An in vivo model to study blood-testis barrier dynamics

研究血睾屏障动力学的体内模型

• 批准号: 7485598
• 负责人: C. Yan Cheng
• 金额: $ 7.09万
• 依托单位: POPULATION COUNCIL
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-15 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7485598
• 关键词: Abbreviations; Address; Adherens Junction; Adult; Amino Acids; Apical; Bacitracin; Biochemical; Biological; Biology; Blood-Testis Barrier; Bos taurus; Bovine Serum Albumin; Butadiene; Cadmium; Cattle; Cells; DNA Nucleotidylexotransferase; Development; Dimethyl Sulfoxide; Disruption; Eagles; Electrical Resistance; Epidermal Growth Factor; Epithelial; Event; Extracellular Signal Regulated Kinases; Fibroblast Growth Factor 2; Fluorescein; Fluorescein-5-isothiocyanate; Fluoresceins; Follicle Stimulating Hormone; Gap Junctions; Gentamicins; Germ Cells; Glycerol; Goals; HEPES; Human; Imidazole; Immunohistochemistry; In Situ Nick-End Labeling; In Vitro; Infertility; Inhibitory Concentration 50; Insulin; Integral Membrane Protein; Interferons; Isothiocyanates; Kinetics; Label; Laboratory Study; Liquid substance; Luteinizing Hormone; MAP Kinase Gene; MDCK cell; Mediating; Mitogen-Activated Protein Kinases; Mitogens; Modeling; Molecular; Movement; Nutrient; Oligodendroglia; Pathway interactions; Peptides; Permeability; Phase; Physiological; Polymerase Chain Reaction; Proteins; Rattus; Recovery; Regulation; Research Personnel; Reverse Transcriptase Polymerase Chain Reaction; Reverse Transcription; STF; Spermatids; Spermatocytes; Spermatogenesis; Spermatogonia; Staging; Structure of rete testis; Study models; Testis; Tight Junctions; TimeLine; Title; Transferrin; Transforming Growth Factors; Tubular formation; Tumor Necrosis Factor-alpha; Tumor Necrosis Factors; U-0126; base; cell motility; cytokine; cytotoxic; extracellular; human MAPK14 protein; human TNF protein; in vivo; in vivo Model; inhibitor/antagonist; innovation; junctional adhesion molecule; kinase inhibitor; membrane-associated guanylate kinase; mitogen-activated protein kinase p38; myotubularin; novel; occludin; outer surface lipoprotein; response; sertoli cell; spermatogenic epithelium structure; synthetic peptide; toxicant; upstream kinase

DESCRIPTION (provided by applicant): During spermatogenesis, developing preleptotene spermatocytes residing in the basal compartment of the seminiferous epithelium must traverse the blood-testis barrier (BTB) at stage VIII of the epithelial cycle in adult rat testes, entering the adluminal compartment for further development. Without this timely movement of developing germ cells across the BTB, spermatogenesis will be disrupted, leading to infertility. While this cellular phenomenon pertinent to spermatogenesis is known for decades, the mechanism(s) that regulates BTB dynamics to facilitate germ cells to traverse the BTB is entirely unknown. This by and large is due to the lack of a suitable in vivo model to study BTB dynamics. In this application, the P.I. proposes to develop and extensively characterize a novel model to meet this need. In brief, local administration of a 22-amino acid synthetic peptide based on the second extracellular loop of occludin, a tight junction (TJ)-integral membrane protein at the BTB, to adult rat testes was shown to induce reversible disruption of BTB in the seminiferous epithelium. Also, this event was associated with changes in the expression of several target proteins (e.g., transforming growth factor ¿-3, TGF-¿3, and tumor necrosis factor a, TNFa) that mimicked the Sertoli cell TJ-barrier restructuring events in vitro. Perhaps the most important of all, this local occludin peptide treatment also induced reversible germ cell loss (in particular spermatids and spermatocytes, but not spermatogonia) from the seminiferous epithelium. The P.I. seeks to extensively characterize this novel in vivo model by delineating the detailed timeline of cellular, molecular and biochemical changes in the seminiferous epithelium correlating with the status of spermatogenesis and the integrity of the BTB. Results of these studies will yield a reliable study model for investigators in the field to understand the mechanism and regulation of BTB restructuring during spermatogenesis. Our primary goal is to develop an innovative in vivo model to study BTB dynamics which is significantly different from currently available study models using toxicants (e.g., cadmium and glycerol). First, the peptide that can induce BTB restructuring is non-cytotoxic. Second and perhaps most importantly, the disrupted BTB can be "resealed", making this model uniquely suitable to study the biology and regulation of BTB re-assembly during spermatogenesis.

描述（由申请人提供）：在精子发生过程中，位于生精上皮基底室中的正在发育的前瘦素精母细胞必须在成年大鼠睾丸上皮周期的第八阶段穿过血睾屏障（BTB），进入腔内室进行进一步发育。如果发育中的生殖细胞不能及时穿过 BTB，精子发生就会受到干扰，从而导致不育。虽然这种与精子发生相关的细胞现象已为人所知数十年，但调节 BTB 动力学以促进生殖细胞穿过 BTB 的机制却完全未知。这总的来说是由于缺乏合适的体内模型来研究 BTB 动力学。在此应用中，P.I.建议开发并广泛表征一种新颖的模型来满足这一需求。简而言之，对成年大鼠睾丸局部施用基于 Occludin（BTB 上的紧密连接 (TJ) 整合膜蛋白）第二个细胞外环的 22 个氨基酸合成肽，可诱导生精上皮中 BTB 的可逆破坏。此外，该事件与几种靶蛋白（例如转化生长因子 ¿-3、TGF-¿3 和肿瘤坏死因子 a、TNFa）表达的变化有关，这些靶蛋白在体外模拟支持细胞 TJ 屏障重组事件。也许最重要的是，这种局部闭合蛋白肽治疗还诱导生精上皮的可逆性生殖细胞损失（特别是精细胞和精母细胞，但不是精原细胞）。 P.I.试图通过描绘与精子发生状态和 BTB 完整性相关的生精上皮细胞、分子和生化变化的详细时间线来广泛描述这种新颖的体内模型。这些研究结果将为该领域的研究人员提供可靠的研究模型，以了解精子发生过程中 BTB 重组的机制和调节。我们的主要目标是开发一种创新的体内模型来研究 BTB 动力学，该模型与目前使用有毒物质（例如镉和甘油）的研究模型显着不同。首先，能够诱导BTB重组的肽是非细胞毒性的。其次，或许也是最重要的一点，被破坏的 BTB 可以被“重新密封”，使得该模型特别适合研究精子发生过程中 BTB 重新组装的生物学和调控。