Assessment of Feng-Liao-Chang-Wei-Kang as a potential inducer of cytochrome P450 3A4 and pregnane X receptors.

Feng-Liao-Chang-Wei-Kang (FLCWK), a traditional Chinese patent medicine, consists primarily of Polygonum hydropiper and Daphniphyllum calycinum roots. As a complex containing several kinds of flavonoids, FLCWK has the potential to impact the drug metabolism enzyme P450 3A4 (CYP3A4) and nuclear receptors. The purpose of this research was to probe the effects of FLCWK on CYP3A1, the homolog of CYP3A4 in rats, and to confirm whether FLCWK interferes with PXR and CAR-mediated transactivation of CYP3A4. The effects of FLCWK on Cyp3a1 mRNA, catalytic activity levels, and protein expression in Sprague-Dawley (SD) rat liver tissues were examined using real-time PCR, western blotting, and high-performance liquid chromatography (HPLC) assays, respectively. The efficacy of PXR and CAR on CYP3A4 transcriptional activity were detected using luciferase reporter assays and further research of the impact of FLCWK on CYP3A4 gene expression mediated by the PXR pathway was examined by transient transfection of PXR siRNA. FLCWK significantly increased Cyp3a1 mRNA, CYP3A1 activity, and protein expression levels in SD rats. FLCWK highly induced CYP3A4 luciferase activity mediated by PXR in PXRCYP3A4 co-transfected cells. A siRNA-mediated drop-off in PXR expression greatly cut the effect of FLCWK on CYP3A4 mRNA expression in HepG2 cells. These findings show that FLCWK up-regulates CYP3A4 levels via the PXR pathway. This effect should be considered being applied in clinical use as FLCWK has the potential to interact with other drugs.

[Puerarin inhibits DNA damage of HaCaT cells induced by UVB via ceramide pathway].

This study is to investigate the effect and mechanism of puerarin on DNA damage of HaCaT cells induced by UVB. Puerarin pre-treated cells were irradiated with UVB at 30 mJ x cm(-2). Twenty four hours after irradiation, DNA damage was detected by comet assay, ceramide was measured by thin layer chromatography and gas chromatography, intracellular free calcium ion was analyzed by flow cytometry, the phosphorylation level of p38 protein was examined by Western blotting method. Levels of DNA damage, ceramide, free calcium ion and p-p38 protein were elevated in UVB model cells. Contrary to the model group, all indicators above were reduced in all groups pre-treated by puerarin. Puerarin restrains the ceramide accumulation to block downstream p38 MAPK pathway and calcium ion rising, therefore reduces DNA damage in HaCaT cells induced by UVB.

Feng-Liao-Chang-Wei-Kang Combined with 5-Fluorouracil Synergistically Suppresses Colitis-Associated Colorectal Cancer via the IL-6/STAT3 Signalling Pathway.

BACKGROUND: Colitis-associated colorectal cancer (CAC) develops from active colonic inflammation, which is characterized by the production of proinflammatory cytokines that can induce mutations. IL-6 is produced by multiple cell types located within the tumor microenvironment including tumor-infiltrating immune cells, stromal cells, and the tumor cells themselves. The aim of our study was to explore the mechanism of Feng-Liao-Chang-Wei-Kang (FLCWK) and 5-fluorouracil (5-FU) in treating CAC. METHOD: HCT116 cells were treated with 5-FU in the absence or presence of FLCWK. Cell proliferation was assayed by MTT assays. Apoptosis and the cell cycle phases were detected by flow cytometry. Western blotting and Q-PCR assays were used to detect the expression levels of proteins and genes related to the IL-6/STAT3 signalling pathway. A mouse model for CAC was established by treating animals with 12.5 mg/kg azoxymethane (AOM) followed by 3 cycles of 2.5% dextran sodium sulphate (DSS). The associated pathological changes were determined after haematoxylin and eosin (H&E) staining. The expression of related proteins and genes in various tissues was examined using immunofluorescence techniques. RESULTS: FLCWK enhanced the ability of 5-FU to promote apoptosis by inhibiting the proliferation of HCT116 cells and blocking the IL-6/STAT3 pathway. FLCWK combined with 5-FU reduced the number and size of colon tumors in mice with CAC and significantly increased their survival rate. In the CAC model, FLCWK synergized with 5-FU to inhibit the phosphorylation of STAT3, preventing IL-6/STAT3 signal transduction and thus further inducing apoptosis and inhibition of colon cancer cell proliferation. CONCLUSION: FLCWK can inhibit the activation of STAT3 by reducing the production of IL-6, thereby increasing the occurrence of colitis-related colorectal cancer with 5-FU.

Improvement and optimization of a T-cell-dependent antibody response (TDAR) method for BALB/c mice using keyhole limpet hemocyanin (KLH) as specific antigen.

Although T-cell-dependent antibody response (TDAR) assays with keyhole limpet hemocyanin (KLH) as specific antigen have many advantages, most experiments produce qualitative results based on antibody titers. It was hypothesized that if experimental conditions (like antigen coating concentration, serum dilution, and detecting [here, horseradish peroxidase-goat anti-mouse IgG] antibody dilution) could be optimized, the resulting measured value (here, optical density) could be used to directly analyze and evaluate the experimental results. This means specifically that the assay OD values could be used for approximate quantitative statistical analysis; it does not require a further conversion of the data into qualitative forms or require obtaining further titer data from additional experiments. As such, the use of this "improved" assay would: greatly reduce the complexity of experimental operations; improve overall sensitivity and practicality of traditional TDAR assays; and, allow for direct assessing of any immunosuppression caused by a test drug in a host. Here, KLH-immunized serum was obtained from BALB/c mice, and the means to detect serum anti-KLH antibodies by an indirect ELISA were optimized. The results indicated that in this system, the optimal KLH coating concentration was 80 µg/ml, the optimal dilution range of the serum (at immunization dose of 5 mg KLH/kg) was 1:200-1:800, and the optimal dilution of HRP-goat anti-mouse IgG antibody was 1:16,000. Methodology verification was performed and a regression model of y = 144.16x + 0.9815 (R2 = 0.9571, indicating very good linearity) was obtained. Intragroup precision was 7.51-9.40%; the intergroup coefficient of variation was 9.83-14.22%. The lower limit of detection was 0.1385. The present results showed this indirect ELISA exhibited very good linearity, accuracy, and precision.

Treatment of unstable angina pectoris with modified nuan gan jian--a report of 33 cases.

OBJECTIVE: To observe the effectiveness of Modified Nuan Gan Jian Jiao Nang (MNGJ [symbol: see text] Modified Liver-Warming Capsule) in treating unstable angina pectoris. METHOD: Sixty-six eligible cases were assigned randomly into a treatment group and a control group and treated for 3 weeks. RESULTS: MNGJ produced an effect in reducing episodes, improving the abnormal findings in electro-cardiogram (ECG) (P<0.05) significantly, and decreasing myocardial oxygen consumption (P<0.05). In addition, it could decrease the plasma TXB2 level and increase 6-keto-prostaglandin F1alpha (6-Keto-PGF1alpha) level (P<0.01). CONCLUSION: MNGJ was quite effective in treating unstable angina pectoris, suggesting that the treatment of the disease could start from the liver.