Tounong Powder () extracts induce G1 cell cycle arrest and apoptosis in LoVo cells.

OBJECTIVE: To further explore the anti-cancer effect of Tounong Powder () extracts (TNSEs) on human colon cancer LoVo cells and examine the possible molecular mechanisms. METHODS: The contents of TNSEs were determined by liquid chromatograph-mass spectrometer (LC-MS) analysis after extraction with water and methanol. Variations of cell morphological features were observed using fluorescence microscopy. Cytotoxicity was determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell cycle distribution and apoptosis were analyzed using flow cytometry at different TNSE doses (0, 62.5, 125, or 250 µg/mL). Protein expressions of phosphatidylinositol 3-kinase (PI3K), phosphate protein kinase B (p-AKT), phosphate mammalian target of rapamycin (p-mTOR), p-p70s6k1, cleaved caspase-9 and -3 were detected using Western blot analysis. RESULTS: TNSEs induced cell growth inhibition in a concentration- and time-dependent manner. Flow cytometric analysis showed apoptotic cells and cell cycle arrest at the G phase after TNSEs treatment compared with controls. Furthermore, TNSEs significantly down-regulated the proteins PI3K, p-AKT, p-mTOR, and p-p70s6k1, and up-regulated the proteins cleaved caspase-9 and -3 dosedependently, as determined by Western blot. CONCLUSIONS: TNSEs reduced LoVo cell proliferation, and caused apoptosis and cell-cycle arrest in LoVo cells. This effect might be associated with regulation of the PI3K/AKT signaling pathway.

Progression of Large Lymphoma Is Significantly Impeded with a Combination of Gemcitabine Chemotherapy and Dendritic Cells Intra-Tumor Vaccination.

Relapsed, refractory lymphoma remains to be a challenge and lacks efficient treatment. Some tumor cells escape from treatment, become resistant to chemotherapeutic agents, and rapidly regenerate into large tumors. Lymphoma cells induce accumulation of Gr-1(+-)CD11b(+) myeloid derived suppressor cells (MDSCs) in lymphatic organs and their vicinity. MDSCs enable tumor cells to escape from immune cells mediated surveillance and attack. Gemcitabine is a chemotherapeutic agent that eliminates both tumor cells and MDSCs, improving the immune environment favorable for subsequent treatment. We evaluated the effects of low dose gemcitabine combined with intra-tumorally delivered dendritic cells (DCs) for the treatment of A20 large-size lymphoma. We showed that MDSCs increased markedly in lymphoma-bearing mice, and that gemcitabine significantly increased the apoptosis of MDSCs. Treatment of lymphoma with either gemcitabine or intra-tumoral DCs alone could not inhibit tumor growth or rescue lymphoma-bearing mice. Treatment of lymphoma with small dose gemcitabine followed by intra-tumorally injected DCs significantly improved the efficacy of either individual treatment by reducing MDSCs, inducing onsite DCs maturation, eliminating tumor cells, inhibiting tumor growth and relapse, and extending the survival of the lymphoma-bearing mice, partly through the induction of the IFNγ secreting cells and the activation of cytotoxic lymphocytes. We showed that NK cells and CD8(+ )T cells were the major effectors to mediate the inhibition of tumor growth. Thus, the observation that gemcitabine synergizes DCs mediated immunotherapy to improve the efficacy of large size lymphoma treatment provides an experimental basis for the combination of chemotherapy and immunotherapy for the efficient treatment of relapsed or refractory lymphoma.

The effect of tou nong san on transplanted tumor growth in nude mice.

Tou Nong San (TNS) is a traditional Chinese medicinal decoction used to treat sores and carbuncles. It contains four herbal drugs and one animal medicine: Radix Astragaliseu Seu Hedysari, Angelica sinensis, Ligustici Chuanxiong, Spina Gleditsiae, and stir-baked Squama Manis. Previous studies have shown that it has anticancer effects. This report validates in vivo antitumor properties of TNS. The compounds contained in TNSE were confirmed by liquid chromatographmass spectrometer (LC-MS) analysis. The in vivo antitumor activity of TNS extract (TNSE) was tested by feeding it to athymic mice harboring a human colonic tumor subcutaneous xenograft. Toxicity was monitored by recording behavior and weight parameters. Seven compounds were detected in TNSE by LC-MS. TNSE was fed to athymic mice for 2 weeks. No adverse reactions were reported. Compared to the control group, administration of TNSE to tumor bearing mice significantly reduced both tumor weight and volume. The expressions of p-PI3K, p-AKT, p-mTOR, p-p70s6k1, VEGF, and CD31 were significantly reduced, the expression levels of cleaved Caspase-9 and cleaved Caspase-3 were significantly increased in the TNSE groups compared to the control group as determined by western blot and immunohistochemistry. TNSE produced anticolonic cancer effects and the underlying mechanisms involved inhibition of the PI3K/AKT signal transduction pathway, inhibition of angiogenesis, and promotion of apoptotic proteins.

Tounongsan extract induces apoptosis in cultured Raji cells.

OBJECTIVE: To investigate the effects of Tounongsan () extract (TNSE) on proliferation and apoptosis of the human lymphoma cell line Raji and its possible mechanism of action. METHODS: The viability of TNSE-treated Raji cells was measured by a 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Cell apoptosis was determined by flow cytometry. The molecular mechanisms of TNSE-mediated apoptosis were further investigated by reverse transcription-polymerase chain reaction (RT-PCR) analysis of the mRNA expression of nuclear factor κB (NF-κB), Bcl-xL, Bcl-2-associated death promoter (Bad), caspase-9 and caspase-3. Western blotting was used to detect the protein expressions of NF-κB, Bad, cleaved caspase-9 and cleaved caspase-3. RESULTS: TNSE inhibited Raji cell proliferation in dose- and time-dependent manners. After 48-h treatment with various concentrations of TNSE (125, 250 and 500 µg/mL), the apoptosis rates of Raji cell were 12.23%±1.98% (P<0.05), 20.97%±3.96% (P<0.01) and 30.4%±4.87% (P<0.01), respectively, compared with those of the control (6.02%±1.01%). RT-PCR demonstrated that NF-κB mRNA expression was significantly downregulated in Raji cells treated with 250 µg/mL TNSE for 48 h (P<0.05), while Bad, caspase-9 and caspase-3 mRNA levels were upregulated (P<0.05). Moreover, TNSE treatment resulted in downregulation of NF-κB protein expression and strikingly upregulated protein expressions of Bad, cleaved caspase-9, cleaved caspase-3 in a dose-dependent manner, as determined by Western blot. CONCLUSION: TNSE exhibits significant anti-proliferative and apoptotic effects in Raji cells, which may be involved in regulation of NF-κB and Bad, and activation of caspase-9 and caspase-3.