Downregulation of cyclin D1 is associated with decreased levels of p38 MAP kinases, Akt/PKB and Pak1 during chemopreventive effects of resveratrol in liver cancer cells.

Resveratrol is a naturally occurring phytoalexin with antioxidant activity. The chemopreventive effects of resveratrol against various types of cancer are well known, though the underlying molecular mechanisms of its action are still not identified. Hepatocellular carcinoma (HCC) is a one of the most lethal malignancies and there is no effective treatment till date. It is known that cyclin D1 is overexpressed in liver cancers. Accordingly we have studied the chemopreventive effects of resveratrol on cyclin D1 expression and the signaling pathways that regulate cyclin D1 in HepG2 cells. Flow cytometry and PCNA western blot data showed that resveratrol inhibits proliferation of HepG2 cells. Also, resveratrol treatment downregulated cyclin D1 as well as p38 MAP kinase, Akt and Pak1 expression and activity in HepG2 cells, suggesting that growth inhibitory activity of resveratrol is associated with the downregulation of cell proliferation and survival pathways. Furthermore, resveratrol treated cells showed increase in ERK activity suggesting possible sensitization to apoptosis. Thus in the present study, we report a three-dimensional relationship between the growth inhibitory effects of resveratrol - decrease in the levels of cyclin D1 - and downregulation of cell proliferation and survival pathways in HepG2 cells leading to cellular degenerative changes. These observations suggest that resveratrol has good potential as effective chemopreventive agent against liver cancer and warrant further studies.

Differential role of MAP kinase isoforms in malachite green transformed Syrian hamster embryo fibroblasts in culture.

Malachite green (MG) induces DNA damage and malignant transformation of Syrian hamster embryo (SHE) cells in primary culture. In the present study, we have studied the role of all the three isoforms of mitogen activated protein (MAP) kinases i.e. ERK (extracellular regulated kinase), JNK (JUN- N- terminal kinase) and p38 kinase during transformation of SHE cells by MG. The results showed that transformed cells were associated with a decreased expression of phosphoactive ERK and JNK and increased expression of p38 kinase as evident from the Western blot, immunofluorescence and flow cytometry studies. Also, a persistent nuclear localization of p38 kinase was observed in the transformed cells. The present study indicated that p38 kinase was present at higher levels and seemed to be associated with transformation, which suggested that inhibitors of p38 kinase could serve in general as potential agents for selective cancer therapy.

Decreased phosphoactive ERKs and JNKs in Malachite-green-transformed Syrian hamster embryo fibroblasts are associated with increased phosphoactive p38 kinase: possible therapeutic importance.

BACKGROUND: Malachite green (MG), consisting of green crystals with a metallic lustre, is highly soluble in water, cytotoxic to various mammalian cells and also acts as a liver tumor promoter. In view of its industrial importance and possible exposure of human beings, MG poses a potential environmental health hazard. We have earlier reported the malignant transformation of Syrian hamster embryo (SHE) cells by MG. METHODS: Cell transformation assays were carried out as described in the literature. Western blotting and flow cytometry were carried out by standard methods. RESULTS: In this study, we have studied the role of all three isoforms of mitogen-activated protein (MAP) kinases, i.e. extracellular regulated kinases (ERKs), Jun N-terminal kinases (JNKs) and p38 kinase in the MG-transformed SHE fibroblasts compared to controls. Our results showed that transformed cells were associated with decreased expression of ERKs and JNKs as evidenced by Western blotting studies. However, the p38 MAP kinase was found to be upregulated. Flow cytometric DNA histogram analysis indicated an increase in the expression of S phase cells in the transformed cell line as compared to their control counterparts. CONCLUSIONS: The present studies indicate that decreased phosphoactive ERKs and JNKs and increased phosphoactive p38 kinase are associated with increased S phase cells during transformation of SHE cells by MG.

Inhibitory effect of cinnamoyl compounds against human malignant cell line.

In the present study, anti-proliferative effects of dietary polyphenolic compounds have been observed and demonstrated the strong anticancer efficacy of curcumin (CMN), an active constituent of dietary spice (turmeric) using human leukemia cancer cell line. CMN inhibited the proliferation of K562 leukemic cells by induction of apoptosis. The current study demonstrated synergy with combination of drug therapy, and suggested that combination of ferulic acid and cisplatin synergistically inhibited cellular proliferation. Cytotoxic synergy was observed independent of the sequence of addition of two drugs to cultured cells. The synergized growth inhibitory effect with cisplatin was probably associated with G2-M arrest in cell cycle progression. These findings suggested that among the cinnamoyl compounds, CMN was most potent and FER appeared to be a better modulating agent on human malignant cell line.

DNA damage and G2/M arrest in Syrian hamster embryo cells during Malachite green exposure are associated with elevated phosphorylation of ERK1 and JNK1.

Malachite Green (MG), consisting of green crystals with a metallic luster, is highly soluble in water, cytotoxic to various mammalian cells and also acts as a liver tumor promoter. In view of its industrial importance and possible exposure to human beings, MG poses a potential environmental health hazard. We have earlier reported the malignant transformation of Syrian hamster embryo (SHE) cells in primary culture by MG. In this study, we have studied the ability of MG to cause DNA damage, cell cycle arrest, apoptosis and possible roles of ERK, JNK and p38 MAP kinases. Exposure of SHE cells to MG causes DNA damage. Flow cytometric analysis showed an increase of G2/M phase and apoptotic cells in MG treated cells compared to control SHE cells. Western blots of MG treated cells with phosphoactive antibodies showed elevated phosphorylation of ERK1 and JNK1 and no change in p38 kinase. However, total forms of ERKs, JNKs and p38 kinases showed similar levels of expression in control and MG treated SHE cells. The present study indicates that elevated phosphorylation of ERK1 and JNK1 and an increase in G2/M phase and apoptotic cells seems to be the changes associated with MG exposure to SHE cells in primary culture.

Hyperphosphorylation of extracellular regulated kinase 2 (ERK2) and inhibition of JNK2 phosphorylation are associated with increased S-phase during transformation of Syrian hamster embryo cells by Malachite Green.

Malachite Green (MG), consisting of green crystals with a metallic lustre, is highly soluble in water, cytotoxic to various mammalian cells and also acts as a liver tumour promoter. In view of its industrial importance and possible exposure to human beings, MG poses a potential environmental health hazard. We have earlier reported the malignant transformation of Syrian hamster embryo (SHE) cells in primary culture by MG. In this study, we have studied the mitogen activated protein (MAP) kinase signal transduction pathway in preneoplastic cells induced by MG. Western blots of MG induced preneoplastic cells showed no phosphorylation of ERK1, an increased phosphoactive ERK2 associated with a decreased expression of phosphoactive JNK2. However, total forms of ERKs, JNKs and p38 Kinases showed similar levels of expression in control and preneoplastic SHE cells. Indirect immunofluorescence studies have shown a distinct nuclear localisation of phosphoactive ERKs in MG induced preneoplastic cells. Flow cytometric analysis showed an increase of S-phase cells in preneoplastic cells compared to control SHE cells. The present study indicates that hyperphosphorylation of ERK2, decreased JNK2 phosphorylation and an increase in S-phase cells seems to be the early changes associated with the MG induced malignant transformation of SHE cells in primary culture.