Effects of pulsatile fluid shear stress gene expression of human smooth muscle cells by using oligo-microarray

Introduction

We investigated the effects of pulsatile flow shear stress on the gene expression profiles of human umbilical artery smooth muscle cells (HUASMCs) in vitro by using the Express ChipTMDNA microarray method; to investigate the difference between pulsatile and steady shear stress on differentially expressed genes of HUASMCs.

Methods

With a modified pulsatile flow chamber system, HUASMCs were respectively exposed to pulsatile and steady fluid shear stress (5.52 dyne/cm2) for 6h, and normal static cultured HUASMCs were selected as a control. The total cellular RNA was extracted by TRlzol Reagent (Life Technologies, Inc) according to the manufacturer’s manual. Conversion of mRNA to single stand cDNA and double stand cDNA template was synthesized by Reverse Transcription from the total RNA. cRNA probe was transcribed with biotin labeling. After hybridization of probe with microarray, binding of streptavidin to biotin was performed to be amplificated with the first antibody, and further amplificated with Cy3-conjugated second antibody. Then detection of Cy3 dye was carried with ScanArray 5000.

Results

The results showed that a total of 1,330 genes revealed differential expression in HUASMCs exposed on pulsatile shear stress (5.52 dyne/cm2, 6 h); however, 2,676 genes revealed differential expression in HUASMCs exposed on steady shear stress. Comparing HUASMCs exposed on pulsatile with steady shear stress, there were 2,297 genes revealed differential expression.

Discussion/Conclusions
The transcriptional profile of fluidally induced genes in HUASMCs suggested a different response to pulsatile and steady shear stress. The ExpressChipTMDNA microarray technology is effective in screening the differentially expressed genes of vascular smooth muscle cells induced by pulsatile and steady fluid and better understanding their pathogenesis.

References

[1]. Lee AA, Graham DA, et al(2002). J Biomech Eng.124(1):37.
[2]. Palumbo R, , et al(2002). Arterioscler Thromb Vasc Biol. 22(3):405.
[3]. Liu SQ, Goldman J(2001). IEEE Trans Biomed Eng. 48(4):474.
[4]. Feng Y, Yang J, et aL(1999). J Circ Res. 85: 1118.
[5]. Shigematsu K, et al (2000). Int Angiol. 19(1):39.
[6]. Zhang JN, et al(2002). Thromb Haemost. 88(5):817.