ceRNA crosstalk mediated by ncRNAs is a novel regulatory mechanism in fish sex determination and differentiation.

Competing endogenous RNAs (ceRNAs) are vital regulators of gene networks in mammals. The involvement of noncoding RNAs (ncRNAs) as ceRNA in genotypic sex determination (GSD) and environmental sex determination (ESD) in fish is unknown. The Chinese tongue sole, which has both GSD and ESD mechanisms, was used to map the dynamic expression pattern of ncRNAs and mRNA in gonads during sex determination and differentiation. Transcript expression patterns shift during the sex differentiation phase, and ceRNA modulation occurs through crosstalk of differentially expressed long ncRNAs (lncRNAs), circular RNAs (circRNAs), microRNAs (miRNAs), and sex-related genes in fish. Of note was the significant up-regulation of a circRNA from the sex-determining gene dmrt1 (circular RNA dmrt1) and a lncRNA, called AMSDT (which stands for associated with male sex differentiation of tongue sole) in Chinese tongue sole testis. These two ncRNAs both share the same miRNA response elements with gsdf, which has an up-regulated expression when they bind to miRNA cse-miR-196 and concurrent down-regulated female sex-related genes to facilitate testis differentiation. This is the first demonstration in fish that ceRNA crosstalk mediated by ncRNAs modulates sexual development and unveils a novel regulatory mechanism for sex determination and differentiation.

Environmentally-induced sex reversal in fish with chromosomal vs. polygenic sex determination.

Sex ratio depends on sex determination mechanisms and is a key demographic parameter determining population viability and resilience to natural and anthropogenic stressors. There is increasing evidence that the environment can alter sex ratio even in genetically sex-determined species (GSD), as elevated temperature can cause female-to-male sex reversal (neomales). Alarmingly, neomales are being discovered in natural populations of several fish, amphibian and reptile species worldwide. Understanding the basis of neomale development is important for conservation biology. Among GSD species, it is unknown whether those with chromosomal sex determination (CSD), the most common system, will better resist the influence of high temperature than those with polygenic sex determination (PSD). Here, we compared the effects of elevated temperature in two wild zebrafish strains, Nadia (NA) and Ekkwill (EKW), which have CSD with a ZZ/ZW system, against the AB laboratory strain, which has PSD. First, we uncovered novel sex genotypes and the results showed that, at control temperature, the masculinization rate roughly doubled with the addition of each Z chromosome, while some ZW and WW fish of the wild strains became neomales. Surprisingly, we found that at elevated temperatures WW fish were just as likely as ZW fish to become neomales and that all strains were equally susceptible to masculinization. These results demonstrate that the Z chromosome is not essential for male development and that the dose of W buffers masculinization at the control temperature but not at elevated temperature. Furthermore, at the elevated temperature the testes of neomales, but not of normal males, contained more spermatozoa than at the control temperature. Our results show in an unprecedented way that, in a global warming scenario, CSD species may not necessarily be better protected against the masculinizing effect of elevated temperature than PSD species, and reveal genotype-by-temperature interactions in male sex determination and spermatogenesis.

The influence of increased water temperature on the duration of spermatogenesis in a neotropical fish, Astyanax altiparanae (Characiformes, Characidae).

In view of the established climate change scenario and the consequent changes in global temperature, it is essential to study its effects on animal spermatogenesis. Therefore, the aim of this study was to verify the duration of spermatogenesis at different temperatures. For this purpose, 96 male and adult specimens of Astyanax altiparanae were kept in a closed circulation system with water temperature stabilized at 27 °C and 32 °C. Subsequently, the specimens received pulses of BrdU (bromodeoxyuridine) at a concentration of 100 mg/kg/day for 2 consecutive days, and the samples were collected daily for a period of 15 days. Their testes were removed, fixed, processed in historesin, and sectioned in 3 µm, submitted to hematoxylin/eosin staining and to bromodeoxyuridine immunodetection. Partial results of the optimum temperature experiments allowed the classification of A. altiparanae spermatogenic cells in Aund, Adiff, and type B spermatogonia, spermatocytes, spermatids, and spermatozoa. The duration of spermatogenesis was determined as approximately 6 days for animals at a temperature of 27 °C and 1 day for animals at 32 °C. The elevated temperature was also responsible for increasing cell proliferation, resulting in an increase in the number of spermatocytes, spermatids, spermatozoa, and cell death (cell pyknotic). The duration of spermatogenesis in A. altiparanae was directly affected by the elevated water temperature, causing a reduction in the estimated time of spermatogenesis.

Reproductive cycle of the tetra Astyanax bimaculatus (Characiformes: Characidae) collected in Amazonian streams.

A histological characterization of gonadal development in the tetra Astyanax bimaculatus was performed, aimed at determining its reproductive cycle in streams localized inside the Amazonian forest. Collections were carried out monthly from August 2017 to July 2018 at the Zoobotânica Foundation of Marabá, PA. Collected specimens were weighed and measured, and their gonads and liver were removed and weighed to calculate gonadosomatic and hepatosomatic indexes. Gonads were fixed and treated for routine histology for light microscopy. Materials were stained with toluidine blue and haematoxylin and eosin. The Amazonian A. bimaculatus species presented two reproductive periods in the year, one at the end of the winter season and another during the summer. Females showed an asynchronous development of their oocytes and only two reproductive phases of development were observed during the whole period 'developing' and 'spawning capable'. Males presented cystic spermatogenesis, with an anastomosing tubular testis containing spermatogonia spread along the germinal epithelium (unrestricted spermatogonial). These morphological characteristics are considered phylogenetically more primitive. Male specimens were observed to have five different phases during the period: immature, initial maturation, mid maturation, final maturation and regression. The huge fluctuations in Amazonian streams was observed, in which water volumes varied considerably across seasons, culminating even in total drought. In spite of this, A. bimaculatus could be found during all seasons, showing its impressive reproductive adaptation to its conditions.

Testis structure, spermatogonial niche and Sertoli cell efficiency in Neotropical fish.

Neotropical icthyofauna represents one of the most diverse and extreme ecosystems in the world. Likewise, reproduction showed enormous diversity with different reproductive systems, modes and behavior. On the other hand, information on Neotropical fish species, in particular on male reproductive physiology is restricted to few species. This mini-review aimed to compile the existing information on spermatogenesis of Neotropical teleosts focusing on testis structure, spermatogonial niche and Sertoli cell efficiency. The first topic covers the histological analysis of the testicular structure, showing a conserved testicular pattern in relation to the phylogenetic position: basal species present anastomosing tubular testis (e.g. Astyanax altiparanae, Conorhynchos conirostris, Pimelodus maculatus, Lophiosilurus alexandri, Rhinelepis aspera, among others), while derived teleosts showed lobular testis (e.g. Cichlasoma dimerus, Cichla kelberi, Odontesthes bonariensis, Synbranchus marmoratus and others). Next to testicular structure, existing data showed that type A undifferentiated spermatogonia (Aund) is differentially distributed among the Neotropical species. Aund can be restricted at the blind-end of the germinal compartment (O. bonariensis), or spread along the germinal epithelium (A. altiparanae), or even distributed along the germinal epithelium but concentrated at the blind-end (C. kelberi and C. intermedia). Moreover, recent studies in A. altiparanae have demonstrated that within the germinal compartment, Aund have a preferential distribution in areas neighboring the interstitial compartment - the spermatogonial niche. The proximity with the interstitium suggests that interstitial cells, such as Leydig cells, are important for Aund maintenance in the testis. Finally, this mini-review highlighted Sertoli cell efficiency, showing that a single Sertoli cell can support a higher number of germ cells (80-140 spermatids) in Neotropical species evaluated at the moment (e.g. A. altiparanae, Hoplias malabaricus, Poecilia reticulata, Serrasalmus spilopleura, C. intermedia). Overall, this review provided basic and functional information on spermatogenesis of Neotropical species. More studies in this field are necessary since Neotropical region is considered one of the hotspot regions to discovery new species providing, therefore, new opportunities to investigate spermatogenesis in fish.

Effects of GnRH and the dual regulatory actions of GnIH in the pituitary explants and brain slices of Astyanax altiparanae males.

The pituitary gonadotropins, Fsh (follicle-stimulating hormone) and Lh (luteinizing hormone), regulate testicular development and functions in all vertebrates. At the pituitary, different signaling systems regulate the synthesis and secretion of the gonadotropins, such as the hypothalamic neuropeptides GnRH (gonadotropin-releasing hormone) and GnIH (gonadotropin-inhibitory hormone). While GnRH exerts stimulatory roles, the actions of GnIH remain controversial for many teleost species. Therefore, the aim of this study was to evaluate the in vitro effects of chicken GnRH2 (cGnRH2) and zebrafish GnIH-3 (zGnIH-3) on the male gonadotropin and GnRH system expression using pituitary explants and brain slices from a neotropical species with economical and ecological relevance, Astyanax altiparanae. Our results showed that in males, cGnRH2 increased fshb and lhb mRNA levels in the pituitary explants. Interestingly, zGnIH-3 has no effect on basal gonadotropin expression, however zGnIH-3 decreased the cGnRH2-induced fshb and lhb transcripts in male pituitary explants. In the male brain slices, zGnIH-3 showed stimulatory effects, increasing gnrh2 mRNA levels. Overall, our results suggested that GnIH seems to have dual regulatory actions on gonadotropin and GnRH2 expression of A. altiparanae males. This study provided basic information on endocrine regulation of A. altiparanae reproduction, and the obtained results will expand our knowledge, improving the reproductive management of this economically important freshwater species.

Thyroid hormone actions on male reproductive system of teleost fish.

Thyroid hormones (THs) play important roles in the regulation of many biological processes of vertebrates, such as growth, metabolism, morphogenesis and reproduction. An increasing number of studies have been focused on the involvement of THs in the male reproductive system of vertebrates, in particular of fish. Therefore, this mini-review aims to summarize the main findings on THs role in male reproductive system of fish, focusing on sex differentiation, testicular development and spermatogenesis. The existing data in the literature have demonstrated that THs exert their roles at the different levels of the hypothalamic-pituitary-gonadal (HPG) axis. In general a positive correlation has been shown between THs and fish reproductive status; where THs are associated with testicular development, growth and maturation. Recently, the molecular mechanisms underlying the role of THs in spermatogenesis have been unraveled in zebrafish testis. THs promote germ cell proliferation and differentiation by increasing a stimulatory growth factor of spermatogenesis produced by Sertoli cells. In addition, THs enhanced the gonadotropin-induced androgen release in zebrafish testis. Next to their functions in the adult testis, THs are involved in the gonadal sex differentiation through modulating sex-related gene expression, and testicular development via regulation of Sertoli cell proliferation. In conclusion, this mini-review showed that THs modulate the male reproductive system during the different life stages of fish. The physiological and molecular mechanisms showed a link between the thyroid and reproduction, suggesting a possibly co-evolution and interdependence of these two systems.

Characterization of gonadotropic cells during continuous and seasonal spermatogenesis of two freshwater fish species: a histochemical and immunohistochemical study.

This work describes gonadotropic (GtH) cells and their morphological and immunohistochemical changes during the spermatogenic cycle of Serrasalmus maculatus (continuous spermatogenesis) and Pimelodus maculatus (seasonal spermatogenesis). GtH cells, widely distributed in the proximal pars distalis of the adenohypophysis, were characterized as round-shaped cells with eccentric nucleus, and cytoplasm with basophilic secretory granules and a variable number of vacuoles for both species. Immunohistochemistry against ß-follicle-stimulating hormone (Fsh) and ß-luteinizing hormone (Lh) in adjacent sections showed two separated GtH-producing cell populations, and a third population where both GtHs are expressed in the same cell for both species. In the seasonal spermatogenesis of P. maculatus, GtH cells seemed to be more abundant during developing and spawning capable phases. In contrast, no cyclic changes were detected in the continuous spermatogenesis of S. maculatus, except for the strong immunoreaction for Fsh and Lh in males with intense spermiogenesis. We conclude that changes reported here might reflect the type of spermatogenic cycle (seasonal or continuous) which are under different regulatory mechanisms (environmental and internal cues) controlling the reproduction in these species.

Growth performance, reproductive status, and chromosomal instability in triploid Nile tilapias.

Reproductive control is one of the biggest challenges in tilapia production and triploidy was developed as an alternative to sterilization. In general, polyploids present chromosomal instability but for triploid Nile tilapia it has yet to be reported. This study evaluated the chromosomal instability from juveniles to adulthood, growth performance and gonadal status of tilapia hatched from eggs submitted or not to heat shock for triploid induction. Nile tilapia oocytes were fertilized (1,476 oocytes), half of the eggs were subjected to a four-minute shock in 41 °C water four minutes after fertilization and the other half were not (Control group). The eggs were incubated (at 27°C) and 160 larvae from the treated group hatched and survived after yolk sac absorption. The determination of ploidy was performed by flow cytometry at 85th (juveniles) and 301st (adults) days of age post yolk sac absorption. At the time of the first cytometry analysis there were 73 surviving juveniles from the treated group, and only 14 were confirmed triploid. However, at the analysis of adult ploidy, one out of 8 surviving adult tilapias from the 14 confirmed triploid juveniles remained triploid. Gonadal histology showed that the non-remaining triploids continued to produce gametes. The growth performance of triploid tilapia was initially superior to that of diploid tilapia during the juvenile phase, but similar in adults. Once the chromosome sets are lost and the tilapias become diploid again, at least in tissues with a high proliferation rate, such as the hematopoietic tissue that was analyzed (and possibly in gonads), all possible advantages of triploids are probably lost. Thus, our results suggest that, due to genomic instabilities, the triploid generation of tilapia has low efficiency.

Time-efficient germ cell transplantation from goldfish (Carassius auratus) into adult common carp (Cyprinus carpio).

Germ cell transplantation in fish is a promising technique for surrogate broodstock parents with broader application in aquaculture and conserving endangered and valuable genetic resources. Herein, we describe the establishment of an intrapapillary xenogeneic transplant of germ cells from sexually mature goldfish (C. auratus) males into common carp (C. carpio) males cytoablated with a thermochemical treatment (two doses of busulfan at 40 mg/kg at 35°C). To analyze the presence and development of donor germ cells in recipient testes, donor germ cells were labeled with PKH26, a fluorescent cell membrane dye, before transplantation. Our results demonstrated that thermochemical treatment caused effective spermatogenesis suppression and pronounced germ cell loss. Moreover, transplanted spermatogonial cells were able to colonize the recipients' testes, resume spermatogenesis, and generate spermatozoa within eight weeks after germ cell transplantation. These findings suggested that recipient testes provided suitable conditions for the survival, colonization, proliferation, and differentiation of donor spermatogonia from a related species. This study indicated that recipients' testes exhibited a high degree of plasticity to accept and support xenogeneic donor germ cells, which were able to form sperm in a short time frame. This approach has significant implications for assisted animal reproduction, biotechnology, conservation, and the production of valuable genetic resources and endangered fish species.

Simulated climate change and atrazine contamination can synergistically impair zebrafish testicular function.

Elements that interfere with reproductive processes can have profound impacts on population and the equilibrium of ecosystems. Global warming represents the major environmental challenge of the 21st century, as it will affect all forms of life in the coming decades. Another coexisting concern is the persistent pollution by pesticides, particularly the herbicide Atrazine (ATZ), which is responsible for a significant number of contamination incidents in surface waters worldwide. While it is hypothesized that climate changes will significantly enhance the toxic effects of pesticides, the actual impact of these phenomena remain largely unexplored. Here, we conducted a climate-controlled room experiment to assess the interactive effects of the projected 2100 climate scenario and environmentally realistic ATZ exposures on the reproductive function of male zebrafish. The gonadosomatic index significantly decreased in fish kept in the extreme scenario. Cellular alterations across spermatogenesis phases led to synergic decreased sperm production and increased germ cell sloughing and death. ATZ exposure alone or combined with climate change effects, disrupted the transcription levels of key genes involved in steroidogenesis, hormone signaling and spermatogenesis regulation. An additive modulation with decreased 11-KT production and increased E2 levels was also evidenced, intensifying the effects of androgen/estrogen imbalance. Moreover, climate change and ATZ independently induced oxidative stress, upregulation of proapoptotic gene and DNA damage in post-meiotic germ cell, but the negative effects of ATZ were greater at extreme scenario. Ultimately, exposure to simulated climate changes severely impaired fertilization capacity, due to a drastic reduction in sperm motility and/or viability. These findings indicate that the future climate conditions have the potential to considerably enhance the toxicity of ATZ at low concentrations, leading to significant deleterious consequences for fish reproductive function and fertility. These may provide relevant information to supporting healthcare and environmental managers in decision-making related to climate changes and herbicide regulation.

Shedding light on the impacts of gestational exposure to polystyrene nanoplastics on the reproductive performance of Poecilia reticulata female and on the biochemical response of embryos.

Although the toxicity of nanoplastics (NPs) has already been reported in experimental aquatic models, their possible effects on the reproductive performance of viviparous freshwater fish and their consequences for embryos, so far, are unknown. Thus, we aimed to evaluate whether the gestational exposure of Poecilia reticulata to polystyrene NPs (PS NPs) impacts the reproductive performance of females, induces teratogenic effects and/or predictive alterations of redox unbalance and cholinesterasic effect. Our results demonstrate that gestational exposure of P. reticulata females (for 30 days) to PS NPs (50 µg/L) affected reproductive aspects of the animals, inferred by the lower percentage of pregnancy and reduced offspring quantity. Although we did not observe teratogenic effect, we observed that the accumulation of PS NPs in embryos was significantly correlated with a redox unbalance, without, however, having a cholinesterasic effect (via evaluation of AChE and BChE activity) in embryos. Thus, by evidencing the accumulation of PS NPs in embryos of P. reticulata females exposed to the pollutant during the gestational period, we confirm not only the plausibility of the maternal transfer of these nanomaterials, but also their consequent physiological impacts on the offspring, which has not yet been demonstrated in live-bearing freshwater fish.

Cylindrospermopsin Disrupts Estrous Cycle and Increases Spermatogenesis in Mice.

Cylindrospermopsin (CYN) is a water-soluble cyanotoxin that has been linked to several cases of poisoning in the world. In vitro studies have shown that CYN acts as an endocrine disruptor by inhibiting progesterone synthesis in primary cell cultures of women, showing estrogenic activity. However, in vivo assessment of CYN in the female and male reproductive systems remains unknown. We thus aimed to evaluate the in vivo effects of CYN in both the female and male reproductive systems of mice. A single intraperitoneal exposure to 64 µg of CYN/kg body weight was performed in females. Estrous cycle was evaluated daily by vaginal cytology, and serum progesterone and estradiol levels were measured after 50 days. We showed an impairment in the estrous cycle as well as a decrease in circulating plasma progesterone levels. In males, weekly intraperitoneal doses of 20 µg of CYN/kg body weight were given and groups were killed after one, two, or four doses. CYN increased the testosterone levels in the groups that received one or two doses of CYN. Additionally, CYN induced a transient increase in spermatozoa in males after four doses. Our results highlight that CYN interferes with both male and female reproductive systems and may lead to infertility. As far as we know, this is the first report showing the impacts of CYN on the mammalian reproductive system, suggesting a threat from this cyanotoxin to human and environmental health.

Cannabidiol improves Nile tilapia cichlid fish welfare.

Cannabidiol (CBD) is a substance derived from Cannabis sativa, widely studied in medicine for controlling neural diseases in humans. Besides the positive effects on humans, it also presents anxiolytic proprieties and decreases aggressiveness and stress in mammals. Therefore, CBD has the potential to increase welfare in reared animals, as it seems to reduce negative states commonly experienced in artificial environments. Here, we tested the effect of different CBD doses (0, 1, 10 and 20 mg/kg) on aggressiveness, stress and reproductive development of the Nile tilapia (Oreochromis niloticus) a fish reared worldwide for farming and research purposes. CBD mixed with fish food was offered to isolated fish for 5 weeks. The 10 mg/kg dose decreased fish's aggressiveness over time, whereas 20 mg/kg attenuated non-social stress. Both doses decreased the baseline cortisol level of fish and increased the gonadosomatic index. However, CBD 1 and 10 mg/kg doses decreased the spermatozoa number. No CBD dose affected feeding ingestion and growth variables, showing that it is not harmful to meat production amount. Despite the effect on spermatozoa, CBD supplementation exhibits high potential to benefit animals' lives in artificial environments. Therefore, we showed for the first time that CBD could be used as a tool to increase non-mammal welfare, presenting a great potential to be explored in other husbandry and captivity species.

Can carbon nanofibers affect anurofauna? Study involving neotropical Physalaemus cuvieri (Fitzinger, 1826) tadpoles.

Although carbon nanotubes' (CNTs) toxicity in different experimental systems (in vivo and in vitro) is known, little is known about the toxic effects of carbon nanofibers (CNFs) on aquatic vertebrates. We herein investigated the potential impact of CNFs (1 and 10 mg/L) by using Physalaemus cuvieri tadpoles as experimental model. CNFs were able to induce nutritional deficit in animals after 48-h exposure to them, and this finding was inferred by reductions observed in body concentrations of total soluble carbohydrates, total proteins, and triglycerides. The increased production of hydrogen peroxide, reactive oxygen species and thiobarbituric acid reactive substances in tadpoles exposed to CNFs has suggested REDOX homeostasis change into oxidative stress. This process was correlated to the largest number of apoptotic and necrotic cells in the blood of these animals. On the other hand, the increased superoxide dismutase and catalase activity has suggested that the antioxidant system of animals exposed to CNFs was not enough to maintain REDOX balance. In addition, CNFs induced increase in acetylcholinesterase and butyrylcholinesterase activity, as well as changes in the number of neuromasts evaluated on body surface (which is indicative of the neurotoxic effect of nanomaterials on the assessed model system). To the best of our knowledge, this is the first report on the impact of CNFs on amphibians; therefore, it broadened our understanding about ecotoxicological risks associated with their dispersion in freshwater ecosystems and possible contribution to the decline in the populations of anurofauna species.

The Use of Menthol in Skin Wound Healing-Anti-Inflammatory Potential, Antioxidant Defense System Stimulation and Increased Epithelialization.

Wound healing involves inflammatory, proliferative, and remodeling phases, in which various cells and chemical intermediates are involved. This study aimed to investigate the skin wound healing potential of menthol, as well as the mechanisms involved in its effect, after 3, 7, or 14 days of treatment, according to the phases of wound healing. Skin wound was performed in the back of Wistar rats, which were topically treated with vehicle cream; collagenase-based cream (1.2 U/g); or menthol-based cream at 0.25%, 0.5%, or 1.0% over 3, 7, or 14 days. Menthol cream at 0.5% accelerated the healing right from the inflammatory phase (3 days) by decreasing mRNA expression of inflammatory cytokines TNF-α and Il-6. At the proliferative phase (7 days), menthol 0.5% increased the activity of antioxidant enzymes SOD, GR, and GPx, as well as the level of GSH, in addition to decreasing the levels of inflammatory cytokines TNF-α, IL-6, and IL-1ß and augmenting mRNA expression for Ki-67, a marker of cellular proliferation. At the remodeling phase (14 days), levels of inflammatory cytokines were decreased, and the level of Il-10 and its mRNA expression were increased in the menthol 0.5% group. Menthol presented skin wound healing activity by modulating the antioxidant system of the cells and the inflammatory response, in addition to stimulating epithelialization.

Environmental impacts of COVID-19 treatment: Toxicological evaluation of azithromycin and hydroxychloroquine in adult zebrafish.

One of the most impact issues in recent years refers to the COVID-19 pandemic, the consequences of which thousands of deaths recorded worldwide, are still inferior understood. Its impacts on the environment and aquatic biota constitute a fertile field of investigation. Thus, to predict the impact of the indiscriminate use of azithromycin (AZT) and hydroxychloroquine (HCQ) in this pandemic context, we aim to assess their toxicological risks when isolated or in combination, using zebrafish (Danio rerio) as a model system. In summary, we observed that 72 h of exposure to AZT and HCQ (alone or in binary combination, both at 2.5 µg/L) induced the reduction of total protein levels, accompanied by increased levels of thiobarbituric acid reactive substances, hydrogen peroxide, reactive oxygen species and nitrite, suggesting a REDOX imbalance and possible oxidative stress. Molecular docking analysis further supported this data by demonstrating a strong affinity of AZT and HCQ with their potential antioxidant targets (catalase and superoxide dismutase). In the protein-protein interaction network analysis, AZT showed a putative interaction with different cytochrome P450 molecules, while HCQ demonstrated interaction with caspase-3. The functional enrichment analysis also demonstrated diverse biological processes and molecular mechanisms related to the maintenance of REDOX homeostasis. Moreover, we also demonstrated an increase in the AChE activity followed by a reduction in the neuromasts of the head when zebrafish were exposed to the mixture AZT + HCQ. These data suggest a neurotoxic effect of the drugs. Altogether, our study demonstrated that short exposure to AZT, HCQ or their mixture induced physiological alterations in adult zebrafish. These effects can compromise the health of these animals, suggesting that the increase of AZT and HCQ due to COVID-19 pandemic can negatively impact freshwater ecosystems.

Multiple endpoints of polylactic acid biomicroplastic toxicity in adult zebrafish (Danio rerio).

Although the toxicity of conventional microplastic types (i.e., petroleum derivatives) in different organisms is already known, knowledge about the effects of alternative biopolymers on aquatic vertebrates remains incipient. Thus, the aim of the present study is to test the hypothesis that the exposure of adult Danio rerio individuals to this pollutant for 30 days is enough to cause polylactic acid biomicroplastics (BioMPs of PLA) accumulation in their bodies, which leads to behavioral/neurotoxic, biochemical, and morphological changes. Based on our results, PLA BioMPs at concentrations of 2.5 and 5 mg/L accumulated in the liver, brain, gills and carcass of the assessed animals. However, such an accumulation was not able to cause locomotor damages or to trigger anxiety-like behavior in them. On the other hand, it was enough to cause behavioral changes (in shoal) predictive of co-specific social interaction and anti-predatory defensive response deficit likely related to cholinergic changes inferred by increased acetylcholinesterase activity and REDOX imbalance. This imbalance was featured by increased production of reactive species. We observed that the treatments have affected animals' pigmentation pattern. Therefore, our study highlights the toxicological potential of the herein assessed biopolymer, and this finding puts in check the innocuousness of this material, as well as expands our knowledge about how PLA BioMPs can affect the ichthyofauna in freshwater environments.

Spermatogenesis in fish.

Spermatogenesis is a developmental process during which a small number of diploid spermatogonial stem cells produce a large number of highly differentiated spermatozoa carrying a haploid, recombined genome. We characterise morphologically the different germ cell stages with particular attention for the spermatogonial generations, including the stem cells and their specific capacity to colonise a recipient's testis after transplantation. We propose a nomenclature for fish germ cells to improve the comparability among different teleost fish but also to higher vertebrates. Survival and development of germ cells depends on their continuous and close contact to Sertoli cells, and we review their multiple roles in the cystic mode of spermatogenesis seen in fish. We then discuss gene expression patterns associated with testis maturation. The endocrine system of vertebrates has evolved as master control system over spermatogenesis. In fish, both pituitary gonadotropins LH and FSH stimulate gonadal sex steroid hormone production directly by activating Leydig cells. Information is reviewed on the effects of progestin, androgens, and estrogens on global testicular gene expression patterns (microarray analysis), and on the molecular mechanisms by which steroids regulate specific candidate genes (identified by subtractive hybridization approaches) during early stages of testis maturation. Moreover, progestin and androgen effects on spermiation and milt hydration are discussed. Sex steroids mainly act via receptors expressed by Sertoli cells. One type of response is that Sertoli cells change growth factor expression, which subsequently modulates germ cell proliferation/differentiation via mechanisms yet to be characterised. Finally, we review data on germ cell autonomous processes, mainly derived from loss-of-function mutant fish lines, before identifying a number of focus areas for future research activities.

Corrigendum to "Lupeol, a Dietary Triterpene, Enhances Wound Healing in Streptozotocin-Induced Hyperglycemic Rats with Modulatory Effects on Inflammation, Oxidative Stress, and Angiogenesis".

[This corrects the article DOI: 10.1155/2019/3182627.].