β-galactosidase and Annexin V/PI Assays as QC Tests for Manufacturing cGMP-Grade Wharton Jelly Derived Mesenchymal Stem Cells

Abstract

Background: According to the guidelines for cGMP, strictly regulated procedures and quality control (QC) tests are required. However, none of the regulations by authorities suggested have β-galactosidase and Annexin V/PI assays as QC tests. In this study, we aimed to demonstrate the feasibility of Annexin V and β-galactosidase assays as valid cGMP-compliant QC tests. Methods: Wharton’s Jelly derived mesenchymal stem cells (WJ-MSCs) were isolated, characterized and screened as valid QC tests. In this study, in addition to routine QC tests, β-gal assays were performed to confirm telomerase activity and Annexin V methods were used to confirm cell viability assays. Results: It was concluded that one donor (WJ-MSC-3) has higher telomerase activity and it has lowest β-gal activity at the same time (p<0,001). Only one donor showed significant difference in annexin V viability tests, indicating the importance of using automatic devices and Annexin V to manual tests for cell viability analysis (Muse: 92,48 ± 0,487% Flow Cytometry: 90,76 ± 1,116% p<0,01). Conclusion: With these existing data, telomerase activity and senescence levels may correlate and affect MSC quality in cell based therapy. In order to determine the viability values by adding Annexin V based automated methods could give more guaranteed results. characteristics and differentiation capacity of MSCs Age related differences are minimal in WJ derived MSCs samples as compared to other adult sources. MSCs harvested from the WJ are considered “young”, much more proliferative, immunosuppressive, and even more therapeutically active than those isolated from ‘’older’’, adult tissue sources 44]. Our results together with the previously reported studies in the literature suggest that adult-derived mesenchymal stem cells will have more significant differences in senescence levels. Surprisingly, although RTA level of 3 of 4 different WJ-MSCs were very close to each other; telomerase activity of WJ-MSC-3 is significantly higher than others. With the present data in mind, telomerase activity and senescence levels may correlate and also may affect MSC quality in cell based therapy.