Key symptoms selection for two major syndromes diagnosis of Chinese medicine in chronic hepatitis B / 中国结合医学杂志

<p><b>OBJECTIVE</b>To identify key symptoms of two major syndromes in chronic hepatitis B (CHB), which can be the clinical evidence for Chinese medicine (CM) doctors to make decisions.</p><p><b>METHODS</b>Standardization scales on diagnosis for CHB in CM were designed including physical symptoms, tongue and pulse appearance. The total of 695 CHB cases with dampness-heat (DH) syndrome or Pi (Spleen) deficiency (SD) syndrome were collected for feature selection and modeling, another 275 CHB patients were collected in different locations for validation. Key symptoms were selected based on modified information gain (IG), and 5 classifiers were applied to assist with models training and validation. Classification accuracy and area under receiver operating characteristic curves (AUC) were evaluated.</p><p><b>RESULTS</b>(1) Thirteen DH syndrome key symptoms and 13 SD syndrome key symptoms were selected from original 125 symptoms; (2) The key symptoms could achieve similar or better diagnostic accuracy than the original total symptoms; (3) In the validation phase, the key symptoms could identify syndromes effectively, especially in DH syndrome, which average prediction accuracy on 5 classifiers could achieve 0.864 with the average AUC 0.772.</p><p><b>CONCLUSION</b>The selected key symptoms could be simple DH and SD syndromes diagnostic elements applied in clinical directly. (Registration N0.: ChiCTR-DCC-10000759).</p>

Regulatory Effect of Qushi Huayu Recipe on Gene Expression Profiles of Fatty Liver Rats / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To observe the intervention and mechanism of Qushi Huayu Recipe (QHR) on gene expression profiles in high lipid diet induced fatty liver rats.</p><p><b>METHODS</b>Fatty liver model was prepared in 20 male SD rats using single high fat diet (88% common forage +2% cholesterol +10% lard). Four weeks after modeling they were divided into the model group and the QHR group according to random digit table, 10 in each group. QHR (at 0. 93 g crude drug/100 g body weight) and distilled water was respectively to rats in the QHR group and the model group by gastrogavage while modeling, once per day. Meanwhile, 10 SD male rats were recruited in a normal group, administered with equal volume of distilled water by gastrogavage. At the end of week 8 all rats were sacrificed, and blood and livers were collected for subsequent analysis. Contents of liver triglyceride (TG) and free fatty acid (FFA) , activities of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were detected using biochemical assay. Pathological changes of liver tissue were observed using H&E and oil red O stain. Liver gene expressions were detected by Affymetrix gene expression profiles. Differentially expressed genes were compared between the QHR group and the model group, functions of differentially expressed genes and signal pathways involved analyzed. Ten differentially expressed genes involved in glycolipid metabolism with fold change more than 2 were selected for verification by real-time PCR.</p><p><b>RESULTS</b>(1) Compared with the normal group, contents of liver TG and FFA, and serum activities of ALT and AST obviously increased in the model group (P <0. 01). Compared with the model group, contents of liver TG and FFA, and activities of ALT and AST obviously decreased in the QHR group (P <0. 05, P <0. 01). QHR could reduce high fat induced fatty degeneration of liver cells , alleviate inflammation, and improve pathological changes of liver tissue. (2) Compared with the model group, there were 80 differentially expressed genes (with fold change > 2, P < 0.05) with clear functions and appointed gene names, including 44 up-regulated and 36 down-regulated genes. Eighty genes were involved in 27 signal pathways with statistical difference, including glycerolipid metabolism, adipocytokine signaling pathway, insulin signal pathway, drug metabolism signal pathway, etc (P < 0.05). (3) RT-PCR results of 10 glycolipids metabolism regulating genes such as Gk, Scd1, Gpat2, G6pc, Irs1, and so on showed that all RT-PCR genes were completely coincide with up-regulated or down-regulated tendency in results of gene chips. 80% genes had approximate fold change.</p><p><b>CONCLUSION</b>QHR could regulate gene expressions related to fat metabolism, carbohydrate metabolism, anti-lipid peroxidation, and drug metabolism in high fat diet induced fatty liver rats, and its comprehensive pharmacological actions could be manifested.</p>

Effect of CKJ recipe containing serum on activation of rat primary hepatic stellate cells, TGF-beta1 and its receptors / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To observe the effect of CKJ Recipe (consisting of Cordyceps sinensis polysaccharide, amygdaloside, and gypenosides) containing serum on the activation of rat primary hepatic stellate cells (rHSCs) and to explore its pharmacological mechanism.</p><p><b>METHODS</b>rHSCs were isolated form liver and cultured for four days. Then they were divided into the normal control group, the model group, and the CKJ group. rHSCs in the model group and the CKJ group were treated with 2.5 ng/mL transforming growth factor beta1 (TGF-beta1) in serum-free DMEM for 24 h. Serum free DMEM (containing no TGF-beta1) was taken as the control for the normal control group. rHSCs in the CKJ group were treated with 5% CKJ-containing serum for 24 h. rHSCs in the other two groups were treated with 5% blank serum for 24 h.The protein expression level of a smooth muscle actin (alpha-SMA) was determined using high throughput screening (HCS) and Western blot. mRNA expression levels of alpha-SMA, collagen I (Col-I), platelet-derived growth factor receptor beta (PDGF-betaR), TGF-beta1, transforming growth factor beta receptor 1 (TGF-betaR1), and transforming growth factor beta receptor 2 (TGF-beta R2) were detected using quantitative RT-PCR.</p><p><b>RESULTS</b>Compared with the normal control group, the protein expression level of alpha-SMA, mRNA expression levels of alpha-SMA, Col-I, PDGF-betaR, TGF-beta1, TGF-betaR1, and TGF-betaR2 significantly increased in the model group (P<0.05, P<0.01). Compared with the model group, the protein expression level of alpha-SMA, mRNA expression levels of alpha-SMA, Col-I, PDGF-betaR, TGF-beta1, TGF-beta1, and TGF-beta R2 significantly decreased in the CKJ group (P<0.05, P<0.01).</p><p><b>CONCLUSION</b>CKJ containing serum could inhibit the protein expression level of o-SMA, which was probably related with inhibiting TGF-beta1 and its related receptors.</p>

Effect of cordyceps polysaccharide on lipid peroxidation of rats with dimethylnitrosamine-induced liver fibrosis / 中国中药杂志

<p><b>OBJECTIVE</b>To observe the pharmacological effect of Cordyceps polysaccharide on dimethylnitrosamine (DMN)-induced liver fibrosis in rats.</p><p><b>METHOD</b>DMN rat liver fibrosis model was established and divided into the normal group (N, n = 6), the model group (M, n = 11), the Cordyceps polysaccharide group (C, n = 8) and the colchicine group (Q, n = 9). During the modeling for four weeks, Cordyceps polysaccharide (60 mg x kg(-1)) and colchicine (0.1 mg x kg(-1)) were orally administered for three weeks, while the model and normal groups were given disinfected water of the same amount.</p><p><b>OBSERVATION</b>serum ALT, AST, GGT and Alb, TBil content; content of hydroxyproline (Hyp) in liver tissues; liver pathology and collagen staining; SOD activity and MDA, GSH, GSH-Px in liver tissues; protein expression of proliferating cell nuclear antigen (PCNA) in liver tissues.</p><p><b>RESULT</b>Serum ALT, AST, GGT, TBil significantly increased, and A1b decreased significantly in the model group. Hepatic Hyp significantly increased in the model group, whereas the index remarkably decreased in the Cordyceps polysaccharide group and the colchicine group. HE staining: the structure of normal hepatic lobules was damaged, with hepatocytes tumefaction and proliferation of connective tissues in portal tracts in the model group, while the Cordyceps polysaccharide group and the colchicine group recorded notable reduction in above pathological changes. Collagen staining: the model group showed hepatic lobule fibrous septum and many intact pseudolobules; while the Cordyceps polysaccharide group and the colchicine group witnessed decrease in collagen deposition. The model group showed significant decrease in SOD, GSH-Px and GSH and increase in MDA, whereas the Cordyceps polysaccharide group and the colchicine group recorded notable growth in GSH and GSH-Px. The model group showed significant decrease in protein expression of PCNA in liver tissues, while the Cordyceps polysaccharide group and the colchicine group showed significant reduction.</p><p><b>CONCLUSION</b>Cordyceps polysaccharide can significantly inhibit DMN-induced liver fibrosis and lipid peroxidation in rats.</p>

Dynamic change of lipid peroxidation-related protein expression and the intervention effects of Yiguanjian decoction in a rat model of CCl4-induced liver fibrosis / 中华肝脏病杂志

To investigate the dynamic change of lipid peroxidation-related protein expression and the intervention effects of Yiguanjian (YGJ) Decoction on liver fibrosis induced by CCl4 in rat. Fifty-seven male Wistar rats were randomly divided into a liver fibrosis group (n = 39) and a normal group (n = 18). The liver fibrosis was treated with peritoneal injection of 50% CCl4 for nine weeks. At the end of weeks 3 and 6 of CCl4 treatment, six rats were sacrificed to assess the status of liver fibrosis. At the end of week 7, the remaining -fibrotic rats were randomly divided into an untreated model group (M, n=15) and a YGJ-treated group (n = 12). The YGJ group was administered daily, oral YGJ Decoction for three weeks, concomitant with continued CCl4 treatment. The M group and normal group received the same treatment oral regimen and volume of distilled water. At the end of week 8, four rats in group M were sacrificed to observed the fibrosis status. At the end of week 9, the fibrotic rats were sacrificed for sampling. Liver function, histological changes, contents of hydroxyproline (Hyp) and malondialdehyde (MDA), activity of super oxidase dismutase (SOD) and L-glutathione (GSH), protein expression of heat shock protein (HSP)70, heme oxygenase (HO)-1, transferrin, peroxiredoxin (Prxd) 6 and liver fatty acid binding protein (L-FABP) were detected. Compared with normal group-, the MDA content was increased significantly in M group at week 6 (M: 4.23+/-0.45 nmol/mg vs. normal: 2.22+/-0.59 nmol/mg, F = 60.13, P less than 0.01) and week 9 (M: 6.29+/-1.23 nmol/mg vs. normal: 2.22+/-0.59 nmol/mg, F = 66.99, P less than 0.01), but the SOD activity was decreased significantly at the same time points [week 6: (M: 196.94+/-39.20 U/mg vs. normal: 264.50+/-30.44 U/mg, F = 11.12, P less than 0.01]); [week 9: (M: 152.2+/-51.65 U/mg vs. normal: 264.50+/-30.44 U/mg, F = 23.11, P less than 0.01)], as were the GSH content [week 6: (M: 48.47+/-7.27 nmol/mg vs. 60.74+/-9.04 nmol/mg, F = 6.71, P less than 0.05]]; [week 9: (M: 37.89+/-9.01 nmol/mg vs. 60.74+/-9.04 nmol/mg, F = 24.06, P less than 0.01]]. Compared with group M at week 9, the YGH-treated model group had markedly decreased MDA (YGJ: 4.25+/-0.86 nmol/mg vs. M: 6.29+/-1.23 nmol/mg, F = 19.52, P less than 0.01], but significantly increased SOD (YGJ: 198.35+/-46.48 U/mg vs. 152.21+/-51.65 U/mg, F = 4.65, P less than 0.05] and GSH (YGJ: 53.73+/-7.54 nmol/mg vs. M: 37.89+/-9.01 nmol/mg, F = 19.23, P less than 0.01). Compared to normal rats at week 9, group M had significantly higher protein levels of HSP70 (normal: 1.21+/-0.06 vs. M: 0.58+/-0.07, F = 166.87, P less than 0.01) and HO-1 (normal: 1.11+/-0.06 vs. M: 0.58+/-0.06, F = 123.96, P less than 0.01), but significantly decreased levels of Prxd6 (normal: 0.04+/-0.05 vs. M: 1.49+/-0.05, F = 1215.85, P less than 0.01), transferrin (normal: 0.67+/-0.03 vs. M: 1.67+/-0.04, F = 301.35, P less than 0.01), and L-FABP (normal: 0.24+/-0.02 vs. M: 1.44+/-0.14, F = 219.05, P less than 0.01). Compared to group M at week 9, the YGJ treatment group showed significantly reduced HSP70 (YGJ: 0.82+/-0.04 vs. M: 1.21+/-0.06, F = 92.31, P less than 0.01) and HO-1 (YGJ: 0.90+/-0.04 vs. 1.11+/-0.06, F = 26.89, P less than 0.01), but significantly increased Prxd6 (YGJ: 0.88+/-0.11 vs. 0.04+/-0.05, F = 150.17, P less than 0.01), transferrin (YGJ: 1.36+/-0.13 vs. 0.24+/-0.02, F = 237.19, P less than 0.01), and L-FABP (YGJ: 1.04+/-0.12 vs. 0.67+/-0.03, F = 27.53, P less than 0.01). YGJ treatment of fibrotic liver rats reduces lipid peroxidation damage by preventing generation of oxidizing substances.

The key target of Chinese medicine treatment on alcoholic and nonalcoholic fatty liver disease: the gut / 中国中西医结合杂志

In recent years, the pathogenesis of "gut-liver axis" in alcoholic and nonalcoholic fatty liver disease has attracted more attention in this field. In this paper, the relationship among fatty liver, gut-permeability, gut-derived endotoxin, and gut microbiota was systematically clarified. Based on the researches of treatment and prevention of fatty liver and gut injury by Chinese medicine, the gut is believed as the curative target for fatty liver disease, which not only is the modern annotation for the Chinese medicine practice, but also might possibly become an important view angle and strategy for fatty liver disease treatment.

Structural shifts of gut flora in rat acute alcoholic liver injury and jianpi huoxue decoction's effect displayed by ERIC-PCR fingerprint / 中国结合医学杂志

<p><b>OBJECTIVE</b>To study the structural shifts of gut flora in rats with acute alcoholic liver injury (AALI), and the effect of jianpi huoxue decoction (JPHXD) on the gut flora.</p><p><b>METHODS</b>Thirty-six Sprague-Dawley rats were randomly allocated to the control, AALI and JPHXD groups equally. The rats in the control group were given water and those in AALI and JPHXD groups were given ethanol by intragastric gavage for 5 days, while rats in the JPHXD group were administered JPHXD simultaneously. The blood and liver tissue were collected at the end of the experiment. The activities of serum alkaline aminotransferase (ALT), aspartate aminotransferase (AST), hepatic γ-glutamyltranspetidase (γ-GT) and hepatic triglyceride (TG) levels were determined. Plasma endotoxin level in the portal vein was measured. Pathological changes of liver tissues were determined by hematoxylin and eosin (HE) staining and oil red O staining. The total DNA of gut flora were extracted from fecal samples by Bead-beating method and determined by ERIC-PCR fingerprint method. The similarity cluster analysis and principal component analysis were performed to analyze the ERIC-PCR fingerprint respectively.</p><p><b>RESULTS</b>In the AALI group, the ratio of liver/body weight, activities of ALT, AST and hepatic γ-GT, amount of hepatic TG were elevated significantly compared with those in the control group (all P<0.01). JPHXD decreased the ratio, activities of ALT, AST, γ-GT and TG significantly compared with those in the AALI group (P<0.05 or P<0.01). HE and oil red O staining showed that fat deposited markedly in liver tissue, while JPHXD alleviated pathological changes markedly. Plasma LPS level in rat portal vein in the AALI group increased significantly (P<0.01), but it was decreased significantly in the JPHXD group (P<0.01). The cluster analysis and principal component analysis of ERIC-PCR fingerprint showed that gut flora in the AALI group changed markedly, and JPHXD could recover gut flora to some extent.</p><p><b>CONCLUSIONS</b>The structure of gut flora shifted markedly during acute alcoholic liver injury, JPHXD had prevention effect through the modification of gut flora.</p>

Qushi Huayu Decoction (祛湿化瘀方) inhibits protein and gene expression of cathepsin B in HepG2 cells induced by free fatty acids / 中国结合医学杂志

<p><b>OBJECTIVE</b>To study the experimental efficacy of Qushi Huayu Decoction (祛湿化瘀方,QHD) on, protein and gene expression of cathepsin B (ctsb) in HepG2 cells induced by free fatty acids (FFAs).</p><p><b>METHODS</b>The model of HepG2 steatosis and tumor necrosis factor-α (TNF-α) secretion was induced by long-chain FFAs. HepG2 cells were divided into 4 groups: control group (group C), model group (group M), low-dose QHD group (group L) and high-dose QHD group (group H). Long-chain FFAs were added to groups M, L and H. The 10% blank-control serum was added to group C and M, while 5% and 10% QHD-containing sera were added to group L and H, respectively. The levels of serum TNF-α and cellular triglyceride (TG) were detected. Cellular p-IκB and ctsb expression were detected using Western blot and PCR. The expression and distribution of ctsb were observed by immunofluorescence.</p><p><b>RESULTS</b>After incubating with FFA for 24 h, TG deposition in HepG2, TNF-α content in cell supernatant, the protein expression of cellular ctsb and P-IκB, as well as mRNA expression of ctsb increased markedly in group M compared with group C (P<0.05, P<0.01). Compared with group M, TG deposition, the expression of cellular ctsb, P-IκB and ctsb mRNA in groups L and H, as well as TNF-α content in group H, decreased significantly (P<0.05). Cell immunochemical fluorescence studies showed that ctsb was released from lysosomes and distributed in the cytoplasm extensively and diffusedly after being stimulated with FFA. In this study, these above-mentioned changes were inhibited markedly in groups L and H.</p><p><b>CONCLUSION</b>QHD might have a direct inhibitory effect on the ctsb target in the FFA-ctsb-TNFα pathway of hepatic lipotoxicity.</p>

The role of adiponectin and adiponectin receptor 2 in the pathology of fatty liver / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the role of adiponectin (ADP) and adiponectin receptor 2 (adipoR2) in pathology of fatty liver, and to investigate the effect of Chinese herbal decoction (Qushi Huayu Decoction, QHD) on fatty liver disease.</p><p><b>METHODS</b>Two experimental fatty liver models were used. One was induced with high-fat diet for ten weeks, and the rats were divided into normal, model and QHD group, the QHD group was administrated with QHD during the last four weeks. The other experimental fatty liver model was induced by subcutaneous injection of carbon tetrachloride (CCl4) in combination with high-fat and low-protein diet for four weeks, and the rats were also divided into normal, model and QHD group, the QHD group was administrated with QHD during the last two weeks. The observation items include: (1) hepatic steatosis (H.E. staining); (2) serum ADP, hepatic triglyceride (TG), free fatty acid (FFA) and adipoR2; (3) correlation among serum ADP content, hepatic TG, FFA and adipoR2.</p><p><b>RESULTS</b>(1) Serious hepatic steatosis, increased hepatic TG and FFA, decreased serum ADP and hepatic adipoR2 were observed in the two models (P less than 0.01). QHD administration significantly reduced the hepatic TG and FFA, and increased serum ADP and hepatic adipoR2 (P less than 0.01) in these two models. (2) Inverse correlation was observed between hepatic TG, FFA and serum ADP, hepatic adipoR2 in these two models.</p><p><b>CONCLUSION</b>(1) Decreased serum ADP and hepatic adipR2 may play important roles in pathological process of fatty liver. (2) QHD administration increased the serum ADP and hepatic adipoR2.</p>

Effect of Jianpi Huoxue decoction-containing serum on tumor necrosis factor-alpha secretion and gene expression of endotoxin receptors in RAW264.7 cells induced by lipopolysaccharide / 中国结合医学杂志

<p><b>OBJECTIVE</b>To evaluate the effect of Jianpi Huoxue decoction (JHD)-containing serum on tumor necrosis factor-alpha (TNF-alpha) secretion and endotoxin receptor gene expression in RAW264.7 cells induced by lipopolysaccharide (LPS).</p><p><b>METHODS</b>The cytotoxicity of blank-control serum and JHD-containing serum at different concentrations were evaluated through the lactate dehydrogenase (LDH) assay in RAW264.7 cells. RAW264.7 cells were divided into six groups: 5% blank-control serum group (C1, n=3), 5% blank-control serum plus LPS group (L1, n=4), 5% JHD-containing serum plus LPS group (J1, n=4), 10% blank-control serum group (C2, n=3), 10% blank-control serum plus LPS group (L2, n=4), and 10% JHD-containing serum plus LPS group (J2, n=4). After cultured with the corresponding serum for 1 h, cells in L1, L2, J1 and J2 were treated with LPS (0.1 microg/mL) for 12 h without rinse. The supernate, cells, protein and RNA were collected for assay. TNF-alpha in the culture supernate was assayed by the enzyme linked immunosorbent assay (ELISA). Protein expression of TNF-alpha in RAW cells was detected by Western-blot. TNF-alpha, Toll-like receptor 2 (TLR2), TLR4 and CD14 mRNA expression in RAW cells were detected by real-time RT-PCR.</p><p><b>RESULTS</b>The LDH assay supported that cultured for 24 h or less with the JHD-containing serum at the concentration of 10% or lower, RAW264.7 cells showed no cytotoxicity. After stimulation with LPS for 2 h, TNF-alpha in the culture supernate of the 5% blank-control serum plus LPS group (L1, P=0.03), 10% blank-control serum plus LPS group (L2, P=0.002) and in the cell layer (P=0.01) of these groups increased remarkably. After stimulation with LPS for 1 h, the mRNA expression of TNF-alpha (P=0.004), TLR (P=0.03), CD14 (P=0.004) was up-regulated obviously. In the 10% JHD-containing serum plus LPS group (J2), the protein expression of TNF-alpha in both supernate (P=0.04) and cell layer (P=0.04), gene expression of TNF-alpha (P=0.03), TLR4 (P=0.001), CD14 (P=0.001) were all inhibited. On the other hand, the TLR2 mRNA expression was not up-regulated after LPS stimulation in the 10% blank-control serum plus LPS group (L2).</p><p><b>CONCLUSION</b>JHD-containing serum inhibited the LPS-induced cytokines expression in RAW264.7 which was probably associated with its inhibitory effect on the mRNA expression of LPS receptors TLR and CD14.</p>