Pulmonary Hypertension Induces Serotonin Hyperreactivity and Metabolic Reprogramming in Coronary Arteries via NOX1/4-TRPM2 Signaling Pathway.

BACKGROUND: Clinical evidence revealed abnormal prevalence of coronary artery (CA) disease in patients with pulmonary hypertension (PH). The mechanistic connection between PH and CA disease is unclear. Serotonin (5-hydroxytryptamine), reactive oxygen species, and Ca2+ signaling have been implicated in both PH and CA disease. Our recent study indicates that NOXs (NADPH [nicotinamide adenine dinucleotide phosphate] oxidases) and TRPM2 (transient receptor potential cation channel subfamily M member 2) are key components of their interplay. We hypothesize that activation of the NOX-TRPM2 pathway facilitates the remodeling of CA in PH. METHODS: Left and right CAs from chronic hypoxia and monocrotaline-induced PH rats were collected to study vascular reactivity, gene expression, metabolism, and mitochondrial function. Inhibitors or specific siRNA were used to examine the pathological functions of NOX1/4-TRPM2 in CA smooth muscle cells. RESULTS: Significant CA remodeling and 5-hydroxytryptamine hyperreactivity in the right CA were observed in PH rats. NOX1/4-mediated reactive oxygen species production coupled with TRPM2-mediated Ca2+ influx contributed to 5-hydroxytryptamine hyperresponsiveness. CA smooth muscle cells from chronic hypoxia-PH rats exhibited increased proliferation, migration, apoptosis, and metabolic reprogramming in an NOX1/4-TRPM2-dependent manner. Furthermore, the NOX1/4-TRPM2 pathway participated in mitochondrial dysfunction, involving mitochondrial DNA damage, reactive oxygen species production, elevated mitochondrial membrane potential, mitochondrial Ca2+ accumulation, and mitochondrial fission. In vivo knockdown of NOX1/4 alleviated PH and suppressed CA remodeling in chronic hypoxia rats. CONCLUSIONS: PH triggers an increase in 5-hydroxytryptamine reactivity in the right CA and provokes metabolic reprogramming and mitochondrial disruption in CA smooth muscle cells via NOX1/4-TRPM2 activation. This signaling pathway may play an important role in CA remodeling and CA disease in PH.

Reduced CircSMOC1 Level Promotes Metabolic Reprogramming via PTBP1 (Polypyrimidine Tract-Binding Protein) and miR-329-3p in Pulmonary Arterial Hypertension Rats.

BACKGROUND: Pulmonary arterial hypertension maintains rapid cell proliferation and vascular remodeling through metabolic reprogramming. Recent studies suggested that circRNAs play important role in pulmonary vascular remodeling and pulmonary arterial smooth muscle cells proliferation. However, the relationship between circRNA, cell proliferation, and metabolic reprogramming in pulmonary arterial hypertension has not been investigated. METHODS: RNA-seq and qRT-PCR reveal the differential expression profile of circRNA in pulmonary arteries of pulmonary arterial hypertension rat models. Transfection was used to examine the effects of circSMOC1 on pulmonary artery smooth muscle cells, and the roles of circSMOC1 in vivo were investigated by adenoassociated virus. Mass spectrometry, RNA pull-down, RNA immunoprecipitation, and dual-luciferase reporter assay were performed to investigate the signaling pathway of circSMOC1 regulating the metabolic reprogramming. RESULTS: CircSMOC1 was significantly downregulated in pulmonary arteries of pulmonary arterial hypertension rats. CircSMOC1 knockdown promoted proliferation and migration and enhanced aerobic glycolysis of pulmonary artery smooth muscle cells. CircSMOC1 overexpression in vivo alleviates pulmonary vascular remodeling, right ventricular pressure, and right heart hypertrophy. In the nucleus, circSMOC1 directly binds to PTBP1 (polypyrimidine tract-binding protein), competitively inhibits the specific splicing of PKM (pyruvate kinase M) premRNA, resulting in the upregulation of PKM2 (pyruvate kinase M2), the key enzyme of aerobic glycolysis, to enhance glycolysis. In the cytoplasm, circSMOC1 acted as a miR-329-3p sponge, and its reduction in pulmonary arterial hypertension suppressed PDHB (pyruvate dehydrogenase E1 subunit beta) expression, leading to the impairment of mitochondrial oxidative phosphorylation. CONCLUSIONS: circSMOC1 is crucially involved in the metabolic reprogramming of pulmonary artery smooth muscle cells through PTBP1 and miR-329-3p to regulate pulmonary vascular remodeling in pulmonary arterial hypertension.

[TRPV4 channel mediates the increase of pulmonary microvascular endothelial permeability in rats with chronic hypoxic pulmonary hypertension].

The purpose of the present study was to investigate the effect of transient receptor potential vanilloid 4 (TRPV4) channel on the permeability of pulmonary microvascular endothelial cells (PMVECs) in rats with chronic hypoxia-induced pulmonary hypertension (CHPH), so as to clarify the mechanism of vascular endothelial dysfunction during the occurrence of pulmonary hypertension (PH). CHPH rat model was established by exposure to chronic hypoxia (CH) for 21 days. Primary PMVECs were cultured by adherent tissue blocks at the edge of the lung. The permeability coefficient of primary cultured PMVECs was detected by fluorescein isothiocyanate (FITC)-dextran. The structure of tight junction (TJ) was observed by transmission electron microscope. The expression of TRPV4 and TJ-related proteins, such as, Occludin, Claudin-5, ZO-1 were examined by real-time fluorescence quantitative PCR and Western blotting. The intracellular calcium concentration ([Ca2+]i) in PMVECs and its effect on PMVECs permeability were observed after the intervention of TRPV4 specific agonist GSK1016790A (GSK, 10 nmol/L) and specific inhibitor HC-067047 (HC, 1 µmol/L, 0.5 µmol/L). The results showed that the CHPH model was successfully established in rats treated with CH for 21 days. In CHPH rats, the structure of TJ was destroyed, the function of PMVECs barrier was decreased, the intercellular permeability was increased, the expression of TJ-related proteins were significantly decreased and the expression of TRPV4 was significantly increased (P < 0.01). The amplitude of [Ca2+]i in PMVECs of CHPH rats was significantly increased after activation of TRPV4. The inhibition ratio of HC on [Ca2+]i in PMVECs of CHPH rats was significantly higher than that in normal PMVECs. TRPV4 specific inhibitor HC reversed the increase of PMVECs permeability and increased the expression of three TJ-related proteins in CHPH rats (P < 0.01, P < 0.05). These results suggest that TRPV4 channel can induce endothelial dysfunction by increasing the [Ca2+]i, resulting in the destruction of TJ structure and the decrease of TJ-related proteins expression on PMVECs in CHPH rats.

Biotin provisioning by horizontally transferred genes from bacteria confers animal fitness benefits.

Insect symbionts are widespread in nature and lateral gene transfer is prevalent in insect symbiosis. However, the function of horizontally transferred genes (HTGs) in insect symbiosis remains speculative, including the mechanism that enables insects to feed on plant phloem deficient in B vitamins. Previously, we found there is redundancy in biotin synthesis pathways from both whitefly Bemisia tabaci and symbiotic Hamiltonella due to the presence of whitefly HTGs. Here, we demonstrate that elimination of Hamiltonella decreased biotin levels but elevated the expression of horizontally transferred biotin genes in whiteflies. HTGs proteins exhibit specific expression patterns in specialized insect cells called bacteriocytes housing symbionts. Complementation with whitefly HTGs rescued E. coli biotin gene knockout mutants. Furthermore, silencing whitefly HTGs in Hamiltonella-infected whiteflies reduced biotin levels and hindered adult survival and fecundity, which was partially rescued by biotin supplementation. Each of horizontally transferred biotin genes are conserved in various laboratory cultures and species of whiteflies with geographically diverse distributions, which shares an evolutionary origin. We provide the first experimental evidence that biotin synthesized through acquired HTGs is important in whiteflies and may be as well in other animals. Our findings suggest that B vitamin provisioning in animal-microbe symbiosis frequently evolved from bacterial symbionts to animal hosts through horizontal gene transfer events. This study will also shed light on how the animal genomes evolve through functional transfer of genes with bacterial origin in the wider contexts of microbial ecology.

Capillary-based microfluidic analysis systems.

An overview of the recent development of high-throughput capillary-based microfluidic analysis systems (CBMAS) is presented. The advantages and applications of these systems are discussed and illustrated by recent results obtained in the authors' group and other groups. A possible outlook for CBMAS is discussed.

An automated electrokinetic continuous sample introduction system for microfluidic chip-based capillary electrophoresis.

An automated and continuous sample introduction system for microfluidic chip-based capillary electrophoresis (CE) was developed in this work. An efficient world-to-chip interface for chip-based CE separation was produced by horizontally connecting a Z-shaped fused silica capillary sampling probe to the sample loading channel of a crossed-channel chip. The sample presentation system was composed of an array of bottom-slotted sample vials filled alternately with samples and working electrolyte, horizontally positioned on a programmable linearly moving platform. On moving the array from one vial to the next, and scanning the probe, which was fixed with a platinum electrode on its tip, through the slots of the vials, a series of samples, each followed by a flow of working electrolyte was continuously introduced electrokinetically from the off-chip vials into the sample loading channel of the chip. The performance of the system was demonstrated in the separation and determination of FITC-labeled arginine and phenylalanine with LIF detection, by continuously introducing a train of different samples. Employing 4.5 kV sampling voltage (1000 V cm(-1) field strength) for 30 s and 1.8 kV separation voltage (400 V cm(-1) field strength) for 70 s, throughputs of 36 h(-1) were achieved with <1.0% carryover and 4.6, 3.2 and 4.0% RSD for arginine, FITC and phenylalanine, respectively (n = 11). Net sample consumption was only 240 nL for each sample.