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387 cAMP RESPONSE ELEMENT BINDING PROTEIN AND COLLAGEN 1 IMPACT ON VASCULAR SMOOTH MUSCLE CELL GENE REGULATION AND PHENOTYPE
  1. Y. Bogaert,
  2. J. Grippa,
  3. R. Nemenoff,
  4. J. E-B Reusch
  1. Denver CO., David Geffen School of Medicine at UCLA

Abstract

Atherosclerosis is a major cause of morbidity and mortality in the U.S. In response to vascular injury, vascular smooth muscle cells (VSMC) undergo phenotypic modulation, a switch in phenotype from quiescent to active (proliferative, migratory and pro-apoptotic). Phenotypic modulation or de-differentiation of VSMC may play an important role in atherosclerotic plaque instability. Structural matrix proteins affect VSMC phenotype, but little is known about the mechanism of this effect. To better understand the impact of matrix protein on VSMC phenotype we cultivated VSMC on native Type 1 collagen fibrils (NC) or denatured collagen (DNC). Adult VSMC showed dramatic alterations in morphology from the flat, lamellar morphology of cells plated on DNC to stellate shaped cells with multiple dendritic processes on NC. We hypothesized that NC was leading to an activated VSMC phenotype by decreasing expression of key transcriptional regulators of VSMC differentiation. Our group has established that the transcriptional regulator CREB (cAMP response element binding protein) plays a key role in the maintenance of the quiescent, differentiated SMC phenotype. Other markers of VSMC differentiation are smooth muscle a-actin (SMA) and Serum Response Factor (SRF). SMA, CREB and SRF are highly expressed in quiescent cells and expressed at low levels in proliferative cells. VSMC plated on NC for 24 or 48 hours had significant decreases in SMA, CREB and SRF protein expression. The decrease in SMA involved transcriptional control, as there was also a significant reduction in SMA promoter activity. We investigated the mechanisms involved in the decreased SMA and CREB expression of adult cells on NC. Growth of adult VSMC on NC resulted in increased activation of the PI3kinase signaling cascade and AKT as compared to cells on DNC. Pharmacological disruption of AKT signaling blocked NC downregulation of SMA and CREB. We next assessed the relationship between vascular collagen 1 expression, CREB, and SRF protein content in vivo. Aortic lysates from lean and fatty Zucker rats were examined for collagen 1, CREB and SRF by Western Analysis. Collagen 1 expression increased in the fatty rats by 200% between 4-14 weeks, CREB content decreased to 60% of baseline and SRF protein decreased to 20% of baseline.

Summary Collagen 1 exposure correlates with VSMC dedifferentiation in vivo and in vitro. Vascular collagen accumulation may promote atherosclerosis. Funding Source: DK19928, DK064741

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