Veterinary Research Forum

Abstract Interstitial cells of Cajal (ICC) play a crucial role in regulating gastrointestinal motility, functioning as pacemakers in the digestive system of equids. This study described and compared the ICC populations in different regions of the stomachs of equids (horses, donkeys, and mules). A total of 30 healthy stomachs from horses, donkeys, and mules were evaluated using immunohistochemistry and high-resolution microscopy to assess various regions of the stomach in these species. The results showed the presence of ICC in all three equid species, with a predominance in the submucosa across the squamous mucosa, glandular mucosa, and pyloric antrum. The density and morphology of ICC varied slightly between species, although no significant differences were found. Among the equids studied, donkeys showed the lowest ICC density, while horses exhibited the highest in the evaluated tissues. Four cellular morphologies were identified, three of which were included in the analysis, as toluidine staining revealed that one corresponded to mast cells. Further studies are needed to elucidate dynamics and possible alterations in response to gastric pathologies.

Abstract Continuous exposure to pesticides may affect ovarian function and folliculogenesis by altered oxidative statute, steroidogenesis, and oocyte quality. Sweet apricot kernels (SAK) have been traditionally used as phytotherapeutic agents known for their broad pharmacological potential, including anti-oxidant, anti-inflammatory, and anti-cancer properties. This study aimed to assess the protective effect of SAK against ovarian toxicity induced by sub-chronic dietary exposure to a pesticide mixture, consisting of trifloxystrobin and thiacloprid, in female rats. Thirty-two female Wistar rats were randomly assigned into four groups of eight. Samples in 0.50 mL were given by oral gavage for 100 consecutive days. The control group received distilled water, SAK group included untreated rats received SAK extract (50.00 mg kg-1), Mix group received trifloxystrobin (0.30 mg kg^-1) + thiacloprid (0.20 mg kg^-1), and SAK + Mix group included Mix treated rats supplemented with SAK extract (50.00 mg kg^-1). Ovarian tissue samples were collected at the end of experiment for biochemical, histopathological, and immunohistochemical analyses. Pesticide mixture induced ovarian toxicity, characterized by a significant decrease in serum estradiol levels, ovarian total anti-oxidant capacity, ovarian follicle counts, Bcl-2 immuno-staining, and ovarian superoxide dismutase and catalase activities. In addition, ovarian malondialdehyde levels, serum interleukin-6 concentrations, and p53 immuno-staining were increased significantly. The co-administration of SAK extract significantly attenuated malondialdehyde, restored total anti-oxidant capacity levels, and modulated inflammation, estrogen levels, apoptosis-related proteins expression, and catalase and superoxide dismutase enzymes activities. Overall, these results suggest that SAK may protect against pesticide-induced ovarian toxicity through antioxidant, anti-inflammatory, and antiapoptotic mechanisms.

Abstract Bisphenol A (BPA), a typical environmental endocrine disruptor, induces trans-generational reproductive injury in animals and humans. This study investigated the protective effects and mechanisms of Semen Cuscutae flavonoids (SCFs) against pre-natal BPA exposure-induced reproductive toxicity in male and female mice offspring. Pregnant Kunming mice were intragastrically administered corn oil (control), 50.00 mg kg^-1 per day BPA (BPA group), or combined 50.00 mg kg^-1 per day BPA and 1.00 mg kg^-1 per day SCFs (BPA+SCF group) from gestation day one to gestation day 14. Serum, testes and ovaries from 8-week-old offspring mice were collected, and multiple reproductive-related indicators were detected via enzyme-linked immunosorbent assay, terminal deoxynucleotidyl transferase-mediated dUTP nick‑end labeling, immunohistochemistry, and RT-qPCR. Results showed that pre-natal BPA exposure significantly reduced serum testosterone in male offspring, and elevated estradiol while suppressed follicle stimulating hormone and luteinizing hormone in female offspring, accompanied by abnormal gonadal histomorphology and sharply increased gonadal apoptotic cells. At the molecular level, BPA down-regulated steroidogenic genes, including steroidogenic acute regulatory protein and CYP11α, and folliculogenesis-related KIT ligand, up-regulated ovarian anti-Müllerian hormone, disturbed pro-apoptotic Bax/ anti-apoptotic Bcl-2 balance. The SCFs intervention reversed BPA-induced hormonal disorders and gonadal histological lesions. Mechanistically, SCFs up-regulated Bcl-2 and down-regulated Bax genes to reduce gonadal apoptosis. Meanwhile, SCFs restored the mRNA and protein expressions of Steroidogenic acute regulatory protein, CYP11α, KIT ligand and anti-Müllerian hormone. In conclusion, SCFs exert protective effects against pre-natal BPA-triggered offspring reproductive toxicity by balancing apoptotic signaling and modulating steroidogenic and folliculogenic genes expressions.

Abstract Abomasal motility disorders, such as displacement and volvulus, represent significant clinical and economic challenges in dairy cattle. The use of synthetic prokinetics is limited by residue concerns. This study investigated the in vitro effects of eugenol on bovine abomasal smooth muscle contractility. Muscle strips from healthy cattle were mounted in organ baths containing oxygenated Krebs solution. Basal tone and contractions induced by carbachol (1.00 and 4.00 µM), barium chloride (30.00 mM), and potassium chloride (30.00 and 80.00 mM) were recorded. Cumulative eugenol concentrations (1.00 - 600 mg L^-1) caused a significant, concentration-dependent inhibition of both basal tone and spasmogen-induced contractions (Friedman test followed by Dunnett's test). The inhibition was particularly abrupt in high- potassium chloride -induced contractions. The effects were fully reversible upon washout. These results demonstrated that eugenol acted as a non-selective spasmolytic agent on bovine abomasal smooth muscle, likely via inhibition of calcium influx. This supported its potential as a natural therapeutic candidate for managing gastrointestinal hypermotility in ruminants, warranting further in vivo studies.

Abstract The epithelial-mesenchymal transition is a key molecular process in the development of metastasis in breast cancer. The aim of this study was to purify low molecular weight peptides derived from scorpion venom (PESV) from the venom of Hottentotta zagrosensis and to evaluate their anti metastatic potential in the human breast cancer cell line Michigan Cancer Foundation. Scorpion venom was fractionated using Fast Performance Liquid Chromatography and ultrafiltration. The separated fractions were loaded onto Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis and stained with silver nitrate. This process identified protein bands with molecular weights below 10.00 kDa, which were subsequently analyzed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and Western blotting. Michigan Cancer Foundation cells were exposed to 10.00 and 20.00 µg mL^-1 of PESV, and their effects on the expression of E cadherin, N cadherin, Vimentin, Snail and Twist proteins were evaluated by Western blotting. The treatment significantly increased the epithelial marker E cadherin, while simultaneously reduced the mesenchymal markers N cadherin and Vimentin as well as the transcription factors Snail and Twist. These changes in PESV treated groups were dose dependent, and at 20.00 µg mL^-1 the effect was comparable to that of cisplatin. In conclusion, these findings suggested that PESV, extracted from the venom of H. zagrosensis, effectively suppressed the aggressive phenotype of cancer cells while promoting their reversion to an epithelial state. The preclinical evidence demonstrated that this peptide markedly inhibited metastasis, underscoring its potential as a promising therapeutic candidate for breast cancer.

Abstract Optimization of dietary lipid (LIP) utilization through dietary lecithin and L-carnitine (L-CA) supplementation is a key nutritional strategy to improve growth performance and feed efficiency of carnivorous ornamental fish. However, limited information is available on the combined effects of dietary LIP, soybean lecithin (SBL), and L-CA content in Oscar fish. The effects of SBA and L-CA at high and low dietary LIP levels were evaluated on Oscar fish. A total number of 225 fish (6.50 ± 0.50 g) were randomly allocated to five dietary groups including: control diet (LIP 10.00%), diet containing 18.00% LIP, diet containing 18.00% LIP + 4.00% SBL, diet containing 18.00% LIP + 500 mg kg^-1 L-CA, and diet containing 18.00% LIP + 4.00% SBL + 500 mg kg^-1 L-CA for a 6-week period. Fish fed the diet containing 18.00% LIP + 4.00% SBL exhibited higher final body weight, specific growth rate, and weight gain, alongside a decreased Feed conversion ratio compared to other groups. No significant differences were detected in carcass moisture or protein content among experimental groups, whereas, LIP content was significantly elevated in the 18.00% LIP + 4.00% SBL diet. Additionally, hepatic antioxidant enzyme activities (e.g., superoxide dismutase, catalase and glutathione peroxidase) as well as intestinal digestive enzymes (e.g., alkaline protease, lipase, and alpha-amylase) did not differ significantly across experiment groups. In conclusion, a diet of 18.00% LIP + 4.00% SBL is recommended to enhance growth performance and feed efficiency in Oscar fish.

Abstract Oxidative stress is a key factor in male infertility and cryopreservation-induced sperm damage, impairing motility, viability and DNA integrity. Antioxidant supplementation has been proposed as a strategy to mitigate cryoinjury, with marine macroalgae such as Padina australis offering potent bioactive compounds. This study investigated the effects of P. australis methanolic extract on bull sperm quality and embryo development in vitro. Semen freezing media were supplemented with P. australis methanolic extract at concentrations of 0.00, 10.00, 20.00, 40.00, 80.00, and 160-ppm. Supplementation produced significant dose-dependent effects across sperm parameters. The 20.00-ppm group consistently showed the highest values for motility, velocity measures: Curvilinear velocity, average path velocity, straight line velocity, kinematic ratios: straightness and linearity, while anterior lateral head displacement and beat cross frequency also increased significantly. Antioxidant activities were markedly elevated at 20.00-ppm, whereas, higher doses (80.00 - 160-ppm) failed to sustain improvements and, in some cases, negatively impacted motility and DNA damage. Live sperm and sperm membrane integrity values were significantly improved at 20.00-ppm, while 160-ppm reduced Hypo-osmotic swelling test and increased DNA damage. Embryo development outcomes mirrored sperm quality, with fertilization, cleavage and blastocyst formation significantly enhanced at 20.00-ppm. Conversely, DNA damage malondialdehyde, activity, unfertilized oocytes and degeneration rates were lowest at this concentration. These findings demonstrated that moderate supplementation with P. australis methanolic extract optimally improved post-thaw sperm function and embryo development. Marine algal extracts, thus, represented promising and natural cryoprotectants for enhancing livestock breeding efficiency and genetic resource conservation.

Abstract Ischemia-reperfusion injury (IRI) is one of the leading causes of cellular necrosis, inflammation, and kidney failure. This study aimed to investigate the combined effects of granulocyte colony-stimulating factor (GCSF) and intravenous lipid emulsion (ILE) in mitigating kidney damage caused by IRI in rats. Twenty-five male Sprague-Dawley rats were divided into five groups, including IRI + ILE + GCSF, IRI + ILE, IRI + GCSF, IRI, and sham. Ischemia was induced by clamping the renal pedicle for 45 min. The GCSF (20.00 µg kg^-1) was administered intraperitoneally on days zero, three, and seven, while ILE (5.00 mL kg^-1) was given intravenously for seven days. After the two-week treatment period, the left kidneys of the rats were excised via nephrectomy. The kidney samples were immediately preserved in 10.00% formalin, processed using standard histological methods, and paraffin-embedded. Sections were prepared, and after Hematoxylin and Eosin staining, histomorphometric analyses were conducted. The TUNEL staining was used to quantify apoptotic cells. The IRI group showed the highest increase in urinary space (33.50 ± 0.49), cellular necrosis (score: 3.40), and apoptosis (score: 29.64). The ILE + GCSF group significantly reduced these parameters (urinary space: 15.30 ± 0.47, necrosis: 1.80, and apoptosis: 5.24%). Additionally, inflammation levels, renal tubular dilation, and protein cast formation were significantly lower in the ILE + GCSF group compared to the IRI group. The combination of GCSF and ILE significantly reduced damage-related parameters. The GCSF's protective effect in reducing tissue damage and cell apoptosis following IRI is further enhanced by ILE, providing a synergistic effect.

Abstract Leptospira interrogans antigens in the uterus, mammary gland and kidney tissues of sheep, employing multiple diagnostic approaches in this study. The research material comprised tissue samples (uterus, mammary gland, kidney) and blood specimens collected from 80 sheep slaughtered in abattoirs. Serum samples were analyzed using the enzyme-linked immunosorbent assay (ELISA), while tissue samples were examined by ELISA, Hematoxylin–Eosin staining, immunohistochemistry and immunofluorescence (IF). The ELISA testing of sera revealed leptospiral antibodies in 16 animals, corresponding to a seroprevalence of 20.00% (16/80). Tissue-based ELISA demonstrated L. interrogans antigen positivity in 11.25% (9/80) of uterus samples, 6.25% (5/80) of mammary gland samples, and 15.00% (12/80) of kidney samples. Immunohistochemistry and immunofluorescence analyses yielded same results, with antigen detection rates of 7.50% (6/80) in uterus, 5.00% (4/80) in mammary gland and 11.25% (9/80) in kidney tissues. These findings confirmed the presence of L. interrogans in reproductive and renal tissues of sheep and highlighted the kidney as the primary site of colonization. The combined use of ELISA, immunohistochemistry, and immunofluorescence provided complementary evidence for antigen detection and localization. Overall, the results contributed to a deeper understanding of leptospirosis pathogenesis in sheep and underscored the importance of comprehensive diagnostic approaches in livestock health management.