胆汁内外引流术对梗阻性黄疸大鼠肝脏Kupffer细胞CD14表达的影响

目的探讨胆汁内外引流术对梗阻性黄疸大鼠肝脏Kupffer细胞CD14表达的影响。方法将60只成年SD雄性大鼠随机分为4组,梗阻性黄疸组(OJ组)、胆汁内引流术组(ID组)、胆汁外引流术组(ED组)及假手术组(SH组)并分别建立模型;采用免疫组织化学染色及半定量分析的方法测定各组大鼠肝脏Kupffer细胞CD14的表达情况。结果 OJ组大鼠肝脏Kupffer细胞CD14表达明显强于SH组(P0.01),ID组与OJ组比较表达明显降低(P0.01),ED组与OJ组比较无显著性差异(P=0.5907)。结论梗阻性黄疸时胆汁内引流术在改善Kupffer细胞CD14表达方面优于胆汁外引流术,提示胆汁内引流术优于外引流术的机制可能与肝脏Kupffer细胞CD14表达过程有关。     

胆汁内外引流术对梗阻性黄疸大鼠肝脏诱导型一氧化氮合酶表达的影响

目的 探讨胆汁引流方法对梗阻性黄疸大鼠肝脏诱导型一氧化氮合酶(iNOS)表达的影响.方法 将42只成年雄性SD大鼠随机分为四组,采用我们既往建立的二次手术方法分别建立梗阻性黄疸(OJ,n=12)、胆汁内引流术(ID,n=10)、胆汁外引流术(ED,n=10)及假手术(SH,n=10)模型.于第二次术后7 天留取肝脏标本.通过SABC免疫组化方法测定肝脏组织iNOS的表达情况.结果 假手术组大鼠肝组织几乎无iNOS表达.梗阻性黄疸时,大鼠肝组织iNOS表达增强,阳性面积增加(30.539±6.255),平均光密度升高(0.296±0.055).通过胆汁内引流术解除黄疸后,肝脏iNOS表达明显受到抑制,阳性表达面积减少(16.571±2.044)(P=0.017),平均光密度下降(0.204±0.029)(P<0.01).但通过胆汁外引流术后,肝组织iNOS表达阳性面积(38.186±7.495)及平均光密度(0.399±0.086)非但不减少,反而显著增加(P<0.01,P＝0.004).结论 梗阻性黄疸时,大鼠肝脏iNOS表达增强,胆汁内引流术可明显抑制肝脏iNOS表达,而胆汁外引流术却使其表达增强,提示在抑制诱导型一氧化氮合酶的表达方面胆汁内引流术优于外引流术.     

胆汁内外引流术对梗阻性黄疸大鼠肺肿瘤坏死因子α、中性粒细胞弹性蛋白酶的影响

目的研究胆汁内外引流方法对梗阻性黄疸大鼠肺肿瘤坏死因子α(TNF-α)、中性粒细胞弹性蛋白酶(NE)水平的影响。方法将64只成年SD雄性大鼠随机分为4组,分别建立梗阻性黄疸(OJ)、胆汁内引流术(ID)、胆汁外引流术(ED)及假手术(SH)4组模型。于2次术后第14天留取肺组织匀浆液标本。采用双抗体夹心酶联免疫吸附法(ELISA)检测10%肺匀浆液TNF-α水平,生化法检测10%肺匀浆液NE水平。结果成功建立了大鼠梗阻性黄疸及内外引流术模型。梗阻性黄疸时大鼠肺TNF-α、NE水平较假手术对照组明显升高(100.893 pg/mL±21.271 pg/mL vs 64.091 pg/mL±13.034 pg/mL,P<0.01;50.396μg/mL±17.388μg/mL vs 39.718μg/mL±9.625μg/mL,P<0.05)。通过胆汁内引流术解除黄疸后,大鼠肺TNF-α浓度(75.141 pg/mL±15.849 pg/mL)与梗阻性黄疸组相比下降明显(P<0.01);而通过胆汁外引流术解除黄疸后,大鼠肺TNF-α浓度仍较高(112.129 pg/mL±36.886 pg/mL),与梗阻性黄疸组相比无差异(P>0.05)。行胆汁内、外引流术后,大鼠肺NE水平均降低(39.390μg/mL±12.410μg/mL、44.790μg/mL±16.681μg/mL),但与梗阻性黄疸组相比内引流明显(P<0.05)、外引流无差异(P>0.05),且内引流恢复至正常水平,与假手术对照组相比无差异(P>0.05)。结论梗阻性黄疸可导致肺组织炎症细胞因子升高,胆汁内引流术可明显改善梗阻性黄疸时肺组织炎症细胞因子水平、甚至接近正常,而胆汁外引流术没有改善肺炎症细胞因子水平,提示术前利用胆汁内引流术解除梗阻性黄疸缓解肺部炎症反应优于外引流术。     

胆道外引流术后胆汁回输对恶性梗阻性黄疸免疫功能的影响

在临床上，恶性梗阻性黄疸病人手术前后常常出现一些严重的并发症，如感染、多脏器功能衰竭等，其原因尚不明确，有作者认为免疫功能损害是重要因素之一。因此，我们观测了胆道外引流手术前后免疫功能的变化，报告如下。     

胆汁内、外引流术对梗阻性黄疸大鼠血清肿瘤坏死因子和肝脏库普弗细胞诱导型一氧化氮合酶表达的影响

目的 研究胆汁内、外引流术对梗阻性黄疸大鼠血清肿瘤坏死因子-α(TNF-α)水平和肝脏库普弗细胞诱导型一氧化氮合酶(iNOS)表达的影响.方法 采用成年雄性Sprague-Dawley大鼠48只,分成梗阻性黄疸、胆汁外引流、内引流和假手术四组,每组12只.采用原位灌注消化肝脏及贴壁培养方法 分离并纯化库普弗细胞,采用逆转录(RT)-PCR方法 检测库普弗细胞iNOS mRNA表达.用ELlSA方法 测定血清TNF-α含量.结果 梗阻性黄疸组血清TNF-α水平为(110.8±26.3)pg/ml,与假手术组的(88.4±17.9)pg/ml比较,差异有统计学意义(P=0.045).内引流组解除胆道梗阻后,血清TNF-α水平受到抑制,为(89.84±28.3)pg/ml,而胆汁外引流组却无此作用,为(118.6±22.7)Pg/ml,后者与梗阻性黄疸组比较差异无统计学意义(P=0.059).胆道梗阻形成后,梗阻性黄疸组库普弗细胞iNOSmRNA表达增强(0.82±0.24),显著高于假手术组(0.38±0.35,P=0.005).胆汁内引流术解除黄疸后,内引流术组的库普弗细胞iNOS mRNA表达受到抑制(0.59±0.35),但与梗阻性黄疸组比较差异无统计学意义(P=0.139),外引流组iNOSmRNA表达并未受抑制(0.974±0.48),与梗阻性黄疸组比较差异无统计学意义(P=0.321),但显著高于胆汁内引流组(P=0.016).结论 胆汁内引流术在逆转梗阻性黄疸大鼠升高的血清TNF-α水平和肝脏库普弗细胞iNOS mRNA的表达方面优于胆汁外引流术.     

术前不同引流方式减黄对梗阻性黄疸大鼠部分肝切除术后肝功能和肝再生的影响

目的 了解术前不同引流方式减黄对梗阻性黄疸(OJ) SD大鼠部分肝切除术(PH)术后肝功能和肝再生的影响.方法 建立OJ不同引流方式减黄70％部分肝切除SD大鼠动物模型.并在术后0、1、2、4、12、24、48和72h收集大鼠血液及肝脏组织标本,测定血清TBIL、ALB、ALT、AST水平,计算残肝重量、肝再生率,免疫组化法观察肝脏组织PCNA表达,ELISA法检测血清TNF-α水平.结果 PH术后各时段内引流(ID)组和外引流(ED)组TBIL、ALT、AST水平较OJ组均偏低.各时段ID组ALB水平较OJ组、ED组偏高.72 h肝再生率ID组高于ED组,ED组高于OJ组.3组PCNA水平均于12 h明显升高,ID组于24 h达高峰,OJ组、ED组高峰延迟至48 h且峰值偏低.PH术后各组血清TNF-α水平均呈上升趋势,ID组于12 h达高峰,OJ组、ED组均于24 h达高峰,各时段OJ组、ED组血清TNF-α水平较ID组均偏高.结论 内外引流术均可改善OJ所致的高胆红素血症和肝功能,并改善OJ大鼠残肝再生能力,但内引流效果更明显,且内引流术可以有效降低血清TNF-α水平.     

Short-term effects of external and internal biliary drainage on liver and cellular immunity in experimental obstructive jaundice

Background/Purpose

The effects of preoperative biliary drainage for obstructive jaundiced patients are controversial. Although experimental studies have proven the benefit of internal biliary drainage (ID) over external biliary drainage (ED), ID has several clinical problems, such as clogging or tube replacement. The aim of this study was to determine whether there were any differences in T-cell function, liver function, and histology, between rats in ID and ED groups in short-term experiments.

Methods

Following bile duct ligation (BDL) for 14 days, rats in the ED and ID groups had 7 days of ED and 7 days of ID, respectively. Normal rats were used as negative controls (control group). For positive controls, we used a group with BDL and no drainage (BDL group). Serum bilirubin, aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) were measured, splenic T-cell proliferation was assayed to check cellular immunity, and liver histology was examined.

Results

Recovery of bilirubin and ALT was similar in the ED and ID groups. Recovery of AST was worse in the ID group than in the ED group, but the difference was not statistically significant. Levels of ALP in the BDL and ID groups were significantly higher than those in the control and ED groups. Rats in the BDL group showed a significant decrease in T-cell function compared to the control group. The ED group showed better recovery of T-cell function than the ID group in the 7 days after relief of obstructive jaundice. The livers in the ID group demonstrated histologically moderate interface hepatitis with periportal inflammation and lymphocyte infiltration, which strongly suggested incomplete tube obstruction, but those in the ED group showed minimal change.

Conclusions

ED is superior to ID concerning the recovery of cellular immunity and liver inflammation in the short-term after relief from biliary obstruction in this model. As the patency of the tube is well maintained in ED compared to ID, patency of the tube is essential to obtain good recovery of cellular immunity, irrespective of the drainage method.

     

不同胆道引流方式对梗阻性黄疸大鼠肠屏障功能的影响

目的:探讨不同胆道引流方式对梗阻性黄疸大鼠肠屏障功能的影响.方法:60只SD大鼠随机分为假手术组(shame operation,SO组)、梗阻性黄疸组(obstructive jaundice,OJ组)、内引流组(internal biliary drainage,ID组)和外引流组(enternal biliary drainage,ED组).制备OJ模型,SO组、OJ组在术后第7天处死大鼠并采集标本,ID组、ED组则行不同引流手术,在引流术后第7天处死大鼠并采集标本.检测血清丙氨酸氨基转移酶(alanine aminotransferase,ALT)、总胆红素(total bilirubin,TBIL)、直接胆红素(direct bilirubin,DBIL)、胆汁酸(totalbileacide,TBA)及小肠黏液sIgA含量,并对结果进行统计学分析.结果:OJ组较SO组血清ALT、TBIL、DBIL、TBA均明显升高,小肠黏液sIgA含量(0.160±0.150)明显减少.ED、ID组血清中ALT、TBIL、DBIL、TBA明显低于OJ组;ID组小肠黏液sIgA含量明显高于OJ组及ED组(0.272±0.182vs0.160±0.150,0.191±0.113,均P<0.05).结论:胆道内、外引流均能有效解除胆道梗阻,改善肝功能.但内引流可明显增加肠道黏液sIgA,其改善肠道屏障功能的效果优于外引流.     

Endotoxin, cytokines, and endotoxin binding proteins in obstructive jaundice and after preoperative biliary drainage

BACKGROUND: Obstructive jaundice is associated with postoperative complications related to increased endotoxaemia and the inflammatory response. In animals obstructive jaundice is associated with endotoxaemia and cytokine induction, which are reversed by internal biliary drainage. AIMS: To study endotoxaemia and the subsequent inflammatory response in obstructive jaundiced patients and after endoscopic biliary drainage. METHODS: In 15 patients with malignant distal obstructive jaundice, inflammatory and bacteriological parameters were assessed before endoscopic stent placement and after three weeks endoscopic drainage. RESULTS: Drainage reduced bilirubin from 252.5 to 45.1 micromol/l. At baseline low level endotoxaemia was detected (4.3 pg/ml) which was not affected after drainage (4.5 pg/ml). Serum interleukin 8 (IL-8) and endotoxin binding proteins were increased in jaundice and reduced after drainage (IL-8 113.6 to 20.7 pg/ml; lipopolysaccharide binding protein 24.2 to 16.5 microg/ml; sCD14 17.4 to 7.6 microg/ml; bactericidal/permeability increasing protein 2.9 to 1.8 ng/ml). Levels of other cytokines, augmented in animals, were only slightly increased and not changed after drainage (tumour necrosis factor (TNF): 21.7 and 18.4 pg/ml; sTNFr p55/75: 2.9/7.0 and 2.7/5.6 ng/ml; IL-6: 4.2 and 6.1 pg/ml; IL-10: 4.5 and 2.7 pg/ml). Elastase and lactoferrin tended towards reduction after drainage. All bile cultures were positive after stenting. CONCLUSIONS: The effects of obstructive jaundice in humans on endotoxin and cytokines are different from those in animal models. Obstructive jaundice causes alterations in circulating endotoxin binding proteins and IL-8. Concentrations of other mediators (TNF, previously suggested as being responsible for systemic endotoxaemia effects) are low and not affected by drainage.     

一氧化氮和内毒素在梗阻性黄疸致病机制方面的作用

梗阻性黄疸（obstructive jaundice，OJ）是临床常见的症状。梗阻性黄疸患者术后并发症和死亡率较高，常死于感染和败血症，提示机体术前的免疫功能受到了抑制。梗阻性黄疸可使网状内皮系统功能降低，细菌清除能力减弱。肠道来源的细菌和内毒素逃脱肝脏枯否细胞的监视，进入血液循环。有研究发现梗阻性黄疸常并发肠源性内毒素血症，内毒素激活枯否细胞，产生大量炎性介质，如一氧化氮（NO）、肿瘤坏死因子α（TNFα）及氧自由基等。     

一氧化氮和内毒素在梗阻性黄疸致病机制方面的作用

梗阻性黄疸（obstructive jaundice）是临床常见的表现。梗阻性黄疸患者术后并发症和死亡率较高，常死于感染和败血症，提示机体术前的免疫功能受到了抑制。梗阻性黄疸可使网状内皮系统功能降低，细菌清除能力减弱。肠道来源的细菌和内毒素逃脱肝脏枯否细胞的监视，进入血液循环。有研究发现梗阻性黄疸常并发肠源性内毒素血症，内毒素激活枯否细胞，产生大量炎性介质，如一氧化氮（NO），肿瘤坏死因子α（TNFα）及氧自由基等。