Role of nitric oxide in hemorrhagic shock-induced bacterial translocation

BACKGROUND: Hemorrhagic shock-induced bacterial translocation is an etiologic factor in the pathogenesis of multiple system organ damage. Excessive production of nitric oxide (NO) during hemorrhagic shock may lead to cellular injury and gut barrier failure that promotes bacterial translocation. We investigated the effect of aminoguanidine (AG) and N(G)-nitro-l-arginine methyl ester (l-NAME), both inhibitors of NO synthase, on hemorrhagic shock- induced bacterial translocation in the rat. MATERIALS AND METHODS: Anesthetized male Sprague-Dawley rats were subjected to a hemorrhagic shock protocol for 30 min followed by intravenous injection (1 mL/kg body wt) with normal saline, AG (100 mg/kg), or l-NAME (10 mg/kg). Tissues/organs were examined histologically for damage and bacterial translocation. Plasma nitrate/nitrite was measured using a procedure based on the Griess reaction, and nitric oxide synthase (NOS) expression was determined immunohistochemically. RESULTS: The shocked animals treated with saline died within 90 min, and deaths were associated with 100% bacterial translocation, increased tissue/organ damage, and elevated nitrate/nitrite production. In contrast, both AG and l-NAME increased the survival time of shocked rats to >72 h, abrogated bacterial translocation, reduced tissue/organ damage, and prevented excessive nitrate/nitrite production and upregulation of expression of endothelial NOS and inducible NOS. CONCLUSIONS: Prevention of bacterial translocation by pharmacologic agents such as aminoguanidine and l-NAME could be an important therapeutic approach to lessen mortality rates following hemorrhagic shock.     

门静脉高压症时细菌移位与内毒素血症和一氧化氮之间关系的实验研究

目的 探讨门静脉高压症时细菌移位与内毒素血症和一氧化氮（NO）之间的关系。方法 采用门静脉部分缩窄（PVS）制成门静脉高压模型的30只大鼠均分为3组：模型组（B）、左旋精氨酸组（C）、单－甲基精氨酸组（D）。另10只大鼠接受假手术作为对照组（A）。术后2周取肠系膜淋巴结（MLN）、脾和血标本进行细菌培养;测定门静脉压力、内毒素和NO2^-水平;并用镧做示踪剂观察肠粘膜屏障通透性变化和粘膜 形态的改变。结果 PVS各组MLN细菌检出率高于A组,血浆内毒素水平均上升。PVS各组NO2^-和门静脉压力均较A组增高,在D组二者均较B组下降。细菌移位率、内毒素和NO2^-水平之间呈显著正相关。镧沉积到PVS各组的粘膜上皮细胞和其上下固有层细胞间,粘膜细胞的微绒毛有形态改变。结论 门静脉高压症可发生细菌移位的内毒素血症,可能与肠粘屏障通透性增强和破坏有关,提示细菌移位引起的内毒素血症并增加NO2^－水平。     

The effects of nitric oxide supplementation and inhibition on bacterial translocation in bile duct ligated rats

Obstructive jaundice promotes bacterial translocation from the gut, but the role of nitric oxide is controversial in this process. We studied the effects of nitric oxide synthase substrate, L-arginine, and nitric oxide synthase inhibitor, N(G)-nitro-L-arginine methyl ester, on bacterial translocation in bile duct ligated rats. The animals were randomized into five groups; control, sham, common bile duct ligation alone, nitric oxide inhibition, and nitric oxide supplementation. Obstructive jaundice was performed with common bile duct ligation. L-arginine or N(G)-nitro-L-arginine methyl ester was injected once daily for 14 days. Blood bilirubin level, liver histology, and bacterial translocation to the mesenteric lymph nodes as well as to the liver were assessed. The L-arginine supplemented group had the lowest bacterial translocation rate, but the most prominent hepatic fibrosis. Nitric oxide inhibition increased bacterial translocation to the mesenteric lymph nodes. Therefore, the administration of nitric oxide donor or inhibitor acts as a significant regulatory factor for bacterial translocation in obstructive jaundice.     

Administration of nitric oxide with caspase inhibitors minimizes bacterial translocation in experimental intestinal transplantation

BACKGROUND: Bacterial translocation (BT) has been suggested to be responsible for the high incidence of infections occurring after small-bowel transplantation (Trp). Nitric oxide (NO) and apoptosis could affect cell demise. The aim of this study was to asses whether supplementation of University of Wisconsin (UW) solution with NO donors and apoptosis inhibitors can abolish BT in Trp. METHODS: The following experimental groups were studied: sham, Trp, intestinal transplantation, Trp+spermine NONOate (NONOs), and Trp+NONOs+caspase inhibitor Z-Val-Ala-Asp(Ome)-fluoromethylketone(Z-VAD-fmk). Histologic analysis, caspase-3 activity, DNA fragmentation, and BT from graft to mesenteric lymph nodes, liver, and spleen were measured in tissue samples after transplantation. RESULTS: During intestinal transplantation, apoptosis and necrosis were increased, showing graft injury and high levels of BT. The rats treated with NONOs showed a histologic protection of transplanted graft and a decrease in BT despite caspase-3 and DNA fragmentation-inducing effects. Administration of caspase inhibitor Z-VAD to NONOs-treated rats reversed the NO apoptosis-inducing effects and showed the lowest levels of BT in all tissues. CONCLUSIONS: Exogenous administration of NO associated with the inhibition of apoptosis maintains the graft in optimal conditions in terms of BT and improves the histology of the graft after intestinal transplantation in rats.     

胆汁性肝硬化猪肝组织中内皮素受体及诱导型一氧化氮合酶mRNA表达的意义

目的　研究胆汁性肝硬化猪肝组织中内皮素受体 (ETAR、ETBR)及诱导型一氧化氮合酶 (iNOS)mRNA的表达及其对肝脏血液循环的影响。方法　湖北白种猪 15头 ,其中实验组 10头 ,对照组 5头。监测门静脉压 (Pp)、门静脉血流量 (Qp)和平均动脉压 (MAP)的变化 ,用原位杂交方法检测肝组织中ETAR、ETBR及iNOSmRNA的表达水平。结果　术后 4周实验组Pp(14 .6±1.7)mmHg (1mmHg =0 .13 3kPa)、Qp(3 2 .3± 2 .8)ml·min-1·kg-1及MAP(80 .5± 2 .5 )mmHg显著高于对照组的 (8.1± 3 .6)mmHg、(14 .2± 3 .7)ml·min-1·kg-1和 (10 8.3± 6.5 )mmHg(P <0 .0 1)。实验组ETAR、ETBR及iNOSmRNA的表达水平分别为 0 .78± 0 .2 0、0 .90± 0 .2 3和 0 .75± 0 .13 ,均显著高于对照组 (分别为 0 .3 8± 0 .2 6、0 .2 4± 0 .15和 0 .3 2± 0 .11,P <0 .0 1)。结论　胆汁性肝硬化猪肝组织中ETAR、ETBR及iNOSmRNA表达水平的变化影响肝脏血液循环的调节。     

Pitavastatin prevents bacterial translocation after nonpulsatile/low-pressure blood flow in early atherosclerotic rat: inhibition of small intestine inducible nitric oxide synthase

BACKGROUND: Cardiopulmonary bypass decreases intestinal mucosal blood flow because of nonpulsatile and low-pressure blood flow resulting in bacterial translocation (BT) and atherosclerosis also has peripheral blood flow deficiency. The risk of nonpulsatile and low-pressure blood flow for atherosclerotic animals and the effect of statin administration, which has pleiotropic effects, were studied. METHODS: Wistar rats were divided into four groups: group N (normal diet), group C (high-cholesterol diet), group S (group C plus pitavastatin therapy), and group I [group C plus inducible nitric oxide (iNOS) inhibitor therapy]. First of all, vascular responses were measured. Then the rats underwent nonpulsatile/low-pressure blood flow in the intestine, and the serum peptidoglycan concentration as a parameter of BT, the small intestinal PO(2) ratio (intestinal PO(2)/PaO(2)) as a parameter of mucosal blood flow, and NO concentrations were measured before surgery (T0), at the end of 90 min of stenosis (T1), and 90 min after the release of stenosis (T2). Immunostaining for nitrotyrosine was also performed at T2. RESULTS: Group C had vascular endothelial dysfunction without histological changes, which indicated early atherosclerosis. The serum peptidoglycan concentration increased significantly at T2 only in group C. The intestinal PO(2) ratio was decreased at T1 in all the groups, and retuned to baseline at T2 in group N and group S, but not in group C or group I. Jejunal NO only in group C was significantly higher at all time points and ileal NO production at T1 and T2. There tended to be a positive stain for nitrotyrosine along the mucosal epithelium in group C. CONCLUSION: In the setting of early atherosclerosis, intestinal blood flow does not only improve after nonpulsatile/low-pressure blood flow but causes BT because of a large amount of NO from high enzymatic intestinal iNOS activity, and pitavastatin treatment can prevent BT by improving both issues.     

一氧化氮/一氧化氮合酶系统在脊髓继发性损伤中的作用

脊髓损伤(spinal cord injury,SCI)是临床亟待解决的一大难题,虽然SCI的实验性治疗在18世纪就已经开始,但迄今尚未取得有意义的突破。原因可能是对SCI后病理发展过程认识不足。目前认为急性脊髓损伤有两种损伤机制,即原发性机械损伤及迟发性结构损害所致的继发性损伤,后者产生的损伤程度更严重、范围更广。因此,虽然原发的脊髓横断伤较少,但由于继发的不可逆的结构的改变,损伤仍然非常严重,故阻断继发性损伤的发生成了SCI研究的焦点。一氧化氮(Nitric Oxide,NO)/一氧化氮合酶(Nitric Oxide Synthase,NOS)系统是近年来被认为在继发…     

一氧化氮(NO)合酶和NO在延迟性异种移植排斥反应中的作用

目的利用小鼠至大鼠异位心脏移植模型,研究诱导性一氧化氮合酶(iNOS)和受体血清一氧化氮(NO)在延迟性异种移植排斥反应(DXR)中的作用.方法将大鼠随机分为4组A组(6只),空白对照;B组(5只),来氟米物(Lef)+环孢素A(CsA);C组(6只),氨基胍;D组(6只),氨基胍+Lef+CsA.利用免疫组织化学染色检测CD68和NOS2,原位杂交技术检测iNOS mRNA表达.于移植前3 d和移植心脏排斥时分别采集血清检测NO含量.结果所有被排斥心脏中均见巨噬细胞(MФ)浸润,Lef+CsA显著延长移植心脏存活(与A和C组相比,P＜0.05),单用氨基胍使移植心脏存活(3 83±1.47)d(与A组比较,P＜0.05),氨基胍联用Lef和CsA使移植心脏存活(8.67±1.76)d(与A、B和C组比较,P＜0.05).发生DXR时浸润的MФ均有NOS2蛋白和mRNA阳性表达,且不受氨基胍影响.发生DXR时大鼠血清NO水平较移植前显著升高(P＜0.01),氨基胍可显著降低排斥时NO水平.结论小鼠至大鼠心脏移植发生DXR时浸润的MФ表达iNOS增多,且血清NO升高.抑制iNOS活性,降低NO水平可显著延长移植物存活时间,提示iNOS和NO是DXR发生的可能机制之一.     

Inhaled nitric oxide decreases the bacterial load in a rat model of Pseudomonas aeruginosa pneumonia

Gaseous nitric oxide (NO) is bactericidal in vitro. However whether and how it can be used for the treatment of bacterial lung infections in patients with cystic fibrosis is unclear. Here we assessed the bactericidal effect of intermittently inhaled 160 ppm NO for 30 min every 4 h in a Pseudomonas aeruginosa pneumonia model in rats. NO significantly reduced P. aeruginosa colony count in rat lungs but did not affect neutrophil myeloperoxidase function methemoglobin percentage nor plasma nitrite/nitrate levels. This regimen warrants exploration in infected patients with cystic fibrosis.     

Kupffer cell:hepatocyte cocultures release nitric oxide in response to bacterial endotoxin

Nitric oxide (NO.) is a short-lived intermediate in a biochemical pathway where L-arginine is converted to L-citrulline and nitrite/nitrate (NO2-/NO3-). This highly reactive molecule is the biologically active component of this inducible pathway in macrophages. Using a rat Kupffer cell:hepatocyte (KC:HC) coculture model, we have previously shown that this combination of cells produces large quantities of both citrulline and NO2-/NO3- if exposed to lipopolysaccharides (LPS) but we did not determine whether nitric oxide was produced or released. We had also shown that this L-arginine metabolism was associated with a profound decrease in total protein synthesis. In these experiments, we show that KC:HC cocultures release nitric oxide into the culture supernatant if exposed to LPS. NO. production by these cells requires L-arginine and is inhibited by NG-mono-methyl-L-arginine. In addition, the time course for NO. release by KC:HC cocultures parallels the previously reported time course for NO2-/NO3- synthesis and the decrease in protein synthesis, supporting the hypothesis that NO. is the reactive nitrogen intermediate of the pathway responsible for this inhibition of protein synthesis. Finally, we show that KC:HC cocultures release more NO. than KC alone in response to LPS, and we propose that the combination of KC and HC acts as a functional unit capable of generating large amounts of NO. from L-arginine in gram-negative sepsis.     

Hypoxia-induced bacterial translocation in the puppy.

Because hypoxia is one of the most common major stresses to which a neonate is exposed, we postulated that it alone might be the cause of intestinal bacterial translocation, which could be the underlying etiology of neonatal sepsis. An animal model, in which hypoxia is the sole stress, was developed in our laboratory and tested in 18 puppies to determine the effect of hypoxia and reoxygenation on intestinal bacterial translocation. In group I (n = 8), following laparotomy and cannulation of the superior mesenteric vein (SMV), the FIO2 was decreased from 21% to 9% for 90 minutes followed by reoxygenation at 21% for 120 minutes. The abdomen was closed and the animals were allowed to recover. After 24 hours the mesenteric lymph nodes (MLNs), spleen, and liver were harvested for bacterial determination and the ileum and jejunum for histological evaluation. Group II (n = 7) was treated the same as group I with the FIO2 maintained at 21%. Group III (n = 3) animals were killed, without intervention, for bacterial analysis. In group I, the systemic PO2 decreased by 75%, SMV PO2 decreased by 64%, and oxygen delivery to the small bowel decreased by 80% in comparison with group II. The mean arterial pressure and cardiac output were not significantly different between group I and group II; however, the mucosal blood flow was decreased by 60% (P less than .001) in group I. Arterial and SMV blood lactic acid levels were unchanged in group I in comparison with group II, suggesting that anaerobic metabolism was not initiated in the splanchnic circulation during hypoxia.(ABSTRACT TRUNCATED AT 250 WORDS)     

The cardiovascular significance of nitric oxide

Nitric oxide (NO) synthesized from L-arginine is a ubiquitous intercellular chemical messenger involved in signal transduction in diverse mammalian cells. The isolation of molecular clones for NO synthases has permitted the characterization of several distinct enzyme isoforms. NO synthesized in vascular endothelial cells plays an important role in the control of vascular tonus and platelet aggregation, through the activation of guanylate-cyclase activity in target tissues mediated by NO. Nitric oxide which is produced by cytokine activated mononuclear cells plays an important role in inflammation and immunity as a cytotoxic effector molecule and as a transducer molecule in immune cells and in oxidative stress as a potential source of intracellular free radicals. An increase in reactive oxygen species can produce damage to lipids, proteins and DNA and induce necrosis or apoptosis. The implication of NO in different pathological processes, such as atherosclerosis, diabetes, ischaemia and reperfusion, or during inflammatory processes and the generation of free radicals contributing to the endothelial injury associated to these processes.     

iNOS/NO对结直肠肿瘤发生发展的影响

一氧化氮（nitric oxide,NO）具有广泛的生物学活性。近年来发现一氧化氮（NO）、一氧化氮合成酶（nitric oxide synthase-2,NOS-2）与结直肠肿瘤的发生、发展密切相关,它与环氧化酶（cy-cloxygenase-2,COX-2）之间存在复杂的调控机制。对NO清除剂、NOS抑制剂和释放NO的非甾体抗炎药的研究为结直肠肿瘤的防治提供了一个思路。     

The role of nitric oxide in renal transplantation

This review discusses the concept that nitric oxide synthase (NOS) may orchestrate both the inflammatory response to the renal allograft and anti-inflammatory defense in the graft itself. NO is produced by endothelial, epithelial, as well as inflammatory cells. In the setting of transplantation, the endothelium is the first lining to be subjected to the early response to injury. In turn, activated endothelial cells facilitate leukocyte recruitment, immune-mediated injury, and angiogenesis. On activation by inflammatory stimuli, endothelial cells up-regulate multiple vasoactive substances, oxygen radicals, cytokines, chemokines, and growth factors. Therefore, endothelial integrity, especially the expression of protecting vasoactive agents, such as NO, may be a key factor in resistance or sensitivity to transplantation-mediated injury. Thus, evaluating the mechanisms by which NO is involved in either protecting or injuring the transplanted allogeneic kidney is important for our understanding of renal allograft rejection. This review focuses on the role of NO in the inflammatory endothelial-leukocyte interactions, which are implicated in acute and chronic rejection of the transplanted kidney.     

The role of nitric oxide in obstructive nephropathy

PURPOSE: Ureteral obstruction leads to tubulointerstitial fibrosis and loss of renal function. Nitric oxide production ameliorates fibrosis due to obstructive uropathy. However, nitric oxide is produced by 3 isoforms of the enzyme, nitric oxide synthase. We evaluated the role of inducible nitric oxide synthase in obstructive uropathy using nitric oxide synthase knockout mice, and determined whether the administration of L-arginine to promote nitric oxide synthesis by alternative nitric oxide synthase isoforms modulates renal fibrosis in these animals. MATERIALS AND METHODS: Complete unilateral ureteral obstruction was created in wild-type C57 and inducible nitric oxide synthase knockout mice. Control animals of each strain underwent sham surgery. Throughout the experiment mice had free access to untreated tap water or water supplemented with 10 gm./l. L-arginine. Animals were sacrificed 1 and 2 weeks, respectively, after creation of unilateral ureteral obstruction. We obtained serum as well as bladder and obstructed renal pelvic urine, and determined the nitrite level in each fluid. Renal cortical thickness was measured in the normal and obstructed kidneys. The degree of tubulointerstitial fibrosis was evaluated by trichrome staining and type I collagen deposition in kidney tissue specimens. RESULTS: Nitrite was significantly decreased in the serum, bladder and renal pelvic urine of inducible nitric oxide synthase knockout mice with unilateral ureteral obstruction compared with that in wild-type C57 mice at 1 and 2 weeks (p<0.05). In knockout mice with unilateral ureteral obstruction 1 week in duration that drank tap or L-arginine supplemented water nitrite in serum and each urine sample was higher than in sham operated knockout controls. The level returned to baseline after 2 weeks of obstruction (p<0.05). After 2 weeks of obstruction there was significantly greater cortical thinning in knockout than in C57 mice (p<0.05). Moreover, knockout mice given L-arginine supplemented water for 2 weeks had even greater cortical thinning than after 1 week or than mice given tap water for 1 to 2 weeks (p<0.05). Decreased renal cortical thickness in knockout mice after 2 weeks of obstruction was associated with less intense trichrome staining and a virtual absence of type I collagen deposition compared with findings in the wild-type C57 strain. CONCLUSIONS: Inducible nitric oxide synthase knockout mice with unilateral ureteral obstruction have significantly lower nitrite in serum and urine than wild-type C57 mice. Knockout mice also have more severe renal cortical thinning than C57 animals after creation of unilateral ureteral obstruction. Providing L-arginine supplemented water to inducible nitric oxide synthase knockout mice exacerbates the loss of cortical thickness. Alterations in cortical thinning that we observed in knockout mice were associated with decreased tubulointerstitial fibrosis and a decreased net renal extracellular matrix accumulation. These data indicate that endothelial or neuronal nitric oxide synthase may be more important than inducible nitric oxide synthase for modulating renal fibrosis in obstructive uropathy.     

NO在创伤愈合过程中的作用

创伤修复是损伤组织恢复其完好结构的必需过程，大致分为三个阶段：①局部炎症反应阶段：②细胞增殖分化及肉芽组织形成阶段：③组织重建阶段。有研究发现包括一氧化氮（nitric oxide，NO）在内的小分子自由基对伤口的良好愈合起到了关键作用。现就NO在创伤修复过程中的作用作以阐述。