Effect of gambogic acid on proliferation and apoptosis of human diffuse large B-cell lymphoma cell line OCI-LY-1

AIM: To investigate the effects of gambogic acid (GA) on the proliferation and apoptosis of human diffuse large B-cell lymphoma cells. METHODS: Diffuse large B-cell lymphoma cell line OCI-LY-19 was treated with GA. The cell viability and proliferation were evaluated by MTS assay. The morphological changes of the cells were observed under inverted microscope and fluorescence microscope with PI staining. The cell apoptosis was detected by flow cytometry. The changes of proteasome-related proteins Ubs, Bax, HSP90 and IκB-α, the apoptosis-related proteins PARP, pro-caspase-3 and caspase-8, and the proliferation-associated molecules ERK and STAT5 were detected by Western blotting. RESULTS: The proliferation of OCI-LY-19 cells was inhibited and the apoptosis was induced by GA. The expression of Ubs, Bax and HSP90 was up-regulated by GA treatment in a dose-dependent manner. The PARP protein was cleaved. The levels of apoptosis-related proteins pro-caspase-3 and pro-caspase-8, and proliferation-associated molecules p-Akt, p-ERK and p-STAT5 were down-regulated by GA treatment in a dose- and time-dependent manner. CONCLUSION: Gambogic acid down-regulates the activity of proteasome, inhibits cell proliferation and promotes cell apoptosis of diffuse large B cell lymphoma.     

Effects of aspirin on apoptosis of homologous nasopharyngeal carcinoma cells with different radioresistance

AIM: To investigate the effects of aspirin on the apoptosis of nasopharyngeal carcinoma cell lines CNE2R and CNE2 with different radioresistance and its potential mechanism. METHODS: The effects of aspirin on the cell viability, apoptosis, and protein levels of procaspase-3, cleaved caspase-3, procaspase-9, procaspase-12, PARP and cleaved PARP, PI3K p110α, Akt, Bcl-2, Bax and p27 in the CNE2R cells and CNE2 cells were detected by the methods of MTT assay, flow cytometry and Western blot. RESULTS: Aspirin inhibited the viability of homologous nasopharyngeal carcinoma cell lines CNE2 and CNE2R (with the IC₅₀ to CNE2 cells of 6.18, 3.92 and 3.06 mmol/L for 24 h, 48 h and 72 h, respectively; and with the IC₅₀ to CNE2R cells of 7.05, 3.90 and 2.20 mmol/L for 24 h, 48 h and 72 h, respectively). After treated with aspirin for 48 h, the apoptotic rate of CNE2R cells was higher than that of CNE2 cells (P<0.05). After treated with aspirin for 48 h, the protein levels of procaspase-3, procaspase-9, procaspase-12 and PARP were decreased, the protein levels of cleaved caspase-3, cleaved PARP and p27 were increased, and the protein levels of PI3K p110α, Akt and Bcl-2/Bax were decreased. CONCLUSION: Aspirin inhibits the viability of homologous nasopharyngeal carcinoma cell lines CNE2R and CNE2 with different radioresistance. Aspirin also induces the apoptosis of CNE2 and CNE2R cells, which is more effective in CNE2R cells. The underlying mechanisms may be involved in affecting PI3K/Akt signaling pathway, Bcl-2/Bax and p27 expression.     

HBx modulates apoptosis by activating JAK2/STAT3 signaling pathway in human renal proximal tubular epithelial cells

AIM: To investigate the correlation of hepatitis B virus X protein (HBx) with renal tubular epithelial cell apoptosis in hepatitis B virus-associated glomerulonephritis (HBVGN) and the possible signaling mechanism. METHODS: The activation of JAK2/STAT3 signal pathway and the expression of apoptosis-related proteins in human kindey proximal tubular epithelial cells (HK-2 cells) were determined by Western blotting after transfection with HBx eukaryotic expression vector. The cell proliferation was observed by CCK-8 assay. The cell apoptosis was analyzed by the imaging of HO33342 staining, transmission electron microscopy and flow cytometry with Annexin V/PI double staining. RESULTS: After transfection of the target gene HBx, the expression levels of both p-JAK2 and p-STAT3 were significantly increased. At the same time, the cell proliferation was obviously inhibited, and the apoptotic rate was increased. After incubation with AG490, the JAK2/STAT3 signal pathway was partially blocked, and the cell apoptosis induced by HBx was reduced. CONCLUSION：HBx up-regulates the activation of JAK2/STAT3 signal pathway to induce renal tubular epithelial cell apoptosis, which is possibly involved in the pathogenic mechanism that HBV directly damages nephridial tissue.     

Sodium aescinate induces apoptosis of HeLa cells by inhibiting Akt/ERK signaling pathways and increasing death receptor expression

AIM:To study the effects of sodium aescinate on the apoptosis of cervical cancer HeLa cells and its molecular mechanism. METHODS:MTT assay was used to detect the growth and proliferation of HeLa cells. The morphological alteration was observed under inverted microscope. Annexin V-FITC/PI double staining and DAPI nuclear staining were used to determine the apoptosis of HeLa cells induced by sodium aescinate. The apoptosis-related proteins PARP, cleaved caspase-8 and pro-caspase-3, and the proliferation-associated molecules Akt and ERK, as well as TRAIL receptors DR4 and DR5 were detected by Western blotting. RESULTS:Sodium aescinate inhibited the growth of HeLa cells in a concentration-dependent manner. Treatment with sodium aescinate induced the typical morphology of apoptotic cells and increased the apoptotic rate significantly. The cleaved PARP, cleaved caspase-8 and cleaved caspase-9 protein expression was observed. The expression of DR4 and DR5 was up-regulated. Meanwhile, pro-caspase-3 was decreased, and the levels of p-Akt and p-ERK were down-regulated by sodium aescinate in a dose-dependent manner. CONCLUSION:Sodium aescinate inhibits the proliferation and promotes the apoptosis of HeLa cells by increasing death receptor expression and repressing proliferation-associated signaling pathways.     

PI3K/Akt signaling pathway regulates autophagy induced by acute kidney injury in septic rats

AIM：To investigate the autophagy induced by sepsis and acute kidney injury, and the regulation of phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway in this process. METHODS：The rats were subjected to cecal ligation and puncture (CLP) or sham operation. Histopathologic changes of the renal tissues were examined by HE staining. Blood urea nitrogen (BUN) and serum creatinine (SCr) were measured by chemical colorimetry. The protein expression of microtubule-associated protein light chain 3 I/II (LC3 I/II), beclin-1 and p-Akt at different time points after CLP was detected by Western blotting. In vitro, human proximal tubular epithelial cell line HK-2 were treated with LPS to induce autophagy. The protein expression of LC3 I/II and p-Akt in the HK-2 cells after LPS treatment at different time points and different concentrations was detected by Western blotting. These molecules in HK-2 cells and apoptosis of HK-2 cells treated with LPS plus PI3K inhibitor or Akt inhibitor were also detected. RESULTS：Compared with sham group, the severe changes of renal histopathological injuries in CLP groups were observed, the levels of BUN and SCr in CLP groups were significantly increased. LC3 I/II, beclin-1 and phosphorylation of Akt gradually increased after CLP. After treatment with LPS, the expression of p-Akt (308) in the HK-2 cells gradually increased in a dose- and time-dependent fashion. The expression of beclin-1 and p-Akt (472) reached a peak at 8 h or 10 mg/L LPS treatment. Treatment with PI3K or Akt inhibitor down-regulated the expression of LC3 and promoted the apoptosis of HK-2 cells. CONCLUSION：Autophagy in the kidney is induced by sepsis and acute kidney injury. PI3/Akt signaling pathway may be involved in this process.     

PI3K/mTOR dual inhibitor PF-04691502 induces apoptosis of human gastric cancer SGC-7901 cells

AIM:To determine the antitumor effect of PF-04691502, a dual inhibitor of phosphatidylinositol 3-kinase (PI3K)/Akt and mammalian target of rapamycin (mTOR), on the viability and apoptosis of human gastric cancer cell line SGC-7901.METHODS:Cell viability was analyzed by MTT assay. Cell cycle was detected by flow cytometry, and Annexin V-FITC/PI dual staining was used to detect cell apoptosis. Protein expression of p21, cyclin D1, caspase-3, caspase-8, caspase-9 and poly(ADP-ribose) polymerase (PARP) was determined by Western blotting. RESULTS:MTT assay and cell cycle analysis results indicated that PF-04691502 inhibited the viability of SGC-7901 cells in a dose-dependent manner, and arrested the cells in G1 phase. PF-04691502 down-regulated the expression of cyclin D1 and up-regulated the expression of p21. In addition, SGC-7901 cells treated with PF-04691502 showed typical characteristics of apoptosis, accompanied by activation of caspases and cleavage of PARP. CONCLUSION: The PI3K/mTOR dual inhibitor PF-04691502 induces the apoptosis and inhibited the growth of SGC-7901 cells, implicating its potential therapeutic value for the treatment of cancer.     

Pseudolaric acid B induces glioblastoma cell line U87 mitotic arrest and apoptosis

AIM: To investigate the effects of pseudolaric acid B on the growth and apoptosis of glioblastoma cell line U87. METHODS: The cell morphological changes were observed under inverted microscope. The cell viability was evaluated by MTS assay. The cell cycle was analyzed by flow cytometry and Western blot. The cell apoptosis was detected by flow cytometry. The changes of apoptosis-related proteins cleaved PARP, caspase-3, procaspase-9 and caspase-8 were determined by Western blot. RESULTS: Pseudolaric acid B inhibited the viability of U87 cells, arrested U87 cells in mitosis. Apoptosis of U87 cells was induced by pseudolaric acid B. The caspase pathway was activated. CONCLUSION: Pseudolaric acid B induces glioblastoma cell line U87 mitotic arrest and apoptosis.     

Effects of baicalin on CA46 cell proliferation inhibition and apoptosis induction

AIM: To investigate the effects of baicalin on proliferation inhibition and apoptosis induction in human Burkitt lymphoma cell line CA46 and to explore its underlying mechanisms. METHODS: CA46 cells were exposed to baicalin at different dosages and its proliferation inhibition was detected by MTT assay. The ability of baicalin to induce CA46 cell apoptosis was examined by Annexin V-FITC/PI double staining analysis, TUNEL labeling method and DNA fragmentation. The mRNA expressions of c-myc and bcl-2 were detected by RT-PCR, and the protein expressions of c-Myc, Bcl-2, caspase-3 precursor (procaspase-3) and poly ADP-ribose polymerase (PARP) were detected by Western blotting. RESULTS: Baicalin remarkably inhibited the CA46 cell proliferation, with an IC50 value of 10 μmol/L. Apoptosis was remarkably induced by baicalin in a dose-dependent manner, and its earlier and later stages were detected by annexin V-FITC/PI double staining analysis, TUNEL labeling method and DNA fragmentation, respectively. Furthermore, RT-PCR showed that the mRNA expressions of c-myc and bcl-2 in treated CA46 cells decreased in a time-dependent manner. Western blotting showed that the protein expressions of c-Myc, Bcl-2, procaspase-3 and PARP (116 kD) in baicalin treated CA46 cells were down-regulated in a time-dependent manner, while the expression of PARP(85 kD) was up-regulated. CONCLUSION: Baicalin efficiently induces proliferation inhibition and apoptosis in CA46 cells, which may be related with the down-regulation of c-Myc and Bcl-2 expressions, as well as the up-regulation of caspase-3 activity.     

Effects of curcumin analogue T83 on apoptosis of homologous nasopharyngeal carcinoma cells with different radioresistance

AIM：To compare the effect of T83 (a 4-arylidene curcumin analogue) on the apoptosis of homologous nasopharyngeal carcinoma cells with different radioresistance. METHODS：The effects of T83 on the viability, apoptosis, mitochondrial membrane potential (MMP), expression of procaspase-3/procaspase-9/Cyt-C proteins and relative PTEN/Akt/p27 mRNA expression in CNE-2R cells and CNE-2 cells were detected and compared by the methods of MTT assay, Hoechst staining, flow cytometry, Western blotting and qRT-PCR. RESULTS：T83 inhibited the viability of CNE-2R cells with the IC50 of 0.9,0.4 and 0.2 μmol/L for 24 h, 48 h and 72 h,respectively, which was more effective than that inhibiting the viability of CNE-2 cells with the IC50 of 1.8,0.5 and 0.4 μmol/L, respectively. After treated with T83 for 48 h, chromatin condensation, margination and splitting into a massive structure were observed in CNE-2R cells and CNE-2 cells,and the late apoptotic rate of CNE-2R cells was increased from 1.57% to 27.26%, which was higher than that of CNE-2 cells (1.74% to 8.15%). After treated with T83 for 36 h, the MMP in CNE-2R cells decreased by 87.71% and that decreased by 50.47% in CNE-2 cells. After treated with T83 for 48 h, the protein levels of procaspase-3 and procaspase-9 were decreased, and the protein level of Cyt-C was increased, which were more susceptible in CNE-2R cells than those in CNE-2 cells. After treated with T83 for 24 h, the relative mRNA expression of PTEN and p27 was significantly up-regulated, and the mRNA expression of Akt was down-regulated, which were more susceptible in CNE-2R cells than those in CNE-2 cells. CONCLUSION：Compared with CNE-2 cells, the inhibitory effect of T83 on the viability of CNE-2R cells is more specific by starting the mitochondrial apoptotic pathway, which is due to the inhibition of PTEN/Akt/p27 signaling pathway.     

Resveratrol inhibits chondrosarcoma via mitochondrial and PI3K/Akt signaling pathways

AIM：To investigate the inhibitory effects of resveratrol on chondrosarcoma and the relation with mitochondrial and PI3K/Akt pathways. METHODS：Chondrosarcoma SW1353 cells were treated with resveratrol at concentrations of 25, 50 and 100 μmol/L for the time intervals of 24 h, 48 h and 72 h. The viability and apoptosis of the SW1353 cells in the presence or absence of resveratrol were analyzed by CCK8 assay and Hoechst 33258 staining, respectively. The protein levels of Bcl-2, Bax, activated caspase-3, Akt and p-Akt were detected by Western blotting. The cell migration ability was determined by wound scratch assay. RESULTS：Exposure of the cells to resveratrol resulted in a decrease in the cell viability in a dose- and time-dependent manner (P<0.05). visible nuclei with apoptotic characteristics in resveratrol group were observed. The protein levels of activated caspase-3 and Bax were increased, and Bcl-2 and p-Akt were decreased compared with control group. The total Akt were not significantly changed. Resveratrol also significantly reduced the migration of tumor cells. CONCLUSION：Resveratrol induces apoptosis of chondrosarcoma, which plays a role of part through mitochondrial and PI3K/Akt signaling pathways.     

Brucine induces apoptosis in colon cancer SW480 cells via inhibiting IL-6/STAT3 signaling pathway

AIM: To observe the effects of brucine on the viability and apoptosis of colon cancer SW480 cells.METHODS: The SW480 cells were divided into control group, 1 μmol/L brucine treatment group, 100 μg/L IL-6 treatment group and IL-6＋brucine treatment group. The cell viability was detected by CCK-8 assay. The apoptotic rate was measured by flow cytometry using fluorescein-labeled Annexin V/PI. The changes of apoptosis-related proteins were determined by Western blot. The protein level of p-STAT3 was also detected by immunofluorescence staining. RESULTS: Brucine inhibited SW480 cell growth, and the viability inhibition rate of the SW480 cells treated with brucine alone was more efficient than using brucine combined with IL-6 (P < 0.05). The apoptotic SW480 cells increased significantly after 1 μmol/L brucine treatment as compared with brucine treatment alone (P < 0.05). The apoptotic SW480 cells were significantly reduced in brucine and IL-6 combination treatment group (P < 0.05). Brucine inhibited the protein level of p-STAT3 significantly. The protein level of p-STAT3 was significantly increased in 100 μg/L IL-6 treatment group. Compared with 1 μmol/L brucine treatment alone, the expression of Bcl-2 was increased and the protein levels of p-STAT3, Bax and cleaved PARP were reduced in brucine and IL-6 combination treatment group (P < 0.05).CONCLUSION: Brucine may inhibit the activation of STAT3 phosphorylation in IL-6/STAT3 pathway to exert an antitumor effect on SW480 cells in vitro.     

Minocycline suppresses sodium nitroprusside-induced apoptosis in PC12 cells by regulating PI3K/Akt signaling

AIM：To explore the role of PI3K/Akt signaling in the anti-apoptotic effect of minocyline (MC) on the apoptosis of PC12 cells induced by sodium nitroprusside (SNP). METHODS：PC12 cells were divided into 4 groups: blank control group, SNP (500 μmol/L) group, MC (10 μmol/L)+SNP group and LY294002+MC+SNP group. The cell viability was observed by MTT assay. The expression of Akt and p-Akt was determined by Western blotting. RESULTS： The viability of the PC12 cells decreased after exposed to 500 μmol/L SNP for 24 h. Meanwhile, MC at concentration of 10 μmol/L significantly blocked the effect of SNP, such as decreasing the cell viability. Pretreatment with LY294002 for 60 min prior to exposure of the PC12 cells to MC and SNP down-regulated the expression of p-Akt induced by SNP. CONCLUSION：Minocycline regulates PI3K/Akt signaling pathway to restrain the apoptosis of PC12 cells induced by SNP.     

Effects of matrine on apoptosis and related protein expression in human medulloblastoma D341 cells

AIM: To investigate matrine-induced apoptosis of human medulloblastoma D341 cells and the expression of Bax, Bcl-2, serine/threonine kinase Akt and phosphorylated Akt (p-Akt) in vitro. METHODS: D341 cells were divided into experimental groups (added with matrine at different concentrations) and control group (under the same conditions without matrine). The proliferation of D341 cells was analyzed by CCK-8 assay. Apoptosis was detected by Annexin V-FITC/PI double staining and the expression of Bax, Bcl-2, Akt and p-Akt was detected by Western blotting. RESULTS: Matrine significantly inhibited the proliferation of D341 cells and increased the apoptosis in a dose- and time-dependent manner. The cell apoptosis was characterized by chromatin condensation with margination of chromatin to the nuclear membrane, increased when and larger cytoplasmic vacuoles, and formation of apoptotic body after treatment with matrine. The expression of Bax increased, while the expression of Bcl-2 and p-Akt decreased when the drug concentration gradually increased. CONCLUSION: Matrine induces the apoptosis of human medulloblastoma D341 cells in vitro by activation of Bax, down-regulation of Bcl-2 and reduction of p-Akt expression level in the PI3K/Akt signaling pathway.     

MK-2206, an inhibitor of Akt,induced cell apoptosis and autophagy in U2OS cells

AIM：To observe the effect of MK-2206, an inhibitor of Akt, on the cell apoptosis and autophagy of U2OS cells. METHODS：The cell viability was detected by MTT assay. The cell apoptosis was analyzed by TdT-mediated dUTP nick end labeling assay. The expression of LC3-II was examined by Western blotting. RESULTS：MK-2206 inhibited the cell viability in a dose-dependent manner. MK-2206 induced the cell apoptosis via activation of caspase-3, caspase-9 and PARP. MK-2206 treatment substantially induced the U2OS cell autophagy by increasing in the levels of LC3-II. Blockage of autophagy using chloroquine magnified MK-2206-induced cell death in U2OS cells. CONCLUSION：The Akt inhibitor MK-2206 induces cell apoptosis and autophagy. Blocking autophagy magnifies MK-2206-induced the inhibition of the viability in U2OS cells.     

Attenuated Salmonella carrying pGRIM-19-si-survivin co-expression plasmid inhibits growth of prostate cancer subcutaneous xenografts in vivo

AIM：To explore the effects of pGRIM-19-si-survivin co-expression plasmid carried by human attenuated Salmonella on prostate cancer subcutaneous xenograft growth in nude mice. METHODS：Prostate cancer xenograft model was established in nude mice. Co-expression plasmids carried by attenuated Salmonella were introduced by intraperitoneal injection. The xenograft volumes were monitored timely. Immunohistochemical staining, RT-PCR and TUNEL assay were applied to investigate the related mechanisms that pGRIM-19-si-survivin inhibited tumor growth in vivo. RESULTS：Compared with psi-survivin and pGRIM-19 carried by attenuated Salmonella (control groups), the tumor volumes were reduced markedly in pGRIM-19-si-survivin plasmid group. The mean shrinkage rates were 2.36 and 3.02 times. pGRIM-19-si-survivin co-expression plasmid carried by attenuated Salmonella inhibited survivin expression but strengthened GRIM-19 expression obviously (P<0.05). The mRNA expression of apoptosis-related proteins such as Bcl-xL, Stat3, cyclin D1 and c-Myc was inhibited, and the vascular endothelial growth factor (VEGF) mRNA and Ki67 protein were also inhibited, but the caspase-3 mRNA expression was up-regulated (P<0.05) with significant cell apoptosis. CONCLUSION： pGRIM-19-si-survivin co-expression plasmid carried by human attenuated Salmonella inhibits the growth of prostate cancer subcutaneous xenografts in nude mice by promoting cell apoptosis and inhibiting prostatic cancer proliferation.     

Inhibition of c-Myc by 10058-F4 overcomes imatinib resistance in chronic myeloid leukemia cells

AIM：To investigate the effect of c-Myc inhibitor 10058-F4 on human chronic myeloid leukemia (CML) K562 cells and imatinib-resistant K562/G cells. METHODS：The protein expression of c-Myc was detected by Western blotting. Cell proliferation was evaluated by MTT assay and colony formation assay. PI staining was used to determine the cell cycle distribution. Annexin V-PI staining was applied for apoptosis detection. RESULTS：Imatinib-resistant K562/G cells displayed lower sensitivity to imatinib than K562 cells with high expression of c-Myc. Treatment with specific c-Myc inhibitor 10058-F4 inhibited the cell proliferation in a dose- and time-dependent manner, and K562/G displayed more sensitivity to 10058-F4 than K562 cells. 10058-F4 also induced cell cycle arrest in G0/G1 phase and induced apoptotic cell death in the 2 cells. Importantly, 10058-F4 suppressed the colony formation ability in K562 and K562/G cells. CONCLUSION：c-Myc is a novel target to overcome imatinib-induced drug resistance, and c-Myc inhibitor provides a new approach in CML therapy.     

Effect of down-regulation of X-box binding protein 1 gene expression on viability and apoptosis of glioma cells

AIM: To investigate the effect of down-regulation of X-box binding protein 1 (XBP1) expression on the viability and apoptosis of glioma cells. METHODS: The mRNA expression of XBP1 in the glioma tissues was detected by qPCR. Small interfering RNA (siRNA) interfering with XBP1 expression (XBP1-siRNA) was transfected into human brain glioma U251 cells. At the same time, control group (the cells without special treatment) and negative control (NC-siRNA) group (transfected with siRNA without any interference) were set up. The mRNA expression of XBP1 in the 3 groups 48 h after transfection was detected by qPCR. The protein levels of XBP1, proliferating cell nuclear antigen (PCNA), B-cell lymphoma/leukemia-2 (Bcl-2), Bcl-2-associated X protein (Bax), cyclin D1 (cyclin D1), phosphatidylinositol 3-kinase (PI3K) and phosphorylated Akt (p-Akt) were determined by Western blot. The cell viability was measured by CCK-8 assay. The cell cycle distribution and apoptosis were analyzed by flow cytometry. RESULTS: The expression level of XBP1 in the glioma tissues was significantly higher than that in the tumor adjacent tissues (P<0.05). The XBP1 expression at mRNA and protein levels was significantly decreased in the cells transfected with XBP1-siRNA (P<0.05). No statistically significant difference of the cell viability, cell cycle, apoptotic rate and the protein levels of PCNA, Bcl-2, Bax, cyclin D1, PI3K and p-Akt between NC-siRNA group and control group was observed. Compared with control group, the cell viability, S-phase cells and the protein levels of PCNA, Bcl-2, cyclin D1, PI3K, and p-Akt in XBP1-siRNA group were decreased significantly, and the apoptotic rate, G₀/G₁-phase cells and Bax protein expression were significantly increased (P<0.05). CONCLUSION: Down-regulation of XBP1 gene expression in brain glioma cells reduces the viability of cancer cells, blocks the cells in G₁ phase and promote apoptosis. The mechanism is related to the inhibition of PI3K/Akt signaling pathway.     

Evodiamine inhibits growth of Huh7 cells and enhances their sensitivity to TRAIL

AIM: To investigate the effects of evodiamine on the growth and apoptosis of human hepatocellular carcinoma Huh7 cells, and to illustrate the molecular mechanism that evodiamine enhances antitumor activity of tumors necrosis factor-related apoptosis-inducing ligand (TRAIL) in Huh7 cells.METHODS: The cell viability was measured by MTT assay. The cell cycle distribution was analyzed by flow cytometry. The apoptosis rate was determined by TUNEL staining. The protein levels of cell cycle-and apoptosis-related proteins were detected by Western blot analysis.RESULTS: Treatment of Huh7 cells with evodiamine reduced the cell viability (P<0.05). Evodiamine induced cell cycle arrest in G₂/M phase by upregulation of p27, cyclin B1, cell division cycle protein 2 (Cdc2) and p-Cdc2. Evodiamine triggered apoptosis accompanied by cleavage of caspase-3 and poly(ADP-ribose) polymerase (PARP). Combination of evodiamine with TRAIL significantly reduced the cell viability and increased cleavage of caspase-3 and PARP as compared with the use of each agent alone. Moreover, evodiamine increased the expression of death receptor 5 (DR5) in the Huh7 cells.CONCLUSION: Evodiamine inhibits the cell growth by reducing the cell viability and inducing cell cycle arrest. Evodiamine also triggers cell apoptosis and enhances the sensitivity of Huh7 cells to TRAIL by upregulating the expression of DR5.     

Effects of baicalin on CA46 cell xenografts in nude mice

AIM: To study the effects of baicalin on CA46 cell xenografts in nude mice. METHODS: The nude mice with CA46 cell xenografts were treated with drugs via intraperitoneal injection daily, and were divided into 5 groups: negative control group, 15 mg/kg baicalin group, 30 mg/kg baicalin group, 60 mg/kg baicalin group and 4 mg/kg etoposide (VP-16) positive control group. After 12-day treatment, the weight of CA46 cell xenografts stripped from some nude mice in the 5 groups was used to evaluate the effect of baicalin on xenograft growth in the nude mice. The apoptosis, necrosis and pathological changes of the xenograft cells were examined under light microscope and transmission electronic microscope respectively. The expression levels of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway-related proteins extracted from xenografts were determined by Western blotting. The other nude mice with CA46 cell xenografts in the 5 groups continued to be treated with the drugs until death in order to evaluate the effect of balcalin on survival time of the nude mice with CA46 cell xenografts. RESULTS: Baicalin remarkably inhibited the growth of CA46 cell xenografts, induced apoptosis and necrosis of xenograft cells, and reduced the protein expression of phospho-Akt (p-Akt), nuclear factor-kappa B (NF-κB), mammalian target of rapamycin (mTOR) and phospho-mTOR (p-mTOR) in the xenografts after 12-day treatment. Furthermore, baicalin prolonged the survival time of the nude mice with CA46 cell xenografts in a dose-dependent manner. CONCLUSION: Baicalin inhibits the growth and induces apoptosis of CA46 cell xenografts in the nude mice, and prolongs the survival time of the nude mice with CA46 cell xenografts through the mechanism of down-regulating PI3K/Akt/NF-κB and PI3K/Akt/ mTOR signaling pathways.     

Cholestane-3β, 5α, 6β-triol induces apoptosis of malignant glioma cells

AIM：To investigate the effect of cholestane-3β, 5α, 6β-triol (Triol) on apoptosis of malignant glioma cells. METHODS：C6 cells and A172 cells were incubated with Triol at different concentrations for different time durations. MTT assay was used to detect the cell viability. Hoechst 3f3342 staining and TUNEL assay were used to analyze the cell apoptosis. The caspase activity was measured. The expression of apoptosis-related proteins, Bcl-2 family members, was determined by Western blotting. RESULTS：Triol decreased the cell viability of C6 and A172 cells in a dose- and time-dependent manner and the IC50 values were (17.8±0.6)μmol/L and (20.6±0.2) μmol/L, respectively. Visible nuclei with apoptotic characteristics, significant increase in TUNEL-positive cells, and the activation of apoptotic execution enzyme caspase-3 indicated that cell apoptosis was induced by Triol in both cell lines. After C6 cells were exposed to Triol for 12 h, 24 h and 48 h, the activity of caspase-8 in extrinsic apoptotic pathway and caspase-9 in intrinsic apoptotic pathway was increased time-dependently. Meanwhile, the levels of anti-apoptotic proteins, Bcl-2 and Bcl-xL, was down-regulated, while pro-apoptotic protein Bak was up-regulated in a time-dependent manner. CONCLUSION：Triol induces apoptosis of malignant glioma cells by activating intrinsic and extrinsic apoptotic pathways, and Bcl-2 family members are involved in Triol-induced apoptosis.