Xeno-Free Cultivation of Mesenchymal Stem Cells From the Corneal Stroma.

PURPOSE

The human cornea has recently been described as a source of corneal stroma-derived mesenchymal stem cells (hMSCs). In vitro expansion of these cells involves basal medium supplemented with fetal bovine serum (FBS). As animal-derived serum can confer a risk of disease transmission and can be subject to considerable lot-to-lot variability, it does not comply with newer Good Manufacturing Practice (GMP)-required animal component-free culture protocols for clinical translation.

METHODS

This study investigated animal-free alternatives to FBS for cultivation of human corneal stromal MSCs. Proliferative capacity was studied for cultures supplemented with different concentrations (2.5%, 5%, and 10%) of FBS, human AB serum, human platelet lysate (HPL), and XerumFree. Unsupplemented basal medium was used as a control. The expression of specific hMSC markers (CD73+, CD90+, CD105+, CD19-, CD34-, CD79α-, CD11b-, CD14-, CD45-, and HLA-DR-) and trilineage differentiation (adipogenesis, osteogenesis, and chondrogenesis) were compared for the two outperforming supplements: 10% FBS and HPL.

RESULTS

HPL is the only consistent non-xeno supplement where hMSC cultures show significantly higher proliferation than the 10% FBS-supplemented cultures. Both FBS- and HPL-supplemented hMSC cultures showed plastic adherence and trilineage differentiation, and no significant differences were found in the expression of the hMSC marker panel. No significant differences in stemness were detected between FBS and HPL cultures.

CONCLUSIONS

We conclude that HPL is the best supplement for expansion of human corneal stromal MSCs. HPL significantly outperforms human AB serum, the chemically defined XerumFree, and even the gold standard, FBS. The xeno-free nature of HPL additionally confers preferred standing for use in GMP-regulated clinical trials using human corneal stromal MSCs.