Suppression of bcl-2 gene by RNA interference increases chemosensitivity to cisplatin in nasopharyngeal carcinoma cell line CNE1

To explore the effect of suppressing BCL-2 expression using RNA interference (RNAi) technique in nasopharyngeal carcinoma cell line CNE1. CNE1 cell lines stably expressing shRNAs targeted bcl-2 and GL3 gene were established and gene expression inhibition was assessed by Western blotting analysis. The effect of suppressing bcl-2 by RNAi on cell growth was studied, the apoptosis induction and the sensitization of CNE1 cells to cisplatin were quantified by MTT assay and flow cytometry. The results showed that: stable transfection of CNE1     

Induction of antiproliferative effect by diosgenin through activation of p53,release of apoptosis-inducing factor （AIF） and modulation of caspase-3 activity in different human cancer cells

Previously, we demonstrated that a plant steroid, diosgenin, altered cell cycle distribution and induced apoptosis in the human osteosarcoma 1547 cell line. The objective of this study was to investigate if the antiproliferative effect of diosgenin was similar for different human cancer cell lines such as laryngocarcinoma HEp-2 and melanoma M4Beu cells. Moreover, this work essentially focused on the mitochondrial pathway. We found that diosgenin had an important and similar antiproliferative effect on different types of cancer cells. In addition, our new results show that diosgenin-induced apoptosis is caspase-3 dependent with a fall of mitochondrial membrane potential, nuclear localization of AIF and poly (ADP-ribose) polymerase cleavage. Diosgenin treatment also induces p53 activation and cell cycle arrest in the different cell lines studied.     

Double role of Fas ligand in the apoptosis of intervertebral disc cells in vitro

Fas ligand （FasL） may play an important role in maintaining the immune privilege of intervertebral disc （IVD）. Besides, it is closely related to the apoptosis of degenerative disc cells. Nowadays, lots of reports have described about the paradoxical effects of FasL, although the effect of FasL on IVD cells is still under debate. In this study, we tried to investigate the effects of FasL on Fas expression and on the apoptosis of nucleus pulposus （NP） cells in Sprague-Dawley rats. The results showed that the expression of Fas in NP cells was significantly increased by the recombinant FasL. Meanwhile, the apoptosis of NP cells increased markedly in a FasL dose-dependent manner. Interestingly, RNA interference results indicated that the increase of Fas expression and the NP cell apoptosis described previously were inhibited by Fas siRNA, suggesting that RNA interference might be one of novel strategies to prevent IVD cells from apoptosis.     

Mechanism of taurine-induced apoptosis in human colon cancer cells

Taurine （Tau） has been shown to possess cancer therapeutic effect through induction of apoptosis, while the underlying molecular mechanism of its anti-cancer effect is not well understood. PUMA （p53-upregulated modulator of apoptosis） plays an important role in the process of apoptosis induction in a variety of human tumor ceils in both p53- dependent and -independent manners. However, whether PUMA is involved in the process of Tau-induced apoptosis in cancer cells has not been well studied. In the present study, we treated human colorectal cancer cells HT-29 （mutant p53） and LoVo （wild-type p53） with different concentrations of Tau, which led to the repression of cell proliferation and induction of apoptosis in both cell fines. Meanwhile, we also observed the increased expression of PUMA and high Bax/Bcl-2 ratios. To determine the role of PUMA in Tau-induced apoptosis, we used small interfering RNA interference to suppress PUMA expression. As a result, apoptosis was decreased in response to Tau treatment. All these results indicated that PUMA plays a critical role in Tauinduced apoptosis pathway in human colorectal cancer ceils. Demonstration of the molecular mechanism involved in the anti-tumor effect of Tau may be useful in the therapeutic target selection for p53-deficient colorectal cancer.     

Effects of inhibition of ubiquitin-proteasome pathway on human primary leukemic cells

Though there were a lot of reports about the totally different responses to the inhibition of ubiquitin-proteasome pathway in different kinds of cell lines, much less has been known about the responses in primary human leukemic cells. In this study, the effects of inhibition of ubiquitin-proteasome pathway on human bone marrow (BM) mononuclear cells (MNCs) obtained from 10 normal persons and 8 leukemia patients were examined. The results showed that the responses obviously varied individually. Among them, BM MNCs in 3 cases of leukemic patients were extremely sensitive, demonstrated by that >90% cells were induced to undergo apoptosis within 24 h, but MNCs in 10 cases of normal persons showed resistance to the inhibition and no apoptosis was observed. Furthermore, Western blots revealed that the Bcl-2 expression was relatively high in the sensitive primary leukemia cells, and especially the cleavage of 26 ku Bcl-2 into a 22 ku fragment occurred during the induction of apoptosis. In contrast, the Bcl-2 e     

Interference effect of epigallocatechin-3-gallate on targets of nuclear factor kappaB signal transduction pathways activated by EB virus encoded latent membrane protein 1

AIM: To elucidate the interference effect of epigallocatechin-3-gallate (EGCG) on targets of nuclear factor kappaB (NF-kappaB) signal transduction pathway activated by EB virus encoded latent membrane protein 1 (LMP1) in nasopharyngeal carcinoma (NPC) cell lines. METHODS: The survival rates of CNE1 and CNE-LMP1 cell lines after the EGCG treatment were determined by MTT assay. NF-kappaB activation in CNE1 and CNE-LMP1 cells after EGCG treatment was analyzed by promoter luciferase reporter system. And then nuclear translocation of NF-kappaB (p65) after the EGCG treatment was analyzed by immunofluorescence and western blotting. Meanwhile, the changes of IkappaBalpha phosphorylation were observed after the EGCG treatment. EGFR promoter activity was analyzed by promoter luciferase reporter system and EGFR phosphorylation was observed by western blotting after the EGCG treatment. RESULTS: EGCG inhibited the survival rates of CNE1 and CNE-LMP1 cells and NF-kappaB activation caused by LMP1 in CNE-LMP1 cells. EGCG also suppressed the nuclear translocation of NF-kappaB (p65) and IkappaBalpha phosphorylation. Meanwhile, EGCG inhibited EGFR promoter activity and EGFR phosphorylation. CONCLUSIONS: EGCG inhibited not only the dose-dependent survival rate of NPC cells, but also the dose-dependent activation of NF-kappaB. This inhibition of LMP1-caused NF-kappaB activation was mediated via the phosphorylative degradation of its inhibitory protein IkappaBalpha, and then EGCG inhibited EGFR activity which was a downstream gene from NF-kappaB. This study suggests that interference effect of EGCG on targets of signal transduction pathway plays an important role in the anticancer function.     

Carboplatin induces apoptotic cell death through downregulation of constitutively active nuclear factor-kappaB in human HPV-18 E6-positive HEp-2 cells

Because the role of nuclear factor kappaB (NF-kappaB) is in cellular growth control and neoplasia, we explored the status of NF-kappaB and investigated its role in survival of human HPV-18 E6-positive HEp-2 cells. We observed accumulation of p65 in the nucleus. Moreover, without any external stimulus constitutive NF-kappaB DNA binding and transactivation activity were detected in HEp-2 cells. Treatment with NF-kappaB inhibitor curcumin (diferuloylmethane) and transient transfection of the mutant form of IkappaBalpha, IkappaBalpha super repressor (IkappaBalpha-SR), suppressed constitutive NF-kappaB activity as well as proliferation, suggesting that constitutive NF-kappaB activity is required for the survival of HEp-2 cells. Carboplatin treatment downregulated constitutive NF-kappaB activity and prevented nuclear retention of p65. Further, carboplatin also suppressed the constitutive IkappaBalpha phosphorylation leading to stabilization of IkappaBalpha protein in the cells. Carboplatin inhibited NF-kappaB binding to its response element present in Bcl-2 promoter resulting in downregulation of antiapoptotic Bcl-2 protein. Thus, our results for the first time indicate that constitutive NF-kappaB has a significant role in the survival of HPV-18 E6-positive HEp-2 cells. Moreover, inactivation of NF-kappaB is one of the mechanisms underlying the induction of carboplatin-mediated apoptosis in HPV-18 E6-positive cancer cells.     

Cell-Specific Association and Shuttling of IκBα Provides a Mechanism for Nuclear NF-κB in B Lymphocytes

Mature B lymphocytes are unique in containing nuclear Rel proteins prior to cell stimulation. This activity consists largely of p50-c-Rel heterodimers, and its importance for B-cell function is exemplified by reduced B-cell viability in several genetically altered mouse strains. Here we suggest a mechanism for the cell specificity and the subunit composition of constitutive B-cell NF-kappaB based on the observed properties of Rel homo- and heterodimers and IkappaBalpha. We show that c-Rel lacks a nuclear export sequence, making the removal of c-Rel-containing complexes from the nucleus less efficient than removal of p65-containing complexes. Second, the nuclear import potential of p65 and c-Rel homodimers but not p50-associated heterodimers was attenuated when they were complexed to IkappaBalpha, leading to a greater propensity of heterodimers to be nuclear. We propose that subunit composition of B-cell NF-kappaB reflects the inefficient retrieval of p50-c-Rel heterodimers from the nucleus. Cell specificity may be a consequence of c-Rel-IkappaBalpha complexes being present only in mature B cells, which leads to nuclear c-Rel due to IkappaBalpha turnover and shuttling of the complex.