| Research Abstract |
Primarily cultured smooth muscle cells (SMCs) rapidly dedifferentiate under normal culture conditions. We investigated to establish a culture system maintaining a differentiated phenotype of SMCs using several extracellular matrices and growth factors or cytokines. From these analyzes, we found that laminin has a potency to maintain a differentiated phenotype of SMCs under serum-free culture conditions. Furthermore, we obtained evidence that insulin-like growth factor I (IGFI), II (IGFII), or insulin among several growth factors and cytokines possesses a remarkable activity to maintain the differentiated phenotype of SMCs for a long culture, and IGFI is a most potent factor for SMC differentiation. These results suggest the involvement of signal transduction via laminin and IGFI receptors in SMC differentiation. We also found that the expression of IGFI-receptor is SMC phenotype-dependent. Therefore, the downregulation of IGFI-recepter might be one reason for dedifferentiation of SMCs
… More
by serum growth factors. Using our SMC culture system, we characterized the transcriptional regulation of the caldesmon (CaD) and the alpha1 integrin promoters. These analyzes revealed that the CArG box plays a vital role for high level transcription of the both genes in SMCs, and the serum response factor (SRF) is a core factor for the CArG box binding. We demonstrated that the expression of alpha-SM actin in visceral SMCs is opposite to that in vascular SMCs ; alpha-SM actin is expressed in undifferentiated and dedifferentiated visceral SMCs, but not in differentiated visceral SMCs. We identified a novel cis-element in the promoter region of alpha-SM actin gene which acts as a negative regulator. In the CaD gene, alternative selection of two 5'-splice sites within exon 3 has determined the expression of h-or l-CaD isoform. We found functional involvement of hnRNPA1 in the selection of distal 5'-splice site in SMCs. We found that expressional change of alpha-tropomyosin (alpha-TM) isoforms during dedifferentiation of SMCs is arisen by a selectional change between mutually exclusive exons, exons 2a and 2b, and such change occurrs coordinately with CaD isoformal change, suggesting a common splicing mechanism for SMC phenotype-dependent expression of alpha-TM and CaD isoforms. Less
|
