Differential regulation of vascular smooth muscle and endothelial cell proliferation in vitro and in vivo by cAMP/PKA-activated p85α^PI3K

Abstract

cAMP inhibits proliferation in most cell types, triggering different and sometimes opposing molecular pathways. p85alpha (phosphatidylinositol 3-kinase regulatory subunit) is phosphorylated by cAMP/PKA in certain cell lineages, but its effects on vascular smooth muscle cells (VSMCs) and endothelial cells (ECs) are unknown. In the present study, we evaluated 1) the role of p85alpha in the integration of cAMP/PKA-dependent signaling on the regulation of VSMC and EC growth in vitro; and 2) the effects of PKA-modified p85alpha on neointimal hyperplasia and endothelial healing after balloon injury in vivo. Plasmid constructs carrying wild-type and PKA-modified p85alpha were employed in VSMCs and ECs in vitro and after balloon injury in rat carotid arteries in vivo. cAMP/PKA reduced VSMC proliferation through p85alpha phosphorylation. Transfected PKA-activated p85alpha binds p21ras, reducing ERK1/2 activation and VSMC proliferation in vitro. In contrast, EC proliferation inhibition by cAMP is independent from PKA modification of p85alpha and ERK1/2 inhibition; indeed, PKA-activated p85alpha did not inhibit per se ERK1/2 activation and proliferation in ECs in vitro. Interestingly, cAMP reduced both VSMC and EC apoptotic death through p85alpha phosphorylation. Accordingly, PKA-activated p85alpha triggered Akt activation, reducing both VSMC and EC apoptosis in vitro. Finally, compared with controls, vascular gene transfer of PKA-activated p85alpha significantly reduced neointimal formation after balloon injury in rats, without inhibiting endothelial regeneration of the injured arterial segment. In conclusions, PKA-activated p85alpha integrates cAMP/PKA signaling differently in VSMCs and ECs. By reducing neointimal hyperplasia without inhibiting endothelial regeneration, it exerts a protective effect against restenosis after balloon injury.

The p85α regulatory subunit of PI3K mediates cAMP-PKA and retinoic acid biological effects on MCF7 cell growth and migration

“…cAMP induces proliferation in FRTL5 thyroid cell (42,43), while it inhibits proliferation in NIH 3T3 (44). A differential regulation of VSM and endothelial cell proliferation, by cAMP/PKA-activated p85α PI3K has also been described (39). In our model cAMP treatment is able to reduce cell proliferation in all conditions examined.…”

“…In fact the phosphorylation of p85α PI3K Ser83 inhibits cell proliferation in fibroblasts and VSMC, while it is essential for the correct cell cycle progression in thyroid cells, and does not affect cell proliferation of endothelial cells (9,10,39). Our results show that phosphorylation of p85α PI3K Ser83 is essential for MCF7 cell survival and proliferation, as demonstrated by the low growth rate observed in cells overexpressing the p85A mutant.…”

Mechanisms of Smooth Muscle Cell Proliferation and Endothelial Regeneration After Vascular Injury and Stenting - Approach to Therapy -

“…In this way, rat carotid arteries were locally transfected with FLAG 49 ) VSMC and Endothelial Regeneration After Stenting tagged plasmid vectors expressing p85 WT , p85 PKA-inactive , and p85 PKA-activated immediately after balloon injury. 48 In this elegant experiment, we demonstrated that local delivery of p85 PKA-activated prevents VSMC proliferation and NIH after experimental balloon injury, and that p85 PKA-activated local delivery does not affect endothelial regeneration in rat balloon-injured arteries.…”

“…Our experimental contribution to understanding the molecular and cellular determinants of smooth muscle and endothelium in restenosis and NIH comes from vascular gene transfer of dominant negative mutants of the regulatory subunit of the phosphatidylinositol 3-kinase, namely p85. 48 Indeed, we recently investigated the effects of overexpression of p85α PI3K plasmids in rat carotid arteries subjected to balloon injury, by using pluronic F127 gel as the sustained-release polymer. In this way, rat carotid arteries were locally transfected with FLAG 49 ) VSMC and Endothelial Regeneration After Stenting tagged plasmid vectors expressing p85 WT , p85 PKA-inactive , and p85 PKA-activated immediately after balloon injury.…”

Differential phosphoinositide 3-kinase signaling: implications for PTCA?

“…Torella et al (6) further investigated signaling events triggered by pSer83-p85␣ overexpression in EC and VSMC cell culture and demonstrated that pSer83-p85␣ binds to Ras and inhibits ERK-1/2 activation in VSMCs but not in ECs. These results suggest a cAMP-and PKA-independent but ERK-1/2-dependent mechanism of EC proliferation, suggesting that EC and VSMC proliferation are regulated by multiple redundant pathways, among which ERK-1/2 activation is inessential for EC, but essential for VSMC, proliferation.…”