Abstract
Background & Aim Induced pluripotent stem cells (iPSCs) are commonly studied for clinical use due to its flexibility in differentiating into any desirable cellular products. iPSCs are commonly derived from fibroblasts or cells from peripheral blood due to its accessibility. Although not common, we have derived iPSCs from mesenchymal stem cells (MSC) for several purposes. We are developing an allogenic combinatory cellular therapy for heart disease using both MSCs with iPSC derived cardiomyocyte (CM) or cardiac progenitor cells (CPCs). We theorize, by utilizing iPSC derived from MSCs, it provides more compatibility when transplanting the two final cellular products from one donor into patients, thus yielding better therapeutic effects. Also, recent changes in donor qualification increase the financial burden when launching new cellular therapeutics due to the increase of required testing on donor and final cellular product. In this scenario, by reprogramming the MSCs into iPSCs then differentiate into CM, we eliminated the additional cost of qualifying the parent cells of iPSCs. Methods, Results & Conclusion Although cardiac differentiation of iPSC is well studied, the optimal differentiation condition needed to be tested for each cell line used. The initial evaluation must be performed to determine the optimal cell density, the concentration of CHIR99021, and the time point of switching to IWP2. After determining these factors, the cardiac differentiation was performed on both IMR90 (lung fibroblast) and MSC derived iPSCs in order to evaluate the efficiency of differentiation of the latter iPSC. The iPSCs were plated onto Vitronectin coated plates until the desired confluence reached using iPSC culture medium. At day 0 of the cardiac differentiation process, the iPSC culture medium was replaced with CHIR99021in differentiation medium (RPMI-1640 with B-27 without insulin). Two or three days later, IWP2 replaced CHIR99021 in the differentiation medium for another two days. After day 7 of the cardiac differentiation process, cells were maintained with CM culture medium (RPMI-1640 with B-27 with insulin) for another seven days. The medium was changed every three to four days. In summary, only the optimized combination of conditions yield beating CM culture at day 14 from both MSC and IMP90 derived iPSCs. CM derived from both iPSCs stained positive for Troponin T. At day 7 of the cardiac differentiation, the cultures were stained positive for GHRHR, which will be used as a marker for CPC selection. Induced pluripotent stem cells (iPSCs) are commonly studied for clinical use due to its flexibility in differentiating into any desirable cellular products. iPSCs are commonly derived from fibroblasts or cells from peripheral blood due to its accessibility. Although not common, we have derived iPSCs from mesenchymal stem cells (MSC) for several purposes. We are developing an allogenic combinatory cellular therapy for heart disease using both MSCs with iPSC derived cardiomyocyte (CM) or cardiac progenitor cells (CPCs). We theorize, by utilizing iPSC derived from MSCs, it provides more compatibility when transplanting the two final cellular products from one donor into patients, thus yielding better therapeutic effects. Also, recent changes in donor qualification increase the financial burden when launching new cellular therapeutics due to the increase of required testing on donor and final cellular product. In this scenario, by reprogramming the MSCs into iPSCs then differentiate into CM, we eliminated the additional cost of qualifying the parent cells of iPSCs. Although cardiac differentiation of iPSC is well studied, the optimal differentiation condition needed to be tested for each cell line used. The initial evaluation must be performed to determine the optimal cell density, the concentration of CHIR99021, and the time point of switching to IWP2. After determining these factors, the cardiac differentiation was performed on both IMR90 (lung fibroblast) and MSC derived iPSCs in order to evaluate the efficiency of differentiation of the latter iPSC. The iPSCs were plated onto Vitronectin coated plates until the desired confluence reached using iPSC culture medium. At day 0 of the cardiac differentiation process, the iPSC culture medium was replaced with CHIR99021in differentiation medium (RPMI-1640 with B-27 without insulin). Two or three days later, IWP2 replaced CHIR99021 in the differentiation medium for another two days. After day 7 of the cardiac differentiation process, cells were maintained with CM culture medium (RPMI-1640 with B-27 with insulin) for another seven days. The medium was changed every three to four days.
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