Ultrastructural observations on the lung of Pelophylax kl. esculentus tadpoles during development

The anatomical organization of the lung in Pelophylax kl. esculentus tadpoles has been described in early and middle-staged tadpoles (Gosner stages 32 and 39). The lung's ontogenic development has been studied using light microscopy and both scanning and transmission electron microscopy. In the early-staged tadpoles, the lung is well vascularized, but it is provided with only two internal septa types and devoid of ciliated epithelium and lamellar bodies. The complete differentiation process is putatively reached in the middle-staged tadpoles (GS 39), and numerous septa of first, second, and third-order deeply protruding into the lung lumen could be recognized. The histological and ultrastructural features at this stage correspond to that described in the adults' lungs, but mucous secreting goblet cells could not be detected. Pneumocytes, with numerous apical microvilli, surround the growing network of capillaries and show a cytoplasm rich in dense bodies and few mature lamellar bodies. On the epithelial surface, a thin layer of mucous covering the underlying epithelium could also be seen. Neuroepithelial bodies, supposed to be involved in chemoreception, are organized in clusters on the first order septa and surrounded by cytoplasmatic processes originating by neighboring cells. The lung arrangement of P. kl. esculentus is compared with that of other Anurans in order to further elucidate the putative role of the lung in gas exchanges. The present studies reduce the information gap existing in the literature regarding lung morphology and development in amphibian larval stages, also contributing to the discussion on the onset of airbreathing respiration in tadpoles

Tissues injury and pathological changes in Hyla intermedia juveniles after chronic larval exposure to tebuconazole.

Abstract Tebuconazole (TBZ), an azole pesticide, is one of the most frequently detected fungicides in surface water. Despite its harmful effects, mainly related to endocrine disturbance, the consequences of TBZ exposure in amphibians remain poorly understood. Here, we investigated the adverse and delayed effects of TBZ chronic exposure on a native anuran species, often inhabiting cultivated areas, the Italian tree frog (Hyla intermedia). To disclose the multiple mechanisms of action through which TBZ exerts its toxicity we exposed tadpoles over the whole larval period to two sublethal TBZ concentrations (5 and 50 μg/L), and we evaluated histological alterations in three target organs highly susceptible to xenobiotics: liver, kidney, and gonads. We also assessed morphometric and gravimetric parameters: snout-vent length (SVL), body mass (BM), liver somatic index (LSI), and gonad-mesonephros complex index (GMCI) and determined sex ratio, gonadal development, and differentiation. Our results show that TBZ induces irreversible effects on multiple target organs in H. intermedia, exerting its harmful effects through several pathological pathways, including a massive inflammatory response. Moreover, TBZ markedly affects sexual differentiation also by inducing the appearance of sexually undetermined individuals and a general delay of germ cell maturation. Given the paucity of data on the effects of TBZ in amphibians, our results will contribute to a better understanding of the environmental risk posed by this fungicide to the most endangered group of vertebrates

Morphological and Functional Alterations in Zebrafish (Danio rerio) Liver after Exposure to Two Ecologically Relevant Concentrations of Lead

Lead (Pb) is a non-essential, highly toxic, and persistent element widely recognized as one of the most concerning pollutants. It is listed on the Priority List of Hazardous Substances. Widespread environmental contamination from Pb is a serious issue for human health and wildlife. In fish, Pb mainly accumulates in the liver, which is a key component for metal detoxification and excretion processes. In this study, we investigated, for the first time, the morphological and functional injuries induced in zebrafish (Danio rerio) liver by two very low and environmentally relevant concentrations of Pb (2.5 and 5 μg/L) after 48, 96, and 192 h of exposure. We observed significant histological alterations in all the exposed samples, and it was demonstrated that the extent of injuries increased with dose and exposure time. The most common modifications observed were congestion of blood vessels and sinusoids, cytoplasmic vacuolizations, parenchyma dyschromia, and macrophage proliferation. Pb administration also resulted in a significant increase in lipid content and the upregulation of key genes that are involved in metal detoxification (mtf1) and the defensive response against oxidative stress (sod1 and cat). We show that even very low doses of Pb can disrupt liver morphology and function.Fil: Macirella, Rachele. Università della Calabria; ItaliaFil: Curcio, Vittoria. Università della Calabria; ItaliaFil: Ahmed, Abdalmoiz I. M.. Università della Calabria; ItaliaFil: Talarico, Federica. Università della Calabria; ItaliaFil: Sesti, Settimio. Università della Calabria; ItaliaFil: Paravani, Enrique Valentin. Universidad Nacional de Entre Ríos. Facultad de Ingeniería; ArgentinaFil: Odetti, Lucia Magdalena. Universidad Nacional del Litoral. Facultad de Bioquímica y Ciencias Biológicas. Cátedra de Toxicología y Bioquímica Legal; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe; ArgentinaFil: Mezzasalma, Marcello. Università della Calabria; ItaliaFil: Brunelli, Elvira. Università della Calabria; Itali

Morpho-Functional Alterations in the Gills of a Seawater Teleost, the Ornate Wrasse (Thalassoma pavo L.), after Short-Term Exposure to Chlorpyrifos

Chlorpyrifos (CPF) is an organophosphorus insecticide commonly used for domestic and agricultural purposes. The risk posed by environmental contamination from CPF is well acknowledged, and it has been detected worldwide in aquatic habitats and coastal areas. In addition, due to its slower degradation in seawater compared to freshwater, CPF is of particular concern for marine environments. Here, we investigated for the first time the morpho-functional alterations induced by CPF on the gills of Thalassoma pavo, a widespread species in the Mediterranean Sea. We tested the effects of two sublethal concentrations (4 and 8 µg/L) after 48 and 96 h. Our study demonstrates that the alterations induced by CPF are dose and time-dependent and highlight the harmful properties of this insecticide. After exposure to the low tested concentration, the more frequent alteration is an intense proliferation of the primary epithelium, whereas after exposure to the high concentration, the primary epithelium proliferation is less extensive, and the most evident effects are the thinning of secondary lamellae and the ectopia of chloride and goblet cells. CPF also modulated the expression of Na+/K+-ATPase. Dilation of lamellar apical tips, pillar cell degeneration, and appearance of aneurysms are often observed

The differential role of Leydig cells in the skin and gills of Lissotriton italicus larvae

Larval urodeles are provided with external gills involved, along with the skin, in gas exchange and osmoregulation. Gills and skin epithelia are different, each showing a peculiar set of specialized cells but both provided with Leydig cells (LCs). Information on LCs in the gills is lacking as the literature has focused primarily on the epidermis. Contradictory and fragmentary results highlight that LCs origin, fate, and functions remain not fully understood. Here, we investigated the morpho‐functional differences of LCs in the skin and gills of Lissotriton italicus larvae for the first time. LCs showed the same morphological and ultrastructural features in both tissues, even if LCs were significantly larger in the epidermis. Despite the uniform morphology within the LCs population, the proliferative ability was different. The putative diversity in the mucus composition was evaluated using a panel of 4 lectins as markers of specific carbohydrate moieties, revealing that sites of specific glycoconjugates were comparable in two tissues. To disclose the involvement of LCs in water storage and transport, immunofluorescence assay for aquaporin‐3 has also been performed, demonstrating the expression of this protein only in gills epithelium. By demonstrating that LCs can multiply by cell division in gills, our results will also contribute to the discussion about their proliferative ability. Finally, we found that the LCs cytoplasm is rich in glycoconjugates, which are involved in many diverse and essential functions in vertebrates. RESEARCH HIGHLIGHTS: In gills LCs can multiply by cell division and express aquaporin‐3 demonstrating a tissue‐specific role of LCs. LCs cytoplasm is rich in glycoconjugates. LCs population show a uniform morphology in both gills and skin

Antioxidant/Anti-Inflammatory Effects of Caloric Restriction in an Aged and Obese Rat Model: The Role of Adiponectin

Caloric restriction (CR) represents a powerful intervention for extending healthspan and lifespan in several animal models, from yeast to primates. Additionally, in humans, CR has been found to induce cardiometabolic adaptations associated with improved health. In this study, we evaluated in an aged and obese rat model the effect of long-term (6 months) caloric restriction (−40%) on the oxidative/inflammatory balance in order to investigate the underlining mechanisms. In plasma, we analyzed the oxidative balance by photometric tests and the adiponectin/tumor necrosis factor-α-induced gene/protein 6 (TSG-6) levels by Western blot analysis. In the white adipose tissue, we examined the protein levels of AdipoR1, pAMPK, NFκB, NRF-2, and glutathione S-tranferase P1 by Western blot analysis. Our results clearly showed that caloric restriction significantly improves the plasmatic oxidative/inflammatory balance in parallel with a major increase in circulating adiponectin levels. Additionally, at the level of adipose tissue, we found a positive modulation of both anti-inflammatory and antioxidant pathways. These adaptations, induced by caloric restriction, with the achievement of normal weight, suggest that inflammatory and redox imbalance in obese aged rats appear to be more linked to obesity than to aging

Morphological and Molecular Alterations Induced by Lead in Embryos and Larvae of Danio rerio

Lead (Pb) is one of the most toxic and persistent elements and may adversely affect both humans and wildlife. Given the risks posed to humans, lead is listed among priority substances of public health importance worldwide. In fish, available studies deal with high doses, and the potential hazard of Pb at low concentrations is largely unknown. Given its well-demonstrated translational value for human toxicity research, we used zebrafish as a model species. Embryos were exposed to two environmentally relevant concentrations of lead (2.5 and 5 µg/L) from 6 h post-fertilization and analyzed after 48, 96, and 144 h. The morphological abnormality arose after 48 h, and the incidence and intensity were dose and time dependent. Spinal and tail deformities were the most frequently detected alterations. Pb also modulated the expression of genes involved in the toxicological responses (sod and mt), thus demonstrating that zebrafish’s early stages are able to mount an adaptive response. Moreover, ldh and β-catenin were significantly upregulated in all groups, whereas wnt3 expression was increased in the high concentration group. Our results confirm that zebrafish embryos and larvae are valuable early warning indicators of pollution and may play a major role in ecosystems and human health monitoring

Morphological and Functional Alterations Induced by Two Ecologically Relevant Concentrations of Lead on Danio rerio Gills

Lead (Pb), due to its high toxicity and bioaccumulation tendency, is one of the top three pollutants of concern for both humans and wildlife and occupies second place in the Priority List of Hazardous Substances. In freshwater fish, Pb is mainly absorbed through the gills, where the greatest accumulation occurs. Despite the crucial role of gills in several physiological functions such as gas exchange, water balance, and osmoregulation, no studies evaluated the effects of environmentally relevant concentrations of Pb on this organ, and existing literature only refers to high levels of exposure. Herein we investigated for the first time the molecular and morphological effects induced by two low and environmentally relevant concentrations of Pb (2.5 and 5 μg/L) on the gills of Danio rerio, a model species with a high translational value for human toxicity. It was demonstrated that Pb administration at even low doses induces osmoregulatory dysfunctions by affecting Na+/K+-ATPase and AQP3 expression. It was also shown that Pb upregulates MTs as a protective response to prevent cell damage. Modulation of SOD confirms that the production of reactive oxygen species is an important toxicity mechanism of Pb. Histological and morphometric analysis revealed conspicuous pathological changes, both dose- and time-dependent

Effects of Two Sublethal Concentrations of Mercury Chloride on the Morphology and Metallothionein Activity in the Liver of Zebrafish (Danio rerio)

Mercury (Hg) is a highly hazardous pollutant widely used in industrial, pharmaceutical and agricultural fields. Mercury is found in the environment in several forms, elemental, inorganic (iHg) and organic, all of which are toxic. Considering that the liver is the organ primarily involved in the regulation of metabolic pathways, homeostasis and detoxification we investigated the morphological and ultrastructural effects in Danio rerio liver after 96 h exposure to two low HgCl2 concentrations (7.7 and 38.5 μg/L). We showed that a short-term exposure to very low concentrations of iHg severely affects liver morphology and ultrastructure. The main effects recorded in this work were: cytoplasm vacuolization, decrease in both lipid droplets and glycogen granules, increase in number of mitochondria, increase of rough endoplasmic reticulum and pyknotic nuclei. Pathological alterations observed were dose dependent. Trough immunohistochemistry, in situ hybridization and real-time PCR analysis, the induction of metallothionein (MT) under stressor conditions was also evaluated. Some of observed alterations could be considered as a general response of tissue to heavy metals, whereas others (such as increased number of mitochondria and increase of RER) may be considered as an adaptive response to mercury

Bergamot Polyphenols Boost Therapeutic Effects of the Diet on Non-Alcoholic Steatohepatitis (NASH) Induced by “Junk Food”: Evidence for Anti-Inflammatory Activity

Wrong alimentary behaviors and so-called “junk food„ are a driving force for the rising incidence of non-alcoholic fatty liver disease (NAFLD) among children and adults. The “junk food„ toxicity can be studied in “cafeteria„ (CAF) diet animal model. Young rats exposed to CAF diet become obese and rapidly develop NAFLD. We have previously showed that bergamot (Citrus bergamia Risso et Poiteau) flavonoids, in the form of bergamot polyphenol fraction (BPF), effectively prevent CAF diet-induced NAFLD in rats. Here, we addressed if BPF can accelerate therapeutic effects of weight loss induced by a normocaloric standard chow (SC) diet. 21 rats fed with CAF diet for 16 weeks to induce NAFLD with inflammatory features (NASH) were divided into three groups. Two groups were switched to SC diet supplemented or not with BPF (CAF/SC±BPF), while one group continued with CAF diet (CAF/CAF) for 10 weeks. BPF had no effect on SC diet-induced weight loss, but it accelerated hepatic lipid droplets clearance and reduced blood triglycerides. Accordingly, BPF improved insulin sensitivity, but had little effect on leptin levels. Interestingly, the inflammatory parameters were still elevated in CAF/SC livers compared to CAF/CAF group after 10 weeks of dietary intervention, despite over 90% hepatic fat reduction. In contrast, BPF supplementation decreased hepatic inflammation by reducing interleukin 6 (Il6) mRNA expression and increasing anti-inflammatory Il10, which correlated with fewer Kupffer cells and lower inflammatory foci score in CAF/SC+BPF livers compared to CAF/SC group. These data indicate that BPF mediates a specific anti-inflammatory activity in livers recovering from NASH, while it boosts lipid-lowering and anti-diabetic effects of the dietary intervention