A study on in-vitro transdifferentiation of rat bone marrow stromal cells into neuroepithelial-like cells

الملخص EN

Background : bone marrow stem cells (BMSCs) are a rich source of stem cells and may represent a valid alternative to neural or embryonic stem cells by replacing the autologous damaged tissues in neurodegenerative diseases.

In this study, we attempted to devise a protocol for the induction of BMSCs into neuroepithelial-like cells (NELCs).

Methods : rat BMSCs were isolated from the long bones of adult Sprague–Dawley rats.

Their purity in the 4th passage was evaluated with fibronectin by immunocytochemistry, and the stemness marker Oct-4 was assessed by RT-PCR technique.

The cells were expanded and induced in the induction stage.

The BMSCs were incubated with either β-mercaptoethanol (βME) (1 mM), dimethyl sulfoxide (DMSO) (2 %) or biotylated hydroxyanisol or butylated hydroxyanisol (BHA) (200 μM) in α-MEM medium without fetal bovine serum (FBS).

They were washed with phosphate buffer saline (PBS) and proceeded to the 2nd phase of induction, where the induction medium was changed with α-MEM and 15% FBS containing all-trans retinoic acid (RA) (1 μM) (for 3 days).

Then, the expression of the markers was assessed with GFAP, nestin and neurofilament 68 antibodies, respectively and the expression of Oct-4 and NeuroD was evaluated by RT-PCR.

Results : the purity of the BMSCs at the 4th passage was more than 92%.

The mRNA of Oct-4 was expressed in these cells.

Induction of BMSCs by DMSO-RA could differentiate NELCs significantly more than βME-RA and BHA-RA.

The transdifferentiation of NELCs was evaluated by nestin antibody and NeuroD mRNA expression; later markers expressed very low detectable level in BMSCs.

But the differentiation of BMSCs into astrocytes was less in all of the experiment groups that is estimated GFAP antibody.

Conclusion : The application of DMSO-RA can transdifferentiate BMSCs into NELCs in-vitro.