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To investigate the therapeutic potential and underlying mechanisms of exogenous nicotinamide adenine dinucleotide+ on postresuscitation myocardial and neurologic dysfunction in a rat model of cardiac arrest.Thirty-eight rats were randomized into three groups: 1) Sham, 2) Control, and 3) NAD. Except for the sham group, untreated ventricular fibrillation for 6 minutes followed by cardiopulmonary resuscitation was performed in the control and NAD groups. Nicotinamide adenine dinucleotide+ (20 mg/kg) was IV administered at the onset of return of spontaneous circulation.University-affiliated research laboratory.Sprague-Dawley rats.Nicotinamide adenine dinucleotide+.Hemodynamic and myocardial function were measured at baseline and within 4 hours following return of spontaneous circulation. Survival analysis and Neurologic Deficit Score were performed up to 72 hours after return of spontaneous circulation. Adenosine triphosphate (adenosine triphosphate) level was measured in both brain and heart tissue. Mitochondrial respiratory chain function, acetylation level, and expression of Sirtuin3 and NADH dehydrogenase (ubiquinone) 1 alpha subcomplex, 9 (NDUFA9) in isolated mitochondrial protein from both brain and heart tissue were evaluated at 4 hours following return of spontaneous circulation. The results demonstrated that nicotinamide adenine dinucleotide+ treatment improved mean arterial pressure (at 1 hr following return of spontaneous circulation, 94.69 ± 4.25 mm Hg vs 89.57 ± 7.71 mm Hg; p0.05), ejection fraction (at 1 hr following return of spontaneous circulation, 62.67% ± 6.71% vs 52.96% ± 9.37%; p0.05), Neurologic Deficit Score (at 24 hr following return of spontaneous circulation, 449.50 ± 82.58 vs 339.50 ± 90.66; p0.05), and survival rate compared with that of the control group. The adenosine triphosphate level and complex I respiratory were significantly restored in the NAD group compared with those of the control group. In addition, nicotinamide adenine dinucleotide+ treatment activated the Sirtuin3 pathway, down-regulating acetylated-NDUFA9 in the isolated mitochondria protein.Exogenous nicotinamide adenine dinucleotide+ treatment attenuated postresuscitation myocardial and neurologic dysfunction. The responsible mechanisms may involve the preservation of mitochondrial complex I respiratory capacity and adenosine triphosphate production, which involves the Sirtuin3-NDUFA9 deacetylation.

Acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) is a severe syndrome lacking efficient therapy and resulting in high morbidity and mortality. Although resveratrol (RES), a natural phytoalexin, has been reported to protect the ALI by suppressing the inflammatory response, the detailed mechanism of how RES affected the immune system is poorly studied. Pulmonary conventional dendritic cells (cDCs) are critically involved in the pathogenesis of inflammatory lung diseases including ALI. In this study, we aimed to investigate the protective role of RES via pulmonary cDCs in lipopolysaccharide (LPS)-induced ALI mice. Murine ALI model was established by intratracheally challenging with 5 mg/kg LPS. We found that RES pretreatment could mitigate LPS-induced ALI. Additionally, proinflammatory-skewed cytokines decreased whereas anti-inflammatory-related cytokines increased in bronchoalveolar lavage fluid by RES pretreatment. Mechanistically, RES regulated pulmonary cDCs’ maturation and function, exhibiting lower level of CD80, CD86, major histocompatibility complex (MHC) II expression, and IL-10 secretion in ALI mice. Furthermore, RES modulated the balance between proinflammation and anti-inflammation of cDCs. Moreover, in vitro RES pretreatment regulated the maturation and function of bone marrow derived dendritic cells (BMDCs). Finally, the adoptive transfer of RES-pretreated BMDCs enhanced recovery of ALI. Thus, these data might further extend our understanding of a protective role of RES in regulating pulmonary cDCs against ALI.

Blocking ferroptosis reduces ischemia-reperfusion injury in some pathological contexts. However, there is no evidence that ferroptosis contributes to post-resuscitation myocardial dysfunction (PRMD). Here, we evaluated the therapeutic performance of ferroptosis inhibitors (UAMC-3203 or/and Deferoxamine) on the PRMD in a rat model of cardiac arrest and surveyed the changes of essential ferroptosis markers in the myocardium. Remarkably, all treatments reduce the severity of cardiac dysfunction and microcirculation hypoperfusion after resuscitation compared with control. Consistently, we observe that the ferroptosis marker Glutathione peroxidase 4, 4-hydroxynonenal and non-heme iron altered (1 ± 0.060 vs. 0.021 ± 0.016, 1 ± 0.145 vs. 3.338 ± 0.221, 52.010 ± 3.587 ug/g vs. 70.500 ± 3.158 ug/g, all P 0.05) in the myocardium after resuscitation. These changes were significantly suppressed by UAMC-3203 [(0.187 ± 0.043, 2.848 ± 0.169, all P 0.05), (72.43 ± 4.920 ug/g, P 0.05)], or Deferoxamine (0.203 ± 0.025, 2.683 ± 0.273, 55.95 ± 2.497 ug/g, all P 0.05). Briefly, UAMC-3203 or/and Deferoxamine improve post-resuscitation myocardial dysfunction and provide evidence of ferroptosis involvement, suggesting that ferroptosis inhibitors could potentially provide an innovative therapeutic approach for mitigating the myocardial damage caused by cardiopulmonary resuscitation.

Background To investigate the therapeutic potential of combined therapy with polyethylene glycol‐20k (PEG‐20k) and MCC950 on post‐resuscitation myocardial function in a rat model of cardiac arrest. Methods and Results Thirty rats were randomized into 5 groups: Sham, Control, PEG‐20k, MCC950, PEG‐20k+ MCC950. Except for sham, animals were subjected to 6 minutes of ventricular fibrillation followed by 8 minutes cardiopulmonary resuscitation. Two milliliters PEG‐20k was administered by intravenous injection coincident with the start of cardiopulmonary resuscitation; MCC950 (10 mg/kg), a highly selective NLRP3 inflammasome inhibitor, was delivered immediately after restoration of spontaneous circulation. Myocardial function, sublingual microcirculation, mitochondrial function, plasma cardiac troponin I, and interleukin‐1β, expression of proteins in SIRT1 (sirtuin 1)/PGC‐1α (peroxisome proliferator‐activated receptor gamma coactivator 1‐alpha) and NLRP3 (the NOD‐like receptor family protein 3) inflammasome pathways were evaluated. Following cardiopulmonary resuscitation, myocardial function was compromised with a significantly decreased cardiac output, ejection fraction, and increased myocardial performance index, cardiac troponin I. Sublingual microcirculation was disturbed with impaired perfused vessel density and microvascular flow index. Cardiac arrest reduced mitochondrial routine respiration, Complex I‐linked respiration, respiratory control rates and oxidative phosphorylation coupling efficiency. PEG‐20k or MCC950 alone restored mitochondrial respiratory function, restituted sublingual microcirculation, and preserved myocardial function, whereas a combination of PEG‐20k and MCC950 further improved these aspects. PEG‐20k restored the expression of SIRT1 and PGC‐1α, and blunted activation of NLRP3 inflammasomes. MCC950 suppressed expression of cleaved‐caspase‐1/pro‐caspase‐1, ASC (apoptosis‐associated speck‐like protein), GSDMD [gasdermin d], and interleukin‐1β. Conclusions Combined therapy with PEG‐20k and MCC950 is superior to either therapy alone for preserving post‐resuscitated myocardial function, restituting sublingual microcirculation at restoration of spontaneous circulation at 6 hours. The responsible mechanisms involve upregulated expression of SIRT1/PGC1‐α in tandem with inhibition of NLRP3 inflammasomes.

Post‑cardiac arrest myocardial dysfunction (PAMD) is a leading cause of death in patients undergoing resuscitation patients following cardiac arrest (CA). Although prostaglandin E1 (PGE1) is a clinical drug used to mitigate ischemia injury, its effect on PAMD remains unknown. In the present study, the protective effects of PGE1 on PAMD were evaluated in a rat model of CA and in a hypoxia‑reoxygenation (H/R) in vitro model. Rats were randomly assigned to CA, CA+PGE1 or sham groups. Asphyxia for 8 min followed by cardiopulmonary resuscitation were performed in the CA and CA+PGE1 groups. PGE1 was intravenously administered at the onset of return of spontaneous circulation (ROSC). PGE1 treatment significantly increased the ejection fraction and cardiac output within 4 h following ROSC and improved the survival rate, compared with the CA group. Moreover, PGE1 inactivated GSK3β, prevented mitochondrial permeability transition pore (mPTP) opening, while reducing cytochrome c and cleaved caspase‑3 expression, as well as cardiomyocyte apoptosis in the rat model. To examine the underlying mechanism, H/R H9c2 cells were treated with PGE1 at the start of reoxygenation. The changes in GSK3β activity, mPTP opening, cytochrome c and cleaved caspase‑3 expression, and apoptosis of H9c2 cells were consistent with those noted in vivo. The results indicated that PGE1 attenuated PAMD by inhibiting mitochondria‑mediated cardiomyocyte apoptosis.

Background: Post-cardiac arrest myocardial dysfunction (PAMD) is a leading cause of death in resuscitated patients after cardiac arrest (CA). Prostaglandin E1 (PGE1) is a clinical drug used to mitigate ischemia injury. However, its effect on PAMD remains unknown. Methods: We investigated the protective effects of PGE1 on PAMD in a rat model of cardiac arrest and a hypoxia-reoxygenation (H/R) H9c2 cell model. Forty-two male Wistar rats were randomly assigned to CA, CA+PGE1, and sham groups. Asphyxia for 8 min followed by cardiopulmonary resuscitation was performed in the CA and CA+PGE1 groups. PGE1 (1 μg/kg) was intravenously administered at the onset of return of spontaneous circulation (ROSC). Ejection fraction (EF) and cardiac output (CO) were measured at baseline, 1, 2, 3, and 4 h after ROSC; survival was monitored for 72 h. Cardiomyocyte apoptosis, mitochondrial permeability transition pore (mPTP) opening, and protein levels of glycogen synthase kinase 3β (GSK3β), cytochrome c, and cleaved caspase-3 were measured 4 h after ROSC. H9c2 cells were treated with PGE1(0.5 μM) at the start of reoxygenation. Apoptosis, mPTP opening, and protein levels of GSK3β, cytochrome c, and cleaved caspase-3 of H9c2 cells were detected. Results: Compared to the CA group, PGE1 treatment significantly increased the EF and CO within 4 h after ROSC and improved the survival rate. It activated GSK3β, prevented mPTP opening, suppressed cytochrome c and cleaved caspase-3 expression, and reduced cardiomyocyte apoptosis in the rat model. In vitro, Changes in GSK3β, mPTP opening, cytochrome c and cleaved caspase-3 expression, and apoptosis in H9c2 cells were consistent with those in the rat model. Conclusions: Our results indicate that PGE1 attenuates PAMD via inhibiting mitochondria-mediated cardiomyocyte apoptosis.

Objective: Acute carbon monoxide （CO）poisoning can cause delayed neurological sequelae (DNS). Glycogen synthase kinase 3β (GSK-3β) /Tau protein pathway is reported to play a key role in neurological abnormalities. In the present study, we aimed to determine the role of GSK-3β/Tau in DNS following acute CO poisoning.Methods: 4-benzyl-2-methyl-1,2,4-thiadiazolidine-3,5-dione (TDZD-8), a specific non-competitive inhibitor of GSK-3β, was used to inhibit GSK-3β. Twenty-four male Sprague-Dawley rats were randomly assigned to the three groups: Control group, CO group and CO-TDZD-8 group. Rats breathed 1000 ppm CO for 40 minutes and then 3000 ppm for up to 20 minutes until they lost consciousness. TDZD-8 (1 mg/kg) was administered intravenously three times after the end of CO exposure at 0, 24, 48 hours late. Learning and memory abilities were observed using the Morris Water Maze (MWM). Brain histological changes were evaluated by hematoxylin-eosin staining. Moreover, the expression levels of Tau and GSK-3β were detected after acute carbon monoxide poisoning.Results: TDZD-8 significantly attenuated the learning and memory dysfunction induced by acute CO poisoning, ameliorated the histology structure of damaged neural cells in cortex and hippocampus CA1 area. TDZD-8 clearly decreased p-Tau expression, reversed the reduction of p-GSK-3β induced by acute CO poisoning.Conclusions: The therapeutic effect of TDZD-8 in alleviating DNS caused by acute CO poisoning is related to the inactivation of Tau by intensifying the level of GSK-3β phosphorylation.

The physiology and physiopathology process of mitochondrial function following cardiac arrest remains poorly understood. We aimed to assess mitochondrial respiratory function on the heart and brain homogenates from cardiac arrest rats. The expression level of SIRT1/PGC-1α pathway was measured by immunoblotting. 30 rats were assigned to the CA group and the sham group. Rats of CA were subjected to 6 min of untreated ventricular fibrillation (VF) followed by 8 min of cardiopulmonary resuscitation (CPR). Mitochondrial respiratory function was compromised following CA and I/R injury, as indicated by CIL (451.46 ± 71.48 vs. 909.91 ± 5.51 pmol/min*mg for the heart and 464.14 ± 8.22 vs. 570.53 ± 56.33 pmol/min*mg for the brain), CI (564.04 ± 64.34 vs. 2729.52 ± 347.39 pmol/min*mg for the heart and 726.07 ± 85.78 vs. 1762.82 ± 262.04 pmol/min*mg for the brain), RCR (1.88 ± 0.46 vs. 3.57 ± 0.38 for the heart and 2.05 ± 0.19 vs. 3.49 ± 0.19, for the brain) and OXPHOS coupling efficiency (0.45 ± 0.11 vs. 0.72 ± 0.03 for the heart and 0.52 ± 0.05 vs. 0.71 ± 0.01 for the brain). However, routine respiration was lower in the heart and comparable in the brain after CA. CIV did not change in the heart but was enhanced in the brain. Furthermore, both SIRT1 and PGC-1α were downregulated concurrently in the heart and brain. The mitochondrial respiratory function was compromised following CA and I/R injury, and the major affected respiratory state is complex I-linked respiration. Furthermore, the heart and the brain respond differently to the global I/R injury after CA in mitochondrial respiratory function. Inhibition of the SIRT1/PGC-1α pathway may be a major contributor to the impaired mitochondrial respiratory function.

Acute pulmonary edema is one of the major outcomes of exposure to high levels of hydrogen sulfide (H2S). However, the mechanisms involved in H2S-induced acute pulmonary edema are still poorly understood. Therefore, the present study is designed to evaluate the role of epithelial sodium channel (ENaC) in H2S-induced acute pulmonary edema. The Sprague-Dawley rats were exposed to sublethal concentrations of inhaled H2S, then the pulmonary histological and lung epithelial cell injury were evaluated by hematoxylin-eosin staining and electron microscopy, respectively. In addition to morphological investigation, our results also revealed that H2S exposure significantly decreased the alveolar fluid clearance and increased the lung tissue wet-dry ratio. These changes were demonstrated to be associated with decreased ENaC expression. Furthermore, the extracellular-regulated protein kinases 1/2 pathway was demonstrated to be implicated in H2S-mediated ENaC expression, because PD98059, an ERK1/2 antagonist, significantly mitigated H2S-mediated ENaC down-regulation. Therefore, our results show that ENaC might represent a novel pharmacological target for the treatment of acute pulmonary edema induced by H2S and other hazardous gases.

Urinary trypsin inhibitor (UTI) decreases inflammatory cytokine levels and mortality in experimental animal models of inflammation. Here, we observed the effect of UTI on lipopolysaccharide (LPS)-induced hyperpermeability in human umbilical vein endothelial cells (HUVECs) and explored the role of vascular endothelial-cadherin (VE-cadherin) in its effect. The effect of UTI on endothelial barrier hyperpermeability was detected by an electrical cell–substrate impedance sensing (ECIS) system and a transwell chamber system. The expression of VE-cadherin in HUVECs was examined by real-time PCR and western blot. We demonstrated that the alleviation of LPS-induced barrier dysfunction could be achieved by pretreatment with 3000 U/mL of UTI. VE-cadherin monoclonal antibody (mAb) could inhibit the protective effects. UTI maintained VE-cadherin expression by increasing protein stability at both the transcriptional and post-transcriptional levels. Meanwhile, VE-cadherin expression on the cell surface increased when the cells were pretreated with UTI. Furthermore, pretreatment with UTI decreased the phosphorylation of VE-cadherin at Tyr658 but not Tyr731. Our data show that prophylactic UTI maintains the endothelial barrier function, increases VE-cadherin expression, and inhibits the phosphorylation of VE-cadherin at Tyr658 under inflammatory conditions. It suggests a scientific and potential clinical therapeutic importance of UTI in treatment of inflammatory disorders.

Acute lung injury (ALI) is one of the fatal outcomes after exposure to high levels of hydrogen sulfide (H2S), and the matrix metalloproteinases (MMPs) especially MMP-2 and MMP-9 are believed to be involved in the development of ALI by degrading the extracellular matrix (ECM) of blood-air barrier. However, the roles of MMP-2 and MMP-9 in H2S-induced ALI and the mechanisms of dexamethasone (DXM) in treating ALI in clinical practice are still largely unknown. The present work was aimed to investigate the roles of MMP-2 and MMP-9 in H2S-induced ALI and the protective effects of DXM. In our study, SD rats were exposed to H2S to establish the ALI model and in parallel, A549 cells were incubated with NaHS (a H2S donor) to establish cell model. The lung HE staining, immunohistochemisty, electron microscope assay and wet/dry ratio were used to identify the ALI induced by H2S, then the MMP-2 and MMP-9 expression in both rats and A549 cells were detected. Our results revealed that MMP-2 and MMP-9 were obviously increased in both mRNA and protein level after H2S exposure, and they could be inhibited by MMP inhibitor doxycycline (DOX) in rat model. Moreover, DXM significantly ameliorated the symptoms of H2S-induced ALI including alveolar edema, infiltration of inflammatory cells and the protein leakage in BAFL via up-regulating glucocorticoid receptor(GR) to mediate the suppression of MMP-2 and MMP-9. Furthermore, the protective effects of DXM in vivo and vitro study could be partially blocked by co-treated with GR antagonist mifepristone (MIF). Our results, taken together, demonstrated that MMP-2 and MMP-9 were involved in the development of H2S-induced ALI and DXM exerted protective effects by alleviating the expression of MMP-2 and MMP-9. Therefore, MMP-2 and MMP-9 might represent novel pharmacological targets for the treatment of H2S and other hazard gases induced ALI.

Acute pulmonary edema (APE) is one of the fatal outcomes after exposure to high levels of hydrogen sulfide (H2S), available evidence suggest that dexamethasone (DXM), a potent anti-inflammatory agent, has been widely used or proposed as a therapeutic approach for H2S-induced APE in clinical practice, however, the underlying mechanism remains poorly understood. Ample evidence suggest that epithelial Na(+) channel, especially for the subunit α-epithelial Na(+) channel (α-ENaC) plays a critical role in alveolar fluid clearance. Therefore, the present study is undertaken to investigate the effects of DXM on α-ENaC following H2S exposure. The Sprague Dawley rats were exposed to H2S to establish APE model, in parallel, A549 cells were treated with NaHS to establish cell model. In vivo study, we found that DXM significantly attenuated H2S-induced lung histopathological changes and alveolar fluid clearance decrement, however, these preventive effects of DXM can be obviously counteracted by the mifepristone (MIF), the glucocorticoid receptor (GR) blocker. Moreover, DXM markedly attenuated H2S-mediated α-ENaC down-regulation, and similarly, the process can be partially retarded by MIF. Furthermore, DXM obviously prevented H2S-mediated ERK1/2 activation both in vitro and in vivo study. These results, taken together, suggested that DXM exerted protective effects on H2S-induced APE, and α-ENaC might be a potential therapeutic target for APE induced by H2S.