Lung Ischemia-reperfusion Injury In Rats Research Articles

Circulating mtNFPs Are Associated with ARDS after CPB and Regulate Endothelial Barrier through FPR2.

Cardiopulmonary bypass (CPB) increases the risk of acute respiratory distress syndrome (ARDS) due to endothelial cell (EC) barrier dysfunction. However, the specific role of mitochondrial N-formyl peptides (mtNFPs) in ARDS following CPB remains unexplored. Here, we investigated the differential expression of circulating mtNFPs in patients after CPB, focusing on the novel role of FPR2 in ECs. Levels of circulating mtNFPs were assessed using enzyme-linked immunosorbent assay (ELISA). Several mtNFPs (ND4, ND5, ND6, and Cox1) were significantly upregulated in patients with ARDS at day 1 post-CPB compared to patients without ARDS. Higher levels of ND6 were correlated with worst PaO2/FiO2 (r=-0.2219 and P<0.0001) and cardiac Troponin T (r=2.107 and P<0.0001). Utilizing patient-derived serum and a rat lung ischemia reperfusion injury (LIRI) model, we observed a positive correlation between serum ND6 concentration and ARDS, which is also associated with EC barrier dysfunction. In vitro experiments, using trans-endothelial electric resistance (TEER) measurements and fluorescence microscopy with FITC-labeled VE-cadherin, demonstrated that ND6 disrupts the EC barrier through FPR2. Furthermore, FPR2 controls the release of ND6 out of mitochondria and cytoplasm under hypoxia reoxygenation (HR). Activated FPR2 leads to upregulation of nuclear transcription factor-kappa B (NF-κB) by inducing IκBα phosphorylation, promoting ICAM1 and VCAM1 expression, thereby compromising EC barrier integrity. Circulating pro-inflammatory and barrier-disruptive mtNFPs, particularly ND6, are associated with ARDS in patients undergoing CPB. The novel ND6-FPR2 axis regulates inflammation and EC permeability through the NF-κB pathway.

Preparation and Evaluation of Preventive Effects of Inhalational and Intraperitoneal Injection of Myrtenol Loaded Nano-Niosomes on Lung Ischemia-Reperfusion Injury in Rats

IntroductionIschemia-reperfusion injury (IRI) is directly related to forming reactive oxygen species, endothelial cell injury, increased vascular permeability, and the activation of neutrophils and cytokines. Niosomes are nanocarriers and an essential part of drug delivery systems. We aimed to investigate the effects of myrtenol's inhaled and intraperitoneal niosomal form, compared to its simple form, on lung ischemia reperfusion injury (LIRI). Material and methodWistar rats were divided into ten groups. Simple and niosomal forms of myrtenol were inhaled or intraperitoneally injected daily for one week prior to LIRI. We evaluated oxidative stress, apoptotic, and inflammatory indices, nitric oxide, inducible nitric oxide synthase (iNOS), endothelial nitric oxide synthase (eNOS) and histopathological indices. ResultsPretreatment with simple and niosomal forms of myrtenol significantly inhibited the indices of pulmonary edema, pro-inflammatory cytokines and proteins, oxidant agents, nitric oxide, iNOS, apoptotic proteins, congestion of capillaries, neutrophil infiltration, and bleeding in the alveoli. Furthermore, myrtenol increased anti-inflammatory cytokines, anti-oxidants agents, eNOS, anti-apoptotic proteins and the survival time of animals. The niosomal form of myrtenol showed a more ameliorative effect than its simple form. ConclusionThe results showed the superior protective effect of the inhalation of myrtenol niosomal form against LIRI compared to its simple form and systemic use.

Post-ischemic treatment of nalmefene hydrochloride attenuated lung ischemia-reperfusion injury in rats via the Sirt1/Nrf2/HO-1 pathway with inhibition of ferroptosis

Objective: To study the effect and mechanism of post-ischemic treatment of nalmefene in alleviating the lung ischemia-reperfusion injury by inhibiting ferroptosis through activation of the Sirt 1/Nrf 2/HO-1 axis. Methods: A total of 60 rats were randomly divided into six groups equally (n=10): the sham group, the model group(I/R), the nalmefene group, the nalmefene+EX527 group, the nalmefene+ML385 group, the nalmefene+Fe-citrate group (nalmefene+Fe group). The sham group without drug treatment was not treated with ischemia-reperfusion. The pulmonary ischemia-reperfusion model was established by occlusion of the left pulmonary hilum in the model group without drug treatment. After ischemic treatment, the nalmefene group was injected with nalmefene (15 μg/kg) via the tail vein at 5 minutes before reperfusion. The nalmefene+EX527 group, the nalmefene+ML385 group, and the nalmefene+Fe group were injected intraperitoneally with EX527 (5 mg/kg), ML385 (30 mg/kg), Fe-citrate(15 mg/kg), respectively, 2 h before moulding and then injected with nalmefene (15 μg/kg) via the tail vein at 5 minutes before reperfusion. All rats were sacrificed three hours after reperfusion, and the specimens from the upper lobe of the left lung tissue were preserved. The degree of lung tissue injury and the wet/dry weight ratio were assessed in each group of rats. Fe 2+, MDA, TNF-α, and IL-6 content, GSH activity and the expression levels of Sirt1, Nrf2, HO-1, ACSL4 and GPX4 were determined. Results: Compared with the sham group, the wet/dry weight ratio, lung tissue injury score, ACSL 4 expression level, Fe 2+, TNF-α, IL-6 and MDA content, Sirt 1, Nrf 2, HO-1 messenger RNA and protein expression levels were significantly increased (P<0.01), while GPX 4 expression level and GSH activity were significantly decreased in the model group (P<0.01). Compared with the model group, wet/dry weight ratio, lung tissue injury score, ACSL 4 expression level, Fe 2+, TNF-α, IL-6, and MDA content decreased significantly (P<0.01), Nrf 2, HO-1 messenger RNA and protein, GPX 4 expression, and GSH activity were significantly increased in the nalmefene group and the nalmefene+EX527 group (P<0.01). Sirt 1 messenger RNA and protein expression increased significantly in the nalmefene (P<0.01) and the nalmefene+EX527 groups (P>0.05). In the nalmefene+ML385 group, the wet/dry weight ratio, lung tissue injury score, TNF-α and IL-6 content were decreased significantly (P<0.01), while Sirt 1 messenger RNA and protein expression levels were significantly increased (P<0.01), but there were no significant changes in Nrf 2, HO-1 messenger RNA and protein expression levels, ACSL 4 and GPX 4 expression levels, Fe 2+, MDA content, and GSH activity (P>0.05). In the nalmefene+Fe group, wet/dry weight ratio, lung-injury score, TNF-α, IL-6, MDA content were decreased significantly (P<0.01), messenger RNA and protein expression levels of Sirt 1, Nrf 2, HO-1, and GSH activity were increased significantly (P<0.01), but there were no significant changes in Fe 2+content, ACSL 4 and GPX 4 expression levels (P>0.05). Compared with the nalmefene group, in the nalmefene+EX527 group, the nalmefene+ML385 group and the nalmefene+Fe group, wet/dry weight ratio, lung tissue damage score, ACSL 4 expression level, TNF-α, IL-6 and MDA content were significantly increased (P<0.01), the expression level of GPX 4 and GSH activity were significantly decreased (P<0.01). The expression levels of Sirt 1, Nrf 2, HO-1 messenger RNA and protein were significantly decreased in the nalmefene+EX527 group (P<0.01). The expression levels of Nrf 2, HO-1 messenger RNA and protein decreased significantly in the namemefene+ML385 group (P<0.01), but there was no significant change in Sirt 1 messenger RNA and protein expression level (P>0.05). Sirt 1, Nrf 2, HO-1 messenger RNA-protein expression levels did not change significantly in the nalmefene+Fe group (P>0.05). Conclusion: Post-ischemic treatment with nalmefene hydrochloride may alleviate pulmonary ischemia-reperfusion injury by inhibiting ferroptosis through activation of the Sirt 1/Nrf 2/HO-1 axis.

Perfluorocarbon restrains inflammation and cell apoptosis in rats with lung ischemia reperfusion injury via down regulation of TLR4/NF κB signaling pathway

Purpose: To exaluate the effect of perfluorocarbon on lung ischemia-reperfusion injury in rats, and to unravel the potential underlying mechanism.Methods: A total of 36 Sprague-Dawley (SD) rats were randomly assigned to sham group, model group, and perfluorocarbon group (12 rats per group). The levels of inflammatory factors (TNF-α and IL1β) were determined using enzyme-linked immunosorbent assay (ELISA). Cell apoptosis was evaluated by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay, while Western blotting was conducted to determine the protein expressions of TLR4 and NF-κB.Results: The levels of inflammatory factors in the model and perfluorocarbon groups were significantly higher than those in operation group (p &lt; 0.05), while their levels in perfluorocarbon group were significantly lower than in model group (p &lt; 0.05). The mRNA expression levels of TNF-α and IL-1β in lung tissues rose significantly in both model and perfluorocarbon groups when compared with those in sham group (p &lt; 0.05), but declined significantly in the perfluorocarbon group in comparison with those in model group (p &lt; 0.05). Furthermore, the perfluorocarbon group exhibited a significantly lower cell apoptosis than model group (p &lt; 0.05). The relative protein expression levels of TLR4 and NF-κBdeclined significantly in perfluorocarbon group than in model group.Conclusions: Perfluorocarbon down-regulates the TLR4/NF-κB signaling pathway, and inhibits inflammation an

Dexmedetomidine Alleviates Lung Oxidative Stress Injury Induced by Ischemia-Reperfusion in Diabetic Rats via the Nrf2-Sulfiredoxin1 Pathway.

Oxidative stress injury (OSI) is an important pathological process in lung ischemia-reperfusion injury (LIRI), and diabetes mellitus (DM) can exacerbate this injury. Dexmedetomidine protects against LIRI by reducing OSI. However, the effect of dexmedetomidine on LIRI under diabetic conditions remains unclear. Therefore, this study is aimed at exploring the effects and mechanisms of dexmedetomidine on OSI induced by LIRI in diabetic rats. Rats were randomly divided into control+sham (CS), DM+sham (DS), control+ischemia-reperfusion (CIR), DM+ischemia-reperfusion (DIR), and DM+ischemia-reperfusion+dexmedetomidine (DIRD) groups (n = 6). In the CS and DS groups, the nondiabetic and diabetic rats underwent thoracotomy only without LIRI. In the CIR, DIR, and DIRD groups, LIRI was induced through left hilum occlusion for 60 min, followed by reperfusion for 120 min in nondiabetic and diabetic rats, and rats in the DIRD group were administered dexmedetomidine (3, 5, and 10 μg/kg). Compared with those in the CS group, the OSI, lung compliance, apoptosis, and oxygenation indices deteriorated in the DS group (P < 0.05), and these indices were further aggravated in the CIR and DIR groups (P < 0.05), being the worst in the DIR group (P < 0.05). Compared to those of the DIR group, the OSI, lung compliance (15.8 ± 2.4 vs. 11.6 ± 1.7 ml/kg), apoptosis (22.5 ± 2.6 vs. 51.8 ± 5.7), oxygenation (381 ± 58 vs. 308 ± 78 mmHg), and caspase-3 and caspase-9 protein expression indices were attenuated, and Nrf2 and sulfiredoxin1 protein expression was increased in the DIRD group (P < 0.05). And the lung injury, oxygenation, OSI, and Nrf2 and sulfiredoxin1 protein expression changed in a concentration-dependent manner. In conclusion, dexmedetomidine alleviated lung OSI and improved lung function in a diabetic rat LIRI model through the Nrf2-sulfiredoxin1 pathway.

Methane Inhalation Protects Against Lung Ischemia-Reperfusion Injury in Rats by Regulating Pulmonary Surfactant via the Nrf2 Pathway.

Methane (CH4) exerted protective effects against lung ischemia-reperfusion (I/R) injury, but the mechanism remains unclear, especially the role of pulmonary surfactant. Therefore, this study aimed to explore the effects of CH4 inhalation on pulmonary surfactant in rat lung I/R injury and to elucidate the mechanism. Rats were randomly divided into three groups (n = 6): the sham, I/R control, and I/R CH4 groups. In the sham group, only thoracotomy was performed on the rats. In the I/R control and I/R CH4 groups, the rats underwent left hilum occlusion for 90 min, followed by reperfusion for 180 min and ventilation with O2 or 2.5% CH4, respectively. Compared with those of the sham group, the levels of large surfactant aggregates (LAs) in pulmonary surfactant, lung compliance, oxygenation decreased, the small surfactant aggregates (SAs), inflammatory response, oxidative stress injury, and cell apoptosis increased in the control group (P < 0.05). Compared to the control treatment, CH4 increased LA (0.42 ± 0.06 vs. 0.31 ± 0.09 mg/kg), oxygenation (201 ± 11 vs. 151 ± 14 mmHg), and lung compliance (16.8 ± 1.0 vs. 11.5 ± 1.3 ml/kg), as well as total antioxidant capacity and Nrf2 protein expression and decreased the inflammatory response and number of apoptotic cells (P < 0.05). In conclusion, CH4 inhalation decreased oxidative stress injury, inflammatory response, and cell apoptosis, and improved lung function through Nrf2-mediated pulmonary surfactant regulation in rat lung I/R injury.

Parecoxib mitigates lung ischemia-reperfusion injury in rats by reducing oxidative stress and inflammation and up-regulating HO-1 expression.

ABSTRACTPurpose:To investigate the protective effect of parecoxib against lung ischemia-reperfusion injury (LIRI) in rats and the mechanism.Methods:Thirty rats were divided into sham-operated, LIRI and LIRI+parecoxib groups. LIRI model (ischemia for 60 min, followed by reperfusion for 120 min) was constructed in LIRI and LIRI+parecoxib groups. In LIRI+parecoxib group, 10 mg/kg parecoxib was given via femoral vein 15 min before ischemia beginning. At the end of the reperfusion, blood gas analysis, lung wet to dry mass ratio measurement, lung tissue biochemical determination and heme oxygenase-1 (HO-1) protein expression determination were performed.Results:Compared with LIRI group, in LIRI+parecoxib group the oxygenation index was significantly increased, the alveolar-arterial oxygen partial pressure difference was significantly decreased, the lung wet to dry mass ratio was significantly decreased, the lung tissue malondialdehyde content was significantly decreased, the lung tissue superoxide dismutase and myeloperoxidase activities were significantly increased, the lung tissue tumor necrosis factor α and interleukin 1β levels were significantly decreased, and the lung tissue HO-1 protein expression level was significantly increased (all P < 0.05).Conclusions:Parecoxib pretreatment can mitigate the LIRI in rats by reducing oxidative stress, inhibiting inflammatory response and up-regulating HO-1 expression in lung tissue.

Dexamethasone on alleviating lung ischemia/reperfusion injury in rats by regulating PI3K/AKT pathway

To investigate the protective effect and mechanism of dexamethasone in lung ischemia/reperfusion injury (LIRI) rats. (1) Part one experiment: 24 Sprague-Dawley (SD) rats were divided into four groups according to the random number method (n = 6): standard ventilation group (N group), normal saline group (NS group), LIRI group, and dexamethasone+LIRI group (DEX group). The rat model of LIRI was established by clamping the left pulmonary hilum for 1 hour and reperfusing it for 2 hours. The DEX group was given dexamethasone 3 mg/kg 5 minutes before reperfusion, and NS group was injected with normal saline. Group N did not receive any treatment. The left lung tissue of the rats in each group were taken alive 2 hours after reperfusion. The lung tissue was harvested for lung wet/dry mass ratio (W/D) measurement. Hematoxylin-eosin (HE) staining and electron microscopy was used to observe the pathological changes of lung tissue and to assess the degree of injury. Ultrastructural changes of lung tissue were observed under electron microscope. The levels of tumor necrosis factor-α (TNF-α), interleukin (IL-1β, IL-6) in lung tissue were detected by enzyme linked immunosorbent assay (ELISA). The expressions of phosphorylated protein kinase B (p-AKT) was detected by Western Blot. (2) Part two experiment: intervention with phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) pathway inhibitor LY294002 to further explore the mechanism of dexamethasone in reducing lung injury induced by LIRI. Twenty-four SD rats were divided into four groups according to the random number method (n = 6): N group, LIRI group, DEX group, and dexamethasone+LY294002+LIRI group (LY group). All the groups except the LY group were treated with membrane and intervention according to part one experiment. The LY group was injected with LY294002 0.3 mg/kg after injection of dexamethasone. The expressions of M1 macrophage polarization markers CD11c, CD16, and M2 macrophage polarization markers CD206, Arg1 were detected by immunohistochemistry. (1) Part one experiment: compared with N group, the morphological and ultrastructural changes of lung tissue in the LIRI group were significantly changed, lung injury score, lung W/D ratio and TNF-α, IL-1β, IL-6 levels were significantly increased, and p-AKT expression was significantly decreased. Compared with the LIRI group, the morphological and ultrastructural changes of the lung tissue in the DEX group were significantly improved, and the lung injury score was reduced (5.00±0.89 vs. 8.83±0.75), lung W/D ratio and TNF-α, IL-1β, IL-6 levels were significantly decreased [lung W/D ratio: 6.25±0.56 vs. 8.27±0.72, TNF-α (ng/L): 93.28±16.42 vs. 205.90±25.30, IL-1β (ng/L): 130.10±10.81 vs. 209.10±19.20, IL-6 (ng/L): 195.80±21.17 vs. 310.50±20.77], p-AKT expression was significantly increased [p-AKT/AKT: (57.58±8.80)% vs. (36.62±9.25)%], and the differences were statistically significant (all P < 0.05). There was no significant difference in each index between NS group and N group. (2) Part two experiment: compared with the N group, the expression of macrophage polarization markers CD11c, CD16, CD206 and Arg1 in the LIRI group were significantly increased. Compared with the LIRI group, the expressions of CD11c and CD16 in the lung tissue of the DEX group were significantly decreased, and the expressions of CD206 and Arg1 were significantly increased. The intervention of PI3K/AKT signaling pathway inhibitor LY294002 significantly blocked the effect of dexamethasone on LIRI-mediated macrophage polarization (CD11c immunohistochemical score: 7.20±0.36 vs. 5.00±0.34, CD16 immunohistochemical score: 8.20±0.48 vs. 7.40±0.64, CD206 immunohistochemical score: 5.80±0.59 vs. 7.40±0.28, Arg1 immunohistochemical score: 7.20±0.72 vs. 8.80±0.48, all P < 0.05). Dexamethasone pretreatment can alleviate the intrapulmonary inflammatory response and lung injury caused by LIRI in rats. The mechanism of action is related to the polarization direction of pulmonary macrophagesvia activation of the PI3K/AKT pathway by dexamethasone.

Kaempferol Improves Lung Ischemia-Reperfusion Injury via Antiinflammation and Antioxidative Stress Regulated by SIRT1/HMGB1/NF-κB Axis.

Trauma, organ transplantation, and thromboembolism are the main causes of lung ischemia-reperfusion injury (LIRI), and new therapies and drugs are urgent to relieve LIRI. In preliminary experiment, authors found that kaempferol could improve LIRI in rats, and the current study further explored its possible mechanism. The model of rat LIRI was established and appropriate research methods were implemented. Results shown that kaempferol could significantly improve LIRI, inhibit release of inflammatory factors including interleukin (IL) 6 and tumor necrosis factor (TNF) α in bronchoalveolar lavage fluid, and reduce oxidative stress reaction. Western blotting was used to detect protein expression levels and found that kaempferol could up-regulate the protein expressions of phosphorylated (p-) p65 and p65, and down-regulate the protein expression of sirtuin (SIRT) 1. Immunofluorescence was used to localize the expression of high mobility group box (HMGB) 1 and found its higher expression in outside of nucleus. However, the above effects of kaempferol on LIRI markedly attenuated by EX 527, a selective inhibitor of SIRT 1. Taken together, we first reported the protective effect of kaempferol on rat LIRI and confirmed that kaempferol’s antiinflammation and antioxidative stress involving the SIRT1/HMGB1/NF-κB axis.

Protective effects of a hydrogen-rich solution during cold ischemia in rat lung transplantation

BackgroundMolecular hydrogen can reduce the oxidative stress of ischemia-reperfusion injury in various organs for transplantation and potentially improve survival rates in recipients. This study aimed to evaluate the protective effects of a hydrogen-rich preservation solution against ischemia-reperfusion injury after cold ischemia in rat lung transplantation. MethodsLewis rats were divided into a nontransplant group (n = 3), minimum-ischemia group (n = 3), cold ischemia group (n = 6), and cold ischemia with hydrogen-rich (more than 1.0 ppm) preservation solution group (n = 6). The rats in the nontransplant group underwent simple thoracotomy, and the rats in the remaining 3 groups underwent orthotopic left lung transplantation. The ischemic time was <30 minutes in the minimum-ischemia group and 6 hours in the cold ischemia groups. After 2-hour reperfusion, we evaluated arterial blood gas levels, pulmonary function, lung wet-to-dry weight ratio, and histologic features of the lung tissue. The expression of proinflammatory cytokines was measured using quantitative polymerase chain reaction assays, and 8-hydroxydeoxyguanosine levels were evaluated using enzyme-linked immunosorbent assays. ResultsWhen compared with the nontransplant and minimum-ischemia groups, the cold ischemia group had lower dynamic compliance, lower oxygenation levels, and higher wet-to-dry weight ratios. However, these variables were significantly improved in the cold ischemia with hydrogen-rich preservation solution group. This group also had fewer signs of perivascular edema, lower interleukin-1β messenger RNA expression, and lower 8-hydroxydeoxyguanosine levels than the cold ischemia group. ConclusionsThe use of a hydrogen-rich preservation solution attenuates ischemia-reperfusion injury in rat lungs during cold ischemia through antioxidant and anti-inflammatory effects.

Role of poly(ADP-ribose) polymerase-1 on lung ischemia-reperfusion injury in rats: the relationship with autophagy

Objective To evaluate the role of poly(ADP-ribose) polymerase-1 (PARP-1) in lung ischemia-reperfusion (I/R) injury in rats and the relationship with autophagy. Methods Twenty-four clean-grade healthy male Sprague-Dawley rats, aged 8-12 weeks, weighing 200-240 g, were divided into 3 groups (n=8 each) using a random number table method: sham operation group (S group), lung I/R group (I/R group) and lung I/R plus PARP-1 inhibitor PJ34 group (I/R+ PJ34 group). The chest was only opened without clamping the left hilum of lung in group S. Lung I/R injury model was established by clamping the left hilum of lung for 45 min followed by 120 min reperfusion in I/R and I/R+ PJ34 groups.PJ34 10 mg/kg was intraperitoneally injected at 30 min before ischemia in group I/R+ PJ34, while the equal volume of normal saline was injected in S and I/R groups.The rats were sacrificed at the end of reperfusion, and lungs were removed for microscopic examination of the pathological changes (with a light microscope) which were scored and for determination of wet to dry weight ratio (W/D ratio), cell apoptosis (by TUNEL), expression of PARP-1 activity markers (PAR), Bcl-2, Bax, microtubule-associated protein 1 light chain 3 Ⅰ (LC3-Ⅰ), LC3-Ⅱ and Beclin-1 (using Western blot). The apoptosis index, Bcl-2/Bax ratio and LC3-Ⅱ/LC3-Ⅰ ratio were calculated. Results Compared with S group, the W/D ratio, pathological scores, apoptosis index and LC3-Ⅱ/LC3-Ⅰ ratio were significantly increased, the expression of PAR and Beclin-1 was up-regulated, and Bcl-2/Bax ratio was decreased in I/R and I/R+ PJ34 groups (P<0.05). Compared with I/R group, the W/D ratio, pathological scores, apoptosis index, and LC3-Ⅱ/LC3-Ⅰ ratio were significantly decreased, the expression of PAR and Beclin-1 was down-regulated, and Bcl-2/Bax ratio was increased in I/R+ PJ34 group (P<0.05). Conclusion PARP-1 activation is involved in lung I/R injury in rats, and the mechanism may be related to increasing autophagy and inducing cell apoptosis. Key words: Poly(ADP-ribose) polymerases; Lung; Reperfusion injury; Autophagy

Dexmedetomidine alleviates lung ischemia-reperfusion injury in rats by activating PI3K/Akt pathway.

This research aims to investigate the role and mechanism of PI3K/Akt pathway in the pathological process of lung ischemia-reperfusion injury in dexmedetomidine-treated rats. Forty-five healthy male Sprague-Dawley rats were divided into three groups: sham operation group, lung ischemia-reperfusion group (IR group) and dexmedetomidine pretreatment group (Dex group). Rats in the sham operation group did not receive other procedures except for opening left chest. The left lung hilar of rats in the IR group was clamped with non-invasive vascular clamp after anesthesia to establish an ischemic model. After 1 hour, the vascular clamp was released and the rats were reperfused for 2 hours. As for rats in the Dex group, 3 μg/kg of dexmedetomidine (pumping time of 10 min) was pumped through the tail vein before releasing the left hilar clamp. After the experiment, blood samples and lung tissues were collected. Serum levels of interleukin 6 (IL-6), tumor necrosis factor-α (TNF-α), IL-10, and IL-1 in rats were examined. Activities of malondialdehyde (MDA), myeloperoxidase (MPO), superoxide dismutase (SOD) and catalase (CAT) in rat lung tissues were also detected. Besides, the expressions of hypoxia-inducible factor-la (HIF-la), p-Akt, Caspase-3, and Caspase-9 in lung tissues were detected by Western blot. The mRNA expression levels of HIF-1a, p-Akt, Caspase-3, and Caspase-9 in lung tissues were evaluated by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Lung ischemia-reperfusion markedly increased the levels of IL-6, TNF-α, IL-10, and IL-1 in the IR group. In contrast, dexmedetomidine pretreatment decreased the expression levels of IL-6, TNF-α, IL-10, and IL-1in the Dex group. Also, the activities of MDA and MPO in lung tissues of rats in the IR group significantly increased after lung ischemia-reperfusion injury, whereas dexmedetomidine pretreatment reversed the elevated activities of MDA and MPO in the Dex group. Furthermore, dexmedetomidine pretreatment also improved the activities of SOD and CAT in rat lung tissues compared with rats with lung ischemia-reperfusion injury. In addition, dexmedetomidine pretreatment increased the expression levels of HIF-1α, p-Akt and HIF- in the Dex group when compared to those in the IR group. The mRNA expressions of HIF-1a, p-Akt, Caspase-3, and Caspase-9 in lung tissue of rats was significantly reduced after dexmedetomidine pretreatment. Rat lung ischemia-reperfusion can induce severe lung injury. Dexmedetomidine treatment can attenuate lung ischemia-reperfusion injury by activating the PI3K/Akt signaling pathway at the transcriptional level.

Low-intensity pulsed ultrasound pretreatment inhibits HMGB1 expression and attenuates lung ischemia-reperfusion injury in rats via the cholinergic anti-inflammatory pathway

To observe the effects of low-intensity pulsed ultrasound (LIPUS) pretreatment on pulmonary expression of high mobility group box-1 (HMGB1) in a rat model of lung ischemia-reperfusion (IR). Thirty-two male SpragueDawley rats weighing 250-300 g were randomly divided (n=8) into sham-operated group, lung IR group, LIPUS pretreatment group and pretreatment with α7-nicotinic cholinergic receptor (α7nAChR) antagonist group. In the sham-operated group, the left pulmonary hilum was dissociated without occlusion; in the other 3 groups, the left pulmonary hilum was occluded for 45 min followed by reperfusion for 180 min; LIPUS pretreatment for 30 min and intraperitoneal injection of methyllycaconitine (2 mg/kg), an α7nAChR antagonist, were administered before the operation. The wet/dry weight ratio (W/D) and pulmonary permeability index (LPI) of the lung tissue were measured, and the lung histopathology was observed and scored. The contents of interleukin-1 (IL-1) and IL-6 in the lung tissues were measured using ELISA, and the pulmonary expression of HMGB1 protein was detected using immunofluorescence assay and Western blotting. Compared with those in the sham-operated group, the W/D of the lung tissue, LPI, pathological scores, IL-1 and IL-6 contents in the lung tissue, and pulmonary HMGB1 expression all significantly increased in the other 3 groups (P &lt; 0.05). LIPUS preconditioning significantly lowered the W/D values, LPI, pathological score, IL-1 and IL-6 contents and HMGB1 expression in the lung tissues following lung IR, and these effects were significantly inhibited by administration of methyllycaconitine. LIPUS preconditioning can reduce lung IR injury possibly by activating α7nAChR-dependent cholinergic anti-inflammatory pathway to reduce lung tissue HMGB1 expression.

Adiponectin protects against lung ischemia-reperfusion injury in rats with type 2 diabetes mellitus.

Adiponectin (APN) has been associated with the pathogenesis of acute brain, liver and heart injury. However, the role of APN in lung ischemia-reperfusion injury (LIRI) in diabetes mellitus remains unclear. To investigate this, the present study evaluated the effects of APN on lung dysfunction and pathological alterations in rats with type 2 diabetes mellitus via lung ischemia/reperfusion (I/R). The lung-protective effects of APN globular domain (gAPN) in rats with type 2 diabetes mellitus were also investigated by measuring the oxygenation index, inflammatory cytokines, lung edema, histopathology, oxidative stress, apoptosis and the protein expression levels of phosphorylated 5′adenosine monophosphate-activated protein kinase (p-AMPK), endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS). The results of the present study demonstrated that the diabetes mellitus rats + I/R (DIR) group exhibited greater concentrations of tumor necrosis factor-α and interleukin-6, and increases in the wet-weight to dry-weight ratio, lung injury score, oxidative stress and cellular apoptosis. These effects were accompanied by lower pulmonary oxygenation compared with the normal rat + I/R (NIR) group (P<0.05). Additionally, all of these alterations were attenuated in the NIR + gAPN and DIR + gAPN groups compared with in the NIR and DIR groups, respectively. In the DIR group, the expression levels of p-AMPK/AMPK and eNOS were significantly downregulated, and the levels of iNOS were upregulated, compared with those of the NIR group. Treatment with APN activated AMPK, increased eNOS expression and attenuated iNOS expression. The results of the present study demonstrated that APN exerted protective effects against LIRI via its anti-inflammatory, antioxidative stress and anti-apoptotic activities. These protective effects of APN were eliminated in rats with type 2 diabetes mellitus, in which LIRI was exacerbated. The present study indicated that APN may be a potential therapeutic agent for LIRI in type 2 diabetes mellitus.

Protective mechanism of cysteinyl aspartate-specific protease-3 inhibitors on lung ischemia-reperfusion injury in rats

Objective To investigate the protective effects of cysteinyl aspartate-specific protease (Caspase)-3 inhibitors on lung ischemia-reperfusion injury in rats. Methods 45 models of lung ischemia-reperfusion injury rats were randomly divided into model group, low dose group and high dose group.15 rats were selected as sham operation group. The rats in low dose group and the high dose group were injected with z-VAD-FMK, 5 mg/kg and 15 mg/kg by tail vein respectively and rats in model group and the sham operation group were injected with the normal saline. After 12 h treatment, the oxidative stress index [malondialdehyde (MDA), superoxide dismutase (SOD) and reactive oxygen species (ROS)] and inflammatory factors [interleukin (IL)-1β, IL-6 and tumor necrosis factor-α (TNF-α) ] were analyzed. The related protein levels of apoptosis [apoptosis inducing factor (AIF), Caspase-3, B cell lymphoma/leukemia-2 (bcl-2) and bcl-2 associated X protein (bax)] were analyzed by Western blotting. The level of apoptosis in lung tissue was analyzed by TdT-mediated dUTP nick end labeling (TUNEL) staining. Results Compared with the control group, the levels of MDA and ROS significantly increased in the model group, low-dose group and high dose group, while the level of SOD significantly decreased. Compared with the model group, the levels of MDA and ROS in the low dose group and the high dose group obviously decreased, while the level of SOD increased significantly. Compared with the sham group, TNF-α, IL-1β and IL-6 in model group, low dose group and high dose group significantly increased. Compared with model group, TNF-α, IL-1β and IL-6 in low dose group and high dose group significantly reduced. Compared with the sham operation group, AIF, Caspase-3 and bax protein level and cell apoptosis index in model group, low dose group and high dose group significantly increased and bcl-2 levels significantly increased; Compared with Sham group (3.43±0.67), cell apoptosis indexs in model group, low dose group and high dose group (25.32±3.21, 17.39±2.97 and 11.33±2.56) significantly increased (P=0.000). Compared with the model group, cell apoptosis indexs in low dose group and high dose groupsignificantly reduced (P=0.000). Compared with low dose group, cell apoptosis indexs in high dose groupsignificantly reduced (P=0.000). Conclusion Caspase-3 inhibitor z-VAD-FMK can significantly reduce oxidative stress, inflammation and lung apoptosis, and play a protective role in lung tissue. Key words: Cysteinyl aspartate-specific protease-3 inhibitors; Ischemia reperfusion injury; Oxidative stress; Inflammation; Apoptosis

Role of α2 adrenergic receptors in dexmedetomidine-induced inhibition of lipid peroxidation during lung ischemia-reperfusion injury in rats

Objective To evaluate the role of α2 adrenergic receptors in dexmedetomidine-induced inhibition of lipid peroxidation during lung ischemia-reperfusion (I/R) injury in rats. Methods Thirty-two isolated rat lungs in which the model of isolated lung perfusion was successfully established, were divided into 4 groups (n=8 each) using a random number table: control group (C group), I/R group, dexmedetomidine group (D group) and dexmedetomidine plus yohimbine group (DY group). The isolated lungs were subjected to 60 min of ischemia and apnea followed by 75 min of reperfusion and ventilation to establish the model of isolated lung I/R injury.From the beginning of reperfusion, 2.3 ng/ml dexmedetomidine was added to the perfusion fluid in D group, and 2.3 ng/ml dexmedetomidine and 0.4 μg/ml yohimbine (an α2 adrenergic receptor blocker) were added to the perfusion fluid in DY group.Lung specimens were obtained immediately after the end of reperfusion for determination of the wet/dry weight ratio (W/D ratio), superoxide dismutase (SOD) activity (by using modified pyrogallol autoxidation method) and malondialdehyde (MDA) content (by thiobarbituric acid method) and for examination of the pathological changes (using haematoxylin and eosin staining). Results Compared with C group, the W/D ratio and MDA content were significantly increased, and the SOD activity was decreased in I/R, D and DY groups (P<0.05). Compared with I/R group, the W/D ratio and MDA content were significantly decreased, and the SOD activity was increased in D group (P<0.05). Compared with DY group, the W/D ratio and MDA content were significantly decreased, and the SOD activity was increased in group D (P<0.05). The pathological changes of lung tissues were significantly attenuated in D group as compared with I/R and DY groups. Conclusion The mechanism by which dexmedetomidine inhibits lipid peroxidation is related to activating α2 adrenergic receptors during lung I/R injury in rats. Key words: Dexmedetomidine; Lung; Reperfusion injury; Receptors, adrenergic, alpha-2; Lipid peroxidation

Biliverdin protects the isolated lungs from ischemia/reperfusion injury via anti-apoptosis

To observe the effect of biliverdin (BV) on the lung ischemia/reperfusion injury (LIRI), and to investigate the mechanism of BV in treatment of LIRI. Thirty-two male Sprague-Dawley (SD) rats were randomly divided into control group, LIRI group, glutathione (GSH) group and BV-treated group, with 8 rats in each group. The rat LIRI model was reproduced by isolated lung perfusion system. Fifteen minutes after perfusion balance, lungs of control group were perfused continuously for 90 minutes, and the lungs in other groups were reperfused for 90 minutes after 1-hour ischemia, while the perfusion was added 10 μmol/L BV in BV-treated group and 4 mmol/L GSH in GSH group respectively. During the perfusion, the respiratory related indicators, such as tidal volume (VT), static compliance (Cst), arterial partial pressure of oxygen (PaO2), airway resistance (Raw), were dynamically observed. After perfusion, the wet/dry weight ratio (W/D) of lungs was determined. Pathological changes in lung tissue were observed under light microscope. The cell apoptosis was observed by TdT-mediated dUTP nick end labeling (TUNEL) method, and the apoptosis index was calculated. The protein expression levels of heme oxygenase-1 (HO-1), phosphorylated c-Jun N-terminal kinase (p-JNK), and caspase-3 were determined by Western Blot. Compared with control group, VT, Cst and PaO2 in LIRI group were significantly decreased from reperfusion for 30 minutes, and Raw was significantly increased. The pathological results showed that there was different degree of hyperemia edema, inflammatory cells infiltration and bronchial endothelium injury in the lung tissue of LIRI group. The W/D ratio of LIRI group was significantly higher than that of control group (8.98±2.34 vs. 5.89±0.52, P < 0.05). TUNEL results showed that the tan apoptosis cells of LIRI group were more than control group, and the apoptosis index was significantly higher than that of the control group [(13.88±2.35)% vs. (2.26±0.60)%, P < 0.05]. Compared with control group, the protein expression levels of HO-1, p-JNK, and caspase-3 in LIRI group were significantly increased [HO-1 (gray value): 0.55±0.13 vs. 0.16±0.02, p-JNK (gray value): 0.46±0.08 vs. 0.16±0.05, caspase-3 (gray value): 0.65±0.13 vs. 0.26±0.03, all P < 0.05]. Compared with LIRI group, the respiratory indicators in BV-treated group were improved significantly, the lung tissue injury was significantly reduced, the W/D ratio was significantly decreased (6.39±0.45 vs. 8.98±2.34, P < 0.05), and the cell apoptosis index was significantly reduced [(4.49±1.10)% vs. (13.88±2.35)%, P < 0.05], as well as the protein expression levels of HO-1, p-JNK, and caspase-3 were significantly lowered [HO-1 (gray value): 0.19±0.03 vs. 0.55±0.13, p-JNK (gray value): 0.31±0.06 vs. 0.46±0.08, caspase-3 (gray value): 0.33±0.05 vs. 0.65±0.13, all P < 0.05], which indicating that BV could alleviate LIRI via anti-apoptosis. The improvement effect of BV on PaO2, apoptosis index, protein expressions of HO-1 and caspase-3, and JNK phosphorylation were better than positive drug GSH, indicating that BV could protect LIRI ideally. BV alleviates LIRI via its anti-JNK pathway and its anti-apoptosis property.

Modulatory effect of silymarin on pulmonary vascular dysfunction through HIF-1α-iNOS following rat lung ischemia-reperfusion injury.

Silymarin is a traditional therapeutic used to protect the liver, acting to oppose lipid peroxidation, to enhance liver regeneration and functioning as an antioxidant. However, the effects of silymarin on pulmonary vascular dysfunction have not been investigated. In the present study, the modulatory effects of silymarin on pulmonary vascular dysfunction and the underlying mechanisms behind this were investigated in a lung ischemia-reperfusion (I/R) injury rat model. Male Sprague Dawley rats were randomly divided into 3 groups, including: i) A control group (n=10); ii) an I/R group (n=10); and iii) a silymarin-treated group (n=10). All experimental rats received 250 mg/kg/day of silymarin for 8 days. Silymarin was demonstrated to markedly improve lung I/R-induced pulmonary vascular dysfunction and lung moisture. Following silymarin treatment, inflammation and oxidative stress in the lung I/R-injury rats were demonstrably suppressed. Treatment with silymarin also inhibited the activation of caspase-3 and -9, and hypoxia inducible factor-1α (HIF-1α) and inducible nitric oxide synthase (iNOS) protein expression in the lung I/R-injury rats. Silymarin was concluded to impact upon pulmonary vascular dysfunction through the HIF-1α-iNOS pathway in the lung I/R injury rat model.

Role of mitochondrial calcium uniporter in lung ischemia-reperfusion injury in rats

Objective To evaluate the role of mitochondrial calcium uniporter in lung ischemia-reperfusion(I/R)injury in rats. Methods Thirty healthy male Wistar rats, weighing 250-300 g, were randomly divided into 3 groups(n=10 each)using a random number table: sham operation group(group S), group I/R and ruthenium red(mitochondrial calcium uniporter inhibitor)group(group RR). Lung I/R was produced by clamping the left pulmonary hilum for 45 min followed by 2 h reperfusion in I/R and RR groups.In addition, ruthenium red 2.5 mg/kg was injected via the tail vein at 1 h before occlusion of the left pulmonary hilum in RR group.Arterial blood samples were obtained at 2 h of reperfusion for blood gas analysis, and oxygenation index(PaO2/FiO2)was calculated.The rats were sacrificed, and then the lungs were removed for determination of wet/dry lung weight ratio(W/D ratio). Results Compared with group S, oxygenation index was significantly decreased, and W/D ratio was increased in I/R and RR groups.Oxygenation index was significantly higher, and W/D ratio was lower in group RR than in group I/R. Conclusion The mechanism underlying lung I/R injury is related to the opening of mitochondrial calcium uniporter in rats. Key words: Mitochondrial membrane transport proteins; Reperfusion injury; Lung

Role of endoplasmic reticulum stress in lung ischemia-reperfusion injury in rats

Objective To investigate the role of endoplasmic reticulum stress in lung ischemia-reperfusion (I/R) injury in rats. Methods Fifty-six healthy male Sprague-Dawley rats, weighing 250-320 g, were randomly divided into 2 groups using a random number table: sham operation group (group S, n=16) and I/R group (n=40). Lung I/R was induced by clamping the left hilum of lung for 60 min followed by reperfusion.Before ischemia (T0) and at 0, 1, 2 and 4 h (T4) of reperfusion, PaO2 and PaCO2 were recorded.The rats were sacrificed, and lungs were removed for examination of pathological changes.Wet/dry lung weight ratio (W/D ratio) was calculated.The expression of molecular chaperones of 78-kDa glucose-regulated protein (GRP78) and C/EBP homologous protein (CHOP) protein and mRNA was detected in lung tissues using Western blot and real-time PCR, respectively. Results Compared with group S, PaO2was significantly decreased, PaCO2 and W/D ratio were increased, and the expression of GRP78 and CHOP protein and mRNA was up-regulated during reperfusion, and the pathological changes of lungs were found in group I/R.With the prolongation of reperfusion, the pathological changes of lungs were gradually aggravated, and the expression of GRP78 and CHOP protein and mRNA was gradually up-regulated in group I/R. Conclusion Excessive endoplasmic reticulum stress may be one of the pathophysiological mechanisms of lung I/R injury in rats. Key words: Endoplasmic reticulum; Stress; Reperfusion injury; Lung