TIMAP promotes angiogenesis by suppressing PTEN-mediated Akt inhibition in human glomerular endothelial cells

Abstract

Obeidat M, Li L, Ballermann BJ. TIMAP promotes angiogenesis by suppressing PTEN-mediated Akt inhibition in human glomerular endothelial cells. Am J Physiol Renal Physiol 307: F623-F633, 2014. First published July 9, 2014 doi:10.1152/ajprenal.00070.2014.-The function of TIMAP, an endothelial cell (EC)-predominant protein phosphatase 1-regulatory subunit, is poorly understood. We explored the potential role of TIMAP in the Akt-dependent regulation of glomerular EC proliferation, survival, and in vitro angiogenesis. To deplete TIMAP, the EC were transfected with TIMAP-specific or nonspecific small interfering (si) RNA. The rate of electrical impedance development across subconfluent EC monolayers, a measure of the time-dependent increase in EC number, was 93 Ϯ 2% lower in TIMAP-depleted than in control EC. This effect on cell proliferation was associated with reduced DNA synthesis and increased apoptosis: TIMAP silencing reduced 5-ethynyl-2=-deoxyuridine incorporation by 38 Ϯ 2% during the exponential phase of EC proliferation, and cleaved caspase 3 as well as caspase 3 activity increased in TIMAPdepleted relative to control cells. Furthermore, TIMAP depletion inhibited the formation of angiogenic sprouts by glomerular EC in three-dimensional culture. TIMAP depletion strongly diminished growth factor-stimulated Akt phosphorylation without altering ERK1/2 phosphorylation, suggesting a specific effect on the PI3K/ Akt/PTEN pathway. Endogenous TIMAP and PTEN colocalized in EC and coimmunoprecipitated from EC lysates. The inhibitory PTEN phosphorylation on S370 was significantly reduced in TIMAP-depleted compared with control EC, while phosphorylation of PTEN on the S380/T382/T383 cluster remained unchanged. Finally, the PTEN inhibitor bpV(phen) fully reversed the suppressive effect of TIMAP depletion on Akt phosphorylation. The data indicate that in growing EC, TIMAP is necessary for Akt-dependent EC proliferation, survival, and angiogenic sprout formation and that this effect of TIMAP is mediated by inhibition of the tumor suppressor PTEN. endothelial cells; PTEN; protein phosphatase 1-regulatory subunit; PPP1R16B; apoptosis; clectric cell-substrate impedance sensing MECHANISMS THAT REGULATE ANGIOGENESIS have received massive attention since Folkman (15) first advanced the theory that tumor growth depends on new blood vessel formation. Angiogenesis involves the sprouting and elongation of new blood vessels from preexisting vessels followed by stabilization and vessel specification (28). This process is crucial for normal blood vessel formation and patterning during embryonic development, for postembryonic blood vessel repair (5), and remodeling in response to hypoxia, in the female reproductive cycle, and for the establishment of the placental vasculature. Pathological retinal and tumor vascularization also depend on angiogenesis. While endothelial cells (EC) at the tip of the angiogenic sprout determine the direction and pattern of new blood vessel growth, elongation of the new vessel stalk requires EC proliferatio...

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“…Several candidate genes are also involved in aspects of physiology and metabolic efficiency. XIRP2 has been shown to play a role in cardiac remodelling (McCalmon et al ., ), whereas TIMAP is involved in the regulation of barrier function in pulmonary endothelial cells (Poirier et al ., ) and promotes angiogenesis (Obeidat et al ., ). ROCK2 is associated with variation in blood pressure and cardiovascular physiology (Riento & Ridley, ; Noma et al ., ; Seasholtz et al ., ; Rankinen et al ., ), and MOGAT2 plays an important role in metabolic efficiency and regulation of energy metabolism (Gao et al ., ).…”

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