OBJECTIVE:To investigate the synergistic effects of polyphyllin Ⅰ(PPⅠ)combined with tumor necrosis factorrelated apoptosis-inducing ligand(TRAIL)on the growth of osteosarcoma cells through downregulating the Wnt/β-...OBJECTIVE:To investigate the synergistic effects of polyphyllin Ⅰ(PPⅠ)combined with tumor necrosis factorrelated apoptosis-inducing ligand(TRAIL)on the growth of osteosarcoma cells through downregulating the Wnt/β-catenin signaling pathway.METHODS:Cell viability,apoptosis and cell cycle distribution were examined using cell counting kit-8 and flow cytometry assays.The morphology of cancer cells was observed with inverted phase contrast microscope.The migration and invasion abilities were examined by xCELLigence real time cell analysis DP system and transwell assays.The expressions of poly(adenosine diphosphate-ribose)polymerase,C-Myc,Cyclin B1,cyclin-dependent kinases 1,N-cadherin,Vimentin,Active-β-catenin,β-catenin,p-glycogen synthase kinase 3β(GSK-3β)and GSK-3βwere determined by Western blotting assay.RESULTS:PPⅠ sensitized TRAIL-induced decrease of viability,migration and invasion,as well as increase of apoptosis and cell cycle arrest of MG-63 and U-2 OS osteosarcoma cells.The synergistic effect of PPⅠwith TRAIL in inhibiting the growth of osteosarcoma cells was at least partially realized through the inactivation of Wnt/β-catenin signaling pathway.CONCLUSION:The combination of PPⅠ and TRAIL is potentially a novel treatment strategy of osteosarcoma.展开更多
Auditory neuropathy spectrum disorder(ANSD)represents a variety of sensorineural deafness conditions characterized by abnormal inner hair cells and/or auditory nerve function,but with the preservation of outer hair ce...Auditory neuropathy spectrum disorder(ANSD)represents a variety of sensorineural deafness conditions characterized by abnormal inner hair cells and/or auditory nerve function,but with the preservation of outer hair cell function.ANSD represents up to 15%of individuals with hearing impairments.Through mutation screening,bioinformatic analysis and expression studies,we have previously identified several apoptosis-inducing factor(AIF)mitochondria-associated 1(AIFM1)variants in ANSD families and in some other sporadic cases.Here,to elucidate the pathogenic mechanisms underlying each AIFM1 variant,we generated AIF-null cells using the clustered regularly interspersed short palindromic repeats(CRISPR)/CRISPR-associated protein 9(Cas9)system and constructed AIF-wild type(WT)and AIF-mutant(mut)(p.T260A,p.R422W,and p.R451Q)stable transfection cell lines.We then analyzed AIF structure,coenzyme-binding affinity,apoptosis,and other aspects.Results revealed that these variants resulted in impaired dimerization,compromising AIF function.The reduction reaction of AIF variants had proceeded slower than that of AIF-WT.The average levels of AIF dimerization in AIF variant cells were only 34.5%-49.7%of that of AIF-WT cells,resulting in caspase-independent apoptosis.The average percentage of apoptotic cells in the variants was 12.3%-17.9%,which was significantly higher than that(6.9%-7.4%)in controls.However,nicotinamide adenine dinucleotide(NADH)treatment promoted the reduction of apoptosis by rescuing AIF dimerization in AIF variant cells.Our findings show that the impairment of AIF dimerization by AIFM1 variants causes apoptosis contributing to ANSD,and introduce NADH as a potential drug for ANSD treatment.Our results help elucidate the mechanisms of ANSD and may lead to the provision of novel therapies.展开更多
Most of the ocular tumors have poor prognosis, and they remain a difficult problem in the area of ophthalmology. With the rapid development of molecular biology and immunologic techniques and the deep research on ocul...Most of the ocular tumors have poor prognosis, and they remain a difficult problem in the area of ophthalmology. With the rapid development of molecular biology and immunologic techniques and the deep research on ocular tumor related genes, it becomes possible to diagnose and treat malignant tumors from the molecular level. The tumor necrosis factor related apoptosis-inducing ligand (TRAIL), a member of the tumor necrosis factor (TNF) super family, is a promising candidate, either alone or in combination with established cancer therapies, since it can initiate apoptosis through the activation of their death receptors. The ability of TRAIL to selectively induce apoptosis of transformed, virus-infected or tumor cells but not normal cells promotes the development of TRAIL-based cancer therapy. Here, we will review TRAIL and its receptors' structure, function, mechanism of action and application in ocular tumors therapy.展开更多
Studies have shown that tumor necrosis factor-related apoptosis-inducing ligand（TRAIL）exhibits strong induction of apoptosis in human glioma cells.It remains unclear whether the mitochondrion pathway,an important ap...Studies have shown that tumor necrosis factor-related apoptosis-inducing ligand（TRAIL）exhibits strong induction of apoptosis in human glioma cells.It remains unclear whether the mitochondrion pathway,an important apoptosis signaling pathway,is involved in TRAIL-induced glioma cell apoptosis.In the present study,in vitro cultured human glioma U87 cells were treated with human recombinant soluble TRAIL.Apoptosis of glioma U87 cells,mitochondrial transmembrane potential（Δψm）,cytoplasmic cytochrome c concentration and changes in caspase-3,-8 and-9 activity following human recombinant soluble TRAIL treatment were investigated to determine the mechanism of glioma U87 cell apoptosis induced by TRAIL.Additionally,blocking caspase-8resulted in TRAIL-induced mitochondrion pathway activation,suggesting that TRAIL,through activating caspase-8,initiated a series of mitochondrial events and resulted in apoptosis of glioma U87 cells.展开更多
hap, a novel human apoptosis-inducing gene which can interact with another newly discovered apoptosis-inducing geneASY, was identified, by cloning its cDNAs from human lung cell line (WI-38) cDNA library. Two major mR...hap, a novel human apoptosis-inducing gene which can interact with another newly discovered apoptosis-inducing geneASY, was identified, by cloning its cDNAs from human lung cell line (WI-38) cDNA library. Two major mRNA species (1.8 and 2.7 kb in length, respectively) were previously identified by Northern blot analysis of poly(A)+ RNA from human multiple tissues using partialhap cDNA as a probe. In the present work, the molecular mechanism accounting for the generation of the twohap transcripts were investigated. The rapid amplification of cDNA 3′-ends (3′-RACE) technique and the sequential Southern blot analysis, in conjunction with the sequencing analysis demonstrated that the twohap transcripts derive from the alternative polyadenylation site selection: a AATAAA signal at position 1 528–1 533 nt for the 1.8 kbhap mRNA: and a AATAAA signal at position 2 375–2 380 nt for the 2.7 kbhap mRNA. Furthermore, a number of regulatory elements withinhap 3′-untranslated region (3′-UTR) were also examined.展开更多
Objective:Structural abnormalities and dysfunction of the placenta contribute to pregnancy-related complications,such as preeclampsia.Syncytin-A(synA)has been reported to be expressed in the placenta.The contribution ...Objective:Structural abnormalities and dysfunction of the placenta contribute to pregnancy-related complications,such as preeclampsia.Syncytin-A(synA)has been reported to be expressed in the placenta.The contribution of synA to developmental abnormalities and dysfunction of the placenta remains elusive.In this study,we aimed to explore the role of synA in placental development and functions.Methods:SynA-knockout mice were generated using the CRISPR-Cas9 method,and the phenotypes of the placenta and fetus ofsynA-knockout mice were observed.Real-time quantitative polymerase chain reaction(PCR)and routine PCR were employed to detect the genotypes of the offspring.CD31 immunohistochemistry was used to evaluate the vessel density of the placenta,and the protein levels of key molecules were measured by western blotting.Results:SynA knockout caused fetal death.Furthermore,synA-knockout mice showed placental developmental abnormalities,indicated by a thinner labyrinth layer,thicker spongiotrophoblast layer,lower blood vessel density,and significantly higher numbers of apoptotic trophoblasts,when compared with wild-type littermates.Mechanistically,synA ablation induced apoptosis-inducing factor(AIF)cleavage and nuclear localization and promoted placental trophoblast apoptosis.In addition,synA knockout increased the calpain1 protein levels.The calpain1 inhibitor calpeptin blockedsynA knockout-induced AIF cleavage,partially restoring the placental structural abnormalities ofsynA-knockout mice.Conclusions:SynA knockout leads to placental developmental abnormalities by inducing trophoblastic apoptosis via the calpain1-AIF pathway.展开更多
Tumor necrosis factor-related apoptosis-inducing ligand （TRAIL） is a promising agent for anticancer therapy. The identification of small molecules that can establish the sensitivity of prostate cancer （PCa） cells ...Tumor necrosis factor-related apoptosis-inducing ligand （TRAIL） is a promising agent for anticancer therapy. The identification of small molecules that can establish the sensitivity of prostate cancer （PCa） cells to TRAIL-induced apoptosis is crucial for the targeted treatment of PCa. PC3, DU145, JAC-1, TsuPrl, and LNCaP cells were treated with Andrographolide （Andro） and TRAIL, and the apoptosis was measured using the Annexin V/PI double staining method. Real time-polymerase chain reaction （PCR） and Western blot analysis were performed to measure the expression levels of target molecules. RNA interference technique was used to down-regulate the expression of the target protein. We established a nude mouse xenograft model of PCa, which was used to measure the caspase-3 activity in the tumor cells using flow cytometry. In this research study, our results demonstrated that Andro preferentially increased the sensitivity of PCa cells to TRAIL-induced apoptosis at subtoxic concentrations, and the regulation mechanism was related to the up-regulation of DR4. In addition, it also increased the p53 expression and led to the generation of reactive oxygen species （ROS） in the cells. Further research revealed that the DR4 inhibition, p53 expression, and ROS generation can significantly reduce the apoptosis induced by the combination of TRAIL and Andro in PCa cells. In conclusion, Andro increases the sensitivity of PCa cells to TRAIL-induced apoptosis through the generation of ROS and up-regulation of p53 and then promotes PCa cell apoptosis associated with the activation of DR4.展开更多
Portal hypertension （PHT） gastropathy is a frequent complication of fiver cirrhosis and one of the leading causes of death from cirrhosis. Apoptosis is widely considered to be an active energy-dependent mode of cell...Portal hypertension （PHT） gastropathy is a frequent complication of fiver cirrhosis and one of the leading causes of death from cirrhosis. Apoptosis is widely considered to be an active energy-dependent mode of cell death and a distinct entity from necrotic cell death. It is unclear whether gastric mucosal apoptosis is involved in PHT gastropa- thy. Prostaglandins （PGs） produced through cyclooxygenase （COX） are thought to play a key role in protection of the gastrointestinal mucosa from injury and apoptosis. However, the role of COX in PHT gastropathy is still not clearly understood. The aims of this study were to investigate whether （1） gastric mucosal apoptosis is involved in PHT gas- tropathy and （2） downregulation of COX contributes to this apoptosis. In this study, we show that gastric mucosal apoptosis was remarkably increased while mucosal proliferation was inhibited in PHT rats. Gastric mucosal COX- 1 was significantly suppressed at both the mRNA and protein levels, and PGE2 was reduced in PHT rats. Further, PGE2 treatment suppressed gastric mucosal apoptosis in PHT rats. However, gastric mucosal COX-2 levels did not differ between sham-operated rats and PHT rats. Gastric mucosal levels of tumor necrosis factor-α （TNF-α） and Fas ligand, but not TNF-related apoptosis-inducing ligand, were increased, and activated caspase-8 and caspase-3 levels were upregulated in PHT rats. The release of cytochrome c from the mitochondria to the cytosol was not observed in PHT rats. Our data indicate that downregulation of COX-1 is involved in gastric mucosal apoptosis via death signal- ing-mediated type-I cell death in PHT rats.展开更多
To examine the influence on apoptotic mechanisms following inhibition of polo-like kinases as therapeutically approach for cholangiocellular cancer treatment.METHODSAs most cholangiocarcinomas are chemotherapy-resista...To examine the influence on apoptotic mechanisms following inhibition of polo-like kinases as therapeutically approach for cholangiocellular cancer treatment.METHODSAs most cholangiocarcinomas are chemotherapy-resistant due to mechanisms preventing tumor cell death, we investigated the effect of Cisplatin on cholangiocellular carcinoma (CCA) cell lines KMCH-1 and Mz-Ch-1. Polo-like kinases (PLK) are important regulators of the cell cycle and their inhibition is discussed as a potential therapy while PLK inhibition can regulate apoptotic mediators. Here, cells were treated with PLK inhibitor BI6727 (Volasertib), Cisplatin, and in combination of both compounds. Cell viability was assessed by MTT; apoptosis was measured by DAPI staining and caspase-3/-7 assay. Western blot and qRT-PCR were used to measure expression levels of apoptosis-related molecules Bax and Bcl-2.RESULTSThe cell viability in the CCA cell lines KMCH-1 and Mz-Ch-1 was reduced in all treatment conditions compared to vehicle-treated cells. Co-treatment with BI6727 and cisplatin could even enhance the cytotoxic effect of cisplatin single treatment. Thus, co-treatment of cisplatin with BI6727 could slightly enhance the cytotoxic effect of the cisplatin in both cell lines whereas there was evidence of increased apoptosis induction solely in Mz-Ch-1 as compared to KMCH-1. Moreover, PLK inhibition decreases protein levels of Bcl-2; an effect that can be reversed by the proteasomal degradation inhibitor MG-132. In contrast, protein levels of Bax were not found to be altered by PLK inhibition. These findings indicate that cytotoxic effects of Cisplatin in Mz-Ch-1 cells can be enhanced by cotreatment with BI6727.CONCLUSIONIn conclusion, BI6727 treatment can sensitize CCA cells to cisplatin-induced apoptosis with proteasomal Bcl-2 degradation as an additional pro-apoptotic effect.展开更多
The rapidly developing resistance of cancers to chemotherapy agents and the severe cytotoxicity of such agents to normal cells are major stumbling blocks in current cancer treatments.Most current chemotherapy agents h...The rapidly developing resistance of cancers to chemotherapy agents and the severe cytotoxicity of such agents to normal cells are major stumbling blocks in current cancer treatments.Most current chemotherapy agents have significant cytotoxicity,which leads to devastating adverse effects and results in a substandard quality of life,including increased daily morbidity and premature mortality.The death receptor of tumor necrosis factor-related apoptosis-inducing ligand(TRAIL)can sidestep p53-dependent pathways to induce tumor cell apoptosis without damaging most normal cells.However,various cancer cells can develop resistance to TRAIL-induced apoptosis via different pathways.Therefore,it is critical to find an efficient TRAIL sensitizer to reverse the resistance of tumor cells to TRAIL,and to reinforce TRAIL’s ability to induce tumor cell apoptosis.In recent years,traditional Chinese medicines and their active ingredients have shown great potential to trigger apoptotic cell death in TRAIL-resistant cancer cell lines.This review aims to collate information about Chinese medicines that can effectively reverse the resistance of tumor cells to TRAIL and enhance TRAIL’s ability to induce apoptosis.We explore the therapeutic potential of TRAIL and provide new ideas for the development of TRAIL therapy and the generation of new anticancer drugs for human cancer treatment.This study involved an extensive review of studies obtained from literature searches of electronic databases such as Google Scholar and PubMed."TRAIL sensitize"and"Chinese medicine"were the search keywords.We then isolated newly published studies on the mechanisms of TRAIL-induced apoptosis.The name of each plant was validated using certified databases such as The Plant List.This study indicates that TRAIL can be combined with different Chinese medicine components through intrinsic or extrinsic pathways to promote cancer cell apoptosis.It also demonstrates that the active ingredients of traditional Chinese medicines enhance the sensitivity of cancer cells to TRAIL-mediated apoptosis.This provides useful information regarding traditional Chinese medicine treatment,the development of TRAIL-based therapies,and the treatment of cancer.展开更多
AIM:To demonstrate the apoptosis-inducing effect of iidocalne on human corneal stromal(HCS)cells fn vitm,and provide experimental basis for safety anesthetic usage In clinic of ophthalmology.METHODS:In vitro cultured ...AIM:To demonstrate the apoptosis-inducing effect of iidocalne on human corneal stromal(HCS)cells fn vitm,and provide experimental basis for safety anesthetic usage In clinic of ophthalmology.METHODS:In vitro cultured HCS cells were treated with lidocaine at different doses and times,and their morphology was monitored successively with inverted phase contrast microscopy.The membrane permeability of them was detected by acridine orange/ethidium bromide(AO/EB)double staining.The DNA fragmentation of them was examined by agarose gel electrophoresis,and their ultrastructure was observed by transmission electron microscopy(TEM),respectively.RESULTS:Exposure to lidocaine at doses from0.3125g/L to 20g/L induced morphological changes of HCS cells such as cytoplasmic vacuolation,cellular shrinkage,and turning round,and elevated membrane permeability of these cells in AO/EB staining.The change of morphology and membrane permeability was doseand time-dependent,while lidocaine at dose below0.15625g/L could not induce these changes.Furthermore,lidocaine induced DNA fragmentation and ultrastructural changes such as cytoplasmic vacuolation,structural disorganization,chromatin condensation,and apoptotic body appearance of the cells.CONCLUSION:Lidocaine has significant cytotoxicity on human corneal stromal cells in vitro in a dose-and time-dependent manner by inducing apoptosis of these cells.The established experimental model and findingsbased on this model here help provide new insight into the apoptosis-inducing effect of local anesthetics in eye clinic.展开更多
Apoptosis induced by tumor necrosis factor-related apoptosis-inducing ligand(TRAIL) can be regulated by the epidermal growth factor(EGF) signaling pathway.In this study,recombinant adenoviral vectors that encode TRAIL...Apoptosis induced by tumor necrosis factor-related apoptosis-inducing ligand(TRAIL) can be regulated by the epidermal growth factor(EGF) signaling pathway.In this study,recombinant adenoviral vectors that encode TRAIL gene from the hTERT/RGD promoter(AdTRAIL) was combined with drugs including gefitinib,elotinib,and cetuximab that inhibit EGFR and the EGF signaling pathway in non-small cell lung cancer(NSCLC) cell lines to investigate their antitumor activity.In vitro,compared to single reagent,AdTRAIL combined with EGFR inhibitors reduced proliferation and enhanced apoptosis in H460,A549,and SW1573 cell lines.Western blot results suggested that these effects were relative to up-regulation of pro-apoptosis protein BAX and down-regulation of p-AKT.In vivo,AdTRAIL combined with cetuximab resulted in a significant growth reduction in H460 xenografts without damage to the main organs of nude mice.Histological examination and TUNEL analyses of xenografts showed that cetuximab enhanced cell apoptosis induced by AdTRAIL.These results indicate that EGFR inhibitors enhanced AdTRAIL anti-tumor activity in NSCLC cell lines and that inhibiting the AKT pathway played an important role in this enhancement.展开更多
Tumor necrosis factor-related apoptosis-inducing ligand （TRAIL） is a promising anti-cancer agent. Epigallocatechin-3-gallate （EGCG） is a polyphenolic constituent of green tea. In this study, inhibitory effect of c...Tumor necrosis factor-related apoptosis-inducing ligand （TRAIL） is a promising anti-cancer agent. Epigallocatechin-3-gallate （EGCG） is a polyphenolic constituent of green tea. In this study, inhibitory effect of combined use of EGCG and TRAIL on human melanoma A375 cells was examined and the possible mechanism investigated. The cells were divided into 4 groups： control group, EGCG group （EGCG： 10, 20 μg/mL）, TRAIL group （TRAIL： 25 ng/mL） and EGCG＋TRAIL group （combined group）. The growth inhibition was measured in the A375 cells treated with different concentrations of TRAIL （（25, 50, 75, 100, 125, 150 ng/mL） by MTT assay. The apoptosis was assessed by flow cytometry. The expressions of DR4 and DR5 were detected by flow cytometry and western blotting. The activities of caspase-8 and caspase-3 were determined by colorimetric assay. The results showed that TRAIL could dose-dependently inhibit the growth of A375 cells and the IC50 of TRAIL was 150 ng/mL. The apoptosis rate was 11.8% in the TRAIL group, 5%–7% in the EGCG group and 48.9%–59.1% in the combined group. Significant difference was found in the apoptosis rate between the combined group and the EGCG or TRAIL group （P〈0.05 for each）. The expression of DR4 instead of DR5 was significantly increased in the EGCG group. The activity of caspase-3 rather than caspase-8 was substantially enhanced in the EGCG group. These results suggest that EGCG is useful for the TRAIL-based treatment for melanoma.展开更多
Objective: To study the expression of caspase-3 and tumor necrosis factor-related apoptosisinducing ligand (TRAIL) receptors in the CD4+ and CD8+ T cells of systemic lupus enythematosus (SLE) patients. Methods: RT-PCR...Objective: To study the expression of caspase-3 and tumor necrosis factor-related apoptosisinducing ligand (TRAIL) receptors in the CD4+ and CD8+ T cells of systemic lupus enythematosus (SLE) patients. Methods: RT-PCR was used to analyze the expression of caspase-3 and TRAIL receptors in CD4+ and CD8+ T cells of SLE patients and normal subjects. Results: The death domain-containing TRAIL-R1/R2 as well as 'decoy' TRAIL-R3/R4 were co-expressed in majority of CD4+ and CD8+ T cells in both SLE patients and normal subjects. The CD8+ T cells from SLE patients showed significantly higher expression of caspase-3 and TRAIL-R2 than those from normal subjects,and the expression was correlated with the activity of the disease. Conclusion: The TRAIL-R2 signal pathway might contribute to the apoptosis of T cells in SLE.展开更多
AIM： To investigate the potential of pigment epitheliumderived factor（PEDF） to protect the immortalized rat retinal ganglion cells-5（RGC-5） exposed to Co Cl2-induced chemical hypoxia. METHODS： After being differ...AIM： To investigate the potential of pigment epitheliumderived factor（PEDF） to protect the immortalized rat retinal ganglion cells-5（RGC-5） exposed to Co Cl2-induced chemical hypoxia. METHODS： After being differentiated with staurosporine（SS）, RGC-5 cells were cultured in four conditions： control group cells cultured in Dulbecco ＇s modified eagle medium（DMEM） supplemented with 10% fetal bovine serum, 100 μmol/m L streptomycin and penicillin（named as normal conditions）; hypoxia group cells cultured in DMEM containing 300 μmol/m L Co Cl2; cells in the group protected by PEDF were first pretreated with 100 ng/m L PEDF for 2h and then cultured in the same condition as hypoxia group cells; and PEDF group cells that were cultured in the presence of 100 ng/m L PEDF under normal conditions. The cell viability was assessed by MTT assay, the percentage of apoptotic cells was quantified using Annexin V-FITC apoptosis kit, and intra-cellar reactive oxygen species（ROS） was measured by dichloro-dihydro-fluorescein diacetate（DCFH-DA） probe. The mitochondria-mediated apoptosis was also examined to further study the underlying mechanism of the protective effect of PEDF. The opening of mitochondrial permeability transition pores（m PTPs） and membrane potential（Δψm） were tested as cellular adenosine triphosphate（ATP） level and glutathione（GSH）. Also, the expression and distribution of Cyt C and apoptosis inducing factor（AIF） were observed.RESULTS： SS induced differentiation of RGC-5 cells resulting in elongation of their neurites and establishing contacts between outgrowths. Exposure to 300 μmol/m L Co Cl2 triggered death of 30% of the total cells in cultures within 24 h. At the same time, pretreatment with 100 ng/m L PEDF significantly suppressed the cell death induced by hypoxia（P〈0.05）. The apoptosis induced by treatment of Co Cl2 was that induced cell death accompanied with increasing intracellar ROS and decreasing GSH and ATP level. PEDF pretreatment suppressed these effects（P〈0.05）. Additionally, PEDF treatment inhibited the opening of m PTPs and suppressed decreasing of Δψm in RGC-5 cells, resulting in blocking of the mitochondrial apoptotic pathway.CONCLUSION： Pretreatment of RGC-5 cells with 100 ng/m L PEDF significantly decreases the extent of apoptosis. PEDF inhibits the opening of m PTPs and suppresses decreasing of Δψm. Moreover, PEDF also reduces ROS production and inhibits cellular ATP level＇s reduction. Cyt C and AIF activation in PEDF-pretreated cultures are also reduced. These results demonstrate the potential for PEDF to protect RGCs against hypoxic damage in vitro by preventing mitochondrial dysfunction.展开更多
Objective: To investigate whether piperlongumine can sensitize prostate cancer cells to tumor necrosis factor-related apoptosisinducing ligand(TRAIL) and trigger apoptosis in prostate cells. Methods: Human prostate ca...Objective: To investigate whether piperlongumine can sensitize prostate cancer cells to tumor necrosis factor-related apoptosisinducing ligand(TRAIL) and trigger apoptosis in prostate cells. Methods: Human prostate cancer cell lines PC3, LNCa P, and VCa P were cultured with piperlongumine and TRAIL. Then, cell proliferation, migration, caspase activation, apoptotic protein expressions, and death receptor expressions were measured.Results: Piperlongumine inhibited cell proliferation at low doses(<10 μM) alone and in combination with TRAIL(25 ng/m L), induced apoptosis, and suppressed cyclooxygenase activation. Additionally, piperlongumine induced expression of death receptors which potentiated TRAIL-induced apoptosis in cancer cells but did not affect decoy receptors. Piperlongumine also downregulated tumor cell-survival pathways, inhibited colony formation and migration of cancer cells alone or in combination with TRAIL. The combination of piperlongumine with TRAIL was found to be synergistic. Conclusions: Our findings indicate that piperlongumine can sensitize cancer cells to TRAIL through the upregulation of death receptors and can trigger apoptosis with the downregulation of antiapoptotic proteins.展开更多
OBJECTIVE TNF-related apoptosis-inducing ligand(TRAIL)is a promising cancer therapeutic agent due to its minimal toxicity to normal tissues and remarkable apoptotic activity in tumors.However,most breast cancer cells ...OBJECTIVE TNF-related apoptosis-inducing ligand(TRAIL)is a promising cancer therapeutic agent due to its minimal toxicity to normal tissues and remarkable apoptotic activity in tumors.However,most breast cancer cells are resistant to TRAIL-induced apoptosis.Our objectives are to investigate the underlying molecular mechanisms and to develop strategies to overcome such resistance.METHODS To identify modulators of TRAIL-induced apoptosis,we carried out a genome wide si RNA screen.To validate the screening result,we either silenced or overexpressed the identified genes in various breast cancer cells and changes in growth and TRAIL-induced cell apoptosis were determined in vitro and in an orthotopic xenograft mouse model.Finally,we investigated whether small molecules targeting the identified genes improve the effectiveness of TRAIL-therapy.RESULTS We unexpectedly identified androgen receptor(AR)to be responsible for TRAIL resistance.While AR is classically viewed as the key factor in prostate cancer progression,we found that AR expression levels were markedly elevated in human invasive breast cancer specimens including triple-negative breast cancers(TNBC)that are highly aggressive with poor prognosis.Importantly,breast cancer cell lines express different levels of AR that correlated with their TRAIL resistance.AR overexpression in MDA-MB-231 and MDA-MB-436 cells suppressed the TRAIL sensitivity whereas knockdown of AR rendered MCF-7 and MDA-MB-453 cells sensitive to TRAIL-induced apoptosis.AR overexpression also induced TRAIL resistance in breast tumors in vivo.Further,we observed an upregulation of the TRAIL receptor,death receptor 5(DR5)in breast cancer cells,following the removal or inhibition of AR by its antagonists Casodex and MDV3100.Treatment with AR antagonists also enhanced TRAIL-induced breast cancer cell apoptosis.CONCLUSION AR signaling suppresses TRAIL-induced breast cancer cell apoptosis,in part,by suppressing DR5 expression,and a combination of AR antagonists together with TRAIL may be a novel and effective therapy for TNBC.展开更多
BACKGROUND The biochemical phenomenon defined as poly adenosine diphosphate(ADP)-ribosylation(PARylation)is essential for the progression of pancreatic cancer.However,the excessive accumulation of poly ADP-ribose(PAR)...BACKGROUND The biochemical phenomenon defined as poly adenosine diphosphate(ADP)-ribosylation(PARylation)is essential for the progression of pancreatic cancer.However,the excessive accumulation of poly ADP-ribose(PAR)induces apoptosis-inducing factor(AIF)release from mitochondria and energy deprivation resulting in the caspase-independent death of cancer cells.AIM To investigate whether sustained calcium supply could induce an anticancer effect on pancreatic cancer by PAR accumulation.METHODS Two pancreatic cancer cell lines,AsPC-1 and CFPAC-1 were used for the study.Calcium influx and mitochondrial reactive oxygen species(ROS)were observed by fluorescence staining.Changes in enzyme levels,as well as PAR accumulation and energy metabolism,were measured using assay kits.AIF-dependent cell death was investigated followed by confirming in vivo anticancer effects by sustained calcium administration.RESULTS Mitochondrial ROS levels were elevated with increasing calcium influx into pancreatic cancer cells.Then,excess PAR accumulation,decreased PAR glycohydrolase and ADP-ribosyl hydrolase 3 levels,and energy deprivation were observed.In vitro and in vivo antitumor effects were confirmed to accompany elevated AIF levels.CONCLUSION This study visualized the potential anticancer effects of excessive PAR accumulation by sustained calcium supply on pancreatic cancer,however elucidating a clear mode of action remains a challenge,and it should be accompanied by further studies to assess its potential for clinical application.展开更多
This study is to examine the effect of human recombinant soluble TRAIL （TNF-related apoptosis-inducing ligand） protein inducing apoptosis in MG-63 human osteosarcoma cells. The inhibitive rates of TRAIL to MG-63 cel...This study is to examine the effect of human recombinant soluble TRAIL （TNF-related apoptosis-inducing ligand） protein inducing apoptosis in MG-63 human osteosarcoma cells. The inhibitive rates of TRAIL to MG-63 cells were detected by MTT assay. The apoptosis induced by TRAIL in MG-63 human osteosarcoma cells was analyzed with FACS and TUNEL and the apoptotic bodies were observed by transmission electron microscope. MTT assay showed that the inhibitive rates of 500, 1 000, 2 000 and 4 000 ng/mL TRAIL for 24 h were 10.1%, 24.3%, 50.6% and 97.7% respectively. Flow cytometric analysis showed that after MG-63 cells were treated with 2 gg/mL TRAIL for 6 h, obvious apoptotic peak would immediately appear before diploid peak. Human soluble TRAIL protein can quickly kill MG-63 osteosarcoma cells selectively, and may have potential value for clinical treatment of osteosarcoma.展开更多
基金National Key R&D Program of China:Cooperating Studies on Measurement Technologies of Human Phenome and Crossscale Correlation of Phenotypic Data(No.2020YFE0201600)National Nature Science Foundation:Study on LncRNA-CCDC18-AS1 Mediated Osteosarcoma Occurrence by Activating YAP/TAZ and Tumor Microenvironment M2 TAM-dependent Lung Metastasis,and Efficacy/mechanism of Removing Blood Stasis/clearing heat/eliminating Toxic Material Principle(No.81973877)+2 种基金Mechanism Study on m6A Methyltransferase RBM15 Mediated YAP Epigenetic Modification to Promote Osteosarcoma Lung Metastasis through Lymphatic System and Management with Qichong Powder(No.82174408)Shanghai Collaborative Innovation Center of Industrial Transformation of Hospital TCM Preparation:Preclinical Study on the Treatment of Osteosarcoma with Qingre Jiedu GranulesResearch Projects within Budget of Shanghai University of Traditional Chinese Medicine:the Research on the Mechanism of the HIPK3 Activation of Wnt/β-catenin Induction the Osteosarcoma and the Intervention of Banmao Decoction(No.2021LK047)。
文摘OBJECTIVE:To investigate the synergistic effects of polyphyllin Ⅰ(PPⅠ)combined with tumor necrosis factorrelated apoptosis-inducing ligand(TRAIL)on the growth of osteosarcoma cells through downregulating the Wnt/β-catenin signaling pathway.METHODS:Cell viability,apoptosis and cell cycle distribution were examined using cell counting kit-8 and flow cytometry assays.The morphology of cancer cells was observed with inverted phase contrast microscope.The migration and invasion abilities were examined by xCELLigence real time cell analysis DP system and transwell assays.The expressions of poly(adenosine diphosphate-ribose)polymerase,C-Myc,Cyclin B1,cyclin-dependent kinases 1,N-cadherin,Vimentin,Active-β-catenin,β-catenin,p-glycogen synthase kinase 3β(GSK-3β)and GSK-3βwere determined by Western blotting assay.RESULTS:PPⅠ sensitized TRAIL-induced decrease of viability,migration and invasion,as well as increase of apoptosis and cell cycle arrest of MG-63 and U-2 OS osteosarcoma cells.The synergistic effect of PPⅠwith TRAIL in inhibiting the growth of osteosarcoma cells was at least partially realized through the inactivation of Wnt/β-catenin signaling pathway.CONCLUSION:The combination of PPⅠ and TRAIL is potentially a novel treatment strategy of osteosarcoma.
基金the National Natural Science Foundation of China(Nos.32070584,81830028,31771398,82222016,and 8207040100)the Zhejiang Provincial Natural Science Foundation of China(No.LZ19C060001)the Fundamental Research Funds for the Central Universities(No.2019QNA6001)。
文摘Auditory neuropathy spectrum disorder(ANSD)represents a variety of sensorineural deafness conditions characterized by abnormal inner hair cells and/or auditory nerve function,but with the preservation of outer hair cell function.ANSD represents up to 15%of individuals with hearing impairments.Through mutation screening,bioinformatic analysis and expression studies,we have previously identified several apoptosis-inducing factor(AIF)mitochondria-associated 1(AIFM1)variants in ANSD families and in some other sporadic cases.Here,to elucidate the pathogenic mechanisms underlying each AIFM1 variant,we generated AIF-null cells using the clustered regularly interspersed short palindromic repeats(CRISPR)/CRISPR-associated protein 9(Cas9)system and constructed AIF-wild type(WT)and AIF-mutant(mut)(p.T260A,p.R422W,and p.R451Q)stable transfection cell lines.We then analyzed AIF structure,coenzyme-binding affinity,apoptosis,and other aspects.Results revealed that these variants resulted in impaired dimerization,compromising AIF function.The reduction reaction of AIF variants had proceeded slower than that of AIF-WT.The average levels of AIF dimerization in AIF variant cells were only 34.5%-49.7%of that of AIF-WT cells,resulting in caspase-independent apoptosis.The average percentage of apoptotic cells in the variants was 12.3%-17.9%,which was significantly higher than that(6.9%-7.4%)in controls.However,nicotinamide adenine dinucleotide(NADH)treatment promoted the reduction of apoptosis by rescuing AIF dimerization in AIF variant cells.Our findings show that the impairment of AIF dimerization by AIFM1 variants causes apoptosis contributing to ANSD,and introduce NADH as a potential drug for ANSD treatment.Our results help elucidate the mechanisms of ANSD and may lead to the provision of novel therapies.
文摘Most of the ocular tumors have poor prognosis, and they remain a difficult problem in the area of ophthalmology. With the rapid development of molecular biology and immunologic techniques and the deep research on ocular tumor related genes, it becomes possible to diagnose and treat malignant tumors from the molecular level. The tumor necrosis factor related apoptosis-inducing ligand (TRAIL), a member of the tumor necrosis factor (TNF) super family, is a promising candidate, either alone or in combination with established cancer therapies, since it can initiate apoptosis through the activation of their death receptors. The ability of TRAIL to selectively induce apoptosis of transformed, virus-infected or tumor cells but not normal cells promotes the development of TRAIL-based cancer therapy. Here, we will review TRAIL and its receptors' structure, function, mechanism of action and application in ocular tumors therapy.
基金the National Natural Science Foundation of China, No. 30672409the Science and Technology Foundation Program of Guangdong Province, No. 2006B36003017
文摘Studies have shown that tumor necrosis factor-related apoptosis-inducing ligand（TRAIL）exhibits strong induction of apoptosis in human glioma cells.It remains unclear whether the mitochondrion pathway,an important apoptosis signaling pathway,is involved in TRAIL-induced glioma cell apoptosis.In the present study,in vitro cultured human glioma U87 cells were treated with human recombinant soluble TRAIL.Apoptosis of glioma U87 cells,mitochondrial transmembrane potential（Δψm）,cytoplasmic cytochrome c concentration and changes in caspase-3,-8 and-9 activity following human recombinant soluble TRAIL treatment were investigated to determine the mechanism of glioma U87 cell apoptosis induced by TRAIL.Additionally,blocking caspase-8resulted in TRAIL-induced mitochondrion pathway activation,suggesting that TRAIL,through activating caspase-8,initiated a series of mitochondrial events and resulted in apoptosis of glioma U87 cells.
基金Sopported by the National Nature Science Foundation grant of P. R. China( 39880 0 31)
文摘hap, a novel human apoptosis-inducing gene which can interact with another newly discovered apoptosis-inducing geneASY, was identified, by cloning its cDNAs from human lung cell line (WI-38) cDNA library. Two major mRNA species (1.8 and 2.7 kb in length, respectively) were previously identified by Northern blot analysis of poly(A)+ RNA from human multiple tissues using partialhap cDNA as a probe. In the present work, the molecular mechanism accounting for the generation of the twohap transcripts were investigated. The rapid amplification of cDNA 3′-ends (3′-RACE) technique and the sequential Southern blot analysis, in conjunction with the sequencing analysis demonstrated that the twohap transcripts derive from the alternative polyadenylation site selection: a AATAAA signal at position 1 528–1 533 nt for the 1.8 kbhap mRNA: and a AATAAA signal at position 2 375–2 380 nt for the 2.7 kbhap mRNA. Furthermore, a number of regulatory elements withinhap 3′-untranslated region (3′-UTR) were also examined.
文摘Objective:Structural abnormalities and dysfunction of the placenta contribute to pregnancy-related complications,such as preeclampsia.Syncytin-A(synA)has been reported to be expressed in the placenta.The contribution of synA to developmental abnormalities and dysfunction of the placenta remains elusive.In this study,we aimed to explore the role of synA in placental development and functions.Methods:SynA-knockout mice were generated using the CRISPR-Cas9 method,and the phenotypes of the placenta and fetus ofsynA-knockout mice were observed.Real-time quantitative polymerase chain reaction(PCR)and routine PCR were employed to detect the genotypes of the offspring.CD31 immunohistochemistry was used to evaluate the vessel density of the placenta,and the protein levels of key molecules were measured by western blotting.Results:SynA knockout caused fetal death.Furthermore,synA-knockout mice showed placental developmental abnormalities,indicated by a thinner labyrinth layer,thicker spongiotrophoblast layer,lower blood vessel density,and significantly higher numbers of apoptotic trophoblasts,when compared with wild-type littermates.Mechanistically,synA ablation induced apoptosis-inducing factor(AIF)cleavage and nuclear localization and promoted placental trophoblast apoptosis.In addition,synA knockout increased the calpain1 protein levels.The calpain1 inhibitor calpeptin blockedsynA knockout-induced AIF cleavage,partially restoring the placental structural abnormalities ofsynA-knockout mice.Conclusions:SynA knockout leads to placental developmental abnormalities by inducing trophoblastic apoptosis via the calpain1-AIF pathway.
文摘Tumor necrosis factor-related apoptosis-inducing ligand （TRAIL） is a promising agent for anticancer therapy. The identification of small molecules that can establish the sensitivity of prostate cancer （PCa） cells to TRAIL-induced apoptosis is crucial for the targeted treatment of PCa. PC3, DU145, JAC-1, TsuPrl, and LNCaP cells were treated with Andrographolide （Andro） and TRAIL, and the apoptosis was measured using the Annexin V/PI double staining method. Real time-polymerase chain reaction （PCR） and Western blot analysis were performed to measure the expression levels of target molecules. RNA interference technique was used to down-regulate the expression of the target protein. We established a nude mouse xenograft model of PCa, which was used to measure the caspase-3 activity in the tumor cells using flow cytometry. In this research study, our results demonstrated that Andro preferentially increased the sensitivity of PCa cells to TRAIL-induced apoptosis at subtoxic concentrations, and the regulation mechanism was related to the up-regulation of DR4. In addition, it also increased the p53 expression and led to the generation of reactive oxygen species （ROS） in the cells. Further research revealed that the DR4 inhibition, p53 expression, and ROS generation can significantly reduce the apoptosis induced by the combination of TRAIL and Andro in PCa cells. In conclusion, Andro increases the sensitivity of PCa cells to TRAIL-induced apoptosis through the generation of ROS and up-regulation of p53 and then promotes PCa cell apoptosis associated with the activation of DR4.
文摘Portal hypertension （PHT） gastropathy is a frequent complication of fiver cirrhosis and one of the leading causes of death from cirrhosis. Apoptosis is widely considered to be an active energy-dependent mode of cell death and a distinct entity from necrotic cell death. It is unclear whether gastric mucosal apoptosis is involved in PHT gastropa- thy. Prostaglandins （PGs） produced through cyclooxygenase （COX） are thought to play a key role in protection of the gastrointestinal mucosa from injury and apoptosis. However, the role of COX in PHT gastropathy is still not clearly understood. The aims of this study were to investigate whether （1） gastric mucosal apoptosis is involved in PHT gas- tropathy and （2） downregulation of COX contributes to this apoptosis. In this study, we show that gastric mucosal apoptosis was remarkably increased while mucosal proliferation was inhibited in PHT rats. Gastric mucosal COX- 1 was significantly suppressed at both the mRNA and protein levels, and PGE2 was reduced in PHT rats. Further, PGE2 treatment suppressed gastric mucosal apoptosis in PHT rats. However, gastric mucosal COX-2 levels did not differ between sham-operated rats and PHT rats. Gastric mucosal levels of tumor necrosis factor-α （TNF-α） and Fas ligand, but not TNF-related apoptosis-inducing ligand, were increased, and activated caspase-8 and caspase-3 levels were upregulated in PHT rats. The release of cytochrome c from the mitochondria to the cytosol was not observed in PHT rats. Our data indicate that downregulation of COX-1 is involved in gastric mucosal apoptosis via death signal- ing-mediated type-I cell death in PHT rats.
基金Supported by DFG/German Research Foundation,No.FI 1630/3-1 and No.IFORES D/107-114400(to CDF)
文摘To examine the influence on apoptotic mechanisms following inhibition of polo-like kinases as therapeutically approach for cholangiocellular cancer treatment.METHODSAs most cholangiocarcinomas are chemotherapy-resistant due to mechanisms preventing tumor cell death, we investigated the effect of Cisplatin on cholangiocellular carcinoma (CCA) cell lines KMCH-1 and Mz-Ch-1. Polo-like kinases (PLK) are important regulators of the cell cycle and their inhibition is discussed as a potential therapy while PLK inhibition can regulate apoptotic mediators. Here, cells were treated with PLK inhibitor BI6727 (Volasertib), Cisplatin, and in combination of both compounds. Cell viability was assessed by MTT; apoptosis was measured by DAPI staining and caspase-3/-7 assay. Western blot and qRT-PCR were used to measure expression levels of apoptosis-related molecules Bax and Bcl-2.RESULTSThe cell viability in the CCA cell lines KMCH-1 and Mz-Ch-1 was reduced in all treatment conditions compared to vehicle-treated cells. Co-treatment with BI6727 and cisplatin could even enhance the cytotoxic effect of cisplatin single treatment. Thus, co-treatment of cisplatin with BI6727 could slightly enhance the cytotoxic effect of the cisplatin in both cell lines whereas there was evidence of increased apoptosis induction solely in Mz-Ch-1 as compared to KMCH-1. Moreover, PLK inhibition decreases protein levels of Bcl-2; an effect that can be reversed by the proteasomal degradation inhibitor MG-132. In contrast, protein levels of Bax were not found to be altered by PLK inhibition. These findings indicate that cytotoxic effects of Cisplatin in Mz-Ch-1 cells can be enhanced by cotreatment with BI6727.CONCLUSIONIn conclusion, BI6727 treatment can sensitize CCA cells to cisplatin-induced apoptosis with proteasomal Bcl-2 degradation as an additional pro-apoptotic effect.
基金supported by the Guangdong Basic and Applied Basic Research Foundation(No.2019A1515110167),China。
文摘The rapidly developing resistance of cancers to chemotherapy agents and the severe cytotoxicity of such agents to normal cells are major stumbling blocks in current cancer treatments.Most current chemotherapy agents have significant cytotoxicity,which leads to devastating adverse effects and results in a substandard quality of life,including increased daily morbidity and premature mortality.The death receptor of tumor necrosis factor-related apoptosis-inducing ligand(TRAIL)can sidestep p53-dependent pathways to induce tumor cell apoptosis without damaging most normal cells.However,various cancer cells can develop resistance to TRAIL-induced apoptosis via different pathways.Therefore,it is critical to find an efficient TRAIL sensitizer to reverse the resistance of tumor cells to TRAIL,and to reinforce TRAIL’s ability to induce tumor cell apoptosis.In recent years,traditional Chinese medicines and their active ingredients have shown great potential to trigger apoptotic cell death in TRAIL-resistant cancer cell lines.This review aims to collate information about Chinese medicines that can effectively reverse the resistance of tumor cells to TRAIL and enhance TRAIL’s ability to induce apoptosis.We explore the therapeutic potential of TRAIL and provide new ideas for the development of TRAIL therapy and the generation of new anticancer drugs for human cancer treatment.This study involved an extensive review of studies obtained from literature searches of electronic databases such as Google Scholar and PubMed."TRAIL sensitize"and"Chinese medicine"were the search keywords.We then isolated newly published studies on the mechanisms of TRAIL-induced apoptosis.The name of each plant was validated using certified databases such as The Plant List.This study indicates that TRAIL can be combined with different Chinese medicine components through intrinsic or extrinsic pathways to promote cancer cell apoptosis.It also demonstrates that the active ingredients of traditional Chinese medicines enhance the sensitivity of cancer cells to TRAIL-mediated apoptosis.This provides useful information regarding traditional Chinese medicine treatment,the development of TRAIL-based therapies,and the treatment of cancer.
基金National High Technology Research and Development Program("863"Program)of China(No.2006AA02A132)
文摘AIM:To demonstrate the apoptosis-inducing effect of iidocalne on human corneal stromal(HCS)cells fn vitm,and provide experimental basis for safety anesthetic usage In clinic of ophthalmology.METHODS:In vitro cultured HCS cells were treated with lidocaine at different doses and times,and their morphology was monitored successively with inverted phase contrast microscopy.The membrane permeability of them was detected by acridine orange/ethidium bromide(AO/EB)double staining.The DNA fragmentation of them was examined by agarose gel electrophoresis,and their ultrastructure was observed by transmission electron microscopy(TEM),respectively.RESULTS:Exposure to lidocaine at doses from0.3125g/L to 20g/L induced morphological changes of HCS cells such as cytoplasmic vacuolation,cellular shrinkage,and turning round,and elevated membrane permeability of these cells in AO/EB staining.The change of morphology and membrane permeability was doseand time-dependent,while lidocaine at dose below0.15625g/L could not induce these changes.Furthermore,lidocaine induced DNA fragmentation and ultrastructural changes such as cytoplasmic vacuolation,structural disorganization,chromatin condensation,and apoptotic body appearance of the cells.CONCLUSION:Lidocaine has significant cytotoxicity on human corneal stromal cells in vitro in a dose-and time-dependent manner by inducing apoptosis of these cells.The established experimental model and findingsbased on this model here help provide new insight into the apoptosis-inducing effect of local anesthetics in eye clinic.
文摘Apoptosis induced by tumor necrosis factor-related apoptosis-inducing ligand(TRAIL) can be regulated by the epidermal growth factor(EGF) signaling pathway.In this study,recombinant adenoviral vectors that encode TRAIL gene from the hTERT/RGD promoter(AdTRAIL) was combined with drugs including gefitinib,elotinib,and cetuximab that inhibit EGFR and the EGF signaling pathway in non-small cell lung cancer(NSCLC) cell lines to investigate their antitumor activity.In vitro,compared to single reagent,AdTRAIL combined with EGFR inhibitors reduced proliferation and enhanced apoptosis in H460,A549,and SW1573 cell lines.Western blot results suggested that these effects were relative to up-regulation of pro-apoptosis protein BAX and down-regulation of p-AKT.In vivo,AdTRAIL combined with cetuximab resulted in a significant growth reduction in H460 xenografts without damage to the main organs of nude mice.Histological examination and TUNEL analyses of xenografts showed that cetuximab enhanced cell apoptosis induced by AdTRAIL.These results indicate that EGFR inhibitors enhanced AdTRAIL anti-tumor activity in NSCLC cell lines and that inhibiting the AKT pathway played an important role in this enhancement.
文摘Tumor necrosis factor-related apoptosis-inducing ligand （TRAIL） is a promising anti-cancer agent. Epigallocatechin-3-gallate （EGCG） is a polyphenolic constituent of green tea. In this study, inhibitory effect of combined use of EGCG and TRAIL on human melanoma A375 cells was examined and the possible mechanism investigated. The cells were divided into 4 groups： control group, EGCG group （EGCG： 10, 20 μg/mL）, TRAIL group （TRAIL： 25 ng/mL） and EGCG＋TRAIL group （combined group）. The growth inhibition was measured in the A375 cells treated with different concentrations of TRAIL （（25, 50, 75, 100, 125, 150 ng/mL） by MTT assay. The apoptosis was assessed by flow cytometry. The expressions of DR4 and DR5 were detected by flow cytometry and western blotting. The activities of caspase-8 and caspase-3 were determined by colorimetric assay. The results showed that TRAIL could dose-dependently inhibit the growth of A375 cells and the IC50 of TRAIL was 150 ng/mL. The apoptosis rate was 11.8% in the TRAIL group, 5%–7% in the EGCG group and 48.9%–59.1% in the combined group. Significant difference was found in the apoptosis rate between the combined group and the EGCG or TRAIL group （P〈0.05 for each）. The expression of DR4 instead of DR5 was significantly increased in the EGCG group. The activity of caspase-3 rather than caspase-8 was substantially enhanced in the EGCG group. These results suggest that EGCG is useful for the TRAIL-based treatment for melanoma.
基金Supported by the National Natural Science Foundation of China (No. 30271199)
文摘Objective: To study the expression of caspase-3 and tumor necrosis factor-related apoptosisinducing ligand (TRAIL) receptors in the CD4+ and CD8+ T cells of systemic lupus enythematosus (SLE) patients. Methods: RT-PCR was used to analyze the expression of caspase-3 and TRAIL receptors in CD4+ and CD8+ T cells of SLE patients and normal subjects. Results: The death domain-containing TRAIL-R1/R2 as well as 'decoy' TRAIL-R3/R4 were co-expressed in majority of CD4+ and CD8+ T cells in both SLE patients and normal subjects. The CD8+ T cells from SLE patients showed significantly higher expression of caspase-3 and TRAIL-R2 than those from normal subjects,and the expression was correlated with the activity of the disease. Conclusion: The TRAIL-R2 signal pathway might contribute to the apoptosis of T cells in SLE.
基金Supported by National Natural Science Foundation of China(No.81100665)
文摘AIM： To investigate the potential of pigment epitheliumderived factor（PEDF） to protect the immortalized rat retinal ganglion cells-5（RGC-5） exposed to Co Cl2-induced chemical hypoxia. METHODS： After being differentiated with staurosporine（SS）, RGC-5 cells were cultured in four conditions： control group cells cultured in Dulbecco ＇s modified eagle medium（DMEM） supplemented with 10% fetal bovine serum, 100 μmol/m L streptomycin and penicillin（named as normal conditions）; hypoxia group cells cultured in DMEM containing 300 μmol/m L Co Cl2; cells in the group protected by PEDF were first pretreated with 100 ng/m L PEDF for 2h and then cultured in the same condition as hypoxia group cells; and PEDF group cells that were cultured in the presence of 100 ng/m L PEDF under normal conditions. The cell viability was assessed by MTT assay, the percentage of apoptotic cells was quantified using Annexin V-FITC apoptosis kit, and intra-cellar reactive oxygen species（ROS） was measured by dichloro-dihydro-fluorescein diacetate（DCFH-DA） probe. The mitochondria-mediated apoptosis was also examined to further study the underlying mechanism of the protective effect of PEDF. The opening of mitochondrial permeability transition pores（m PTPs） and membrane potential（Δψm） were tested as cellular adenosine triphosphate（ATP） level and glutathione（GSH）. Also, the expression and distribution of Cyt C and apoptosis inducing factor（AIF） were observed.RESULTS： SS induced differentiation of RGC-5 cells resulting in elongation of their neurites and establishing contacts between outgrowths. Exposure to 300 μmol/m L Co Cl2 triggered death of 30% of the total cells in cultures within 24 h. At the same time, pretreatment with 100 ng/m L PEDF significantly suppressed the cell death induced by hypoxia（P〈0.05）. The apoptosis induced by treatment of Co Cl2 was that induced cell death accompanied with increasing intracellar ROS and decreasing GSH and ATP level. PEDF pretreatment suppressed these effects（P〈0.05）. Additionally, PEDF treatment inhibited the opening of m PTPs and suppressed decreasing of Δψm in RGC-5 cells, resulting in blocking of the mitochondrial apoptotic pathway.CONCLUSION： Pretreatment of RGC-5 cells with 100 ng/m L PEDF significantly decreases the extent of apoptosis. PEDF inhibits the opening of m PTPs and suppresses decreasing of Δψm. Moreover, PEDF also reduces ROS production and inhibits cellular ATP level＇s reduction. Cyt C and AIF activation in PEDF-pretreated cultures are also reduced. These results demonstrate the potential for PEDF to protect RGCs against hypoxic damage in vitro by preventing mitochondrial dysfunction.
基金supported by the Turkish Scientific Council(TUBITAK),Grant#115S942.
文摘Objective: To investigate whether piperlongumine can sensitize prostate cancer cells to tumor necrosis factor-related apoptosisinducing ligand(TRAIL) and trigger apoptosis in prostate cells. Methods: Human prostate cancer cell lines PC3, LNCa P, and VCa P were cultured with piperlongumine and TRAIL. Then, cell proliferation, migration, caspase activation, apoptotic protein expressions, and death receptor expressions were measured.Results: Piperlongumine inhibited cell proliferation at low doses(<10 μM) alone and in combination with TRAIL(25 ng/m L), induced apoptosis, and suppressed cyclooxygenase activation. Additionally, piperlongumine induced expression of death receptors which potentiated TRAIL-induced apoptosis in cancer cells but did not affect decoy receptors. Piperlongumine also downregulated tumor cell-survival pathways, inhibited colony formation and migration of cancer cells alone or in combination with TRAIL. The combination of piperlongumine with TRAIL was found to be synergistic. Conclusions: Our findings indicate that piperlongumine can sensitize cancer cells to TRAIL through the upregulation of death receptors and can trigger apoptosis with the downregulation of antiapoptotic proteins.
基金supported by National Institutes of Health(R21CA193271 and R01HL116849)National Natural Science Foundation of China(31100595 and 31300683)
文摘OBJECTIVE TNF-related apoptosis-inducing ligand(TRAIL)is a promising cancer therapeutic agent due to its minimal toxicity to normal tissues and remarkable apoptotic activity in tumors.However,most breast cancer cells are resistant to TRAIL-induced apoptosis.Our objectives are to investigate the underlying molecular mechanisms and to develop strategies to overcome such resistance.METHODS To identify modulators of TRAIL-induced apoptosis,we carried out a genome wide si RNA screen.To validate the screening result,we either silenced or overexpressed the identified genes in various breast cancer cells and changes in growth and TRAIL-induced cell apoptosis were determined in vitro and in an orthotopic xenograft mouse model.Finally,we investigated whether small molecules targeting the identified genes improve the effectiveness of TRAIL-therapy.RESULTS We unexpectedly identified androgen receptor(AR)to be responsible for TRAIL resistance.While AR is classically viewed as the key factor in prostate cancer progression,we found that AR expression levels were markedly elevated in human invasive breast cancer specimens including triple-negative breast cancers(TNBC)that are highly aggressive with poor prognosis.Importantly,breast cancer cell lines express different levels of AR that correlated with their TRAIL resistance.AR overexpression in MDA-MB-231 and MDA-MB-436 cells suppressed the TRAIL sensitivity whereas knockdown of AR rendered MCF-7 and MDA-MB-453 cells sensitive to TRAIL-induced apoptosis.AR overexpression also induced TRAIL resistance in breast tumors in vivo.Further,we observed an upregulation of the TRAIL receptor,death receptor 5(DR5)in breast cancer cells,following the removal or inhibition of AR by its antagonists Casodex and MDV3100.Treatment with AR antagonists also enhanced TRAIL-induced breast cancer cell apoptosis.CONCLUSION AR signaling suppresses TRAIL-induced breast cancer cell apoptosis,in part,by suppressing DR5 expression,and a combination of AR antagonists together with TRAIL may be a novel and effective therapy for TNBC.
文摘BACKGROUND The biochemical phenomenon defined as poly adenosine diphosphate(ADP)-ribosylation(PARylation)is essential for the progression of pancreatic cancer.However,the excessive accumulation of poly ADP-ribose(PAR)induces apoptosis-inducing factor(AIF)release from mitochondria and energy deprivation resulting in the caspase-independent death of cancer cells.AIM To investigate whether sustained calcium supply could induce an anticancer effect on pancreatic cancer by PAR accumulation.METHODS Two pancreatic cancer cell lines,AsPC-1 and CFPAC-1 were used for the study.Calcium influx and mitochondrial reactive oxygen species(ROS)were observed by fluorescence staining.Changes in enzyme levels,as well as PAR accumulation and energy metabolism,were measured using assay kits.AIF-dependent cell death was investigated followed by confirming in vivo anticancer effects by sustained calcium administration.RESULTS Mitochondrial ROS levels were elevated with increasing calcium influx into pancreatic cancer cells.Then,excess PAR accumulation,decreased PAR glycohydrolase and ADP-ribosyl hydrolase 3 levels,and energy deprivation were observed.In vitro and in vivo antitumor effects were confirmed to accompany elevated AIF levels.CONCLUSION This study visualized the potential anticancer effects of excessive PAR accumulation by sustained calcium supply on pancreatic cancer,however elucidating a clear mode of action remains a challenge,and it should be accompanied by further studies to assess its potential for clinical application.
基金Supported by the Natural Science Foundation of HubeiProvince (2003ABA163)Specialized Research Fund for the Doctoral Pro-gram of Higher Education (20060486049)
文摘This study is to examine the effect of human recombinant soluble TRAIL （TNF-related apoptosis-inducing ligand） protein inducing apoptosis in MG-63 human osteosarcoma cells. The inhibitive rates of TRAIL to MG-63 cells were detected by MTT assay. The apoptosis induced by TRAIL in MG-63 human osteosarcoma cells was analyzed with FACS and TUNEL and the apoptotic bodies were observed by transmission electron microscope. MTT assay showed that the inhibitive rates of 500, 1 000, 2 000 and 4 000 ng/mL TRAIL for 24 h were 10.1%, 24.3%, 50.6% and 97.7% respectively. Flow cytometric analysis showed that after MG-63 cells were treated with 2 gg/mL TRAIL for 6 h, obvious apoptotic peak would immediately appear before diploid peak. Human soluble TRAIL protein can quickly kill MG-63 osteosarcoma cells selectively, and may have potential value for clinical treatment of osteosarcoma.
