Textile dyeing wastewater negatively influences the hematological profile and reproductive health of male Swiss albino mice

Objective:To determine the effects of textile dyeing industrial wastewater on the hematological parameters and reproductive health including histoarchitecture of male gonad(testes)of mice.Methods:Twenty-four Swiss albino mice at 4-weeks old were divided into four groups(n=6 per group).Mice of group 1 supplied with normal drinking water were served as the control group.Mice of group 2,3 and 4 were supplied normal drinking water mixed with textile dyeing wastewater at 5%,10% and 20% concentration,respectively.After completing 24 weeks of treatment,different hematological profile,weight of testes,gonadosomatic index(GSI),sperm concentration and morphology were measured.Moreover,histopathological changes in testes were examined.Results:Hematocrit value and hemoglobin concentrations were decreased in all groups of wastewater-treated mice compared to the control group.Likewise,weight of testes,GSI and sperm concentration were decreased significantly in wastewater-treated mice in comparison to the control group.The percentage of morphologically healthy epididymal sperm was significantly reduced in wastewater-treated mice.Histopathological examination revealed degenerative changes in seminiferous tubules,a smaller number of spermatogenic cells,elongation of seminiferous tubules and degenerative changes of seminiferous tubules in wastewater-treated mice.Conclusions:Textile dyeing wastewater has harmful effects on hematological profile and reproductive health of male mice.

Engineering of ovarian tissue for ovarian dysfunctions:A review

This review explores tissue engineering as a potential solution for reproductive health issues in women caused by genetic or acquired diseases,such as premature ovarian failure or oophorectomy.The loss of ovarian function can lead to infertility,osteoporosis,and cardiovascular disease.Hormone replacement therapy is a common treatment,but it has limitations and risks.The review focuses on two main approaches in tissue engineering:scaffold-based(3D printing,electrospinning,decellularization)and scaffold-free(stem cell transplantation,organoid cultivation).Both approaches show promise in preclinical studies for creating functional ovarian tissue.Challenges include vascularization,innervation,long-term function,and safety.Despite these challenges,tissue engineering offers a potential avenue for restoring fertility and hormone balance in women with ovarian dysfunction.

Sex-biased autophagy as a potential mechanism mediating sex differences in ischemic stroke outcome

Stroke is the second leading cause of death and a major cause of disability worldwide,and biological sex is an important determining factor in stroke incidence and pathology.From childhood through adulthood,men have a higher incidence of stroke compared with women.Abundant research has confirmed the beneficial effects of estrogen in experimental ischemic stroke but genetic factors such as the X-chromosome complement can also play an important role in determining sex differences in stroke.Autophagy is a self-degrading cellular process orchestrated by multiple core proteins,which leads to the engulfment of cytoplasmic material and degradation of cargo after autophagy vesicles fuse with lysosomes or endosomes.The levels and the activity of components of these signaling pathways and of autophagy-related proteins can be altered during ischemic insults.Ischemic stroke activates autophagy,however,whether inhibiting autophagy after stroke is beneficial in the brain is still under a debate.Autophagy is a potential mechanism that may contribute to differences in stroke progression between the sexes.Furthermore,the effects of manipulating autophagy may also differ between the sexes.Mechanisms that regulate autophagy in a sex-dependent manner in ischemic stroke remain unexplored.In this review,we summarize clinical and pre-clinical evidence for sex differences in stroke.We briefly introduce the autophagy process and summarize the effects of gonadal hormones in autophagy in the brain and discuss X-linked genes that could potentially regulate brain autophagy.Finally,we review pre-clinical studies that address the mechanisms that could mediate sex differences in brain autophagy after stroke.

Gonad traits and age structure of the sea urchin Heliocidaris crassispina in central range:a comparison with rangeextenders

The sea urchin Heliocidaris crassispina has northern expanded in geographic range to Toga Bay,Oga Peninsula in the Sea of Japan due to the global warming.To demonstrate the gonad traits and age structure of H.crassispina in the central range and compare the difference with those in northern extended range,we collected 100 H.crassispina individuals from a barren off Shitsumi,Fukui(central range),in August 2018.Their growth,gonad traits(size,development and color),and sex ratio were investigated and compared with those in Toga Bay,in August 2014(extended range).The successive 2012–2016 year classes of H.crassispina indicated successive juvenile recruitment in the central range,as found in the extended range.The individuals at three years old in central range were obviously larger than those in the extended range,showing large test diameter and body weight,possibly due to the higher temperature in central range than that in northern extended range.In addition,a balanced sex ratio in the central range was found,in contrast to the female-skewed sex ratio found in the extended range.Gonadal development was sex-and site-dependent.The testicular development in the central range was not only delayed in comparison to ovarian development,but also less delayed than those in the extended range,which possibly attribute to the higher water temperature in the central range.A significantly lower male gonad index and L*(lightness)value was recorded in central ranges compared to that in extended range,possibly due to the releasement of sperm of H.crassispina in central ranges.

Poultry genetic heritage cryopreservation and reconstruction:advancement and future challenges

Poultry genetics resources,including commercial selected lines,indigenous breeds,and experimental lines,are now being irreversibly lost at an alarming rate due to multiple reasons,which further threats the future livelihood and academic purpose.Collections of germplasm may reduce the risk of catastrophic loss of genetic diversity by guaranteeing that a pool of genetic variability is available to ensure the reintroduction and replenishment of the genetic stocks.The setting up of biobanks for poultry is challenging because the high sensitiveness of spermatozoa to freezing–thawing process,inability to cryopreserve the egg or embryo,coupled with the females being heterogametic sex.The progress in cryobiology and biotechnologies have made possible the extension of the range of germplasm for poultry species available in cryobanks,including semen,primordial germ cells,somatic cells and gonads.In this review,we introduce the state-of-the-art technologies for avian genetic resource conservation and breed reconstruction,and discuss the potential challenges for future study and further extending of these technologies to ongoing and future conservation efforts.

Single-cell RNA sequencing reveals gonadal dynamic changes during sex differentiation in hermaphroditic protogynous orange-spotted grouper(Epinephelus coioides)

DEAR EDITOR,Sex differentiation is a complex process that requires many factors to regulate gonadal proliferation, differentiation,development, and organization. In teleosts, the molecular mechanisms of sex differentiation are diverse and unclear.

Male sex determination： insights into molecular mechanisms

Disorders of sex development often arise from anomalies in the molecular or cellular networks that guide the differentiation of the embryonic gonad into either a testis or an ovary, two functionally distinct organs. The activation of the Y-linked gene Sry （sex- determining region Y） and its downstream target Sox9 （Sry box-containing gene 9） triggers testis differentiation by stimulating the differentiation of Sertoli cells, which then direct testis morphogenesis. Once engaged, a genetic pathway promotes the testis development while actively suppressing genes involved in ovarian development. This review focuses on the events of testis determination and the struggle to maintain male fate in the face of antagonistic pressure from the underlying female programme.

The m6A methylation regulates gonadal sex differentiation in chicken embryo

Background:As a ubiquitous reversible epigenetic RNA modification,N6-methyladenosine(m6A)plays crucial regulatory roles in multiple biological pathways.However,its functional mechanisms in sex determination and differentiation during gonadal development of chicken embryos are not clear.Therefore,we established a transcriptome-wide m6A map in the female and male chicken left gonads of embryonic day 7(E7)by methylated RNA immunoprecipitation sequencing(MeRIP-seq)to offer insight into the landscape of m6A methylation and investigate the post-transcriptional modification underlying gonadal differentiation.Results:The chicken embryonic gonadal transcriptome was extensively methylated.We found 15,191 and 16,111 m6A peaks in the female and male left gonads,respectively,which were mainly enriched in the coding sequence(CDS)and stop codon.Among these m6A peaks,we identified that 1013 and 751 were hypermethylated in females and males,respectively.These differential peaks covered 281 and 327 genes,such as BMP2,SMAD2,SOX9 and CYP19A1,which were primarily associated with development,morphogenesis and sex differentiation by functional enrichment.Further analysis revealed that the m6A methylation level was positively correlated with gene expression abundance.Furthermore,we found that YTHDC2 could regulate the expression of sex-related genes,especially HEMGN and SOX9,in male mesonephros/gonad mingle cells,which was verified by in vitro experiments,suggesting a regulatory role of m6A methylation in chicken gonad differentiation.Conclusions:This work provided a comprehensive m6A methylation profile of chicken embryonic gonads and revealed YTHDC2 as a key regulator responsible for sex differentiation.Our results contribute to a better understanding of epigenetic factors involved in chicken sex determination and differentiation and to promoting the future development of sex manipulation in poultry industry.

Characterization, Expression and Function Analysis of DAX1 Gene of Scallop(Chlamys farreri Jones and Preston 1904) During Its Gametogenesis

DAX1, a member of nuclear receptor superfamily, has a function in the sex determination and gonadal differentiation of several vertebrate species. However, little information about DAX1 of invertebrates is available. Here we cloned a homolog of scallop(Chlamys farreri Jones and Preston 1904) dax1, Cf-dax1, and determined its expression characteristics at m RNA and protein levels. The c DNA sequence of Cf-dax1 was 2093 bp in length, including 1404 bp open reading frame(ORF) encoding 467 amino acids. Unlike those of vertebrates, no conserved LXXLL-related motif was found in the putative DNA binding region of Cf-DAX1. Fluorescence in situ hybridization showed that Cf-dax1 located on the short arm of a pair of subtelocentric chromosomes. Tissue distribution analysis using semi-quantitative RT-PCR revealed that Cf-dax1 expressed widely in adult scallop tissues, with the highest expression level found in adductor muscle, moderate level in mantle, gill and testis, and low level in kidney, ovary and hepatopancreas. The result of quantitative real-time PCR indicated that the expression of Cf-dax1 was significantly higher(P<0.05) in testis than in ovary at the same stage, showing a sex-dimorphic expression pattern. Furthermore, immunohistochemical detection found that Cf-DAX1 mainly located in spermatogonia and spermatocytes of testis and in oogonia and oocytes of ovary, implying that DAX1 may involve in gametogenesis of bivalves.

Effects of spectral composition,photoperiod and light intensity on the gonadal development of Atlantic salmon Salmo salar in recirculating aquaculture systems(RAS)

Artificial lighting regimes have been successfully used to inhibit sexual maturity of Atlantic salmon in confinement.However,when these operations are applied in commercial recirculating aquaculture systems(RAS) using standard lighting technology,sexual maturation is not suppressed.In this study,an L_9(3~3) orthogonal design was used to determine the effects of three factors(spectral composition,photoperiod,and light intensity) on the gonadal development of Atlantic salmon in RAS.We demonstrated that the photoperiod at the tested levels had a much greater effect on the gonadosomatic index and female Fulton condition factor than spectral composition and light intensity.The photoperiod had a significant effect on the secretion of sex steroids and melatonin(P<0.05),and a short photoperiod delayed sex steroid and melatonin level increases.The three test factors had no significant effects on the survival rate,specific growth rate,relative weight gain,and male Fulton condition factor(P>0.05).The optimum lighting levels in female and male Atlantic salmon were LD 8:16,455 nm(or 625 nm),8.60 W/m^2;and LD 8:16,8.60 W/m^2,455 nm respectively.These conditions not only delayed gonadal development,but also had no negative effects on Atlantic salmon growth in RAS.These results demonstrate that a combination of spectral composition,photoperiod and light intensity is effective at delaying the gonadal development of both male and female salmon in RAS.

Phenotypic correlations of somatic and gonad traits of sea urchins Glyptocidaris crenularis in two sampled periods:first insight into its breeding and aquaculture

Prior knowledge of the correlations among commercially important traits in Glyptocidaris crenularis can be used to guide development of breeding and aquaculture programs for this species.We evaluated the phenotypic correlations between somatic and gonadal traits in G.crenularis during two sample periods(October 2010 and April 2011).The coeffi cients of variation(CV%)for body weight(BW),redness(a*),yellowness(b*),gonad wet weight(GW),and gonad index(GI)were>20% in both periods,while those of other traits were very low.GW was signifi cantly correlated with somatic traits(diameter,height,and body weight)in both periods,although the correlation coeffi cients ranged from 0.473 to 0.636(P<0.01).Gonad moisture content(GC),a*,and lightness(L*)were not signifi cantly correlated with somatic traits(P>0.05).The color trait b* was negatively correlated with somatic traits(test diameter,test height,and body weight)during the period when the gonads were well developed(April 2011,P<0.05).We observed little difference in the nature of the phenotypic correlations between the two sample periods.The high variation in the somatic and gonad weight traits in G.crenularis suggests there is considerable potential for selective breeding.However,our results suggest that gonad quality traits cannot be selected indirectly by selecting for body weight.Our study provides an insight into the method for improving the breeding and aquaculture of G.crenulari.

Recovery of gonadal development in tiger puffer Takifugu rubripes after exposure to 17β-estradiol during early life stages

The aim of the present study was to investigate the long-term effects of 17l）-estradiol （E2） exposure on gonadal development in the tiger puffer （Taktfugu rubripes）, which has a genetic sex determination system of male homogametic XY-XX. Tiger puffer larvae were exposed to 1, 10 and 100 μg/L E2 from 15 to 100 days post-hatch （dph） and then maintained in clean seawater until 400 dph. Changes in sex ratio, gonadal structure and gonadosomatic index （GSI） were monitored at 100, 160, 270 and 400 dph. Sex-associated single nucleotide polymorphism （SNP） markers were used to analyze the genetic sex of samples, except those at 100 dph. Exposure had a positive effect on the conversion of genetically male gonads into phenotypically female gonads at 100 dph. However, gonads from 60% of genetic XY males in the 1-μg/L E2 group and 100% in the 10-μg/L E2 group developed intersexual gonads at 160 dph; gonads of all genetic XY males in the two treatment groups reverted to testis by 270 dph. While 38%, 57% and 44% of gonads of XY fish in the 100-gg/L E2 group reverted to intersexual gonads at 160, 270 and 400 dph, respectively, none reverted to testis after E2 treatment. In addition, E2 exposure inhibited gonadal growth of both genetic sexes, as indicated by the clear dose-dependent decrease in GSI at 270 and 400 dph. The results showed that exposure to E2 during the early life stages of tiger puffer disrupted gonadal development, but that fish recovered after migration to clean seawater. The study suggests the potential use of tiger puffer as a valuable indicator species to evaluate the effects of environmental estrogens on marine fish, thereby protecting valuable fishery resources.

Higher Dietary Arachidonic Acid Levels Improved the Growth Performance, Gonad Development, Nutritional Value, and Antioxidant Enzyme Activities of Adult Sea Urchin(Strongylocentrotus intermedius)

The gonads of sea urchins(Strongylocentrotus intermedius) are characterized by high levels of arachidonic acid(ARA, 20:4 n-6) and eicosapentaenoic acid(EPA, 20:5 n-3). However, to our knowledge, little information is available regarding the physiological response of adult sea urchins to dietary ARA. In the present study, four dietary feeds were formulated with graded ARA(0, 0.5%, 1%, and 2% dry diet). Each diet was randomly allocated to three cages during a 56-day feeding experiment. The results showed that the sea urchin weight gain rate(WGR) and the gonadosomatic index(GI) significantly increased as ARA was equal to or above 1.0% of dry diet(P < 0.05). The activities of superoxide dismutase(SOD), catalase(CAT), and total anti-oxidative capacity(T-AOC) were the highest in the coelomic fluid of sea urchins that were fed diets with 1% ARA. The total essential amino acid(TEAA) and its ratio to total non-essential amino acid(TNEAA) showed a similar tendency to WGR and GI as dietary ARA increased, and the highest TEAA and TEAA/TNEAA were observed in the gonads of sea urchins that were fed diets with 1% ARA. Levels of ARA and ARA/EPA of the gonads increased while n-3/n-6 polyunsaturated fatty acid(PUFA) decreased with the increase of dietary ARA(P < 0.05). EPA in the gonads of experimental animals fed with formulated feeds showed no significant differences(P> 0.05), but was significantly lower than those fed with kelp(P < 0.05). These results suggested that relatively higher levels of ARA(1% dry diet) significantly promoted growth, gonad development, activities of antioxidant enzymes, as well as nutritional values(TEAA, TEAA/TNEAA, and PUFA) of adult S. intermedius.

Heshouwu decoction, a Chinese herb for tonifying kidney, ameliorates hypothalamic-pituitary-testicular axis secretion in aging rats

An increasing amount of evidence demonstrates the anti-aging effect of Heshouwu in pill form. In this study, a subacute aging rat model was established by continuous intraperitoneal injection of D-galactose and treated with Heshouwu decoction （a Chinese herb for tonifying the kidney, comprising Heshouwu pill, Herba Epimedii, Radix Salviae Miltiorrhiae, and Poria）. Heshouwu pill treated rats were the positive control group. Radioimmunoassay, immunohistochemical staining, and western blot assay showed hypothalamic gonadotropin-releasing hormone, hypothalamic substance P, and serum gonadotropin levels to be significantly increased in the model rats; the concentrations of hypothalamic ~3-endorphin, and serum levels of insulin-like growth factor I and testosterone were significantly decreased. 1713- and 3[3-hydroxysteroid dehydrogenase expression in testicular tissue was also decreased. Intragastric administration of Heshouwu decoction at high （9.6 g/mL/100 g）, medium （4.8 g/mL/100 g）, and low （2.4 g/mlJ100 g） doses, Heshouwu decoction pretreatment at a medium dose （4.8 g/mL/100 g）, and Heshouwu pill （2.06 g/mL/100 g） significantly reversed these changes. Heshouwu decoction pretreatment and high-dose Heshouwu decoction had the greatest anti-aging effects. These experimental findings indicate that Heshouwu decoction can improve hypothalamic-pituitary-testicular axis secretion in a subacute aging rat model, and prevent and delay gonadal axis aging, with an effect superior to that of Heshouwu pill.

Characterization of Bacterial Community Associated with Four Organs of the Yesso Scallop (Patinopecten yessoensis) by High-Throughput Sequencing

We used Illumina high-throughput sequencing of PCR-amplified V3-V4 16 S rRNA gene regions to characterize bacterial communities associated with the adductor muscles, gills, gonads and intestines of the Yesso scallop(Patinopecten yessoensis) from waters around Zhangzidao, Dalian, China. Overall, 421,276 optimized reads were classified as 25 described bacterial phyla and 308 genera. Firmicutes, Proteobacteria, Tenericutes, Bacteroidetes, Chlamydiae and Spirochaetae accounted for > 97% of the total reads in the four organs. The bacterial 16 S rDNA sequences assigned to Firmicutes and Proteobacteria were abundant in the adductor muscles, gills and gonads; while reads from Tenericutes were dominant in the intestines, followed by those from Firmicutes, Chlamydiae, Proteobacteria and Bacteroidetes. At the genus level, the dominant genera in the adductor muscles, gills and gonads appeared to be Bacillus, Enterococcus and Lactococcus, whereas Mycoplasma was dominant in the intestines. The relative abundances of Bacillus, Enterococcus, Lactococcus, Alkaliphilus, Raoultella, Paenibacillus and Oceanobacillus were significantly lower in the intestine than in the other three organs. Cluster analysis and principal coordinates analysis of the operational taxonomy units profile revealed significant differences in the bacterial community structure between the intestine and the other three organs. Taken together, these results suggest that scallops have intestine-specific bacterial communities and the adductor muscles, gills and gonads harbor similar communities. The difference in the bacterial community between organs may relate to unique habitats, surroundings, diet and their respective physiological functions.

PRODUCTION OF FERTILE EGGS AND SPERMS IN LABORATORY-REARED AMPHIOXUS BRAN- CHIOSTOMA BELCHERI TSINGTAUNESE

Although amphioxus has been studied embryologically for over 130 years,little is known concerning the gonadal development of the laboratory reared animal.The present study in which the amphioxus were successfully maintained in the laboratory for more than one year showed that they could produce fertile eggs and sperms like their wild counterparts.

Functional analysis of the promoter of the dmrt1 gene in Chinese tongue sole,Cynoglossus semilaevis

The dmrt1 (doublesex and mab-3 related transcription factor 1) gene is considered to play an essential role in testis differentiation and development among metazoan species. As an economically important marine fish in China, Chinese tongue sole (Cynoglossus semilaevis) possess a ZW/ZZ sex-determining system, and the females usually grow two to four times bigger than males. Previous studies have confirmed that Z-linked dmrt1 gene is the male determining gene in C. semilaevis and TALENs-mediated knocking-out individuals showed increased growth phenotype. In order to investigate the function of dmrt1 regulatory regions and its potential application, we have cloned the promoter of C. semilaevis dmrt1. The acquired sequence consisted of a 5'-upstream sequence and a partial exon1;functional analysis showed that there were two positive regulatory regions in the promoter, as well as one negative region and one neutral region. These positive regions were then combined to obtain several newly constructed promoters. Both in vitro and in vivo analysis confirmed one of them (dmrt1-Δ3+Δ4) showed elevated promoting activity compared to the full-length promoter and could drive exogenous gene expression in tongue sole gonads. Our results will provide useful information for understanding the regulatory mechanism of dmrt1 gene during C. semilaevis sex determination, and the endogenous promoter will facilitate the transgenic research for analyzing sex- and growth-related genes.

Characterization and differential expression of three GnRH forms during reproductive development in cultured turbot Schophthalmus maximus

Turbots( Schophthalmus maximus), one of the most important economic marine flatfish species, fail to undergo final spawning and spermiation naturally under artificial farming conditions. In vertebrates, reproduction is regulated by the brain-pituitary-gonadal axis(BPG-axis), and gonadotropin releasing hormone(GnRH) is one of its key components. Therefore, to better understand the physiology of reproduction in the turbot, three of the genes encoding GnRH subtypes— sbGnRH, c GnRH-II and sGnRH —were cloned and sequenced by isolating the cDNA sequences. The localizations and patterns of expression of their mRNAs were also evaluated during seasonal gonadal development. All three mRNAs were expressed abundantly in the brain; sbGnRH and sGnRH mRNAs were also detected in the gonads and pituitary gland, and sbGnRH expression was much higher than that of sGnRH, indicating the critical role of sbGnRH in regulating the BPG-axis. Moreover, the brain expression patterns of sbGnRH and sGnRH mRNAs showed an increased trend during gonadal development, peaking in mature stages. This indicated the direct regulation of gonadal development by the GnRH system. In addition, c GnRH-II mRNA expression showed no significant variations, suggesting that c GnRH-II is not critically involved in the control of reproduction. Further, the mRNA abundances of the three GnRH forms in the breeding season were significantly higher than those in immature and post-breeding stages in all analyzed brain areas. Therefore, we propose that sbGnRH is the most important hormone for the regulation of reproduction in turbot via the BPG-axis. These results will help in better understanding the reproductive endocrine mechanisms of turbots and lay the groundwork for additional studies aimed at comparing the reproductive physiology of wild individuals with those raised under artificial conditions.

Anti-diabetic and anti-hyperlipidemic effects of sea cucumber(Cucumaria frondosa) gonad hydrolysates in type Ⅱ diabetic rats

The present study investigated the anti-diabetic activity and potential mechanisms of sea cucumber gonad hydrolysates(SCGH)in a rat model of type II diabetes induced by streptozotocin(STZ)combined with high-fat diet(HFD).Results showed that SCGH significantly reduced water intake,fasting blood glucose level and glycated hemoglobin level.Moreover,the oral glucose tolerance,insulin resistance and plasma lipid level in diabetic rats were also alleviated.Furthermore,histological analysis showed that SCGH effectively protected the tissue structure of liver.In addition,mechanism studies showed that SCGH improved glucose metabolism via activating the IRS/Akt signaling pathway,and promoted lipid metabolism via activating the AMPK signaling pathway.In summary,these findings suggested that SCGH have potential anti-diabetic effects by improving insulin resistance and lipid metabolism disorders.

Female Reproductive Cycles of Phrynocephalus przewalskii (Lacertilia:Agamidae) in the Tengger Desert,China

The timing of reproduction can significantly affect an offspring＇s fitness, thereby also influencing the fitness of the parents, especially in species inhabiting extreme environments, such as deserts. Female reproductive cycles in Phrynocephalus przewalskii were studied from April to September 2008. Significant cycles of gonadal volume were found in all studied populations and the cycles were similar among the various populations. Females began vitellogenesis in April and contained oviductal eggs form May to June. Gonad volume decreased significantly in July and reached minimum volume from August to September. The follicular growth was negatively correlated with increasing precipitation and temperature in all populations. Hatching occurs during summer and early fall, when most of the annual rainfall occurs. Mean clutch size based on all populations was 2.7 ± 0.9 SE （n = 71）.