Induction of heparin-binding epidermal growth factor-like growth factor mRNA by phorbol ester and angiotensin II in rat aortic smooth muscle cells.

D.H Temizer,M Yoshizumi,M.A Perrella,E.E Susanni,T Quertermous,M.E Lee

doi:10.1016/s0021-9258(18)35847-2

Abstract

To determine whether the gene encoding the recently identified heparin-binding epidermal growth factor-like growth factor (HB-EGF), a potent smooth muscle cell (SMC) mitogen of macrophage origin, is transcribed and regulated in vascular SMC, we isolated cDNA clones encoding rat HB-EGF from a macrophage library. Using the rat HB-EGF cDNA as a probe for RNA blot analysis, we detected low levels of HB-EGF mRNA in rat aortic SMC in culture. However, 20 nM 12-O-tetradecanoylphorbol-13-acetate (TPA) and 10(-6) M angiotensin II (AII) induced a marked increase in HB-EGF mRNA levels in rat aortic SMC (11- and 4.6-fold, respectively) that was both dose- and time-dependent. In response to TPA and AII, HB-EGF mRNA levels increased rapidly, peaked at 2 h, and returned to base line at 7 h. This effect of AII on HB-EGF induction was specific, as evidenced by the fact that it could be completely blocked by the AII antagonist saralasin. This is the first demonstration that HB-EGF is transcribed and regulated in SMC. The inducible transcription of this potent SMC mitogen gene in vascular SMC suggests that HB-EGF may have an important autocrine role in the proliferation of SMC in vascular diseases such as atherosclerosis and hypertension.

Highlights

This is the first demonstrationthat heparin-binding epidermal growth factorlike growth factor (HB-EGF) is transcribed and regulatedin smooth muscle cell (SMC).The inducibletranscription of this potent SMC mitogen gene invascular SMC suggests that HB-EGF may have an important autocrine role in the proliferation of SMC in vascular diseasessuch as atherosclerosis and hypertension
To test whether SMC transcribe the HB-EGF gene, we first amplified an HB-EGF cDNA fragment from rat lung RNA by the reverse transcription polymerase chain reaction (PCR). Using this fragment as a probe, we identified a low level of HB-EGF mRNA inrat aortic SMC (RASMC) in culture byRNA blot analysis. 12-O-Tetradecanoylphorbol13-acetate (TPA) and angiotensin I1 (AII) induced a marked increase in the level of HB-EGF mRNA in RASMC that was bothtime-and dose-dependent
Vascular SMC proliferationis a critical event in the development of atherosclerotic lesions [1,2,3,4,5], which commonly feature macrophages, T cells, and endothelial cells

Summary

RAT human

101 LGKKRDPCLKKYKDFCIHGECKYLKELRIPSCHCLPGYHGQRCHGLSLW 150 IIIIIIIII : IIIIIIIIIlIII : I I I II llIIIIII : IIIlIIIII. RAT 151 ENPLYTYDHTIlrLAWAVVLSSVCLLVIVGLLMFRYHRRGGYDLESEW 200 II.IIIIIIII:IIIIIIIIIIIIIIIIIIIIIIIIIIIIIII:I.IIII human 151 enhlyty ~ awawlssvcllvivgllrnfryhrrggydveneek v200

DNA fragment from rat lung RNA by reverse transcription

TPA h M l

DISCUSSION