Human chorionic-plate-derived mesenchymal stem cells and Wharton's jelly-derived mesenchymal stem cells: a comparative analysis of their potential as placenta-derived stem cells.

Placenta-derived stem cells (PDSCs) have gained interest as an alternative source of stem cells for regenerative medicine because of their potential for self-renewal and differentiation and their immunomodulatory properties. Although many studies have characterized various PDSCs biologically, the properties of the self-renewal and differentiation potential among PDSCs have not yet been directly compared. We consider the characterization of chorionic-plate-derived mesenchymal stem cells (CP-MSCs) and Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs) among various PDSCs and the assessment of their differentiation potential to be important for future studies into the applicability and effectiveness of PDSCs in cell therapy. In the present study, the capacities for self-renewal and multipotent differentiation of CP-MSCs and WJ-MSC isolated from normal term placentas were compared. CP-MSCs and WJ-MSCs expressed mRNAs for the pluripotent stem cell markers Oct-4, Nanog, and Sox-2. Additionally, HLA-G for immunomodulatory effects was found to be expressed at both the mRNA and protein levels in both cell types. The CP-MSCs and WJ-MSCs also had the capacities to differentiate into cells of mesodermal (adipogenic and osteogenic) and endodermal (hepatogenic) lineages. Expression of adipogenesis-related genes was higher in CP-MSCs than in WJ-MSCs, whereas WJ-MSCs accumulated more mineralized matrix than CP-MSCs. The WJ-MSCs expressed more of CYP3A4 mRNA, a marker for mature hepatocytes, than CP-MSCs. Thus, we propose that CP-MSCs and WJ-MSCs are useful sources of cells for appropriate clinical applications in the treatment of various degenerative diseases.

Characterization of Fetal Tissue-derived Mesenchymal Stem Cells.

Mesenchymal stem cells (MSCs) have unique immunologic properties that may someday prove useful in cell-based therapy for various degenerative diseases. Its potential is limited, however, by several factors, including the rarity of these cells and difficulty in isolating them. To evaluate their potential as new sources for cell therapy, we isolated MSCs from human fetal tissue (hfMSC) derived from spontaneous abortus (8∼10 weeks) then studied their cell cycle and cell surface marker expression using a fluorescence-activated cell sorter (FACS), as well as the expression of differentiation markers using real-time polymerase chain reaction (RT-PCR). The hfMSCs were able to undergo PCR up to 20 times without displaying significant changes in morphology or expression of various stemness markers (Nanog and human telomerase reverse transcriptase [hAFP]), including germ layer markers (hNF68, alpha-cardiac actin, and hAFP). Also, teratomas were not seen in mice with severe combined immunodeficiency syndrome (SCID) that received a transplantation of hfMSCs with hTERT activity. The FACS analysis revealed that the majority of hfMSCs express mesenchymal markers CD13, CD44, CD71, CD90, CD105, CD253a, and HLA-ABC, but did not express CD31, CD34, CD38, CD45, and HLA-DR. Interestingly, hfMSCs derived from the cell membrane during early passages were negative for both HLA-ABC and HLA-DR, although HLA-ABC expression was detected during later passages (>20 passages). We found that hfMSCs could be differentiated into an osteogenic lineage; this was indicated by modulation of osteoblast markers specific for mRNA. We conclude that hfMSCs could be used as a new source of cells to treat patients with osteogenic diseases, as well as to understand the mechanisms of immunosuppression by MSCs.

Efficient cryopreservative conditions for cultivated limbal and conjunctival epithelial cells.

PURPOSE: To examine the effects of cryopreservation on the viability of cultivated corneal limbal and conjunctival epithelial cells and to evaluate the optimal conditions for cryopreservation. METHODS: The cultivated human limbal epithelial cells (HLECs) were stored in media including 20%, 50%, and 90% fetal bovine serum (FBS) and 10% dimethyl sulfoxide (DMSO) at -196 degrees C for 1 week. The cultivated rabbit conjunctival epithelial cells were stored in 10%, 20%, and 50% FBS with 10% glycerol or DMSO as a cryoprotectant at -196 degrees C for 1 week. After thawing, cell viability was assessed using the trypan blue vital staining and 3-[4,5-dimethylthiazol-2-yl]-2,5-dephenyl tetrazolium bromide (MTT) assay. Immunofluorescent staining was performed with cytokeratin 3/12 antibody. Colony-forming efficiency (CFE) was evaluated 2 weeks after culture. RESULTS: HLECs cryopreserved with 50% FBS showed the highest cell viability, whereas those with 20% FBS revealed the lowest survival rate (87.1% +/- 0.8% and 79.8% +/- 4.01%, respectively; P = 0.030). CFE of HLECs was 2.13 +/- 1.35%, 2.31 +/- 2.23%, and 1.94 +/- 0.72% in cells with 20%, 50%, and 90% FBS, respectively (P > 0.05). For conjunctival epithelial cells, the cell viability was the highest with 50% FBS and 10% glycerol (95.0% +/- 4.27%), and the lowest survival rate was observed in the condition of 10% FBS and 10% DMSO (80.0% +/- 5.49%). CFE of cryopreserved conjunctival epithelial cells was 14.1% +/- 1.9% in cells with 20% FBS and glycerol and 13.5% +/- 2.0% in those with 20% FBS and DMSO (P > 0.05). HLECs expressed CK3/12 after cryopreservation in all conditions examined. CONCLUSIONS: The best results were yielded by 50% FBS for cell viability in HLECs. Glycerol seems to be superior to DMSO in cell viability of the rabbit conjunctival epithelium after cryopreservation.