Androgen-related sperm storage in oviduct of Chinese Soft-Shelled Turtle in vivo during annual cycle

Liu, Tengfei; Chu, Xia; Huang, Yufei; Yang, Ping; Li, Quanfu; Hu, Lisi; Chen, Hong; Chen, Qiusheng

doi:10.1038/srep20456

Cited by 22 publications

(21 citation statements)

References 47 publications

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“…Paraffin-embedded blocks were cut to 6 μm slices using a microtome device (Leica, RM 2245, Germany). The sections were dewaxed and stained with a haematoxylin-eosin staining method that was previously described by Liu et al (2016a). At this point, the samples were observed under a light microscope (Olympus DP73).…”

Section: Methodsmentioning

confidence: 99%

Molecular and Cellular Mechanisms of Apoptosis during Dissociated Spermatogenesis

et al. 2017

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Apoptosis is a tightly controlled process by which tissues eliminate unwanted cells. Spontaneous germ cell apoptosis in testis has been broadly investigated in mammals that have an associated spermatogenesis pattern. However, the mechanism of germ cell apoptosis in seasonally breeding reptiles following a dissociated spermatogenesis has remained enigmatic. In the present study, morphological evidence has clearly confirmed the dissociated spermatogenesis pattern in Pelodiscus sinensis. TUNEL and TEM analyses presented dynamic changes and ultrastructural characteristics of apoptotic germ cells during seasonal spermatogenesis, implying that apoptosis might be one of the key mechanisms to clear degraded germ cells. Furthermore, using RNA-Seq and digital gene expression (DGE) profiling, a large number of apoptosis-related differentially expressed genes (DEGs) at different phases of spermatogenesis were identified and characterized in the testis. DGE and RT-qPCR analysis revealed that the critical anti-apoptosis genes, such as Bcl-2, BAG1, and BAG5, showed up-regulated patterns during intermediate and late spermatogenesis. Moreover, the increases in mitochondrial transmembrane potential in July and October were detected by JC-1 staining. Notably, the low protein levels of pro-apoptotic cleaved caspase-3 and CytC in cytoplasm were detected by immunohistochemistry and western blot analyses, indicating that the CytC-Caspase model might be responsible for the effects of germ cell apoptosis on seasonal spermatogenesis. These results facilitate understanding the regulatory mechanisms of apoptosis during spermatogenesis and uncovering the biological process of the dissociated spermatogenesis system in reptiles.

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Section: Methodsmentioning

confidence: 99%

Molecular and Cellular Mechanisms of Apoptosis during Dissociated Spermatogenesis

et al. 2017

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“…Immunohistochemical staining for LC3 and ATG7 was performed according to the manufacturer's recommendations and as suggested in previous studies [ 37 , 38 ]. The sections were processed using a standard immunohistochemistry protocol as previously described [ 39 ]. Briefly, after deparaffinization, blocking endogenous peroxidase, microwave antigen retrieval, and BSA (bovine serum albumin) blocking, a primary rabbit anti-LC3B polyclonal antibody (ab48394, Abcam, Cambridge, UK) or mouse anti-ATG7 monoclonal antibody (SAB4200304, Sigma Aldrich, St. Louis, USA) was applied for 1 h at room temperature.…”

Section: Methodsmentioning

confidence: 99%

In vivo autophagy and biogenesis of autophagosomes within male haploid cells during spermiogenesis

et al. 2017

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Autophagy is a unique catabolic pathway that is linked to several physiological processes. However, its role in the process of spermiogenesis is largely unknown. The aim of the current study was to determine the in vivo role of autophagy and the origin of autophagosome membrane biogenesis within male haploid cells. Our immunohistochemistry results demonstrated that LC3 and ATG7 localization were increased dramatically in round to elongated spermatids (haploid cells) towards the lumen of seminiferous tubules, however, poorly expressed in the early stages of germ cells near the basal membrane. Moreover, transmission electron microscopy revealed that the numbers of lysosomes and autophagosomes increased in the elongated spermatids as spermiogenesis progressed. However, no evidence was found for the presence of autophagosomes in the Sertoli cells, spermatogonia or early primary spermatocytes (diploid cells). Furthermore, TEM showed that many endoplasmic reticula were transformed into a “chrysanthemum flower center,” from which a double-layered isolation membrane appeared to develop into an autophagosome. This study provides novel evidence about the formation of autophagosomes through the chrysanthemum flower center from the endoplasmic reticulum, and suggests that autophagy may have an important role in the removal of extra cytoplasm within male haploid cells during spermiogenesis.

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“…Spermatogenesis, copulation and ovulation are seasonal and segregational in turtles 5 . Our earlier studies clearly provided structural and ultrastructural evidence for sperm storage in the oviduct of female P. sinensis , and spermatozoa could survive in the oviduct during the hibernation season (from November to the following April) 9 20 . Recently, the draft genome sequences of P. sinensis have been published 21 , providing a useful database for genomic and functional investigations on some important biological traits in Chinese soft-shelled turtle.…”

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confidence: 65%

Global analysis of differential gene expression related to long-term sperm storage in oviduct of Chinese Soft-Shelled Turtle Pelodiscus sinensis

Liu

Yang

Chen

et al. 2016

Sci Rep

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Important evolutionary and ecological consequences arise from the ability of female turtles to store viable spermatozoa for an extended period. Although previous morphological studies have observed the localization of spermatozoa in Pelodiscus sinensis oviduct, no systematic study on the identification of genes that are involved in long-term sperm storage has been performed. In this study, the oviduct of P. sinensis at different phases (reproductive and hibernation seasons) was prepared for RNA-Seq and gene expression profiling. In total, 2,662 differentially expressed genes (DEGs) including 1,224 up- and 1,438 down-regulated genes were identified from two cDNA libraries. Functional enrichment analysis indicated that many genes were predominantly involved in the immune response, apoptosis pathway and regulation of autophagy. RT-qPCR, ELISA, western blot and IHC analyses showed that the expression profiles of mRNA and protein in selected DEGs were in consistent with results from RNA-Seq analysis. Remarkably, TUNEL analysis revealed the reduced number of apoptotic cells during sperm storage. IHC and TEM analyses found that autophagy occurred in the oviduct epithelial cells, where the spermatozoa were closely attached. The outcomes of this study provide fundamental insights into the complex sperm storage regulatory process and facilitate elucidating the mechanism of sperm storage in P. sinensis.

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Androgen-related sperm storage in oviduct of Chinese Soft-Shelled Turtle in vivo during annual cycle

Cited by 22 publications

References 47 publications

Molecular and Cellular Mechanisms of Apoptosis during Dissociated Spermatogenesis

Molecular and Cellular Mechanisms of Apoptosis during Dissociated Spermatogenesis

In vivo autophagy and biogenesis of autophagosomes within male haploid cells during spermiogenesis

Global analysis of differential gene expression related to long-term sperm storage in oviduct of Chinese Soft-Shelled Turtle Pelodiscus sinensis

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