Molecular mechanisms of downregulated Kv channels in IPAH: Role of microRNA

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DESCRIPTION (provided by applicant): Idiopathic pulmonary arterial hypertension (IPAH) is a fatal and progressive disease that predominantly affects women. Pulmonary vascular remodeling, sustained vasoconstriction and in situ thrombosis are the major causes for the elevated pulmonary vascular resistance in IPAH patients. Pulmonary vascular remodeling is characterized in part by significant medial and intimal hypertrophy, due to increased pulmonary arterial smooth muscle cell (PASMC) proliferation and decreased PASMC apoptosis. A rise in cytosolic Ca2+ ([Ca2+]cyt) in PASMC is a major trigger for pulmonary vasoconstriction and an important stimulus for PASMC proliferation. Downregulation of voltage-gated K+ (Kv) channel expression and decrease in Kv currents (/K(V)) contribute to a) increasing PASMC proliferation by raising [Ca2+]cyt via membrane depolarization and b) decreasing PASMC apoptosis by inhibiting apoptotic volume decrease and maintaining sufficient K+ in the cytoplasm to inhibit caspases and nucleases. Our data showed that the expression and activity of Kv channels are reduced, while proliferation is enhanced and apoptosis is inhibited, in IPAH-PASMC compared to normal PASMC. MicroRNAs (miRNAs) are short RNAs that bind to complementary sequences in the 3'-untranslated regions (3'-UTR) of target mRNAs to inhibit mRNA translation and/or induce mRNA degradation, thereby inhibiting the expression of specific mRNA targets. miRNAs are thus posttranscriptional regulators that potentially regulate the level of Kv channel mRNAs in IPAH-PASMC. We recently identified several miRNAs (e.g., miR-29b, miR- 138 and miR-222) that are highly expressed, whereas several Kv channels (e.g., KCNA1-7/10) are significantly downregulated, in IPAH-PASMC compared to normal PASMC. Overexpression of miR-29b, miR-138 or miR- 222 in normal PASMC decreases whole-cell /K(V), whereas inhibition of miR-29b rescues (i.e., significantly enhances) /K(V) in IPAH-PASMC. These data suggest that miR-138, miR-222 and miR-29b are sufficient to decrease /K(V) in normal PASMC, while miR-29b is necessary for the decreased /K(V) in IPAH-PASMC. Based on these observations, we hypothesize that selectively upregulated miRNAs are involved in the posttranscriptional inhibition of Kv channels in IPAH-PASMC and the miRNA-mediated Kv channel inhibition contributes to increasing proliferation and decreasing apoptosis in IPAH-PASMC. Three Specific Aims are proposed to test this hypothesis: 1) To identify the miRNAs that are upregulated and the Kv channels that are downregulated in IPAH-PASMC, and to determine which Kv channel subunits are posttranscriptionally regulated by the upregulated miRNAs in IPAH-PASMC; 2) To determine whether upregulated miRNAs in IPAH-PASMC regulate the stability and translation of target Kv channel mRNAs; and 3) To determine the role of miRNA-mediated posttranscriptional inhibition of Kv channels in PASMC proliferation and apoptosis. The long-term goal of this study is to define the mechanism underlying the inhibition of Kv channels in IPAH- PASMC and to explore the possibility to target miRNA for developing therapeutic approaches for IPAH. PUBLIC HEALTH RELEVANCE: Idiopathic pulmonary arterial hypertension (IPAH) is a rare and fatal disease that predominantly affects young women. Abnormalities in pulmonary arteries, the blood vessels in the lungs, have been shown to relate to the elevated blood pressure in the lungs. This study is designed to determine the role of microRNA- mediated regulation of potassium channels in the cellular processes that lead to the high blood pressure in the lungs of patients with this devastating disease, and to reveal potential novel targets for therapeutic approaches to treat IPAH.