title: Reprogramming of fibroblasts from older women with pelvic floor disorders alters cellular behavior associated with donor age.
abstract: We aimed to derive induced pluripotent stem cell (iPSC) lines from vaginal fibroblasts from older women with pelvic organ prolapse. We examined the effect of donor age on iPSCs and on the cells redifferentiated from these iPSCs. Vaginal fibroblasts were isolated from younger and older subjects for reprogramming. iPSCs were generated simultaneously using an excisable polycistronic lentiviral vector expressing Oct4, Klf4, Sox2, and cMyc. The pluripotent markers of iPSCs were confirmed by immunocytochemistry and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Spectral karyotyping was performed. The ability of the iPSCs to differentiate into three germ layers was confirmed by embryoid body and teratoma formation. Senescence marker (p21, p53, and Bax) expressions were determined by qRT-PCR and Western blot. The iPSCs were redifferentiated to fibroblasts and were evaluated with senescence-associated β-galactosidase (SA) activity and mitotic index using time-lapse dark-field microscopy. iPSCs derived from both the younger and older subjects expressed pluripotency markers and showed normal karyotype and positive teratoma assays. There was no significant difference in expression of senescence and apoptosis markers (p21, p53, and Bax) in iPSCs derived from the younger subject compared with the older subject. Furthermore, fibroblasts redifferentiated from these iPSCs did not differ in SA activity or mitotic index. We report successful derivation of iPSCs from women with pelvic organ prolapse. Older age did not interfere with successful reprogramming. Donor age differences were not observed in these iPSCs using standard senescence markers, and donor age did not appear to affect cell mitotic activity in fibroblasts redifferentiated from iPSCs.
authors: Department of Obstetrics/Gynecology, Stanford University School of Medicine, Stanford, California 94305-5317, USA. yanwen@stanford.edu
authors: Yan
authors: Y
authors: Wen
authors: Prachi
authors: P
authors: Wani
authors: Lu
authors: L
authors: Zhou
authors: Tom
authors: T
authors: Baer
authors: Smruti Madan
authors: SM
authors: Phadnis
authors: Renee A
authors: RA
authors: Reijo Pera
authors: Bertha
authors: B
authors: Chen
mesh_terms: D053687:Adult Stem Cells / Q000166:cytology* / Q000502:physiology; D000367:Age Factors; D000368:Aged; D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D016922:Cellular Senescence / Q000502:physiology; D058732:Embryoid Bodies / Q000166:cytology; D005260:Female; D005347:Fibroblasts / Q000166:cytology* / Q000502:physiology; D005822:Genetic Vectors; D006801:Humans; D007621:Karyotyping; D000090062:Kruppel-Like Factor 4; D016086:Lentivirus / Q000235:genetics; D051379:Mice; D008875:Middle Aged; D008938:Mitosis / Q000502:physiology; D059952:Pelvic Floor Disorders / Q000473:pathology*; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000502:physiology; D061251:Primary Cell Culture; D013724:Teratoma / Q000473:pathology; D016159:Tumor Suppressor Protein p53 / Q000235:genetics; D014621:Vagina / Q000166:cytology*
chemical_list: C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53