Effect of Induction Time on the Proliferation and Differentiation of Induced Schwann-Like Cells from Adipose-Derived Stem Cells

Abstract

To compare how different induction time takes effect on the proliferation and secretion ability of adipose-derived stem cell (ADSC)-induced Schwann-like cells (iSCs), ADSCs were isolated from healthy adult female rats. Flow cytometry (FCM) was performed to detect the ADSC-positive markers CD29, CD44, and CD90 and the negative marker CD45. iSC induction medium was used to culture the ADSCs. S-100, GFAP, MBP, and P75 were detected by immunofluorescence staining to identify iSC differentiation. Cell morphological changes were observed by an inverted microscope after induction. An MTS assay was used to evaluate the cell proliferation ability. Western blot analyses of caspase-3/cleaved caspase-3 and FCM were applied to assess cell apoptosis. Co-culture system of PC12 and ADSCs or iSCs was established to analyse the biological function of iSCs. Among the examined proteins, S-100, GFAP, MBP, and P75 were expressed in iSCs. After day 7, the cell proliferation rate was significantly lower than that before induction, and on day 19, the proliferation rate of iSCs was lower than 50% of the proliferation rate before induction (OD value = 0.016 ± 0.003 vs. 0.400 ± 0.004, p < 0.01). Starting from day 19, P21, P53, Apoj, S100, Gdnf, and Mbp all consistently showed a trend toward increased expression. Secretion of NGF, MBP, and BDNF was more enhanced at 19 days than that at 7 days. In co-culture system, the induction effect of iSCs was more pronounced at 19 days than that at 7 days, and the difference was statistically significant (55.40 ± 4.50 μm vs 37.15 ± 3.75 μm, p < 0.01). In conclusion, the proliferation ability of ADSC-derived iSCs was negatively correlated with the induction time, while the expression of SC marker proteins was positively correlated. Therefore, iSCs are suitable for use at 19 days after induction.

In Vitro Cultures of Adipose-Derived Stem Cells: An Overview of Methods, Molecular Analyses, and Clinical Applications

“…According to the results, different induction times negatively impacted proliferation but positively impacted the expression of SCs. Application of induced SCs for nerve repair and functional reconstruction upon nerve injury appeared to be the most beneficial after 19 days of induction [44].…”

Adipose tissue stem cells in peripheral nerve regeneration—In vitro and in vivo

“…(2007), which uses 5 ng/ml platelet‐derived growth factor, 10 ng/ml basic fibroblast growth factor, 14 μM forskolin, and GGF‐2 252 ng/ml (Kingham et al., 2007). Also, other differentiation media with glial growth factors are described (Wong et al., 2020). Alternative protocols have detected that SCLC differentiation of ASCs is possible by treatment with SC‐conditioned medium, olfactory ensheathing cell‐conditioned medium, the addition of insulin and progesterone, or nerve leachate (Fu et al., 2016; Kang et al., 2019; Liu et al., 2020; Xie et al., 2017).…”

“…A frequently used differentiation protocol is described by Kingham et al (2007), which uses 5 ng/ml platelet-derived growth factor, 10 ng/ml basic fibroblast growth factor, 14 μM forskolin, and GGF-2 252 ng/ml (Kingham et al, 2007). Also, other differentiation media with glial growth factors are described (Wong et al, 2020).…”

Adipose-derived stem cell-derived extracellular vesicles inhibit neuroblastoma growth by regulating GABBR1 activity through LINC00622-mediated transcription factor AR

“…In a previous study, the human cells were cocultured with mouse Evs. 43 Evs isolated from the intestinal organoids of mouse and human regulate inflammatory responses of a variety of the immune cells such as microglia cells, dendritic cells, mouse bone-marrow derivedmacrophages, and human monocytes. 44 Therefore, referring to the role of different types of EVs, 45 we chose to use murine Evs over human cells.…”

“…Furthermore, referring to a previous study, we selected female rats for experiments. 43 However, the ADSCs from male rats also have the ability of adipogenic and osteogenic differentiation; ADSC-EVs CD63 and Alix are positive and the diameter isolated from ADSC is 30-100 nm. 50,51 Due to the limitation of experimental time, we have not carried out relevant research in male rats, and whether ADSCs isolated from inguinal adipose of male rats and ADSC isolated from female rats have similar phenotypes still remains to be explored.…”