scholarly journals Drug transporters and blood–testis barrier function

2011 ◽  
Vol 209 (3) ◽  
pp. 337-351 ◽  
Author(s):  
Linlin Su ◽  
Dolores D Mruk ◽  
Will M Lee ◽  
C Yan Cheng
Keyword(s):  
Sertoli Cell ◽  
Barrier Function ◽  
Drug Transporters ◽  
Organic Anion ◽  
Permeability Barrier ◽  
Host Immune System ◽  
Slc Transporters ◽  
Apical Compartment ◽  
Immunological Barrier ◽  
Blood Testis Barrier

The blood–testis barrier (BTB) creates an immunological barrier that segregates the seminiferous epithelium into the basal and apical compartment. Thus, meiosis I/II and post-meiotic germ cell development take place in a specialized microenvironment in the apical compartment behind the BTB and these events are being shielded from the host immune system. If unwanted drugs and/or chemicals enter the apical compartment from the microvessels in the interstitium via the basal compartment, efflux pumps (e.g. P-glycoprotein) located in Sertoli cells and/or spermatids can actively transport these molecules out of the apical compartment. However, the mechanism(s) by which influx pumps regulate the entry of drugs/chemicals into the apical compartment is not known. In this study, a solute carrier (SLC) transporter organic anion transporting polypeptide 3 (Oatp3, Slco1a5) was shown to be an integrated component of the N-cadherin-based adhesion complex at the BTB. However, a knockdown of Oatp3 alone or in combination with three other major Sertoli cell drug influx pumps, namely Slc22a5, Slco6b1, and Slco6c1, by RNAi using corresponding specific siRNA duplexes failed to perturb the Sertoli cell tight junction (TJ) permeability barrier function. Yet, the transport of [3H]adjudin, a potential male contraceptive that is considered a toxicant to spermatogenesis, across the BTB was impeded following the knockdown of either Oatp3 or all the four SLC transporters. In short, even though drug transporters (e.g. influx pumps) are integrated components of the adhesion protein complexes at the BTB, they are not involved in regulating the Sertoli cell TJ permeability barrier function, instead they are only involved in the transport of drugs, such as adjudin, across the immunological barrier at the BTB.

scholarly journals MiR-142-3p Inhibits TGF-β3-Induced Blood-Testis Barrier Impairment by Targeting Lethal Giant Larvae Homolog 2

2018 ◽  
Vol 46 (1) ◽  
pp. 253-268 ◽  
Author(s):  
Ying Xu ◽  
Weixing Wu ◽  
Yunxia Fan ◽  
Shuyi Jiang ◽  
Xiaoyu Jia ◽  
...  
Keyword(s):  
Barrier Function ◽  
Transforming Growth Factor ◽  
Luciferase Assay ◽  
Luciferase Reporter ◽  
Sodium Fluorescein ◽  
Permeability Barrier ◽  
Junction Protein ◽  
Lethal Giant Larvae ◽  
Blood Testis Barrier

Background/Aims: Transforming growth factor-β3 (TGF-β3) has been proved to perturb the blood-testis barrier (BTB) by accelerating junction protein endocytosis in Sertoli cells (SCs) to accommodate the traversing of preleptotene spermatocytes across the BTB around stage VIII in rat. Yet the molecular network underlying the impairment of TGF-β3 on BTB integrity is not fully elucidated. Our study herein was designed to investigate the participation of microRNA-142-3p (miR-142-3p), which has been reported to affect TGF-β3 signaling via different pathways, during BTB dynamics and the corresponding mechanisms. Methods: MiRNA mimic or agomiRNA was co-administered with or without TGF-β3 in the cultured SCs or in the rat testis. The SC permeability barrier function was reflected by measuring the transepithelial resistance (TER) and the permeability of the sodium fluorescein (Na-F). The BTB integrity was detected by the permeation of biotin. A luciferase reporter assay was used to testify the potential target of miR-142-3p, lethal giant larvae 2 (Lgl2). Laser capture microdissection (LCM) was applied to acquire cell components of different stages of seminiferious tubules, followed by detection of the expression levels of miR-142-3p, TGF-β3, and Lgl2 by qPCR. The SC barrier function was also detected as above in the presence of TGF-β3 after Lgl2 knockdown. Results: We revealed a reversion of TGF-β3-induced BTB impairment after miR-142-3p treatment both in vitro and in vivo. Meanwhile, the activation of Cdc42 and reduction in occludin aroused by TGF-β3 were also reversed by miR-142-3p. The predicted binding of miR-142-3p with 3’-untranslated region (3’-UTR) of Lgl2, was verified by the luciferase assay. Moreover, an increased Lgl2 level in TGF-β3-treated SCs was found and correlated stage-specific expressions of TGF-β3, miR-142-3p, and Lgl2 were revealed. Knockdown of Lgl2 in SCs was shown to partially antagonize the BTB disruption mediated by TGF-β3. Conclusions: Collectively, our results suggest a resistance of miR-142-3p on the BTB impairment caused by TGF-β3 during the seminiferous epithelial cycle by targeting Lgl2.

scholarly journals c-Yes regulates cell adhesion at the apical ectoplasmic specialization-blood-testis barrier axis via its effects on protein recruitment and distribution

2013 ◽  
Vol 304 (2) ◽  
pp. E145-E159 ◽  
Author(s):  
Xiang Xiao ◽  
Dolores D. Mruk ◽  
C. Yan Cheng
Keyword(s):  
Cell Adhesion ◽  
Sertoli Cell ◽  
Germ Cells ◽  
Early Stage ◽  
Seminiferous Epithelium ◽  
Permeability Barrier ◽  
Cell Interface ◽  
Ectoplasmic Specialization ◽  
Cell Cell ◽  
Blood Testis Barrier

During spermatogenesis, extensive restructuring takes place at the cell-cell interface since developing germ cells migrate progressively from the basal to the adluminal compartment of the seminiferous epithelium. Since germ cells per se are not motile cells, their movement relies almost exclusively on the Sertoli cell. Nonetheless, extensive exchanges in signaling take place between these cells in the seminiferous epithelium. c-Yes, a nonreceptor protein tyrosine kinase belonging to the Src family kinases (SFKs) and a crucial signaling protein, was recently shown to be upregulated at the Sertoli cell-cell interface at the blood-testis barrier (BTB) at stages VIII–IX of the seminiferous epithelial cycle of spermatogenesis. It was also highly expressed at the Sertoli cell-spermatid interface known as apical ectoplasmic specialization (apical ES) at stage V to early stage VIII of the epithelial cycle during spermiogenesis. Herein, it was shown that the knockdown of c-Yes by RNAi in vitro and in vivo affected both Sertoli cell adhesion at the BTB and spermatid adhesion at the apical ES, causing a disruption of the Sertoli cell tight junction-permeability barrier function, germ cell loss from the seminiferous epithelium, and also a loss of spermatid polarity. These effects were shown to be mediated by changes in distribution and/or localization of adhesion proteins at the BTB (e.g., occludin, N-cadherin) and at the apical ES (e.g., nectin-3) and possibly the result of changes in the underlying actin filaments at the BTB and the apical ES. These findings implicate that c-Yes is a likely target of male contraceptive research.

scholarly journals Regulation of spermiogenesis, spermiation and blood–testis barrier dynamics: novel insights from studies on Eps8 and Arp3

2011 ◽  
Vol 435 (3) ◽  
pp. 553-562 ◽  
Author(s):  
C. Yan Cheng ◽  
Dolores D. Mruk
Keyword(s):  
Sertoli Cell ◽  
Morphological Changes ◽  
Growth Factor Receptor ◽  
Adhesion Protein ◽  
Apical Compartment ◽  
Epidermal Growth ◽  
Actin Regulatory Proteins ◽  
Filament Network ◽  
Ectoplasmic Specialization ◽  
Blood Testis Barrier

Spermiogenesis in the mammalian testis is the most critical post-meiotic developmental event occurring during spermatogenesis in which haploid spermatids undergo extensive cellular, molecular and morphological changes to form spermatozoa. Spermatozoa are then released from the seminiferous epithelium at spermiation. At the same time, the BTB (blood–testis barrier) undergoes restructuring to facilitate the transit of preleptotene spermatocytes from the basal to the apical compartment. Thus meiotic divisions take place behind the BTB in the apical compartment to form spermatids. These germ cells enter spermiogenesis to transform into elongating spermatids and then into spermatozoa to replace those that were released in the previous cycle. However, the mole-cular regulators that control spermiogenesis, in particular the dynamic changes that occur at the Sertoli cell–spermatid interface and at the BTB, are not entirely known. This is largely due to the lack of suitable animal models which can be used to study these events. During the course of our investigation to develop adjudin [1-(2,4-dichlorobenzyl)-1H-indazole-3-carbohydrazide] as a potential male contraceptive, this drug was shown to ‘accelerate’ spermiation by inducing the release of premature spermatids from the epithelium. Using this model, we have identified several molecules that are crucial in regulating the actin filament network and the unique adhesion protein complex at the Sertoli cell–spermatid interface known as the apical ES (ectoplasmic specialization). In the present review, we critically evaluate these and other findings in the literature as they relate to the restricted temporal and spatial expression of two actin regulatory proteins, namely Eps8 (epidermal growth factor receptor pathway substrate 8) and Arp3 (actin-related protein 3), which regulate these events.

Barrière hématotesticulaire (blood–testis barrier) et variations de perméabilité du compartiment gonial dans le follicule testiculaire de Locusta migratoria implanté en milieu dépourvu d'ecdystéroïdes

1986 ◽  
Vol 64 (9) ◽  
pp. 2053-2061 ◽  
Author(s):  
C. Marcaillou ◽  
S. Lauverjat
Keyword(s):  
Locusta Migratoria ◽  
Permeability Barrier ◽  
Adult Males ◽  
Septate Junctions ◽  
Implantation Time ◽  
Fourth Larval Stage ◽  
Apical Compartment ◽  
Normal Spermatogenesis ◽  
Blood Testis Barrier

In Locusta migratoria, the testicular follicle is made on an apical compartment that contains the youngest germ cells and a basal one in which the spermatids differentiate. As soon as it is formed during the fourth larval stage, the latter compartment becomes impervious to exogenous macromolecules. This permeability barrier, which corresponds to the blood–testis barrier defined in mammals, depends on extensive septate junctions. In this paper, testicular follicles were cultured in vivo in adult males known to be free of hemolymphatic ecdysteroids. The permeability of the apical compartment to horseradish peroxidase, used as a tracer, varied as a function of implantation time. This apical compartment became impermeable as soon as the basal compartment cells were postmeiotic. Once implanted into an adult host, a follicle that contained only gonia completed normal spermatogenesis and established a blood–testis barrier, providing a tight compartment in which meiotic processes occurred. Septate junctions developed normally. These results show that ecdysterone controls neither the differentiation of septate junctions nor the establishment of the blood–testis barrier, which appear both to be conditioned by the initiation of meiosis.

scholarly journals Regulation of the blood-testis barrier by coxsackievirus and adenovirus receptor

2012 ◽  
Vol 303 (8) ◽  
pp. C843-C853 ◽  
Author(s):  
Linlin Su ◽  
Dolores D. Mruk ◽  
C. Yan Cheng
Keyword(s):  
Sertoli Cell ◽  
Sertoli Cells ◽  
Regulatory Protein ◽  
Integral Membrane Protein ◽  
Permeability Barrier ◽  
Structural Protein ◽  
Coxsackievirus And Adenovirus Receptor ◽  
Adenovirus Receptor ◽  
Blood Testis Barrier

The blood-testis barrier (BTB) divides the seminiferous epithelium into the basal and the adluminal compartment. It restricts paracellular diffusion of molecules between Sertoli cells, confers cell polarity, and creates a unique microenvironment in the adluminal compartment for spermatid development. However, it undergoes restructuring during the epithelial cycle so that preleptotene spermatocytes differentiated from type B spermatogonia residing in the basal compartment can traverse the BTB at stage VIII of the cycle, while the immunological barrier is maintained. Herein, coxsackievirus and adenovirus receptor (CAR), a tight junction (TJ) integral membrane protein in the testis and multiple epithelia and endothelia, was found to act as a regulatory protein at the BTB, besides serving as a structural adhesion protein. RNAi-mediated knockdown of CAR in a Sertoli cell epithelium with an established TJ-permeability barrier that mimicked the BTB in vivo resulted in a disruption of the TJ barrier and an increase in endocytosis of the TJ-protein occludin. Furthermore, such an enhancement in occludin endocytosis was accompanied by a downregulation of Thr-phosphorylation in occludin and an increase in the association of endocytosed occludin with early endosome antigen-1. These findings were confirmed by overexpressing CAR in Sertoli cells, which was found to “tighten” the Sertoli cell TJ barrier, promoting BTB function. These findings support the emerging concept that CAR is not only a structural protein, it is involved in conferring the phosphorylation status of other adhesion proteins at the BTB (e.g., occludin) possibly mediated via its structural interactions with nonreceptor protein kinases, thereby modulating endocytic vesicle-mediated protein trafficking.

scholarly journals rpS6 Regulates Blood-Testis Barrier Dynamics By Affecting F-Actin Organization and Protein Recruitment

Endocrinology ◽  
2012 ◽  
Vol 153 (10) ◽  
pp. 5036-5048 ◽  
Author(s):  
Ka-Wai Mok ◽  
Dolores D. Mruk ◽  
Bruno Silvestrini ◽  
C. Yan Cheng
Keyword(s):  
Sertoli Cell ◽  
Actin Filament ◽  
Sertoli Cells ◽  
Plasma Membranes ◽  
Permeability Barrier ◽  
Actin Organization ◽  
Filament Bundles ◽  
Mtorc1 Pathway ◽  
Protein Recruitment ◽  
Blood Testis Barrier

Abstract During spermatogenesis, preleptotene spermatocytes residing near the basement membrane of the seminiferous tubule must traverse the blood-testis barrier (BTB) at stage VIII–IX of the epithelial cycle to continue their development in the adluminal compartment. Unlike other blood-tissue barriers (e.g. the blood-brain barrier) that are created by the endothelial tight junction (TJ) barrier of capillaries, the BTB is created by specialized junctions between Sertoli cells in which TJ coexists with basal ectoplasmic specialization (basal ES, a testis-specific adherens junction). The basal ES is typified by the presence of tightly packed actin filament bundles sandwiched between cisternae of endoplasmic reticulum and the apposing plasma membranes of Sertoli cells. These actin filament bundles also confer unusual adhesive strength to the BTB. Yet the mechanisms by which these filamentous actin (F-actin) networks are regulated from the bundled to the debundled state to facilitate the transit of spermatocytes remain elusive. Herein, we provide evidence that ribosomal protein S6 (rpS6), the downstream signaling molecule of the mammalian target of rapamycin complex 1 (mTORC1) pathway, is a major regulator of F-actin organization and adhesion protein recruitment at the BTB. rpS6 is restrictively and spatiotemporally activated at the BTB during the epithelial cycle. An activation of rpS6 led to a disruption of the Sertoli cell TJ barrier and BTB integrity. Its silencing in vitro or in vivo by using small interfering RNA duplexes or short hairpin RNA was found to promote the Sertoli cell TJ permeability barrier by the recruitment of adhesion proteins (e.g. claudin-11 and occludin) to the BTB. Thus, rpS6 in the mTORC1 pathway regulates BTB restructuring via its effects on the F-actin organization and protein recruitment at the BTB.

Fluorochloridone perturbs blood-testis barrier/Sertoli cell barrier function through Arp3-mediated F-actin disruption

2018 ◽  
Vol 295 ◽  
pp. 277-287 ◽  
Author(s):  
Luqing Liu ◽  
Yubin Zhang ◽  
Xiuli Chang ◽  
Rui Li ◽  
Chunhua Wu ◽  
...  
Keyword(s):  
Sertoli Cell ◽  
Barrier Function ◽  
Blood Testis Barrier

scholarly journals Differential effects of c-Src and c-Yes on the endocytic vesicle-mediated trafficking events at the Sertoli cell blood-testis barrier: an in vitro study

2014 ◽  
Vol 307 (7) ◽  
pp. E553-E562 ◽  
Author(s):  
Xiang Xiao ◽  
Dolores D. Mruk ◽  
Elissa W. P. Wong ◽  
Will M. Lee ◽  
Daishu Han ◽  
...  
Keyword(s):  
Sertoli Cell ◽  
Germ Cells ◽  
Sertoli Cells ◽  
Protein Trafficking ◽  
In Vitro Study ◽  
Underlying Mechanism ◽  
Endocytic Vesicle ◽  
Immunological Barrier ◽  
Blood Testis Barrier

The blood-testis barrier (BTB) is one of the tightest blood-tissue barriers in the mammalian body. However, it undergoes cyclic restructuring during the epithelial cycle of spermatogenesis in which the “old” BTB located above the preleptotene spermatocytes being transported across the immunological barrier is “disassembled,” whereas the “new” BTB found behind these germ cells is rapidly “reassembled,” i.e., mediated by endocytic vesicle-mediated protein trafficking events. Thus, the immunological barrier is maintained when preleptotene spermatocytes connected in clones via intercellular bridges are transported across the BTB. Yet the underlying mechanism(s) in particular the involving regulatory molecules that coordinate these events remains unknown. We hypothesized that c-Src and c-Yes might work in contrasting roles in endocytic vesicle-mediated trafficking, serving as molecular switches, to effectively disassemble and reassemble the old and the new BTB, respectively, to facilitate preleptotene spermatocyte transport across the BTB. Following siRNA-mediated specific knockdown of c-Src or c-Yes in Sertoli cells, we utilized biochemical assays to assess the changes in protein endocytosis, recycling, degradation and phagocytosis. c-Yes was found to promote endocytosed integral membrane BTB proteins to the pathway of transcytosis and recycling so that internalized proteins could be effectively used to assemble new BTB from the disassembling old BTB, whereas c-Src promotes endocytosed Sertoli cell BTB proteins to endosome-mediated protein degradation for the degeneration of the old BTB. By using fluorescence beads mimicking apoptotic germ cells, Sertoli cells were found to engulf beads via c-Src-mediated phagocytosis. A hypothetical model that serves as the framework for future investigation is thus proposed.

scholarly journals An Occludin-Focal Adhesion Kinase Protein Complex at the Blood-Testis Barrier: A Study Using the Cadmium Model

Endocrinology ◽  
2009 ◽  
Vol 150 (7) ◽  
pp. 3336-3344 ◽  
Author(s):  
Erica R. Siu ◽  
Elissa W. P. Wong ◽  
Dolores D. Mruk ◽  
K. L. Sze ◽  
Catarina S. Porto ◽  
...  
Keyword(s):  
Focal Adhesion Kinase ◽  
Sertoli Cell ◽  
Focal Adhesion ◽  
Adhesion Molecule ◽  
Protein Complex ◽  
Permeability Barrier ◽  
Ectoplasmic Specialization ◽  
Blood Testis Barrier

Several integral membrane proteins that constitute the blood-testis barrier (BTB) in mammalian testes, in particular rodents, are known to date. These include tight junction (TJ) proteins (e.g. occludin, junctional adhesion molecule-A, claudins), basal ectoplasmic specialization proteins (e.g. N-cadherin), and gap junction proteins (e.g. connexin43). However, the regulators (e.g. protein kinases and phosphatases) that affect these proteins, such as their interaction with the cytoskeletal actin, which in turn confer cell adhesion at the TJ, remain largely unknown. We report herein that focal adhesion kinase (FAK) is a putative interacting partner of occludin, but not claudin-11 or junctional adhesion molecule-A. Immunohistochemistry and fluorescence microscopy studies illustrated that the expression of FAK in the seminiferous epithelium of adult rat testes was stage specific. FAK colocalized with occludin at the BTB in virtually all stages of the seminiferous epithelial cycle but considerably diminished in stages VIII–IX, at the time of BTB restructuring to facilitate the transit of primary leptotene spermatocytes. Using Sertoli cells cultured in vitro with established TJ-permeability barrier and ultrastructures of TJ, basal ectoplasmic specialization and desmosome-like junction that mimicked the BTB in vivo, FAK was shown to colocalize with occludin and zonula occludens-1 (ZO-1) at the Sertoli-Sertoli cell interface. When these Sertoli cell cultures were treated with CdCl2 to perturb the TJ-barrier function, occludin underwent endocytic-mediated internalization in parallel with FAK and ZO-1. Thus, these findings demonstrate that FAK is an integrated regulatory component of the occludin-ZO-1 protein complex, suggesting that functional studies can be performed to study the role of FAK in BTB dynamics.

scholarly journals Filamin A Is a Regulator of Blood-Testis Barrier Assembly during Postnatal Development in the Rat Testis

Endocrinology ◽  
2012 ◽  
Vol 153 (10) ◽  
pp. 5023-5035 ◽  
Author(s):  
Wenhui Su ◽  
Dolores D. Mruk ◽  
Pearl P. Y. Lie ◽  
Wing-yee Lui ◽  
C. Yan Cheng
Keyword(s):  
Postnatal Development ◽  
Sertoli Cell ◽  
Actin Filament ◽  
Permeability Barrier ◽  
Filamin A ◽  
Rat Testis ◽  
Filament Network ◽  
Blood Testis Barrier

Abstract The blood-testis barrier (BTB) is an important ultrastructure in the testis. A delay in its assembly during postnatal development leads to meiotic arrest. Also, a disruption of the BTB by toxicants in adult rats leads to a failure in spermatogonial differentiation. However, the regulation of BTB assembly remains unknown. Herein, filamin A, an actin filament cross-linker that is known to maintain and regulate cytoskeleton structure and function in other epithelia, was shown to be highly expressed during the assembly of Sertoli cell BTB in vitro and postnatal development of BTB in vivo, perhaps being used to maintain the actin filament network at the BTB. A knockdown of filamin A by RNA interference was found to partially perturb the Sertoli cell tight junction (TJ) permeability barrier both in vitro and in vivo. Interestingly, this down-regulating effect on the TJ barrier function after the knockdown of filamin A was associated with a mis-localization of both TJ and basal ectoplasmic specialization proteins. Filamin A knockdown also induced a disorganization of the actin filament network in Sertoli cells in vitro and in vivo. Collectively, these findings illustrate that filamin A regulates BTB assembly by recruiting these proteins to the microenvironment in the seminiferous epithelium to serve as the building blocks. In short, filamin A participates in BTB assembly by regulating protein recruitment during postnatal development in the rat testis.

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