Effects of Pearl Hydrolysate on Hepatic Sinusoidal Endothelial Cell Viability and Capillarization in Liver Fibrosis / 中国医学科学院学报

Objective To study the effect and mechanism of pearl hydrolysate on hepatic sinusoidal capillarization in liver fibrosis. Methods Hepatic sinusoidal endothelial cells (HSEC) and hepatic stellate cells (HSC-LX2) were incubated with Hepu pearl hydrolysate.The proliferation of HSEC and HSC-LX2 was examined by MTT colorimetry.The cell cycle and apoptosis of HSC-LX2 were measured by flow cytometry.The changes of the microstructures such as fenestra and basement membrane of HSEC were observed by transmission electron microscopy. Results The intervention with leptin increased the viability of HSC-LX2 (P=0.041),decreased the viability of HSEC (P=0.004),and caused capillarization signs such as decreased number and diameter of fenestrae and formation of continuous basement membrane.The treatment with pearl hydrolysate at different doses increased and expanded the fenestrae of HSEC (low dose:P=0.020;medium dose:P=0.028;high dose:P=0.032),disintegrated the extracellular basement membrane of HSEC (low dose:P=0.020;medium dose:P=0.028;high dose:P=0.032),decreased the viability of HSC-LX2 (low dose:P=0.018;medium dose:P=0.013;high dose:P=0.009),and induced the apoptosis of HSC-LX2 (low dose:P=0.012;medium dose:P=0.006;high dose:P=0.005).Pearl hydrolysate exerted therapeutic effect on capillarization in a dose-dependent manner (low dose:P=0.020;medium dose:P=0.028;high dose:P=0.032).Moreover,high-dose pearl hydrolysate showed stronger effect on capillarization of hepatic sinuses than colchicine (P=0.034) and salvianolic acid B (P=0.038). Conclusion Hepu pearl hydrolysate can increase the viability of HSEC,restore the area of fenestrae,disintegrate the basement membrane,and decrease the viability and induce the apoptosis of HSC-LX2,demonstrating significant pharmacological effects on the capillarization of HSEC and HSC-LX2.

Identification of the metabolites from co-cultures of marine Streptomyces sp. IMB18-531 and Cladosporium sp. IMB19-099 / 药学学报

A new siderophore chelate (1) and 8 known compounds were identified from the liquid co-cultures of the marine-derived Streptomyces sp. IMB18-531 and Cladosporium sp. IMB19-099 by a combination of chromatography methods, including C18 reversed-phase medium pressure chromatography, gel column chromatography and HPLC. Their structures were determined by spectroscopic analysis and chemical methods as aluminioxamine E (1), desferrioxamine E (2), ferrioxamine E (3), terragine E (4), capsimicin (5), cyclo(L-prolinyl-L-tyrosine) (6), anthranilic acid (7), (Z)-14-methylpentadec-9-enoic acid (8), and (Z)-hexadec-8-enoic acid (9). Compound 2 showed inhibitory activities against the expression of liver fibrosis related genes COL1A1, MMP2, and TIMP2. Compounds 5, 8, and 9 displayed antibacterial activities against methicillin-resistant Staphylococcus aureus, S. epidermidis and Bacillus subtilis, with MICs of 16-64 μg·mL-1. Compound 5 showed cytotoxicities against human pancreatic cancer MIA Paca-2 and human colon cancer HT-29 cell lines with IC50 of 2.9 and 6.3 μmol·L-1, respectively.

Preparation of evodiamine-glycyrrhizic acid micelles with glycyrrhizic acid as carrier and their anti-hepatic fibrosis activity / 中国中药杂志

This study aimed to prepare evodiamine-glycyrrhizic acid(EVO-GL) micelles to enhance the anti-hepatic fibrosis activity of evodiamine. Firstly, EVO-GL micelles were prepared with use of thin film dispersion method. With particle size, encapsulation efficiency, loading capacity of micelles and the solubility of evodiamine as the indexes, the effect of different factors on micelles was observed to screen the optimal preparation methods and process. Then the pharmaceutical properties and the therapeutic effects of EVO-GL micelles prepared by optimal process were evaluated on CCl_4-induced hepatic fibrosis. The results showed that the micelles prepared by the thin film dispersion method had an even size, with an average particle size of(130.80±12.40)nm, Zeta potential of(-41.61±3.12) mV, encapsulation efficiency of 91.23%±1.22%, drug loading of 8.42%±0.71%, high storage stability at 4 ℃ in 3 months, and slow in vitro release. Experimental results in the treatment of CCl_4-induced hepatic fibrosis in rats showed that EVO-GL micelles had a synergistic anti-hepatic fibrosis effect, which significantly reduced the liver function index of hepatic fibrosis rats. In conclusion, the EVO-GL micelles prepared with glycyrrhizic acid as a carrier would have a potential application prospect for the treatment of hepatic fibrosis.

Effects of surfactants on water solubility and anti-hepatic fibrosis of curcuminoids / 中草药

Objective To increase the water solubility of curcumin constituents by three kinds of food grade surfactants, and to study the effect of surfactants on the quality standard and stability and anti-hepatic fibrosis in vivo of curcumin solution. Methods The high-speed dispersion method was used to prepare the solution of curcuminoids-surfactants. The qualitative analysis of curcuminoids was confirmed by LC-MS/MS. HPLC-DAD fingerprinting was established and quantitative analysis was conducted. And the effects on anti-liver fibrosis of different concentrations curcumin ratio solution were evaluated by rat model. Results The maximum water solubility of curcumin constituents was 1.387 mg/mL. LC-MS/MS analysis showed that the structure of curcuminoids were stable; The results of fingerprinting analysis demonstrated that the similarity of 18 batches of samples were higher than 99%. The contents of curcumin, demethoxycurcumin and bisdemethoxycurcumin in different batches of samples were stable with the RSD of 1.60%, 2.24%, and 3.74%. The rat model of hepatic fibrosis showed that the contents of AST and ALT in free curcumin suspension group were (64.1 ± 20.3) and (45.1 ± 13.9) U/L, respectively, while that solution of curcuminoids-surfactants group was (19.4 ± 8.7) and (11.8 ± 4.9) U/L. Conclusion The solubility of curcuminoids was increased 517 times compared with the free state (2.677 μg/mL) after solubilization with surfactants, and repeated tests indicated that the structure and contents of curcuminoids were stable; the solubilized curcuminoids exhibited better anti-liver fibrosis effects than the free.

Research progress in drugs for anti-hepatic fibrosis / 药学学报

Hepatic fibrosis is an important pathological process in the development of liver cirrhosis and liver cancer from chronic liver damage. So far there is no effective chemical drug in clinic for treatment of hepatic fibrosis. Therefore, the research in anti-hepatic fibrosis drugs is a hot topic. For drugs currently under research and development, most of the mechanisms of action are related to inhibition of factors that could cause or deteriorate liver fibrosis, including activation and proliferation of hepatic stellate cells, inflammation, oxidative stress and production of extracellular matrix, et al. In this review, we briefly analyze targets and drugs related to the mechanisms mentioned above in order to provide a reference to the future research and development of anti-hepatic fibrosis drugs.

Discrimination of proliferation inhibiting ingredients in Corydalis soxicola on rat hepatic stellate cell-T6 based on composition-activity relationship / 中草药

Objective: To establish the composition-activity relationship (CAR) model based on study of chemical composition and relating proliferation inhibitory rate of Corydalis soxicola and recognize the anti-hepatic fibrosis active compounds of C. soxicola. Methods: Nine orthogonal C. soxicola extract were analyzed by HPLC, and 21 characteristic peaks were profiled. Anti-hepatic fibrosis activity was investigated by MTT assays on HSC-T6, and the potential active components were identified by scores plot and variable importance in projection (VIP) values by means of orthogonal partial least squares (OPLS) analysis, and the activities of identified components were verified by MTT and flow cytometry. LDH kit was used to detect the effect of dehydrocavidine, palmatine, and berberine on LDH activity. Results: The results showed that six peaks including peaks 13, 14, 16, 18, 19, and 20 were significantly related to anti-hepatic fibrosis activity among nine orthogonal extract from C. soxicola. Peaks 18, 19, and 20 were characterized as dehydrocavidine, palmatine, and berberine, respectively. MTT assay showed that dehydrocavidine, palmatine, and berberine with various concentration significantly inhibited the proliferation of HSC-T6 cells. It was also found in flow cytometry that the apoptotic rates of dehydrocavidine, palmatine, and berberine (0.10 mg/mL) on HSC-T6 were 42.12%, 42.22%, and 36.73%, respectively, which were obviously higher than that of control (1.69%, P < 0.01). No obvious cytotoxic effect was found when the final concentration of dehydrocavidine, palmatine, and berberine were less than 0.15, 0.10, and 0.10 mg/mL, respectively. Conclusion: In this study, it was for the first time found that dehydrocavidine, palmatine, and berberine in C. soxicola extract can effectively inhibit the proliferation and induce apoptosis of HSC-T6 based on composition-activity relationship, and there was no obvious cytotoxic effect of the effective concentration, which showed that they may be the active components with potential anti-hepatic fibrosis effect and have a certain security in the application in C. soxicola. At the same time, it also suggests that the research ideas based on CAR can provide an effective method for the identification of active components in natural plants.

Role of Collagen Metabolism in Liver Fibrosis and Research Progress in Traditional Chinese Medicine on the Regulation of Collagen Metabolism / 中国药师

The abnormal accumulation of extracellular matrix (ECM) in liver is the main feature of hepatic fibrosis,and collagen is the most important component in ECM.Collagen plays an important role in the occurrence and development of liver fibrosis.Therefore,to find traditional Chinese medicine with significant effect on collagen metabolism has become a critical approach for the treatment of hepatic fibrosis.This review summarized the role of collagen metabolism in liver fibrosis,and the research progress in traditional Chinese medicines with anti-hepatic fibrosis effect by regulating collagen metabolism was explored as well.

Effect of Loureirin A on Proliferation and Frizzled-4 Expression of Rat Hepatic Stellate Cells in vitro / 昆明医科大学学报

Objective To investigate the molecular mechanism of Loureirin A mediated anti-hepatic fibrosis by evaluting its effects on proliferation , secretion ofα-smooth muscle actin (α-SMA) and transforming growth factor-beta1 (TGF-β1), and expression of rat hepatic stellate cells in vitro . Methods Primary hepatic stellate cells were isolated and cultured from Sprague-Dawley rats. After activating and inducing primary hepatic stellate cells from qHSC to aHSC, the activated hepatic stellate cells model in vitro was established. Then we observed the morphological changes of static hepatic stellate cells and activated hepatic stellate cells with inverted phase contrast microscope. Cultured hepatic stellate cells were treated with different concentrations of loureirin A and the inhibitory rate of HSCs proliferation was measured by MTT assay. The expression of Frizzled-4 was measured by western blot analysis. The content ofα-SMA and TGF-β1 in the cultured HSCs'supernatant were measured by enzyme-linked immunosorbent assay (ELISA) . Results Loureirin A the proliferation of inhibited activated hepatic stellate cells in a time-dose-dependent manner compared with the control group,IC50=0.30 μg/μL. After loureirinA treatment of the HSCs, western blot analysis showed that Frizzled-4 expression level was obviously lower than control group. Loureirin A also inhibitedα-SMA and TGFβ1 (P<0.05) secretion in the cultured HSCs'supernatant in different degree by the assay of ELISA. Conclusions The molecular mechanism of Loureirin A and Wnt signaling pathway mediated anti-hepatic fibrosis and anti-angiogenesis may involve down-regulation the expression of Frizzled-4, inhibiting the synthesis and secretion ofα-SMA,TGF-β1and the proliferation of HSCs.

Spectrum-activity relationship on anti-hepatic fibrosis efficacy of Trionycis Carapax / 中草药

Objective: To investigate the spectrum-activity relationship between HPLC of Trionycis Carapax from different regions and its anti-hepatic fibrosis efficacy, in order to reveal the "active component group" for anti-hepatic fibrosis efficacy of Trionycis Carapax. Methods: The samples from 12 regions were determined with the HPLC-DAD method. The representative standard fingerprint was calculated using the Similarity Evaluation System for Chromatographic Fingerprint of Traditional Chinese Medicine (Version 2004A). The common patten and the principal component analysis (PCA) were established. Using the LX-2 hepatic stellate cell as a hepatic fibrosis model of Trionycis Carapax to study its inhibition on the hepatic fibrosis cells. The spectrum-effect relationship was studied by SPSS 19.0 software. Results: Seven common peaks in the HPLC-fingerprint of Trionycis Carapax were obtained. It was tentatively concluded that peaks 2 and 5 related better to the inhibitory effect of LX-2 cells (OD value) in the seven characteristic peaks. Peak 4 had the strongest correlation. And the quality of Trionycis Carapax was influenced by the concentration of peak 4 obtained by PCA. Conclusion: All the samples could inhibit the proliferation of LX-2 hepatic stellate cell in some extent. There may be a certain relationship between HPLC fingerprint and anti-hepatic fibrosis efficacy. In addition, the research could be used as the quality control method of Trionycis Carapax.

Experimental Study on Bupleuri in Combination With Saliva Miltiorrhiza in Treatment of Hepatic Fibrosis in Rats / 中国药房

OBJECTIVE:To investigate the effects of Bupleuri plus Saliva miltiorrhiza on preventing the formation of hepatic fibrosis in rats METHODS:To establish an animal model of hepatic fibrosis by feeding high fat diet and 10% alcohol and subcutaneously injecting CCl4 and to detect the zymogram and biochemical parameters and observe the pathological changes of liver tissue RESULTS:Bupleuri and Saliva miltiorrhiza could significantly depress the activities of ALT,AST,MAO,ALP and had no influence on the ?-GT in serum,the serum TG was decreased and TG in tissue was obviously raised and serum GSH was elevated in hepatic fibrosis rat The extract could significantly decreased the levels of PC Ⅰ and PC Ⅲ in hepatic fibrosis rat The histopathological examination showed that the extract could diminish liver degeneration and necrosis,and block the formation of fiber diazoma,the degree of collagen proliferation was significantly lower than that of the control group CONCLUSION:Bupleuri and Saliva miltiorrhea extract could effectively protect the liver cell,and could retard the formation of hepatic fibrosis as well