Dataset Information

Regulation of BTB dynamics in spermatogenesis - insights from the adjudin toxicant model.

ABSTRACT: During spermatogenesis, cell organelles and germ cells, most notably haploid spermatids, are transported across the seminiferous epithelium so that fully developed spermatids line-up at the edge of the tubule lumen to undergo spermiation at stage VIII of the cycle. Studies have suggested that the microtubule (MT)-based cytoskeleton is necessary to support these cellular events. However, the regulatory molecule(s) and underlying mechanism(s) remain poorly understood. Herein, we sought to better understand this event by using an adjudin-based animal model. Adult rats were treated with adjudin at low-dose (10?mg/kg b.w.) which by itself had no notable effects on spermatogenesis. Rats were also treated with low-dose adjudin combined with overexpression of two endogenously produced blood-testis barrier (BTB) modifiers, namely rpS6 [ribosomal protein S6, the downstream signaling protein of mammalian target of rapamycin complex 1 (mTORC1)] and F5-peptide (a biological active peptide released from laminin-?3 chain at the Sertoli-spermatid interface) versus the two BTB modifiers alone. Overexpression of these two BTB modifiers in the testis was shown to enhance delivery of adjudin to the testis, effectively inducing disruptive changes in MT cytoskeletons, causing truncation of MT conferred tracks that led to their collapse across the epithelium. The net result was massive germ cell exfoliation in the tubules, disrupting germ cell transport and cell adhesion across the seminiferous epithelium that led to aspermatogenesis. These changes were the result of disruptive spatial expression of several MT-based regulatory proteins. In summary, MT cytoskeleton supported by the network of MT regulatory proteins is crucial to maintain spermatogenesis.

SUBMITTER: Mao BP

PROVIDER: S-EPMC6813747 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Regulation of BTB Dynamics in Spermatogenesis-Insights From the Adjudin Model.

Mao Bai-Ping BP Li Linxi L Yan Ming M Ge Renshan R Lian Qingquan Q Cheng C Yan CY

Toxicological sciences : an official journal of the Society of Toxicology 20191101 1

During spermatogenesis, cell organelles, and germ cells, most notably haploid spermatids, are transported across the seminiferous epithelium so that fully developed spermatids line-up at the edge of the tubule lumen to undergo spermiation at stage VIII of the cycle. Studies have suggested that the microtubule (MT)-based cytoskeleton is necessary to support these cellular events. However, the regulatory molecule(s) and underlying mechanism(s) remain poorly understood. Herein, we sought to better ...[more]

PMID: 31397872

Similar Datasets

Project description:In adult mammals such as rats, the blood-testis barrier (BTB) conferred by adjacent Sertoli cells in the seminiferous epithelium segregates post-meiotic germ cell development from the systemic circulation and is one of the tightest blood-tissue barriers. Yet it must "open" transiently at stages VIII to IX of the epithelial cycle to accommodate the migration of preleptotene/leptotene spermatocytes. While this is a vital event of spermatogenesis, the mechanism(s) that regulates BTB dynamics is virtually unknown. Recent studies have suggested that transforming growth factor-beta3 (TGF-beta3) and tumor necrosis factor alpha (TNFalpha) secreted by Sertoli and germ cells into the microenvironment of the BTB are capable of inducing reversible BTB disruption in vivo, apparently by reducing the steady-state levels of occludin and zonula occludens-1 (ZO-1) at the BTB via the p38 mitogen activated protein (MAP) kinase signaling pathway. In this study, local administration of TGF-beta3 (200 ng/testis) to the testis was shown to reversibly perturb the BTB integrity in vivo. We next sought to delineate the mechanism by which these cytokines maintain the steady-state level of integral membrane proteins: occludin, junctional adhesion molecule-A (JAM-A) and N-cadherin at the BTB. Primary Sertoli cells cultured in vitro were shown to establish intact tight junctions and functional BTB within two days when assessed by transepithelial electrical resistance (TER) measurement across the cell epithelium. Sertoli cell integral membrane protein internalization at the BTB was assessed by biotinylation of cell surface proteins, to be followed by tracking the endocytosed/biotinylated proteins by using specific antibodies. Both TGF-beta3 (3 ng/ml) and TNFalpha (10 ng/ml) were shown to significantly accelerate the kinetics of internalization of JAM-A, N-cadherin, and occludin versus controls. Treatment of cells with phenylarsine oxide (PAO) at 10 microM that blocks clathrin-mediated endocytosis was shown to inhibit the TGF-beta3-induced protein internalization. This inhibition of TGF-beta3-mediated protein endocytosis was further validated by silencing of clathrin. The specific effect of TGF-beta3 on protein internalization was further confirmed by RNAi using specific TGF-beta receptor I (TbetaR1) siRNA duplexes. When TbetaR1 was knocked down, the TGF-beta3-induced increase in the kinetics of JAM-A and occludin endocytosis was abolished, making them indistinguishable from controls, illustrating the specificity of the TGF-beta3 effects on protein endocytosis. In summary, this report demonstrates for the first time that BTB dynamics are regulated by TGF-beta3 and TNFalpha via an enhancement of protein endocytosis at the BTB.

Project description:For non-hormonal male contraceptives that exert their effects in the testis locally instead of via the hypothalamic-pituitary-testicular axis, such as adjudin that disrupts germ cell adhesion, a major hurdle in their development is to improve their bioavailability so that they can be efficiently delivered to the seminiferous epithelium by transporting across the blood-testis barrier (BTB). If this can be done, it would widen the gap between their efficacy and general toxicity. However, Sertoli cells that constitute the BTB, peritubular myoid cells in the tunica propria, germ cells at different stages of their development, as well as endothelial cells that constitute the microvessels in the interstitium are all equipped with multiple drug transporters, most notably efflux drug transporters, such as P-glycoprotein, multidrug resistance-related protein 1 (MRP1) and breast cancer resistance protein (BCRP) that can actively prevent drugs (e.g., adjudin) from entering the seminiferous epithelium to exert their effects. Recent studies have shown that BCRP is highly expressed by endothelial cells of the microvessels in the interstitium in the testis and also peritubular myoid cells in tunica propria even though it is absent from Sertoli cells at the site of the BTB. Furthermore, BCRP is also expressed spatiotemporally by Sertoli cells and step 19 spermatids in the rat testis and stage-specifically, limiting to stage VII?VIII of the epithelial cycle, and restricted to the apical ectoplasmic specialization [apical ES, a testis-specific F-actin-rich adherens junction (AJ)]. Interestingly, adjudin was recently shown to be capable of downregulating BCRP expression at the apical ES. In this Opinion article, we critically discuss the latest findings on BCRP; in particular, we provide some findings utilizing molecular modeling to define the interacting domains of BCRP with adjudin. Based on this information, it is hoped that the next generation of adjudin analogs to be synthesized can improve their efficacy in downregulating BCRP and perhaps other drug efflux transporters in the testis to improve their efficacy to traverse the BTB by modifying their interacting domains.

Dataset Information

Regulation of BTB dynamics in spermatogenesis - insights from the adjudin toxicant model.

Publications

Regulation of BTB Dynamics in Spermatogenesis-Insights From the Adjudin Model.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets