Calcium Homeostasis Modulator (CALHM1/2) in Pulmonary Arterial Hypertension

Abstract

Calcium Homeostasis Modulator in Pulmonary Arterial Hypertension byMarisela Rodriguez Master of Science (M.S.) in Clinical Translational Sciences University of Arizona, Tucson 2020 Professor Jason Yuan, Co-Chair Professor Joe G.N. Garcia, Co-Chair Pulmonary arterial hypertension (PAH) is a progressive and fatal disease that predominantly affects women. The increased pulmonary arterial pressure (PAP) in patients with PAH is mainly generated by increased pulmonary vascular resistance (PVR) (18, 70, 103). Sustained pulmonary vasoconstriction, excessive pulmonary vascular remodeling, in situ thrombosis, and increased pulmonary vascular wall stiffness are the major causes for the elevated PVR and PAP in patients with PAH. Concentric pulmonary vascular remodeling is among one of the major causes for the elevated pulmonary vascular resistance (PVR) and pulmonary arterial pressure (PAP), also one the major causes for increasing afterload of right ventricle (RV) and inducing right heart failure leading to death if untreated (103). Excessive pulmonary artery smooth muscle cell (PASMC) proliferation and inhibited PASMC apoptosis have been implicated in the development and progression of pulmonary vascular wall thickening in patients with PAH and animals with severe experimental pulmonary hypertension (PH). An increase in cytosolic free Ca2+ concentration ([Ca2+]cyt) in PASMCs is not only a trigger for PASMC contraction and pulmonary vasoconstriction, but also an important stimulus for PASMC proliferation, migration, and pulmonary vascular remodeling (16, 83, 85). [Ca2+]cyt in PASMCs is increased by Ca2+ influx through Ca2+-permeable cation channels in the plasma membrane (PM) and Ca2+ release or mobilization from the intracellular Ca2+ stores, mainly the sarcoplasmic (SR) or endoplasmic (ER) reticulum. There are at least three classes of Ca2+-permeable cation channels identified in human and animal PASMCs that are responsible for Ca2+ influx associated with excitation-concentration coupling (EC-coupling) and Ca2+-mediated PASMC proliferation and migration: (i) voltage-dependent Ca2+ channels (VDCC), (ii) receptor-operated Ca2+ channels (ROCC), and (iii) store-operated Ca2+ channels (SOCC) (50, 57). VDCC are opened or activated by membrane depolarization due to, for example, decreased activity or downregulated K+ channels (57) while ROCC is opened or activated by ligand-mediated binding to membrane receptors including G protein-coupled receptors (GPCR) and tyrosine kinase receptors (TKR). Activation of GPCR or TKR upon binding to respective ligands increases production of diacylglycerol (DAG) and inosital 1,4,5-triphosphate (IP3), two important intracellular second messengers. DAG then activates ROCC and introduces receptor-operated Ca2+ entry (ROCE), while IP3 activates IP3 receptors, also referred to as Ca2+ release channels, in the SR/ER membrane and induces Ca2+ release from the intracellular stores to the cytosol contributing to increasing [Ca2+]cyt. Depletion or significant reduction of Ca2+ levels in the ER/SR due to Ca2+ mobilization or release leads to Ca2+ influx through SOCC, commonly referred to as store-operated Ca2+ entry (SOCE). Active depletion of intracellularly stored Ca2+ in the SR/ER then also results in the dimerization and translocation of STIM1 (and/or STIM2) in the SR/ER membrane and forms STIM protein puncta close to the SR/ER-plasma membrane junctions. Then the multimer STIM1/2 proteins in the ER-PM recruit Orai proteins in the plasma membrane to form SOCC responsible for SOCE (16). It has been shown that transient receptor potential (TRP) channels are involved in forming ROCC in PASMCs and can be activated directly by DAG. In our previous publishing, we have determined that the proliferative phenotype of PASMCs employs SOCC leading to increased expression levels of STIM2, not STIM1, and also Orai2 and TRPC6 expressions from IPAH patients- altogether providing an underlying mechanism for enhanced SOCE. TRP channels are also reported to participate in the regulation of SOCE in many cell types (21). In addition to TRP channels, many other types of Ca2+-permeable cation channels may also participate in ROCE and SOCE. Calcium homeostasis modulators (CALHM) including CALHM1 and CALHM2, have been identified as a family of physiologically important plasma membrane ion channels that are permeable to both cations and anions. These channels are allosterically regulated by membrane voltage (or membrane potential) and extracellular Ca2+; CALHM1 and CALHM2 channels are closed at the resting membrane potential but can be opened by strong membrane depolarization. Reduction of extracellular [Ca2+] increases the probability for CALHM channels to open, which allow the channels to be activated at a negative potential. Ultimately, it is widely known that the increased [Ca2+]cyt due to upregulated and activated Ca2+-permeable cation channels contribute to pulmonary vasoconstriction and excessive proliferation of PASMCs (and other cell types, for example fibroblasts and myofibroblasts) in patients with PAH, eventually this leads to concentric pulmonary vascular remodeling (17). Therefore, a rise in intracellular [Ca2+] and activated Ca2+ in PASMC via upregulated and/or activated Ca2+-permeable cation channels play a major role. CALHM1 and CALHM2 have a significant impact on the pathogenesis that lead to the development and progression of PAH. As discussed earlier, sustained pulmonary vasoconstriction and excessive pulmonary vascular remodeling comprise of two major causes for the elevated PVR and PAP in patients with PAH and animals with experimental PH (50). Pulmonary vasoconstriction is certainly a major cause for increasing PVR and PAP at the early stage of disease development, while concentric pulmonary vascular remodeling and obliterative intima and plexiform lesions are made up of the late state pathological changes that contribute to maintaining high PVR and PAP (57). The transition from the contractile or differentiated phenotype to the synthetic or proliferative phenotype of PASMC is thus an important pathogenic process that promotes vascular remodeling (62) in which we aimed at investigating. In this study, I hypothesized that CALHM1 and/or CALHM2 are involved in PASMC phenotypical transition from the contractile or differentiated phenotype to the synthetic or proliferative phenotype, while CALHM1/2 are upregulated in PASMC from patients with PAH and animals with experimental PH.