Characterization of PKCα-rutin interactions and their application as a treatment strategy for pulmonary arterial hypertension by inhibiting ferroptosis.

As a common plant-derived dietary flavonoid, rutin receives widespread attention because of its good antioxidant bioactivities. Protein kinase Cα (PKCα) is a serine/threonine kinase that is involved in uncountable cellular processes, among which ferroptosis, a novel form of cell death, is triggered by lipid peroxidation and has been reported to be associated with pulmonary arterial hypertension (PAH). But it is still not well appreciated how rutin inhibits ferroptosis in PAH and what function PKCα has in this process. In this study, we first observed whether rutin could prevent PAH by attenuating ferroptosis with a PAH animal model and pulmonary artery smooth muscle cells (PASMCs) under hypoxia. Mitochondrial metabolomics and network pharmacology were employed to clarify the metabolic alterations and screen target proteins, and the results showed that PKCα was a vital node in rutin regulating mitochondrial metabolism related to ferroptosis in PAH. Based on molecular docking and multispectral analysis, we found that rutin could directly interact with PKCα through hydrogen bonds, which could induce static quenching, and then influence the secondary structure of PKCα. In conclusion, these findings mainly point to a novel mechanism that rutin protects PAH rats by modifying the structure and altering the activity of PKCα, and thus suppressing ferroptosis. This work reveals that the interaction behaviors between small molecules and bio-macromolecules are a critical factor to develop natural biological active ingredients and gives an insight into the potential applications of flavonoids in health and disease.

Insights into the interaction of cyclooxygenase and lipoxygenase with natural compound 3,4',5,7-Tetrahydroxyflavone based on multi-spectroscopic and metabolomics.

Hypoxia induce right ventricular dysfunction in human heart, but the molecular mechanism remains limited. As known, cyclooxygenases (COX) and lipoxygenases (LOX) play a key role in the cardiovascular system under hypoxia. 3,4',5,7-Tetrahydroxyflavone (THF), which widely exists in a variety of plants and vegetables, is famous for good ability to relieve cardiac injury, but the mechanism remains to be further understood. In this study, we firstly estimated the preventive role of THF against hypoxia-induced right ventricular dysfunction. Metabolomics analysis showed there were differential metabolites involved in above process, which helped us to screen the crucial regulated enzymes of these metabolites. Molecular docking and multi-spectroscopic revealed the molecular mechanism of interaction between THF and COX/LOX. Results suggested that THF bound to COX/LOX through static quenching and these bindings were driven by hydrogen bonds. After binding with THF, the secondary structure of COX/LOX was changed. In general, this study indicated that THF inhibited COX/LOX by spontaneously forming complexes with them.

CTRP9 mitigates vascular endothelial cell injury in patients with hypertensive heart disease by inhibiting PI3K/Akt/mTOR axis.

OBJECTIVE: To investigate the mechanism of factor-alpha-related protein 9 (CTRP9) in mitigating the vascular endothelial cell (VEC) injury in patients with hypertensive heart disease (HHD) by the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) axis. METHODS: 43 patients with HHD admitted to our hospital from February 2018 to February 2019 were included in the study group, and another 39 healthy controls from the same period were the reference group. The total protein of transfected VECs was detected by western blotting, and the proliferation rate of the VECs was determined by Cell Counting Kit-8 (CCK-8). The levels of CTRP9, high sensitivity C-reactive protein (hs-CRP), thrombomodulin (TM), and von Willebrand factor (vWF) were detected by ELISA. The mechanism of CTRP9 in alleviating VEC injury in HHD patients by inhibiting the PI3K/Akt/mTOR axis was analyzed. RESULTS: The two groups did not differ in terms of their general data (P>0.05). The CTRP9 level in the study group was higher than in the reference group (P<0.001). Study group had higher levels of endothelin-1 (ET-1), hs-CRP, TM, vWF (P<0.001), and markedly lower phospho-PI3K (p-PI3K) and phospho-protein kinase B (p-AKT) protein levels (P<0.05). Compared to the reference group, the proliferation capacity of trophoblast cells in the study group was sharply decreased (P<0.05). The study group had lower phosphorylation levels of PI3K, Akt, and mTOR proteins than the reference group (P<0.05). Phosphorylation of Akt occurred at 15 min and reached its peak at 30 min. A drastically reduced invasion capacity of VECs was observed in the study group compared to the reference group (P<0.05). CONCLUSIONS: CTRP9 mitigates VEC injury in patients with HHD by inhibiting the PI3K/Akt/mTOR axis.

Targeted metabolomics combined with network pharmacology to reveal the protective role of luteolin in pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is a progressive disease that significantly endangers human health, where metabolism may drive pathogenesis: a shift from mitochondrial oxidation to glycolysis occurs in diseased pulmonary vessels and the right ventricle. An increase in pulmonary vascular resistance in patients with heart failure with a preserved ejection fraction portends a poor prognosis. Luteolin exists in numerous foods and is marketed as a dietary supplement assisting in many disease treatments. However, little is known about the protective effect of luteolin on metabolism disorders in diseased pulmonary vessels. In this study, we found that luteolin apparently reversed the pulmonary vascular remodeling of PAH rats by inhibiting the abnormal proliferation of pulmonary artery smooth muscle cells (PASMCs). Moreover, network pharmacology and metabolomics results revealed that the arachidonic acid pathway, amino acid pathway and TCA cycle were dysregulated in PAH. A total of 14 differential metabolites were significantly changed during the PAH, including DHA, PGE2, PGD2, LTB4, 12-HETE, 15-HETE, PGF2α, and 8-iso-PGF2α metabolites in the arachidonic acid pathway, and L-asparagine, oxaloacetate, N-acetyl-L-ornithine, butane diacid, ornithine, glutamic acid metabolites in amino acid and TCA pathways. However, treatment with luteolin recovered the LTB4, PGE2, PGD2, 12-HETE, 15-HETE, PGF2α and 8-iso-PGF2α levels close to normal. Meanwhile, we showed that luteolin also downregulated the gene and protein levels of COX 1, 5-LOX, 12-LOX, and 15-LOX in the arachidonic acid pathway. Collectively, this work highlighted the metabolic mechanism of luteolin-protected PAH and showed that luteolin would hold great potential in PAH prevention.

Integrated metabolomics and mechanism to reveal the protective effect of kaempferol on pulmonary arterial hypertension.

Currently, there have been breakthroughs in the study of the underlying mechanisms of pulmonary arterial hypertension, but treatment is still challenging. The aim of this study was explore the effect of kaempferol in PAH and discover the mechanism of this process. First, we assessed the effect of kaempferol in animal models of PAH. The echocardiography and pressure-volume admittance catheter data showed that kaempferol could improve right ventricular function and alleviate the progression of PAH. Moreover, morphometric analysis of the lung vasculature indicated that pulmonary vascular remodeling could be reduced by kaempferol. The expression of autophagy-related proteins and autophagic processes were evaluated by western blotting and mRFP-GFP-LC3 fluorescence microscopy, the results indicated that autophagy played an important role in the process by which kaempferol prevents PAH. Moreover, we performed untargeted and targeted plasma metabolomics analysis on rat models of PAH and used multivariate analysis to reveal multiple pathways and targets. Consequently, we identified 247 plasma biomarkers that were tightly linked to the preventive effects of kaempferol, which were mainly related to amino acid metabolism and arachidonic acid metabolism. In conclusion, kaempferol could effectively alleviate the development of PAH by regulating abnormal autophagy and metabolic disorders, providing a novel perspective on the treatment of PAH.

microRNA-103a-3p confers protection against lipopolysaccharide-induced sepsis and consequent multiple organ dysfunction syndrome by targeting HMGB1.

BACKGROUND: Sepsis and subsequent multiple organ dysfunction syndrome (MODS) have high global incidence and mortality rate, imposing tremendous health burden. microRNAs (miRNAs or miRs) are implicated in the pathogenesis of sepsis and MODS. The aim of this study is to explore the potential mechanisms of miR-103a-3p targeted high mobility group box 1 (HMGB1) involvement in the pathogenesis of sepsis complicated with multiple organ dysfunction syndrome (MODS). METHODS: A mouse sepsis model was induced by lipopolysaccharide (LPS). Bone marrow-derived macrophages were collected and LPS was used to establish a cellular inflammation model. Targeted binding between miR-103a-3p and HMGB1 was verified by a double luciferase assay and their roles in LPS-induced sepsis were further explored using gain-of-function experiments. RESULTS: miR-103a-3p was decreased while HMGB1 was increased in sepsis. In LPS-induced mouse sepsis models, the downregulation of HMGB1 was found to result in reductions in NO, TNF-α, IL-1ß, IL-6, lung myeloperoxidase activity, pulmonary microvascular albumin leakage, serum alanine aminotransferase, aspartate aminotransferase activity, and lung and liver tissue apoptosis. Additionally, decreased HMGB1 blunted the inflammatory response and increased survival rate of modeled mice. Importantly, HMGB1 was confirmed to a target gene of miR-103a-3p. In cellular inflammation models, miR-103a-3p was found to alleviate LPS-induced sepsis and MODS in vitro by decreasing HMGB1. CONCLUSIONS: Taken together, our results demonstrated the inhibitory role of miR-103a-3p in sepsis via inhibiting HMGB1 expression.