Journal of Tissue Engineering and Reconstructive Surgery ›› 2024, Vol. 20 ›› Issue (3): 275-.

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Preliminary study on modRNA transfection of human adipose-derived stem cells with different transfection methods

  

  • Online:2024-06-01 Published:2024-07-05

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Abstract:

Objective To select a transfection method with better efficiency and to determine its feasibility in modRNA
transfection of human adipose-derived stem cell （hADSC） as well as specific protein expression pattern. Methods ModGFP
was transfected into hADSC using two kinds of electrotransfection methods （Optimization4 and Optimization6） and liposome
transfection reagent （RNAiMAX）.The transfection efficiency and protein expression of different transfection methods were
analyzed by flow cytometry and fluorescence microscope. Cell survival rate at 24 h after transfection was evaluated by AM/PI
staining. Electrotransformation mode with better effect and RNAiMAX were used to transfect modVEGFA into hADSC.
Culture supernatant of hADSC after being transfected for 24 h was collected to incubate human umbilical vein endothelial
cells （hUVECs） followed by tube formation and transwell migration experiments. hADSCmodVEGFA mixed with matrigel was
injected subcutaneously in nude mice. The ability to stimulate angiogenesis in vivo was analyzed by gross observation， HE
staining， and immunofluorescence staining. Results The transfection efficiency was higher in Optimization6 group （87.75%
±1.52%）， compared with Optimization4 group （80.85% ±1.48%） and RNAiMAX group （81.8%±1.08%）.Fluorescence
microscopy and flow analysis showed that the protein expression pattern was better in electrotransfection groups than that of
RNAiMAX group. The toxicity of these three transfection methods in cells was minimal， no statistical difference was found.
Compared with RNAiMAX， Optimization6 can transfect modVEGFA more efficiently into hADSC and possess better ability of
promoting angiogenesis both in vitro and in vivo. Conclusion Compared with RNAiMAX and Optimization4， Optimization6
has higher transfection efficiency， better protein expression pattern， and stronger angiogenesis promoting effect both in vitro
and in vivo.

Key words:

adipose-derived stem cell