A Robust and Highly Efficient Approach for Isolation of Mesenchymal Stem Cells From Wharton's Jelly for Tissue Repair.

Mesenchymal stem cells derived from umbilical cord Wharton's Jelly (WJ-MSCs) are emerging as promising therapeutics for a variety of diseases due to their ability of regeneration and immunomodulation, and their non-tumorigenic and non-immunogenic properties. Although multiple protocols have been developed for WJ-MSC isolation, insufficient cell numbers, heterogeneous cell population, and variations in procedures between different laboratories impede further clinical applications. Here, we compared six widely used WJ-MSC isolation methods regarding cell morphology, yield, purity, proliferation rate, and differentiation potential. Based on these analyses, we identified that the inefficiency of the extracellular matrix digestion results in low cell yield. Thus, we developed a new method called "Mince-Soak-Digest (MSD)" to isolate MSCs from WJ by incorporating a soaking step to facilitate the digestion of the extracellular matrix and release of the cells. Our newly developed method generates significantly higher cell yield (4- to 10-fold higher) than six widely used methods that we tested with high purity and consistency. Importantly, by transplantation of WJ-MSCs to the rat uterus, we repair the endometrial injury and restore the fertility of the rats. In conclusion, our results provide a robust and highly efficient approach for the isolation of WJ-MSCs to restore injured tissue. The higher efficiency of MSD assures the abundance of WJ-MSCs for clinical applications. Furthermore, the reliability of MSD contributes to the standardization of WJ-MSC isolation, which eliminates the discrepancies due to isolation procedures, thus facilitating the evaluation of the efficacy of WJ-MSCs across various human clinical applications.