Maintenance Of Testes Research Articles

Denovo RNA-Seq analysis of ovary and testis reveals potential differentially expressed transcripts associated with gonadal unsynchronization development in Onychostoma macrolepis

The Onychostoma macrolepis (O. macrolepis) is a rare and endangered wild species. Their endangered extinction might be due to their low fertility. To further illustrate the molecular mechanism of gonad development of the male and female O. macrolepis, the present study carried out de novo testicular and ovarian transcriptome sequencing. By comparing ovary and testis, 30,869 differentially expressed unigenes (9870 in female, 20999 in male) were identified. In addition, KEGG and GO analysis suggested that the Hedgehog signaling pathway have important roles in testis maintenance and spermatogenesis, whereas the Hippo signaling pathway and Wnt signaling pathway are likely to participate in ovary maintenance. RT-qPCR analysis results were consistent with transcriptome sequencing that all of gender differentiation-related genes (FOXL2, GDF9, WNT4, CYP19A1, SOX9 and GATA4), temperature-enriched genes (NOVA1, CTGF and NR4A1), clock-related genes (PER2, PER3, CRY1, CRY2, BMAL1 and CIPC) were significantly differential expression in testis compared with ovaries. Taken together, these results revealed a potential molecular mechanism that low fertility of the O. macrolepis might strong correlate with the gonadal dyssynchrony development of the male and female, which might provide theoretical basis and technical support for artificial reproduction and germplasm resource protection of the O. macrolepis.

Cloning, expression, and function of the Spdmrt-like gene in Scylla paramamosain.

The mud crab Scylla paramamosain is an economically important species for aquaculture in China and has sexually dimorphic between females and males. Understanding sex differentiation in this species is essential for the development of monosex aquaculture. The Dmrt genes play a vital role in sex differentiation in animals. In this study, two dmrt-like transcript variants, Spdmrt-like-tv1 and Spdmrt-like-v2, were cloned. SpDmrt-like-tv1 contained a DM domain, while SpDmrt-like-tv2 contained a DM and a DMA domain. Spdmrt-like-tv1 and Spdmrt-like-tv2 were both specifically expressed in testis. During testicular development, the expression level of Spdmrt-like-tv1 increased from stage I to stage II (P > 0.05) and then decreased from stage II to stage III (P < 0.05). The expression level of Spdmrt-like-tv2 in stages I and II was significantly higher than that in stage III (P < 0.05). During embryonic development, the expression level of Spdmrt-like-tv1 was higher in the mid-embryonic stage compared with the early and late stages, but the differences were not significant. Moreover, the expression level of Spdmrt-like-tv2 was stable and remained high throughout embryonic development. Furthermore, the expression level of Spdmrt-like-tv2 was significantly higher than that of Spdmrt-like-tv1. Knockdown of Spdmrt-like variants indicated that the regulative target gene of Spdmrt-like-tv1 was Spsox21, and the regulative target genes of Spdmrt-like-tv2 were Spfoxl2 and Spsox21. Combined with the results in our previously published peer-reviewed articles that the expression of Spfoxl2 in the testis was significantly higher than that in the ovary, and Spfoxl2 negatively regulated Spvtg expression. Spsox21 played a role in the development and maintenance of testis as well as in the process of neural development and regulation of body segmentation. Therefore, we suggest that Spdmrt-like-tv1 and Spdmrt-like-tv2 might be involved in testicular development and embryonic development, and Spdmrt-like-tv2 might play more important roles in these two developmental processes by regulating the expression of Spfoxl2 and Spsox21 due to its high expression.

Amh dominant expression in Sertoli cells during the testicular differentiation and development stages in the olive flounder Paralichthys olivaceus.

The olive flounder Paralichthys olivaceus, an important marine fish, shows gender differences in growth. The mechanism on its gonadal differentiation direction affected with exogenous factors still needs to be clarified. The anti-Müllerian hormone (amh) gene is involved in fish testicular differentiation and maintenance. The aim of this study was to investigate the expression of the flounder amh in tissues and the gonads. The quantitative expression analysis results showed that it was highly expressed in the testis, especially in the testis at stages I - IV (P<0.05). Also, amh was detected in Sertoli cells surrounding spermatogonia and peripheral seminiferous lobule of the testis with in situ hybridization (ISH) and immunohistochemistry (IHC). During the differentiation period, the amh expression in the testis of the tamoxifen treatment group (100ppm) was higher than that in the ovary of the 17β-estradiol (E2, 5ppm) group, and the expression levels of amh during process of the male differentiation in the tamoxifen group were higher than those in the 17ɑ-methyltestosterone (MT, 5ppm) group (P<0.05). ISH results also exhibited that amh was expressed in the somatic cells that surrounded the germ cells of juvenile flounder similar to adult ones. Furthermore, the flounder gonads in the tamoxifen group maintained more germ cells and somatic cells than those in the MT group from 20 to 80mm total length (TL). Especially, at 60 and 80mm TL, the numbers of germ and somatic cells in the tamoxifen group were significantly higher than those in the MT group (P<0.05). In summary, amh might initiate the process of testicular differentiation, and is involved in the early development and maintenance of testis.

Characterization of gonad differentially expressed SoxB2 genes in mud crab Scylla paramamosain.

Members of sox gene family play critical roles in development, and some of them have crucial functions in sexual dimorphism. To understand the role of two SoxB2 genes, Sox14b and Sox21 of mud crab Scylla paramamosain, the full-length 1939bp SpSox14b cDNA sequence and 861bp SpSox21 cDNA sequence were obtained from the crab's transcriptome database, which encode 397 and 259 amino acids respectively. The results of sq-PCR showed that SpSox14b was expressed in all tissues, while SpSox21 was only expressed in the testis and brain. qRT-PCR showed that the expression level of SpSox14b in ovary was significantly higher than that of testis, and during the gonad development its expression was the highest in O2 (previtellogenesis) stage. The expression level of SpSox21 in testis was much higher than in brain, and was significantly higher in T3 (the mature sperm stage) than in other stages of testis development. Meanwhile, in different stages of larval development, SpSox21 was low expressed in zoea, then increased significantly in megalopa. Therefore we speculated that SpSox14b and SpSox21 may play different roles in the gonad development of mud crab, especially SpSox21 may be involved in the development and maintenance of testis. The expression level of SpSox14b and SpSox21 during the eye-pigment formation was significantly higher than that in other embryonic development stages, the results of whole-mount in situ hybridization showed that SpSox14b and SpSox21 were mainly located near the head and the compound eyes in eye-pigment formation stage and hatching. It suggested that they may be involved in the formation of brain nerves and are related to the regulation of body segments, and play different roles in sexual dimorphism.

Functional analysis of a SoxE gene in the oriental freshwater prawn, Macrobrachium nipponense by molecular cloning, expression pattern analysis, and in situ hybridization (de Haan, 1849).

In this study, a full-length cDNA sequence of SoxE (subgroup E within the Sox family of transcription factors) was cloned from Macrobrachium nipponense and named MnSoxE1. The full-length cDNA of MnSoxE1 is 1748bp, consisting of a 110bp 5' UTR, a 105bp 3' UTR, and a 1533bp ORF that encodes 510 amino acids. Conserved domains showed that MnSoxE1 has a high similarity to the SoxE gene of Penaeus vannamei. Phylogenetic tree analysis classified that MnSoxE1 with the SoxE gene of other arthropods into one clade. These results suggested that MnSoxE1 belongs to the SoxE subgroup. During embryonic development, MnSoxE1 was mainly expressed in the gastrula stage, implicating its involvement in tissue cell differentiation and formation. In the post-larval stages, the expression of MnSoxE1 continued to increase on days 1-10. The expression level in males was significantly higher than that in females. Males are clearly distinguishable from females on post-larval day 25, showing that MnSoxE1 may play a role in promoting early development and germ cell and gonadal differentiation, especially for males. qPCR analysis showed that MnSoxE1 may also be involved in oogonium proliferation during ovary development. Further in situ hybridization analysis revealed that MnSoxE1 was mainly located in oocytes and spermatocytes, especially in sertoli cells, and implies that it may be involved in the development of oocytes and spermatocytes, as well as the maintenance of testes in mature prawns. These results indicate that MnSoxE1 is involved in gonadal differentiation and development in M. nipponense, especially testis development.

Transcriptome sequencing and comparative analysis of adult ovary and testis identify potential gonadal maintenance-related genes in Mauremys reevesii with temperature-dependent sex determination.

Mauremys reevesii is a classical organism with temperature-dependent sex determination (TSD). Gonad development in early life has recently received considerable attention but gonadal maintenance after sex differentiation in turtles with TSD remains a mystery. In this study, we sequenced the transcriptomes for the adult testis and ovary using RNA-seq, and 36,221 transcripts were identified. In total, 1,594 differentially expressed genes (DEGs) were identified where 756 DEGs were upregulated in the testis and 838 DEGs were upregulated in the ovary. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis suggested that the TGF-beta signaling pathway and Hedgehog signaling pathway have important roles in testis maintenance and spermatogenesis, whereas the Hippo signaling pathway and Wnt signaling pathway are likely to participate in ovary maintenance. We determined the existence of antagonistic networks containing significant specific-expressed genes and pathways related to gonadal maintenance and gametogenesis in the adult gonads of M. reevesii. The candidate gene Fibronectin type 3 and ankyrin repeat domains 1 (FANK1) might be involved with the regulation of testis spermatogenesis.

Cloning the Dmrt1 and DmrtA2 genes of ayu (Plecoglossus altivelis) and mapping their expression in adult, larval, and embryonic stages.

The Dmrt family of genes are involved in sex differentiation in different species of invertebrates, and some vertebrates including human. In this study, we cloned the full-length cDNA of ayu (Plecoglossus altivelis) Dmrt1 and DmrtA2. Sequence and phylogenetic tree analyses showed ayu Dmrt1 showed highest similarity to that of Oncorhynchus mykiss while ayu DmrtA2 is most similar to that of Oryzias latipes. Fluorescence-based quantitative reverse transcription PCR (qRT-PCR) revealed the Dmrt1 was predominantly expressed in the testis. At the larval stages, Dmrt1 mRNA expression level was highest during 52-64 days post hatching (dph) and at the gastrula stage during embryonic development. DmrtA2, meanwhile, was specifically expressed in the ovary and was highly expressed in the female brain tissue, but not male brain tissue. During the larval stages, DmrtA2 expression remained high before day 34, and then fluctuated while generally decreasing. During embryonic development, DmrtA2 expression increased gradually and peaked at the hatching stage. Our data suggest that ayu Dmrt1 might participate in the differentiation and maintenance of testis while DmrtA2 may play a role in ovary-differentiation and mature-ovary maintenance. DmrtA2 might also participate in brain development.

Maintenance of spermatogenesis and seminal vesicles in the hypophysectomized catfish, Heteropneustes fossilis (Bloch): effects of ovine and salmon gonadotropin, and testosterone.

The effects of 20-day treatment with ovine luteinizing (LH) and follicle-stimulating hormone (FSH), partially purified salmon gonadotropin (SG-G100), and testosterone were investigated by evaluating the maintenance of spermatogenesis in the testes and secretory activity in the seminal vesicles of the catfish, Heteropneustes fossilis, after hypophysectomy during the prespawning period. Low doses of LH (0.1 or 1 μg per fish) neither maintained spermatogenesis in the testes nor secretory activity in the seminal vesicles, whereas a higher dose of LH (10 μg per fish) maintained spermatogenesis qualitatively, and sustained the seminal vesicles in a moderately secretory state. One microgram of SG-G100 per fish partially maintained spermatogenesis but the seminal vesicles showed poor maintenance. Ten micrograms of SG-G100 per fish not only sustained spermatogenesis qualitatively and quantitatively, but also induced spermiation; the seminal vesicles, though enlarged, contained little secretion, indicating possible reduction through spermiation. FSH did not maintain spermatogenesis in the testes or secretory activity in the seminal vesicles. Testosterone (10, 100, or 500 μg per fish) had a dose-dependent maintenance response in the testes and seminal vesicles; with 500 μg of testosterone the maintenance of testes and seminal vesicles was similar to that in initial intact controls, with no indication of spermiation.

Effects of chronic administration of sheep stalk-median eminence extract on organ weights of immature hypophysectomized rats with pituitary grafts.

SummarySingle pituitary grafts from donors of the same sex maintained some growth and led to slight maintenance of testes in male and uteri in female hypophysectomized rats. There was slight maintenance of adrenal weight in both sexes. Injection of crude sheep stalk-median eminence (SME) extracts resulted in a further significant increase in testes and ventral prostate weights in hypophysectomized, grafted males and in uterine and ovarian weights in grafted females on comparison to the weights of these organs in grafted animals injected with cerebral cortical extract. Adrenal weight was increased in rats of both sexes by SME extract. The results provide evidence that chronic injection of hypothalamic extract can increase the weight of pituitary target organs in both male and female rats presumably via stimulation of both release and synthesis of pituitary hormones.