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We studied the apoptosis-inducing properties of the antimicrobial peptide cecropin of Musca domestica in human hepatocellular carcinoma cell line BEL-7402 and its underlying mechanism. Proliferation inhibition of the human hepatocellular carcinoma BEL-7402 cells and the human normal liver cells were determined by the MTT assay, and the cell viability was determined by trypan blue dye exclusion assay. The apoptotic tumor cells treated with cecropin were examined by transmission electron microscopy and terminal-deoxynucleotidyl transferase mediated nick end labeling. The apoptosis rate was measured by flow cytometry (FCM) with PI/Annexin-V double staining. Western blot analysis and RT-PCR were used to determine the expression levels of proteins involved in apoptosis, such as Fas, Fas-L, caspase-8, and caspase-3. The experimental results showed that Musca domestica cecropin inhibited the proliferation of human hepatocellular carcinoma BEL-7402 cells in dosedependent and time-dependent manners, without affecting the proliferation of normal liver cells. FCM showed that the cell apoptosis rates were 5.1+/-0.11%, 8.1+/-0.04%, and 10.9+/-0.15% after the treating with 100 mM cecropin for 24, 48, and 72 h, respectively. The rates of apoptosis were 5.4+/-0.14% and 8.0+/-0.13% after the treating with 25 and 50 microM cecropin for 72 h, respectively. Western blot analysis and RT-PCR showed that the apoptosisrelated molecules including Fas, Fas-L, caspase-8 and caspase-3 were activated. This study showed that the antimicrobial peptide cecropin-inducing apoptosis in human hepatocellular carcinoma BEL-7402 cells, which might be associated with upregulation of Fas, Fas-L, and caspase-8 and caspase-3 and triggering extrinsic apoptotic pathway.

To determine istanbulin A in Sarcandra glabra (Thunb.) Nakai.An RP-HPLC method was established and an hypersil ODS C18 column (4.6 mm x 200 mm, 5 microm) was used. The detective wavelength was at 218nm.Istanbulin A had a good linearity in the range of 0. 135 to approximately 2. 16 microg. The recovery rate was 98.54%.This method is simple, accurate and repeatable. It can be used to determine istanbulin A in Sarcandra glabra.

β-defensin 2 is a small antimicrobial peptide of the innate immune system and has been thought to regulate anti-tumor immunity. However, little is known on whether β-defensin 2 could modulate melanoma-specific NK and T cell responses. In this study, we first cloned the murine β-defensin 2 gene by RT-PCR and generated the β-defensin 2 stably expressing B16 cells (B16-mBD2). Subsequently, we evaluated whether vaccination with irradiated B16-mBD2 could modulate the growth of implanted B16 cells and determined the potential mechanisms underlying the action of B16-mBD2 vaccine in modulating the growth of B16 tumors in C57BL/6. We found that vaccination with irradiated B16-mBD2, but not with control B16-p or parental B16, inhibited the development and progression of B16 tumors, and prolonged the survival of tumor-bearing mice. However, vaccination with irradiated B16-mBD2 failed to inhibit the development of B16 tumors in the CD4(+)- or CD8(+)-depleted recipients. Furthermore, vaccination with irradiated B16-mBD2 stimulated strong NK activity and promoted potent B16-specific CTL responses, accompanied by augmenting IFN-γ and IL-12, but not IL-4, responses in the recipient mice. Moreover, vaccination with irradiated B16-mBD2 promoted the infiltration of CD8(+) and CD4(+) T, NK cells and macrophages in the tumor tissues. These data suggest β-defensin 2 may act as a positive regulator, promoting anti-tumor NK and T cell responses in vivo. Therefore, β-defensin 2 may be used for the development of immunotherapy for the intervention of melanoma.