Chronic hypoxia preconditioning increases survival in rats suffering from heatstroke

1. In the present study, we assessed the protective effects of chronic hypoxia preconditioning against heatstroke-induced injury in urethane-anaesthetized rats. Heatstroke was induced by exposing the animals to an ambient temperature of 42 degrees C. The time at which both the mean arterial pressure (MAP) and local cerebral blood flow (CBF) in the striatum began to decrease from peak levels was taken as the onset of heatstroke. Control rats were exposed to a temperature of 24 degrees C. 2. Mean arterial pressure, CBF, blood pH, PaO2, PaCO2 and survival time (the interval between onset of heatstroke and cardiac arrest) after heat stress were all lower than in control rats (in which 'survival time' was defined as > 360 min). However, blood lactate concentrations were greater in rats exposed to heat. Rats placed at high altitude (HA), when exposed to the same heat stress (42 degrees C) survived much longer (113 +/- 26 min; n = 8) than rats maintained at sea level (SL; 20 +/- 2 min; n = 8). 3. After the onset of heatstroke, blood pH and lactate concentrations were found to be significantly higher and lower, respectively, in HA rats than in SL rats. 4. Western blot assay revealed that chronic hypoxia preconditioning induced heat shock protein (HSP) 72 expression in both the kidneys and lungs. 5. Thus, it appears that the observed benefit of chronic hypoxia preconditioning is related to attenuation of tissue acidification and elevations of HSP72 expression in both kidneys and lungs during heatstroke.     

黄芩素调控热休克蛋白70表达对溃疡性结肠炎大鼠的影响

目的研究黄芩素调控热休克蛋白70(HSP70)表达对溃疡性结肠炎大鼠的影响。方法用三硝基苯磺酸(TNBS)法构建溃疡性结肠炎大鼠模型,将建模成功的大鼠随机分为模型组、实验组及对照组,各15只,另外选取10只大鼠为正常组。实验组建模后第2天灌胃50 mg·kg^-1黄芩素,对照组灌胃0.42 g·kg^-1美沙拉嗪缓释颗粒,正常组与模型组则灌胃等量生理盐水,每天1次,连续干预2周。每天记录各组大鼠体重变化,进行疾病活动指数(DAI)评分;以苏木精-伊红染色观察大鼠结肠组织病理学形态;以免疫组化、蛋白质印迹法及实时荧光定量聚合酶链式反应检测结肠黏膜组织HSP70表达。结果治疗后,正常组、模型组、实验组和对照组大鼠DAI评分分别为0,(3.15±0.21),(0.89±0.14),(0.75±0.12)分,结肠黏膜组织损伤评分(CMIS)分别为(0.14±0.03),(2.86±0.23),(1.59±0.11),(1.35±0.10)分,结肠黏膜组织HSP70蛋白相对表达量分别为0.93±0.16,0.43±0.11,0.86±0.12,0.75±0.11,HSP70 mRNA相对表达量分别为1.32±0.23,0.21±0.06,0.78±0.11,0.65±0.08,差异均有统计学意义(均P<0.05)。结论黄芩素可有效减轻溃疡性结肠炎大鼠炎性反应及结肠黏膜组织炎性病理损伤,其作用机制可能与有效上调结肠黏膜组织HSP70表达相关。     

Protective effects of heat shock protein70 induced by geranylgeranylacetone in atrophic gastritis in rats

Aim： To investigate the effect of geranylgeranylacetone （GGA） on the progression of atrophic gastritis in rats and its potential mechanism. Methods： Atrophic gastritis was induced in Sprague-Dawley rats with 0.1% ammonia solution, 60% ethanol, and 20 mmol/L deoxycholic acid for 24 weeks. Accompanied by the induction of atrophic gastritis, 200 mg/kg GGA was administered by oral gavage for 8 weeks （weeks 17-24）. The histological changes in gastric mucosa were quantitated by the index of inflammation, the gastric mucosal thickness, and the amount of glands of 1 mm horizontal length in antrum. Endogenous heat shock protein （HSP）70 levels and distribution were determined by immunoblotting and immunohistochemistry in gastric mucosa. Results： GGA alleviated the pathological progression of atrophic gastritis with inflammation relief （inflammation index： 1.40 in the GGA group and 1.65 in the atrophic gastritis group） and glandular restoration （mucosal thickness and quantity of glands： 194.3 μn and 38.7 mm in the GGA group; 123.3 lain and 32.7 mm in the atrophic gastritis group; P〈0.05）. GGA significantly induced HSP70 synthesis （P〈0.05）. Moreover, quercetin, an inhibitor of HSP70 expression, aggravated the infiltration of inflammatory cells and glandular loss in the antrum. Conclusion： GGA prevented the progression of atrophic gastritis in rats via the induction of HSP70 expression.     

Elevated vascular γ‐butyrobetaine levels attenuate the development of high glucose‐induced endothelial dysfunction

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Norepinephrine‐induced diastolic dysfunction with aortic valve opening under calcium‐loading in rats

Heart failure associated with a high plasma level of norepinephrine (NE) has an extremely poor prognosis with NE being the most powerful predictor of all‐cause mortality. An increase in the diastolic intracellular calcium level (Ca²⁺) occurs in left ventricular dysfunction; however, the cause‐and‐effect relationships among Ca²⁺loading, high plasma NE, and an increase in diastolic ventricular pressure is unclear. Here, we examined the relationship between diastolic dysfunction and NE with and without Ca²⁺loading in rats. Animals were studied in four groups: Ca²⁺loading for 45 min (Ca²⁺group), NE alone for 25 min (30 µg/kg/min NE for 25 min; NE group), Ca²⁺loading and NE for 25 min after Ca²⁺loading (Ca²⁺‐NE group), and a vehicle group. Hemodynamics were examined using a micromanometer‐tipped pressure catheter, and diastolic function was studied using Doppler echocardiography. Significantly increased left ventricular end‐diastolic pressure (LVEDP) and decreased E and Ea waves and deceleration time (DCT) were found in the Ca²⁺‐NE group, compared with the Ca²⁺and NE groups. There were no changes in left ventricular pressure (LVP) and LV ejection fraction (EF) among the four groups. NE‐induced diastolic contracture (NEIDC) with aortic valve opening occurred in the diastole when LVP overshot the aortic pressure after co‐administration of NE and Ca²⁺after Ca²⁺loading, and pulmonary hemorrhage was observed in all animals of the Ca²⁺‐NE group. The results support the suggestion that NE may be an important factor in the development of diastolic dysfunction in ischemic heart disease. Drug Dev. Res. 67:511–518, 2006. © 2006 Wiley‐Liss, Inc.     

热休克蛋白70在内毒素血症大鼠心肌中的表达及已酮可可碱的影响

目的：探讨热休克蛋白70在内毒素血症大鼠心肌组织中的表达及已酮可可碱的影响。方法：采用直接注射内毒素的方法建立大鼠急性内毒素血症模型。18只Wistar大鼠随机分成3组，假手术组，内毒素组和PTX组。分别采用免疫组织化学法和免疫印迹法检测心肌组织HSP70蛋白。结果：内毒素组免疫组织化学法与免疫印迹法检测心肌组织HSP70蛋白水平与假手术组比较都明显增强（P〈0．01）；予PTX干预后，免疫组织化学法与免疫印迹法检测心肌组织HSP70蛋白水平与内毒素组比较也都明显增强（P〈0．01）。结论：内毒素血症大鼠心肌组织内的HSP70蛋白表达可明显增高，PTX干预可以进一步增高内毒素血症大鼠心肌组织内的HSP70蛋白表达，从而进一步加强对心肌的保护作用。     

Association between perfluorooctanoic acid exposure and degranulation of mast cells in allergic inflammation

Perfluorooctanoic acid (PFOA) has wide applications, including as a raw material for converted paper and packaging products. With the widespread use of PFOA, concerns regarding its potential environmental and health impacts have increased. In spite of the known hepatotoxicity and genotoxicity of PFOA, correlation with PFOA and allergic inflammation is not well known. In this study, the effect of PFOA on the degranulation of mast cells and mast cell‐mediated allergic inflammation in the presence of FcεRI cross‐linking was evaluated. In immunoglobulin (Ig) E‐stimulated mast cells, PFOA increased the release of histamine and β‐hexosaminidase by the up‐regulation of intracellular calcium levels. PFOA enhanced gene expression of several pro‐inflammatory cytokines, including tumor necrosis factor (TNF)‐α, interleukin (IL)‐1β, IL‐6, and IL‐8 by the activation of nuclear factor (NF)‐κB in IgE‐stimulated mast cells. Also, PFOA exacerbated allergic symptoms via hypothermia, and an increase of serum histamine, TNF‐α, IgE and IgG₁ in the ovalbumin‐induced systemic anaphylaxis. The present data indicate that PFOA aggravated FcɛRI‐mediated mast cell degranulation and allergic symptoms. Copyright © 2016 John Wiley & Sons, Ltd.     

Carbohydrate‐derived Fulvic acid (CHD‐FA) inhibits Carrageenan‐induced inflammation and enhances wound healing: efficacy and Toxicity study in rats

The objectives of this study were to evaluate the safety and anti‐inflammatory and wound‐healing characteristics of carbohydrate‐derived fulvic acid (CHD‐FA) in rats. CHD‐FA (≥100 mg/kg p.o.) effectively reduced carrageenan‐induced paw edema in rats, which was comparable to 10 mg/kg p.o. indomethacin. Topical application of CHD‐FA, formulated to contain 1.75% active product in a cetomicrogol cream at pH 1.98, compared favorably with fusidic acid cream (10 mg/g) in accelerating the healing of excised wounds infected with Staphylococcus aureus. No signs of toxicity were observed in rats during the 6‐day acute and 6‐month chronic treatment with CHD‐FA (100 mg/kg p.o.). Topical application of CHD‐FA, formulated in UEA cream and applied to the right ears of mice at 400 mg/g body weight on days 1 and 7–38, produced no adverse events. No signs of toxicity were observed in the teratogenicity study, in which CHD‐FA was administered at 100 mg/kg p.o. to pregnant female mice 3 days before fertilization to 14 days of pregnancy. In conclusion, CHD‐FA is a safe compound with anti‐inflammatory and wound‐healing properties and merits further evaluation in the treatment of patients suffering from similar conditions. Drug Dev Res 73: 18–23, 2012. © 2011 Wiley Periodicals, Inc.     

热应激和短暂心肌缺血对大鼠心肝组织热休克蛋白70表达的影响

目的探讨热应激及短暂心肌缺血对大鼠心、肝组织损伤及热休克蛋白（HSP70）表达的影响。方法24只雄性SD大鼠随机分为：对照组、假手术组、热休克组和短暂心肌缺血组,每组6只,应激后20h取心、肝组织,采用Western blotting、免疫组化及HE染色方法,观察大鼠心、肝组织的病理变化和HSP70的表达。结果（1）热应激及短暂心肌缺血后大鼠心、肝组织均有病理改变且HSP70表达与对照相比均显著增加（P〈0．05）;（2）短暂心肌缺血后心肌中HSP70表达量显著高于热应激后的表达量（P〈0．05）,而肝脏在两种刺激后HSP70表达量差异无统计学意义（P〉0．05）。结论短暂心肌缺血对心脏组织中HSP70表达的影响强于热应激,而对肝脏的影响两者无显著差异,这可能与不同刺激引起心、肝组织中HSP70表达机制不同有关。     

NF‐κB/p53‐activated inflammatory response involves in diquat‐induced mitochondrial dysfunction and apoptosis

Inflammation generated by environmental toxicants including pesticides could be one of the factors underlying neuronal cell damage in neurodegenerative diseases. In this study, we investigated the mechanisms by which inflammatory responses contribute to apoptosis in PC12 cells treated with diquat. We found that diquat induced apoptosis, as demonstrated by the activation of caspases and nuclear condensation, inhibition of mitochondrial complex I activity, and decreased ATP level in PC12 cells. Diquat also reduced the dopamine level, indicating that cell death induced by diquat is due to cytotoxicity of dopaminergic neuronal components in these cells. Exposure of PC12 cells to diquat led to the production of reactive oxygen species (ROS), and the antioxidant N‐acetyl‐cystein attenuated the cytotoxicity of caspase‐3 pathways. These results demonstrate that diquat‐induced apoptosis is involved in mitochondrial dysfunction through production of ROS. Furthermore, diquat increased expression of cyclooxygenase‐2 (COX‐2) and tumor necrosis factor‐α (TNF‐α) via inflammatory stimulation. Diquat induced nuclear accumulation of NF‐κB and p53 proteins. Importantly, an inhibitor of NF‐κB nuclear translocation blocked the increase of p53. Both NF‐κB and p53 inhibitors also blocked the diquat‐induced inflammatory response. Pretreatment of cells with meloxicam, a COX‐2 inhibitor, also blocked apoptosis and mitochondrial dysfunction. These results represent a unique molecular characterization of diquat‐induced cytotoxicity in PC12 cells. Our results demonstrate that diquat induces cell damage in part through inflammatory responses via NF‐κB‐mediated p53 signaling. This suggests the potential to generate mitochondrial damage via inflammatory responses and inflammatory stimulation‐related neurodegenerative disease.     

一氧化氮供体预处理对乳鼠心肌细胞的延迟保护作用及其机制

目的　探讨一氧化氮模拟预处理延迟保护作用的机制。方法　体外培养新生大鼠心肌细胞 ,实验分为 :①阴性对照组 (Normal组 ) ;②SNAP组 :一氧化氮 (nitricoxide ,NO)供体S 亚硝基 N 乙酰青霉胺 (S nitroso N acetyl 1 ,1 penicil lamine ,SNAP ,5 0 0 μmol·L-1 )与心肌细胞共育 2 4h ;③Che +SNAP组 :蛋白激酶C拮抗剂白屈菜季铵碱 (chelerythrinechlo ride ,Che ,1 0 μmol·L-1 )处理心肌细胞 30min后 ,加入SNAP与心肌细胞共育 2 4h ;④PDTC +SNAP组 :核因子κB(NF κB)特异性阻断剂PDTC(1 0 0 μmol·L-1 )处理心肌细胞 30min后 ,加入SNAP与心肌细胞共育 2 4h ;⑤缺氧复氧损伤组 (H/R组 ) :心肌细胞缺氧 6h ,复氧 3h。②～④组部分取细胞爬片以免疫组织化学法观察SNAP对热休克蛋白 70 (HSP70 )表达的影响 ;其余细胞经历H/R损伤后 ,检测心肌细胞存活率及乳酸脱氢酶 (LDH)活性。结果　在正常心肌细胞免疫组织化学法未检测到HSP70的表达 ,心肌细胞经过H/R损伤后 ,可检测到少量HSP70阳性细胞 ,其阳性染色A值为 94 6± 9 1 ,心肌细胞培养上清LDH活性为 (2 1 90 5± 1 5 1 7)U·L-1 ,细胞存活率为 5 1 7%± 4 6 % ,细胞损伤较正常组加重 (P <0 0 1 ) ;SNAP处理细胞后 2 4h ,HSP70阳性细胞数增多 ,     

大黄对肠缺血再灌注大鼠肠黏膜和通透性的影响及其机制

目的：观察肠缺血再灌注损伤后,肠内补充大黄对大鼠肠黏膜结构和通透性的影响,并探讨其作用机制。方法：雄性SD大鼠30只,随机分为3组（n=10）。大黄组：夹闭肠系膜上动脉建立肠缺血再灌注损伤模型,肠外营养＋肠内大黄液;模型组：建立肠缺血再灌注损伤模型,肠外营养;对照组：分离肠系膜上动脉但不夹闭,自由饮食。术后第3天收集尿液,采用双糖法（乳果糖/甘露醇）反应肠黏膜通透性,第6天处死大鼠留取小肠检测黏膜结构和热休克蛋白70（HSP70）含量。结果：对照组和大黄组肠黏膜通透性均低于模型组（P〈0.01）。大黄组肠黏膜通透性虽高于对照组,但无统计学意义（P〉0.05）;模型组小肠绒毛高度、黏膜厚度显著低于对照组和大黄组（P〈0.01）,大黄组接近于对照组（P〉0.05）;小肠全层厚度各组间无统计学差异;对照组和大黄组HSP70含量均高于模型组（P〈0.01）。大黄组HSP70含量虽略高于对照组,但差异无统计学意义（P〉0.05）。结论：大黄可减少肠缺血再灌注损伤引起的肠黏膜通透性增加,保护肠黏膜结构。大黄的肠道保护作用与多种机制有关,诱导小肠HSP70表达增加可能是其中之一。     

Effect of dimethylnitrosamine‐induced liver dysfunction on the pharmacokinetics of 5‐fluorouracil after administration of S‐1, an antitumour drug,to rats

Objectives The anti‐tumour agent S‐1 comprises tegafur (a prodrug of 5‐fluorouracil; 5‐FU), gimeracil (2‐chloro‐2,4‐dihydroxypyridine (CDHP); a competitive inhibitor of 5‐FU metabolism) and oteracil potassium. The effect of hepatic dysfunction induced by dimethylnitrosamine (DMN) on the pharmacokinetics of 5‐FU after administration of S‐1 to rats was investigated. Methods S‐1 (5 mg/kg) was administered intravenously and orally to rats with DMN‐induced liver dysfunction. Plasma concentrations of S‐1 components and 5‐FU were measured by HPLC and LC/MS‐MS. Blood tests and in‐vitro enzymatic investigations were also conducted. Key findings DMN treatment induced hepatic dysfunction and decreased the conversion of tegafur to 5‐FU in the liver without altering renal function or dihydropyrimidine dehydrogenase activity. Following intravenous administration of S‐1, the blood concentration‐time profiles of CDHP were similar between control rats and rats with hepatic dysfunction, but the half‐life of tegafur was significantly prolonged. The maximum plasma concentration (C_max) of 5‐FU was significantly reduced and the area under the blood concentration‐time curve (AUC) was reduced by 22%. Following oral administration, the C_max of tegafur, 5‐FU and CDHP were significantly decreased and half‐lives significantly increased. Hepatic dysfunction had a less pronounced effect on the AUC of 5‐FU (13.6% reduction). Conclusions The pharmacokinetic profiles of tegafur, 5‐FU and CDHP were altered by changes in the elimination rate of tegafur induced by a decrease in the conversion of tegafur to 5‐FU. However, hepatic dysfunction had less of an effect on the AUC of 5‐FU, which correlates with anti‐tumour effect, after the oral administration of S‐1.     

Curcumin ameliorates asthmatic airway inflammation by activating nuclear factor‐E2‐related factor 2/haem oxygenase (HO)‐1 signalling pathway

Previous studies have shown that curcumin alleviates asthma in vivo. However, the relationship between curcumin and the nuclear factor‐E2‐related factor 2 (Nrf2)/haem oxygenase (HO)‐1 pathway in asthma treatment remains unknown. The aim of the present study was to investigate the mechanisms of curcumin involved in the amelioration of airway inflammation in a mouse asthma model. Curcumin was administrated to asthmatic mice, and bronchoalveolar lavage fluid was collected. Inflammatory cell infiltration was measured by Giemsa staining. Immunoglobulin E production in bronchoalveolar lavage fluid was measured by enzyme–linked immunosorbent assay. Histological analyses were evaluated with haematoxylin‐eosin and periodic acid‐Schiff staining. Airway hyperresponsiveness was examined by whole‐body plethysmography. Nuclear factor‐E2‐related factor 2, HO‐1, nuclear factor‐κB and inhibitory κB/p‐inhibitory κB levels in lung tissues were detected by western blot, and Nrf2 activity was measured by electrophoretic mobility shift assay. Tumour necrosis factor‐α, interleukin (IL)‐1β, and IL‐6 levels in the small interfering RNA‐transfected cells were detected by enzyme–linked immunosorbent assay. Curcumin treatment significantly reduced immunoglobulin E production, attenuated inflammatory cell accumulation and goblet cell hyperplasia, and ameliorated mucus secretion and airway hyperresponsiveness. Nuclear factor‐E2‐related factor 2 and HO‐1 levels in lung tissues were significantly increased. Meanwhile, Nrf2 activity was enhanced. Nuclear factor‐κB and p‐inhibitory κB levels were elevated in the lung tissue of ovalbumin‐challenged mice. Both were restored to normal levels after curcumin treatment. Haem oxygenase‐1 and nuclear Nrf2 levels were enhanced in dose‐ and time‐dependent manners in curcumin‐treated RAW264.7 cells. Curcumin blocked lipopolysaccharide‐upregulated expression of tumour necrosis factor‐α, IL‐1β, and IL‐6. After the cells were transfected with HO‐1 or Nrf2 small interfering RNA, lipopolysaccharide‐induced pro‐inflammation cytokine expression was significantly restored. In summary, curcumin might alleviate airway inflammation in asthma through the Nrf2/HO‐1 pathway, potentially making it an effective drug in asthma treatment.     

Overexpressed Hsp70 alleviated formaldehyde‐induced apoptosis partly via PI3K/Akt signaling pathway in human bronchial epithelial cells

Formaldehyde (FA) is a ubiquitous environmental pollutant, which can induce apoptosis in lung cell and is related to the pathogenesis of asthma, pneumonia, and chronic obstructive pulmonary disease. Heat shock protein 70 (Hsp70) is an ATP‐dependent molecular chaperone and exhibits an anti‐apoptosis ability in a variety of cells. Previous studies reported that the expression of Hsp70 was induced when organisms were exposed to FA. Whether Hsp70 plays a role in the FA‐induced apoptosis and the involved cell signaling pathway remain largely unknown. In this study, human bronchial epithelial cells with overexpressed Hsp70 and the control were exposed to different concentrations of FA (0, 40, 80, and 160 μmol/L) for 24 hours. Apoptosis and the expression levels of PI3K, Akt, p‐Akt, MEK, p‐MEK, and GLI2 were detected by Annexin‐APC/7AAD double‐labeled flow cytometry and western blot. The results showed that overexpression of Hsp70 decreased the apoptosis induced by FA and alleviated the decline of PI3k and p‐Akt significantly. Inhibitor (LY 294002, a specific inhibitor of PI3K‐Akt) test result indicated that PI3K‐Akt signaling pathway was involved in the inhibition of FA‐induced apoptosis by Hsp70 overexpression and also active in the maintenance of GLI2 level. However, it also suggested that other signaling pathways activated by overexpressed Hsp70 participated in this process, which was needed to be elucidated in further research.     

Chronic activation of AMP-activated protein kinase prevents 20-hydroxyeicosatetraenoic acid-induced endothelial dysfunction

1. 20-Hydroxyeicosatetraenoic acid (20-HETE) is a potent vasoconstrictor involved in vascular dysfunction and blood pressure regulation. Studies have revealed strong associations between 20-HETE and endothelial dysfunction; however, the signalling mechanisms are largely unknown. Therefore, the aim of the present study was to investigate the effect of 20-HETE on the association between endothelial nitric oxide synthase (eNOS) and heat shock protein 90 (Hsp90). 2. In mouse aortic rings, 20-HETE significantly enhanced the constriction to phenylephrine and inhibited the relaxation to acetylcholine (P=0.05 vs control rings). In mice with chronic AMP-activated protein kinase (AMPK) activation, this protected against the negative effects of 20-HETE (P<0.05). Immunoprecipitation of eNOS in human umbilical vein endothelial cells treated with 20-HETE revealed a decrease in basal and vascular endothelial growth factor-stimulated Hsp90 association with eNOS (P<0.05). Pretreatment of cells with 5'-aminoimidazole-4-carboxyamide-ribonucleoside (AICAR; a chronic activator of AMPK) prevented the loss of Hsp90 association with eNOS following 20-HETE treatment. Treatment with 20-HETE for 24 h induced an increase in eNOS phosphorylation that was not seen following acute treatment (30 min). The increased eNOS phosphorylation was accompanied by transient changes in Akt phosphorylation. 3. In conclusion, 20-HETE impairs eNOS-Hsp90 association, which can be reversed by chronic activation of AMPK. This provides a mechanism for reduced nitric oxide bioactivity and endothelial dysfunction in diseases with elevated 20-HETE levels, such as hypertension.