The effects of exogenous hyaluronic acid with different molecular weight on chondrogenic differentiation of rabbit bone marrow mesenchymal stem cells

Abstract

Objective To study the effects of exogenous hyaluronic acid (HA) with different molecular weight on chondrogenic differentiation of rabbit bone marrow-derived mesenchymal stem cells (MSCs).Methods The un-induced MSCs in passage 3 were divided into five groups with different suplements:Group A,basic medium + HA 0.1 g/L (average molecular weight:1000 × 103 ) ; Group B,basic medium + HA 0.1 g/L ( average molecular weight 1800 × 103 ) ; group C,basic medium + HA 0.1 g/L ( average molecular weight 2000 × 103 ) ; Group D,basic medium + transforming growth factor-33 ( TGFβ3) 10 μg/L (positive control) ; Group E,basic medium only (negative control).The morphologic features and proliferation of MSCs were observed under a light microscope.The chondrogenic differentiation was identified by toluidine blue staining as well as immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR) were used to detect the expression of collagen Ⅱ respectively.Results The speed of cell proliferation was decelerated after HA was added.The cell morphology changed from long spindle to polygon,oval.The extracellular matrix showed metachromasia with toluidine blue and positive type Ⅱ collagen immunohistochemical staining.The mRNA expression of type Ⅱ collagen was detected by RT-PCR.The relative gene expression of type Ⅱ collagen in group A,B and C was 0.64 ± 0.06,0.72 ±0.03,0.75 ±0.01,respectively,and that in groups D and E was 0.96 ±0.15 and 0.09 ±0.03 respectively.There was no statistically significant difference in the expression of type Ⅱ collagen between groups B and group C.There was significant difference between experimental group and control groups.Conclusion The exogenous HA with different molecular weight can induce chondrogenic differentiation of MSCs with different ability.HA with higher average molecular weight enhances the chondrogenesis of MSCs effectively.But all HA shows weaker ability of inducement than TGF-33. Key words: Hyaluronic acid; Molecular weight; Mesenchymal stem cell; Chondrogenesis