Novel augmentation by bufalin of protein kinase C-induced cyclooxygenase-2 and IL-8 production in human breast cancer cells.

Cyclooxygenase-2 (COX-2) and IL-8 are two inflammatory mediators induced by protein kinase C (PKC) via various stimuli. Both contribute significantly to cancer progression. Bufalin, a major active component of the traditional Chinese medicine Chan Su, is known to induce apoptosis in various cancer cells. This study clarifies the role and mechanism of bufalin action during PKC regulation of COX-2/IL-8 expression and investigates the associated impact on breast cancer. Using MB-231 breast cancer cells, bufalin augments PKC induction of COX-2/IL-8 at both the protein and mRNA levels, and the production of prostaglandin E2 (PGE2) and IL-8. The MAPK and NF-κB pathways are involved in both the PKC-mediated and bufalin-promoted PKC regulation of COX-2/IL-8 production. Bufalin increases PKC-induced MAPKs phosphorylation and NF-κB nuclear translocation. PGE2 stimulates the proliferation/migration of breast cancer cells. Furthermore, PKC-induced matrix metalloproteinase 3 expression is enhanced by bufalin. Bufalin significantly enhances breast cancer xenograft growth, which is accompanied by an elevation in COX-2/IL-8 expression. In conclusion, bufalin seems to promote the inflammatory response in vitro and in vivo, and this occurs, at least in part, by targeting the MAPK and NF-κB pathways, which then enhances the growth of breast cancer cells.

Attenuation of LPS-induced cyclooxygenase-2 and inducible NO synthase expression by lysophosphatidic acid in macrophages.

LPS can activate the inflammatory cascades by inducing various inflammatory mediators, such as prostaglandin E(2) (PGE(2)) resulting from cyclooxygenase-2 (COX-2), and NO produced by inducible NO synthase (iNOS). Lysophosphatidic acid (LPA) has been demonstrated to participate in inflammation. This study aimed to clarify the impact and the involving mechanisms of LPA on LPS-incurred inflammation in macrophages. First, LPA appeared to attenuate LPS-induced protein and mRNA expression of COX-2 and iNOS genes, as well as production of PGE(2) and NO. By using selective inhibitors targeting various signaling players, the inhibitory G protein alpha subunit (Gα(i)) seemed to be involved in the effect of LPA; p38, ERK and NF-κB were involved in the LPS-mediated COX-2/PGE(2) pathway; and p38, JNK, phosphoinositide-3-kinase and NF-κB were involved in the LPS-mediated iNOS/NO pathway. LPA was able to diminish LPS-induced phosphorylation of p38 and Akt, as well as NF-κB p65 nuclear translocation. By utilization of inhibitors of COX-2 and iNOS, there appeared to be no modulation between the COX-2/PGE(2) and the iNOS/NO signaling pathways. Our findings demonstrate a clear anti-inflammatory role of LPA acting via Gα(i) in LPS-mediated inflammatory response in macrophages, owing, at least in part, to its suppressive effect on LPS-induced activation of p38, Akt and NF-κB.

Inhibition of PKC-Induced COX-2 and IL-8 Expression in Human Breast Cancer Cells by Glucosamine.

Breast cancer is a common cancer leading to many deaths among females. Cyclooxygenase-2 (COX-2) and interleukin-8 (IL-8) are two highly expressed inflammatory mediators to be induced by the protein kinase C (PKC) signaling via various inflammatory stimuli and both contribute significantly to cancer metastasis/progression. Glucosamine has been shown to act as an anti-inflammation molecule. The aim of this study was to clarify the role and acting mechanism of glucosamine during the PKC-regulation of COX-2/IL-8 expression and the associated impact on breast cancer. In MCF-7 breast cancer cells, glucosamine effectively suppresses the PKC induction of COX-2 and IL-8 promoter activity, mRNA and protein levels, as well as the production of prostaglandin E(2) (PGE(2)) and IL-8. Glucosamine is able to promote COX-2 protein degradation in a calpain-dependent manner and IL-8 protein degradation in calpain-dependent and proteasome-dependent manners. The MAPK and NF-κB pathways are involved in PKC-induced COX-2 expression, but only the NF-κB pathway is involved in PKC-induced IL-8 expression. Glucosamine attenuates PKC-mediated IκBα phosphorylation, nuclear NF-κB translocation, and NF-κB reporter activation. Both PGE(2) and IL-8 promote cell proliferation and IL-8 induces cell migration; thus, glucosamine appears to suppress PKC-induced cell proliferation and migration. Furthermore, glucosamine significantly inhibits the growth of breast cancer xenografts and this is accompanied by a reduction in COX-2 and IL-8 expression. In conclusion, glucosamine seems to attenuate the inflammatory response in vitro and in vivo and this occurs, at least in part by targeting to the NF-κB signaling pathway, resulting in an inhibition of breast cancer cell growth.

Depletion of SUMO ligase hMMS21 impairs G1 to S transition in MCF-7 breast cancer cells.

BACKGROUND: hMMS21 is a human SUMO ligase required for DNA damage repair and mitotic progression in HeLa cervical cancer cells. Owing to the diversity of cancer, we further investigated the effect of hMMS21-depletion on MCF-7 breast cancer cells. METHODS: hMMS21-depletion was achieved by RNA interference. Cellular hMMS21 and E2F1 mRNA levels were estimated by RT-PCR and real-time PCR. Cell cycle profile was assessed by flow cytometry. Western blot and co-immunoprecipitation were used to determine the protein levels of various factors involved in G1-S transition and CDK2- or CDK4-associated p21 and p27. Kinase activity of cyclin E/CDK2 was measured in anti-cyclin E immunoprecipitate. RESULTS: hMMS21-depletion induced slower cell growth and G1-S transition. While it had no effect on cyclin D1 or phospho-Rb (S807/811) levels, hMMS21-depletion provoked lower E2F1 levels and cyclin E/CDK2 activity. The decreased cyclin E/CDK2 activity correlated with increased cellular p21(CIP1) levels and CDK2-p21 association. Moreover, ectopic expression of Flag-hMMS21 but not its ligase-inactive mutant rescued the decreased growth rates of hMMS21-depletd cells. Thus, depletion of hMMS21 seems to impair G1-S transition due to lowered E2F1 protein levels and cyclin E/CDK2 activity. The decreased cyclin E/CDK2 activity is probably attributable to its greater association with p21 as a result of increased p21 levels. In addition, hMMS21-mediated sumoylation appears to be involved. GENERAL SIGNIFICANCE: This study demonstrates that hMMS21 is required for G1-S transition in breast cancer cells and implies that manipulation of hMMS21-mediated sumoylation may alter the growth rates of breast cancer cells.