interleukin-8 and cepharanthine

Cepharanthine is a biscoclaurine alkaloid extracted from Stephania cepharantha Hayata, which is widely used for the treatment of many acute and chronic diseases, and can exert antitumor effects on several human cancer cell lines. However, little is known about the detailed mechanisms of the antitumor activity of Cepharanthine. In the present study, we determined whether Cepharanthine could suppress angiogenesis and growth of human oral squamous cell carcinoma (OSCC) cells in vitro and in vivo. Cepharanthine significantly inhibited expression of two major pro-angiogenic molecules, vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8), in cultured cells and in cells implanted into the subcutaneous tissue of nude mice. Also, Cepharanthine inhibited the nuclear factor-kappaB (NF-kappaB) activity in human OSCC cells in vitro and in vivo. The decreased expression of VEGF and IL-8 correlated with decreased tumor cell growth and decreased vascularization in vitro and in vivo. These findings suggest that Cepharanthine can suppress angiogenesis and growth of OSCC cells by inhibiting expression of VEGF and IL-8 involved in the blockade of NF-kappaB activity.

Topics: Animals; Antineoplastic Agents, Phytogenic; Benzylisoquinolines; Blotting, Western; Carcinoma, Squamous Cell; Cell Movement; Cell Proliferation; Cells, Cultured; Enzyme-Linked Immunosorbent Assay; Female; Humans; Immunohistochemistry; Interleukin-8; Mice; Mice, Nude; Mouth Neoplasms; Neovascularization, Pathologic; NF-kappa B; Reverse Transcriptase Polymerase Chain Reaction; Vascular Endothelial Growth Factor A; Xenograft Model Antitumor Assays

We have already demonstrated that human head and neck cancer cells have significantly enhanced levels of transcription factor nuclear factor (NF)-kappaB activity compared to their normal counterparts, suggesting that NF-kappaB plays an important role in the development of head and neck cancer. However, it has been reported that chemotherapeutic agents and radiation activate NF-kappaB activity in cancer cells, thus making the cells radioresistant and chemoresistant. In addition, we have shown that the suppression of NF-kappaB activity enhanced apoptosis in oral squamous cell carcinoma cells. In this study, we examined whether cepharanthin-induced inhibition of NF-kappaB activity enhances radiosensitivity in human oral carcinoma cells. Cepharanthin is a biscoclaurine alkaloid extracted from the roots of Stephania cepharantha hayata, and is widely used in Japan for the treatment of patients with leucopenia, nasal allergy, and venomous snakebites. gamma-irradiation (IR) induces NF-kappaB activity in oral carcinoma cells through the activation of upstream molecules, including Akt and IkappaB kinase. However, a luciferase assay revealed that cepharanthin suppresses IR-induced NF-kappaB activity in oral squamous cell carcinoma cells, thereby enhancing the radio-sensitivity. Western blot analysis showed an enhanced cleavage of poly-(ADP-ribose) polymerase protein in carcinoma cells by both cepharanthin treatment and IR exposure compared to IR or cepharanthin alone. In an in vivo study, B88 cells were s.c. inoculated into the backs of nude mice. Tumor-bearing nude mice received either cepharanthin, IR alone, or a combination of cepharanthin and IR. The combined treatment suppressed tumor growth significantly more than either cepharanthin or IR alone. Cepharanthin inhibited the production of IR-induced IL-6 and IL-8, which are downstream targets of NF-kappaB. In quantitative real-time RT-PCR, IR also induced the expression of anti-apoptotic proteins [cellular inhibitor of apoptosis protein (cIAP)-1 and -2] in carcinoma cells. Treatment of cancer cells with cepharanthin combined with exposure to IR decreased cIAP-1 and -2 mRNA expression. These findings suggested that the combination of radiotherapy and cepharanthin could enhance radiosensitivity in the treatment of human oral cancer.

Topics: Alkaloids; Animals; Apoptosis; Benzylisoquinolines; Blotting, Western; Carcinoma, Squamous Cell; Cell Line, Tumor; Fluorescent Antibody Technique; Gamma Rays; Humans; I-kappa B Kinase; Inhibitor of Apoptosis Proteins; Interleukin-6; Interleukin-8; Luciferases; Mice; Mice, Inbred BALB C; Mice, Nude; Mouth Neoplasms; NF-kappa B; Radiation-Protective Agents; Radiation, Ionizing; RNA, Messenger

The antitumor effect of biscoclaurine alkaloids, at a distant site was examined in the double grafted tumor system, in which mice received simultaneous intradermal inoculations of Meth-A in both right (10(6) cells) and left (2 x 10(5) cells) flanks and were then injected with 0.5 mg of CR in the right tumor on days 3, 4 and 5. CR inhibited the growth of not only the right but also the left, non-treated tumor. In order to examine the role of lymph nodes in the antitumor activity of CR, regional (axillary and inguinal) lymph nodes were resected. Since in resected mice the antitumor activity of CR against the left tumor was weakened, the regional lymph nodes have a very important role in the antitumor effect of intratumoral administration of CR at a distant site. Spleen cells prepared from CR treated mice were examined for Lyt-1, Lyt-2 and L3T4 phenotypes. The number of Lyt-1 positive lymphocytes increased in the spleen after intratumoral administration of CR. Isolated tumor cells obtained from the right tumor treated with CR and the left side tumor on day 6 were cultured for 24 h. The culture supernatants were harvested and tested for the presence of chemotactic activity for neutrophil or macrophage. Significant neutrophil chemotactic factor (NCF) activity was detected in the culture media from CR-treated right tumor tissue, and macrophage chemotactic factor (MCF) activity was detected in the culture media from left tumor tissue. CR-induced NCF was partially neutralized by treatment with anti-human IL-8 IgG, and might be murine IL-8 factor. These results suggest that intratumoral administration of CR first induces neutrophils in the right tumor, then Lyt-1 positive cells in the spleen, and subsequently induces cytotoxic macrophages in the left, non-treated tumor, thus bringing about the regression of distant tumors.

Topics: Alkaloids; Animals; Antineoplastic Agents; Benzylisoquinolines; Chemotactic Factors; Injections, Intralesional; Interleukin-8; Lymph Node Excision; Macrophages; Mice; Mice, Inbred BALB C; Sarcoma, Experimental