Toxicity attenuation processing technology and mechanism of Rhizoma Dioscoreae Bulbiferae stir-fried with Paeoniae Radix Alba decoction / 中国中药杂志

This study explored toxicity attenuation processing technology of Rhizoma Dioscoreae Bulbiferae stir-fried with Paeoniae Radix Alba decoction for the first time, and further explored its detoxification mechanism. Nine processed products of Rhizoma Dioscoreae Bulbiferae stir-fried with Paeoniae Radix Alba decoction were prepared by orthogonal experiment with three factors and three levels. Based on the decrease in the content of the main hepatotoxic component diosbulbin B before and after processing of Rhizoma Dioscoreae Bulbiferae by high-performance liquid chromatography, the toxicity attenuation technology was preliminarily screened out. On this basis, the raw and representative processed products of Rhizoma Dioscoreae Bulbiferae were given to mice by gavage with 2 g·kg~(-1)(equival to clinical equivalent dose) for 21 d. The serum and liver tissues were collected after the last administration for 24 h. The serum biochemical indexes reflecting liver function and liver histopathology were combined to further screen out and verify the proces-sing technology. Then, the lipid peroxidation and antioxidant indexes of liver tissue were detected by kit method, and the expressions of NADPH quinone oxidoreductase 1(NQO1) and glutamate-cysteine ligase(GCLM) in mice liver were detected by Western blot to further explore detoxification mechanism. The results showed that the processed products of Rhizoma Dioscoreae Bulbiferae stir-fried with Paeoniae Radix Alba decoction reduced the content of diosbulbin B and improved the liver injury induced by Rhizoma Dioscoreae Bul-biferae to varying degrees, and the processing technology of A_2B_2C_3 reduced the excessive levels of alanine transaminase(ALT) and aspartate transaminase(AST) induced by raw Rhizoma Dioscoreae Bulbiferae by 50.2% and 42.4%, respectively(P<0.01, P<0.01). The processed products of Rhizoma Dioscoreae Bulbiferae stir-fried with Paeoniae Radix Alba decoction reversed the decrease protein expression levels of NQO1 and GCLM in the liver of mice induced by raw Rhizoma Dioscoreae Bulbiferae to varying degrees(P<0.05 or P<0.01), and it also reversed the increasing level of malondialdehyde(MDA) and the decreasing levels of glutathione(GSH), glutathione peroxidase(GPX), and glutathione S-transferase(GST) in the liver of mice(P<0.05 or P<0.01). In summary, this study shows that the optimal toxicity attenuation processing technology of Rhizoma Dioscoreae Bulbiferae stir-fried with Paeoniae Radix Alba decoction is A_2B_2C_3, that is, 10% of Paeoniae Radix Alba decoction is used for moistening Rhizoma Dioscoreae Bulbiferae and processed at 130 ℃ for 11 min. The detoxification mechanism involves enhancing the expression levels of NQO1 and GCLM antio-xidant proteins and related antioxidant enzymes in the liver.

Gender differences in antidepressant effect of raw Rehmanniae Radix / 中国中药杂志

For the first time, this study evaluated the gender differences and mechanisms of the antidepressant effects of raw Rehmanniae Radix(RRR) based on the classic depression model with traditional Chinese medicine syndrome of Yin deficiency and internal heat. The depression model with Yin deficiency and internal heat was established by the widely recognized and applied method of thyroxine induction of the classic depression model with Yin deficiency and internal heat(chronic unpredictable mild stress). Male and female mice were simultaneously treated with RRR. The study analyzed indicators of nourishing Yin and clearing heat, conventional antidepressant efficacy test indicators, and important biomolecules reflecting the pathogenesis and prevention and treatment mechanisms of depression, and conducted a correlation analysis of antidepressant efficacy, Yin-nourishing and heat-clearing efficacy, and biological mechanism in different genders, thereby comprehensively assessing the antidepressant effects of RRR on depression of Yin deficiency and internal heat, as well as its gender differences and mechanisms. RRR exhibited antidepressant effects in both male and female mouse models, and its antidepressant efficacy showed gender differences, with a superior effect observed in females. Moreover, the effects of RRR on enhancing or improving hippocampal neuronal pathology, nucleus-positive areas, postsynaptic dense area protein 95, and synaptophysin protein expression were more significant in females than in males. In addition, RRR significantly reversed the abnormal upregulation of nuclear factor(NF)-κB/cyclooxygenase 2(COX2)/NOD-like receptor thermal protein domain associated protein 3(NLRP3) pathway proteins in the hippocampus of both male and female mouse models. The antidepressant effects of RRR were more pronounced in depression female mice with Yin deficiency and internal heat syndrome, possibly due to the improvement of neuronal damage and enhancement of neuroplasticity. The antidepressant mechanisms of RRR for depression with Yin deficiency and internal heat syndrome may be associated with the downregulation of the NF-κB/COX2/NLRP3 pathway to reduce neuronal damage and enhance neuroplasticity.

Hepatitis B virus mutations associated with in situ expression of hepatitis B core antigen, viral load and prognosis in chronic hepatitis B patients.

In this retrospective study, we investigated the prevalence and significance of mutations in part of the hepatitis B virus (HBV) x gene, and tried to clarify their relationship with clinicopathological or histopathological characteristics and prognosis in patients with chronic hepatitis B (CHB). A total of 83 consecutive CHB patients (1986-1994) were chosen for the present study. Sequence analysis was performed using polymerase chain reaction (PCR) and the direct sequencing method. The histological activity index was described using Scheuer scores. Two-step immunohistochemical staining showed the expression of viral antigens in situ. Tissue HBV DNA levels were determined by fluorescence quantitative real-time PCR. For the prognostic study, all the patients were followed up using clinical and laboratory data. Mutation at nt1726-1730 correlated significantly with decreased expression of HBcAg in situ (P = 0.006) and with lower HBV DNA levels in the liver (P = 0.004). In particular, the CTGAC mutation showed the strongest decrease of the viral load (P = 0.007). By contrast, nt1762/1764 mutation correlated with increased HBcAg (P = 0.005) and higher HBV DNA levels (P = 0.006). The mutants with the wild-type of nt1726-1730 or nt1762/1764 mutation were more prevalent in hepatocellular carcinoma (HCC) patients than in CHB patients. Although the mutations did not correlate with cirrhosis, the frequency of nt1762/1764 mutation in patients with hepatocarcinogenesis was significantly higher than in those without hepatocarcinogenesis (P = 0.011). Mutations at nt1726-1730 and nt1762/1764 are associated with in situ expression of HBcAg and viral load. Higher HBV DNA levels in the liver may be associated with hepatocarcinogenesis. Mutation at nt1762/1764 remarkably increases the risk of hepatocarcinogenesis.

[Chronic hepatitis B virus infection and the methylation status of p16INK4A promoter].

OBJECTIVE: To explore the relationship among HBV-associated histopathological indexes, x gene mutations and the methylation status of p16INK4A promoter in liver with chronic hepatitis B virus infection, in order to illustrate their role in p16INK4A hypermethylation and HCC progression. METHODS: Twenty-three cases of surgically resected HBV-associated hepatocellular carcinoma and twenty-five fine needle aspiration biopsy cases of chronic hepatitis B were chosen for this study. The methylation status of the p16INK4A promoter in tumors, their corresponding peritumoral samples and chronic hepatitis B cases was determined by methylation-specific polymerase chain reaction (MSP). EnVision two-step immunohistochemical staining showed the expression of viral antigens in situ. Tissue HBV DNA levels were determined by real-time fluorescence quantitative PCR. Polymerase chain reaction and the direct sequencing method was used for mutation analysis of HBV x gene. RESULTS: In peritumoral samples (P = 0. 025) and chronic hepatitis B cases (P = 0.029), the expression of HBx protein in methylated groups was all significantly higher than that in unmethylated groups of p16INK4A gene. But in tumors, there was no such significant difference. Other HBV antigens including HBsAg and HBcAg, tissue HBV DNA levels and point mutations of HBV x gene did not show a relationship with the methylation status of p16INK4A gene. CONCLUSION: The data suggest that p16INK4A hypermethylation correlated closely with higher HBx expression in precancerous lesions. HBx may play an important role in the early stage of HBV-associated hepatocarcinogenesis via induction of hepermethylation of p16INK4A promoter.

Association of p16INK4A hypermethylation with hepatitis B virus X protein expression in the early stage of HBV-associated hepatocarcinogenesis.

The aim of the present study was to explore the relationship between methylation status of the p16(INK4A) promoter and some HBV-related factors, and the role of these factors in p16(INK4A) hypermethylation and hepatocellular carcinoma (HCC) progression. Twenty-three cases of surgically resected HBV-associated HCC and 25 fine-needle aspiration biopsy cases of chronic hepatitis B (CHB) were studied. The methylation status of the p16(INK4A) promoter was determined by methylation-specific polymerase chain reaction (PCR). Two-step immunohistochemical staining showed the expression of viral antigens in situ. Tissue HBV-DNA levels were determined by fluorescence quantitative real-time PCR. PCR and the direct sequencing method were used for mutation analysis. In peritumoral tissues (P = 0.025) and CHB samples (P = 0.029), the expression of hepatitis B virus X protein (HBx) was higher in methylated groups of p16(INK4A) promoter than in unmethylated groups. Other HBV factors including hepatitis B surface antigen and hepatitis B core antigen, tissue HBV-DNA levels and HBV x gene mutations had no relation to the methylation status of p16(INK4A) promoter. The data indicate that p16(INK4A) promoter hypermethylation correlated closely with higher HBx expression in the precancerous lesions, suggesting that HBx may play an important role in the early stage of HBV-associated hepatocarcinogenesis via induction of hypermethylation of p16(INK4A) promoter.