title: In vitro spermatogenesis by three-dimensional culture of rat testicular cells in collagen gel matrix.
abstract: In an effort to improve in vitro spermatogenesis by potentiating the interactions between developing germ cells, somatic cells, and the extracellular matrix (ECM), the efficiency of the germ cell-somatic cell coculture in a three-dimensional (3D) collagen gel matrix was examined. Cells isolated from rat seminiferous tubules 18 days after birth were cultured for 22 days in a monolayer without ECM, collagen gel (CG), or collagen+Matrigel (CGM). After culture, the viabilities of the cultured cells in the monolayer, CG, and CGM culture were 42.8%, 70.7% and 76.1%, respectively. Occludin-positive cells in a cyst-like structure were found in the ECM gel matrix together with 3beta hydroxysteroid dehydrogenase-positive cells, suggesting the presence of functional Sertoli cells and Leydig cells, respectively. Flow cytometric analysis of DNA content revealed a significant increase in the haploid cell population in the CG and CGM compared to the monolayer culture. Transition protein 2 (TP2) and protamine 2-positive cells were found together with a significant increase in TP2 mRNA levels in cells cultured in CG and CGM over those in monolayer culture, suggesting the occurrence of the post-meiotic differentiation of spermatogenetic cells. Taken together, a 3D in vitro culture system for testicular cells using a collagen gel matrix could enhance viability, meiosis, and post-meiotic differentiation of germ cells to presumptive differentiating spermatids.
authors.affiliation: Department of Biotechnology, College of Natural Sciences, Seoul Women's University, Seoul 139-774, Korea.
authors.forename: Jae Ho
authors.initials: JH
authors.lastname: Lee
authors.forename: Hyun Joo
authors.initials: HJ
authors.lastname: Kim
authors.forename: Haekwon
authors.initials: H
authors.lastname: Kim
authors.forename: Sang Jin
authors.initials: SJ
authors.lastname: Lee
authors.forename: Myung Chan
authors.initials: MC
authors.lastname: Gye
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics; D003094:Collagen; D005434:Flow Cytometry; D008297:Male; D016133:Polymerase Chain Reaction; D051381:Rats; D017207:Rats, Sprague-Dawley; D013091:Spermatogenesis; D013737:Testis / Q000166:cytology* / Q000378:metabolism
chemical_list: D002868:Chromosomal Proteins, Non-Histone; C053886:spermatid transition proteins; D003094:Collagen