Veterinary Research Forum

Abstract Aging markedly increases vulnerability to cardiovascular disorders, including acute coronary syndrome and coronary artery disease. Although melatonin (M) and nicotinamide mono-nucleotide (N) have each shown cardioprotective properties, their combined effects on ischemia-reperfusion (IR) injury in the aged heart remain unclear. This study investigated co-administration of M and N enhances cardiac functional recovery and reduces inflammatory responses following myocardial IR injury in aged rats. Forty adult male Wistar rats (n = 8/group) were randomly assigned to five groups: Sham, IR, M (IR + M), N (IR + N), and N + M (IR + N + M). Cardiac function was evaluated after IR by measuring coronary flow, heart rate, left ventricular end-diastolic pressure, and left ventricular developed pressure. Melatonin was administered at 50.00 µM in the reperfusion solution for 15 min after 30 min of ischemia, N was administered intraperitoneally at 100 mg kg-1 every other day for 28 days. Creatine kinase and inflammatory markers, including interleukin IL-1β, IL-6, tumor necrosis factor-alpha, and nuclear factor kappa B, were also assessed. IR significantly impaired coronary flow and heart rate, both of which were improved by M and N treatment. These agents also reduced left ventricular end-diastolic pressure and improved left ventricular developed pressure recovery, with the combined therapy producing the greatest effect. Likewise, creatine kinase and inflammatory marker levels were significantly elevated in the IR group but markedly reduced by treatment, especially by combined administration. These findings suggest that M and N exert additive cardioprotective effects against IR injury in aged rats.

Abstract This study aimed to explore whether melatonin protects TM3 Leydig cells from cobalt (II) chloride (CoCl₂)-induced hypoxia through the transforming growth factor beta (TGF-β) signaling pathway. Cells were divided into four groups: a control group without treatment (Group 1), a melatonin group (10.00 ng mL-1; Group 2), a group treated with CoCl2 (100 µM) to induce hypoxia (Group 3), and a melatonin + CoCl₂ group (Group 4). After 96 hr of incubation, cell viability was assessed using the MTT assay, and transforming growth factor beta 1, activin receptor-like kinase-5, and bone morphogenetic protein 4 gene and protein expressions were measured through RT‑PCR and western blotting. The CoCl₂ and melatonin + CoCl₂ groups exhibited significantly diminished cell viability compared to the control. However, melatonin treatment enhanced survival in the CoCl₂-exposed cells. Notably, transforming growth factor beta 1 expression was elevated in all groups. Activin receptor-like kinase-5 (gene and protein expression increased in CoCl₂-treated groups but was lower in the melatonin + CoCl₂ group. Melatonin treatment reduced bone morphogenetic protein 4 expression compared to the control, while CoCl₂ groups showed increased bone morphogenetic protein 4 levels. These findings suggest melatonin's potential as a therapeutic agent against oxidative stress and hypoxia in TM3 cells through its antioxidant properties.

Abstract Spinal cord injury (SCI) results in the demise of neural and glial cells, as well as extensive neuro- inflammation. Hydrogel formulation for prolonged release of melatonin (Mel) has demonstrated enhanced effectiveness and safety. In this study, SCI was induced in rats by contusion at the T₉ vertebrae. Chitosan (CH) /Mel hydrogel was fabricated and characterized using scanning electron microscopy (SEM) and Fourier transform infra-red to examine its specific effects on the apoptotic and histopathological markers of SCI. The scanning electron microscopy images revealed the presence of porosity in the CH/Mel hydrogel. Forty male Wistar rats were randomly divided into five groups (n = 8), including sham, control (SCI-induced treated locally with 100 µL CH hydrogel), and groups 3, 4, and 5 (treated locally immediately after SCI induction with 100 µL CH hydrogel containing 50.00, 100, and 200 mg kg^-1 Mel, respectively). The CH/Mel hydrogel at a dose of 25.00 mg mL^-1 significantly increased cell viability in the U87 cell line after 24 hr of exposure. However, at 48 and 72 hr after exposure, Bax and Bcl2 expressions were significantly increased and reduced in the SCI group, respectively, and CH/Mel hydrogel could alleviate their expressions, especially in higher doses. In addition, S100 protein expression was up-regulated in the SCI group. However, CH/Mel hydrogel down-regulated it in a dose-dependent manner. The histopathological findings demonstrated that CH/Mel hydrogel dramatically improved SCI outcomes, like vacuolar degeneration, necrosis, and severe cystic and axonal degenerations. In conclusion, CH/Mel hydrogel induced neuroprotection and it had the potential to be used as a therapeutic agent for the treatment of SCI.

Abstract This study investigated carbendazim (CBZ)-induced hepatic dysfunction and the mechanistic pathway regarding the protective effect of melatonin (MEL). Twenty-eight male rats were grouped as follows: Control, CBZ (150 mg kg^-1), MEL (20.00 mg kg^-1), and CBZ + MEL. The experiment was conducted for 60 days. Tissue samples were stained with Hematoxylin and Eosin and immuno-fluorescence methods to examine apoptotic pathway. Also, hepatic enzymes and miR-122 expression were evaluated. The findings indicated that the CBZ group exhibited an increase in degenerated hepatocytes, hyperemia of sinusoids, and leukocyte infiltration, accompanied by elevated levels of aspartate aminotransferase and alanine aminotransferase, as well as up-regulation of miR-122. Also, there was a significant increase in the fluorescence intensities of caspase-3 and Bax in the CBZ group, whereas a substantial reduction in the fluorescence intensity of Bcl-2 was recorded. In contrast, the simultaneous administration of MEL alongside CBZ was shown to be effective in improving histological structure, decreasing levels of aspartate aminotransferase and alanine aminotransferase, reducing the apoptosis index, and modulating the expression of miR-122 in comparison with the CBZ-only group. The increased expression of miR-122 noted in the CBZ group may correlate with an elevation in the immunoreactivity of apoptosis markers and alterations in liver architecture. Additionally, MEL seems to alleviate CBZ-induced hepatotoxicity by down-regulating miR-122 expression, diminishing the fluorescence intensity of caspase-3 and Bax, and enhancing the immunoreactivity of Bcl-2. Collectively, the regulation of miR-122 may serve as a potential mechanism by which MEL confers its protective effects against liver damage induced by CBZ.

Abstract Leydig cells play a crucial role in male reproductive physiology, and their dysfunction is often associated with male infertility. Hypoxia negatively affects the structure and function of Leydig cells. This study aimed to investigate the impact of melatonin on the c-Jun N-terminal kinase (Jnk), P38, and extra-cellular signal-regulated kinases 1 and 2 (Erk1/2) mitogen-activated protein kinase (MAPK) signaling pathways in TM3 mouse Leydig cells under hypoxia induced by cobalt (II) chloride (CoCl₂). The TM3 cell line was utilized as a subject of research, and 100 μM CoCl₂ was employed to induce hypoxia. Following the addition of 10.00 ng mL^-1 melatonin, quantitative reverse transcription-polymerase chain reaction and western blot analyses were conducted to assess the gene expression and protein level of Jnk, p38, and Erk1/2, while enzyme-linked immunosorbent assay was used to measure testosterone secretion. The results showed that melatonin significantly increased testosterone production in the CoCl₂ + melatonin group compared to the CoCl₂-treated group. Furthermore, melatonin elevated both the protein level and mRNA expression of Erk1/2, Jnk, and p38 genes in the CoCl₂ + melatonin group compared to the CoCl₂ group. In conclusion, melatonin activated the Jnk, p38, and Erk1/2 MAPK signaling pathways and enhanced testosterone production in the presence of CoCl₂ in TM3 cells.

Abstract The aim of the present study was to evaluate the effect of melatonin as an antioxidant on spatial navigation memory in male diabetic rats. Thirty-two male white Wistar rats weighing 200 ± 20 g were divided into four groups, randomly: control, melatonin, diabetic and melatonin-treated diabetic. Experimental diabetes was induced by intraperitoneal injection of 50 mg kg-1 streptozotocin. Melatonin was injected (10 mg kg-1 day-1, ip) for 2 weeks after 21 days of diabetes induction. At the end of administration period, the spatial navigation memory of rats was evaluated by cross-arm maze. In this study lipid peroxidation levels, glutathione-peroxidase and catalase activities were measured in hippocampus. Diabetes caused to significant decrease in alternation percent in the cross-arm maze, as a spatial memory index, compared to the control group (p < 0.05), whereas administration of melatonin prevented the spatial memory deficit in diabetic rats. Also melatonin injection significantly increased the spatial memory in intact animals compared to the control group (p < 0.05). Assessment of hippocampus homogenates indicated an increase in lipid peroxidation levels and a decrease in GSH-Px and CAT activities in the diabetic group compared to the control animals, while melatonin administration ameliorated these indices in diabetic rats. In conclusion, diabetes induction leads to debilitation of spatial navigation memory in rats, and the melatonin treatment improves the memory presumably through the reduction of oxidative stress in hippocampus of diabetic rats.