Acute lymphoblastic leukemia (ALL) is the most common malignant hematological disease in childhood. Glucocorticoids are frequently used in the chemoradiotherapy regimen for ALL and can induce the apoptosis of ALL cells through several signaling pathways, but about 10% of ALL children have poor response to glucocorticoids. Studies have revealed that glucocorticoids induce the apoptosis of ALL cells by upregulating the expression of BIM gene, and BIM gene is associated with glucocorticoid resistance in childhood ALL. This article reviews the recent studies on glucocorticoid resistance in childhood ALL, especially the role of BIM and its expression products in this process.
Apoptosis ; Bcl-2-Like Protein 11 ; genetics ; Child ; Drug Resistance ; Glucocorticoids ; therapeutic use ; Humans ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; drug therapy ; genetics ; pathology

OBJECTIVETo investigate the changes in SiHa cell apoptosis after inhibition of CD147 expression.

METHODSRNA interference (RNAi) technique was used to down-regulate CD147 expression in SiHa cells, and RT-PCR and Western blotting were used to detect expression of CD147, Bcl-2, Bim and caspase-3; the percentage of cell apoptosis were detected by flow cytometry.

RESULTSSiRNA sequence 1, 2 inhibited CD147 expression in SiHa cells effectively (P<0.05), resulting also in down-regulated expression of Bcl-2 (P<0.05) and up-regulated expression of caspase-3 and Bim(P<0.05). The percentage of apoptotic cells increased significantly, and early apoptosis was the most obvious in the cells (P<0.05).

CONCLUSIONSilencing of CD147 expression induces SiHa cell apoptosis partially through the Bcl-2 pathway .

Apoptosis ; Apoptosis Regulatory Proteins ; metabolism ; Basigin ; metabolism ; Bcl-2-Like Protein 11 ; Caspase 3 ; metabolism ; Cell Line, Tumor ; Down-Regulation ; Humans ; Membrane Proteins ; metabolism ; Proto-Oncogene Proteins ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; RNA Interference

OBJECTIVETo establish osteoblast apoptosis model induced by gingipains, and to examine the expression of pro-apoptotic protein Bcl-2 interacting mediator (Bim), Bcl-2 associated X protein (Bax) and Bcl-2 antagonist/killer (Bak).

METHODSGingipain and gingipain acticity were extracted and measured. Mouse osteoblast cell line MC3T3-E1 cells were cultured in the presence of 0.453, 0.906, 1.812 U/L gingipains for 0, 16, 24 and 48 h. Apoptosis was examined by 4',6-diamidino-2-phenylindole (DAPI) staining or annexin V/propidine iodide (PI) staining.Protein expression of Bim, Bax and Bak was determined by Western blotting after osteoblasts were cultured with 1.812 U/L gingipain for 0, 4, 8, 16, 24 and 48 h. Osteoblasts were cultured with 1.812 U/L gingipain which had been inhibited with N-alpha-tosyl L-lysyl-chlorom ethylketone (TLCK). Western blotting was used to detect Bim expression and DAPI staining to measure apoptosis.

RESULTSArginine-specific proteinases (Rgp) activity was (18.11 ± 2.11) U/L and specific proteinases (Kgp) was (1.02 ± 0.25) U/L. Percentage of osteoblast apoptosis induced by 1.812 U/L gingipain rose to (6.31 ± 0.37)% after 16 h, and reached (11.20 ± 0.35)% at 24 h and (10.80 ± 0.46)% after 48 h with DAPI staining. Annexin V/PI staining supported the result from DAPI staining.Bim protein level increased during osteoblast apoptosis, the relative fold rose to (0.31 ± 0.03) after 4 h (about 2 fold compared to control), peaking at 24 h (0.57 ± 0.05, 3-4 fold compared to control). Proteinase inhibitor TLCK effectively blocked the activity of gingipain and inhibited up-regulation of Bim induced by gingipains from (0.58 ± 0.04) to (0.14 ± 0.03). The percentage of osteoblast apoptosis decreased from (11.20 ± 0.35)% to (4.31 ± 0.38)% in the presence of TLCK. Expression of Bax remained unchanged when cells were cultured with or without gingipains. Bak was under the detectable level in MC3T3-E1.

CONCLUSIONS1.812 U/L gingipains induced osteoblast apoptosis. Protein expression of Bim was up-regulated during cell apoptosis and was down-regulated when gingipain inhibited with TLCK, suggesting that Bim was involved in osteoblast apoptosis induced by gingipain. Inhibition of Bim protein expression protected osteoblast from apoptosis.

Adhesins, Bacterial ; pharmacology ; Animals ; Apoptosis ; drug effects ; Apoptosis Regulatory Proteins ; metabolism ; Bcl-2-Like Protein 11 ; Cell Line ; Cysteine Endopeptidases ; pharmacology ; Humans ; MCF-7 Cells ; Membrane Proteins ; metabolism ; Mice ; Osteoblasts ; cytology ; metabolism ; Proto-Oncogene Proteins ; metabolism ; Tosyllysine Chloromethyl Ketone ; pharmacology ; bcl-2 Homologous Antagonist-Killer Protein ; metabolism ; bcl-2-Associated X Protein ; metabolism

Although the anti-malaria drug chloroquine (CQ) has been shown to enhance chemotherapy and radiation sensitivity in clinical trials, the potential mechanisms underlying this enhancement are still unclear. Here, we examined the relevant mechanisms by which the multipotent CQ enhanced the cytotoxicity of topotecan (TPT). The lung cancer cell line A549 was treated with TPT alone or TPT combined with CQ at non-cytotoxic concentrations. Cell viability was assessed using the MTT assay. The percentage of apoptotic cells and the presence of a side population of cells were both determined by flow cytometry. Autophagy and the expression of Bcl-2 family proteins were examined by Western blotting. The accumulation of YFP-LC3 dots and the formation of acidic vesicular organelles were examined by confocal microscopy. CQ sensitized A549 cells to TPT and enhanced TPT-induced apoptosis in a Bcl-2 family protein-independent fashion. CQ inhibited TPT-induced autophagy, which modified the cytotoxicity of TPT. However, CQ failed to modify the transfer of TPT across the cytoplasmic membrane and did not increase lysosomal permeability. This study showed that CQ at non-cytotoxic concentrations potentiated the cytotoxicity of TPT by interfering with autophagy, implying that CQ has significant potential as a chemotherapeutic enhancer.
Apoptosis ; drug effects ; Apoptosis Regulatory Proteins ; metabolism ; Autophagy ; drug effects ; Bcl-2-Like Protein 11 ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Chloroquine ; pharmacology ; Drug Synergism ; Humans ; Lung Neoplasms ; metabolism ; pathology ; Membrane Proteins ; metabolism ; Proto-Oncogene Proteins ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Topoisomerase I Inhibitors ; pharmacology ; Topotecan ; pharmacology ; bcl-2-Associated X Protein ; metabolism

OBJECTIVETo study p53 up-regulated modulator of apoptosis (PUMA) and bcl-2 interacting mediator of cell death (BIM) of the BH3-only protein family expression in colorectal cancer tissues and its relationship with colorectal cancer invasion, metastasis and prognosis.

METHODSImmunohistochemical staining (EnVision) was used to detect PUMA/BIM expression in 30 cases of normal mucosa, 30 cases of colorectal adenoma and 142 cases of colorectal cancer tissues.

RESULTSPUMA in colorectal cancer tissues was positive expressed (82.4%), which was significantly lower than in the normal mucosa colorectal adenomas (96.7%) and normal mucosa tissues (96.7%) (both χ(2) = 3.93, P < 0.05). Positive expression rate of BIM in colorectal cancer tissues (62.7%) was significantly lower than that in colorectal adenomas and normal mucosa (96.7% and 90.0%) (χ(2) = 8.42 and 13.29, P < 0.01). PUMA and BIM in colorectal cancer tissues were positively correlated (r = 0.747, P = 0.000). PUMA expression was related to tumor differentiation (χ(2) = 11.87), invasion depth (χ(2) = 11.59), lymph node metastasis (χ(2) = 12.82), TNM stage (χ(2) = 33.47) and P-gp expression (χ(2) = 18.30), all P < 0.05, but not related to the patients' age, gender, tumor size, tumor histological type and GST-π expression (P > 0.05). BIM expression was related to tumor differentiation (χ(2) = 16.19), lymph node metastasis (χ(2) = 14.95), TNM stage (χ(2) = 52.66) and P-gp expression (χ(2) = 10.60) (P < 0.05), but not related to patients' age, sex, tumor size, tumor histological type, invasion depth and GST-π expression (P > 0.05). 1-, 3-, 5-year survival rates of the positive expression of PUMA/BIM in patients with colorectal cancer were significantly higher than that of PUMA/BIM in patients with negative expression (χ(2) = 6.10 and 27.6, P < 0.05). Cox multivariate analysis showed that lymph node metastasis (RR = 0.238), TNM stage (RR = 7.895), PUMA (RR = 1.691) and BIM (RR = 0.440) could be used as independent prognostic indicators (P < 0.05).

CONCLUSIONSPUMA and BIM expressions in colorectal cancer are related to the tumor invasion, metastasis and prognosis. Low expressions of PUMA and BIM were related to the late period and poor prognosis of colorectal cancer patients.

Adult ; Aged ; Aged, 80 and over ; Apoptosis Regulatory Proteins ; metabolism ; Bcl-2-Like Protein 11 ; Biomarkers, Tumor ; metabolism ; Colorectal Neoplasms ; metabolism ; pathology ; Female ; Humans ; Lymphatic Metastasis ; Male ; Membrane Proteins ; metabolism ; Middle Aged ; Neoplasm Invasiveness ; Prognosis ; Proto-Oncogene Proteins ; metabolism ; Survival Rate

OBJECTIVETo transfer pro-apoptotic BIM directly into tumor cells bypass the complicated biological processes of BIM activation so as to reverse the chemoresistance of cancer cells.

METHODSBIMS was specifically amplified from HL-60 cells by RT-PCR, confirmed to be correct by sequencing and cloned into shuttle vector pAdTrack-CMV carrying a green fluorescence protein gene to generate a recombinant plasmid pAdTrack-CMV-BIMS. This plasmid and adenovirus backbone plasmid pAdEasy-1 were linearized and electroporated into E.coli BJ5183 host bacteria to mediate homologous recombination. The positive clone was identified by restrict endonuclease digestion. The recombinant pAdEasy-CMV-BIMS was transferred into HEK293 cells for packaging and amplification. The successful construction of recombinant human BIMS adenovirus (Ad-BIMS) was demonstrated by Western blot. To test whether Ad-BIMS has the capability of inducing apoptosis of tumor cells, Ad-BIMS was used to infect GC resistant Burkitt lymphoma Raji cells.

RESULTSAfter infected for 2-5 days, BIMS expression in Raji cells was detected by RT-PCR and Western blot. The significant growth retardation and apoptosis of Raji cells were also observed by MTT and flow cytometry.

CONCLUSIONThese results indicated that BIMS might be a potential candidate of gene therapy for chemoresistant tumor cells.

Adenoviridae ; Apoptosis ; Apoptosis Regulatory Proteins ; genetics ; Bcl-2-Like Protein 11 ; Burkitt Lymphoma ; therapy ; Genetic Therapy ; Genetic Vectors ; HEK293 Cells ; HL-60 Cells ; Humans ; Membrane Proteins ; genetics ; Proto-Oncogene Proteins ; genetics

OBJECTIVETo evaluate the effect of Foxo3a gene over-expression on the development of rat ovarian granulosa cells and in prevention of cisplatin-induced ovarian damage in rats.

METHODSRat ovarian granulose cells released mechanically from the ovaries were cultured in vitro and identified with HE staining and immunohistochemical staining for FSHR. A recombinant adenovirus carrying Foxo3a gene was constructed for infecting the granulose cells, and the cell growth and expressions of cyclin D1, p27, Bax, and Bim were detected; the cell apoptosis and cell cycle changes were detected using Hoechst/PI 33342 staining and flow cytometry, respectively. The transfected cells were challenged with cisplatin and the cell apoptosis was detected with flow cytometry.

RESULTSOver 90% of the cultured cells survived and contained more than 95% ovarian granulose cells. Infection of the cells with the recombinant adenovirus resulted in over-expressions of Foxo3a at the mRNA and protein levels at 36 h and 48 h after the infection, respectively. The infected cells showed suppressed proliferation, increased apoptotic rate and cell cycle arrest in G1 phase with increased expressions of Bim, p27, and cyclin D1 but without significant changes in Bax expression. Cisplatin exposure caused a significantly higher apoptosis rate in the infected cells than in the control cells.

CONCLUSIONOver-expression of Foxo3a gene can promote granulose cell apoptosis by increasing Bim expression and cause cell cycle arrest in G1 phase by increasing cyclin D1 and p27 expressions, but can not prevent the toxic effects of cisplatin on ovarian granulosa cells.

Animals ; Apoptosis ; Apoptosis Regulatory Proteins ; metabolism ; Bcl-2-Like Protein 11 ; Cell Cycle Checkpoints ; Cell Proliferation ; Cells, Cultured ; Cisplatin ; adverse effects ; Cyclin D1 ; metabolism ; Cyclin-Dependent Kinase Inhibitor p27 ; metabolism ; Female ; Forkhead Box Protein O3 ; Forkhead Transcription Factors ; genetics ; metabolism ; Gene Expression ; Granulosa Cells ; cytology ; drug effects ; Membrane Proteins ; metabolism ; Proto-Oncogene Proteins ; metabolism ; Rats ; Transfection ; bcl-2-Associated X Protein ; metabolism

OBJECTIVETo study the role of BH3-only gene in oxaliplatin-induced apoptosis of human colon cancer cell line, and to explore the associated mechanisms.

METHODSTwo strains of human colon cancer cell line SW480 and HT29 were selected, and treated respectively with different concentrations of oxaliplatin (0.3, 0.6, 1.25, 2.5, 5, 10 and 20 mg/L). Cell growth and inhibition were detected by MTT method. Apoptosis was measured by flow cytometry. Bim and PUMA expressions were examined by fluorescence quantitative PCR.

RESULTSAfter treatment of different oxaliplatin concentrations in human colon carcinoma cells SW480 line, the cell growth was inhibited in a dose-dependent manner, while Bim and PUMA expressions were significantly up-regulated. While HT29 cell lines received the same treatment, no obvious inhibition of cell growth and up-regulation of Bim and PUMA expression were found. When SW480 cells were exposed to 5 mg/L and 10 mg/L of oxaliplatin for 24 h, the early apoptotic rates were (4.87±0.55)% and (12.10±1.04)%; for 48 h, the early apoptotic rates were (11.47±0.85)% and (30.07±2.01)%; for 72 h, the early apoptotic rates were (28.99±2.12)% and (38.32±3.15)% respectively, which were all significantly higher than those in control group [(0.30±0.10)%, (0.40±0.10)% and (0.50±0.20)%, all P<0.01]. In HT29 cells, the differences of apoptotic rates between oxaliplatin treatment group and control group were not statistically significant (all P>0.05).

CONCLUSIONSOxaliplatin can inhibit colon cancer cell line SW480 growth and induce apoptosis. Induction of apoptosis of colon cancer cells by oxaliplatin may be associated with the up-regulation of BH3-only proteins, Bim and PUMA.

Apoptosis ; drug effects ; Apoptosis Regulatory Proteins ; metabolism ; Bcl-2-Like Protein 11 ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Colonic Neoplasms ; pathology ; Humans ; Membrane Proteins ; metabolism ; Organoplatinum Compounds ; pharmacology ; Proto-Oncogene Proteins ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism

This study was purposed to investigate the effect of bortezomib (Bor) and arsenic trioxide (As(2)O(3)) combination on multiple myeloma cell line KM3 and its mechanisms. KM3 cells were cultured with different concentration of Bor or As(2)O(3) as well as both for a certain time. The cell proliferation was analysed by MTT assay and the concentration of 50% proliferation inhibition (IC(50)) was calculated. Early apoptosis and late apoptosis of KM3 cells were detected by Annexin-V-FITC Kit, and the change of transmembrane potential was measured by flow cytometry. mRNA of Caspase-3, Bim and Bcl-xL were detected by RT-PCR. The results showed that the proliferation inhibitory rate of KM3 cells treated by Bor plus As(2)O(3) was much higher than that of KM3 cells treated by Bor only for 72 h [ (27.64 ± 0.81)% vs (21.67 ± 2.20)%, P < 0.05]. There were more KM3 cells treated by Bor plus As(2)O(3) in early apoptosis at 48 h and late apoptosis at 72 h than that of KM3 cells treated only by Bor [ (53.20 ± 3.70)% vs (35.40 ± 2.58)%, P < 0.01; (63.96 ± 2.97)% vs (54.08 ± 3.76)%, P < 0.01]. Transmembrane potential (Δψm) of KM3 cells treated by Bor plus As(2)O(3) decreased more at 48 h, as compared with Bor alone. The expression levels of caspase-3 mRNA and Bim mRNA in KM3 cells treated with Bor plus As(2)O(3) were higher than that in KM3 cells treated with Bor alone. But the expression level of Bcl-xL mRNA was lower than that in KM3 cells treated with Bor alone. It is concluded that As(2)O(3) can enhance the apoptosis-inducing effect of Bor on multiple myeloma cell line KM3, which is associated with decreasing the expression of Bcl-xl mRNA and increasing the expression of Caspase-3 and Bim mRNA.
Apoptosis ; drug effects ; Apoptosis Regulatory Proteins ; metabolism ; Arsenicals ; administration & dosage ; pharmacology ; Bcl-2-Like Protein 11 ; Boronic Acids ; administration & dosage ; pharmacology ; Bortezomib ; Caspase 3 ; metabolism ; Cell Line, Tumor ; Humans ; Membrane Proteins ; metabolism ; Multiple Myeloma ; metabolism ; pathology ; Oxides ; administration & dosage ; pharmacology ; Proto-Oncogene Proteins ; metabolism ; Pyrazines ; administration & dosage ; pharmacology ; bcl-X Protein ; metabolism

The present study was to investigate whether Toll-like receptor 4 (TLR4)-mediated Akt/FoxO3a/Bim signaling pathway participated in lipopolysaccharide (LPS)-induced apoptosis in hippocampal neurons. The primarily cultured rat hippocampal neurons were treated with LPS, TLR4 antibody+LPS, and LY294002+LPS, respectively. Cell vitality was assayed by CCK-8. Expressions of p-Akt, Akt, p-FoxO3a, FoxO3a, Bim and active-Caspase-3 of each group were detected by Western blot analysis; the mRNA expression of Bim was detected by real-time quantitative PCR; FoxO3a nuclear translocation was detected by fluorescence microscope. The rate of cell apoptosis was assayed by flow cytometry. The results showed that cell vitality of hippocampal neurons decreased after being treated with LPS in a time-dependent way. Compared with the control group, the expressions of p-Akt and p-FoxO3a decreased significantly, FoxO3a translocated into the nucleus, meanwhile, the expression of Bim and active-Caspase-3, and the apoptotic ratio of hippocampal neurons increased in LPS treated neurons. Pretreatment with TLR4 antibody significantly blocked, while PI3K antagonist LY294002 further strengthened these changes induced by LPS. In conclusion, the present study suggests that Akt/FoxO3a/Bim signaling pathways mediated by TLR4 participate in the apoptotic processes of primarily cultured hippocampal neurons treated with LPS, and the activation of TLR4 causes neuronal apoptosis.
Animals ; Apoptosis ; Apoptosis Regulatory Proteins ; metabolism ; Bcl-2-Like Protein 11 ; Caspase 3 ; metabolism ; Chromones ; pharmacology ; Forkhead Box Protein O3 ; Forkhead Transcription Factors ; metabolism ; Hippocampus ; cytology ; Lipopolysaccharides ; Membrane Proteins ; metabolism ; Morpholines ; pharmacology ; Neurons ; cytology ; metabolism ; Phosphatidylinositol 3-Kinases ; antagonists & inhibitors ; Proto-Oncogene Proteins ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Rats ; Signal Transduction ; Toll-Like Receptor 4 ; metabolism