Isolation and culture of rat adipose-derived stem cells and differentiation into oligodendrocyte precursor cells

ZHANG Ya-Qun; LI -Fu; REN Yi-Yan; QIAO Yan-Zi; ZHAO Dong-Mei

doi:10.16098/j.issn.0529-1356.2022.05.003

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Acta Anatomica Sinica ›› 2022, Vol. 53 ›› Issue (5) : 557-562. DOI: 10.16098/j.issn.0529-1356.2022.05.003

Isolation and culture of rat adipose-derived stem cells and differentiation into oligodendrocyte precursor cells

ZHANG Ya-qun FU Li REN Yi-yan QIAO Yan-zi ZHAO Dong-mei*

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Abstract

Objective To explore the possibility of rat adiposederived stem cells (ADSCs) to differentiate into oligodendrocyte precursor cells(OPCs) and find an effective way to treat demyelinating disease. Methods ADSCs from the inguinal region of SD rats were isolated, digested with collagenase type Ⅰ and trypsin, collagenase type Ⅰ digestion method as control, counted and compared; Cultured in vitro and observed the growth characteristics. After ADSCs subcultured 3 times of passages, CD29, CD90 and CD45 were detected by flow cytometry; After differentiation into adipocyte, the cells were identified by the staining of oil red O; After differentiation into OPCs by stem cell differentiation medium and OPCs induced differentiation medium, the expression of α-N-acetylneuraminic acid α-2, 8-sialyltransferase Ⅰ(A2B5) and NG2 was detected by immunofluorescent staining. Results The number of ADSCs in the combined enzyme group was higher than the collagenase type 1 group (P<0.05, n=7); ADSCs grew in a long shuttle type and their morphology tended to be stable after passage. The surface marker CD29,CD90 were positive, and CD45 was negative. After adipogenic induction, oil red O staining showed red lipid droplets of varying sizes in the cells. After OPCs induction, immunofluorescence detection showed that positive reaction of cell surface fluorescence was seen with antibody to A2B5 and NG2,(87.03±0.94)% expressed A2B5, (90.07±0.96)% expressed NG2. After cultured for 3 days, immunofluorescence detection showed that positive reaction of cell surface fluorescence was seen with antibody to myelin basic protein (MBP). Conclusion ADSCs are obtained by combined enzyme digestion and the cells are much more than collagenase alone and can be induced to OPCs in vitro.

Key words

Adipose-derived stem cell / Isolation / Cultivation / Induced differentiation / Oligodendrocyte Precursor cell / Demyelinating disease / Flow cytometry / Rat

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ZHANG Ya-qun FU Li REN Yi-yan QIAO Yan-zi ZHAO Dong-mei. Isolation and culture of rat adipose-derived stem cells and differentiation into oligodendrocyte precursor cells[J]. Acta Anatomica Sinica. 2022, 53(5): 557-562 https://doi.org/10.16098/j.issn.0529-1356.2022.05.003

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