【作者】 胡明秋；

【导师】 王鹏志； 刘彤； 李卫东；

【作者基本信息】 天津医科大学 ， 外科学， 2010， 博士

【摘要】 目的：探讨胆囊动力学异常在胆囊结石形成中的作用,为明确保胆取石手术适应证提供依据。方法：50例胆囊结石病人(病例组)和20例健康人(正常对照组),采用B超测定空腹胆囊壁厚度、空腹胆囊容积、餐后30min、60 min、90 min胆囊餐后残余容积,计算胆囊排空容积EV、胆囊残余指数RF和胆囊排空率E。病例组以正常对照组平均RF+2SD值(40.18%)为界限,分为胆囊收缩“正常”组(n=12)和胆囊收缩“减弱”组两个亚组(n=38)。结果：1)正常对照组胆囊壁厚度0.15±0.03cm,胆囊排空率(71.88±6.03)%。根据参数可信区间的计算方法,正常健康人胆囊壁厚度95%的可信区间为0.09-0.21 cm,胆囊排空率95%的可信区间为59.82%-83.94%。2)病例组胆囊壁厚度0.31±0.05cm,胆囊排空率(48.65±15.81)%,与正常对照组比较均有显著差异(P<0.05)。3)胆囊结石患者胆囊收缩“正常”组胆囊壁厚度0.28±0.05 cm,胆囊收缩“减弱”组胆囊壁厚度0.32±0.05 cm,与正常对照组比较,均有显著差异(P<0.05),胆囊收缩“正常”组与胆囊收缩“减弱”组胆囊壁厚度比较也有显著差异(P<0.05)。4)胆囊结石患者胆囊收缩“减弱”组与胆囊收缩“正常”组比较,虽然FV没有明显差异(P>0.05),但胆囊收缩“减弱”组最小残余容积增大(P<0.05),胆囊排空率降低(P<0.05)。胆囊结石患者胆囊收缩“正常”组与正常对照组比较,虽然排空率没有明显差异,但胆囊收缩“正常”组空腹胆囊容积明显增大(P<0.05)。5)病例组胆囊结石患者的胆囊排空率与胆囊壁厚度二者呈显著负相关,Y=106.695-185.270X,r=-0.617,P=0.000；对照组胆囊排空率与胆囊壁厚度二者无相关,Y=77.846-38.986X,r=-0.184,P=0.437。结论：胆囊排空障碍是胆囊结石形成的动力学基础之一,为临床用药防止胆囊结石形成及复发、保胆取石术手术适应证的选择提供了理论依据。我们认为保胆取石术应具备的基本条件是胆囊排空率>50%,胆囊壁厚度≤0.3cm。目的：探讨胆囊结石患者胆囊收缩素(CCK)-A受体mRNA转录水平和受体分布、定位及蛋白表达水平。方法：50例胆囊结石患者(病例组)和20例无胆囊结石肝移植供体(供体组)胆囊标本,采用RT-PCR方法测定胆囊标本CCK-A受体分子mRNA转录水平,采用酶免疫组织化学法研究胆囊CCK-A受体的分布和定位,测定胆囊标本CCK-A受体分子蛋白表达水平。结果：1)供体组胆囊CCK-A受体分子mRNA转录水平0.8182±0.0481,病例组mRNA转录水平0.6465±0.0910,两组比较有明显差异(P<0.01)。2)胆囊CCK-A受体分子不仅在胆囊平滑肌细胞表达,而且在腺上皮细胞也有明显表达。蛋白表达阳性细胞中,细胞核与细胞浆均有表达。3)与供体组相比,病例组免疫组化染色5个高倍视野中腺上皮细胞、平滑肌细胞蛋白表达阳性细胞数明显减少,两组比较有显著差异(P<0.01)。4)供体组胆囊CCK-A受体分子蛋白表达水平(蛋白表达阳性细胞总数)和mRNA转录水平呈正相关,Y=-458.027+975.883X,r=0.741,P=0.000；病例组胆囊CCK-A受体分子蛋白表达水平(蛋白表达阳性细胞总数)和mRNA转录水平也呈正相关,Y=-43.193+414.018X,r=0.644,P=0.000。结论：1)胆囊收缩素与胆囊CCK-A受体结合,不仅调节胆囊平滑肌收缩,而且还调节胆囊腺上皮细胞的分泌功能。2)胆囊CCK-A受体属于胞核和胞质受体(胞内受体)。3)胆囊结石患者CCK-A受体分子mRNA转录水平和蛋白表达水平较低。4)胆囊结石患者胆囊CCK-A受体分子蛋白表达水平降低与mRNA转录水平下调是一致的。目的：探讨胆囊结石患者胆囊CCK-A受体分子mRNA转录水平和蛋白表达水平对胆囊收缩功能的影响。方法：同第二部分研究。结果：1)将病例组患者的两个亚组(胆囊收缩“正常”组12例和收缩“减弱”组38例)进行比较,胆囊收缩“减弱”组CCK-A受体mRNA转录水平显著低于胆囊收缩“正常”组(0.6279±0.0900比0.7055±0.0680,P<0.01)；即使是胆囊收缩“正常”的胆石病患者,胆囊CCK-A受体mRNA转录水平也较无胆囊结石的供体组减少(0.7055±0.0680比0.8182±0.0481,P<0.01)。2)供体组20例,胆囊平滑肌细胞蛋白表达阳性7例、强阳性13例,而病例组50例,平滑肌细胞蛋白表达弱阳性24例、阳性22例、强阳性4例,二者比较有显著差异(P<0.01)。病例组和供体组胆囊平滑肌CCK-A受体蛋白表达阳性细胞数比较也有显著差异(P<0.01)。将病例组的两个亚组进行比较,胆囊收缩“减弱”组胆囊平滑肌CCK-A受体分子蛋白表达阳性强度和阳性细胞数显著低于无胆囊结石的供体组(P<0.01);即使是胆囊收缩“正常”的胆囊结石患者,胆囊平滑肌CCK-A受体分子蛋白表达阳性强度和阳性细胞数也较无胆囊结石的供体组低(P<0.05)。胆囊收缩“减弱”组和胆囊收缩“正常”组胆囊平滑肌CCK-A受体分子蛋白表达水平比较,表达阳性强度有显著差异(P<0.05),但蛋白表达阳性细胞数无差异(P>0.05)。3)病例组胆囊CCK-A受体mRNA转录水平与胆囊壁厚度二者呈显著负相关,Y=0.965-1.019X,r=-0.582,P=0.000;病例组胆囊平滑肌CCK-A受体蛋白表达水平(平滑肌蛋白表达阳性细胞总数)与胆囊壁厚度二者呈显著负相关,Y=193.784-246.306X,r=-0.419,P=0.002.4)病例组胆囊排空率与胆囊CCK-A受体mRNA转录水平二者呈显著正相关,Y=-37.558+133.493X,r=0.778,P=0.000；病例组胆囊排空率与胆囊平滑肌CCK-A受体蛋白表达水平(平滑肌蛋白表达阳性细胞总数)二者呈显著正相关Y=13.725+0.300X,r=0.587,P=0.000。结论：胆囊结石患者CCK-A受体分子1nRNA转录水平和蛋白表达水平与胆囊壁厚度、胆囊排空率存在数量上的相关关系,CCK-A受体分子mRNA转录水平和蛋白表达水平的差异可能是影响胆囊收缩功能的关键因素。目的：探讨胆囊结石患者胆汁胆固醇浓度对胆囊CCK-A受体分子mRNA转录水平和蛋白表达水平的影响。方法：采用酶法测定胆汁胆固醇浓度,其余方法同第二、三部分研究。结果：1)供体组胆汁胆固醇浓度6.68±1.75mmol/L,病例组胆汁胆固醇浓度9.40±2.33mmol/L,二者比较有显著差异(P<0.01)。将病例组患者的两个亚组进行比较,胆囊收缩“减弱”组胆汁胆固醇浓度显著低于胆囊收缩“正常”组,P<0.05,但胆囊收缩“正常”组胆汁胆固醇浓度和无胆囊结石的肝移植供体供体组比较,无显著差异,P>0.05。2)胆囊壁厚度与胆汁胆固醇浓度二者呈显著正相关,Y=0.186+0.014X,r=0.608,P=0.000.3)病例组胆囊排空率与胆汁胆固醇浓度二者呈显著负相关,Y=98.686-5.314X,r=-0.795,P=0.000。4)供体组CCK-A受体mRNA转录水平与胆汁胆固醇浓度二者呈显著负相关,Y=0.961-0.021X,r=-0.776,P=0.000:病例组CCK-A受体mRNA转录水平与胆汁胆固醇浓度二者也呈显著负相关,Y=0.934-0.031X,r=-0.786,P=0.000。5)供体组胆囊CCK-A受体总蛋白表达水平(腺上皮和平滑肌细胞蛋白表达阳性细胞总数)与胆汁胆固醇浓度呈负相关,Y=523.268-27.358X,r=-0.757,P=0.000；病例组胆囊CCK-A受体总蛋白表达水平(腺上皮和平滑肌细胞蛋白表达阳性细胞总数)与胆汁胆固醇浓度也呈负相关,Y=365.440-15.000X,r=-0.599,P=0.000.6)供体组胆囊平滑肌CCK-A受体蛋白表达水平(平滑肌蛋白表达阳性细胞数)与胆汁胆固醇浓度呈负相关,Y=301.824-19.120X,r=-0.667,P=0.001;病例组胆囊平滑肌CCK-A受体蛋白表达水平(平滑肌蛋白表达阳性细胞数)与胆汁胆固醇浓度也呈负相关,Y=190.866-7.887X,r=-0.603,P=0.000.结论：1)胆囊结石患者的胆汁胆固醇浓度高于无胆囊结石的人,尤其是胆囊收缩功能差的胆囊结石患者,胆汁胆固醇浓度明显升高。2)胆囊的慢性炎症与胆汁胆固醇浓度有关。3)无论是胆囊结石患者,还是非胆囊结石的肝移植供体,胆囊CCK-A受体分子mRNA转录水平和蛋白表达水平随着胆汁胆固醇浓度的高低而下调或上调,表明胆汁胆固醇可能是胆囊收缩功能障碍的始动因素,降低胆汁高胆固醇浓度可以促进胆囊动力功能的恢复,为预防结石的形成或复发提供了理论基础。更多还原

【Abstract】 Objective:To find the role of abnormal gallbladder dynamics in gallstone formation and to provide evidence for choosing indications of gallstone removal with gallbladder preservation surgery. Methods:50 cases of gallstone patients (patient group) and 20 healthy individuals (control group) were chosen. Type-B ultrasonic was adopted to measure gallbladder wall thickness and gallbladder full volume (FV) for patients under starvation condition, and measure gallbladder residual volume (RV) for patients at 30 minutes (min),60 min, and 90 min post-meal. The minimum RV (RVmin) was chosen. Gallbladder emptying volume (EV)=FV-RVmin. Gallbladder residual factor (RF)=(RV/FV)×100%. Gallbladder emptying efficiency (E)=(1-RF). By defining average RF+2SD (40.18%) in control group as a baseline, patient group had been divided into "normal" gallbladder contraction group (n=12) and "weakening" gallbladder contraction group (n=38). Results:1) Gallbladder wall thickness of control group was 0.15±0.03 cm. Gallbladder emptying efficiency in control group was (71.88±6.03)%. According to confidence interval calculation of parameter, gallbladder wall thickness in control group was 0.09～0.21 cm with 95% confidence interval. Gallbladder emptying efficiency in control group was 59.82%～83.94% with 95% confidence interval.2) Gallbladder wall thickness of patient group was 0.31±0.05 cm. Gallbladder emptying efficiency in patient group was (48.65±15.81)%. Gallbladder wall thickness and emptying efficiency were significantly different between patient group and control group (P<0.05).3) Gallbladder wall thickness of "normal" gallbladder contraction group was 0.28±0.05 cm,and wall thickness of "weakening" gallbladder contraction group was 0.31±0.05cm. In comparison of two sub-groups, "normal" and "weakening" groups, with control group, there was significant difference (P<0.05). There was also significant difference between "weakening" gallbladder contraction group and "normal" group (P<0.05).4) Compare to "normal" group, FV had no significant change, but RV increased (P<0.05), gallbladder emptying efficiency (E%) decreased (P<0.05), EV decreased significantly (P<0.05) in "weakening" group. Comparing the "normal" group to control group, although E had no significant change, FV increased dramatically (P<0.05).5)There is significant negative correlation between gallbladder emptying efficiency and gallbladder wall thickness in patient group (Y=106.695-185.270X, r=-0.617, P=0.000),while there was no significant correlation between them within control group (Y=77.846-38.986X, r=-0.184, P=0.437). Conclusion:The gallbladder emptying dysfunction is the fundamental for gallstone formation, and offered theoretical evidence for choosing chemotherapy to prevent gallstone formation and relapse and selecting indications of gallstone removal with gallbladder preservation surgery. Thus, we concluded that the prerequisite for gallstone removal with gallbladder preservation surgery are gallbladder emptying efficiency>50%, and gallbladder wall thickness≤0.3 cm.Objective:To study the CCK-A receptor’s location and its protein production and mRNA expression levels in gallstone patients. Methods:50 cholesterol calculus patients (patient group) and 20 liver transplant provider (provider group) were chosen. CCK-A receptor mRNA transcription level between patient and provider groups had been tested by reverse transcription (RT)-PCR, and CCK-A receptor’s location and its protein production level between patient and provider groups had been tested by immunohistology (IH). Results:1) The level of CCK-A receptor mRNA transcription is 0.6465±0.0910 in patient group, and the mRNA expression is 0.8182±0.0481 in provider group; There is significant difference between the two groups (P<0.01).2) Gallbladder CCK-A receptor was not only expressed in gallbladder smooth muscle cells, but also expressed in gallbladder epithelial cells. CCK-A receptor was present in both nucleus and cytoplasm within CCK-A receptor positive expressed cell.3) Compare to provider group, number of CCK-A receptor positive expressed gallbladder epithelial and gallbladder smooth muscle cells had decreased dramatically under 5 high power field (hpf) within patient group.4) There is positive correlation between CCK-A receptor protein production level (total CCK-A positive expressed cell number) and mRNA transcription level in provider group (Y=-458.027+ 975.883X, r=0.741, P=0.000), and there is the same type of correlation in patient group as well(Y=-43.193+414.018X, r=0.644, P=0.000). Conclusion:1) The binding of CCK-A and CCK-A receptor can not only adjust gallbladder smooth muscle contraction, but also secretory function of gallbladder epithelial cells.2) CCK-A receptor belongs to both nucleus and cytoplasm receptor.3) CCK-A receptor protein production level and mRNA transcription level are both low in gallstone patient.4) Decreased level of CCK-A receptor protein production level is consistent with decreased mRNA transcription level within gallstone patient.Objective:To study the effect of levels of CCK-A receptor mRNA transcription and protein production on gallbladder contraction in gallstone patients. Methods: The methods are the same as partⅠand partⅡ. Results:1) CCK-A receptor mRNA transcription level in "weakening" gallbladder contraction group was significantly lower than "normal" gallbladder contraction group (0.6279±0.0900 vs 0.7055±0.0680, P<0.01); CCK-A receptor mRNA transcription level was still lower than that of provider group (0.7055±0.080 vs 0.8182±0.0481, P<0.01) even in "normal" group.2) Within 20 cases in provider group, there were 7 regular CCK-A receptor positive expressed smooth muscle cells, and 13 strong CCK-A receptor positive expressed smooth muscle cells, within 50 cases in patient group, there were 24 weak CCK-A receptor positive expressed smooth muscle cells,22 regular CCK-A receptor positive expressed smooth muscle cells, and 4 strong CCK-A receptor positive expressed smooth muscle cells, while there are significant difference between patient and provider groups (P<0.01). CCK-A positive expressed cell number between patient and provider groups are also significantly different (P<0.01). CCK-A receptor protein production level and positive expressed cell number of gallbladder smooth muscle in "weakening" gallbladder contraction group are significantly lower than that of provider group(P<0.01); Even within "normal" gallbladder contraction group, CCK-A receptor protein production level and positive expressed cell number are significantly lower than that of provider group (P<0.05). There are significant difference of CCK-A receptor protein production level (P<0.05),but there are no significant difference of CCK-A receptor protein positive expressed cell number between "normal" and "weakening" groups (P>0.05).3)There is negative correlation between CCK-A receptor mRNA transcription level and gallbladder wall thickness in patient group (Y=0.965-1.019X, r=-0.582, P=0.000).4) There is negative correlation between CCK-A receptor protein production level and gallbladder wall thickness in patient group (Y=193.784-246.306X, r=-0.419, P=0.002).5) There is positive correlation between gallbladder emptying efficiency and CCK-A receptor mRNA transcription level in patient group (Y=-37.558+133.493X, r=0.778, P= 0.000); There is positive correlation between gallbladder emptying efficiency and CCK-A receptor protein production level in gallbladder smooth muscle (CCK-A positive expressed cell number in gallbladder smooth muscle) in patient group (Y=13.725+0.300X, r=0.587, P=0.000). Conclusion:1) There is quantitative relationship between levels of CCK-A receptor protein production and mRNA transcription and gallbladder thickness and emptying efficiency within gallstone patients, the difference between CCK-A receptor protein production level and mRNA transcription level may determine the extent of gallbladder contraction dysfunction.Objective:to study the effect of biliary cholesterol concentration on CCK-A receptor mRNA transcription and protein production in gallstone patients. Methods: The biliary cholesterol concentration had been tested by biochemistry. The other methods are the same as partⅠand partⅡ. Results:1)Biliary cholesterol concentration is 6.68±1.75 mmol/L, in provider group, and is 9.40±2.33 mmol/L, in patient group; There is significant difference between the two groups (P<0.01). In comparison of two sub-groups, "normal" and "weakening" groups, with provider group, there was significant difference between "weakening" gallbladder contraction group and provider group (P<0.05), but no difference between "normal" gallbladder contraction group and provider group (P>0.05); 2)There is significant positive correlation between gallbladder wall thickness and biliary cholesterol concentration (Y=0.186+0.014X, r=0.608, P=0.000) in patient group; 3) There is significant negative correlation between gallbladder evacuation efficiency and biliary cholesterol concentration in patient group (Y=98.686-5.314X, r=-0.795, P=0.000); 4) there is significant negative correlation between CCK-A receptor mRNA transcription level and biliary cholesterol concentration in both provider group (Y=0.961-0.021X, r=-0.776, P=0.000), and patient group (Y=0.934-0.031X, r=-0.786, P=0.000); 5)There is significant negative correlation between CCK-A receptor protein production(total CCK-A positive expressed cell number)and biliary cholesterol concentration in provider group (Y=523.268-27.358X, r=-0.757,P=0.000); There is significant negative correlation between CCK-A receptor protein production(total CCK-A positive expressed cell number)and biliary cholesterol concentration in patient group (Y=365.440-15.000X, r=-0.599, P=0.000).6) There is significant negative correlation between CCK-A receptor protein production level in gallbladder smooth muscle (CCK-A positive expressed cell number in gallbladder smooth muscle) and biliary cholesterol concentration in provider group (Y=301.824-19.120X, r=-0.667, P=0.001); There is significant negative correlation between CCK-A receptor protein production level in gallbladder smooth muscle (CCK-A positive expressed cell number in gallbladder smooth muscle) and biliary cholesterol concentration in patient group (Y=190.866-7.887X, r=-0.603, P=0.000). Conclusion:1) Biliary cholesterol concentration from gallstone patients is significantly higher than that of healthy individuals, especially the patients with weak gallbladder contraction function, whose biliary cholesterol concentrations increase dramatically.2) Gallbladder chronic inflammation is related to biliary cholesterol concentration.3) Within both gallstone patients and liver transplant provider, CCK-A receptor mRNA expression and protein production levels increase or decrease as biliary cholesterol concentration increases or decreases. Thus, biliary cholesterol is the initiative factor for gallbladder contraction dysfunction, decreased biliary cholesterol concentration can improve the recovery of gallbladder dynamic function, and provides theoretical basis for preventing gallstone formation and relapse.更多还原