In vitro differentiation of germ cells from nonobstructive azoospermic patients using three-dimensional culture in a collagen gel matrix

• Published in: Fertility and sterility Vol. 87; no. 4; pp. 824 - 833
• Main Authors: Lee, Jae-Ho, Ph.D, Gye, Myung C., Ph.D, Choi, Kyoo Wan, Ph.D, Hong, Jae Yup, M.D., Ph.D, Lee, Yong Bok, M.D, Park, Dong-Wook, Ph.D, Lee, Seung Jae, M.D, Min, Churl K., Ph.D
• Format: Journal Article
• Language: English
• Published: New York, NY Elsevier Inc 01-04-2007; Elsevier Science
• Subjects: Animals; Azoospermia - physiopathology; Biological and medical sciences; Birth control; Cell Differentiation; Cells, Cultured; Collagen - physiology; collagen gel matrix; Flow Cytometry; Follicle Stimulating Hormone - blood; Gels; Gynecology. Andrology. Obstetrics; Humans; Immunohistochemistry; In vitro spermatogenesis; Internal Medicine; Male; Medical sciences; Obstetrics and Gynecology; Rats; Rats, Sprague-Dawley; Spermatogenesis; Spermatogonia - cytology; Sterility. Assisted procreation; three-dimensional culture; three-dimensional culture; collagen gel matrix; In vitro spermatogenesis; Human; Spermatogenesis; Glycoprotein; Cell differentiation; In vitro; Azoospermia; Three dimensional culture; Fertility; Collagen; Male sterility; Extracellular matrix; Male genital diseases
• ISSN: 0015-0282; 1556-5653
• DOI: 10.1016/j.fertnstert.2006.09.015

Objective To assess the effectiveness of the three-dimensional culture of spermatogenic cells in a collagen gel matrix from nonobstructive azoospermic patients and examine the relation between the success rate of in vitro spermatogenesis and serum FSH level as a diagnostic prediction. Design Prospective study using radioimmunoassay, immunocytochemistry, and flow cytometry with primary cultured cells. Setting Gynecologic clinics and human reproduction research laboratory. Patient(s) Primary culture of spermatogenic cells established from 18 nonobstructive azoospermic patients who underwent histologic diagnoses. Intervention(s) Primary culture of spermatogenic cells in a collagen-based gel matrix, subjected to immunological and flow cytometric analyses. Main Outcome Measure(s) In vitro culture of spermatogenic cells was established in an extracellular milieu that more closely resembled the in vivo condition. The number of chromosomes in newly generated cells during culture was determined by fluorescence-activated cell sorter (FACS) and immunocytochemical analysis. Effects of FSH on the differentiation of the spermatogenic cells were measured. Result(s) Results of histologic studies indicated that 8 of 18 patients showed the spermatocyte arrest. Immunocytochemical and FACS analysis indicated that after 12 days in culture, haploid cells comprised 11%–37% of the cultured cell population with a characteristic expression of a cellular marker for spermatids. The serum level of FSH appeared to be closely correlated with an increase in the number of haploid cells in culture. Conclusion(s) The present three-dimensional culture in a collagen gel matrix provides a suitable means by which spermatocytes could be induced to differentiate into presumptive spermatids in vitro. In addition, the plasma FSH level could be a good indicator for the success of differentiation of cultured spermatogenic cells obtained from patients with spermatogenic arrest.