小鼠内毒素肝损伤中TLR4及SOCS1/3 mRNA的表达变化

脂多糖(LPS)信号的转导在炎症反应过程中起重要作用.近年来发现Toll样受体4(TLR4)是LPS识别与信号转导过程中的重要受体[1].细胞因子信号转导抑制因子(SOCS)可抑制多种信号通路,可能发挥负性调控作用[2].我们通过观察野生型小鼠(C3h/Heouj)内毒素肝损伤中TLR4及SOCS1/3 mRNA表达变化,并以TLR4缺损小鼠(C3h/Hej)为对照,探讨LPS肝损伤的信号通路及调控机制.     

肝门上径路肝切除术治疗右侧肝内胆管结石病

目的评价肝门上径路肝切除术在治疗肝胆管结石病中的价值。方法回顾性分析近两年来我院对1 8例右侧肝胆管结石病患者所行"肝门上径路"肝切除术治疗的临床资料。结果 18例"肝门上径路"肝切除术治疗的患者中,术后2例存在残余结石,其他患者术后均未见胆管炎症发作和结石复发。结论肝门上径路肝切除术可以作为一种有效的治疗右侧肝内胆管结石的方法。     

不同临床分型肝胆管结石病手术方式选择的分析

目的：探讨不同分型肝胆管结石病手术方式的合理选择。 方法：回顾性分析2005年1月—2012年12月手术治疗的667例肝胆管结石病患者的临床资料,患者分别行胆道探查取石术（BDE）、肝切除术（HT）、胆肠吻合术（HJS）中一种或联合多种手术方式处理,参照新的分型方法进行分型后,比较同种类型肝胆管结石病不同手术方式的结石清除率、术后并发症发生率及随访优良率。 结果：I型129例中,HT与BDE+HT的结石清除率、随访优良率均优于其他术式（均P<0.05）,术后并发症发生率各术式间无统计学差异（P>0.05）;IIa型72例中,BDE+HT结石清除率及随访优良率都优于其他术式（P<0.05）,BDE+HJS术后并发症发生率最低（P<0.05）;IIb型 98例中,BDE+HT的结石清除率及随访优良率最高（均P<0.05）,HT术后并发症发生率最高（均P<0.05）;IIc型25例中,各术式各指标间无统计学差异（均P>0.05）;Ea型251例中,BDE+HT结石清除率及随访优良率都优于其他术式,BDE术后并发症发生率较其他术式低（均P<0.05）;Eb型55例与Ec型37例中,BDE+HT+HJS的术后结石清除率及随访优良率优于其他术式（均P<0.05）,术后并发症各术式间无统计学差异（P>0.05）。 结论：根据肝胆管结石病合理分型选择合理的手术方式,加上各种有效的辅助手段,个体化治疗有助于提高结石清除率,减少术后并发症发生。

     

Toll样受体4在胆道损伤修复过程中的表达及意义

目的:探讨Toll样受体4(TLR4)在胆道损伤修复过程中的作用。方法:用免疫组化检测12例良性胆道狭窄胆管组织与4例正常胆管组织中TLR4的表达。用钳夹法在TLR4基因缺陷(TLR4~(-/-))小鼠和野生(TLR4~(+/+))小鼠上建立胆道损伤修复模型,并设各自的假手术对照,术后48h观察各组肝脏及胆管组织病理学变化及肝功能情况。结果:TLR4主要表达于胆管内皮细胞,良性胆道狭窄胆管壁表达阳性率明显高于正常胆管组织(83.33%vs.25.00%,P0.01)。两个假手术组小鼠术后肝脏及胆管组织均未见病理改变,TLR4~(-/-)与TLR4~(+/+)模型组小鼠均出现明显的胆道及肝脏损伤,但前者较后者明显减轻;与各自的对照组比较,TLR4~(-/-)与TLR4~(+/+)模型组小鼠血清谷丙转氨酶、总胆红素、直接胆红素水平均明显升高(均P0.05),但前者各项指标的升高程度均明显低于后者(均P0.05)。结论:TLR4可能通过参与胆管上皮细胞的天然免疫应答,并启动一系列炎症因子的表达,促使成纤维细胞增殖,在胆管良性狭窄中起到重要作用。     

肝胆管结石合并肝脓肿和胆管癌：附14例报告

目的：探讨肝胆管结石合并肝脓肿和胆管癌的诊断和治疗方法。
方法：对2004年7月—2009年12月收治的14例肝胆管结石并肝脓肿和胆管癌的临床资料进行回顾性分析。
结果：全组肝胆管结石并肝脓肿和胆管癌的发生率为0.58%（14/2 432）,术前确诊5例（5/14,35.7%）;另术中快速病检发现7例,术后病检发现2例。肿瘤切除7例（7/14,50.0%）。随访的5例行肿瘤切除者生存期均超过1年,其中1例存活5年3个月;4例非切除性姑息性手术分别存活1,6,7,13个月。
结论：有多年肝胆管结石病史患者合并肝脓肿时要考虑胆管癌的可能。早期诊断及早行肿瘤根治切除是提高疗效的关键。

     

肝胆管结石病人皮下永久性胆管通路建立及其应用价值分析

目的 总结在肝胆管结石外科治疗中建立皮下永久性胆管通路的方法及临床价值。方法 回顾性分析1984年12月至2008年12月成都军区总医院全军普通外科中心收治的2278例肝胆管结石病人的临床资料。均通过外科手术治疗并在术中设置了皮下永久性胆管通路。对128例区域性肝胆管结石病人采用皮下通道型胆囊肝总管吻合建立皮下永久性胆管通路。结果 2278例病人中,获随访792例。随访中,胆管结石复发者184例（占23.2%）,均施行了皮下胆管通路切开、胆道镜取石术。其中,切开皮下空肠袢175例,切开皮下胆囊9例。对于同时伴有吻合口狭窄的43例病人,在胆道镜下采用胆道气囊进行扩张,狭窄得到了不同程度的纠正。全部病人均病愈出院。结论 在肝胆管结石病人手术中,建立皮下永久性胆管通路是治疗术后复发结石的重要步骤。对于区域性肝胆管结石病人,术中利用胆囊建立皮下永久性胆管通路不失为简单、可靠的方法。     

Ki-67蛋白和p53蛋白在肝胆管上皮内瘤变及癌变诊断中的意义

目的：研究增殖指标Ki-67蛋白和p53蛋白在人肝胆管上皮内瘤变及癌变中的表达及其诊断意义。
方法：运用免疫组织化学S-P方法检测70例肝胆管上皮内瘤变及癌变（低级别肝胆管上皮内瘤变42例,高级别上皮内瘤变28例,高级别中癌变16例）组织中Ki-67蛋白和p53蛋白的表达情况。
结果：（1）肝胆管上皮癌变组织中Ki-67的增殖指数为20.65±7.62,明显高于肝胆管上皮内瘤变（P＜0.05）,并随肝胆管上皮增生程度的增加其增值指数也增高。（2）在16例肝胆管癌变组织中p53蛋白的增值指数为14.73±5.57,明显高于胆管上皮内瘤变组织的蛋白表达（P＜0.05）。
结论：从肝胆管上皮内瘤变到癌变这一发展过程中,Ki-67及p53蛋白增值指数随着细胞增生程度而增高,为临床判断病变程度提供了较具体可靠的参考依据。

     

肝胆管结石病的诊治原则

肝胆管结石病在我国是常见病。诊断主要是在临床表现基础上结合影像学检查,超声、CT和磁共振常联合使用。肝胆管结石病的治疗是普通外科良性疾病中的难题,手术是治疗肝胆管结石病的主要手段,基本原则是去除病灶、取尽结石、矫正狭窄、通畅引流、防治复发。随着微创外科迅猛发展,腹腔镜治疗肝胆管结石病也日趋成熟。运用影像学手段术前精确评估,制定个体化治疗方案,才能为肝胆管结石病患者带来最佳治疗。     

Toll样受体在介导免疫及炎症反应中的作用

Toll样受体是近年发现的在天然免疫及炎症反应中起重要作用的一类跨膜受体,对Toll样受体的认识主要源自 对TLR2和TLR4在介导内毒素LPS信号转导中作用的研究。本文就Toll和Toll样受体的结构、成员、信号通路及其在免疫及 炎症反应中的作用作一综述。     

Toll样受体在介导免疫及炎症反应中的作用

Toll样受体是近年发现的在天然免疫及炎症反应中起重要作用的一类跨膜受体，对Toll样受体的认识主要源自对TLR2和TLR4在介导内毒素LPS信号转导中作用的研究。本文就Toll和Toll样受体的结构、成员、信号通路及其在免疫及炎症反应中的作用作一综述。     

小剂量脂多糖诱导血管内皮细胞TLR4的表达

目的:观察小剂量脂多糖(LPS)对人脐静脉内皮细胞(ECV304)TOLL样受体4(TLR4)表达的影响。方法:体外培养ECV304细胞,分别与LPS及LPS+TLR4抗体进行孵育。MTT法检测细胞的增殖活性,免疫组化染色法检测细胞表面TLR4的表达,实时荧光定量聚合酶链反应(RT-PCR)检测细胞核核内TLR4-mRNA及IL-8mRNA的表达。结果:LPS(10～50ng/mL)刺激ECV304细胞24h内,细胞增殖活性无明显变化(P>0.05);而以100ng/mL刺激24h后,细胞增殖活性明显降低(P<0.05),TLR4抗体对此无明显拮抗作用。LPS能明显上调ECV304表达TLR4、TLR4-mRNA及IL-8mRNA,其中10ng/mL的LPS在24h时、50ng/mLLPS在6～24h时,TLR4表达具有统计学意义(P<0.05);50ng/mL的LPS刺激ECV304细胞在4h及8h时,细胞核内TLR4mRNA表达均明显升高(P<0.05),而IL-8mRNA表达在8h时明显升高(P<0.05)。TLR4抗体对ECV304表达TLR4及TLR4-mRNA有拮抗作用(P<0.05),对IL-8mRNA表达无明显拮抗作用(P>0.05)。结论:小剂量LPS可诱导ECV304细胞表达TLR4并引起细胞活化,TLR4抗体可抑制TLR4的表达,但不能抑制细胞的活化。     

Heme oxygenase-1 induction mitigates burn-associated early acute kidney injury via the TLR4 signaling pathway

ObjectivesEarly acute kidney injury (AKI) after burn contributes to disastrous prognoses for severely burned patients. Burn-induced renal oxidative stress and secondary proinflammatory mediator release contribute to early AKI development, and Toll-like receptor (TLR) 4 regulates inflammation. Heme oxygenase-1 (HO-1) is a stress-responsive enzyme that plays a vital role in protecting against ischemia-induced organ injury via its antioxidant properties and regulation of inflammation. We investigated the potential effect of HO-1 induction in preventing burn-induced early AKI and its related mechanism.MethodsA classic major-burn rat model was established using a 100 °C water bath, and hemin was injected intraperitoneally immediately after the injury to induce HO-1. Histological staining and blood tests were used to assess AKI progression based on structural changes and function. Renal levels of HO-1, oxidative stress, proinflammatory mediators and TLR4-related signals were detected using ELISA, immunostaining, qRT-PCR, and western blotting. The selective TLR4 inhibitor TAK242 and TLR4 inducer LPS were introduced to determine the roles of HO-1 in burn-related renal inflammation and the TLR4 pathway.ResultsHemin improved burn-induced renal histological damage and dysfunction, and this beneficial effect was related to reduced renal oxidative stress and the release of proinflammatory mediators, such as tumor necrosis factor-alpha (TNF-α), interleukin (IL)-1β, IL-6 and intracellular adhesion molecule-1 (ICAM-1). Hemin downregulated the expression of TLR4 and the subsequent phosphorylation of IKKα/β, IκBα, and NF-κB p65;. TAK242 exerted an effect similar to but weaker than hemin; and LPS reversed the antiinflammatory effect of hemin and the regulation of TLR4 signals. These results suggested that the TLR4 signaling pathway mediated the HO-1-facilitated regulation of renal inflammation after burn.ConclusionThe present study demonstrated that HO-1 induction prevented burn-induced early AKI by targeting renal inflammation, which was mediated via regulation of the TLR4/NF-κB signaling pathway.     

Toll‐like receptors and their adaptors are regulated in macrophages after phagocytosis of lipopolysaccharide‐coated titanium particles

Macrophages phagocytose metallic wear particles and produce mediators, which can induce cellular host response and aseptic implant loosening. Lipopolysaccharide (LPS) on the wear debris can stimulate macrophages via Toll‐like receptor 4 (TLR4) and enhance the response. However, the precise functional role and interaction of TLRs and their adaptor molecules is still unclear. Rat bone marrow macrophages were stimulated with titanium particle (Ti) coated by LPS (Ti/LPS+) and LPS‐free Ti (Ti/LPS?). mRNA levels of cytokines, TLRs and their adaptor molecules were measured using real time PCR. mRNA levels of TNF‐α, IL‐1β, and IL‐6 increased in Ti/LPS+ than Ti/LPS?. In contrast, mRNA levels of TLR4, TLR5, and TLR9 decreased in Ti/LPS+ compared to Ti/LPS?. mRNA levels of MyD88, IRAK1, IRAK4 decreased gradually, and TRAF6 underwent an initial transient increase, followed by suppression in Ti/LPS+. However, mRNA levels of TLR2 and IRAK2 increased after phagocytosis of Ti/LPS+ than Ti/LPS?. The increased expressions of proinflammatory cytokines found in Ti/LPS+ indicated that their productions cytokines could be enhanced by phagocytosis of LPS‐coated particles. Subsequent down‐regulation of TLR4, TLR5, TLR9, MyD88, IRAK1, and IRAK4 suggests that self‐protective mechanisms to regulate excessive host responses are activated in macrophages. Increase of TLR2 and IRAK2 and a transient increase of TRAF6 in Ti/LPS+ suggest that another possible pathway to modulate TLR‐mediated cellular response to prolong inflammatory response in foreign body reaction of aseptic loosening. This down‐ and/or up‐regulation of the potential TLR‐mediated responses to LPS‐coated particles reflects the proactive behavior of effector cells. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29: 984–992, 2011     

Atorvastatin attenuates experimental contrast-induced acute kidney injury: a role for TLR4/MyD88 signaling pathway

Objectives: To investigate the protective effect of different atorvastatin doses on contrast-induced acute kidney injury and the related mechanism.

Methods: Healthy male Sprague–Dawley (SD) rats were randomly divided into the blank control group, experimental control group and different-dose atorvastatin groups. A rat model of contrast-induced acute kidney injury was established. We detected changes in serum creatinine (Scr) and blood urea nitrogen (BUN) before and after model establishment, observed and scored renal tubular injury, analyzed rat renal cell apoptosis, and measure the expression of signal pathway proteins and downstream inflammatory factors.

Results: After contrast agent injection, the Scr and BUN levels of the experimental control group were significantly increased, the different doses applied in the atorvastatin group significantly reduced the Scr and BUN levels (p?p?p?p?Conclusion: Different atorvastatin doses have protective effects on contrast-induced acute renal tubular injury in rats, possibly by targeting TLR4, suppressing TLR4 expression, regulating the TLR4/Myd88 signaling pathway, and inhibiting the expression of downstream inflammatory factors.     

Curcumin modulates TLR4/NF-κB inflammatory signaling pathway following traumatic spinal cord injury in rats

Abstract

Background

Curcumin, a polyphenolic compound extracted from the plant turmeric, has protective effects on spinal cord injury (SCI) through attenuation of inflammatory response. This study was designed to detect whether curcumin modulates toll-like receptor 4 (TLR4) and the nuclear factor-kappa B (NF-κB) inflammatory signaling pathway in the injured rat spinal cord following SCI.

Methods

Adult male Sprague–Dawley rats were subjected to laminectomy at T8–T9 and compression with a vascular clip. There were three groups: (a) sham group; (b) SCI group; and (g) SCI + curcumin group. We measured TLR4 gene and protein expression by real-time polymerase chain reaction and western blot analysis; NF-κB activity by electrophoretic mobility shift assay, inflammatory cytokines tumor necrosis factor-α, interleukin-1β, and interleukin-6 levels by enzyme-linked immunosorbent assay, hindlimb locomotion function by Basso, Beattie, and Bresnahan rating, spinal cord edema by wet/dry weight method, and apoptosis by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) analysis.

Results

The results showed that SCI induced the up-regulation of TLR4, NF-κB, and inflammatory cytokines in the injured rat spinal cord. Treatment with curcumin following SCI markedly down-regulated the levels of these agents related to the TLR4/NF-κB inflammatory signaling pathway. Administration of curcumin also significantly ameliorated SCI induced hind limb locomotion deficits, spinal cord edema, and apoptosis.

Conclusions

Post-SCI curcumin administration attenuates the TLR4/NF-κB inflammatory signaling pathway in the injured spinal cord, and this may be a mechanism whereby curcumin improves the outcome following SCI.     

Ischemic post-conditioning attenuates renal ischemic reperfusion injury via down-regulation of toll-like receptor 4 in diabetic rats

Background: Ischemia/reperfusion (I/R) injury, which is commonly seen in the field of renal surgery or transplantation, is a major cause of acute renal failure (ARF). The ischemic ARF in diabetic rats is much more severe than that in the normal rats exposed to as same ischemic time. Ischemic post-conditioning (IPO) is a phenomenon by which intermittent interruptions of blood flow in the early phase of reperfusion can protect organs from I/R injury. To determine whether the renal protection effect of IPO mediates by toll-like receptor 4 (TLR4) signaling pathway in diabetic rats.

Methods: Streptozotocin-induced diabetic rats were randomly divided into three groups: sham operation group, I/R group, and IPO group. Except sham operation group, rats were subjected to 30?min of renal ischemia, both with and without treatment with IPO, then reperfusion 24?h. Light microscope and transmission electronic microscope were used to observe structural changes of renal tubule. RT-PCR was used to measure TLR4 and tumor necrosis factor-alpha (TNF-α) mRNA expression level, renal TLR4 and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) protein expression was detected by Western blot.

Results: The results demonstrated that IPO markedly decreased renal ischemic injury caused by I/R and inhibited the proinflammatory expression levels of TLR4, TNF-α, and NF-κB, all of which up-regulated by I/R in diabetic rats.

Conclusion: Taken together, our results suggest that proper IPO may have protective effect on the ischemic injury mediated by renal I/R, which might be associated with inhibition of TLR4 signaling pathway in diabetic rats.     

拮抗Toll样受体4的表达对内毒素血症小鼠肾脏损伤的保护作用

目的观察拮抗Toll样受体4（TLR4）的表达对内毒素血症小鼠肾脏损伤的保护作用以及对细胞核因子-κB（NF-κB）p65的影响。 方法采用内毒素（LPS）诱导的小鼠急性肾功能衰竭模型,并应用TLR4单克隆抗体进行干预,观察TLR4单克隆抗体对内毒素血症小鼠的肾脏组织学、血清肌酐（Cr）、尿素氮（BUN）、胱抑素C、白细胞介素（IL）-1β、IL-6水平以及肾组织中TLR4及NF-κB水平的影响。 结果与模型组比较,TLR4单克隆抗体能够明显改善内毒素血症小鼠肾脏组织病理学损害;降低血清中Cr、BUN、胱抑素C、IL-β和IL-6水平（t = 7.20、7.86、9.99、8.79、3.92,P均< 0.05）,并且降低肾组织中TLR4及NF-κB p65水平（t = 20.94、11.21,P均< 0.05）。 结论TLR4单克隆抗体能够保护内毒素血症小鼠肾脏组织损伤,其作用机制可能与调节TLR4/NF-κB信号转导通路有关。