脂多糖致心肌细胞损伤的Kir6.2机制
摘要
感染性休克也称败血症性休克或中毒性休克;是由病原微生物及其毒素在人体引起的一种微循环障碍状态,致组织缺氧、代谢紊乱、细胞损害甚至多器官功能衰竭;是一严重危害人类健康的综合征群,是外科危重病治疗中最常见的死亡原因,其死亡率高达30%-50%。众所周知,感染性休克时冠状动脉灌注不足,心肌缺血缺氧;心肌纤维变性、坏死或断裂、间质水肿、心肌收缩力减弱;脂多糖(lipopolysaccharide,LPS)可损伤心肌线粒体、肌浆网、肌膜、收缩蛋白,亚细胞结构发生改变;三磷酸腺苷(ATP)产生和利用障碍,最终引发心力衰竭。ATP敏感钾通道(ATP-sensitive potassium channels, KATP)在心血管系统广泛存在,是一个大分子复合体,其分子组成是SUR和Kir6.x。SUR是受体单位,是钾离子通道激动剂或拮抗剂作用的靶点;Kir6.x是内向整流钾离子通道单位,其开放与否受SUR调节。在心肌细胞,细胞膜组成是Kir6.2/SUR2A;线粒体膜组成是Kir6.1/SUR1。研究认为开放KATP对于心肌缺血再灌注损伤具有保护作用;有趣的是SUR2基因缺失小鼠却具有抗缺血再灌注损伤作用;同样令人吃惊的是SUR1基因缺失小鼠也具有抗缺血再灌注损伤作用。但是,关于Kir6.2在LPS休克或血症对心肌细胞的影响未见报道。鉴于此,我们拟研究内毒素血症心肌细胞细胞损伤的Kir6.2机制。本课题主要研究内容包括:
第一部分,整体动物水平研究Kir6.2对LPS心肌细胞损伤和凋亡的影响整体动物水平,以C57小鼠(WT)和Kir6.2基因敲除小鼠(Kir6.2-KO)为模型,两种动物均分为:生理盐水对照组和LPS刺激90min、180min、360min组,共8组。利用HE染色技术检测各组心肌细胞组织病理学变化;利用透射电子显微镜(Transmission Electron Microscopy ,TEM)技术检测各组心肌细胞超微结构变化;利用TUNEL法检测各组心肌细胞凋亡程度。发现对照组WT小鼠和Kir6.2-KO小鼠心肌细胞组织结构正常,超微结构正常,心肌细胞有轻微凋亡;在LPS刺激90min、180min、360min时间点,WT小鼠和Kir6.2-KO小鼠心肌细胞均出现了明显的组织病理学改变,超微结构改变和细胞凋亡。但是各时间点Kir6.2-KO小鼠心肌细胞的组织病理学改变、超微结构改变和凋亡程度均较WT小鼠明显。提示:Kir6.2对LPS心肌细胞损伤具有保护作用,具有抗LPS心肌细胞凋亡的作用。
第二部分,研究Kir6.2对炎症因子表达水平的影响
一、整体动物水平以C57小鼠(WT)和Kir6.2基因敲除小鼠(Kir6.2-KO)为模型,两种动物均分为:生理盐水对照组和LPS刺激90min、180min、360min组,共8组。利用酶联免疫吸附测定(ELISA)技术检测各组动物的血清TNF-α、IL-1β表达水平。生理盐水组未检测到血清炎症因子表达,LPS刺激90min后,炎症因子表达最高,WT小鼠和Kir6.2-KO小鼠之间TNF-α和IL-1β的表达没有显著性差异;LPS刺激180min后,炎症因子表达有所下降,WT小鼠和Kir6.2-KO小鼠之间TNF-α的表达没有显著性差异,WT小鼠IL-1β的表达显著高于Kir6.2-KO小鼠;LPS刺激360min后,炎症因子表达明显下降,WT小鼠TNF-α和IL-1β的表达都显著低于Kir6.2-KO小鼠。结果表明Kir6.2影响小鼠血清炎症因子的表达。提示:Kir6.2缺失使血清炎症因子的表达增加。
二、细胞水平以C57小鼠(WT)和Kir6.2基因敲除小鼠(Kir6.2-KO)为模型,以小鼠腹腔来源的原代巨噬细胞为研究对象,利用酶联免疫吸附测定(ELISA)技术检测细胞上清液的TNF-α表达水平。利用C57小鼠来源的腹腔巨噬细胞检测了生理盐水对照组和LPS刺激30min、60min、90min、180min和360min后各时间点的炎症因子表达水平。发现:在LPS刺激60min之前,炎症因子几乎无表达;在LPS刺激90min后,炎症因子开始大量表达;在LPS刺激180min后,炎症因子表达速率最大;360min的表达速率已降至较低水平。所以我们选取90min时间点研究Kir6.2不同功能状态时内毒素诱导的巨噬细胞炎症因子表达差异。发现:KATP开放剂吡那地尔显著降低WT小鼠腹腔巨噬细胞炎症因子的表达,KATP阻断剂格列本脲显著升高WT小鼠腹腔巨噬细胞炎症因子的表达。提示:激活KATP降低巨噬细胞炎症因子的表达;反之,阻断KATP增加巨噬细胞炎症因子的表达。对Kir6.2-KO小鼠来源的巨噬细胞炎症因子表达的影响,吡那地尔和格列本脲显示了类似的作用。提示:除Kir6.2外,Kir6.1也参与了KATP对内毒素诱导的巨噬细胞炎症因子表达的影响。
第三部分,研究Kir6.2和α7nAChR抗炎通路的相互影响。
细胞水平以C57小鼠(WT)和Kir6.2基因敲除小鼠(Kir6.2-KO)为模型,以小鼠腹腔来源的原代巨噬细胞为研究对象,利用酶联免疫吸附测定(ELISA)技术检测细胞上清液的TNF-α表达水平。发现:在WT小鼠和Kir6.2-KO小鼠组,PNU-282987均能显著降低巨噬细胞炎症因子的表达水平。提示:Kir6.2对胆碱能抗炎通路并无影响。
本课题围绕KATP的Kir6.2亚单位,在整体动物及细胞水平对内毒素心肌细胞损伤进行了探索。发现:LPS致心肌细胞损伤,与WT小鼠比较,Kir6.2基因敲除小鼠心肌细胞组织损伤加重,血清炎症因子表达增加;开放巨噬细胞KATP可使LPS诱导的炎症因子表达水平降低,阻断巨噬细胞KATP可使LPS诱导的炎症因子表达水平增高。结论:KATP亚单位Kir6.2在LPS致心肌细胞损伤过程中具有重要作用;巨噬细胞Kir6.2缺失使LPS诱导的炎症因子表达水平增高,参与了LPS心肌细胞损伤过程。
Severe sepsis, the leading cause of fatality in critically ill patients, is characterized by an immunologically driven systemic response that underlies host protection against microbial infection.Central to this syndrome is a pronounced cytokine-induced cardiovascular state defined by a hypercontractile heart and altered vascular tone. Despite diverse infectious pathogens, the outcome in severe sepsis is governed by the nature and degree of the host response, with the cardiovascular status an ultimate determinant of survival. The Gram-negative bacterial cell wall endotoxin, LPS (lipopolysaccharide), evokes a surge in tumor necrosis factor-alpha (TNF-α), precipitating an inflammatory mediated cascade of pathophysiologic events that imposes a high demand on the hyperdynamic septic heart.
ATP-sensitive potassium (KATP) channels have emerged as protein complexes with the capability to decode signals of metabolic distress. By virtue of an intimate integration with intracellular energetic networks and a capacity for high-fidelity processing of incoming metabolic signals, KATP channels regulate cellular excitability-dependent functions. These channels are situated in high density within metabolically active tissues, where changes in the energetic state are conveyed through the ATP-binding cassette sulfonylurea receptor (SUR) regulatory subunit to the K+ channel pore, Kir6.x of the channel complex. It is the tissue-dependent channel pore that, requiring the regulatory SUR chaperone to function, adjusts membrane potential to match demand and maintain cellular wellbeing. In the vasculature, the pore-forming subunit of the KATP channel has been identified as Kir6.1, and Kir6.1-containing channels have been implicated in the regulation of arterial smooth muscle tone. In the myocardium, where the pore-forming isoform is Kir6.2, defects in KATP channel subunits jeopardize cellular stress tolerance and predispose to injury.
Recent studies indicate that the nervous system, through vagus nerve, can modulate circulating TNF-α, IL-1βa?nd IL-6 levels induced by endotoxin. This new mechanism termed“cholinergic anti-inflammatory pathway”is based on the release of acetylcholine (ACh), the principal neurotransmitter of the vagus nerve that inhibits the production of pro-inflammatory cytokines via itsα7 nicotinic acetylcholine receptor (α7nAChR) in resident tissue macrophages. Nicotine receptors belong to the family of ligand-gated ion channels and consist of twelve subunits, nineαand threeβknown to exist so far. Alpha7 nicotinic acetylcholine receptor (α7nAChR) is a type of nicotinic acetylcholine receptor, constructed by five homomericα7 subunits. This receptor, most abundantly distributing in mammalian brain, has been demonstrated to control excitability and neurotransmitter release and to mediate neuroprotective properties. Recent studies indicated thatα7nAChR expressed on macrophages played an important role in the cholinergic anti-inflammatory pathway. During acute condition,α7nAChR attenuated renal failure induced by ischemia/reperfusion through inhibiting pro-inflammatory cytokines expression, and subsequently decreasing cell apoptosis. During chronic inflammation process,α7nAChR was related to the pathogenesis of many diseases such as allergies, Alzheimer’s disease, diabetes, hypertension, and hormonal imbalances. Therefore as a potential therapeutic target against inflammation and cognitive disorder, selective agonists forα7nAChR attract a great deal of attention.
The first aim of this thesis is to investigate the role of Kir6.2 subunit of ATP-sensitive potassium channels in myocardial damage induced by lipopolysaccharide. Thus, light microscopy was performed on paraffin-embedded myocardial sections stained with hematoxylin-eosin from 4% formalin-fixed left ventricles (LV) taken from WT and Kir6.2-KO mice 90min,180min and 360min after LPS or saline vehicle administration. And TUNEL staining was performed on these sections. Transmitted electron microscopy (EM) was performed on ultramicrotome-cut, lead citrate-stained LV sections with a JEOL 1200 EXII electron microscope. All the results showed that the myocardial damage in LPS-challenged Kir6.2-KO mice were aggravated compared to LPS-challenged WT mice.
We postulated that at least there were two major factors involved in myocardial damage in LPS-challenged mice. One might be the direct injury from LPS, and the other was the inflammatory impairment from cytokines, such as TNF-αinduced by LPS. Here we assessed the possible inflammatory impairment from cytokines induced by LPS.
To assess whether Kir6.2 was able to modulate cytokines expression induced by LPS, Serum cytokine levels were quantified at baseline, 90,180 min and 360min after LPS administration by ELISA. Expression of TNF-αincreased significantly at 360min after LPS. The results suggested that Kir6.2 decreased the expression of cytokines. Meanwhile, cytokine levels in peritoneal macrophages from WT mice and Kir6.2-KO mice were quantified at baseline and 90min after LPS administration by ELISA. Glibenclamide significantly increased TNF-αexpression in peritoneal macrophages from WT mice. And pinacidil significantly decreased TNF-αexpression in peritoneal macrophages from WT mice. These changes were also found in peritoneal macrophages from Kir6.2-KO mice. These results suggested that both Kir6.2 and Kir6.1 were involved in the regulation of the cytokines expression.
The second aim of this study was to investigate interaction between Kir6.2 andα7nAChR. Cytokine levels in peritoneal macrophages from WT mice and Kir6.2-KO mice were quantified at baseline and 90min after LPS administration by ELISA. The selectiveα7nAChR agonist PNU-282987 significantly inhibited TNF-αexpression in peritoneal macrophages from WT mice. PNU- 282987 also significantly decreased TNF-αexpression in peritoneal macrophages from Kir6.2-KO mice. These suggested that Kir6.2 might not involve in the cholinergic anti-inflammatory pathway.
第一部分,整体动物水平研究Kir6.2对LPS心肌细胞损伤和凋亡的影响整体动物水平,以C57小鼠(WT)和Kir6.2基因敲除小鼠(Kir6.2-KO)为模型,两种动物均分为:生理盐水对照组和LPS刺激90min、180min、360min组,共8组。利用HE染色技术检测各组心肌细胞组织病理学变化;利用透射电子显微镜(Transmission Electron Microscopy ,TEM)技术检测各组心肌细胞超微结构变化;利用TUNEL法检测各组心肌细胞凋亡程度。发现对照组WT小鼠和Kir6.2-KO小鼠心肌细胞组织结构正常,超微结构正常,心肌细胞有轻微凋亡;在LPS刺激90min、180min、360min时间点,WT小鼠和Kir6.2-KO小鼠心肌细胞均出现了明显的组织病理学改变,超微结构改变和细胞凋亡。但是各时间点Kir6.2-KO小鼠心肌细胞的组织病理学改变、超微结构改变和凋亡程度均较WT小鼠明显。提示:Kir6.2对LPS心肌细胞损伤具有保护作用,具有抗LPS心肌细胞凋亡的作用。
第二部分,研究Kir6.2对炎症因子表达水平的影响
一、整体动物水平以C57小鼠(WT)和Kir6.2基因敲除小鼠(Kir6.2-KO)为模型,两种动物均分为:生理盐水对照组和LPS刺激90min、180min、360min组,共8组。利用酶联免疫吸附测定(ELISA)技术检测各组动物的血清TNF-α、IL-1β表达水平。生理盐水组未检测到血清炎症因子表达,LPS刺激90min后,炎症因子表达最高,WT小鼠和Kir6.2-KO小鼠之间TNF-α和IL-1β的表达没有显著性差异;LPS刺激180min后,炎症因子表达有所下降,WT小鼠和Kir6.2-KO小鼠之间TNF-α的表达没有显著性差异,WT小鼠IL-1β的表达显著高于Kir6.2-KO小鼠;LPS刺激360min后,炎症因子表达明显下降,WT小鼠TNF-α和IL-1β的表达都显著低于Kir6.2-KO小鼠。结果表明Kir6.2影响小鼠血清炎症因子的表达。提示:Kir6.2缺失使血清炎症因子的表达增加。
二、细胞水平以C57小鼠(WT)和Kir6.2基因敲除小鼠(Kir6.2-KO)为模型,以小鼠腹腔来源的原代巨噬细胞为研究对象,利用酶联免疫吸附测定(ELISA)技术检测细胞上清液的TNF-α表达水平。利用C57小鼠来源的腹腔巨噬细胞检测了生理盐水对照组和LPS刺激30min、60min、90min、180min和360min后各时间点的炎症因子表达水平。发现:在LPS刺激60min之前,炎症因子几乎无表达;在LPS刺激90min后,炎症因子开始大量表达;在LPS刺激180min后,炎症因子表达速率最大;360min的表达速率已降至较低水平。所以我们选取90min时间点研究Kir6.2不同功能状态时内毒素诱导的巨噬细胞炎症因子表达差异。发现:KATP开放剂吡那地尔显著降低WT小鼠腹腔巨噬细胞炎症因子的表达,KATP阻断剂格列本脲显著升高WT小鼠腹腔巨噬细胞炎症因子的表达。提示:激活KATP降低巨噬细胞炎症因子的表达;反之,阻断KATP增加巨噬细胞炎症因子的表达。对Kir6.2-KO小鼠来源的巨噬细胞炎症因子表达的影响,吡那地尔和格列本脲显示了类似的作用。提示:除Kir6.2外,Kir6.1也参与了KATP对内毒素诱导的巨噬细胞炎症因子表达的影响。
第三部分,研究Kir6.2和α7nAChR抗炎通路的相互影响。
细胞水平以C57小鼠(WT)和Kir6.2基因敲除小鼠(Kir6.2-KO)为模型,以小鼠腹腔来源的原代巨噬细胞为研究对象,利用酶联免疫吸附测定(ELISA)技术检测细胞上清液的TNF-α表达水平。发现:在WT小鼠和Kir6.2-KO小鼠组,PNU-282987均能显著降低巨噬细胞炎症因子的表达水平。提示:Kir6.2对胆碱能抗炎通路并无影响。
本课题围绕KATP的Kir6.2亚单位,在整体动物及细胞水平对内毒素心肌细胞损伤进行了探索。发现:LPS致心肌细胞损伤,与WT小鼠比较,Kir6.2基因敲除小鼠心肌细胞组织损伤加重,血清炎症因子表达增加;开放巨噬细胞KATP可使LPS诱导的炎症因子表达水平降低,阻断巨噬细胞KATP可使LPS诱导的炎症因子表达水平增高。结论:KATP亚单位Kir6.2在LPS致心肌细胞损伤过程中具有重要作用;巨噬细胞Kir6.2缺失使LPS诱导的炎症因子表达水平增高,参与了LPS心肌细胞损伤过程。
Severe sepsis, the leading cause of fatality in critically ill patients, is characterized by an immunologically driven systemic response that underlies host protection against microbial infection.Central to this syndrome is a pronounced cytokine-induced cardiovascular state defined by a hypercontractile heart and altered vascular tone. Despite diverse infectious pathogens, the outcome in severe sepsis is governed by the nature and degree of the host response, with the cardiovascular status an ultimate determinant of survival. The Gram-negative bacterial cell wall endotoxin, LPS (lipopolysaccharide), evokes a surge in tumor necrosis factor-alpha (TNF-α), precipitating an inflammatory mediated cascade of pathophysiologic events that imposes a high demand on the hyperdynamic septic heart.
ATP-sensitive potassium (KATP) channels have emerged as protein complexes with the capability to decode signals of metabolic distress. By virtue of an intimate integration with intracellular energetic networks and a capacity for high-fidelity processing of incoming metabolic signals, KATP channels regulate cellular excitability-dependent functions. These channels are situated in high density within metabolically active tissues, where changes in the energetic state are conveyed through the ATP-binding cassette sulfonylurea receptor (SUR) regulatory subunit to the K+ channel pore, Kir6.x of the channel complex. It is the tissue-dependent channel pore that, requiring the regulatory SUR chaperone to function, adjusts membrane potential to match demand and maintain cellular wellbeing. In the vasculature, the pore-forming subunit of the KATP channel has been identified as Kir6.1, and Kir6.1-containing channels have been implicated in the regulation of arterial smooth muscle tone. In the myocardium, where the pore-forming isoform is Kir6.2, defects in KATP channel subunits jeopardize cellular stress tolerance and predispose to injury.
Recent studies indicate that the nervous system, through vagus nerve, can modulate circulating TNF-α, IL-1βa?nd IL-6 levels induced by endotoxin. This new mechanism termed“cholinergic anti-inflammatory pathway”is based on the release of acetylcholine (ACh), the principal neurotransmitter of the vagus nerve that inhibits the production of pro-inflammatory cytokines via itsα7 nicotinic acetylcholine receptor (α7nAChR) in resident tissue macrophages. Nicotine receptors belong to the family of ligand-gated ion channels and consist of twelve subunits, nineαand threeβknown to exist so far. Alpha7 nicotinic acetylcholine receptor (α7nAChR) is a type of nicotinic acetylcholine receptor, constructed by five homomericα7 subunits. This receptor, most abundantly distributing in mammalian brain, has been demonstrated to control excitability and neurotransmitter release and to mediate neuroprotective properties. Recent studies indicated thatα7nAChR expressed on macrophages played an important role in the cholinergic anti-inflammatory pathway. During acute condition,α7nAChR attenuated renal failure induced by ischemia/reperfusion through inhibiting pro-inflammatory cytokines expression, and subsequently decreasing cell apoptosis. During chronic inflammation process,α7nAChR was related to the pathogenesis of many diseases such as allergies, Alzheimer’s disease, diabetes, hypertension, and hormonal imbalances. Therefore as a potential therapeutic target against inflammation and cognitive disorder, selective agonists forα7nAChR attract a great deal of attention.
The first aim of this thesis is to investigate the role of Kir6.2 subunit of ATP-sensitive potassium channels in myocardial damage induced by lipopolysaccharide. Thus, light microscopy was performed on paraffin-embedded myocardial sections stained with hematoxylin-eosin from 4% formalin-fixed left ventricles (LV) taken from WT and Kir6.2-KO mice 90min,180min and 360min after LPS or saline vehicle administration. And TUNEL staining was performed on these sections. Transmitted electron microscopy (EM) was performed on ultramicrotome-cut, lead citrate-stained LV sections with a JEOL 1200 EXII electron microscope. All the results showed that the myocardial damage in LPS-challenged Kir6.2-KO mice were aggravated compared to LPS-challenged WT mice.
We postulated that at least there were two major factors involved in myocardial damage in LPS-challenged mice. One might be the direct injury from LPS, and the other was the inflammatory impairment from cytokines, such as TNF-αinduced by LPS. Here we assessed the possible inflammatory impairment from cytokines induced by LPS.
To assess whether Kir6.2 was able to modulate cytokines expression induced by LPS, Serum cytokine levels were quantified at baseline, 90,180 min and 360min after LPS administration by ELISA. Expression of TNF-αincreased significantly at 360min after LPS. The results suggested that Kir6.2 decreased the expression of cytokines. Meanwhile, cytokine levels in peritoneal macrophages from WT mice and Kir6.2-KO mice were quantified at baseline and 90min after LPS administration by ELISA. Glibenclamide significantly increased TNF-αexpression in peritoneal macrophages from WT mice. And pinacidil significantly decreased TNF-αexpression in peritoneal macrophages from WT mice. These changes were also found in peritoneal macrophages from Kir6.2-KO mice. These results suggested that both Kir6.2 and Kir6.1 were involved in the regulation of the cytokines expression.
The second aim of this study was to investigate interaction between Kir6.2 andα7nAChR. Cytokine levels in peritoneal macrophages from WT mice and Kir6.2-KO mice were quantified at baseline and 90min after LPS administration by ELISA. The selectiveα7nAChR agonist PNU-282987 significantly inhibited TNF-αexpression in peritoneal macrophages from WT mice. PNU- 282987 also significantly decreased TNF-αexpression in peritoneal macrophages from Kir6.2-KO mice. These suggested that Kir6.2 might not involve in the cholinergic anti-inflammatory pathway.
引文
[1] Sands KE.Epidemiology of sepsis syndrome in 8 academic medical centers. JAMA. 1997,278:234-40
[2] Ulloa L, Tracey K J. The‘cytokine profile’: a code for sepsis. Trends Mol Med 2005,11: 56–63
[3] Riedemann NC, Guo RF, Ward PA. Novel strategies for the treatment of sepsis. Nature Med, 2003,9: 517–524
[4] Tracey KJ, Cerami A. Tumor necrosis factor: a pleiotropic cytokine and therapeutic target. Annu Rev Med,1994,45:491–503
[5] Tracey KJ, Cerami A. Tumor necrosis factor, other cytokines and disease. Annu Rev Cell Biol,1993,9:317–343
[6] Van der Poll T, Lowry SF. Tumor necrosis factor in sepsis: mediator of multiple organ failure or essential part of host defense? Shock,1995; 3:1–12
[7] Bryan J, Aguilar-Bryan L. Sulfonylurea receptors: ABC transporters that regulate ATP-sensitive K+ channels. Biochim Biophys Acta,1999,1461:285–305
[8] Seino S, Miki T. Physiological and pathophysiological roles of ATP-sensitive K+ channels. Prog Biophys Mol Biol,2003,81:133–76
[9] Rodrigo GC, Standen NB. ATP-sensitive potassium channels. Curr Pharm Des,2005,11:1915–40
[10] Wilson AJ, Jabr RI, Clapp LH. Calcium modulation of vascular smooth muscle ATP-sensitive K+ channels: Role of protein phosphatase-2B. Circ Res,2000,87:1019–25
[11] Orie NN, Perrino BA, Tinker A, Clapp LH. Calcineurin Aαregulates Kir6.1/SUR2B in HEK293 cells through an interaction with protein kinase A. J Physiol,2005,567P:C24
[12] Miki T, Seino S. Roles of KATP channels as metabolic sensors in acute metabolic changes. J Mol Cell Cardiol 2005,38:917–25
[13] Landry DW, Oliver JA. The ATP-sensitive K+ channel mediates hypotension in endotoxemia and hypoxic lactic acidosis in dog. J Clin Invest,1992,89:2071–4
[14] Vanelli G, Hussain SN, Dimori M, Aguggini G. Cardiovascular responses to glibenclamide during endotoxaemia in the pig. Vet Res Commun,1997,21:187–200
[15] Sorrentino R, Bianca R, Lippolis L, Sorrentino L, Autore G, Pinto A. Involvement of ATP-sensitive potassium channels in a model of a delayed vascular hyporeactivity induced by lipopolysaccharide in rats. Br J Pharmacol,1999,127:1447–53
[16] Shi NQ, Ye B, Makielski JC. Function and distribution of the SUR isoforms and splice variants. J Mol Cell Cardiol,2005,39:51–60
[17] Vanelli G, Hussain SNA, Aguggini G. Glibenclamide, a blocker of ATP-sensitive potassiumchannels,reverses endotoxin-induced hypotension in pig. Exp Physiol,1995,80:167–70
[18] Simonson SG, Welty-Wolf K, Huang YT, Griebel JA, Caplan MS, Fracica PJ, et al. Altered mitochondrial redox responses in gram negative septic shock in primates. Circ Shock, 1994,43:34–43
[19] Kantrow SP, Taylor DE, Carraway MS, Piantadosi CA. Oxidative metabolism in rat hepatocytes and mitochondria during sepsis. Arch Biochem Biophys,1997,345:278–88
[20] Vanhorebeek I, De Vos R, Mesotten D, Wouters PJ, Wolf-Peeters C, Van den Berghe G. Protection of hepatocyte mitochondrial ultrastructure and function by strict blood glucose control with insulin in critically ill patients. Lancet,2005,365:53–9
[21] Garlid KD, Paucek P. Mitochondrial potassium transport: the K+ cycle. Biochim Biophys Acta ,2003,1606:23–41
[22] Costa AD, Quinlan CL, Andrukhiv A, West IC, Jaburek M, Garlid KD. The direct physiological effects of mitoKATP opening on heart mitochondria. Am J Physiol,2006,290:H406–H415
[23] Huang L, Li W, Li B, Zou F. Activation of ATP-sensitive K channels protects hippocampal CA1 neurons from hypoxia by suppressing p53 expression. Neurosci Lett,2006,398:34–8
[24] Sharshar T, Blanchard A, Paillard M, Raphael JC, Gajdos P, Annane D. Circulating vasopressin levels in septic shock. Crit Care Med,2003,31:1752–8
[25] Ahluwalia J, Tinker A, Clapp LH, Duchen MR, Abramov AY, Pope S, et al. The large-conductance Ca2+-activated K+ channel is essential for innate immunity. Nature, 2004,427:853–8
[26] Haslberger A, Romanin C, Koerber R. Membrane potential modulates release of tumor necrosis factor in lipopolysaccharide-stimulated mouse macrophages. Mol Biol Cell,1992,3:451–60
[27] Blunck R, Scheel O, Muller M, Brandenburg K, Seitzer U, Seydel U. New insights into endotoxininduced activation of macrophages: involvement of a K+ channel in transmembrane signaling. J Immunol,2001,166:1009–15
[28] Muller M, Scheel O, Lindner B, Gutsmann T, Seydel U. The role of membrane-bound LBP,endotoxin aggregates, and the MaxiK channel in LPS-induced cell activation. J Endotoxin Res,2003,9:181–6
[1] Sands KE, Bates DW, Lanken PN, Graman PS, Hibberd PL, Kahn KL, Parsonnet J, Panzer R, Orav EJ, Snydman DR, Black E, Schwartz JS, Moore R, Johnson BL Jr, Platt R. Epidemiology of sepsis syndrome in 8 academic medical centers. JAMA,1997,278:234-40
[2] Ulloa L,Tracey K J. The‘cytokine profile’: a code for sepsis. Trends Mol Med, 2005, 11: 56–63
[3] Riedemann NC, Guo RF, Ward PA. Novel strategies for the treatment of sepsis. Nature Med, 2003, 9: 517–524
[4] Tracey KJ, Cerami A. Tumor necrosis factor: a pleiotropic cytokine and therapeutic target. Annu Rev Med, 1994, 45: 491–503
[5] Tracey KJ, Cerami A. Tumor necrosis factor, other cytokines and disease. Annu Rev Cell Biol, 1993, 9: 317–343
[6] Van der Poll T, Lowry SF. Tumor necrosis factor in sepsis: mediator of multiple organ failure or essential part of host defense? Shock, 1995, 3: 1–12
[7] Bryan J, Aguilar-Bryan L. Sulfonylurea receptors: ABC transporters that regulate ATP-sensitive K+ channels. Biochim Biophys Acta, 1999, 1461: 285–305
[8] Seino S, Miki T. Physiological and pathophysiological roles of ATP-sensitive K+ channels. Prog Biophys Mol Biol, 2003, 81: 133–76
[9] Rodrigo GC, Standen NB. ATP-sensitive potassium channels. Curr Pharm Des, 2005,11:1915–40.
[10] Baukrowitz T, Fakler B . KATP channels gated by intracellular nuclentides and phospholipids.Eur J Biochem, 2000, 267: 5842—5848
[11] Baigar R, Seetharaman S, Kowahowski AJ. Identification and properties of a novel intracellar (mitochondria1) ATP-sensitive potassium channel in brain. J Biol Chem, 2001, 276: 33369—33374
[12] Mirenova GD,Nngoa AE,Marinov BS.Functional distinctions between the mitochondrial ATP-dependent K+channel (mitoKATP) and its inward rectifier subunit(mitoKIR).J Biol Chem, 2004, 279: 32562—32568
[13] Garlid KD,Pancek P. Mitochondrial potassium transport: the K(+) cycle.Biochim Biophys Acta, 2003, 1606: 23—41
[14] Bryan J, Aguilar-Bryan L. Sulfonylurea receptors: ABC transporters that regulate ATP-sensitive K+ channels. Biochim Biophys Acta, 1999, 1461: 285–305
[15] Seino S, Miki T. Physiological and pathophysiological roles of ATP-sensitive K+ channels. Prog Biophys Mol Biol, 2003, 81: 133–76
[16] Rodrigo GC, Standen NB. ATP-sensitive potassium channels. Curr Pharm Des, 2005, 11: 1915–40
[17] Wilson AJ, Jabr RI, Clapp LH. Calcium modulation of vascular smooth muscle ATP-sensitive K+ channels: Role of protein phosphatase-2B. Circ Res, 2000, 87: 1019–25
[18] Orie NN, Perrino BA, Tinker A, Clapp LH. Calcineurin Aαregulates Kir6.1/SUR2B in HEK293 cells through an interaction with protein kinase A. J Physiol, 2005, 567P: C24
[19] Miki T, Seino S. Roles of KATP channels as metabolic sensors in acute metabolic changes. J Mol Cell Cardiol , 2005, 38: 917–25
[20] Od JW, Harrell M, Flagg TP. Role of sulfonylurea receptor type 1 subunits of ATP-sensitive potassium channels in myocardial ischemia/reperfusion injury.Circulation, 2008, 117: 1405-13
[21] Hilgers KF, Hartner A, Porst M, Veelken R, Mann JF. Angiotensin II type 1 receptor blockade prevents lethal malignant hypertension: relation to kidney inflammation. Circulation, 2001, 104: 1436-40
[22] Nandakumar KS, Holmdahl R. Therapeutic cleavage of IgG: new avenues for treating inflammation. Trends Immunol, 2008, 29: 173-8
[23] Gallowitsch-Puerta M, Tracey KJ. Immunologic role of the cholinergic anti-inflammatory pathway and the nicotinic acetylcholine alpha 7 receptor. Ann N Y Acad Sci, 2005, 1062: 209-19
[24] Fodale V, Santamaria LB. Cholinesterase inhibitors improve survival in experimental sepsis: a new way to activate the cholinergic anti-inflammatory pathway. Crit Care Med, 2008, 36: 622-3
[25] Kox M, Hoedemaekers AW, Pickkers P, van der Hoeven JG, Pompe JC. A possible role for the cholinergic anti-inflammatory pathway in increased mortality observed in critically ill patients receiving nicotine replacement therapy. Crit Care Med, 2007, 35: 2468-9
[26] Nair MG, Du Y, Perrigoue JG, Zaph C, Taylor JJ, Goldschmidt M, Swain GP, Yancopoulos GD, Valenzuela DM, Murphy A, Karow M, Stevens S, Pearce EJ, Artis D. Alternatively activated macrophage-derived RELM-{alpha} is a negative regulator of type 2 inflammation in the lung. J Exp Med, 2009, 206: 937-52
[27] Munitz A, Seidu L, Cole ET, Ahrens R, Hogan SP, Rothenberg ME. Resistin-like molecule alpha decreases glucose tolerance during intestinal inflammation. J Immunol, 2009, 182:2357-63
[28] Qu P, Du H, Li Y, Yan C. Myeloid-specific expression of Api6/AIM/Sp alpha induces systemic inflammation and adenocarcinoma in the lung. J Immunol, 2009, 182: 1648-59
[29] Patel NS, di Paola R, Mazzon E, Britti D, Thiemermann C, Cuzzocrea S. Peroxisome proliferator-activated receptor-alpha contributes to the resolution of inflammation after renal ischemia/reperfusion injury. J Pharmacol Exp Ther, 2009, 328: 635-43
[30] Li Z, Daniel EE, Lane CG, Arnaout MA, O'Byrne PM. Effect of an anti-Mo1 MAb on ozone-induced airway inflammation and airway hyperresponsiveness in dogs. Am J Physiol, 1992, 263: L723-6
[31] Czura CJ, Friedman SG, Tracey KJ. Neural inhibition of inflammation: the cholinergic anti-inflammatory pathway. J Endotoxin Res, 2003, 9: 409-13
[32] Invitti C. Obesity and low-grade systemic inflammation. Minerva Endocrinol, 2002, 27: 209-14
[33] Nicolussi EM, Huck S, Lassmann H, Bradl M. The cholinergic anti-inflammatory system limits T cell infiltration into the neurodegenerative CNS, but cannot counteract complex CNS inflammation. Neurobiol Dis, 2009
[34] Karim R, MatstnnotoS, Fragashi H. The molecular pathogenesis of endotoxic shock and organ failure. Mol Med To, 1999, 5:123
[35] Li Z, Daniel EE, Lane CG, Arnaout MA, O'Byrne PM. Effect of an anti-Mo1 MAb on ozone-induced airway inflammation and airway hyperresponsiveness in dogs. Am J Physiol, 1992, 263: L723-6
[36] Czura CJ, Friedman SG, Tracey KJ. Neural inhibition of inflammation: the cholinergic anti-inflammatory pathway. J Endotoxin Res, 2003, 9: 409-13
[37] Invitti C. Obesity and low-grade systemic inflammation. Minerva Endocrinol, 2002, 27: 209-14
[38] Nicolussi EM, Huck S, Lassmann H, Bradl M. The cholinergic anti-inflammatory system limits T cell infiltration into the neurodegenerative CNS, but cannot counteract complex CNS inflammation. Neurobiol Dis, 2009
[2] Ulloa L, Tracey K J. The‘cytokine profile’: a code for sepsis. Trends Mol Med 2005,11: 56–63
[3] Riedemann NC, Guo RF, Ward PA. Novel strategies for the treatment of sepsis. Nature Med, 2003,9: 517–524
[4] Tracey KJ, Cerami A. Tumor necrosis factor: a pleiotropic cytokine and therapeutic target. Annu Rev Med,1994,45:491–503
[5] Tracey KJ, Cerami A. Tumor necrosis factor, other cytokines and disease. Annu Rev Cell Biol,1993,9:317–343
[6] Van der Poll T, Lowry SF. Tumor necrosis factor in sepsis: mediator of multiple organ failure or essential part of host defense? Shock,1995; 3:1–12
[7] Bryan J, Aguilar-Bryan L. Sulfonylurea receptors: ABC transporters that regulate ATP-sensitive K+ channels. Biochim Biophys Acta,1999,1461:285–305
[8] Seino S, Miki T. Physiological and pathophysiological roles of ATP-sensitive K+ channels. Prog Biophys Mol Biol,2003,81:133–76
[9] Rodrigo GC, Standen NB. ATP-sensitive potassium channels. Curr Pharm Des,2005,11:1915–40
[10] Wilson AJ, Jabr RI, Clapp LH. Calcium modulation of vascular smooth muscle ATP-sensitive K+ channels: Role of protein phosphatase-2B. Circ Res,2000,87:1019–25
[11] Orie NN, Perrino BA, Tinker A, Clapp LH. Calcineurin Aαregulates Kir6.1/SUR2B in HEK293 cells through an interaction with protein kinase A. J Physiol,2005,567P:C24
[12] Miki T, Seino S. Roles of KATP channels as metabolic sensors in acute metabolic changes. J Mol Cell Cardiol 2005,38:917–25
[13] Landry DW, Oliver JA. The ATP-sensitive K+ channel mediates hypotension in endotoxemia and hypoxic lactic acidosis in dog. J Clin Invest,1992,89:2071–4
[14] Vanelli G, Hussain SN, Dimori M, Aguggini G. Cardiovascular responses to glibenclamide during endotoxaemia in the pig. Vet Res Commun,1997,21:187–200
[15] Sorrentino R, Bianca R, Lippolis L, Sorrentino L, Autore G, Pinto A. Involvement of ATP-sensitive potassium channels in a model of a delayed vascular hyporeactivity induced by lipopolysaccharide in rats. Br J Pharmacol,1999,127:1447–53
[16] Shi NQ, Ye B, Makielski JC. Function and distribution of the SUR isoforms and splice variants. J Mol Cell Cardiol,2005,39:51–60
[17] Vanelli G, Hussain SNA, Aguggini G. Glibenclamide, a blocker of ATP-sensitive potassiumchannels,reverses endotoxin-induced hypotension in pig. Exp Physiol,1995,80:167–70
[18] Simonson SG, Welty-Wolf K, Huang YT, Griebel JA, Caplan MS, Fracica PJ, et al. Altered mitochondrial redox responses in gram negative septic shock in primates. Circ Shock, 1994,43:34–43
[19] Kantrow SP, Taylor DE, Carraway MS, Piantadosi CA. Oxidative metabolism in rat hepatocytes and mitochondria during sepsis. Arch Biochem Biophys,1997,345:278–88
[20] Vanhorebeek I, De Vos R, Mesotten D, Wouters PJ, Wolf-Peeters C, Van den Berghe G. Protection of hepatocyte mitochondrial ultrastructure and function by strict blood glucose control with insulin in critically ill patients. Lancet,2005,365:53–9
[21] Garlid KD, Paucek P. Mitochondrial potassium transport: the K+ cycle. Biochim Biophys Acta ,2003,1606:23–41
[22] Costa AD, Quinlan CL, Andrukhiv A, West IC, Jaburek M, Garlid KD. The direct physiological effects of mitoKATP opening on heart mitochondria. Am J Physiol,2006,290:H406–H415
[23] Huang L, Li W, Li B, Zou F. Activation of ATP-sensitive K channels protects hippocampal CA1 neurons from hypoxia by suppressing p53 expression. Neurosci Lett,2006,398:34–8
[24] Sharshar T, Blanchard A, Paillard M, Raphael JC, Gajdos P, Annane D. Circulating vasopressin levels in septic shock. Crit Care Med,2003,31:1752–8
[25] Ahluwalia J, Tinker A, Clapp LH, Duchen MR, Abramov AY, Pope S, et al. The large-conductance Ca2+-activated K+ channel is essential for innate immunity. Nature, 2004,427:853–8
[26] Haslberger A, Romanin C, Koerber R. Membrane potential modulates release of tumor necrosis factor in lipopolysaccharide-stimulated mouse macrophages. Mol Biol Cell,1992,3:451–60
[27] Blunck R, Scheel O, Muller M, Brandenburg K, Seitzer U, Seydel U. New insights into endotoxininduced activation of macrophages: involvement of a K+ channel in transmembrane signaling. J Immunol,2001,166:1009–15
[28] Muller M, Scheel O, Lindner B, Gutsmann T, Seydel U. The role of membrane-bound LBP,endotoxin aggregates, and the MaxiK channel in LPS-induced cell activation. J Endotoxin Res,2003,9:181–6
[1] Sands KE, Bates DW, Lanken PN, Graman PS, Hibberd PL, Kahn KL, Parsonnet J, Panzer R, Orav EJ, Snydman DR, Black E, Schwartz JS, Moore R, Johnson BL Jr, Platt R. Epidemiology of sepsis syndrome in 8 academic medical centers. JAMA,1997,278:234-40
[2] Ulloa L,Tracey K J. The‘cytokine profile’: a code for sepsis. Trends Mol Med, 2005, 11: 56–63
[3] Riedemann NC, Guo RF, Ward PA. Novel strategies for the treatment of sepsis. Nature Med, 2003, 9: 517–524
[4] Tracey KJ, Cerami A. Tumor necrosis factor: a pleiotropic cytokine and therapeutic target. Annu Rev Med, 1994, 45: 491–503
[5] Tracey KJ, Cerami A. Tumor necrosis factor, other cytokines and disease. Annu Rev Cell Biol, 1993, 9: 317–343
[6] Van der Poll T, Lowry SF. Tumor necrosis factor in sepsis: mediator of multiple organ failure or essential part of host defense? Shock, 1995, 3: 1–12
[7] Bryan J, Aguilar-Bryan L. Sulfonylurea receptors: ABC transporters that regulate ATP-sensitive K+ channels. Biochim Biophys Acta, 1999, 1461: 285–305
[8] Seino S, Miki T. Physiological and pathophysiological roles of ATP-sensitive K+ channels. Prog Biophys Mol Biol, 2003, 81: 133–76
[9] Rodrigo GC, Standen NB. ATP-sensitive potassium channels. Curr Pharm Des, 2005,11:1915–40.
[10] Baukrowitz T, Fakler B . KATP channels gated by intracellular nuclentides and phospholipids.Eur J Biochem, 2000, 267: 5842—5848
[11] Baigar R, Seetharaman S, Kowahowski AJ. Identification and properties of a novel intracellar (mitochondria1) ATP-sensitive potassium channel in brain. J Biol Chem, 2001, 276: 33369—33374
[12] Mirenova GD,Nngoa AE,Marinov BS.Functional distinctions between the mitochondrial ATP-dependent K+channel (mitoKATP) and its inward rectifier subunit(mitoKIR).J Biol Chem, 2004, 279: 32562—32568
[13] Garlid KD,Pancek P. Mitochondrial potassium transport: the K(+) cycle.Biochim Biophys Acta, 2003, 1606: 23—41
[14] Bryan J, Aguilar-Bryan L. Sulfonylurea receptors: ABC transporters that regulate ATP-sensitive K+ channels. Biochim Biophys Acta, 1999, 1461: 285–305
[15] Seino S, Miki T. Physiological and pathophysiological roles of ATP-sensitive K+ channels. Prog Biophys Mol Biol, 2003, 81: 133–76
[16] Rodrigo GC, Standen NB. ATP-sensitive potassium channels. Curr Pharm Des, 2005, 11: 1915–40
[17] Wilson AJ, Jabr RI, Clapp LH. Calcium modulation of vascular smooth muscle ATP-sensitive K+ channels: Role of protein phosphatase-2B. Circ Res, 2000, 87: 1019–25
[18] Orie NN, Perrino BA, Tinker A, Clapp LH. Calcineurin Aαregulates Kir6.1/SUR2B in HEK293 cells through an interaction with protein kinase A. J Physiol, 2005, 567P: C24
[19] Miki T, Seino S. Roles of KATP channels as metabolic sensors in acute metabolic changes. J Mol Cell Cardiol , 2005, 38: 917–25
[20] Od JW, Harrell M, Flagg TP. Role of sulfonylurea receptor type 1 subunits of ATP-sensitive potassium channels in myocardial ischemia/reperfusion injury.Circulation, 2008, 117: 1405-13
[21] Hilgers KF, Hartner A, Porst M, Veelken R, Mann JF. Angiotensin II type 1 receptor blockade prevents lethal malignant hypertension: relation to kidney inflammation. Circulation, 2001, 104: 1436-40
[22] Nandakumar KS, Holmdahl R. Therapeutic cleavage of IgG: new avenues for treating inflammation. Trends Immunol, 2008, 29: 173-8
[23] Gallowitsch-Puerta M, Tracey KJ. Immunologic role of the cholinergic anti-inflammatory pathway and the nicotinic acetylcholine alpha 7 receptor. Ann N Y Acad Sci, 2005, 1062: 209-19
[24] Fodale V, Santamaria LB. Cholinesterase inhibitors improve survival in experimental sepsis: a new way to activate the cholinergic anti-inflammatory pathway. Crit Care Med, 2008, 36: 622-3
[25] Kox M, Hoedemaekers AW, Pickkers P, van der Hoeven JG, Pompe JC. A possible role for the cholinergic anti-inflammatory pathway in increased mortality observed in critically ill patients receiving nicotine replacement therapy. Crit Care Med, 2007, 35: 2468-9
[26] Nair MG, Du Y, Perrigoue JG, Zaph C, Taylor JJ, Goldschmidt M, Swain GP, Yancopoulos GD, Valenzuela DM, Murphy A, Karow M, Stevens S, Pearce EJ, Artis D. Alternatively activated macrophage-derived RELM-{alpha} is a negative regulator of type 2 inflammation in the lung. J Exp Med, 2009, 206: 937-52
[27] Munitz A, Seidu L, Cole ET, Ahrens R, Hogan SP, Rothenberg ME. Resistin-like molecule alpha decreases glucose tolerance during intestinal inflammation. J Immunol, 2009, 182:2357-63
[28] Qu P, Du H, Li Y, Yan C. Myeloid-specific expression of Api6/AIM/Sp alpha induces systemic inflammation and adenocarcinoma in the lung. J Immunol, 2009, 182: 1648-59
[29] Patel NS, di Paola R, Mazzon E, Britti D, Thiemermann C, Cuzzocrea S. Peroxisome proliferator-activated receptor-alpha contributes to the resolution of inflammation after renal ischemia/reperfusion injury. J Pharmacol Exp Ther, 2009, 328: 635-43
[30] Li Z, Daniel EE, Lane CG, Arnaout MA, O'Byrne PM. Effect of an anti-Mo1 MAb on ozone-induced airway inflammation and airway hyperresponsiveness in dogs. Am J Physiol, 1992, 263: L723-6
[31] Czura CJ, Friedman SG, Tracey KJ. Neural inhibition of inflammation: the cholinergic anti-inflammatory pathway. J Endotoxin Res, 2003, 9: 409-13
[32] Invitti C. Obesity and low-grade systemic inflammation. Minerva Endocrinol, 2002, 27: 209-14
[33] Nicolussi EM, Huck S, Lassmann H, Bradl M. The cholinergic anti-inflammatory system limits T cell infiltration into the neurodegenerative CNS, but cannot counteract complex CNS inflammation. Neurobiol Dis, 2009
[34] Karim R, MatstnnotoS, Fragashi H. The molecular pathogenesis of endotoxic shock and organ failure. Mol Med To, 1999, 5:123
[35] Li Z, Daniel EE, Lane CG, Arnaout MA, O'Byrne PM. Effect of an anti-Mo1 MAb on ozone-induced airway inflammation and airway hyperresponsiveness in dogs. Am J Physiol, 1992, 263: L723-6
[36] Czura CJ, Friedman SG, Tracey KJ. Neural inhibition of inflammation: the cholinergic anti-inflammatory pathway. J Endotoxin Res, 2003, 9: 409-13
[37] Invitti C. Obesity and low-grade systemic inflammation. Minerva Endocrinol, 2002, 27: 209-14
[38] Nicolussi EM, Huck S, Lassmann H, Bradl M. The cholinergic anti-inflammatory system limits T cell infiltration into the neurodegenerative CNS, but cannot counteract complex CNS inflammation. Neurobiol Dis, 2009