value, behavioral changes were observed during 18 days of experimental phase . To determine haematological alterations, RBCs, haematocrit (HCT%), haemoglobin (Hb) , platelets count , WBC, MCH , MCHC , MCV values are determined and significant alterations ( p<0.05) found at different salinity concentration . To determine biochemical alterations, ALT (Alanine aminotransferase) or SGPT (Serum glutamate pyruvate transaminase), AST (Aspartate aminotransferase) or SGOT (serum glutamic oxalo acetic transaminase), SOD (superoxide dismutase) and Catalase (CAT) are examined collecting the blood serum. The increasing value of liver enzyme ALT and AST in blood indicated hepatic damages in higher salinities. However, all the biochemical parameters were found as almost normal up to 8 ppt compared with control. This evidence suggests that pengba can sustain and grow well below 8 ppt and can be a potential candidate for culture in coastal areas after heavy downpour when the salinities level falls to 8 ppt or lower.
