ROCK2 mediates the proliferation of pulmonary arterial endothelial cells induced by hypoxia in the development of pulmonary arterial hypertension

Abstract

Abstract. It has been reported that RhoA activation and Rho-kinase (ROCK) expression are increased in chronic hypoxic lungs, and the long-term inhibition of ROCK markedly improves the survival of patients with pulmonary arterial hypertension (PAH). However, whether Rho-kinase α (ROCK2) participates in regulation of the growth of pulmonary arterial endothelial cells (PAECs) remains unknown. The aim of the present study was to investigate the effect of hypoxia on the proliferation of PAECs and the role of ROCK2 in the underlying mechanism. The results of western blotting and reverse transcription-quantitative polymerase chain reaction analysis showed that hypoxia increased the activity and expression of ROCK2 in PAECs, and the stimulating effects of hypoxia on the proliferation of PAECs were attenuated by either the ROCK inhibitor Y27632 or transfection with ROCK2 small interfering RNA. Moreover, analysis of cyclin A and cyclin D1 mRNA expression indicated that ROCK2 mediates the cell cycle progression promoted by hypoxia. These results indicate that hypoxia promotes the proliferation of pulmonary arterial endothelial cells via activation of the ROCK2 signaling pathway. IntroductionPulmonary arterial hypertension (PAH) is a fatal condition characterized by increased pulmonary vascular resistance and finally leading to right heart failure and mortality (1,2). Endothelial injury, prolonged vasoconstriction and the proliferation and migration of vascular smooth muscle cells (VSMCs), are causes of increased pulmonary vascular resistance (3). Multiple pharmacological agents, such as vasodilators and anticoagulants, have been developed for the treatment of PAH; however, the long-term prognosis of patients with severe PAH remains poor (3). Therefore, there is an urgent requirement for the development of more effective treatments for PAH.Rho-kinase (ROCK) is a member of the serine/threonine kinase family that is an important downstream effector of the small GTP-binding protein RhoA. The Rho/ROCK pathway plays an important role in various fundamental cellular functions, including contraction, motility, proliferation and migration (4,5). There are two isoforms of ROCK, namely ROCK1 (Rho-kinase β) and ROCK2 (Rho-kinase α) (6). ROCK1 and ROCK2 are highly homologous with regard to amino acid sequence and kinase domains, sharing ~65% homology in amino acid sequence and 92% homology in their kinase domains (6). Although the two isoforms are ubiquitously expressed in invertebrates and vertebrates, ROCK1 is expressed mainly in circulating inflammatory cells and ROCK2 is expressed in vascular cells (7,8). Homozygous ROCK1-deficient mice show open eyelids at birth and omphalocele, whereas homozygous ROCK2-deficient mice die embryonically because of placental dysfunction, suggesting that ROCK1 and ROCK2 mediate different functions in different types of cells (9,10). To date, to the best of our knowledge, whether ROCK is responsible for the growth of pulmonary arterial endothelial cells (PAECs) has not yet been evaluated.The present ...

ROCK Inhibition as Potential Target for Treatment of Pulmonary Hypertension

“…Proliferation in pulmonary artery EC cultures induced by hypoxia involves the increase in cyclin A and cyclin D1 in order to promote cell cycle progression. Under these conditions, the use of Y-27632 or a small-interfering RNA targeting ROCK2 abolished EC proliferation without compromising cell viability, indicating that ROCK inhibition may also control EC phenotypic changes seen in PH [ 68 ]. These effects could reflect ROCK activation by vascular growth factor and EC stiffening, as ROCK inhibitors also block EC migration and angiogenesis [ 69 , 70 , 71 ].…”

Rho-Associated Coiled-Coil Kinase (ROCK) in Molecular Regulation of Angiogenesis

“…In addition, ROCK regulates EC proliferation by its targeted gene expression associated with cell cycle. In the hypoxic condition, ROCK2 promotes proliferation of pulmonary arterial ECs though upregulating the expression of cyclin A and cyclin D1 101. Rho/ROCK signaling mediates oxLDL-induced EC proliferation through downregulation of cell cycle inhibitor p27 102.…”

The level of ROCK1 and ROCK2 in patients with pulmonary hypertension in plateau area

“…FENG QIAO et al . 14 showed that the activity of endothelial cells decreased after using siRNA to silence the ROCK2 gene. At the same time, they found that under low oxygen, Rho/Rho-kianse pathway was activated, ROCK2 promoted cell division phase G0/G1 phase to S phase by Up-regulated A/D1 cell cycle protein expression to shorten cell cycle and promote endothelial cell proliferation.…”