Immediate-early gene expression in response to hypertrophic and proliferative stimuli in pulmonary arterial smooth muscle cells

J Biol Chem. 1994 Mar 4;269(9):6399-404.

Abstract

Remodeling of the pulmonary vascular tree in pulmonary hypertension is associated with hypertrophy and proliferation of smooth muscle cells. Since the stimuli and signaling pathways for these processes are not well understood, we used a rat pulmonary arterial smooth muscle cell line (PAC1) to examine the effects of thrombin and platelet-derived growth factor (PDGF) on cellular growth and immediate-early gene expression. Over 72 h, thrombin (1 unit/ml) caused hypertrophy as reflected by a 102 +/- 12% increase in protein synthesis and a 49 +/- 11% increase in protein content per cell, but no change in cell number. PDGF (2.5 ng/ml) stimulated proliferation as evidenced by an increase in cell number (doubling in 5 days), but no significant change in protein content per cell. Immediate-early gene expression was examined by Northern blotting: both thrombin and PDGF induced egr-1, c-fos, c-jun, junB, and fra-1 mRNAs within 15 min; the response was maximal at 30-60 min (increases ranging from 2.9- to 9.3-fold over control serum-deprived cells) and returned to base-line levels within 2-4 h. Neither agent affected junD mRNA levels. However, thrombin but not PDGF, caused an increase in fosB mRNA levels (7.7 +/- 4.0-fold higher than control, n = 12, p < 0.0005). The immediate-early gene response to both agonists was generally dependent on extracellular Ca2+, Na2+/H+ exchange, and protein kinase C activation, but not on cAMP. The exception was c-jun mRNA, the levels of which were not affected by inhibition of protein kinase C, but decreased significantly by prevention of cAMP formation. Thapsigargin-sensitive intracellular Ca2+ stores were necessary for the response to thrombin, but not to PDGF. These results demonstrate that thrombin is a hypertrophic agent and that PDGF is a proliferative agent in PAC1 cells. These two agonists stimulate increases in a variety of immediate-early gene mRNAs, but only thrombin induces fosB mRNA.

MeSH terms

  • Animals
  • Calcium / metabolism
  • Cell Division / drug effects
  • Cell Line
  • Egtazic Acid / pharmacology
  • Gene Expression / drug effects*
  • Genes, Immediate-Early*
  • Genes, fos / drug effects
  • Genes, jun / drug effects
  • Hypertrophy
  • Kinetics
  • Muscle, Smooth, Vascular / cytology
  • Muscle, Smooth, Vascular / drug effects
  • Muscle, Smooth, Vascular / metabolism*
  • Platelet-Derived Growth Factor / pharmacology*
  • Protein Biosynthesis
  • Protein Kinase C / metabolism
  • Pulmonary Artery / cytology
  • Pulmonary Artery / drug effects
  • Pulmonary Artery / metabolism*
  • RNA, Messenger / biosynthesis
  • Rats
  • Sodium / pharmacology
  • Sodium-Hydrogen Exchangers / metabolism
  • Thrombin / pharmacology*
  • Transcription, Genetic / drug effects

Substances

  • Platelet-Derived Growth Factor
  • RNA, Messenger
  • Sodium-Hydrogen Exchangers
  • Egtazic Acid
  • Sodium
  • Protein Kinase C
  • Thrombin
  • Calcium