Melatonin Preserves Fluidity in Cell and Mitochondrial Membranes against Hepatic Ischemia–Reperfusion

We evaluated the in vivo effects of melatonin treatment on oxidative damage in the liver in an experimental model of ischemia–reperfusion. A total of 37 male Sprague-Dawley rats were randomly divided into four groups: control, ischemia, ischemia + reperfusion, and ischemia + reperfusion + melatonin. Hepatic ischemia was maintained for 20 min, and the clamp was removed to initiate vascular reperfusion for 30 min. Melatonin (50 mg/kg body weight) was intraperitoneally administered. Fluidity was measured by polarization changes in 1-(4-trimethylammoniumphenyl)- 6-phenyl-1,3,5-hexatriene-p-toluene sulfonate). After 20 min of ischemia, no significant changes were observed in cell and mitochondrial membrane fluidity levels, lipid peroxidation, and protein carbonylation. However, after 30 min of reperfusion, membrane fluidity decreased compared to controls. Increases in lipid and protein oxidation were also seen in hepatic homogenates of animals exposed to reperfusion. Melatonin injected 30 min before ischemia and reperfusion fully prevented membrane rigidity and both lipid and protein oxidation. Livers from ischemia–reperfusion showed histopathological alterations and positive labeling with antibodies to oxidized lipids and proteins. Melatonin reduced the severity of these morphological changes and protected against in vivo ischemia–reperfusion-induced toxicity in the liver. Therefore, melatonin might be a candidate for co-treatment for patients with hepatic vascular occlusion followed by reperfusionGobierno de Aragón (grant No. B56_23D

Feline Leishmaniosis in Northwestern Italy: Current Status and Zoonotic Implications

Leishmaniasis remains one of the major neglected tropical diseases. The epidemiological profile of the disease comprises a wide range of hosts, including dogs and cats. Despite several studies about feline Leishmaniosis, the role of cats in disease epidemiology and its clinical impact is still debated. The present study raises awareness about the impact of leishmaniasis in cats from an endemic region in of Northwestern Italy (Liguria). A total number of 250 serum and 282 blood samples were collected from cats, then assessed for Leishmania infantum (L. infantum) serologically using western blot (WB) and molecularly using polymerase chain reaction (PCR). We also tested the association of Leishmania infection with some infectious agents like haemotropic Mycoplasma, Feline immunodeficiency virus (FIV) and Feline leukemia virus (FeLV) together with the hematobiochemical status of the examined animals. Interestingly, all tested animals were asymptomatic and out of 250 examined serum samples, 33 (13.20%) samples (confidence interval (CI) 95% 9.56–17.96%) were positive at WB for L. infantum, whereas of the 282 blood samples, 80 (28.36%) returned a positive PCR (CI 95% 23.43–33.89%). Furthermore, there was a statistical association between PCR positivity for L. infantum and some hematological parameters besides FIV infection as well as a direct significant correlation between Mycoplasma infection and WB positivity. Taken together, the present findings report high prevalence of L. infantum among cats, which reinforces the significance of such positive asymptomatic animals and confirms the very low humoral response in this species. In addition, the laboratory values provide evidence that infection by the parasite is linked to alteration of some hematological parameters and is correlated to some infectious agents. These data are of interest and suggest future research for accurate diagnosis of such zoonosis

S-Methylcysteine (SMC) Ameliorates Intestinal, Hepatic, and Splenic Damage Induced by Cryptosporidium parvum Infection Via Targeting Inflammatory Modulators and Oxidative Stress in Swiss Albino Mice

Cryptosporidiosis has been proposed to be one of the major causes of diarrhoeal disease in humans worldwide that possesses zoonotic concern. Thereby, this study investigated the potential effects of s-Methylcysteine (SMC) on the parasite in vivo followed by the measurement of cytokines, oxidative stress parameters, and an investigation of the major histopathological changes. Sixty male Swiss albino mice weighing 20–25 g were allocated equally into five groups and orally administered saline only (control), SMC only (SMC50) (50 mg/kg b.w.), and 104 Cryptosporidium parvum oocysts per mouse via an esophageal tube (C + ve untreated). The fourth and fifth groups (C + SMC25, C + SMC50) administrated 104 C. parvum oocysts combined with SMC25 (low dose) and 50 (high dose) mg/kg b.w., respectively. At days 7 and 14 post-infection (PI), the feces was collected from each group in order to count C. parvum oocysts. After two weeks of treatment, the animals were euthanized and the serum was collected for biochemical analysis. Next, the intestinal, spleen, and liver sections were dissected for histopathological examination. The results revealed lower oocyst numbers in the C + SMC25 and C + SMC50 groups compared to the infected untreated group. Moreover, higher doses of SMC treatment significantly reduced the enteritis induced by C. parvum in a dose-dependent manner. The hepatic lesions were also mitigated as demonstrated in C + SMC25 and C + SMC50 groups unlike the infected group via lowering the serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP) enzymes and increasing albumin and globulin serum levels. SMC administration also reduced cytokines production (SAP, TNF-α, IL-6, and IFN-γ) mediated by Cryptosporidium infection in contrast to the infected untreated group. There were marked lymphoid depletion and amyloidosis observed in the infected untreated group, while the treated groups showed obvious increase in the lymphoid elements. Moreover, the scoring of intestinal parasites, hepatic, and splenic lesions in the SMC-treated groups exhibited significantly lower pathological lesions in different organs in a dose-dependent manner, compared to the infected untreated group. Our results also revealed a significant change in the malondialdehyde content with an elevation of glutathione and superoxide dismutase in the intestines collected from C + SMC25 and C + SMC50 mice relative to the untreated group. Taken together, our results indicated that SMC could be a promising effective compound for treating and declining C. parvum infestation via restoring structural alterations in different tissues, enhancing antioxidant enzymes, and suppressing the cytokines liberation

Changes in plasma susceptibility to lipid peroxidation and vitamin C in preterm and full-term neonates

Introduction: This study was designed to compare the plasma lipid peroxidation (LPO) levels in preterm and full-term neonates and their respective mothers, to assess their relationship with the degree of oxidative stress and the levels of vitamin C, an important antioxidant of the body. Material and methods: The studied groups included 70 neonates, 30 preterm (24-36 weeks of gestation) and 40 full-term (37-42 weeks) neonates. Blood samples were obtained from the cord blood in neonates and from the antecubital vein in their mothers at the time of delivery. Plasma susceptibility to LPO was fluorometrically measured before and after its incubation with 2,2'-azobis-2- amidinopropane hydrochloride (AAPH). Plasma vitamin C level was measured by HPLC. Results: The basal LPO levels were similar in all groups of patients. After AAPH incubation, however, plasma LPO significantly (P<0.0001) increased in all groups, although maternal plasma (full-term, 6.62±0.14 and preterm, 8.76±0.03 mmol/l) showed higher (P<0.001) levels of LPO than their respective babies (full-term, 5.11±0.03 and preterm, 7.74±0.15 mmol/l). AAPH-induced LPO was higher in both maternal and preterm neonates’ plasma than in full-term ones (P<0.001). Vitamin C levels were similar in maternal plasma of both groups, but preterm neonates showed higher levels than full-term ones (171.65±9.38 vs. 118.25±2.75 mmol/l respectively, P<0.001). Conclusions: The results suggest that the preterm group was more prone to LPO than the full-term group, whereas vitamin C was not correlated with the degree of oxidative stress.This work was partially supported by grants G03/137, PI02/1447 and PI03/0817 from Instituto de Salud Carlos III, Spain, and CTS-101 from Consejería de Educación, Junta de Andalucía, Spain

Fortification of Fermented Camel Milk with Salvia officinalis L. or Mentha piperita Leaves Powder and Its Biological Effects on Diabetic Rats

Researchers Supporting Project number (RSPD2023R655), King Saud University, Riyadh, Saudi Arabia.Supplementary Materials: The following supporting information can be downloaded from: https://www.mdpi.com/article/10.3390/molecules28155749/s1The incorporation of fermented camel milk with natural additives possesses numerous benefits for the treatment of various pathological and metabolic conditions. The present study investigated the impact of fortification of fermented camel milk with sage or mint leaves powder (1 and 1.5%, respectively) on glucose and insulin levels, lipid profile, and liver and kidney functions in alloxan-induced diabetic rats. The gross chemical composition of sage and peppermint leaves powder was studied. The chemical composition of sage and mint extracts was performed using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-MS) of sage and mint extracts. Furthermore, a total of forty-two adult normal male albino rats were included in this study, whereas one group was kept as the healthy control group (n = 6 rats) and diabetes was induced in the remaining animals (n = 36 rats) using alloxan injection (150 mg/kg of body weight). Among diabetic rats groups, a control group (n = 6 rats) was kept as the diabetic control group whereas the other 5 groups (6 rats per group) of diabetic rats were fed fermented camel milk (FCM) or fermented camel milk fortified with 1 and 1.5% of sage or mint leaves powder. Interestingly, the oral administration of fermented camel milk fortified with sage or mint leaves powder, at both concentrations, caused a significant decrease in blood glucose level and lipid profile, and an increase in insulin level compared to the diabetic control and FCM groups. Among others, the best results were observed in the group of animals that received fermented camel milk fortified with 1.5% sage powder. In addition, the results revealed that the fermented camel milk fortified with sage or mint leaves powder improved the liver and kidney functions of diabetic rats. Our study concluded that the use of sage and mint leaves powder (at a ratio of 1.5%) with fermented camel milk produces functional food products with anti-diabetic activity.Researchers Supporting Project, King Saud University, Riyadh, Saudi Arabia RSPD2023R655European Union-NextGenerationE

Dose-Dependent Effect of Melatonin on BAT Thermogenesis in Zücker Diabetic Fatty Rat: Future Clinical Implications for Obesity

Experimental data have revealed that melatonin at high doses reduced obesity and improved metabolic outcomes in experimental models of obesity, mainly by enhancing brown adipose tissue (BAT) thermogenesis. A potential dose-response relationship has yet to be performed to translate these promising findings into potential clinical therapy. This study aimed to assess the effects of different doses of melatonin on interscapular BAT (iBAT) thermogenic capacity in Zücker diabetic fatty (ZDF) rats. At 6 wk of age, male ZDF rats were divided into four groups (n = 4 per group): control and those treated with different doses of melatonin (0.1, 1, and 10 mg/kg of body weight) in their drinking water for 6 wk. Body weight (BW) was significantly decreased at doses of 1 and 10 mg/kg of melatonin, but not at 0.1 mg/kg compared with the control, with a similar rate of BW decrease being reached at the dose of 1 mg/kg (by ~11%) and 10 mg/kg (by ~12%). This effect was associated with a dose-dependent increase in the thermal response to the baseline condition or acute cold challenge in the interscapular area measurable by infrared thermography, with the highest thermal response being recorded at the 10 mg/kg dose. Upon histology, melatonin treatment markedly restored the typical brownish appearance of the tissue and promoted a shift in size distribution toward smaller adipocytes in a dose-dependent fashion, with the most pronounced brownish phenotype being observed at 10 mg/kg of melatonin. As a hallmark of thermogenesis, the protein level of uncoupled protein 1 (UCP1) from immunofluorescence and Western blot analysis increased significantly and dose-dependently at all three doses of melatonin, reaching the highest level at the dose of 10 mg/kg. Likewise, all three doses of melatonin modulated iBAT mitochondrial dynamics by increasing protein expression of the optic atrophy protein type 1 (OPA1) fusion marker and decreasing that of the dynamin-related protein1 (DRP1) fission marker, again dose-dependently, with the highest and lowest expression levels, respectively, being reached at the 10 mg/kg dose. These findings highlight for the first time the relevance of the dose-dependency of melatonin toward BW control and BAT thermogenic activation, which may have potential therapeutic implications for the treatment of obesity. To clinically apply the potential therapeutic of melatonin for obesity, we consider that the effective animal doses that should be extrapolated to obese individuals may be within the dose range of 1 to 10 mg/kg.Ministrio de Ciencia e Innovacion (Spain) SAF2016-79794-REuropean CommissionUniversity of Granada & FEDER Andalucia-UGR 2020 B-CTS-102-UGR2

Melatonin Improves Skeletal Muscle Structure and Oxidative Phenotype by Regulating Mitochondrial Dynamics and Autophagy in Zücker Diabetic Fatty Rat

Obesity-induced skeletal muscle (SKM) inflexibility is closely linked to mitochondrial dysfunction. The present study aimed to evaluate the effects of melatonin on the red vastus lateralis (RVL) muscle in obese rat models at the molecular and morphological levels. Five-week-old male Zücker diabetic fatty (ZDF) rats and their age-matched lean littermates (ZL) were orally treated either with melatonin (10 mg/kg body weight (BW)/24 h) (M–ZDF and M–ZL) or non-treated (control) (C–ZDF and C–ZL) for 12 weeks. Western blot analysis showed that mitochondrial fission, fusion, and autophagy were altered in the C-ZDF group, accompanied by reduced SIRT1 levels. Furthermore, C-ZDF rats exhibited depleted ATP production and nitro-oxidative stress, as indicated by increased nitrites levels and reduced SOD activity. Western blotting of MyH isoforms demonstrated a significant decrease in both slow and fast oxidative fiber-specific markers expression in the C-ZDF group, concomitant with an increase in the fast glycolytic fiber markers. At the tissue level, marked fiber atrophy, less oxidative fibers, and excessive lipid deposition were noted in the C-ZDF group. Interestingly, melatonin treatment partially restored mitochondrial fission/fusion imbalance in the RVL muscle by enhancing the expression of fission (Fis1 and DRP1) markers and decreasing that of fusion (OPA1 and Mfn2) markers. It was also found to restore autophagy, as indicated by increased p62 protein level and LC3BII/I ratio. In addition, melatonin treatment increased SIRT1 protein level, mitochondrial ATP production, and SOD activity and decreased nitrites production. These effects were associated with enhanced oxidative phenotype, as evidenced by amplified oxidative fiber-specific markers expression, histochemical reaction for NADH enzyme, and muscular lipid content. In this study, we showed that melatonin might have potential therapeutic implications for obesity-induced SKM metabolic inflexibility among patients with obesity and T2DMPID2021-125900OB-I00 from the Ministerio de Ciencia e Innovación (Spain)European Regional Development Fund (ERDF)B-CTS102-UGR from European Regional Development Fund (ERDF

Melatonin Improves Mitochondrial Dynamics and Function in the Kidney of Zücker Diabetic Fatty Rats

Obesity and associated diabetes (diabesity) impair kidney mitochondrial dynamics by augmenting fission and diminishing fusion, which results in mitochondrial and renal dysfunction. Based on available evidence, the antioxidant activities of melatonin may improve impaired renal mitochondrial function in obese diabetic animals by restoring the imbalanced dynamics through inhibiting fission and promoting fusion. Male Zücker diabetic fatty (ZDF) rats and lean littermates (ZL) were orally treated either with melatonin (10 mg/kg BW/day) (M-ZDF and M-ZL) or vehicle (C-ZDF and C-ZL) for 17 weeks. Kidney function was evaluated by measurement of total urine volume, proteinuria, creatinine clearance, and assessment of kidney mitochondrial dynamics and function. C-ZDF exhibited impaired dynamics and function of kidney mitochondria in comparison to C-ZL. Melatonin improved nephropathy of ZDF rats and modulated their mitochondrial dynamics by reducing expression of Drp1 fission marker and increasing that of fusion markers, Mfn2 and Opa1. Furthermore, melatonin ameliorated mitochondrial dysfunction by increasing respiratory control index and electron transfer chain complex IV activity. In addition, it lowered mitochondrial oxidative status. Our findings show that melatonin supplementation improves nephropathy likely via modulation of the mitochondrial fission/fusion balance and function in ZDF rats.Ministerio de Economia y Competitividad (Spain) SAF2016-79794-

Withania coagulans Fruit Extract Reduce Oxidative Stress and Inflammation in Kidneys of Streptozotocin-Induced Diabetic Rats

The present study was carried out to investigate the changes in oxidative and inflammatory status in streptozotocin-induced diabetic rat’s kidneys and serum following treatment with Withania coagulans, a popular herb of ethnomedicinal significance. The key markers of oxidative stress and inflammation such as inflammatory cytokines (IL-1β, IL-6, and TNF-α) and immunoregulatory cytokines (IL-4 and IFN-γ) were increased in kidneys along with significant hyperglycemia. However, treatment of four-month diabetic rats with Withania coagulans (10 mg/kg) for 3 weeks significantly attenuated hyperglycemia and reduced the levels of proinflammatory cytokines in kidneys. In addition, Withania coagulans treatment restored the glutathione levels and inhibited lipid peroxidation along with marked reduction in kidney hypertrophy. The present study demonstrates that Withania coagulans corrects hyperglycemia and maintained antioxidant status and reduced the proinflammatory markers in kidneys, which may subsequently reduce the development and progression of renal injury in diabetes. The results of the present study are encouraging for its potential use to delay the onset and progression of diabetic renal complications. However, the translation of therapeutic efficacy in humans requires further studies.This study was supported by grants from College of Medicine & Health Sciences, UAE University, UAE

Melatonin induces fat browning by transdifferentiation of white adipocytes and de novo differentiation of mesenchymal stem cells

This research was partially supported by grant SAF2016-79794R from the Ministerio de Ciencia e Innovacion (Spain), European Regional Development Fund (ERDF), University of Granada and FEDER Funds grant number B-CTS-102-UGR20. The authors thank Antonio Tirado for their technical assistance.The role of melatonin in obesity control is extensively accepted, but its mechanism of action is still unclear. Previously we demonstrated that chronic oral melatonin acts as a brown-fat inducer, driving subcutaneous white adipose tissue (sWAT) into a brown-fat-like function (beige) in obese diabetic rats. However, immunofluorescence characterization of beige depots in sWAT and whether melatonin is a beige-fat inducer by de novo differentiation and/or transdifferentiation of white adipocytes are still undefined. Lean (ZL) and diabetic fatty (ZDF) Zücker rats were subdivided into two groups, control (C) and oral melatonin-supplemented (M, 10 mg kg−1 day−1) for 6 weeks. Mesenchymal stem cells (MSCs) were isolated from both rat inguinal fat and human lipoaspirates followed by adipogenesis assays with or without melatonin (50 nM for 12 h in a 24 h period, 12 h+/12 h−) mimicking the light/dark cycle. Immunofluorescence and western-blot assays showed the partial transdifferentiation of white adipocytes in both ZL and ZDF rats, with increasing thermogenic and beige markers, UCP1 and CITED1 and decreasing white adipocyte marker ASC-1 expression. In addition, melatonin increased UCP1, CITED1, and PGC1- α expression in differentiated adipocytes in both rats and humans. These results demonstrate that melatonin increases brown fat in obese diabetic rats by both adipocyte transdifferentiation and de novo differentiation. Furthermore, it promotes beige MSC adipogenesis in humans. This may contribute to the control of body weight attributed to melatonin and its metabolic benefits in human diabesity.Spanish Government SAF2016-79794REuropean Commission B-CTS-102-UGR20University of Granad