Time-course Transcriptional Profiling of Human Amniotic Fluid-derived Stem Cells Using Microarray / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: To maintain the homeostasis of stem cells and prevent their ability to initiate tumorigenesis, it is important to identify and modify factors that prevent or accelerate stem cell senescence. We used microarrays to attempt to identify such factors in human amniotic fluid (HAF)-derived stem cells. MATERIALS AND METHODS: To identify gene expression changes over a time course, we compared gene expression profiles of HAF-derived stem cells in different passages (1st, 2nd, 4th, 6th, 8th, and 10th) using a Sentrix Human illumina microarray. RESULTS: Of the 25,804 genes in the microarray chip, 1,970 showed an over 2-fold change relative to the control (the 1st passage)-either upregulated or downregulated. Quantitative real-time PCR validated the microarray data for selected genes: markedly increased genes were CXCL12, cadherin 6 (CDH6), and folate receptor 3 (FOLR3). Downregulated genes included cyclin D2, keratin 8, insulin-like growth factor 2 (IGF2), natriuretic peptide precursor B (NPPB) and cellular retinoic acid binding protein 2 (CRABP2). The expression pattern of the selected genes was consistent with the microarray data except for CXCL12 and IGF2. Interestingly, the expression of NPPB was dramatically downregulated along the time course; it was almost completely shut-down by the 10th passage. In contrast, FOLR3 mRNA expression was dramatically increased. CONCLUSION: Taken together, although a function for NPPB and FOLR3 in stem cell senescence has not been reported, our results strongly suggest that NPPB and/or FOLR3 play a significant role in the regulation of stem cell senescence.

Implication of ADAM-8, -9, -10, -12, -15, -17, and ADAMTS-1 in Implantational Remodeling of a Mouse Uterus

In the present study, whether the ADAM-8, -9, -10, -12, -15, -17, and ADAMTS-1 proteins might play a role in mouse uterus during periimplantation period was investigated. Immunoblotting analyses demonstrated that all ADAM proteins consistently appeared throughout days 1 to 8 of pregnancy but with a variation depending on the species of ADAM gene, the progression of pregnancy, and the site of the uterus. Immunohistochemical analyses indicated that ADAM proteins were localized in the luminal or glandular epithelial layers with a varying intensity depending on the species of ADAM and the progression of pregnancy. Particularly ADAM-8, -12, and -15, were predominantly located in the implantation site of the uterine tissues, whereas little or no protein was localized in the interimplantation site. Based upon these observations, it is suggested that the ADAMs might play an important role in the remodeling of the mouse uterus during the periimplantation period.

Expression of ADAM-8, 9, 10, 12, 15, 17 and ADAMTS-1 Genes in Mouse Uterus During Periimplantation Period / 대한불임학회지

No abstract available.

Successful Birth after Transfer of Re-frozen Blastocysts Developed from Immature Oocytes Retrieved from a Woman with Polycystic Ovarian Syndrome / 대한불임학회지

No abstract available.

Protein Expression of Matrix Metalloproteinases of Mouse Reproductive Organs During Estrous Cycle / 대한불임학회지

Protein expression patterns of matrix metalloproteinases (MMPs) were examined in mouse reproductive organs during estrous cycle. Estrous cycle was classified into diestrus, proestrus, estrus or metestus and MMP expression was analyzed by zymography using gelatin as a substrate. Uterine fluid (UF) obtained both at diestrus and proestrus exhibited 4 major MMPs including 106kDa, 64kDa, 62kDa and 59kDa gelatinases. However, in UF at estrus, the gelatinolytic activity of 64kDa MMP disappeared and that of 106kDa and 62kDa MMPs dramatically decreased. At metestrus, 64kDa MMP activity reappeared and 106kDa and 62kDa MMP exhibited increased activities such that the band intensity of 106kDa was comparable to that in UF at diestrus. Gelatinolytic activity of 59kDa MMP was not changed throughout the cycle. Both ovarian and oviductal tissue homogenate revealed 4 MMPs which corresponded to the 4 MMPs of UF. However, unlike UF MMPs, gelatinolytic activity of these MMPs did not show distinct changes throughout the cycle. Either an inhibitor of MMP, 1, 10-phenanthroline, or a metal chelator, EDTA, abolished the appearance of the above MMP activities in gelatinated gel whereas a serine proteinase inhibitor, phenylmethylsulfonyl fluoride, failed to inhibit the appearance of MMP activities, proving that gelatinolytic activity of the above reproductive tissues were due to the enzymatic activity of MMP. When gelatinolytic activity of mouse serum was examined, it revealed 5 MMPs (131kDa, 106kDa, 89kDa, 64kDa and 62kDa bands) and one gelatinase (84kDa) band. From these results, it is concluded that the protein expression of MMPs of mouse reproductive organs, particularly uterus, is temporally regulated during estrous cycle and uterine 106kDa, 64kDa and 62kDa MMP,3 are suggested to play an important role in cyclic tissue remodeling of mouse uterus.

Effects of Somatic Cell Conditioned Medium on the Chymotrypsin Resistance of Mouse Oocytes / 대한불임학회지

Certain types of somatic cells, as well as follicular cumulus cells associating with mammalian oocytes, are known to produce beneficial effects on in vitro fertilization and preimplantation development of mammalian eggs when they are present in oocyte culture medium. To investigate the nature of the effects of somatic cells, the resistance of mouse oocytes against chymotrypsin treatment was examined after culture within various cell conditioned media. When mouse oocytes matured for 17-18 hr in the presence of cumulus cells were treated with 1% chymotrypsin, half of them remained still alive even after 240 min (t50>240.0). In contrast half of mouse oocytes cultured without cumulus cells underwent degeneration within 65.0 min (t50= 65.0+/-13.2 min) of the same treatment. To see if the effects were due to the secretory products of cumulus cells, mouse cumulus cells were cultured for 20 hr in medium containing 0.4% BSA and the supernatant of culture medium (conditioned medium) was taken. After maturation in the cumulus cell conditioned medium, mouse oocytes exhibited t50 = 190.0 +/-10.8 min upon chymotrypsin treatment whereas half of oocytes cultured without conditioned medium degenerated within 25.5 min. Human granulosa cell conditioned medium gave similar effects such that oocytes matured in conditioned medium exhibited t50 = 183.3+/-19.1 min while t50 of control group oocytes was 60.0+/-6.8 min. Oocytes matured in vero cell conditioned medium exhibited t50 = 196.7+/-8.8 min. On the other hand, amniotic cell conditioned medium resulted in the chymotrypsin resistance of t50 = 80.0+/-8.4 min which was not statistically different from the control value of t50 = 48.0+/-13.2 min. Based upon these results, it is suggested that certain somatic cell types including cumulus cells might change the biochemical properties of mouse oocyte membrane during meiotic maturation as revealed by the enhanced resistance against chymotrypsin treatment. Such effects of somatic cells appear to be mediated via the secretory products rather than direct communication between somatic cells and oocytes.