Pregnane receptor gene polymorphisms, pathogenic bacteria distribution and drug sensitivity, and TCM syndrome differentiation in patients with cholelithiasis.

OBJECTIVE: To investigate the distribution of pathogens and drug resistance in bile and the association between the pregnane X receptor (PXR) gene polymorphisms, traditional Chinese medicine (TCM) syndromes and the risk of cholesterol gallstone disease (CGD). METHODS: A total of 392 samples were enrolled in this study from January 2014 to February 2015, among which 192 patients were with CGD, and 200 samples were healthy. Strains were isolated and susceptibility testing was the disk diffusion method susceptibility testing. The patients were divided into hepatochlic hygropyrexia, stagnation of liver-qi, and the accumulation of damp. The PXR gene polymorphisms were analyzed by polymerase chain reaction-restriction fragment length polymorphism. The association between the PXR gene polymorphisms and the risk of CGD was examined by logistic regression analysis. RESULTS: A total of 192 cases were detected in 230 of bile culture pathogens, including Gram-negative bacteria 133 (57.83%), Gram-positive bacteria 76 (33.04%), and fungi 21 (9.13%). The top five pathogens were Escherichia coli, Klebsiella pneumoniae, Enterococcus faecalis, Candida albicans, and Enterococcus feces, of which 110 cases was of single infection, 48 cases of mixed infection of two strains, eight cases of mixed infection of three bacteria. Among 59 Escherichia coli, the yield extended-spectrum beta-lactamases had 40 (67.80%). The hepatochlic hygropyrexia was the most TCM syndrome, followed by stagnation of liver-qi, and the accumulation of damp was least. Different pathogens and the rs6785049 genotypes distributed differently in cholelithiasis patients with different TCM syndromes (P < 0.05). In hepatochlic hygropyrexia patients the Gram-negative bacteria was most. There was significant differences between CGD group and control group in rs6785049 (P < 0.001). Comparison with wild-type portable GG, GA genotype increased the risk of the occurrence of gallstones (OR = 0.40, 95%CI: 0.16-0.79); likewise, carrying the GA+AA genotype also increased the risk (OR = 0.38, 95%CI: 0.19-0.81). There was no significant differences in rs2276707, rs3814055 site polymorphic loci alleles in CGD group and control group. CONCLUSIONS: In the treatment of cholelithiasis, bile samples should be collected for bacterial culture and sensitivity test, and drugs should be strictly chosen based on the results. The rs6785049 polymorphisms in PXR gene may increase the risk of gallstones ontogeny, and gallstones can be early detected and prevented by detecting genotypes. rs6785049 polymorphisms in PXR gene may has relationship with TCM syndromes.

[Shengqing Capsule down-regulates estrogen and progesterone receptors in epithelial tissue of gallbladder in guinea pigs with gallstone].

OBJECTIVE: To study the role of estrogen receptor (ER) and progesterone receptor (PR) in the formation of cholesterol calculus and investigate the effects of Shengqing Capsule (SQC), a Chinese patent herbal medicine with the function of soothing liver and draining gallbladder, on ER and PR expressions. METHODS: A total of 80 female guinea pigs were divided into normal control group, untreated group, ursodeoxycholic acid group (UDCA group) and SQC group. The cholesterol gallstone was induced by feeding the guinea pigs with high-fat lithogenic diet. SQC and UDCA were separately administered to the guinea pigs in the SQC group and UDCA group. After 7-week administration, all the animals were sacrificed to calculate the incidence of calculus formation and detect the expressions the ER and PR in the epithelial tissue of gallbladder by immunohistochemical method. RESULTS: Gallstone was cholesterol calculus detected by infrared spectrum. The incidence of calculus formation in the SQC group (27.78%) was significantly lower than that in the untreated group (81.25%) (X(2)=9.721 5, P=0.001 8). On the basis of Reiner standard, the expression distribution of ER and PR increased gradually from the normal control group through the SQC group and UDCA group to the untreated group. Except for the former two groups and the latter two groups, the differences between the other groups and UDCA group were statistically significant (P<0.05). Besides, the differences of positive expression rates between groups were statistically significant (P<0.05). CONCLUSION: Increased expressions of ER and PR are closely related to the formation of cholesterol stone. And Shengqing Capsule can down-regulate the expressions of ER and PR.

[Effects of Shengqing Capsule on biochemical parameters in mice with cholesterol gallstone].

OBJECTIVE: To observe the effects of Shengqing Capsule (SQC), a compound of traditional Chinese herbal medicine, on biochemical parameters in C57BL/6J mice with cholesterol gallstone. METHODS: Thirty-eight C57BL/6J mice were randomly divided into normal control group (n=10), untreated group (n=15) and SQC group (n=13). Cholesterol gallstone was induced in mice of the latter two groups by feeding high cholesterol diet. Mice in the SQC group were intragastricly administered SQC 0.22 g/(kg.d). After 8-week treatment, animals were sacrificed and sampled to calculate the incidences of stone formation. The contents of serum cholesterols and Ca(2+), and the malonaldehyde (MDA) content and superoxide dismutase (SOD) activity in liver tissues were detected. RESULTS: The incidences of stone formation were 73.33% in untreated group, 0% in normal control group, and 23.08% in the SQC group. And the INCIDENCE in untreated group was significantly higher than those in the normal control group and the SQC group (P<0.01). Contents of serum total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) in the SQC group were lower than that in the untreated group (P<0.01), and high-density lipoprotein cholesterol (HDL-C) in the SQC group was higher than that in the untreated group (P<0.01). SQC could increase the SOD activity and decrease the MDA content in liver tissues, showing significant differences as compared with those in the untreated group (P<0.05). CONCLUSION: SQC can decrease the incidence of stone formation and improve the biochemical parameters, which may be one of the mechanisms in the treatment and prevention of cholesterol gallstone disease.

[Effects of different Chinese herbal medicines on biochemical parameters in guinea-pig with pigment gallstones].

OBJECTIVE: To observe the effects of Qingdan Capsule (QDC) and Yanggan Lidan Granule (YGLDG), two kinds of compound traditional Chinese herbal medicines, on biochemical parameters in guinea-pigs with pigment gallstones. METHODS: An animal model of pigment gallstones was established in male guinea-pigs by hypodermic injection of lincomycin. The guinea-pigs were randomly divided into blank control group, untreated group, QDC group and YGLDG group. There were 8 guinea-pigs in each group. After ten-day treatment, animals were sacrificed and sampled to calculate the rate of stone formation, total bilirubin (TB), unconjugated bilirubin (UCB) and Ca2+ density in bile of the four groups. RESULTS: In comparison with the untreated group, the rates of stone formation in the QDC and YGLDG groups were significantly decreased (P<0.01). TBIL, UCB and Ca2+ content of bile in both QDC and YGLD groups was also significantly decreased (P<0.05, P<0.01). CONCLUSION: QDC and YGLD have good effects on biochemical changes of animal model of pigment gallstone in reversing the lithogenesity of bile by reducing the content of TB, UCB and Ca2+, hence resulting in clinical treatment and prevention of pigment gallstone disease.

[Clinical early intervention of Tongxia Huayu Decoction on pancreatic microcirculatory disturbance in severe acute pancreatitis].

OBJECTIVE: To study the mechanisms of Tongxia Huayu Decoction (a Chinese herbal decoction for purgation and removing blood stasis) in prognostic improvement for severe acute pancreatitis by early intervention on pancreatic microcirculatory disturbance. METHODS: Fifty-three patients with severe acute pancreatitis were divided randomly into treatment group (n=28) and control group (n=25). Tongxia Huayu Decoction was given to the patients in treatment group in addition to the normal treatment in control group for one week. The clinical symptoms and signs, hemodiastase, urinary amylase, C-reactive protein (CRP) and endothelin (ET) of the patients in the two groups before and after treatment were observed and detected. RESULTS: The total response rate of the treatment group was 98.4%, while that of the control group was 80%, with significant difference between them (P<0.05). There was no significant difference of the contents of hemodiastase, urinary amylase, CRP and ET between the two groups before treatment, while they were significantly decreased after treatment (P<0.01) with more obvious change in treatment group (P<0.01). CONCLUSION: Tongxia Huayu Decoction brings satisfied therapeutic effect on severe acute pancreatitis. The mechanisms may associate with its reducing function on ET releasing so as to inhibit the pancreatic microcirculatory disturbance and acinar cell injury induced by ET.

[Impact of Jinhong Tablet on thymus of rats with cholangitis].

OBJECTIVE: To study the changes of the center of cellular immune thymus and the interference of Jinhong Tablet, a compound traditional Chinese herbal medicine, during the course of cholangitis. METHODS: Twenty-four male Wistar rats were randomly divided into three groups. Ligation group, simple ligation of bile duct without producing infection and fed with normal saline; untreated group, reproduce acute cholangitis and fed with normal saline; Jinhong Tablet-treated group, reproduce cholangitis and treated with Jinhong Tablet. Five days after modeling, the rats were sacrificed and the thymus were sampled to observe the morphological changes with an electronic microscope, count the index of thymus and apoptosis and assay the expressions of Bcl-2 gene coding protein and Fas gene coding protein. RESULTS: The frequency of apoptosis of thymus in the untreated group was higher than that in the Jinhong Tablet-treated group and the ligation group, and the frequency of apoptosis of thymus in the Jinhong Tablet-treated group was higher than that in the ligation group. Compared with the Jinhong Tablet-treated group and the ligation group, the weight and index of thymus were lower and the index of apoptosis of thymus was higher significantly in the untreated group (P<0.01). The expression of Bcl-2 gene coding protein in the ligation group and the Jinhong Tablet-treated group were much higher than that in the untreated group (P<0.01), but there were no significant differences of the expression of Fas gene coding protein among the three groups. CONCLUSION: There exists a promotion of non-physiological apoptosis of cellular immune center such as decrease of the weight and the index of thymus and increase of the index of apoptosis of thymus during acute cholangitis, which may be inhibited by up-regulation of Bcl-2 gene coding protein with antipyretic and purgative herbs Jinhong Tablet.

[Effect of traditional Chinese herbs for nourishing the liver on intracellular free calcium level in gallbladder cells of guinea pigs with gallstones].

OBJECTIVE: To observe the changes of intracellular free calcium level ([Ca(2+)]i) in gallbladder cells of guinea pigs with gallstones so as to study the mechanisms of gallstone formation and the prevention and treatment function of traditional Chinese herbs for nourishing the liver. METHODS: Eighty guinea pigs were randomly divided into four groups, which were normal control group, untreated group, nourishing-liver Chinese drug (NLCD) group and ursodeoxycholic acid (UDCA) group, with 20 guinea pigs in each group. Gallstones were induced in the guinea pigs of the latter 3 groups by the feed of diet inducing cholelithiasis with high cholesterol, while the corresponding medicines were used in NLCD group and UDCA group for prevention and treatment for 7 weeks. Then the state of the guinea pigs, the formation of gallstones, and the changes of [Ca(2+)]i in gallbladder cells were observed. RESULTS: The [Ca(2+)]i in gallbladder cells of guinea pigs in the untreated group was decreased significantly. NLCD improved the behavioral signs of the guinea pigs, significantly decreased the formative rate of gallstones and increased the [Ca(2+)]i in gallbladder cells. CONCLUSIONS: The [Ca(2+)]i in gallbladder cells is the important factor for contractile function of gallbladder and the information of gallstones. Traditional Chinese herbs for nourishing the liver may significantly increase the [Ca(2+)]i in gallbladder cells to facilitate contraction of the smooth muscle cells of gallbladder and relieve the cholestatis. It may be one of the mechanisms of traditional Chinese herbs for nourishing the liver in preventing and treating cholelithiasis.

[Effects of Shengqing Capsules on expression of nuclear factor-kappaB protein in liver cells of guinea pigs with pigment gallstone].

OBJECTIVE: To study the mechanisms of Shengqing Capsules, a compound Chinese herbal medicine for dispersing stagnated liver qi and promoting bile flow, in prevention and treatment of pigment gallstone. METHODS: Liver cells from guinea pigs with pigment gallstone were primarily cultured in vitro. The serums containing different concentrations of Shengqing Capsules were prepared by serum pharmacological method. The primary cultured hepatocytes with excessive expression of nuclear factor-kappaB (NF-kappaB) were stimulated with lipopolysaccharides (LPS) and the effects of the drug-containing serums on the NF-kappaB expression were detected by Western-blotting. RESULTS: Shengqing Capsules down-regulated the NF-kappaB expression increased by LPS stimulation. The effect is dose-dependent. CONCLUSION: Shengqing Capsules can down-regulate the NF-kappaB expression increased by LPS stimulation. It may be one of the important mechanisms of this Chinese herbal medicine in prevention and treatment for pigment gallstone formation and the inflammatory reaction in biliary tract.

[Inhibiting effects of recipe for dispersing stagnated liver qi to promote bile flow on cholesterol gallstone formation in guinea pigs].

OBJECTIVE: To explore the mechanism of the inhibiting effects of recipe for dispersing stagnated liver qi to promote bile flow (Danshijing Tablets) on cholesterol gallstone formation and provide experimental evidence for its clinical application. METHODS: Eighty guinea pigs were randomly divided into 4 groups, which were normal control group, untreated group, ursodeoxycholic acid (UDCA)-treated group and Danshijing Tablets-treated group. The gallstones in guinea pigs were induced by high-cholesterol diet. UDCA and Danshijing Tablets were given orally to guinea pigs in the corresponding group respectively for seven weeks. Then the physical signs of the guinea pigs, the rates of gallstone formation and the histomorphological changes of the gallbladder were examined. RESULTS: The behavior of guinea pigs in the Danshijing Tablets-treated group was obviously improved and the rate of gallstone formation was significantly decreased as compared with those in both untreated and UDCA-treated groups (P<0.05). The inflammation reaction of mucous membrane in gallbladder was evidently reduced in the Danshijing Tablets-treated group and its morphological appearance turned to be approximately normal. CONCLUSION: Recipe for dispersing stagnated liver qi to promote bile flow may decrease gallstone formation. Its mechanism may be related to reducing pathologic changes in gallbladder tissues, which will reduce the damages of cholesterol to the smooth muscle in gallbladder and enhance the contractility of gallbladder.

[Effects of Shengqing Capsules on cholelithiasis-related genes in guinea pigs].

OBJECTIVE: To explore the molecular mechanisms of Shengqing Capsules in treating cholelithiasis. METHODS: Sixty female guinea pigs were randomized into 3 groups: group I (fed with normal diet), group II (fed with low-protein diet) and group III (fed with low-protein diet and Shengqing Capsules). After six-week feeding, the gallstone formation and the expressions of bilirubin UDP-glucuronosyltransferase (B-UGT) mRNA and cholesterol 7alpha-hydroxylase (CYP7A1) mRNA were observed. RESULTS: The proportions of stone-formed in groups I, II and III were 2/14, 9/12 and 4/14, respectively. There were significant differences among the three groups (P<0.05). The expressions of B-UGT and CYP7A1 mRNAs were higher in both group I and group III as compared with those in the group II (P<0.05). CONCLUSION: Shengqing Capsules can reduce the rate of stone-formation, which may be due to its interference of metabolism of bilirubin and cholesterol and up-regulation of the expressions of B-UGT and CYP7A1 mRNAs.

[Impact of antipyretic and purgative herbs on intestinal mucosal barrier and inflammatory response in treatment of acute cholangitis in rats].

OBJECTIVE: To investigate the effects of antipyretic and purgative herbs on intestinal mucosal barrier and inflammatory response in the treatment of acute cholangitis. METHODS: Sixty SD rats were randomly divided into group A (untreated group, acute cholangitis was induced, n=20), group B (treatment group, acute cholangitis was induced and treated with antipyretic and purgative herbs, n=20) and group C (sham operation group, n=20). At the third or fifth day after operation, the rats were sacrificed and sampled. The serum endotoxin, cytokines and inflammatory mediators were tested and the numbers of labeled bacteria in the liver, spleen and mesenteric lymph nodes translocated from the gut were assayed. RESULTS: As compared with group A, the serum content of endotoxin, IL-6, IL-8, TNF-alpha, CRP and NO was significantly lower and that of IL-2 was significantly higher, and the translocated numbers of labeled bacteria from gut were reduced in both group B and group C (P<0.01). CONCLUSION: Antipyretic and purgative herbs can play therapeutic roles in the treatment of acute biliary tract infections, including the protection of intestinal mucosal barrier from bacterial translocation, reduction of serum endotoxin content and regulation of inflammatory response.

[Experimental study on effects of herbs for nourishing and smoothing the liver in reversing bile lithogenicity of guinea pig].

OBJECTIVE: To further probe the mechanisms of herbs for nourishing and smoothing the liver in reversing bile lithogenicity of guinea pig. METHODS: Sixty guinea pigs were divided randomly into control group (fed with normal diet, n=20), model group (fed with lithogenic diet, n=20) and treatment group (fed with lithogenic diet plus herbal medicine, n=20). After four-week feeding, the animals were sacrificed and sampled, the rates of gallstone formation in each group were estimated, and the total bile acid (TBA), total bilirubin (TBIL), conjugated bilirubin (CB), unconjugated bilirubin (UCB), and calcium ion in the bile were determined, and the different bilirubins were analyzed by HPLC. RESULTS: (1) The rate of gallstone formation was 5% in normal group, 81.25% in model group and 31.25% in treatment group (P<0.05). (2) The bile TBIL, CB, UCB and Ca(2+) were higher and the bile TBA was lower significantly in model group than that in the other two groups (P<0.05). (3) HPLC analysis revealed that MCB was higher and DCB was lower significantly in model group (P<0.01), and there were no significant differences of UCB and IPA among the three groups. (4) The percentages of MCB and UCB were much higher and the percentage of DCB was remarkably lower in model group (P<0.05). CONCLUSION: Herbs for nourishing and smoothing the liver can significantly reduce the rate of gallstone formation and has effect of reversing lithogenicity of bile in guinea pigs fed with lithogenic diet.