The biology of blood-testis barrier dynamics

血睾屏障动力学的生物学

• 批准号: 8081158
• 负责人: C. Yan Cheng
• 金额: $ 7.16万
• 依托单位: POPULATION COUNCIL
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8081158
• 关键词: Address; Adult; Androgen Receptor; Androgens; Apical; Back; Basement membrane; Biochemical; Biochemistry; Biological; Biology; Blood-Testis Barrier; Cell Adhesion; Cell physiology; Cell surface; Cells; Cellular biology; Cicatrix; Coculture Techniques; Collagen; Endocytosis; Endosomes; Epithelial; Epithelium; Extracellular Matrix; Extracellular Matrix Proteins; Gelatinase B; Germ Cells; Homeostasis; In Vitro; Infertility; Integral Membrane Protein; Kinetics; Knockout Mice; Laboratories; Male Contraceptions; Male Contraceptive Agents; Male Infertility; Matrix Metalloproteinases; Mediating; Meiosis; Membrane Proteins; Metalloproteases; Molecular; Molecular Biology; Mus; Peptide Hydrolases; Permeability; Pharmaceutical Preparations; Production; Property; Protease Inhibitor; Proteins; Rattus; Recombinants; Recycling; Regulation; Research; Research Personnel; Site; Spermatocytes; Spermatogenesis; Staging; Study models; TNF gene; Techniques; Testing; Testis; Testosterone; Tight Junctions; Time; Tissue Inhibitor of Metalloproteinase-1; Tissues; Transforming Growth Factors; Tumor Necrosis Factor-alpha; Tumor Necrosis Factors; Work; base; cell motility; cytokine; design; in vitro Model; in vivo; inhibitor/antagonist; innovation; insight; late endosome; leydig interstitial cell; migration; mouse model; novel; novel strategies; occludin; public health relevance; sertoli cell; spermatogenic epithelium structure; tool; transcytosis

DESCRIPTION (provided by applicant): The long-term objective of this proposal is to understand the biology and regulation of blood-testis barrier (BTB) dynamics during spermatogenesis using the adult rat testis as a study model. Specifically, it seeks to unravel the mechanism(s) that maintains BTB integrity as preleptotene and leptotene spermatocytes traverse the BTB that occurs at stage VIII of the seminiferous epithelial cycle of spermatogenesis. This study is important since it answers an open question in the field regarding the biochemical and/or molecular mechanism(s) utilized by the testis that regulates the timely 'restructuring' of the BTB to permit spermatocyte transit while maintaining the immunological barrier function at the BTB. This information is also helpful to develop innovative approaches to design new drugs (e.g., male contraceptives that exert their effects by disrupting germ cell function in the seminiferous epithelium) that can be targeted to the seminiferous epithelium behind the BTB. Recent studies have shown that cytokines (e.g., tumor necrosis factor ?, TNF?, and transforming growth factor-?2/-3, TGF-?2 or -?3) have a disruptive effect on BTB integrity, whereas testosterone promotes BTB function, illustrating that the opposing effects of these molecules are crucial to maintain the timely "opening" and "closing" of the BTB during spermatogenesis while facilitating germ cell movement across the barrier. Since both TNF? and TGF-?3 are products of Sertoli and germ cells, and testosterone is a product of Leydig cells (but the androgen receptor resides mostly on the Sertoli cell), in the testis, it is likely that these molecules are released into the BTB microenvironment during spermatogenesis to regulate BTB dynamics. In this application, the P.I. will examine the hypothesis that TNF? regulates BTB dynamics via its effects on basement membrane proteins, such as collagens, proteases and protease inhibitors, which, in turn, regulates the steady-state levels of integral membrane proteins at the BTB. This will determine the status of the BTB to facilitate or prohibit the transit of preleptotene/leptotene spermatocytes across the barrier (Specific Aim 1). Furthermore, the P.I. will test the hypothesis that cytokines and testosterone mediate their opposing effects on the BTB via their differential actions on endocytosis, recycling, endosome-mediated intracellular degradation and transcytosis/relocation of integral membrane proteins at the BTB (Specific Aim 2). The proposed studies will be performed using cutting-edge techniques of biochemistry, molecular biology, and cell biology. The P.I. will also develop and characterize a novel in vitro model using Sertoli-germ cell cocultures to investigate the regulation of germ cell movement across the BTB, which can become a valuable research tool for investigators in the field. In short, this proposal addresses a long-standing mystery in spermatogenesis regarding the biology and regulation of the BTB during the epithelial cycle. PUBLIC HEALTH RELEVANCE: This project seeks to understand the mechanism(s) that regulates blood-testis barrier (BTB) dynamics during the seminiferous epithelial cycle of spermatogenesis using the adult rat testis as a study model. The BTB must `open' or `restructure' transiently during spermatogenesis to facilitate the transit of spermatocytes across the barrier. This, in turn, poses a unique opportunity to transport a drug, such as a male contraceptive that disrupts cell adhesion, behind the BTB to exert its effects, if the mechanism(s) that regulates the timely opening of the BTB is known. In short, this proposal seeks to understand the biology of BTB dynamics during spermatogenesis. It also offers new insight into: (i) developing novel approaches for male contraception, and (ii) the causes of unexplained male infertility.

描述（由申请人提供）：该提案的长期目标是使用成年大鼠睾丸作为研究模型，了解精子发生过程中血睾屏障（BTB）动力学的生物学和调节。具体来说，它试图揭示维持 BTB 完整性的机制，因为前细线期和细线期精母细胞穿过发生在生精上皮周期第八阶段的 BTB。这项研究很重要，因为它回答了该领域的一个悬而未决的问题，即睾丸利用的生化和/或分子机制调节 BTB 的及时“重组”，以允许精母细胞运输，同时维持 BTB 的免疫屏障功能。这些信息还有助于开发创新方法来设计新药（例如，通过破坏生精上皮中的生殖细胞功能发挥作用的男性避孕药），这些药物可以靶向 BTB 后面的生精上皮。最近的研究表明，细胞因子（例如肿瘤坏死因子α、TNFα和转化生长因子-β2/-3、TGF-β2或-β3）对BTB完整性具有破坏性作用，而睾酮则促进BTB功能，这说明这些分子的相反作用对于维持精子发生过程中BTB的及时“打开”和“关闭”至关重要 同时促进生殖细胞穿过屏障的运动。既然都有TNF？ TGF-β3和TGF-β3是支持细胞和生殖细胞的产物，而睾酮是间质细胞的产物（但雄激素受体主要驻留在支持细胞上），在睾丸中，这些分子很可能在精子发生过程中释放到BTB微环境中以调节BTB动态。在此应用中，P.I.将检验TNF？通过对基底膜蛋白（例如胶原蛋白、蛋白酶和蛋白酶抑制剂）的影响来调节 BTB 动态，而基底膜蛋白反过来又调节 BTB 内膜蛋白的稳态水平。这将确定 BTB 的状态，以促进或阻止前细线期/细线期精母细胞穿过屏障（具体目标 1）。此外，P.I.将检验以下假设：细胞因子和睾酮通过对 BTB 内吞作用、再循环、内体介导的细胞内降解和整合膜蛋白的转胞吞作用/重新定位的不同作用来介导对 BTB 的相反作用（具体目标 2）。拟议的研究将利用生物化学、分子生物学和细胞生物学的尖端技术进行。 P.I.还将开发和表征一种新型体外模型，使用支持细胞共培养来研究生殖细胞在 BTB 中运动的调节，这可以成为该领域研究人员的宝贵研究工具。简而言之，该提案解决了精子发生中长期存在的一个谜团，即上皮周期中 BTB 的生物学和调节。公共健康相关性：该项目旨在以成年大鼠睾丸为研究模型，了解生精上皮周期生精过程中血睾屏障 (BTB) 动态的调节机制。 BTB 必须在精子发生过程中短暂“打开”或“重组”，以促进精母细胞穿过屏障。如果调节 BTB 及时打开的机制已知，这反过来又提供了一个独特的机会，可以在 BTB 后面运输药物（例如破坏细胞粘附的男性避孕药）以发挥其作用。简而言之，该提案旨在了解精子发生过程中 BTB 动力学的生物学原理。它还提供了以下方面的新见解：(i) 开发男性避孕的新方法，以及 (ii) 无法解释的男性不育的原因。