Correspondence: Professor Dr Abdul Rahman Hayati, MBChB (Alexandria), DCP (London), FAM (Malaysia), Department of Pathology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Jalan Yaacob Latif, Bandar Tun Razak, 56000 Cheras, Kuala Lumpur, Malaysia. E-mail: hayati_ar@ymail.com (Received 20 May 2010; accepted 15 December 2010) Human chorion-derived stem cells: changes in stem cell properties during serial passage MOHD-MANZOR NUR FARIHA 1 , KIEN-HUI CHUA 2 , GEOK-CHIN TAN 1 , AY-EENG TAN 3 & ABDUL-RAHMAN HAYATI 1 1 Department of Pathology, 2 Department of Physiology and 3 Department of Obstetrics and Gynecology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia Abstract Background aims. Fetal membrane from human placenta tissue has been described as a potential source of stem cells. Despite abundant literature on amnion stem cells, there are limited studies on the stem cell properties of chorion-derived stem cells. Methods. The main aim was to determine the stemness properties of serial-passaged human chorion-derived stem cells (hCDSC). Quantitative polymerase chain reaction (PCR) was performed to reveal the following stemness gene expression in serial-passaged hCDSC: Oct-4, Sox-2, FGF-4, Rex-1, TERT, Nanog (3), Nestin, FZD-9, ABCG-2 and BST-1. Cell growth rate was evaluated from passage (P) 1 until P5. The colony-forming unit–fibroblast (CFU-F) frequency of P3 and P5 cells and multilineage differentiation potential of P5 cells were determined. The immunophenotype of hCDSC was compared using the surface markers CD9, CD31, CD34, CD44, CD45, CD73, CD90, CD117, HLA-ABC and HLA-DR, -DP and -DQ. Immunostaining for trophoblast markers was done on P0, P1, P3 and P5 cells to detect the contamination of trophoblasts in culture, while chromosomal abnormality was screened by cytogenetic analysis of P5 cells. Results. The surface markers for mesenchymal lineage in hCDSC were more highly expressed at P5 compared with P3 and P0, indicating the increased purity of these stem cells after serial passage. Indeed, all the stemness genes except TERT were expressed at P1, P3 and P5 hCDSC. Furthermore, human chorion contained high clonogenic precursors with a 1:30 CFU-F frequency. Successful adipogenic, chondrogenic and osteogenic differentiation demonstrated the multilineage potential of hCDSC. The karyotyping analysis showed hCDSC maintained chromosomal stability after serial passage. Conclusions. hCDSC retain multipotent potential even at later passages, hence are a promising source for cell therapy in the future. Key Words: cell differentiation, fetal stem cells, gene expression, human placenta, immunophenotype, quantitative polymerase chain reaction, serial passage Introduction Adult stem cells have been considered to be devel- opmentally committed and therefore restricted to produce specific cell lineages, namely those from the tissue in which the stem cell resides (1). Adult stem cells have been isolated from a variety of sources, such as bone marrow (BM) (2), lipoaspirate (3,4) and dif- ferent parts of the placenta, the amnion (5), chorion, chorionic villi (6,7) and umbilical cord (8). Although many types of stem cells are available, the sources of stem cells selected for clinical use should possess the ability to renew, be easily isolated and be pluripotent, and have minimal ethical controversies. These proper- ties are seen in human term placental-derived stem cells. These stem cells are available in abundance and do not involve any invasive procedure when harvesting. Because the placentas are mostly discarded anyway, isolating stem cells from these ‘waste’ tissues will not be of serious ethical concerns. Recent reports have shown that mesenchymal stromal cells (MSC) from the placenta do not elicit any immunologic reaction and could be used as autologous grafts for fetuses and newborns in peripartum tissue regen- eration as well as in utero transplantation to treat genetic disorders (9,10). The chorion is the membrane that is attached to the amnion by a spongy layer of loosely arranged collagen fibers. It consists of the mesodermal and trophoblastic region and is easily separated from the amnion (11). Some studies have characterized the stromal-associated surface markers, revealing the neurogenic, chondrogenic, osteogenic, adipogenic, myogenic (6) and angiogenic potency of chorion cells (12,13). However, the immunophenotypes of Cytotherapy, 2011; 13: 582–593 ISSN 1465-3249 print/ISSN 1477-2566 online © 2011 Informa Healthcare DOI: 10.3109/14653249.2010.549121 Cytotherapy Downloaded from informahealthcare.com by Universiti Kebangsaan Malaysia on 05/20/11 For personal use only.