Structural and immunohistochemical characterization of pancreas of Molly fish (Poecilia sphenops), with a special reference to its immune role.

Recently, teleost species have been considered important model systems for investigating different research areas including immunologic one. The available literature provides poor data about the localization and the structure of pancreas in Molly fish. Moreover, little attention has been paid to the immunologic role of pancreatic tissue of teleost, particularly Molly fish; therefore, this study aimed to highlights the description of pancreatic tissue in Molly fish using light- and electron- microscopy, focusing on the role of pancreatic immune cells and pancreatic acinar cells in immune responses. Microscopic analysis revealed that the pancreas of Molly fish was composed of intrahepatic, disseminated and compact parts. Exocrine pancreatic tissue was diffusely extended within the hepatic tissue forming hepatopancreas. The disseminated pancreas appeared as several irregular nodules of pancreatic tissue localized within the mesenteric adipose tissue. The compact pancreas appeared as an oval shaped body embedded within the mesenteric adipose tissue between the spleen and the intestinal loops. Several telocytes and melanomacrophages were detected within the disseminated pancreatic nodules. Moreover, dendritic cells were found in a close association to the exocrine pancreatic acini. The pancreatic acinar cells showed strong immunoreactivity to APG5, TGF-ß, IL-1ß, NF-κB, Nrf2, and SOX9 in both hepatopancreas and disseminated pancreas of Molly fish. S100 protein revealed a strong expression in the exocrine pancreatic acinar cells of disseminated pancreas and also in the endocrine cells of the compact pancreas. In conclusion, findings of this study suggest the potential role of the pancreas of the Molly fish in cell proliferation and differentiation, proinflammatory cytokines stimulation, and regulation of both innate and adaptive immunity. RESEARCH HIGHLIGHTS: Telocytes and melanomacrophages were detected in the disseminated pancreatic nodules of the Molly fish. In Molly fish, dendritic cells were found in a close association to the exocrine pancreatic acini. Strong immunoreactivity of the pancreatic acinar cells of the Molly fish to APG5, TGF-ß, IL-1ß, NF-κB, Nrf2, SOX9, and S100.

Structural and immunohistochemical analysis of the cellular compositions of the liver of molly fish (Poecilia sphenops), focusing on its immune role.

The liver of fish is considered an ideal model for studying the collaboration between environmental agents and the health state of the fish, where it gives good indications about aquatic ecosystem status. Therefore, this study presented immune roles for the liver in molly fish (Poecilia sphenops), using immunohistochemistry and transmission electron microscopy (TEM). The hepatocytes' sinusoidal structures of molly fish livers had taken two different forms; cord-like and tubular, while the biliary tract system showed two different types: isolated and biliary venous tract. The TEM showed that the hepatocytes possessed well-developed cytoplasmic organelles and numerous glycogen and lipid droplets of different sizes. Kupffer cells, Ito cells, aggregation of intrahepatic macrophages and melanomacrophages were also recognized. Melanomacrophages contained numerous phagosomes, many lysosomes, cytoplasmic vacuoles, and melanin pigments. Hepatocytes and Kupffer cells expressed immunoreactivity to APG5, indicating that these cells were involved in the process of autophagy. Telocytes (TCs) were also recognized in the liver of molly fish, and they shared the same morphological characteristics as those in mammals. However, TCs expressed strong immunoreactivity to APG5, TGF-ß, and Nrf2, suggesting their possible role in cellular differentiation and regeneration, in addition to phagocytosis and autophagy. Both IL-1ß and NF-KB showed immunoreactivity in the hepatocytes and in inflammatory cells (including intrahepatic macrophages and melanomacrophage center). Nrf2 and SOX9 showed immunoreactivity in hepatocytes, stem cells, and macrophages. The present study showed the spatial distribution of hepatic vascular-biliary tracts in molly fish. The liver of molly fish has unique functions in phagocytosis, autophagy, and cell regeneration. The expression of APG5 in hepatocytes, Kupffer cells, melanomacrophages, and telocytes supports the role of the liver in lymphocyte development and proliferation. The expression of TGF-ß and NF-κB in hepatocytes, Kupffer cells, telocytes, and macrophages suggests the role of the liver in regulation of cell proliferation and immune response suppression. The expression of IL-1ß and Sox9 in macrophages and melanomacrophages suggests the role of the liver in regulation of both innate and adaptive immunity, cell proliferation and apoptosis, in addition to stem cell maintenance.

Expression of Antimicrobic Peptide Piscidin1 in Gills Mast Cells of Giant Mudskipper Periophthalmodon schlosseri (Pallas, 1770).

The amphibious teleost Giant mudskipper (Periophthalmodon schlosseri, Pallas 1770) inhabit muddy plains and Asian mangrove forests. It spends more than 90% of its life outside of the water, using its skin, gills, and buccal-pharyngeal cavity mucosa to breathe in oxygen from the surrounding air. All vertebrates have been found to have mast cells (MCs), which are part of the innate immune system. These cells are mostly found in the mucous membranes of the organs that come in contact with the outside environment. According to their morphology, MCs have distinctive cytoplasmic granules that are released during the degranulation process. Additionally, these cells have antimicrobial peptides (AMPs) that fight a variety of infections. Piscidins, hepcidins, defensins, cathelicidins, and histonic peptides are examples of fish AMPs. Confocal microscopy was used in this study to assess Piscidin1 expression in Giant Mudskipper branchial MCs. Our results demonstrated the presence of MCs in the gills is highly positive for Piscidin1. Additionally, colocalized MCs labeled with TLR2/5-HT and Piscidin1/5-HT supported our data. The expression of Piscidin1 in giant mudskipper MCs highlights the involvement of this peptide in the orchestration of teleost immunity, advancing the knowledge of the defense system of this fish.

Structural and Functional Aspects of the Spleen in Molly Fish Poecilia sphenops (Valenciennes, 1846): Synergistic Interactions of Stem Cells, Neurons, and Immune Cells.

In fish, the spleen is the prime secondary lymphoid organ. It has a role in the induction of adaptive immune responses, in addition to its significance in the elimination of immune complexes. This study was conducted on 18 randomly obtained adult molly fish (Poecilia sphenops) of both sexes using histological, immunohistochemical, and ultrastructural studies to highlight the cellular components of the spleen and their potential role in the immune system. The spleen of molly fish was characterized by the presence of well-distinct melanomacrophage centers, and other basic structures present in higher vertebrates including red and white pulps, blood vessels, and ellipsoids. Some mitotic cells could also be identified in the red pulp. Mast cells with characteristic metachromatic granules could be seen among the splenic cells. Rodlet cells were randomly distributed in the spleen and were also observed around the ellipsoids. The white pulp of the spleen expressed APG5. The expressions were well distinct in the melanomacrophages, leukocytes, and macrophages. Myostatin was expressed in leukocytes and epithelial reticular cells. IL-1ß showed immunoreactivity in monocytes and macrophages around the ellipsoids. NF-κB and TGF-ß were expressed in macrophages and epithelial reticular cells. Nrf2 expression was detected in stem cells and rodlet cells. Sox-9 had a higher expression in epithelial reticular cells and stem cells. The high frequency of immune cells in the spleen confirmed its role in the regulation of both innate and adaptive immunity, cell proliferation, and apoptosis.

Melatonin alleviates sepsis-induced heart injury through activating the Nrf2 pathway and inhibiting the NLRP3 inflammasome.

Melatonin improved the outcome of septic cardiomyopathy by inhibiting NLRP3 priming induced by reactive oxygen species. To get insights into these events, we studied the melatonin/Nrf2 antioxidant pathways during sepsis in the heart of NLRP3-deficient mice. Sepsis was induced by cecal ligation and puncture and melatonin was given at a dose of 30 mg/kg. Nuclear turnover of Nrf2 and p-Ser40 Nrf2 and expression of ho-1 were enhanced in nlrp3+/+ and nlrp3-/- mice during sepsis. Sepsis caused higher mitochondria impairment, apoptotic and autophagic events in nlrp3+/+ mice than in nlrp3-/- animals. These findings were accompanied by greater levels of Parkin and PINK-1, and lower Mfn2/Drp-1 ratio in nlrp3+/+ than in nlrp3-/- mice during sepsis, supporting less mitophagy in the latter. Ultrastructural analysis of myocardial tissue further confirmed these observations. The activation of NLRP3 inflammasome accounted for most of the deleterious effects of sepsis, whereas the Nrf2-dependent antioxidative response activation in response to sepsis was unable to neutralize these events. In turn, melatonin further enhanced the Nrf2 response in both mice strains and reduced the NLRP3 inflammasome activation in nlrp3+/+ mice, restoring myocardial homeostasis. The data support that the anti-inflammatory efficacy of melatonin against sepsis depends, at least in part, on Nrf2 activation.

Lack of NLRP3 Inflammasome Activation Reduces Age-Dependent Sarcopenia and Mitochondrial Dysfunction, Favoring the Prophylactic Effect of Melatonin.

To investigate the role of NLRP3 inflammasome in muscular aging, we evaluated here the morphological and functional markers of sarcopenia in the NLRP3-knockout mice, as well as the beneficial effect of melatonin supplementation. The gastrocnemius muscles of young (3 months), early-aged (12 months), and old-aged (24 months) NLRP3-knockout female mice were examined. Moreover, locomotor activity and apoptosis were assessed. The results revealed early markers of sarcopenia at the age of 12 months, including reduction of lactate, ratio of muscle weight to body weight, muscle fibers number, and mitochondrial number. Increased interstitial tissues, apoptosis, and muscle fibers area, as well as mitochondrial damage were detected, with little muscular activity effects. In the old-aged, these alterations progressed with a reduction in locomotor activity, mitochondrial cristae destruction, nuclear fragmentation, tubular aggregates (TAs) formation, and increased frailty index. Oral melatonin supplementation preserved the normal muscular structure, muscle fibers number, and muscular activity in old age. Melatonin enhanced lactate production, recovered mitochondria, inhibited TAs formation, reduced apoptosis, and normalized frailty index. The fewer sarcopenic changes as well as the highly detectable prophylactic effects of melatonin treatment reported here in the muscle of NLRP3-knockout mice comparing with that previously detected in wild-type mice, confirming NLRP3 inflammasome implication in muscular aging and sarcopenia onset and progression.

Melatonin protects rats from radiotherapy-induced small intestine toxicity.

Radiotherapy-induced gut toxicity is among the most prevalent dose-limiting toxicities following radiotherapy. Prevention of radiation enteropathy requires protection of the small intestine. However, despite the prevalence and burden of this pathology, there are currently no effective treatments for radiotherapy-induced gut toxicity, and this pathology remains unclear. The present study aimed to investigate the changes induced in the rat small intestine after external irradiation of the tongue, and to explore the potential radio-protective effects of melatonin gel. Male Wistar rats were subjected to irradiation of their tongues with an X-Ray YXLON Y.Tu 320-D03 irradiator, receiving a dose of 7.5 Gy/day for 5 days. For 21 days post-irradiation, rats were treated with 45 mg/day melatonin gel or vehicle, by local application into their mouths. Our results showed that mitochondrial oxidative stress, bioenergetic impairment, and subsequent NLRP3 inflammasome activation were involved in the development of radiotherapy-induced gut toxicity. Oral treatment with melatonin gel had a protective effect in the small intestine, which was associated with mitochondrial protection and, consequently, with a reduced inflammatory response, blunting the NF-κB/NLRP3 inflammasome signaling activation. Thus, rats treated with melatonin gel showed reduced intestinal apoptosis, relieving mucosal dysfunction and facilitating intestinal mucosa recovery. Our findings suggest that oral treatment with melatonin gel may be a potential preventive therapy for radiotherapy-induced gut toxicity in cancer patients.

The benefit of a supplement with the antioxidant melatonin on redox status and muscle damage in resistance-trained athletes.

Previous data showed that the administration of high doses of melatonin improved the circadian system in athletes. Here, we investigated in the same experimental paradigm whether the antioxidant properties of melatonin has also beneficial effects against exercise-induced oxidative stress and muscle damage in athletes. Twenty-four athletes were treated with 100 mg·day-1 of melatonin or placebo 30 min before bedtime during 4 weeks in a randomized double-blind scheme. Exercise intensity was higher during the study that before starting it. Blood samples were collected before and after treatment, and plasma was used for oxygen radical absorption capacity (ORAC), lipid peroxidation (LPO), nitrite plus nitrate (NOx), and advanced oxidation protein products (AOPP) determinations. Glutathione (GSH), glutathione disulphide (GSSG) levels, and glutathione peroxidase (GPx) and reductase (GRd) activities, were measured in erythrocytes. Melatonin intake increased ORAC, reduced LPO and NOx levels, and prevented the increase of AOPP, compared to placebo group. Melatonin was also more efficient than placebo in reducing GSSG·GSH-1 and GPx·GRd-1 ratios. Melatonin, but not placebo, reduced creatine kinase, lactate dehydrogenase, creatinine, and total cholesterol levels. Overall, the data reflect a beneficial effect of melatonin treatment in resistance-training athletes, preventing extra- and intracellular oxidative stress induced by exercise, and yielding further skeletal muscle protection against exercise-induced oxidative damage.