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Expression profile of genes identified in human spermatogonial stem cell‐like cells using suppression subtractive hybridization
Author(s) -
Yoo Jung Ki,
Lim Jung Jin,
Ko Jung Jae,
Lee Dong Ryul,
Kim Jin Kyeoung
Publication year - 2010
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.22588
Subject(s) - suppression subtractive hybridization , biology , gene , spermatogenesis , stem cell , cell growth , microbiology and biotechnology , gene expression , genetics , cdna library , endocrinology
Spermatogenesis is the process by which testicular spermatogonial stem cells (SSCs) self‐renew and differentiate into mature sperm in the testis. Maintaining healthy spermatogenesis requires proper proliferation of SSCs. In this study, we sought to identify factors that regulate the proliferation of SSCs. Human SSC (hSSC)‐like cells were isolated from azoospermic patients by a modified culture method and propagated in vitro. After four to five passages, the SSC‐like cells spontaneously ceased proliferating in vitro, so we collected proliferating (P)‐hSSC‐like cells at passage two and senescent (S)‐hSSC‐like cells at passage five. Suppression subtractive hybridization (SSH) was used to identify genes that were differentially expressed between the P‐hSSC‐like and S‐hSSC‐like cells. We selected positive clones up‐regulated in P‐hSSC‐like cells using SSH and functionally characterized them by reference to public databases using NCBI BLAST tools. Expression levels of genes corresponding to subtracted clones were analyzed using RT‐PCR. Finally, we confirmed the differential expression of 128 genes in positive clones of P‐hSSC‐like cells compared with S‐hSSC‐like cells and selected 23 known and 39 unknown clones for further study. Known genes were associated with diverse functions; 22% were related to metabolism. Fifteen of the known genes and two of the unknown genes were down‐regulated after senescence of hSSC‐like cells. A comparison with previous reports further suggests that known genes selected, SPP1 , may be related to germ cell biogenesis and cellular proliferation. Our findings identify several potential novel candidate biomarkers of proliferating‐ and senescencet‐hSSCs, and they provide potentially important insights into the function and characteristics of human SSCs. J. Cell. Biochem. 110: 752–762, 2010. © 2010 Wiley‐Liss, Inc.