Ectopic expression of clusterin／apolipoprotein J or Bcl-2 decreases thesensitivity of HaCaT cells to toxic effects of ropivacaine

Local anesthetics inhibit cell proliferation and induce apoptosis in various cell types. Ropivacaine, a unique, novel tertiary amine-type anesthetic, was shown to inhibit the proliferation of several cell types including keratinocytes. We found that Ropivacaine could inhibit the proliferation and induce apoptosis in an immortalized human keratinocyte line,HaCaT, in a dose- and time-dependent manner and with the deprivation of serum. The dose-dependent induction of apoptosis by ropivacaine was demonstrated by DNA fragmentation analysis and the proteolytic cleavage of a caspase-3 substrate—poly (ADP-ribose) polymerase (PARP). In addition, ropivacaine downregulated the expression of clusterin/ apoliporotein J, a protein with anti-apoptotic properties, in a dose-dependent manner, which well correlated with the induction of apoptosis of HaCaT cells. To investigate the role of clusterin/apoliporotein J in ropivacaine-induced apoptosis,HaCaT cells overexpressing clusterin/apoliporotein J were generated and compared to cells expressing the well established anti-apoptotic Bcl-2 protein. Ectopic overexpression of the secreted form of clusterin/apoliporotein J or Bcl-2decreased the sensitivity of HaCaT cells to toxic effects of ropivacaine as demonstrated by DNA fragmentation, the proteolytic cleavage of PARP and by a reduction in procaspase-3 expression. Furthermore, the downregulation of endogenous clusterin/apolipoprotein J levels by ropivacaine suggested that this might be one mechanism by which ropivacaine induced cell death in HaCaT cells. In conclusion, the ability of ropivacaine to induce antiproliferative responses and to suppress the expression of the anti-apoptotic protein clusterin/apolipoprotein J, combined with previously reported anti-inflammatory activity and analgesic property of the drug, suggests that ropivacaine may have potential utility in the local treatment of tumors.     

Overexpression of Bax sensitizes prostate cancer cells to TGF-β induced apoptosis

NRP-154 is a tumorigenic epithelial cell line derived from the preneoplastic dorsal-lateral prostate of rats. These cells are exquisitely sensitive to TGF-β induced apoptosis. In contrast, we find that NRP-154 cells can sustain overexpression of exogenous Bax protein, which is different from non-tumor cells where Bax functions as a ubiquitous stimulator of apoptosis. NRP-154 cells stably overexpressing Bax show increased sensitivity to TGF-β induced apoptosis. The degree of TGF-β induced apoptosis displays high correlation with cleavage of Bax at the amino-terminus. Our data indicate that prostate cancer cells can host high levels of latent Bax which can be activated through post-translational modification.     

Inducible resistance to Fas—mediated apoptosis in B cells

Apoptosis produced in B cells through Fas(APO-1,CD95) triggering is regulated by signals derived from other surface receptors:CD40 engagement produces upregulation of Fas expression and marked susceptibility to Fas-induced cell death,whereas antigen receptor engagement,or IL-4R engagement,inhibits Fas killing and in so doing induces a state of Fas-resistance,even in otherwise sensitive,CD40-stimulated targets.Surface immunoglobulin and IL-4R utilize at least partially distinct path ways to produce Fas-resistance that differentially depend on PKC and STAT6,respectively.Further,surface immunoglobulin signaling for inducible Fas-resistance bypasses Btk,requires NF-κB,and entails new macromolecular synthesis.Terminal effectors of B cell Fas-resistance include the known anti-apoptotic gene products,Bcl-XL and FLIP,and a novel anti-apoptotic gene that encodes FAIM (Fas Apoptosis Inhibitory Molecule).faim was identified by differential display and exists in two alternatively spliced forms;faim-S is broadly expressed,but faim-L expression is tissue-specific.The FAIM sequence is highly evolu tionarily conserved,suggesting an important role for this molecule throughout phylogeny.Inducible resistance to Fas killing is hypothesized to protect foreign antigen-specific B cells during potentially hazardous interactions with FasL-bearing T cells,whereas autoreactive B cells fail to become Fas-resistant and are deleted via Fas-dependent cytotoxicity.Inadvertent or aberrant acquisition of Fas-resistance may permit autoreactive B cells to escape Fas deletion,and malignant lymphocytes to impede anti-tumor immunity.     

Altered expression of nuclear matrix proteins in etoposide induced apoptosis in HL－60 cells

INTRODUCTIIONThe nuclear matrix is an essential component ofthe nucleus which is important for the nuclear structural integrity and specific genomic functions[1, 2].Several articles have reported that the nuclear matrix, as a higher order framework structures, mightbe disassembled du-ring the apoptotic process[3-5].Accordingly3 nuclear lamins A/C or B have beenfound to decrease in apoptotic thymocytes[6], Tcells[7], and carcinoma cell line[8, 9]. The nucleolar protein B23, an obscure ma…     

Thapsigargin increases apoptotic cell death in human hepatoma BEL—7404 cells

Effects of thapsigargin,an inhibitor of Ca^2 -ATPase in surface of endoplasmic reticulum,on apoptotic cell death were studied in human hepatoma cells of BEL-7404 cell line by using both flow cytometry and electron microscopy.Propidium iodide staining and flow cytometry revealed that in the serum-free condition,thapsigargin increased the rate of apoptosis of BEL-7404 cells in a dose-dependent manner.Prolongation of the period of serum-free condition enhanced the apoptosis induced by thapsigargin treatment.Morphological observation with electron microscope further demonstrated that chromatin condensation and fragmentation,apoptotic bodies existed in TG-treated cells,supporting that thapsigargin is a potent activator of apoptosis in the cells.     

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.     

Bcl-2 down modulation in WEHI-3B/CTRES cells resistant to Cholera Toxin （CT）-induced apoptosis

The very different effects of Cholera Toxin （CT） on cell growth and proliferation may depend on the type of ganglioside receptors in cell membranes and different signal transduction mechanisms triggered, but other functions related to the drug resistance mechanisms can not be excluded. The effect of CT treatment on the ＂in vitro＂ clonogenicity, the Population Doubling Time （PDT）, apoptosis, PKA activation and Bax and Bcl-2 expression was evaluated in WEHI-3B cell line and its CT-resistant subclone （WEHI-3B/CTRES）. In WEHI-3B parental cells the dramatic accumulation of cAMP induced by CT correlated well with PKA activation, increased PDT value, inhibition of clonogenicity and apoptosis. H-89 treatment inhibited PKA activation by CT but did not protect the cells from apoptosis and growth inhibition. In WEHI-3B/CTRES no significant CT-dependent accumulation of cAMP occurred with any increase of PKA activity and PDT. In CT resistant cells （WEHI-3B/CTRES）, Bcl-2 expression was down regulated by both CT or drug treatment （eg, ciprofloxacin, CPX） although these cells were protected from CT-dependent apoptosis but not from drug-induced apoptosis. Differently from other cell models described, down regulation of Bcl-2 is proved to be independent on cAMP accumulation and PKA activation. Our observations support the implication of cAMP dependent kinase （PKA） in the inhibition of WEHI-3B cells growth and suggest that, in WEHI-3B/CTRES, Bcl-2 expression could be modulated by CT in the absence of cAMP accumulation. Also in consideration of many contradictory data reported in literature, our cell models （of one sensitive parental cell strain and two clones with different uncrossed specific resistance to CT and CPX） provides a new and interesting tool for better investigating the relationship between the CT signal transduction mechanisms and Bcl-2 expression and function.     

细胞凋亡基因的发现

细胞凋亡基因的发现李雨民,孙元明（中国医学科学院放射医学研究所天津３００１９２）细胞凋亡是一种具有特征性的形态变化和生化改变的细胞死亡过程,机体启动这一机制以清除无用和有害的细胞。近来细胞凋亡基因的研究取得了很大进展,８０年代以来Ｈｏｒｖｉｔｚ等在研...     

Apoptosis of human leukemic HL—60 cells induced to differentiate by treatment with RA or DMSO

In present study,we studied the effect of all-trans retinoic acid(ATRA)and dimethylsulfoxide(DMSO)on the induction of apoptosis in HL-60 cell line.Based on morphological changes by Hochest 33342 staining and identification of internuclesomal NDA celeavage by gel electrophoresis,we observed aberrant nuclear chromatin condensation and ladder-like pattern of DNA degradation. Using Flow Cytometric method.We found sub-G1 peak in RA-treated HL-60 cells starting 5 to 6d after the initiation of the treatment However,Such an obvious apoptotic peak was not identified in DMSO-differentiated cells.Combining the research accomplished before.our study approves further that apoptosis could be a common mode of death of terminally differentiated HL-60 cells.     

Expression of Bcl—2 inhibited Fas—mediated apoptosis in human hepatocellular carcinoma BEL—7404 cells

Apoptosis plays an important role in embryonic development,tissue remodeling,immune regulation and tumor regression.Two groups of molecules(Bcl-2 family and “Death factor” family) are involved in regulating apoptosis.In order to know about the effect of Bcl-2 on apoptosis induced by Fas,a typical member of “Death factor” family,the transfection experiments with expression vectors pcDNA3-fl and pcDNA3-bcl-2 were performed in BEL-7404 cells,a human hepatocellular carcinoma cell line which expresses endogenous Fas,but not FasL and Bcl-2.The data showed that the expression of FasL in pcDNA3-fl transfected hepatoma cells obviously induced the apoptosis of the cells.However,the overexpression of Bcl-2 in pcDNA3-bcl-2 transfected 7404/b-16 cells counteracted pcDNA3-fl transient transfection mediated apoptosis.Further study by cotransfection experiments indicated that Bid but not Bax (both were pro-apoptotic proteins of Bcl-2 family) blocked the inhibitory effect of Bcl-2 on Fas-mediated apoptosis.These results suggested that Fas-mediated apoptosis in human hepatoma cells is possibly regulated by Bcl-2 family proteins via mitochondria pathway.     

Blockage of IGF－1R signaling sensitizes urinary bladder cancer cells to mitomycin－mediated cytotoxicity

INTRODUCTIONtransitional cell carcinoma (TCC) of the bladder represents the fifth most preValent malignancy inwestern population. A major problem in the management of TCC is the low sensitivity to chemotherapy and the high recu-rrence after transurethral resection, which occupies a large proportion (approximately 40%) among bladder cancer patients[1, 21. Sodrug resistance remains a major and difficult problem to resolye in TCC chemotherapy. This phenomenon has often been ascribed to so…