β-葡萄糖醛酸酶在胆囊结石形成中的意义
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摘要
目的 胆汁中有两种不同来源的β-葡萄糖醛酸酶(β-G)：内源性β-G和外源性β-G。两者均可以水解胆汁中的结合胆红素(BDG)为游离胆红素(UCB)，后者是胆石的主要成分之一。目前普遍认为β-G在胆管胆色素结石的形成中有重要作用。本研究通过对不同组别胆囊胆汁中的β-G活性、总胆红素浓度(TB)、UCB浓度等指标进行分析，探讨β-G是否影响胆囊胆固醇结石的形成，并分析内、外源性β-G在结石形成中的关系。方法 收集2003年10月-12月在安徽医科大学第一附属医院行胆囊切除术患者的胆囊及其胆汁，根据病理报告分为胆囊息肉组(PO)和胆囊结石组；后者再根据病理及临床分为慢性炎症合并结石组(CⅠ)和急性炎症合并结石组(AⅠ)。PO组：共11例，其中男5例，女6例；年龄范围30-71岁。CⅠ组：共11例，其中男5例，女6例；年龄范围28-72岁。AⅠ组：共6例，其中男3例，女3例；年龄范围32-68岁。全部患者排除恶性肿瘤及各型肝炎。CⅠ组的病理切片表现为炎症细胞以淋巴、单核细胞为主；AⅠ组则主要以中性粒细胞为主，且在临床上有急性胆囊炎症状、体征，术中也证实胆囊急性炎症明显。收集的胆汁标本检测TB、UCB以及内、外源性β-G活性；收集的胆囊结石进行化学分类；术中取胆囊底部组织一块，行HE染色，并确定胆囊病理类型；计算内、外源性β-G活性总值以及UCB／TB，其结果作为统计指标进行分析；收集患者的年龄、性别以及肝肾功能资料，与上述指标进行相关分析。结果PO、CⅠ和AⅠ三组之间的β-G活性总值、内源性β-G活性、外源性β-G活性、TB浓度以及UCB／TB有差异(P＜0.05)，三组之间UCB浓度的差异没有显著性意义(P＞0.05)。CⅠ组与PO组相比，胆汁中内、外源性β-G活性、β-G活性总值以及UCB／TB显著增高(P＜0.05)，TB浓度显著降低(P＜0.05)；两组之间UCB浓度的差异没有显著性意义；AⅠ组与CⅠ组相比，胆汁中内、外源性β-G活性、β-G活性总值显著增高(P＜0.05)；TB浓度显著降低(P＜0.05)；两组之间UCB浓度以及UCB／TB的差异没有显著性意义；由于存在感染，内、外源性
安徽医科大学硕士学位论文
卜G的活性均显著增高(尸<0.05)，并且，内、外源性p一G活性与p一G活性总值
三者之间均呈正相关(P<0.05);本研究组所有的结石均为胆固醇结石;患者的
谷丙转氨酶(ALT)与p一G活性总值以及内、外源性p一G活性均呈正相关。结论
在胆囊结石患者的胆汁中，存在病理性增多的p一G。因为存在感染，内、外源性
p一G的活性均显著升高并呈正相关;p一G的水解作用致使胆汁中TB浓度下降;
UCB浓度也呈下降趋势。患者的ALT与p一G活性相关，此结果是肝脏在炎症情
况下的生理性反应。总之，p一G加速BDG水解并生成大量UCB，后者通过相关
机制促进胆石形成。p一G对于胆囊胆固醇结石的形成有重要意义。
Objective: There are endogenous and bacterial β-Glucuronidases(β-G) in the bile. β-G can split the conjugated bilirubin into glucuronic acid and unconjugated bilirubin (UCB) in bile, the latter is one of the major components of gallstone. β-G play an important role in the formation of pigment stone in bile duct. The authors investigated the difference of total bilirubin content (TB) , UCB content and the activities of β-G in the gallbladder bile of three groups to confirm whether β-G is responsible for the formation of gallbladder cholesterol stone ,and also to analyse the relationship between endogenous and bacterial β-G in the formation of the stone. Methods The gallbladder bile was collected from 28 patients who underwent cholecystectomy in the department of general surgery in the First Affiliated Hospital of AnHui Medical University. Based on the diagnosis, they were divided into three groups: gallbladder polypus (PO), chronic cholecystitis with calculus (CI) and acute cholecystitis with calculus (AI) .F
ive were male and six were female, the age range was 30~71 years in PO. Five were male and six were female ,the age range was 28~ 72 years in CI. Three were male and three were female, the age range was 32~68 years in CI. All the patients with hepatitis or malignant tomour were excluded. The pathological finding showed that lymphocytes and monocytes were obviously seen in CI,but neutrophils in AI. Bile were collected for the detection of TB, UCB and the activities of β-G.The gallstone were collected for the distinction of stone-types.The bottom of gallbladder were sampled for HE dye to determine pathological changes of gallbladder. UCB/TB and the total activities of β-G were Calculated for analysis. Age,sex and liver-kidney function were collected for analysis with data above. Resu I ts TB,UCB/TB and the activities of endogenous ,bacterial and total β-G were significantly different in three groups (P<0.05) , UCB was not different. UCB/TB,the
activities of endogenous , bacterial and total -G in the bile from CI was obviously higher than that in PO (P<0.05 ) , but the level of total bilirubin was reverse in the two groups (P<0.05) ,UCB was not different, and so did the changes in AI and CI except that UCB/TB was not different in two groups. The levels of endogenous ,bacterial and total β-G were positively correlated respectively (P<0.05) .All gallstone is cholesterol stone. ALT were positively correlated to the activities of endogenous , bacterial and total β-G in the bile. Conclusion There are pathological elevation of β-G in gallbladder bile with calculus (P<0.05) , and the levels of endogenous β-G were positive related to the levels of bacterial β-G because of infection (P<0.05) .TB was decreased by the hydrolyzation of β-G (P<0.05 ) ,and UCB has a descending tendency. ALT were positively correlated with the activities of -G,which is the physiological reaction of liver to infection.In conclusion, -Glucuronidase could accelerate the hydrolyzatio
n of BDG and the formation of UCB,the latter could promote the formation of gallstone by certain unknown mechanism. β-G may be responsible for the formation of gallbladder cholesterol stone.
安徽医科大学硕士学位论文
卜G的活性均显著增高(尸<0.05)，并且，内、外源性p一G活性与p一G活性总值
三者之间均呈正相关(P<0.05);本研究组所有的结石均为胆固醇结石;患者的
谷丙转氨酶(ALT)与p一G活性总值以及内、外源性p一G活性均呈正相关。结论
在胆囊结石患者的胆汁中，存在病理性增多的p一G。因为存在感染，内、外源性
p一G的活性均显著升高并呈正相关;p一G的水解作用致使胆汁中TB浓度下降;
UCB浓度也呈下降趋势。患者的ALT与p一G活性相关，此结果是肝脏在炎症情
况下的生理性反应。总之，p一G加速BDG水解并生成大量UCB，后者通过相关
机制促进胆石形成。p一G对于胆囊胆固醇结石的形成有重要意义。
Objective: There are endogenous and bacterial β-Glucuronidases(β-G) in the bile. β-G can split the conjugated bilirubin into glucuronic acid and unconjugated bilirubin (UCB) in bile, the latter is one of the major components of gallstone. β-G play an important role in the formation of pigment stone in bile duct. The authors investigated the difference of total bilirubin content (TB) , UCB content and the activities of β-G in the gallbladder bile of three groups to confirm whether β-G is responsible for the formation of gallbladder cholesterol stone ,and also to analyse the relationship between endogenous and bacterial β-G in the formation of the stone. Methods The gallbladder bile was collected from 28 patients who underwent cholecystectomy in the department of general surgery in the First Affiliated Hospital of AnHui Medical University. Based on the diagnosis, they were divided into three groups: gallbladder polypus (PO), chronic cholecystitis with calculus (CI) and acute cholecystitis with calculus (AI) .F
ive were male and six were female, the age range was 30~71 years in PO. Five were male and six were female ,the age range was 28~ 72 years in CI. Three were male and three were female, the age range was 32~68 years in CI. All the patients with hepatitis or malignant tomour were excluded. The pathological finding showed that lymphocytes and monocytes were obviously seen in CI,but neutrophils in AI. Bile were collected for the detection of TB, UCB and the activities of β-G.The gallstone were collected for the distinction of stone-types.The bottom of gallbladder were sampled for HE dye to determine pathological changes of gallbladder. UCB/TB and the total activities of β-G were Calculated for analysis. Age,sex and liver-kidney function were collected for analysis with data above. Resu I ts TB,UCB/TB and the activities of endogenous ,bacterial and total β-G were significantly different in three groups (P<0.05) , UCB was not different. UCB/TB,the
activities of endogenous , bacterial and total -G in the bile from CI was obviously higher than that in PO (P<0.05 ) , but the level of total bilirubin was reverse in the two groups (P<0.05) ,UCB was not different, and so did the changes in AI and CI except that UCB/TB was not different in two groups. The levels of endogenous ,bacterial and total β-G were positively correlated respectively (P<0.05) .All gallstone is cholesterol stone. ALT were positively correlated to the activities of endogenous , bacterial and total β-G in the bile. Conclusion There are pathological elevation of β-G in gallbladder bile with calculus (P<0.05) , and the levels of endogenous β-G were positive related to the levels of bacterial β-G because of infection (P<0.05) .TB was decreased by the hydrolyzation of β-G (P<0.05 ) ,and UCB has a descending tendency. ALT were positively correlated with the activities of -G,which is the physiological reaction of liver to infection.In conclusion, -Glucuronidase could accelerate the hydrolyzatio
n of BDG and the formation of UCB,the latter could promote the formation of gallstone by certain unknown mechanism. β-G may be responsible for the formation of gallbladder cholesterol stone.
引文
1 Ho YC, Ho LH, Ho KJ. Human hepatic beta-glucuronidase: an enzyme kinetic study. Enzyme, 1985,33(1):9-17.
2 Suzuki N, Takahashi W, Sato T. Types and chemical composition of intrahepatic stones. Prog Clin Biol Res, 1984,152: 71-80.
3 杨波，张弘，朱善德，等．人肝细胞内源性β-葡萄糖醛酸酶与胆红素结石关的免疫电镜观察．中华医学杂志，1999，79(7)：513-514．
4 Leung JW, Liu YL, Lau GC, et al. Bacteriologic analyses of bile and brown pigment stones in patients with acute cholangitis. Gastrointest Endosc, 2001,54(3).340-345.
5 刘鹏飞，肖路加．不同β-葡萄糖醛酸酶活性大肠杆菌对胆红素结石形成的影响．华西医学，1994，9(4)：480-481．
6 Chen CY, Shiesh SC, Tsao HC, et al. Human biliary beta-glucuronidase activity before and after relief of bile duct obstruction: is it the major role in the formation of pigment gallstones? J Gastroenterol Hepatol, 2000,15(9):1071-1075.
7 Ho KJ, Hsu SC, Chen JS, et al. Human biliary beta-glucuronidase: correlation of its activity with deconjugation of bilirubin in the bile. Eur J Clin Invest, 1986, 16(5):361-7
8 Kaufman HS, Magnuson TH, Pitt HA, et al. The distribution of calcium salt precipitates in the core, periphery and shell of cholesterol, black pigment and brown pigment gallstones. Hepatology, 1994,19(5):1124-1132.
9 Ostrow JD. Unconjugated bilirubin and cholesterol gallstone formation. Hepatology, 1990,12(3 Pt 2) :219S-224S.
10 Nakai K, Tazuma S, Ochi H, et al. Does bilirubin play a role in the pathogenesis of both cholesterol and pigment gallstone formation? Direct and indirect influences of bilirubin on bile lithogenicity. Biochim Biophys Acta, 2001, 1534(2-3):78-84.
11 Oyabu H, Tabata M, Nakayama F. Nonbacterial transformation of bilirubin in bile. Dig Dis Sci, 1987, 32(8): 809-816.
12 Kalra J, Lautner D, Massey KL, et al. Oxygen free radicals induced release of lysosomal enzymes in vitro. Mol Cell Biochem, 1988,84(2):233-8.
13 Osnes T, Sandstad O, Skar V, et al. Lipopolysaccharides and beta-glucuronidase activity in choledochal bile in relation to choledocholithiasis. Digestion, 1997, 58(5):437-43.
14 谢群，姚敏，王萍。细菌L型与胆囊炎关系的探讨．临床肝胆病杂志，1992，8(4)：203-204．
15 Leung JW, Liu YL, Leung PS, et al. Expression of bacterial beta-glucuronidase in human bile: an in vitro study. Gastrointest Endosc, 2001, 54(3):346-50.
16 Finch PJ, Ryan FP, Rogers K, et al. Gastric enzymes as a screening test for gastric cancer. Gut, 1987, 28(3):319-22.
17 Maki T. Pathogenesis of calcium bilirubinate gallstone: role of E. coli, beta-glucuronidase and coagulation by inorganic ions, polyelectrolytes and agitation. Ann Surg, 1966, 164(1):90-100.
18 Vitetta L, Sali A. Primary bile duct stones and bacterial activity. HPB Surg, 1992, 6(1):23-32.
19 Ho KJ. Human beta-glucuronidase. Studies on the effects of pH and bile acids in regard to its role in the pathogenesis of cholelithiasis. Biochim Biophys Acta, 1985, 827(3):197-206.
20 黄志强，杨可桢，孟宪钧，等．胆汁β-葡萄糖醛酸酶活性的意义．中华外科杂志，1982，20(1)：49-52．
21 Ho KJ. Biliary electrolytes and enzymes in patients with and without gallstones.
Dig Dis Sci. 1996, 41(12):2409-16.
22 李宁，肖路加，陈世纬，等．胆红素钙结石兔模型β-葡萄糖醛酸酶的变化．华西医科大学学报。1990，21(2)：185-187
23 Rege RV, Webster CC, Ostrow JD. Interactions of unconjugated bilirubin with bile salts. J Lipid Res, 1988, 29(10):1289-1296.
24 Ostrow JD. The etiology of pigment gallstones. Hepatology, 1984, 4(5 Suppl): 215S-222S.
25 Higashijima H, Ichimiya H. Deconjugation of bilirubin accelerates coprecipitation of cholesterol, fatty acids, and mucin in human bile--in vitro study. J Gastroenterol, 1996, 31(6):828-835.
26 Swidsinski A, Khilkin M, Pahlig H, et al. Time dependent changes in the concentration and type of bacterial sequences found in cholesterol gallstones. Hepatology, 1998, 27(3):662-5.
27 王俊田，陈允清，牛树凯．氧自由基对豚鼠胆色素结石形成的影响．山西医药杂志．1999，28(2)：116-118．
28 Moore JR, Turner BS, LaMont JT. Hydrocortisone inhibits mucin secretion from guinea pig gallbladder. Am J Physiol, 1984 247(4 Pt 1):G427-31.
29 邹一平，萧荫祺，李树林．结石性胆囊上皮细胞超微结构和胆汁糖蛋白定量分析．解放军医学杂志，1996，21(1)：33-34．
30 王学军，李玉民，李世雄．细菌代谢产物在胆囊结石成石中的作用．中国普通外科杂志，2002，11(2)：76-79．
31 Dutt MK, Murphy GM, Thompson RP. Unconjugated bilirubin in human bile: the nucleating factor in cholesterol cholelithiasis?. J Clin Pathol, 2003, 56(8):596-608.
32 Lesma A, Monti D, Mezzabotta M, Cristaldi M, et al. Monoconjugated bilirubin as a possible factor in cholesterol gallstone pathogenesis. Minerva Chir, 1997, 52(6): 771-775.
33 Zhu X, Tu X, Wang R, et al. Observation on solubility and pro-nucleative activity of monoconjugated bilirubin. Chin Med J, 1996, 109(7):542-546.
2 Suzuki N, Takahashi W, Sato T. Types and chemical composition of intrahepatic stones. Prog Clin Biol Res, 1984,152: 71-80.
3 杨波，张弘，朱善德，等．人肝细胞内源性β-葡萄糖醛酸酶与胆红素结石关的免疫电镜观察．中华医学杂志，1999，79(7)：513-514．
4 Leung JW, Liu YL, Lau GC, et al. Bacteriologic analyses of bile and brown pigment stones in patients with acute cholangitis. Gastrointest Endosc, 2001,54(3).340-345.
5 刘鹏飞，肖路加．不同β-葡萄糖醛酸酶活性大肠杆菌对胆红素结石形成的影响．华西医学，1994，9(4)：480-481．
6 Chen CY, Shiesh SC, Tsao HC, et al. Human biliary beta-glucuronidase activity before and after relief of bile duct obstruction: is it the major role in the formation of pigment gallstones? J Gastroenterol Hepatol, 2000,15(9):1071-1075.
7 Ho KJ, Hsu SC, Chen JS, et al. Human biliary beta-glucuronidase: correlation of its activity with deconjugation of bilirubin in the bile. Eur J Clin Invest, 1986, 16(5):361-7
8 Kaufman HS, Magnuson TH, Pitt HA, et al. The distribution of calcium salt precipitates in the core, periphery and shell of cholesterol, black pigment and brown pigment gallstones. Hepatology, 1994,19(5):1124-1132.
9 Ostrow JD. Unconjugated bilirubin and cholesterol gallstone formation. Hepatology, 1990,12(3 Pt 2) :219S-224S.
10 Nakai K, Tazuma S, Ochi H, et al. Does bilirubin play a role in the pathogenesis of both cholesterol and pigment gallstone formation? Direct and indirect influences of bilirubin on bile lithogenicity. Biochim Biophys Acta, 2001, 1534(2-3):78-84.
11 Oyabu H, Tabata M, Nakayama F. Nonbacterial transformation of bilirubin in bile. Dig Dis Sci, 1987, 32(8): 809-816.
12 Kalra J, Lautner D, Massey KL, et al. Oxygen free radicals induced release of lysosomal enzymes in vitro. Mol Cell Biochem, 1988,84(2):233-8.
13 Osnes T, Sandstad O, Skar V, et al. Lipopolysaccharides and beta-glucuronidase activity in choledochal bile in relation to choledocholithiasis. Digestion, 1997, 58(5):437-43.
14 谢群，姚敏，王萍。细菌L型与胆囊炎关系的探讨．临床肝胆病杂志，1992，8(4)：203-204．
15 Leung JW, Liu YL, Leung PS, et al. Expression of bacterial beta-glucuronidase in human bile: an in vitro study. Gastrointest Endosc, 2001, 54(3):346-50.
16 Finch PJ, Ryan FP, Rogers K, et al. Gastric enzymes as a screening test for gastric cancer. Gut, 1987, 28(3):319-22.
17 Maki T. Pathogenesis of calcium bilirubinate gallstone: role of E. coli, beta-glucuronidase and coagulation by inorganic ions, polyelectrolytes and agitation. Ann Surg, 1966, 164(1):90-100.
18 Vitetta L, Sali A. Primary bile duct stones and bacterial activity. HPB Surg, 1992, 6(1):23-32.
19 Ho KJ. Human beta-glucuronidase. Studies on the effects of pH and bile acids in regard to its role in the pathogenesis of cholelithiasis. Biochim Biophys Acta, 1985, 827(3):197-206.
20 黄志强，杨可桢，孟宪钧，等．胆汁β-葡萄糖醛酸酶活性的意义．中华外科杂志，1982，20(1)：49-52．
21 Ho KJ. Biliary electrolytes and enzymes in patients with and without gallstones.
Dig Dis Sci. 1996, 41(12):2409-16.
22 李宁，肖路加，陈世纬，等．胆红素钙结石兔模型β-葡萄糖醛酸酶的变化．华西医科大学学报。1990，21(2)：185-187
23 Rege RV, Webster CC, Ostrow JD. Interactions of unconjugated bilirubin with bile salts. J Lipid Res, 1988, 29(10):1289-1296.
24 Ostrow JD. The etiology of pigment gallstones. Hepatology, 1984, 4(5 Suppl): 215S-222S.
25 Higashijima H, Ichimiya H. Deconjugation of bilirubin accelerates coprecipitation of cholesterol, fatty acids, and mucin in human bile--in vitro study. J Gastroenterol, 1996, 31(6):828-835.
26 Swidsinski A, Khilkin M, Pahlig H, et al. Time dependent changes in the concentration and type of bacterial sequences found in cholesterol gallstones. Hepatology, 1998, 27(3):662-5.
27 王俊田，陈允清，牛树凯．氧自由基对豚鼠胆色素结石形成的影响．山西医药杂志．1999，28(2)：116-118．
28 Moore JR, Turner BS, LaMont JT. Hydrocortisone inhibits mucin secretion from guinea pig gallbladder. Am J Physiol, 1984 247(4 Pt 1):G427-31.
29 邹一平，萧荫祺，李树林．结石性胆囊上皮细胞超微结构和胆汁糖蛋白定量分析．解放军医学杂志，1996，21(1)：33-34．
30 王学军，李玉民，李世雄．细菌代谢产物在胆囊结石成石中的作用．中国普通外科杂志，2002，11(2)：76-79．
31 Dutt MK, Murphy GM, Thompson RP. Unconjugated bilirubin in human bile: the nucleating factor in cholesterol cholelithiasis?. J Clin Pathol, 2003, 56(8):596-608.
32 Lesma A, Monti D, Mezzabotta M, Cristaldi M, et al. Monoconjugated bilirubin as a possible factor in cholesterol gallstone pathogenesis. Minerva Chir, 1997, 52(6): 771-775.
33 Zhu X, Tu X, Wang R, et al. Observation on solubility and pro-nucleative activity of monoconjugated bilirubin. Chin Med J, 1996, 109(7):542-546.