Abstract
ObjectiveThis study aimed to determine the effects of emodin on the viability, proliferation and apoptosis of human pulmonary artery smooth muscle cells (PASMCs) under hypoxia and to explore the underling molecular mechanisms.MethodsPASMCs were cultured in a hypoxic environment (1% oxygen) and then treated with emodin. Cell viability, proliferation and apoptosis were evaluated using CCK-8 assay, EdU staining assay, western blot and Mito-tracker red CMXRos and Annexin V-FITC apoptosis detection assay. The microRNA (miRNA)/mRNA and protein expression levels were assessed by quantitative real-time PCR and western blotting, respectively. Based on transcriptomics and proteomics were used to identify potential signaling pathways. Luciferase reporter assay was utilized to examine the interaction between miR-244-5p and DEGS1.ResultsEmodin at 40 and 160 µM concentration-dependently suppressed cell viability, proliferation and migration, but enhanced cell apoptosis of PASMCs under hypoxia. Transcriptomic and proteomic analysis revealed that emodin could attenuate the activity of PI3K/Akt signaling in PASMCs under hypoxia. In addition, delta 4-desaturase, sphingolipid 1 (DEGS1) was found to be a direct target of miR-244-5p. Emodin could significantly up-regulated miR-244-5p expression and down-regulated DEGS1 expression in PASMCs under hypoxia. Furthermore, emodin-mediated effects on cell viability, migration, apoptosis and PI3K/Akt signaling activity of PASMCs under hypoxia were significantly attenuated by miR-244-5p knockdown.ConclusionsOur results indicated that emodin suppressed cell viability, proliferation and migration, promoted cell apoptosis of PASMCs under hypoxia via modulating miR-244-5p-mediated DEGS1/PI3K/Akt signaling pathway. MiR-244-5p/DEGS1 axis was initially investigated in this current study, which is expected to further the understanding of the etiology of pulmonary arterial hypertension.
Highlights
Patients with pulmonary arterial hypertension (PAH) mainly present with shortness of breath and progressive right heart failure
This study indicated that treatment with emodin may achieve therapeutic effect by targeting the miR244-5p/delta 4-desaturase, sphingolipid 1 (DEGS1) axis, so as to inhibit cell viability and proliferation, and promote cell apoptosis of pulmonary artery smooth muscle cells (PASMCs), introducing a new potential approach to treat PAH
Proliferation and promoted apoptosis of PASMCs under hypoxia As shown in Additional file 1: Fig. S2, the Cell Counting Kit-8 (CCK-8) assay results demonstrated that hypoxia treatment increased the cell viability of PASMCs notably at 24 h when compared with the normoxic treatment
Summary
Patients with pulmonary arterial hypertension (PAH) mainly present with shortness of breath and progressive right heart failure. Despite of the attempts in current pharmacotherapy which targets several signaling pathways including nitric oxide, endothelin and prostacyclin [3], PAH is a devastating and lethal cardio-pulmonary disease without effective treatment [4]. It has been revealed by histopathologic observations that in patients with PAH, the proliferation of pulmonary artery smooth muscle cells (PASMCs) were promoted, which were refractory to routine treatments [5]. Studies have demonstrated that protein expression of hypoxia-inducible factors (HIF) including HIF-1α and HIF-2α in PASMCs were up-regulated in PAH patients [7,8,9]. The hyperproliferation of PASMCs is recognized as a promising target for intervention in PAH-related vascular remodeling, it is of great scientific significance to explore the molecular mechanisms underlying PASMC hyperproliferation, in order to develop potential new therapies for PAH treatment
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