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Cardiac ischemia–reperfusion injury (IRI) occurs when blood flow is restored to the myocardium after a period of ischemia, leading to oxidative stress and subsequent myocardial cell damage, primarily due to the accumulation of reactive oxygen species (ROS). In our previous research, we identified that miR-25 is significantly overexpressed in pressure overload-induced heart failure, and its inhibition improves cardiac function by restoring the expression of SERCA2a, a key protein involved in calcium regulation. In this study, we aimed to investigate the role of miR-25 in the context of ischemia–reperfusion injury. We found that miR-25 was markedly upregulated under hypoxic conditions in both in vitro and in vivo models. Through in silico analysis, we identified Sestrin3 (SESN3), an antioxidant protein known for its protective effects against oxidative stress, as a novel target of miR-25. Based on these findings, we hypothesized that inhibiting miR-25 would restore Sestrin3 expression, thereby reducing ROS-induced myocardial cell damage and improving cardiac function. To test this hypothesis, we employed two model systems: a hypoxia/reoxygenation (H/R) stress model using H9c2 myoblasts and a surgically induced ischemia–reperfusion injury mouse model. Our results demonstrated that the use of miR-25 inhibitors significantly improved cardiac function and reduced myocardial damage in both models through the restoration of SESN3 expression. In conclusion, our findings suggest that targeting miR-25 may serve as a novel therapeutic modality to alleviate oxidative damage in the heart. [ABSTRACT FROM AUTHOR]

Hypoxia/reoxygenation (HR) often occurs under cardiac pathological conditions, and HR-induced oxidative stress usually leads to cardiomyocyte damage. Carvedilol, a non-selective β-blocker, is used clinically to treat cardiac ischemia diseases. Moreover, Carvedilol has also been reported to have an antioxidant ability by reducing lipid peroxidation. However, the mechanism of Carvedilol to inhibit lipid peroxidation is still elusive. To explore the protective mechanism of Carvedilol to resist lipid peroxidation on cardiomyocytes, HL-1 cells were cultured under normoxia, hypoxia, and HR and treated with Carvedilol to investigate the alteration on metabolism, protein expression, and mRNA level to explain its oxidative mechanism. The study found that Carvedilol upregulated glutathione peroxidase 4 (GPX4) protein expression to resist HR-induced lipid peroxidation by metabolic remodeling under HR. Also, Carvedilol promoted ferroptosis-related genes, ferritin heavy chain 1 (FTH1) and ferritin light chain 1 (FTL1) mRNA levels, to reduce lipid peroxidation under both hypoxia and HR. In conclusion, our study explores a mechanism by which Carvedilol inhibits ferroptosis by upregulating GPX4, FTH1, and FTL1 levels to downregulate lipid peroxidation under HR. The study provides a potential strategy for using Carvedilol in clinical applications, inspiring further research and development in the area of heart diseases. [ABSTRACT FROM AUTHOR]

Background: Sevoflurane (Sevo) prevents hypoxia/reoxygenation (H/R)-induced cardiomyocytes injury. The expression of miR-4,454 was increased in individuals experiencing an acute myocardial infarction. Objective: The purpose of current investigation was to delved into whether the effects of Sevo on cardiomyocytes are mediated through regulation of miR-4,454 expression. Method: In this study, the expression levels of miR-4,454 and BAG5 were detected by real-time quantitative polymerase chain reaction (RT-qPCR). Cell viability was detected by cell counting kit-8 (CCK-8). The levels of creatine kinase-MB (CK-MB), lactate dehydrogenase (LDH) and cardiac troponin I (cTnI) were detected by enzyme-linked immunosorbent assay (ELISA). Reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD) and catalase (CAT) were detected using various commercially available kits to assess the level of oxidative stress in the cells. The luciferase reporter gene assay was used to verify the interaction of miR-4,454 with downstream target genes. Results: There was a notable upregulation of miR-4,454 expression in H/R-induced cardiomyocyte models. This was accompanied by a decrease in the viability of myocardial cells induced by H/R and an intensification of the extent of myocardial injury and oxidative stress. However, the detrimental effects were mitigated by the administration of Sevo. miR-4,454 had a target site for binding to BAG5, and its expression was negatively modulated by miR-4,454. An increase in the expression of BAG5 was shown to directly offset the exacerbation of cardiomyocyte damage induced by the overexpression of miR-4,454. Conclusion: Sevo may attenuate H/R-induced cardiomyocyte injury by regulating miR-4454. [ABSTRACT FROM AUTHOR]

Objective To explore the role and mechanism of pumilio homolog 1 （PUM1） in hypoxia/reoxygenation induced cell injury in human renal tubular epithelial cells （HK-2）. Methods A hypoxia/reoxygenation （H/R） model of HK-2 cells was established in vitro. PUM1 expression was knocked down through small interfering RNA （siRNA）. Cells were randomized into three groups of control， hypoxia/reoxygenation （H/R） and H/R+siRNA. Western blot （WB） method was used for detecting the expression level of PUM1 protein. Cell Count Kit 8 （CCK-8） was employed for detecting cell viability. Hydrogen peroxide （H2O2）， malondialdehyde （MDA） and superoxide dismutase （SOD） were used for evaluating the levels of oxidative stress. Flow cytometry was utilized for detecting the level of cell apoptosis. Results As compared with control group， protein expression level of PUM1 in H/R 3 h group （1.76 ± 0.11 vs 0.98 ± 0.05）， H/R 6 h group （2.89 ± 0.14 vs 0.98 ± 0.05） and H/R 12 h group （3.78 ± 0.08 vs 0.98 ± 0.05） gradually spiked with the prolongation of hypoxic time. As compared with H/R group， knocking down the expression of PUM1 significantly improved the cell viability （73.67 ± 3.42 vs 29.60 ± 2.94）， oxidative stress [H2O2:（13.53 ± 0.85）μmol/L vs （22.43 ± 1.12）μmol/L， MDA: （16.03 ± 0.70）μmol/L vs （31.20 ± 1.50）μmol/L， SOD: （34670 ± 1800）U/L vs （5730 ± 1220）U/L] and apoptotic level [（14.89 ± 1.65）% vs （39.71 ± 1.94）%] after H/R in H/R+si-PUM1 group. Conclusion PUM1 is up-regulated in H/R induced HK-2 cells and its inhibition may alleviate H/R injury through reducing oxidative stress and lowering cell apoptosis levels.

Objective To explore the therapeutic mechanism of cordycepin for acute kidney injury （AKI） based upon gene chips in gene expression omnibus （GEO） database, network pharmacology, molecular docking and in vitro experiments. Methods Targeted genes of AKI were obtained from two databases, GEO （GSE87025） and GeneCards. Cordycepin-related targets were retrieved from three databases of TargetNet, BATMAN and GeneCards. The common targets of AKI and cordycepin were screened by taking the intersection of two sets. Molecular docking was performed for cordycepin and key targets after gene ontology （GO） annotation and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis. Cell experiments were performed for control, hypoxia/reoxygenation and hypoxia/reoxygenation+cordycepin groups for experimental verifications of the relevant targets. Results There were 12 common targets of AKI and cordycepin. GO functional annotation results indicated that these targets participated predominantly in negative regulation of apoptotic signaling pathway and necroptosis process. KEGG enrichment analysis results showed that they played some roles in signaling pathways of Toll-like receptor, necroptosis, nucleotide-binding oligomerization domain （NOD）-like receptor, nuclear factor-κB （NF-kB）, tumor necrosis factor （TNF） and cell apoptosis, etc. Optimal binding energy between cordycepin and receptor interacting protein kinase 1 （RIPK1） in molecular docking was -7.1, denoting potent binding activity. Western blot indicated that cordycepin down-regulated the expressions of RIPK1, bcl-2 associated x protein （Bax） and caspase 3 and up-regulated B cell lymphoma/lewkmia-2（Bcl-2）. Cell immunofluorescence results revealed that cordycepin down-regulated the expression of RIPK1. Conclusion Cordycepin may alleviate hypoxia/reoxygenation-induced renal tubular epithelial cell injury through regulating RIPK1-mediated apoptosis. Thus it achieves the therapeutic efficacy for AKI.

Calcium influx into neurons triggers neuronal death during cerebral ischemia/reperfusion injury. Various calcium channels are involved in cerebral ischemia/reperfusion injury. Cav3.2 channel is a main subtype of T-type calcium channels. T-type calcium channel blockers, such as pimozide and mibefradil, have been shown to prevent cerebral ischemia/reperfusion injury-induced brain injury. However, the role of Cav3.2 channels in cerebral ischemia/reperfusion injury remains unclear. Here, in vitro and in vivo models of cerebral ischemia/reperfusion injury were established using middle cerebral artery occlusion in mice and high glucose hypoxia/reoxygenation exposure in primary hippocampal neurons. The results showed that Cav3.2 expression was significantly upregulated in injured hippocampal tissue and primary hippocampal neurons. We further established a Cav3.2 gene-knockout mouse model of cerebral ischemia/reperfusion injury. Cav3.2 knockout markedly reduced infarct volume and brain water content, and alleviated neurological dysfunction after cerebral ischemia/reperfusion injury. Additionally, Cav3.2 knockout attenuated cerebral ischemia/reperfusion injury-induced oxidative stress, inflammatory response, and neuronal apoptosis. In the hippocampus of Cav3.2-knockout mice, calcineurin overexpression offset the beneficial effect of Cav3.2 knockout after cerebral ischemia/reperfusion injury. These findings suggest that the neuroprotective function of Cav3.2 knockout is mediated by calcineurin/nuclear factor of activated T cells 3 signaling. Findings from this study suggest that Cav3.2 could be a promising target for treatment of cerebral ischemia/reperfusion injury. [ABSTRACT FROM AUTHOR]

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Background: Hypoxia is one of the most significant pathogenic factors in cardiovascular diseases. Preclinical studies suggest that nonpsychoactive cannabidiol (CBD) and β-adrenoceptor stimulation might possess cardioprotective potential against ischemia-reperfusion injury. The current study evaluates the influence of hypoxia-reoxygenation (H/R) on the function of atria and ventricular papillary muscles in the presence of CBD and the nonselective β-adrenoceptor agonist isoprenaline (ISO). Methods: The concentration curves for ISO were constructed in the presence of CBD (1 µM) before or after H/R. In chronic experiments (CBD 10 mg/kg, 14 days), the left atria isolated from spontaneously hypertensive (SHR) and their normotensive control (WKY) rats were subjected to H/R following ISO administration. Results: Hypoxia decreased the rate and force of contractions in all compartments. The right atria were the most resistant to hypoxia regardless of prior β-adrenergic stimulation. Previous β-adrenergic stimulation improved recovery in isolated left atria and right (but not left) papillary muscles. Acute (but not chronic) CBD administration increased the effects of ISO in left atria and right (but not left) papillary muscles. Hypertension accelerates left atrial recovery during reoxygenation. Conclusions: H/R directly modifies the function of particular cardiac compartments in a manner dependent on cardiac region and β-adrenergic prestimulation. The moderate direct cardioprotective potential of CBD and β-adrenergic stimulation against H/R is dependent on the cardiac region, and it is less than in the whole heart with preserved coronary flow. In clinical terms, our research expands the existing knowledge about the impact of cannabidiol on cardiac ischemia, the world′s leading cause of death. [ABSTRACT FROM AUTHOR]

Objective：To investigate the impact of pumpkin polysaccharide (PP) on hypoxia/reoxygenation (H/R) -induced myocardial cell (H9C2) injury by regulating Nrf2/γ-GCS pathway. Methods：HR cell models were established, and the concentration of pumpkin polysaccharide (PP) was determined. H9C2 cells were grouped into Control group, H/R group, pumpkin polysaccharide group (PP group, 2 mg/L PP), Nrf2 inhibitor group (ML385 group, 2.0 μmol/L ML385), and PP+ML385 group (2.0 mg/L PP+ 2.0 μmol/L ML385), the apoptosis, mitochondrial membrane potential changes, oxidative stress (LDH, MDA, SOD, ROS), the levels of inflammatory factors (TNF-α, IL-6, IL-1β) and the protein expressions of Nrf2 and γ-GCS in H9C2 cells were detected, respectively. Results：Compared with Control group, H/R group had changes in cell morphology, the cell injury and mitochondrial membrane potential decreased, the H9C2 cell viability, SOD and protein expression levels of Nrf2 and γ -GCS were significantly decreased, while the apoptosis rate, LDH, MDA, ROS, and levels of TNF-α, IL-6 and IL-1β were significantly increased (P<0.05) ; compared with H/R group, the cell injury in PP group was reduced, the mitochondrial membrane potential was restored, the H9C2 cell viability, SOD and protein expression levels of Nrf2 and γ-GCS were significantly increased, and the apoptosis rate, LDH, MDA, ROS, and levels of TNF-α, IL-6 and IL-1β were significantly decreased (P<0.05), the effect of ML385 was opposite to that of PP, and ML385 could reverse the antioxidant and anti-inflammatory effects of PP. Conclusion：PP up-regulates the Nrf2/γ-GCS pathway to exert antioxidant and anti-inflammatory effects, and reduces H/R-induced H9C2 cell injury. [ABSTRACT FROM AUTHOR]

The occurrence of myocardial ischemia/reperfusion injury is commonly observed during cardiac surgery; however, there remains a dearth of effective therapeutic strategies to mitigate this injury. The a disintegrin and metallopeptidase domain 10 (ADAM10) is a transmembrane protein anchored on the cell membrane surface, and its precise mechanism of action in myocardial ischemia/reperfusion injury remains incompletely understood. This study aims to investigate the impact of ADAM10 on cardiomyocyte injury induced by hypoxia/reoxygenation (H/R) and elucidate the underlying mechanisms. The ADAM10 overexpression plasmid was transfected into H9c2 cells, which were subsequently treated with the Notch signaling pathway inhibitor DAPT and cultured under H/R conditions. Cell proliferation activity was assessed using the CCK-8 assay. The levels of LDH, SOD, and MDA were quantified through colorimetric analysis. The levels of ROS and the rate of apoptosis were measured using flow cytometry. The morphological changes in the nucleus of H9c2 cells were observed by employing Hoechst 33258 staining. The mRNA expression levels of ADAM10, Notch1, NICD, and Hes1 in H9c2 cells were determined using qRT-PCR. The expressions of Notch signaling pathway and apoptosis-related proteins were analyzed by Western blot. Overexpression of ADAM10 provided protection to H9c2 cells against injury induced by H/R, leading to an increase in SOD levels and alleviation of oxidative stress caused by the accumulation of ROS and the decrease of SOD activity. Meanwhile, overexpression of ADAM10 inhibited apoptosis in H9c2 cells exposed to H/R by regulating the expression of apoptosis-related proteins, such as Bax, Bcl-2 and Cleaved-caspase-3. Additionally, overexpression of ADAM10 facilitated the activation of the Notch1 signaling pathway in H9c2 cells exposed to H/R by upregulating the protein expression of Notch1, NICD, and Hes1. However, the protective effect of ADAM10 on H/R-induced H9c2 cells was partially reversed by DAPT. Our findings demonstrate that ADAM10 exerts protective effects in H/R-induced H9c2 cells by suppressing oxidative stress and apoptosis via the activation of the Notch signaling pathway. [ABSTRACT FROM AUTHOR]

Although our previous studies have established the crucial role of RP105 in myocardial ischemia/reperfusion injury (MI/RI), its involvement in regulating oxidative stress induced by MI/RI remains unclear. To investigate this, we conducted experiments using a rat model of ischemia/reperfusion (I/R) injury. Adenovirus carrying RP105 was injected apically at multiple points, and after 72 h, the left anterior descending coronary artery was ligated for 30 min followed by 2 h of reperfusion. In vitro experiments were performed on H9C2 cells, which were transfected with recombinant adenoviral vectors for 48 h, subjected to 4 h of hypoxia, and then reoxygenated for 2 h. We measured oxidative stress markers, including superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities, as well as malondialdehyde (MDA) concentration, using a microplate reader. The fluorescence intensity of reactive oxygen species (ROS) in myocardial tissue was measured using a DHE probe. We also investigated the upstream and downstream components of the signal transducer and activator of transcription 3 (STAT3). Upregulation of RP105 increased SOD and GSH-Px activities, reduced MDA concentration, and inhibited ROS production in response to I/R injury in vivo and hypoxia reoxygenation (H/R) stimulation in vitro. The overexpression of RP105 led to a decrease in the myocardial enzyme LDH in serum and cell culture supernatant, as well as a reduction in infarct size. Additionally, left ventricular fraction (LVEF) and fractional shortening (LVFS) were improved in the RP105 overexpression group compared to the control. Upregulation of RP105 promoted the expression of Lyn and Syk and further activated STAT phosphorylation, which was blocked by PP2 (a Lyn inhibitor). Our findings suggest that RP105 can inhibit MI/RI-induced oxidative stress by activating STAT3 via the Lyn/Syk signaling pathway. [ABSTRACT FROM AUTHOR]

To investigate the effects of Cirsilineol on cell viability, pyroptosis, and inflammation in hypoxia/reoxygenation-induced renal tubular cells, we utilized the renal tubular epithelial cell line HK2. The in vitro renal injury model was induced by hypoxia/reoxygenation. Cell viability in hypoxia/reoxygenation-induced HK2 cells was determined using a methylthiazolyldiphenyltetrazolium bromide assay and lactate dehydrogenase releasing kit. The effects of Cirsilineol on inflammation and pyroptosis in HK2 cells were evaluated using enzyme-linked immunosorbent assay, flow cytometry, and immunoblot assays. Our results demonstrated that Cirsilineol promoted survival in hypoxia/reoxygenation-induced tubular cells, inhibited inflammation, and reduced pyroptosis in these cells. Further investigation revealed that the protective effects of Cirsilineol are mediated by the suppression of the Toll-like receptor 4/nuclear factor kappa B pathway, which mitigates the cellular response to hypoxia/ reoxygenation injury. Our study suggests that Cirsilineol protects renal tubular epithelial cells from hypoxia/reoxygenation injury by inhibiting inflammation and pyroptosis, highlighting its potential therapeutic value in renal conditions characterized by ischemic injury. [ABSTRACT FROM AUTHOR]

Objective: To investigate the protective effects and mechanisms of salvianolic acid B (Sal B) on hypoxia/reoxygenation H/R-induced renal tubular epithelial cell damage. Methods In vitro experimental cells were divided into the normal control Con groupthe Sal B+Con groupthe H/R groupthe Sal B+H/R group as well as the Con groupthe H/R groupSal B+H/R groupthe H/R+CC group and the H/R+Sal B+CC group. HK-2 cell viability was detected by the CCK-8 method. Cell lysates were collected to measure the levels of GSH and malondialdehyde MDA using an enzyme-linked immunosorbent assay. The expressions of p-AMPK GPX4 ACSL4 and FSP1 were detected by Western blot. The expression of ferroptosis markers GPX4 and ACSL4 proteins was detected by cell immunofluorescence while mitochondrial membrane potential and cell apoptosis were measured by flow cytometry.Results The results of the in vitro experiments showed that Sal B had no significant toxicity on normal HK-2 cell viability within the concentration range of 20-160 μmol/L P>0.05. Compared with the model group Sal B at concentrations of 20 40 and 80 μmol/L dose-dependently increased the viability of HK-2 cells P<0.01. Compared with the model group MDA content and GSH content of the H/R+Sal B group were decreased p-AMPK GPX4 and FSP1 protein expression levels were increased ACSL4 protein expression was decreased mitochondrial membrane potential was significantly increased and apoptosis rate was significantly decreased P<0.05. Compared with the H/R+Sal B group the FSP1 and GPX4 protein expression level in the H/R+AMPK group was decreased the ACSL4 protein expression level was increased P< 0.05 and the mitochondrial membrane potential was significantly decreased P<0.05 and the addition of Sal B could reverse this situation P<0.05.Conclusion Salvianolic acid B can alleviate I/R-induced AKI and its mechanism may involve the activation of the AMPK pathway to mitigate ferroptosis in renal tubular epithelial cells. [ABSTRACT FROM AUTHOR]

BACKGROUND: There are currently no effective drugs to mitigate the ischemia/reperfusion injury caused by fluid resuscitation after hemorrhagic shock (HS). The aim of this study was to explore the potential of the histone deacetylase 6 (HDAC6)-specific inhibitor tubastatin A (TubA) to suppress nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) inflammasome activation in macrophages under hypoxia/reoxygenation (H/R) conditions. METHODS: The viability of RAW264.7 cells subjected to H/R after treatment with different concentrations of TubA was assessed using a cell-counting kit-8 (CCK8) assay. Briefly, 2.5 µmol/L TubA was used with RAW264.7 cells under H/R condition. RAW264.7 cells were divided into three groups, namely the control, H/R, and TubA groups. The levels of reactive oxygen species (ROS) in the cells were detected using fluorescence microscopy. The protein expression of HDAC6, heat shock protein 90 (Hsp90), inducible nitric oxide synthase (iNOS), NLRP3, gasdermin-D (GSDMD), Caspase-1, GSDMD-N, and Caspase-1 p20 was detected by western blotting. The levels of interleukin-1β (IL-1β) and IL-18 in the supernatants were detected using enzyme-linked immunosorbent assay (ELISA). RESULTS: HDAC6, Hsp90, and iNOS expression levels were significantly higher (P<0.01) in the H/R group than in the control group, but lower in the TubA group than in the H/R group (P<0.05). When comparing the H/R group to the control group, ROS levels were significantly higher (P<0.01), but significantly reduced in the TubA group (P<0.05). The H/R group had higher NLRP3, GSDMD, Caspase-1, GSDMD-N, and Caspase-1 p20 expression levels than the control group (P<0.05), however, the TubA group had significantly lower expression levels than the H/R group (P<0.05). IL-1β and IL-18 levels in the supernatants were significantly higher in the H/R group compared to the control group (P<0.01), but significantly lower in the TubA group compared to the H/R group (P<0.01). CONCLUSION: TubA inhibited the expression of HDAC6, Hsp90, and iNOS in macrophages subjected to H/R. This inhibition led to a decrease in the content of ROS in cells, which subsequently inhibited the activation of the NLRP3 inflammasome and the secretion of IL-1β and IL-18. [ABSTRACT FROM AUTHOR]

A new benzofuran, named as (2S, 3S)-5,6-dimethoxy-3-methyl-2-(3'-hydroxyphenyl)-2,3-dihydrobenzofuran (1), along with the six known isoflavonoids, 2'-hydroxy-4',7-dimethoxyisoflavone (2), 2' ²-methoxybiochanin A (3), tectorigenin (4), calycosin (5), 7-hydroxy-2',4',5'-trimethoxyisoflavone (6), orobol (7) were isolated from the heartwood of Dalbergia odorifera T.Chen. The structure of compounds was characterised by NMR spectroscopic data and comparisons with relevant literature data. The absolute structural configuration of compound 1 was determined through X-ray single crystal diffraction. Moreover, compounds 1-7 have no significant cytotoxic effects on H9c2 cells (IC50 > 200 μM). Compound 1-7 exhibit a significant protective effect against H/R (hypoxia/reoxygenation) induced H9c2 cell damage at 10~40, 5~40, 5~40, 5~40, 5~40, 5~40 and 5~10 μM (P≩0.05). [ABSTRACT FROM AUTHOR]

This research is concentrated on investigating the role and mechanism of miR-652-3p in the protective effects of isoflurane (ISO) against myocardial ischemia–reperfusion (I/R) injury. H9c2 cells underwent pretreatment with varying concentrations of ISO, and subsequently, a hypoxia/reoxygenation (H/R) model was constructed. The levels of miR-652-3p, ISL LIM homeobox 1 (ISL1), and inflammatory cytokines interleukin (IL)-6 and tumor necrosis factor-alpha (TNF-α) were evaluated through reverse transcription polymerase chain reaction (RT-qPCR). Enzyme-linked immunosorbent assay was employed to investigate concentrations of myocardial injury markers, such as creatine kinase-MB (CK-MB) and cardiac troponin I (cTnI). Cell counting kit-8 was used to evaluate cell viability, while flow cytometry was utilized to measure apoptosis. Additionally, a dual luciferase reporter assay was conducted to validate the targeting relationship between ISL1 and miR-652-3p. Herein, we confirmed that the level of miR-652-3p was gradually increased with prolonged hypoxia; nevertheless, this increase was suppressed by ISO pretreatment (P < 0.05). Additionally, ISO pretreatment prevented the decrease in cell viability, increase in apoptosis, and overproduction of IL-6, TNF-α, CK-MB, and cTnI induced by H/R (P < 0.05). However, the inhibitory effects of ISO were counteracted by the increased levels of miR-652-3p (P < 0.05). ISL1 is a potential target of miR-652-3p. H/R induction suppressed ISL1 levels compared to the control, but ISO treatment increased its expression (P < 0.05). Overexpression of ISL1 inhibited the elimination of the protective effect of ISO on myocardial damage induced by the elevation of miR-652-3p (P < 0.05). The findings of this research confirm that miR-652-3p attenuated the protective effect of ISO on cardiomyocytes in myocardial ischemia by targeting ISL1. [ABSTRACT FROM AUTHOR]

Aims: Ischaemic heart disease remains a significant cause of mortality globally. A pharmacological agent that protects cardiac mitochondria against oxygen deprivation injuries is welcome in therapy against acute myocardial infarction. Here, we evaluate the effect of large‐conductance Ca2+‐activated K+ channels (BKCa) activator, Compound Z, in isolated mitochondria under hypoxia and reoxygenation. Methods: Mitochondria from mice hearts were obtained by differential centrifugation. The isolated mitochondria were incubated with a BKCa channel activator, Compound Z, and subjected to normoxia or hypoxia/reoxygenation. Mitochondrial function was evaluated by measurement of O2 consumption in the complexes I, II, and IV in the respiratory states 1, 2, 3, and by maximal uncoupled O2 uptake, ATP production, ROS production, transmembrane potential, and calcium retention capacity. Results: Incubation of isolated mitochondria with Compound Z under normoxia conditions reduced the mitochondrial functions and induced the production of a significant amount of ROS. However, under hypoxia/reoxygenation, the Compound Z prevented a profound reduction in mitochondrial functions, including reducing ROS production over the hypoxia/reoxygenation group. Furthermore, hypoxia/reoxygenation induced a large mitochondria depolarization, which Compound Z incubation prevented, but, even so, Compound Z created a small depolarization. The mitochondrial calcium uptake was prevented by the BKCa activator, extruding the mitochondrial calcium present before Compound Z incubation. Conclusion: The Compound Z acts as a mitochondrial BKCa channel activator and can protect mitochondria function against hypoxia/reoxygenation injury, by handling mitochondrial calcium and transmembrane potential. [ABSTRACT FROM AUTHOR]

Renal ischemia/reperfusion is a serious condition that not only causes acute kidney injury, a severe clinical syndrome with high mortality, but is also an inevitable part of kidney transplantation or other kidney surgeries. Alterations of oxygen levels during ischemia/reperfusion, namely hypoxia/reoxygenation, disrupt mitochondrial metabolism and induce structural changes that lead to cell death. A signature mitochondrial phospholipid, cardiolipin, with many vital roles in mitochondrial homeostasis, is one of the key players in hypoxia/reoxygenation-induced mitochondrial damage. In this study, we analyze the effect of hypoxia/reoxygenation on human renal proximal tubule epithelial cell (RPTEC) cardiolipins, as well as their metabolism and mitochondrial functions. RPTEC cells were placed in a hypoxic chamber with a 2% oxygen atmosphere for 24 h to induce hypoxia; then, they were replaced back into regular growth conditions for 24 h of reoxygenation. Surprisingly, after 24 h, hypoxia cardiolipin levels substantially increased and remained higher than control levels after 24 h of reoxygenation. This was explained by significantly elevated levels of cardiolipin synthase and lysocardiolipin acyltransferase 1 (LCLAT1) gene expression and protein levels. Meanwhile, hypoxia/reoxygenation decreased ADP-dependent mitochondrial respiration rates and oxidative phosphorylation capacity and increased reactive oxygen species generation. Our findings suggest that hypoxia/reoxygenation induces cardiolipin remodeling in response to reduced mitochondrial oxidative phosphorylation in a way that protects mitochondrial function. [ABSTRACT FROM AUTHOR]

Gas-loaded nanocarriers (G-LN) show promise in improving heart transplantation (HTx) outcomes. Given their success in reducing cell death during normothermic hypoxia/reoxygenation (H/R) in vitro, we tested their integration into cardioplegic solutions and static cold storage (SCS) during simulated HTx. Wistar rat hearts underwent four hours of SCS with four G-LN variants: O2- or N2-cyclic-nigerosyl-nigerose-nanomonomers (CNN), and O2- or N2-cyclic-nigerosyl-nigerose-nanosponges (CNN-NS). We monitored physiological-hemodynamic parameters and molecular markers during reperfusion to assess cell damage/protection. Hearts treated with nanomonomers (N2-CNN or O2-CNN) showed improvements in left ventricular developed pressure (LVDP) and a trend towards faster recovery of the rate pressure product (RPP) compared to controls. However, nanosponges (N2-CNN-NS or O2-CNN-NS) did not show similar improvements. None of the groups exhibited an increase in diastolic left ventricular pressure (contracture index) during reperfusion. Redox markers and apoptosis/autophagy pathways indicated an increase in Beclin 1 for O2-CNN and in p22phox for N2-CNN, suggesting alterations in autophagy and the redox environment during late reperfusion, which might explain the gradual decline in heart performance. The study highlights the potential of nanomonomers to improve early cardiac performance and mitigate cold/H/R-induced stunning in HTx. These early improvements suggest a promising avenue for increasing HTx success. Nevertheless, further research and optimization are needed before clinical application. [ABSTRACT FROM AUTHOR]

Mitochondrial homeostasis is crucial for maintaining cellular energy production and preventing oxidative stress, which is essential for overall cellular function and longevity. Mitochondrial damage and dysfunction often occur concomitantly in myocardial ischemia–reperfusion injury (MIRI). Notoginsenoside R1 (NGR1), a unique saponin from the traditional Chinese medicine Panax notoginseng, has been shown to alleviate MIRI in previous studies, though its precise mechanism remains unclear. This study aimed to elucidate the mechanisms of NGR1 in maintaining mitochondrial homeostasis in hypoxia/reoxygenation (H/R) H9c2 cells. The results showed that NGR1 pretreatment effectively increased cell survival rates post-H/R, reduced lactate dehydrogenase (LDH) leakage, and mitigated cell damage. Further investigation into mitochondria revealed that NGR1 alleviated mitochondrial structural damage, improved mitochondrial membrane permeability transition pore (mPTP) persistence, and prevented mitochondrial membrane potential (Δψm) depolarization. Additionally, NGR1 pretreatment enhanced ATP levels, increased the activity of mitochondrial respiratory chain complexes I–V after H/R, and reduced excessive mitochondrial reactive oxygen species (mitoROS) production, thereby protecting mitochondrial function. Further analysis indicated that NGR1 upregulated the expression of mitochondrial biogenesis-related proteins (PGC-1α, Nrf1, Nrf2) and mitochondrial fusion proteins (Opa1, Mfn1, Mfn2), while downregulating mitochondrial fission proteins (Fis1, Drp1) and reducing mitochondrial autophagy (mitophagy) levels, as well as the expression of mitophagy-related proteins (Pink1, Parkin, BNIP3) post-H/R. Therefore, this study showed that NGR1 can maintain mitochondrial homeostasis by regulating mitophagy, mitochondrial fission–fusion dynamics, and mitochondrial biogenesis, thereby alleviating H9c2 cell H/R injury and protecting cardiomyocytes.

Hypoxia/reoxygenation (HR) often occurs under cardiac pathological conditions, and HR-induced oxidative stress usually leads to cardiomyocyte damage. Carvedilol, a non-selective β-blocker, is used clinically to treat cardiac ischemia diseases. Moreover, Carvedilol has also been reported to have an antioxidant ability by reducing lipid peroxidation. However, the mechanism of Carvedilol to inhibit lipid peroxidation is still elusive. To explore the protective mechanism of Carvedilol to resist lipid peroxidation on cardiomyocytes, HL-1 cells were cultured under normoxia, hypoxia, and HR and treated with Carvedilol to investigate the alteration on metabolism, protein expression, and mRNA level to explain its oxidative mechanism. The study found that Carvedilol upregulated glutathione peroxidase 4 (GPX4) protein expression to resist HR-induced lipid peroxidation by metabolic remodeling under HR. Also, Carvedilol promoted ferroptosis-related genes, ferritin heavy chain 1 (FTH1) and ferritin light chain 1 (FTL1) mRNA levels, to reduce lipid peroxidation under both hypoxia and HR. In conclusion, our study explores a mechanism by which Carvedilol inhibits ferroptosis by upregulating GPX4, FTH1, and FTL1 levels to downregulate lipid peroxidation under HR. The study provides a potential strategy for using Carvedilol in clinical applications, inspiring further research and development in the area of heart diseases.

BACKGROUND: Ischemic postconditioning is one of the effective ways to reduce ischemia-reperfusion injury and has been more and more widely used in clinical practice in recent years, but its specific molecular mechanism has yet to be studied. OBJECTIVE: To investigate the role and mechanism of piRNA-005854 in the aging cardiomyocytes caused by hypoxic postconditioning. METHODS: In vitro, cardiomyocytes were administered 8 mg/mL D-galactose for 9 days to induce their aging. β-Galactosidase staining was used to observe the aging of cardiomyocytes. Senescent cells were treated with hypoxia/reoxygenation and hypoxic postconditioning. ELISA was utilized to detect changes in myocardial injury markers creatine kinase isoenzyme MB and lactate dehydrogenase levels. Western blot assay was applied to detect the expression changes of autophagy-related proteins LC3II, p62, ULK1 and phosphorylated ULK1 in aging cardiomyocytes. qRT-PCR was employed to determine the expression level of piRNA-005854. piRNA-005854 inhibitor and piRNA-005854 mimics were transferred into aging cardiomyocytes and followed with hypoxic postconditioning. Western blot assay was used to examine the expression of LC3II, p62, ULK1 and phosphorylated ULK1. RESULTS AND CONCLUSION: (1) D-galactose induced obvious senescence of cardiomyocytes 9 days later. (2) Compared with the normoxia group, creatine kinase isoenzyme MB and lactate dehydrogenase levels increased in the hypoxia/reoxygenation group (P < 0.01); LC3 II/I expression was increased; p62 expression was decreased; ULK1 phosphorylation level was increased, and piRNA-005854 expression was increased (P < 0.01). (3) Compared with the hypoxia/ reoxygenation group, creatine kinase isoenzyme MB and lactate dehydrogenase levels significantly reduced in the hypoxic postconditioning group (P < 0.01); LC3 II/I expression significantly decreased (P < 0.05); p62 expression increased (P < 0.01); ULK1 phosphorylation level decreased (P < 0.05), and piRNA-005854 expression decreased (P < 0.01). (4) After transfection of piRNA-005854 inhibitor, LC3II/I expression was decreased (P < 0.01); the expression of p62 was increased significantly (P < 0.05); the phosphorylation level of ULK1 was decreased significantly (P < 0.01). After transfection of piRNA-005854 mimics, LC3II/ I expression was increased significantly; the expression of p62 was decreased, and the phosphorylation level of ULK1 was increased significantly (P < 0.01). (5) The results show that piRNA-005854-mediated reduction of ULK1-dependent autophagy level is a possible mechanism that hypoxic postconditioning exerts its protective effect on aging cardiomyocytes. [ABSTRACT FROM AUTHOR]

Neurodegeneration is linked to the progressive loss of neural function and is associated with several diseases. Hypoxia is a hallmark in many of these diseases, and several therapies have been developed to treat this disease, including gene expression therapies that should be tightly controlled to avoid side effects. Cells experiencing hypoxia undergo a series of physiological responses that are induced by the activation of various transcription factors. Modulation of microRNA (miRNA) expression to alter transcriptional regulation has been demonstrated to be beneficial in treating multiple diseases, and in this study, we therefore explored potential miRNA candidates that could influence hypoxia-induced nerve cell death. Our data suggest that in mouse neuroblasts Neuro-2a cells with hypoxia/reoxygenation (H/R), miR-337-3p is downregulated to increase the expression of Potassium channel tetramerization domain containing 11 (KCTD11) and subsequently promote apoptosis. Here, we demonstrate for the first time that KCTD11 plays a role in the cellular response to hypoxia, and we also provide a possible regulatory mechanism by identifying the axis of miR-337-3p/KCTD11 as a promising candidate modulator of nerve cell survival after H/R exposure. [ABSTRACT FROM AUTHOR]

Background: The purpose of this study was to investigate the regulatory role of G coupled-protein receptor 43 (GPR43) during myocardial ischaemia/reperfusion (I/R) injury and to explore the relevant molecular mechanism. Materials and methods: AC16 hypoxia/reoxygenation (H/R) model was established to simulate I/R injury in vitro. Gain- and loss-of-function experiments were conducted to regulate GPR43 or nesfatin1 expression. Cell viability and apoptosis was examined adopting CCK-8 and TUNEL assays. Commercial kits were applied for detecting ROS and inflammatory cytokines. Quantitative real-time PCR (qRT-PCR) and western blotting were conducted to measure the expression level of critical genes and proteins. Results: GPR43 was downregulated in H/R-mediated AC16 cells. GPR43 over-expression or the GPR43 agonist greatly inhibited H/R-induced cell viability loss, cell apoptosis, and excessive production of ROS and pro-inflammatory cytokines in AC16 cardiomyocytes. Co-immunoprecipitation (Co-IP) assay identified an interaction between GPR43 and nesfatin1, and GPR43 could positively regulate nesfatin1. In addition, the protective role of GPR43 against H/R injury was partly abolished upon nesfatin1 knockdown. Eventually, GPR43 could inhibit H/R-stimulated JNK/P38 MAPK signalling in AC16 cells, which was also hindered by nesfatin1 knockdown. Conclusions: Our findings demonstrated the protective role of GPR43 against H/R-mediated cardiomyocytes injury through up-regulating nesfatin1, providing a novel target for the prevention and treatment of myocardial I/R injury. [ABSTRACT FROM AUTHOR]

Calcium influx into neurons triggers neuronal death during cerebral ischemia/reperfusion injury. Various calcium channels are involved in cerebral ischemia/reperfusion injury. Cav3.2 channel is a main subtype of T-type calcium channels. T-type calcium channel blockers, such as pimozide and mibefradil, have been shown to prevent cerebral ischemia/reperfusion injury-induced brain injury. However, the role of Cav3.2 channels in cerebral ischemia/reperfusion injury remains unclear. Here, in vitro and in vivo models of cerebral ischemia/reperfusion injury were established using middle cerebral artery occlusion in mice and high glucose hypoxia/reoxygenation exposure in primary hippocampal neurons. The results showed that Cav3.2 expression was significantly upregulated in injured hippocampal tissue and primary hippocampal neurons. We further established a Cav3.2 gene-knockout mouse model of cerebral ischemia/reperfusion injury. Cav3.2 knockout markedly reduced infarct volume and brain water content, and alleviated neurological dysfunction after cerebral ischemia/reperfusion injury. Additionally, Cav3.2 knockout attenuated cerebral ischemia/reperfusion injury-induced oxidative stress, inflammatory response, and neuronal apoptosis. In the hippocampus of Cav3.2-knockout mice, calcineurin overexpression offset the beneficial effect of Cav3.2 knockout after cerebral ischemia/reperfusion injury. These findings suggest that the neuroprotective function of Cav3.2 knockout is mediated by calcineurin/nuclear factor of activated T cells 3 signaling. Findings from this study suggest that Cav3.2 could be a promising target for treatment of cerebral ischemia/reperfusion injury.

AbstractContext Although Tongguan capsule (TGC) is used in the treatment of coronary atherosclerotic disease, the exact mechanism remains unclear.Objective Network pharmacology and experimental validation were applied to examine the mechanism of TGC for treating myocardial ischemia-reperfusion injury (MIRI).Materials and methods The components and candidate targets were searched based on various databases such as TCMSP, TCMID, BATMAN-TCM. The binding ability was determined by molecular docking. The ischemia-reperfusion (I/R) model was constructed by ligating the left anterior descending (LAD) coronary artery. APOE-/- mice were divided into three groups (n = 6): Sham group, I/R group, and TGC group (1 g/kg/d). To further verification, HCAEC cells were subjected to hypoxia-reoxygenation (H/R) to establish in vitro model.Results The compounds, such as quercetin, luteolin, tanshinone IIA, kaempferol and bifendate, were obtained after screening. The affinity values of the components with GSK-3β, mTOR, Beclin-1, and LC3 were all

Objective To explore the regulative mechanism of hypoxia/reoxygen (H/R) for invasion ability of extravillous trophoblast (EVT). Methods Immortalized human EVT cell line HTR-8/SVneo in early pregnancy, primary EVT cells isolated from the villi of placenta in early pregnancy, and villi explants were cultured respectively. A simulation model of H/R environment in EVT cells invading the uterus was established under the conversion conditions of 1% O2 for 24 h to 20% O2 for another 24 h. Western blotting and immunofluorescence assay were used to detect the expression of key factors such as cell invasion, and Transwell chamber test was employed to observe cell migration and invasion. Results After H/R induction, the migration and invasion abilities of HTR-8/SVneo cells were weakened, and the epitaxial ability of cultured villi explants and the migration and invasion abilities of isolated primary EVT cells were also weakened. Under H/R conditions, the expression levels of MMP-2 and MMP-9, integrin-β1 and integrin-α5 were decreased in HTR-8/SVneo cells (P < 0.05), and the protein expression of DNA methyltransferase (DNMTs) was also down-regulated (P < 0.01). In the primary EVT cells under H/R condition, the expression levels of epithelial-mesenchymal transition (EMT) positive regulatory factors, Slug and Snails, were inhibited (P < 0.05), that of epithelial cell marker E-cadherin was increased (P < 0.01), protein levels of MMP-2 and MMP-9, integrin-β1 and integrin-α5 were decreased (P < 0.05), and cell invasion ability was reduced along with decreased DNMT1 and DNMT3A expression (P < 0.05). However, overexpression of DNMT1, DNMT3A and DNMT3B in HTR-8/SVneo cells promoted the up-regulation of interstitial markers N-cadherin, and MMP-2 and MMP-9 (P < 0.05). Conclusion H/R regulates the expression of EMT-related factors such as MMP-2/-9 through DNMTs, and thereby reduces the migration and invasion abilities of EVT cells.

Background: Ischemia–reperfusion injury to the central nervous system often causes severe complications. The activation of endogenous neural stem cells (NSCs) is considered a promising therapeutic strategy for nerve repair. However, the specific biological processes and molecular mechanisms of NSC activation remain unclear, and the role of N6-methyladenosine (m6A) methylation modification in this process has not been explored. Methods: NSCs were subjected to hypoxia/reoxygenation (H/R) to simulate ischemia–reperfusion in vivo. m6A RNA methylation quantitative kit was used to measure the total RNA m6A methylation level. Quantitative real-time PCR was used to detect methyltransferase and demethylase mRNA expression levels. Methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA sequencing (RNA-seq) were conducted for NSCs in control and H/R groups, and the sequencing results were analyzed using bioinformatics. Finally, the migration ability of NSCs was identified by wound healing assays, and the proliferative capacity of NSCs was assessed using the cell counting kit-8, EdU assays and cell spheroidization assays. Results: Overall of m6A modification level and Mettl14 mRNA expression increased in NSCs after H/R treatment. The m6A methylation and expression profiles of mRNAs in NSCs after H/R are described for the first time. Through the joint analysis of MeRIP-seq and RNA-seq results, we verified the proliferation of NSCs after H/R, which was regulated by m6A methylation modification. Seven hub genes were identified to play key roles in the regulatory process. Knockdown of Mettl14 significantly inhibited the proliferation of NSCs. In addition, separate analysis of the MeRIP-seq results suggested that m6A methylation regulates cell migration and differentiation in ways other than affecting mRNA expression. Subsequent experiments confirmed the migration ability of NSCs was suppressed by knockdown of Mettl14. Conclusion: The biological behaviors of NSCs after H/R are closely related to m6A methylation of mRNAs, and Mettl14 was confirmed to be involved in cell proliferation and migration. [ABSTRACT FROM AUTHOR]

Hypoxia (20 min) and reoxygenation (30 min) were simulated on isolated rat cardiomyocytes to evaluate the cytoprotective effect of selective δ2-opioid receptor agonist deltorphin II, opioid receptor antagonist naloxone methiodide, μ-opioid receptor antagonist CTAP, κ-opioid receptor antagonist nor-binaltorphimine, ε1-opioid receptor antagonist BNTX, and δ2-opioid receptors naltriben. Deltorphin II was administered 5 min before reoxygenation, antagonists were administered 10 min before reoxygenation. The cytoprotective effect of deltorphin II was assessed by the number of cardiomyocytes survived after hypoxia/reoxygenation, as well as by the lactate dehydrogenase content in the incubation medium. It has been established that the cytoprotective effect of deltorphin II occurs at a concentration of 64 nmol/liter and is associated with activation of δ2-opioid receptors. [ABSTRACT FROM AUTHOR]

AIM: To investigate the regulatory role of retinoid X receptor （RXR） in oxidative stress response of rat type II alveolar epithelial cells（ AECII） induced by hypoxia/reoxygenation（ HR）. METHODS: The AECII were divided into control （C） group, HR group, HR+solvent dimethyl sulfoxide （DMSO） group （HD group）, HR+RXR agonist 9-cis-retinoic acid （9-RA） group （RA group）, and HR+RXR antagonist HX531 group （HX group）. Cell Counting Kit-8 （CCK-8） method was used to measure the cell viability. Immunofluorescence staining was used to detect the expression of surfactant protein A （SP-A） and RXRα in AECII. Kits were detected to the levels of superoxide dismutase （SOD） and malondialdehyde （MDA） in cells. Transmission electron microscopy was used to observe the ultrastructural changes of the cells. Western blot was used to detect the protein level of nuclear factor E2-related factor 2 （Nrf2）. RT-PCR was used to detect the expression level of Nrf2 mRNA. RESULTS: Compared with C group, the cell viability and SOD activity in HR, HD, RA and HX groups were decreased significantly （P<0. 05）, the MDA content were increased significantly （P<0. 05）, the Nrf2 mRNA and protein expression levels were decreased significantly （P<0. 05 or P<0. 01）, and the immunofluorescence expression of RXRα was significantly increased （P<0. 01）. Compared with HR and HX groups, the cells in RA group showed significantly increased cell viability （P<0. 05）, increased SOD activity（ P<0. 05）, decreased MDA content （P<0. 05）, increased Nrf2 mRNA and protein expression levels （P<0. 01）, and significantly increased immunofluorescence expression of RXRα （P<0. 01）. CONCLUSION: Hypoxia/reoxygenation can aggravate the oxidative stress response of rat AECII, and RXR agonist intervention can alleviate HR-induced rat AECII injury by inhibiting oxidative stress. [ABSTRACT FROM AUTHOR]

To explore the effect and possible mechanism of Allium mongolicum Regel flavonoids on cardiomyocyte injury induced by hypoxia/reoxygenation. H9C2 cardiomyocytes were induced by hypoxia/reoxygenation and treated with different doses of Allium mongolicum Regel flavonoids. H9C2 cells were transfected with si-negative control/ si-TALNEC2 and treated with hypoxia/reoxygenation. Besides, cells transfected with plasmid cloning deoxyribonucleic acid/plasmid cloning deoxyribonucleic acid-TALNEC2 were treated with Allium mongolicum Regel flavonoids and induced with hypoxia/reoxygenation. Malondialdehyde, glutathione peroxidase and superoxide dismutase levels were tested to evaluate oxidative stress. Apoptosis rate was analyzed by flow cytometry. TALNEC2 expression was examined using quantitative reverse transcription-polymerase chain reaction, and cleaved caspase-3 and cleaved caspase-9 protein levels were tested by Western blot. Allium mongolicum Regel flavonoids could reduce malondialdehyde level, apoptosis rate, cleaved caspase-3 level, cleaved caspase-9 level, and TALNEC2 expression, while enhanced glutathione peroxidase and superoxide dismutase levels in hypoxia/reoxygenation-induced H9C2 cells in a dose-dependent manner. After transfection of si-TALNEC2, malondialdehyde level, apoptosis rate, cleaved caspase-3 level, and cleaved caspase-9 level were reduced, while superoxide dismutase and glutathione peroxidase levels were enhanced. Transfection of plasmid cloning deoxyribonucleic acid-TALNEC2 could abolish the effect of Allium mongolicum Regel flavonoids on cardiomyocyte injury. Allium mongolicum Regel flavonoids could inhibit hypoxia/reoxygenation-induced cardiomyocyte apoptosis and oxidative stress via reducing TALNEC2 expression. [ABSTRACT FROM AUTHOR]

Objective: This study aimed to investigate the function and mechanism of lncRNA NORAD in Sevoflurane (Sev) protection against myocardial hypoxia-reoxygenation (H/R). Methods: Preprocess rat cardiomyocytes H9c2 cells with Sev at concentrations of 0.5%, 1.0%, and 1.5%, and subjected them to H/R treatment. qRT-PCR was used to detect levels of NORAD and miR-144-3p. Measure concentrations of the inflammatory cytokines IL-6, TNF-α, and IL-10, as well as cardiac injury markers cTnI, CK-MB, and LDH using ELISA. Assess cell proliferation and apoptosis using CCK-8 and flow cytometry. Perform dual-luciferase reporter assay and RIP assay to validate the targeting relationship between NORAD and miR-144-3p. Results: H/R induced inhibition of cell proliferation, increase in apoptosis, and production of IL-6, TNF-α, CK-MB, LDH, and cTnI were significantly attenuated by Sev. As hypoxic treatment time lengthened, the NORAD levels in myocardial cells showed an increase, with Sev pretreatment being able to suppress the NORAD levels elevation. The overexpression of NORAD notably weakened the cardioprotective effect of Sev. NORAD targetedly binds to miR-144-3p and negatively regulates miR-144-3p. Increased miR-144-3p levels inhibited the antagonistic effect of NORAD on the cardioprotective effects of Sev. Conclusion: The current study confirmed that sevoflurane attenuated H/R-induced cardiomyocyte injury via the NORAD/miR-144-3p axis. [ABSTRACT FROM AUTHOR]

Objective: To study the effect of Long non-coding RNA (LncRNA) small nucleolar RNA host gene 12 (SNHG12) regulating miR-138-5p/hypoxia inducible factor-1 (HIF-1α) axis on improving the damage of hypoxia/reoxygenation (H/R) human vascular endothelial cells. Methods: Human umbilical vein endothelial cells (HUVECs) were cultured in vitro and randomly divided into control group, H/R model group, H/R+LncRNA SNHG12 overexpression group, H/R+miR-138-5p mimics group, H/R+co-transfection group and H/R+co-transfection negative control group, each transfection group was transfected separately, and except for control group, the remaining groups were given hypoxia for 5 hours and then reoxygenated for 1 hour to induce the cell models, and then the cell viability of each group was detected by CCK-8 experiment; the cell apoptosis in each group was detected by flow cytometry experiment, and the apoptosis rate of each group was compared; the levels of reactive oxygen species (ROS), lactate dehydrogenase (LDH) and inflammatory factors IL-6, IL-17 and IL-18 in each group were measured by the kit; the expressions of miR-138-5p and HIF-1α mRNA in cells of each group were measured by real-time quantitative PCR (qRT-PCR) experiment; the expressions of apoptotic proteins caspase-9, Bcl-2-associated X protein (Bax) and HIF-1α in each group were evaluated by Western blot. Results: Compared with control group, the apoptosis rate, cellular ROS, LDH, IL-6, IL-17 and IL-18 levels, cellular HIF-1α mRNA and protein levels, cellular caspase-9, Bax and HIF-1α protein levels were increased in H/R model group (P<0.05), the cell viability and miR-138-5p level were decreased (P<0.05). Compared with H/R model group and H/R+co-transfection group, the cell viability, cell HIF-1α mRNA and protein levels were increased in H/R+LncRNA SNHG12 overexpression group (P<0.05), the apoptosis rate，cellular ROS, LDH, IL-6, IL-17 and IL-18 levels，cellular caspase-9 and Bax protein levels, and miR-138-5p level were decreased (P<0.05); the cell viability，cellular HIF-1α mRNA and protein levels were decreased in H/R+miR-138-5p mimics group (P<0.05), the apoptosis rate, cellular ROS, LDH, IL-6, IL-17 and IL-18 levels，cellular caspase-9 and Bax protein levels were increased (P<0.05). Compared with H/R model group, there was no significant difference in cell index levels between the H/R+co-transfection negative control group and the H/R+co-transfection group (P>0.05). Conclusion: LncRNA SNHG12 can upregulate HIF-1α expression by downregulating miR-138-5p expression, inhibit H/R-induced inflammation and oxidative stress in HUVECs, and reduce cell damage and apoptosis. [ABSTRACT FROM AUTHOR]

Background: Mitsugumin 53 (MG53) is a membrane repair factor that is associated with acute myocardial infarction. This study aimed to investigate the effects of MG53 on cardiomyocyte injury and the posttranslational modification of MG53. Methods: Cardiomyocyte injury was evaluated by enzyme-linked immunosorbent assay and flow cytometry. The succinylation and ubiquitination levels of MG53 were examined by immunoprecipitation (IP) and western blot. The relationship between MG53 and KAT3B or SIRT7 was assessed by co-IP and immunofluorescence. Results: The results showed that overexpression of MG53 inhibited inflammation response and apoptosis of cardiomyocytes induced by hypoxia/reoxygenation (H/R). Succinylation and protein levels of MG53 were downregulated in H/R-induced cells, which was inhibited by SIRT7 and promoted by KAT3B. SIRT7 aggravated and KAT3B alleviated MG53-mediated cardiomyocyte injury. Moreover, MG53 was succinylated and ubiquitinated at K130. Conclusion: SIRT7 inhibited/KAT3B promoted succinylation of MG53 at K130 sites, which suppressed ubiquitination of MG53 and upregulated its protein levels, thereby alleviating H/R-induced cardiomyocyte injury. The findings suggested that MG53 may be a potential therapy for myocardial infarction. [ABSTRACT FROM AUTHOR]

Based on the inhibition of mitochondrial permeability transition pore (mPTP) opening, puerarin (PUE) has a good potential to reduce myocardial ischemia/reperfusion injury (MI/RI). However, the lack of targeting of free PUE makes it difficult to reach the mitochondria. In this paper, we constructed matrix metalloproteinase-targeting peptide (MMP-TP) and triphenylphosphonium (TPP) cation co-modified liposomes loaded with PUE (PUE@T/M-L) for mitochondria-targeted drug delivery. PUE@T/M-L had a favorable particle size of 144.9 ± 0.8 nm, an encapsulation efficiency of 78.9 ± 0.6%, and a sustainedrelease behavior. The results of cytofluorimetric experiments showed that MMP-TP and TPP doublemodified liposomes (T/M-L) enhanced intracellular uptake, escaped lysosomal capture, and promoted drug targeting into mitochondria. In addition, PUE@T/M-L enhanced the viability of hypoxia-reoxygenation (H/R) injured H9c2 cells by inhibiting mPTP opening and reactive oxygen species (ROS) production, reducing Bax expression and increasing Bcl-2 expression. It was inferred that PUE@T/M-L delivered PUE into the mitochondria of H/R injured H9c2 cells, resulting in a significant increase in cellular potency. Based on the ability of MMP-TP to bind the elevated expression of matrix metalloproteinases (MMPs), T/M-L had excellent tropism for Lipopolysaccharide (LPS) -stimulated macrophages and can significantly reduce TNF-a and ROS levels, thus allowing both drug accumulation in ischemic cardiomyocytes and reducing inflammatory stimulation during MI/RI. Fluorescence imaging results of the targeting effect using a DiR probe also indicated that DiR@T/M-L could accumulate and retain in the ischemic myocardium. Taken together, these results demonstrated the promising application of PUE@T/M-L for mitochondria-targeted drug delivery to achieve maximum therapeutic efficacy of PUE. [ABSTRACT FROM AUTHOR]

Although Tongguan capsule (TGC) is used in the treatment of coronary atherosclerotic disease, the exact mechanism remains unclear. Network pharmacology and experimental validation were applied to examine the mechanism of TGC for treating myocardial ischemia-reperfusion injury (MIRI). The components and candidate targets were searched based on various databases such as TCMSP, TCMID, BATMAN-TCM. The binding ability was determined by molecular docking. The ischemia-reperfusion (I/R) model was constructed by ligating the left anterior descending (LAD) coronary artery. APOE-/- mice were divided into three groups (n = 6): Sham group, I/R group, and TGC group (1 g/kg/d). To further verification, HCAEC cells were subjected to hypoxia-reoxygenation (H/R) to establish in vitro model. The compounds, such as quercetin, luteolin, tanshinone IIA, kaempferol and bifendate, were obtained after screening. The affinity values of the components with GSK-3β, mTOR, Beclin-1, and LC3 were all <-5 kcal/mol. In vivo, TGC improved LVEF and FS, reducing infarct size. In vitro, Hoechst 33258 staining result showed TGC inhibited apoptosis. Compare with the H/R model, TGC treatment increased the levels of GSK-3β, LC3, and Beclin1, while decreasing the expression of mTOR and p62 (p < 0.05). The findings revealed that TGC exerted a cardioprotective effect by up regulating autophagy-related proteins through the mTOR pathway, which may be a therapeutic option for MIRI. However, there are still some limitations in this research. It is necessary to search more databases to obtain information and further demonstrated through randomized controlled trials for generalization. [ABSTRACT FROM AUTHOR]

Objective To study the protective effect and related mechanisms of short-chain fatty acid sodium acetate on hypoxia/reoxygenation (H/R)-induced injury of human renal tubular epithelial cell line HK2. Methods The H/R model was established, and HK2 cells were incubated with sodium acetate (SA) for 2 h. The survival rate of HK2 cells was detected by CCK8 assay; Enzyme activity kit was used for detection of inflammatory factors, cellular reactive oxygen species and ATP production; Flow cytometry was used to measure mitochondrial membrane potential (MMP) and mitochondrial oxidative stress products (mitoSOX) levels; Electron microscope was used to observe mitochondrial ultrastructural damage; Western blot was used to detected the expression of inflammatory signaling pathway IκB-α/NF-κB and mitochondrial energy disorder signaling pathway AMPK/PGC-1α. Results Compared with the control group, the survival rate of HK2 cells in the H/R group was significantly decreased(P＜0.05); the expression of intracellular ROS, mitoSOX and inflammatory factors was significantly increased(P＜0.05); the ultrastructure of mitochondria was severely damaged, and the content of ATP and MMP was significantly decreased(P＜0.05); it was further found that the protein expression of p-IκB-α and NF-κB-p-P65 was significantly increased while the protein expression of p-AMPK and PGC-1α was significantly decreased(P＜0.05). Compared with H/R group, SA significantly enhanced the survival rate of HK2 cells (P＜0.05); SA inhibited the release of intracellular ROS, mitoSOX and inflammatory factors; SA inhibited mitochondrial ultrastructural damage, decreased ATP and MMP (P＜0.05); SA promoted the expression of p-AMPK and PGC-1α and inhibited the expression of p-IκB-α and NF-κB-p-P65 (P＜0.05). Conclusions Sodium acetate plays a protective role with potential mechanisms of anti-inflammation, anti-oxidative stress, protection of mitochondrial structure and function of HK2 cells induced by H/R through inhibiting IκB-α/ NF-κB and activating AMPK/PGC-1α signal pathway.

Background: Myocardial infarction is one of the leading causes of mortality worldwide; hence, there is an urgent need to discover novel cardioprotective strategies. Kynurenic acid (KYNA), a metabolite of the kynurenine pathway, has been previously reported to have cardioprotective effects. However, the mechanisms by which KYNA may be protective are still unclear. The current study addressed this issue by investigating KYNA's cardioprotective effect in the context of myocardial ischemia/reperfusion. Methods: H9C2 cells and rats were exposed to hypoxia/reoxygenation or myocardial infarction, respectively, in the presence or absence of KYNA. In vitro, cell death was quantified using flow cytometry analysis of propidium iodide staining. In vivo, TTC-Evans Blue staining was performed to evaluate infarct size. Mitochondrial respiratory chain complex activities were measured using spectrophotometry. Protein expression was evaluated by Western blot, and mRNA levels by RT-qPCR. Results: KYNA treatment significantly reduced H9C2-relative cell death as well as infarct size. KYNA did not exhibit any effect on the mitochondrial respiratory chain complex activity. SOD2 mRNA levels were increased by KYNA. A decrease in p62 protein levels together with a trend of increase in PARK2 may mark a stimulation of mitophagy. Additionally, ERK1/2, Akt, and FOXO3α phosphorylation levels were significantly reduced after the KYNA treatment. Altogether, KYNA significantly reduced myocardial ischemia/reperfusion injuries in both in vitro and in vivo models. Conclusion: Here we show that KYNA-mediated cardioprotection was associated with enhanced mitophagy and antioxidant defense. A deeper understanding of KYNA's cardioprotective mechanisms is necessary to identify promising novel therapeutic targets and their translation into the clinical arena. [ABSTRACT FROM AUTHOR]

Background: Anisodine hydrobromide (AT3), an anti-cholinergic agent, could be delivered to the brain across the blood-brain barrier and has been used clinically for the treatment of cerebral ischemia/reperfusion injury. Endothelial dysfunction can be caused by hypoxia/reoxygenation (H/R) via oxidative stress and metabolic alterations. The present study investigated whether AT3 regulates the production of nitric oxide (NO) and reactive oxygen species (ROS), and the HIF-1α pathway via regulation of muscarinic acetylcholine receptors (mAChRs) in brain microvascular endothelial cells after H/R exposure. Methods: Under H/R conditions, hCMEC/D3 cerebral microvascular endothelial cells were treated with AT3. Specific inhibitors of M2- and M4- mAChRs were used to explore the mechanism by which AT3 influences oxidative stress in endothelial cells. Then, mAChRs expression was detected by western blotting and NO production was detected by Greiss reaction. The intracellular ROS level was measured using DCFH-DA probes. The expression of hypoxia-inducible transcription factor 1α (HIF-1α) was also detected. Results: While H/R induced the expression of M2- and M4-mAChRs, AT3 suppressed the H/R-upregulated M2- and M4-mAChRs. H/R also induced the production of NO, ROS, and apoptosis. AT3 and M4-mAChR inhibitors inhibited the H/R-induced production of NO and ROS and apoptosis. HIF-1α was induced by H/R, but was suppressed by AT3. Conclusion: Thus, the in vitro evidence shows that AT3 protects against H/R injury in cerebral microvascular endothelial cells via inhibition of HIF-1α, NO and ROS, predominantly through the downregulation of M4-mAChR. The findings offer novel understandings regarding AT3-mediated attenuation of endothelial cell apoptosis and cerebral ischemia/ reperfusion injury. [ABSTRACT FROM AUTHOR]

Acute myocardial infarction is regarded as myocardial necrosis resulting from myocardial ischemia/reperfusion (I/R) damage and retains a major cause of mortality. Neferine, which was extracted from the green embryos of mature seeds of Nelumbo nucifera Gaertn., has been reported to possess a broad range of biological activities. However, its underlying mechanism on the protective effect of I/R has not been fully clarified. A hypoxia/reoxygenation (H/R) model with H9c2 cells closely simulating myocardial I/R injury was used as a cellular model. This study intended to research the effects and mechanism underlying neferine on H9c2 cells in response to H/R stimulation. Cell Counting Kit‐8 and lactate dehydrogenase (LDH) release assays were employed to measure cell viability and LDH, respectively. Apoptosis and reactive oxygen species (ROS) were determined by flow cytometry analysis. Oxidative stress was evaluated by detecting malondialdehyde, superoxide dismutase, and catalase. Mitochondrial function was assessed by mitochondrial membrane potential, ATP content, and mitochondrial ROS. Western blot analysis was performed to examine the expression of related proteins. The results showed that hypoxia/reoxygenation (H/R)‐induced cell damage, all of which were distinctly reversed by neferine. Moreover, we observed that neferine inhibited oxidative stress and mitochondrial dysfunction induced by H/R in H9c2 that were concomitant with increased sirtuin‐1 (SITR1), nuclear factor erythroid 2‐related factor 2 (Nrf2), and heme oxygenase‐1 expression. On the contrary, silencing the SIRT1 gene with its small interferingRNA eliminated the beneficial effects of neferine. It is concluded that neferine preconditioning attenuated H/R‐induced cardiac damage via suppressing apoptosis, oxidative stress, and mitochondrial dysfunction, which may be partially ascribed to the activation of SIRT1/Nrf2 signaling pathway. [ABSTRACT FROM AUTHOR]

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Thiopental sodium (TPTS) is a barbiturate general anesthetic, while its effects on hypoxia/reoxygenation (H/R)-induced injury are still unclear. This study aimed to investigate whether TPTS exerts protective effects against the H/R-induced osteoblast cell injury and explore the underlying mechanisms. Osteoblast cell injury model was induced by the H/R condition, which was treated with or without TPTS. Cell viability and lactate dehydrogenase (LDH) release were determined by the corresponding commercial kits. The levels of oxidative stress were determined in the experimental groups. Cell apoptosis and Caspase-3 activities were determined by propidium iodide staining and substrate-based assay, respectively. Western blotting and qRT-PCR were performed to measure the mRNA and protein levels, respectively. Treatment with TPTS was able to increase cell viability and reduce LDH release in H/R-induced osteoblasts. Additionally, TPTS regulated oxidative stress in H/R-induced osteoblasts by suppressing malondialdehyde (MDA) and reactive oxygen species (ROS) as well as boosting superoxide dismutase (SOD). TPTS was able to suppress cell apoptosis by suppressing Caspase-3 activity and cleavage. TPTS exerted protective effects against cell injury and apoptosis induced by the H/R conditions, which were associated with its regulation of Akt signaling. Moreover, TPTS induced osteoblast differentiation under the H/R condition. In summary, TPTS attenuates H/R-induced injury in osteoblasts by regulating AKT signaling. [ABSTRACT FROM AUTHOR]

Objective: To discuss the effect of down-regulation of miR-320a expression on the myocardial hypoxia/reoxygenation (H/R)injury model, and to clarify its related mechanism. Methods: Real-time fluorescence quantitative PCR (RT-qPCR)method was used to detect the expression levels of miR-320a in serum of the patients with actue myocardial infarction (AMI)and the myocardial H9C2 cells induced by H/R. The miR-320a inhibitor, inhibitor NC, small interference Janus kinase 2 (si-JAK2), and si-NC plasmids were transfected into the H9C2 cells respectively, and blank control group was set up. After successful transfection, the H/R treatment was performed. The H9C2 cells were divided into control group, H/R group, H/R + inhibitor NC group, H/R + miR-320a inhibitor group, H/R + miR-320a inhibitor + si-NC group and H/R + miR-320a inhibitor + si-JAK2 group. The targeting relationship between miR-320a and Janus kinase 2 (JAK2)was detected by double luciferase reporter gene; the proliferation rate of cells in various groups were detected by CCK-8 assay; the activities of superoxide dismutase (SOD)and levels of malonaldehyde (MDA)in cells and the levels of lactate dehydrogenase (LDH)in cell culture supernanant in various groups were detected by biochemical method; the apoptotic rates of cells in various groups were detected by flow cytometry; the expression levels of miR-320a and JAK2 mRNA in cells in various groups were detected by RT-qPCR method; the expression levels of B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax), cleaved-cysteinyl aspartate specific proteinase-3 (cleaved-caspase-3), JAK2, signal transducers and activator of transcription 3 (STAT3), and phosphorylated STAT3 (p-STAT3)proteins in cells in various groups were detected by Western blotting method. Results: The expression levels of miR-320a in serum of the patients with AMI and the myocardial H9C2 cells in H/R group were significantly higher than those in control group (P<0. 05). The results of double Luciferase reporter gene detection suggested that miR-320a could targetedly bind with JAK2. Compared with control group, the proliferation rate of the cells and SOD activity in the cells in H/R group were decreased significantly (P<0. 05), the apoptotic rate of the cells, MDA level and LDH activity in the cells were significantly increased (P<0. 05), the expression levels of Bcl-2 and JAK2 proteins and ratio of p-STAT3/STAT3 in the cells were significantly decreased (P<0. 05), and the expression levels of Bax and cleaved caspase-3 proteins in the cells were significantly increased (P<0. 05). Compared with H/R group, the proliferation rate of the cells and SOD activity in the cells in H/R+miR-320a inhibitor group were increased (P<0. 05), while the apoptotic rate of the cells, MDA level in the cells, LDH activity in the cell culture supernanant were decreased (P<0. 05), the expression levels of Bcl-2 and JAK2 proteins and ratio of p-STAT3/STAT3 in the cells were significantly increased (P<0. 05), the expression levels of Bax and cleaved- caspase-3 proteins in the cells were significantly decreased (P<0. 05). Compared with H/R+miR-320a inhibitor+ si-NC group, the proliferation rate of the cells and SOD activity in the cells in H/R+miR-320a inhibitor+ si-JAK2 group were decreased (P<0. 05), and the apoptotic rate of the cells, MDA level in the cells, and LDH activity in the cell culture supernanant were increased (P<0. 05). Conclusion: Down-regulation of miR-320a expression can inhibit the apoptosis of the cardiomyocytes induced by H/R and increase the proliferation activity of cells, and its mechanism is related to the targeted regulation of JAK2/STAT3 signaling pathway. [ABSTRACT FROM AUTHOR]

RhoGDIα is an inhibitor of RhoGDP dissociation that involves in Aβ metabolism and NFTs production in Alzheimer's disease (AD) by regulating of RhoGTP enzyme activity. Our previous research revealed that RhoGDIα, as the target of Polygala saponin (Sen), might alleviate apoptosis of the nerve cells caused by hypoxia/reoxygenation (H/R). To further clarify the role of RhoGDIα in the generation of NFTs, we explored the relationship between RhoGDIα and Tau. We found out that RhoGDIα and Tau can bind with each other and interact by using coimmunoprecipitation (Co-IP) and GST pulldown methods in vitro. This RhoGDIα-Tau partnership was further verified by using immunofluorescence colocalization and fluorescence resonance energy transfer (FRET) approaches in PC12 cells. Using the RNA interference (RNAi) technique, we found that the RhoGDIα may be involved in an upstream signaling pathway for Tau. Subsequently, in Aβ25-35- and H/R-induced PC12 cells, forced expression of RhoGDIα via cDNA plasmid transfection was found to reduce the hyperphosphorylation of Tau, augment the expression of bcl-2 protein, and inhibit the expression of Bax protein (reducing the Bax/bcl-2 ratio) and the activity of caspase-3. In mouse AD and VaD models, forced expression of RhoGDIα via injection of a viral vector (pAAV-EGFP-RhoGDIα) into the lateral ventricle of the brain alleviated the pathological symptoms of AD and VaD. Finally, GST pulldown confirmed that the binding sites on RhoGDIα for Tau were located in the range of the ΔC33 fragment (aa 1–33). These results indicate that RhoGDIα is involved in the phosphorylation of Tau and apoptosis in AD and VaD. Overexpression of RhoGDIα can inhibit the generation of NFTs and delay the progress of these two types of dementia. [ABSTRACT FROM AUTHOR]

Hypoxia-inducible factor 1α (HIF-1α) plays a critical role in the apoptotic process during cardiac ischaemia/reperfusion (I/R) injury. This study aimed to investigate whether post-treatment with dexmedetomidine (DEX) could protect against I/R-induced cardiac apoptosis in vivo and in vitro via regulating HIF-1α signalling pathway. Rat myocardial I/R was induced by occluding the left anterior descending artery for 30 minutes followed by 6-hours reperfusion, and cardiomyocyte hypoxia/reoxygenation (H/R) was induced by oxygen-glucose deprivation for 6 hours followed by 3-hours reoxygenation. Dexmedetomidine administration at the beginning of reperfusion or reoxygenation attenuated I/R-induced myocardial injury or H/R-induced cell death, alleviated mitochondrial dysfunction, reduced the number of apoptotic cardiomyocytes, inhibited the activation of HIF-1α and modulated the expressions of apoptosis-related proteins including BCL-2, BAX, BNIP3, cleaved caspase-3 and cleaved PARP. Conversely, the HIF-1α prolyl hydroxylase-2 inhibitor IOX2 partly blocked DEX-mediated cardioprotection both in vivo and in vitro. Mechanistically, DEX down-regulated HIF-1α expression at the post-transcriptional level and inhibited the transcriptional activation of the target gene BNIP3. Post-treatment with DEX protects against cardiac I/R injury in vivo and H/R injury in vitro. These effects are, at least in part, mediated via the inhibition of cell apoptosis by targeting HIF-1α signalling.

Abstract Background Our previous data demonstrated that miR-19b expression was increased in human lung microvascular endothelial cells in-vitro-, in-vivo and in patients with hemorrhagic shock, leading to a decrease in syndecan-1 mRNA and protein and resulting in loss of endothelial barrier function. However, the mechanism underlying increased miR-19b expression remains unclear. The objective of the current study was to determine if c-Jun mediates the early responsive microRNA, miR-19b, to cause endothelial barrier dysfunction. Method Human lung microvascular endothelial cells (HLMEC) or HEK293T cells were transfected with c-Jun overexpressing vector, c-Jun siRNA, miR-19b promoter vector, miR-19b mutated promoter vector, miR-19b oligo inhibitor, then subjected to hypoxia/reoxygenation as in-vitro model of hemorrhagic shock. Levels of protein, miRNA, and luciferase activity were measured. Transwell permeability of endothelial monolayers were also determined. Plasma levels of c-Jun were measured in injured patients with hemorrhagic shock. Result Hypoxia/reoxygenation induced primary (pri-)miR-19b, mature miR-19b, and c-Jun expression over time in a comparable timeframe. c-Jun silencing by transfection with its specific siRNA reduced both pri-miR-19b and mature miR-19b levels. Conversely, c-Jun overexpression enhanced H/R-induced pri-miR-19b. Studies using a luciferase reporter assay revealed that in cells transfected with vectors containing the wild-type miR-19b promoter and luciferase reporter, c-Jun overexpression or hypoxia/ reoxygenation significantly increased luciferase activity. c-Jun knockdown reduced the luciferase activity in these cells, suggesting that the miR-19b promoter is directly activated by c-Jun. Further, chromatin immunoprecipitation assay confirmed that c-Jun directly bound to the promoter DNA of miR-19b and hypoxia/reoxygenation significantly increased this interaction. Additionally, c-Jun silencing prevented cell surface syndecan-1 loss and endothelial barrier dysfunction in HLMECs after hypoxia/reoxygenation. Lastly, c-Jun was significantly elevated in patients with hemorrhagic shock compared to healthy controls. Conclusion Transcription factor c-Jun is inducible by hypoxia/reoxygenation, binds to and activates the miR-19b promoter. Using an in-vitro model of hemorrhagic shock, our findings identified a novel cellular mechanism whereby hypoxia/ reoxygenation increases miR-19b transcription by inducing c-Jun and leads to syndecan-1 decrease and endothelial cell barrier dysfunction. This finding supports that miR-19b could be a potential therapeutic target for hemorrhage shock.