Aquaporin 1 controls the functional phenotype of pulmonary smooth muscle cells in hypoxia-induced pulmonary hypertension.

Vascular remodelling in hypoxia-induced pulmonary hypertension (PH) is driven by excessive proliferation and migration of endothelial and smooth muscle cells. The expression of aquaporin 1 (AQP1), an integral membrane water channel protein involved in the control of these processes, is tightly regulated by oxygen levels. The role of AQP1 in the pathogenesis of PH, however, has not been directly addressed so far. This study was designed to characterize expression and function of AQP1 in pulmonary vascular cells from human arteries and in the mouse model of hypoxia-induced PH. Exposure of human pulmonary vascular cells to hypoxia significantly induced the expression of AQP1. Similarly, levels of AQP1 were found to be upregulated in lungs of mice with hypoxia-induced PH. The functional role of AQP1 was further tested in human pulmonary artery smooth muscle cells demonstrating that depletion of AQP1 reduced proliferation, the migratory potential, and, conversely, increased apoptosis of these cells. This effect was associated with higher expression of the tumour suppressor gene p53. Using the mouse model of hypoxia-induced PH, application of GapmeR inhibitors targeting AQP1 abated the hypoxia-induced upregulation of AQP1 and, of note, reversed PH by decreasing both right ventricular pressure and hypertrophy back to the levels of control mice. Our data suggest an important functional role of AQP1 in the pathobiology of hypoxia-induced PH. These results offer novel insights in our pathogenetic understanding of the disease and propose AQP1 as potential therapeutic in vivo target.

Voluntary exercise does not always suppress lung cancer progression.

Physical exercise can lower lung cancer incidence. However, its effect on lung cancer progression is less understood. Studies on exercising mice have shown decreased ectopic lung cancer growth through the secretion of interleukin-6 from muscles and the recruitment of natural killer (NK) cells to tumors. We asked if exercise suppresses lung cancer in an orthotopic model also. Single-housed C57Bl/6 male mice in cages with running wheels were tail vein-injected with LLC1.1 lung cancer cells, and lung tumor nodules were analyzed. Exercise did not affect lung cancer. Therefore, we also tested the effect of exercise on a subcutaneous LLC1 tumor and a tail vein-injected B16F10 melanoma model. Except for one case of excessive exercise, tumor progression was not influenced. Moderately exercising mice did not increase IL-6 or recruit NK cells to the tumor. Our data suggest that the exercise dose may dictate how efficiently the immune system is stimulated and controls tumor progression.

Analysis of hypoxia-induced noncoding RNAs reveals metastasis-associated lung adenocarcinoma transcript 1 as an important regulator of vascular smooth muscle cell proliferation.

Vascular remodeling, a pathogenic hallmark in pulmonary hypertension, is mainly driven by a dysbalance between proliferation and apoptosis of human pulmonary artery smooth muscle cells. It has previously been shown that microRNAs are involved in the pathogenesis of pulmonary hypertension. However, the role of long noncoding RNAs has not been evaluated. long noncoding RNA expression was quantified in human pulmonary artery smooth muscle cells using PCR arrays and quantitative PCR. Knockdown of genes was performed by transfection of siRNA or GapmeR. Proliferation and migration were measured using BrdU incorporation and wound healing assays. The mouse model of hypoxia-induced PH was used to determine the physiological meaning of identified long noncoding RNAs. The expression of 84 selected long noncoding RNAs was assessed in hypoxic human pulmonary artery smooth muscle cells and the levels of metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) were significantly increased. Depletion of hypoxia-inducible factor 1α abolished the hypoxia-induced upregulation of metastasis-associated lung adenocarcinoma transcript 1 expression. Silencing of MALAT1 significantly decreased proliferation and migration of human pulmonary artery smooth muscle cells. In vivo, MALAT1 expression was significantly increased in lungs of hypoxic mice. Of note, targeting of MALAT1 by GapmeR ameliorated heart hypertrophy in mice with pulmonary hypertension. This is the first report on functional characterization of MALAT1 in the pulmonary vasculature. Our data provide evidence that MALAT1 expression is significantly increased by hypoxia, probably by hypoxia-inducible factor 1α. Intervention experiments confirmed that MALAT1 regulates the proliferative phenotype of smooth muscle cells and silencing of MALAT1 reduced heart hypertrophy in mice with pulmonary hypertension. These data indicate a potential role of MALAT1 in the pathogenesis of pulmonary hypertension. Impact statement Metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is a long noncoding RNA that mediates several biological processes. In the context of vascular biology, MALAT1 has been shown to be inducible by hypoxia and to control cell proliferation. These processes are of major importance for the pathophysiology of hypoxia-induced pulmonary hypertension (PH). Until now, the physiological role of MALAT1 in PH remains unclear. By using smooth muscle cells and by employing an established PH mouse model, we provide evidence that hypoxia induces MALAT1 expression. Moreover, depletion of MALAT1 inhibited migration and proliferation of smooth muscle cells, probably by the induction of cyclin-dependent kinase inhibitors. Of note, MALAT1 was significantly increased in mice exposed to hypoxia and silencing of MALAT1 ameliorated heart hypertrophy in mice with hypoxia-induced PH. Since vascular remodeling and right heart failure as a consequence of pulmonary pressure overload is a major problem in PH, these data have implications for our pathogenetic understanding.

The hypoxia-induced microRNA-130a controls pulmonary smooth muscle cell proliferation by directly targeting CDKN1A.

Excessive proliferation of human pulmonary artery smooth muscle cells (HPASMC) is one of the major factors that trigger vascular remodeling in hypoxia-induced pulmonary hypertension. Several studies have implicated that hypoxia inhibits the tumor suppressor p21 (CDKN1A). However, the precise mechanism is unknown. The mouse model of hypoxia-induced PH and in vitro experiments were used to assess the impact of microRNAs (miRNAs) on the expression of CDKN1A. In these experiments, the miRNA family miR-130 was identified to regulate the expression of CDKN1A. Transfection of HPASMC with miR-130 decreased the expression of CDKN1A and, in turn, significantly increased smooth muscle proliferation. Conversely, inhibition of miR-130 by anti-miRs and seed blockers increased the expression of CDKN1A. Reporter gene analysis proved a direct miR-130-CDKN1A target interaction. Exposure of HPASMC to hypoxia was found to induce the expression of miR-130 with concomitant decrease of CDKN1A. These findings were confirmed in the mouse model of hypoxia-induced pulmonary hypertension showing that the use of seed blockers against miR-130 restored the expression of CDKN1A. These data suggest that miRNA family miR-130 plays an important role in the repression of CDKN1A by hypoxia. miR-130 enhances hypoxia-induced smooth muscle proliferation and might be involved in the development of right ventricular hypertrophy and vascular remodeling in pulmonary hypertension.