Adenosine A1 Receptor (A1R) Behavior During Mesenchymal Stromal Cell Osteogenic Differentiation

Objectives: The aim of the study was to evaluate the different behaviour of Adenosine A1 Receptor (A1R) in Mesenchymal Stem Cells (MSCs) from human Adypose and Dental Pulp tissue during osteogenesis proliferation and differentiation.

Methods: Homogeneous pool of human Adypocite and Dental Pulp stem cells were selected with FACS analysis after surgery procedures. Samples was optical and SEM analyzed. The specimes were cultured in osteogenic and adipogenic medium and the multipotency differentiation was evaluated with Alizarin Red and Oil Red Oil staining and the hAS-SCs/hDPSCs clusters were evaluated with RT-PCR, MTS assay, Western Blot and Microarray analysis.

Results: All samples from human adypocite and dental pulp showed an homogenous population of mesenchymal stem cells. At 21 days the osteogenic differentiation were confirmed with positive Alizarine Red staining analysis. RT-PCR analysis in undifferentiated/differentiated cell clusters were observed the different behaviour of ALP, OC and MEPE markers. The MTS Assay showed an increase after 3-7 days of 70% of the proliferation speed in the differentiated cells. The positive gene reaction with microarray, the increase of ALP and the absence of PECAM-1 markers. confirmed the osteogenic differentiation with different behavior between adypocite and dental pulp tissue. A1R stimulation by the agonist 2-chloro-N6-cyclopentyl-adenosine, caused a greater and earlier enhancement of extracellular matrix mineralization in hDPSCs than in hAS-SCs.

Conclusions: The ALP increase and the absence of PECAM-1 protein confirmed the differentiation in osteogenic cluster and not in hematopoietic sense. The ALP high levels expression and the decrease of CD90 marker suggests that the proliferation mechanism not reduced the differentiation processes. The positive Alizarine red analysis and the A1R higher positive expression in hAS-SCs than hDPSCs suggest a positive impact of the adipose tissue and the purine locally released or exogenous added in regenerative processes.