Curcumin Induces Cycle Arrest of Colon Cancer HCT116 Cells via JAK1/STAT1/p21 Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effect of curcumin on the cycle arrest of human colon cancer HCT116 cells and decipher the possible molecular mechanism. MethodThe methyl thiazolyl tetrazolium （MTT） method was employed to examine the effects of curcumin （0， 12.5， 25， 50， 75， 100 μmol·L-1） and 5-fluorouracil （5-FU， 600 μmol·L-1） on the proliferation of HCT116 cells at different time points （24， 48， 72 h）. Flow cytometry was employed to examine the cycle of HCT116 cells treated with curcumin （0， 25， 50， 75 μmol·L-1） and 5-FU. Western blot was employed to determine the expression of proteins in the Janus kinase 1 （JAK1）/signal transducer and activator of transcription 1 （STAT1） /cyclin-dependent kinase inhibitor 1A （p21） pathway in HCT116 cells. The binding of STAT1 to p21 promoter region was detected by chromatin immunoprecipitation （ChIP）. Small interfering RNA （siRNA） was employed to measure the role of STAT1 in regulating the expression of p21 and that of JAK1 in regulating the activation of STAT1 by Western blot and cellular immunofluorescence， respectively. ResultCompared with the blank group， the HCT-116 cells treated with curcumin and 5-FU showed decreased viability （P<0.05）， increased proportions of cells in the G0/G1 phase （P<0.05）， decreased proportions of cells in the S phase and G2/M phase （P<0.05）， down-regulated protein level of phosphorylated p21 （P<0.05）， and up-regulated protein level of p21 （P<0.05）. Compared with the curcumin group， the p21 siRNA+ curcumin group presented decreased proportion of cells in G0/G1 phase （P<0.05）. Compared with the blank group， curcumin elevated the level of phosphorylated STAT1 （p-STAT1） （P<0.05）. Compared with the curcumin group， the curcumin + STAT1 siRNA group showcased up-regulated protein level of p21 in HCT116 cells （P<0.05）. The mechanism study showed that curcumin treatment enhanced the enrichment of STAT1 in the p21 promoter region （P<0.05） compared with the blank group. Compared with the blank group， curcumin up-regulated the level of phosphorylated JAK1 （p-JAK1） （P <0.05）. Compared with the curcumin group， the curcumin + STAT1 siRNA group demonstrated up-regulated protein levels of p-STAT1 and p21 in HCT116 cells （P<0.05）. ConclusionCurcumin may induce the cycle arrest of human colon cancer HCT116 cells by activating the JAK1/STAT1/p21 signaling pathway.

Effect of astragalin on apoptosis of human colon cancer ISHIKAWA cells / 中华内分泌外科杂志

Objective:To investigate the apoptosis-inducing effect of baohuoside I (BI) on endometrial cancer Ishikawa cells and its related molecular mechanism.Methods:With 0 μ M and 0 h treatment were used as blank control group, and BI treatment was used as experimental group. The inhibitory effect of BI on the proliferation of Ishikawa cells was detected by CCK-8 assay. The apoptosis-inducing effect of BI on Ishikawa cells and the changes of mitochondrial membrane potential were detected by flow cytometry. The expressions of apoptosis-related proteins and signaling pathway-related proteins were detected by Western blot.Results:CCK-8 experiment showed that BI could be expressed in concentration gradient (3, 10, 20, 30, 40 μM). It could effectively inhibit the proliferation of Ishikawa cells (the survival rates were 89.56±0.96, 74.69±1.21, 60.28±1.09 and 43.51±2.17 respectively). Its toxic and side effects on normal cells were lower than that of 5-FU. The results of flow cytometry showed that BI could effectively induce the apoptosis of Ishikawa cells by reducing the level of mitochondrial membrane potential. The proportion of apoptotic cells in each group was (9.92±0.77) %, (14.01±0.83) %, (17.05±1.41) %, (28.21±1.73) % and (44.55±3.11) %. Western blot showed that BI could up-regulate the level of p-p38 and reduce the level of p-STAT3.Conclusions:BI can effectively inhibit the proliferation of Ishikawa cells, and induce apoptosis by reducing the mitochondrial membrane potential and activating the mitochondria-dependent pathway. Its regulatory mechanism is achieved by activating the p38 signaling pathway and inhibiting the STAT3 pathway.

Effects of Shikonin on Autophagy and Apoptosis of Human Colon Cancer HCT 116 Cells / 中国药房

OBJECTIVE：To study the effects of shikonin on autophagy and apoptosis of human colon cancer HCT 116 cells. METHODS：After treating HCT 116 cells for 48 h with shikonin at 0（blank control ）10，20，40 μg/mL，MTT method was used to detect inhibitory rate of cell proliferation. Flow cytometry was used to detect cell apoptosis rate. RT-qPCR assay and Western blotting were respectively used to detect mRNA and protein expressions of microtubule associated protein light chain 3（LC3）and autophagy-related protein Beclin- 1 and p 62. RESULTS ：Compared with blank control ，after treated with 10，20，40 μ g/mL shikonin for 48 h，proliferation inhibitory rate and apoptosis rate of HCT 116 cells were increased significantly （P＜0.05 or P＜ 0.01）. After treated with 10，20，40 μg/mL shikonin for 48 h，mRNA and protein expressions of LC 3，Beclin-1 and p 62 in HCT116 cells were increased to different extents ；except that mRNA expression of LC 3 was not increased significantly after treated with 10 μg/mL shikonin，the difference were statistically significant in other indexes ，compared with blank control （P＜0.05 or P＜0.01）. CONCLUSIONS ：Shikonin can induce the apoptosis of human colon cancer HCT 116 cells and activate its autophagy pathway.

Human Colon Cancer Derived Mesenchymal Stem Cells Promote the Tumorigenicity of Colon Cancer by Secreting Exosomes / 中国药学杂志

OBJECTIVE: To explore the effect of exosome from human colon cancer derived mesenchymal stem cells on the biological function of colon cancer cells. METHODS: Human colon cancer mesenchymal stem cells (hCC-MSCs) were induced from human colon cancer stem cells, and verified by flowcytometry, osteogenesis and adipogenesis differentiation. The exosomes were purified by ExoQuick Exosome Precipitation kit and characterized by electron microscopy, nanoparticle tracking analysis and Western blot. PKH67-labeled hCC-MSCs-Exo was co-cultured with HCT-116 and HT-29. The effects of hCC-MSCs-Exo on the biological function of colon cancer cell lines were investigated by CCK8 assay, wound healing assay and Transwell invasion assay. RESULTS: The hCC-MSCs showed monolayer adherent growth, fibrocyte-like with positive expression of CD29, CD90, CD105 and negative expression of CD34, CD45, HLA-DR, which have the abilities of osteoblasts and adipocytes. hCC-MSCs-Exo has an average diameter of 135 nanometers and positive expression of Alix, CD63 and CD81. After co-culture with colon cancer cell lines, hCC-MSCs-Exo was uptaken into the colon cancer cells. CCK8, wound healing and Transwell assays showed that the proliferation, migration and invasion ability of co-culture group were significantly enhanced compared with control group (P<0.05). CONCLUSION: hCC-MSCs-Exo can promote the proliferation, migration and invasion of colon cancer cells. It is expected to be a new target for colon cancer treatment.

Mechanism of ginsenoside CK inhibiting proliferation of human colon cancer SW480 cells / 中草药

Objective: To study the effects of ginsenoside CK on proliferation and apoptosis of human colon cancer cell line SW480, and further explore the mechanism. Methods: Cell viability was measured by CCK-8 assay. Cell cycle, apoptosis, reactive oxygen species (ROS) levels and changes in mitochondrial membrane potential were measured by flow cytometry. Hoechst staining further detected apoptosis. Western blotting was used to detect the release of cytochrome C and the expression of apoptosis-related proteins such as Bcl-2, Bax and cleaved Caspase-3. Results: Ginsenoside CK had a significant inhibitory effect on the proliferation of human colon cancer cell line SW480. Ginsenoside CK induced SW480 cells arrest in G0/G1 phase, promoted early apoptotic cells, significantly increased intracellular ROS levels and reduced the MMP level. Ginsenoside CK promoted the expression of Bax and cleaved-Caspase-3 and inhibited the expression of Bcl-2. In addition, ginsenoside CK released a large amount of cytochrome C in SW480 cells. Conclusion: Ginsenoside CK has a significant inhibitory effect on the proliferation of human colon cancer cell line SW480. The mechanism may be through the promotion of mitochondrial superoxide elevation, resulting in a significant increase in intracellular ROS levels and a significant decrease in MMP level, further leading to the release of cytochrome C, the up-regulated expression of Bax, the down-regulated expression of Bcl-2, and ultimately leading to apoptosis of cells.

Shikonin Exerts Cytotoxic Effects in Human Colon Cancers by Inducing Apoptotic Cell Death via the Endoplasmic Reticulum and Mitochondria-Mediated Pathways

The apoptotic effects of shikonin (5,8-dihydroxy-2-[(1R)-1-hydroxy-4-methylpent-3-enyl]naphthalene-1,4-dione) on the human colon cancer cell line SNU-407 were investigated in this study. Shikonin showed dose-dependent cytotoxic activity against SNU-407 cells, with an estimated IC50 value of 3 µM after 48 h of treatment. Shikonin induced apoptosis, as evidenced by apoptotic body formation, sub-G1 phase cells, and DNA fragmentation. Shikonin induced apoptotic cell death by activating mitogen-activated protein kinase family members, and the apoptotic process was mediated by the activation of endoplasmic reticulum (ER) stress, leading to activation of the PERK/elF2α/CHOP apoptotic pathway, and mitochondrial Ca2+ accumulation. Shikonin increased mitochondrial membrane depolarization and altered the levels of apoptosis-related proteins, with a decrease in B cell lymphoma (Bcl)-2 and an increase in Bcl-2-associated X protein, and subsequently, increased expression of cleaved forms of caspase-9 and -3. Taken together, we suggest that these mechanisms, including MAPK signaling and the ER-and mitochondria-mediated pathways, may underlie shikonin-induced apoptosis related to its anticancer effect.

Study on Synthesis and Anti-tumor Activity of 6 Kinds of Tectorigenin Mannich Base Derivatives / 中国药房

OBJECTIVE： To conduct structural modification of tectorigenin to search for new compounds with anti-tumor activity. METHODS： Tectorigenin was used as a lead compound， and then added into amine reagents as ethanolamine， methylamine， ethylamine， dimethylamine， diethylamine， n-propylamine and formaldehyde solution. Tectorigenin Mannich base derivatives were synthesized by mannich reaction with as the lead compound. The structures of the derivatives were identified according to IR， UV， MS and NMR data. Solubility of tectorigenin and its derivatives were investigated by solubility test method. MTT assay was used to investigate the inhibitory effects of tectorigenin and its derivatives on the proliferation of human colon cancer cell line HCT116， human lung cancer cell line A549 and human hepatoma cell line HepG2， and half inhibitory concentration （IC50） was calculated. The inhibition rate of tectorigenin and its derivatives （100 mg/kg） on H22 hepatoma-bearing mice in vivo was studied. RESULTS： Totally of 6 kinds of tectorigenin mannich base derivatives were synthesized， such as 8-（N-hydroxyethyl）-methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N-methyl）-methyleneamino-5，7，4′-trihydroxy-6- methoxyisoflavone， 8-（N， N-diethyl）-methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N， N-dimethyl）-methyleneamino- 5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N-ethyl）-methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N-propyl）- methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone （compounds 1-6 in turn）. Compared with tectorigenin， the water solubility of six derivatives was significantly improved， and the solubility was 5-20 times higher than that of tectorigenin. IC50 of compounds 1， 3 and 5 to HCT116 cells were （34.82±3.27）， （16.21±4.13）， （33.12±3.25） μmol/L， which were stronger than that of tectorigenin [（45.23±5.74） μmol/L]； IC50 of compounds 1， 3 and 5 to A549 cells were （37.05±5.74）， （26.88±4.52）， （30.13±6.23） μmol/L， which were stronger than that of tectorigenin [（53.24±6.34） μmol/L]； IC50 of compounds 1， 3 and 5 to HepG2 cells were （23.74±1.45）， （18.96±2.34）， （30.95±2.87） μmol/L， which were stronger than that of tectorigenin [（48.98±2.58） μmol/L]. Compounds 1， 3 and 5 showed higher inhibition rates （55.51％， 57.20％ and 49.15％） than tectorigenin （33.05％） on H22 hepatoma-bearing mice， respectively. The other three compounds had no obvious advantage over tectorigenin in anti-tumor activity. CONCLUSIONS： In this study， compounds 1， 3 and 5 of six tectorigenin mannich base derivatives synthesized in this study have stronger antitumor activity than tectorigenin.

Induction Study of Laminarin Sulfate LAMS-2 on Human Colon Cancer Cell Line LOVO Apoptosis through a Mitochondrial Pathway / 中国药学杂志

OBJECTIVE: To investigate the effects and mechanisms of LAMS-2 on the proliferation and apoptosis of human colon cancer cell line LOVO. METHODS: LOVO cells were treated with different concentrations of LAMS-2,3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay and morphology observation were performed with Hoechst 33258 dye to determine the effects of LAMS-2 on the proliferation and apoptosis of LOVO cells. Flow cytometry (FCM) was used to detect the level of reactive oxygen species (ROS), mitochondrion permeability transition pore (MPTP) and mitochondrial membrane potential (△ψm). Laser scanning confocal microscope (LSCM) was used to analyze intracellular calcium ion concentration, Western blot was performed to analyze the expressions of Cyt-C, caspase-9 and caspase-3. Spectrophotometer was applied to quantify the activity of caspase-9 and caspase-3. RESULTS: LAMS-2 inhibited LOVO cell proliferation and induced apoptosis, the apoptosis morphology was obvious. LAMS-2 treatment increased the intracellular level of ROS and Ca2+; activated intracellular MPTP and decreased △ψm; it also induced the release of Cyt-C and the activation of caspase-9 and caspase-3, increased the activity of caspase-9 and caspase-3. CONCLUSION: LAMS-2 can effectively inhibit the proliferation of LOVO cells and induce apoptosis through a mitochondria-mediated pathway. Keywords:

Anti-cancer effect of ethylacetate fraction from Orostachys japonicus on HT-29 human colon cancer cells by induction of apoptosis through caspase-dependent signaling pathway

Objective: To investigate the anti-colon cancer effects of ethylacetate fraction from Orostachys japonicus (O. japonicus) on HT-29 cancer cells. Methods: The viability of HT-29 cells was assayed by the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) method. Apoptosis induction and cell cycle inhibition were confirmed by fluorescein isothiocyanate and propidium iodide staining using flow cytometry. Morphological changes in the nucleus were observed, using a fluorescence microscope with 4′,6-diamidino-2-phenylindole (DAPI) nuclear staining. The expression levels of the upstream and downstream proteins involved in the anti-cancer mechanism were confirmed by Western blotting. Results: After treating HT-29 cells with different concentrations of ethylacetate fraction from O. japonicus, the viability of cells decreased in a concentration-dependent manner, while apoptosis induction and apoptotic body formation increased. Cell cycle analysis showed that the arrest occurred at the sub-G

Anti-cancer effect of ethylacetate fraction from Orostachys japonicus on HT-29 human colon cancer cells by induction of apoptosis through caspase-dependent signaling pathway

Objective:To investigate the anti-colon cancer effects of ethylacetate fraction from Orostachys japonicus (O. japonicus) on HT-29 cancer cells.Methods:The viability of HT-29 cells was assayed by the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) method. Apoptosis induction and cell cycle inhibition were confirmed by fluorescein isothiocyanate and propidium iodide staining using flow cytometry. Morphological changes in the nucleus were observed, using a fluorescence microscope with 4′,6-diamidino-2-phenylindole (DAPI) nuclear staining. The expression levels of the upstream and downstream proteins involved in the anti-cancer mechanism were confirmed by Western blotting.Results:After treating HT-29 cells with different concentrations of ethylacetate fraction from O. japonicus, the viability of cells decreased in a concentration-dependent manner, while apoptosis induction and apoptotic body formation increased. Cell cycle analysis showed that the arrest occurred at the sub-GConclusions:Combining the above results, it is thought that the survival of HT-29 cells is suppressed by ethylacetate fraction from O. japonicus through mitochondrial regulation-induced caspase cascade activation, induction of apoptosis and cell cycle arrest.

Study on the Effect and Its Mechanism of Fuzheng Jiedu Quyu Formula Combined with Oxaliplatin on Proliferation and Apoptosis of Human Colon Cancer HT-29 Cell / 中国药房

OBJECTIVE:To study the effect and its mechanism of Fuzheng Jiedu Quyu formula(short forFuzheng formula) combined with oxaliplatin (L-OHP) on human colon cancer HT-29 cell proliferation and apoptosis. METHODS:HT-29 cells were divided into blank control group(without drugs),Fuzheng formula group(1000 mg/L),L-OHP group(31.25 mg/L)and combi-nation group (1000 mg/L Fuzheng formula+31.25 mg/L L-OHP). After cultured with corresponding drug for 48 h,MTT method was used to detect the cell proliferation;the changes of cellular morphology were observed by invert microscope;flow cytometry was used to detect the cell cycle and apoptosis rate. Proapoptotic gene Bax,apoptotic gene Bcl-2 mRNA expressions were deter-mined by real-time fluorescence quantitative polymerase chain reaction method;Bax,Bcl-2 protein expressions were assayed by Western blot. RESULTS:Compared with blank control group,cell proliferation was inhibited in L-OHP group and combination group;cell proportion was increased in S stage,G2/M stage and decreased in G0/G1 stage(P<0.05). Cell apoptosis rate in L-OHP group,Fuzheng formula group and combination group was increased;Bax mRNA and protein expression were up-regulated,Bcl-2 mRNA expression was downregulated(P<0.05);and combination group changed more obviously than the single drug groups(P<0.05). CONCLUSIONS:Fuzheng formula combined with L-OHP can inhibit HT-29 cell proliferation and promote its apoptosis, showing better effects than either of the two drugs alone. The mechanism may be associated with up-regulation of Bax gene and pro-tein expressions and down-regulation of Bcl-2 gene expressions in cells.

Induced apoptosis by quercetin in human colon cancer HT-29 cells through Traf6/TAK1 signaling pathway / 国际中医中药杂志

Objective To study the effect of quercetin on the apoptosis of human colon cancer HT-29 cells and observe the possible mechanism.Methods Human colon cancer HT-29 cells were cultured in vitro and divided into four groups, including the control group, the 16μg/ml quercetin group, the Traf6 inhibitor group and the 16μg/ml quercetin + inhibitor group. The cells in control group were cultured with complete medium and other groups were treated with quercetin or/and Traf6 inhibitor. Flow cytometry was used to observe the impact of quercetin on the apoptosis of HT-29 cells; Western Blot technology was used to detect the expression levels of Traf6, TAK1, p-TAK1, caspase-3, Bax and Bcl-2; RT-PCR was used to investigate the expression level of Traf6 mRNA after treating for 24 h.Results Compared with the 16μg/ml quercetin group, the expression levels of Traf6 (0.59 ± 0.03 vs. 0.96 ± 0.04), p-TAK1 (0.43 ± 0.02vs. 0.72 ± 0.04), caspase-3 (0.59 ± 0.03vs. 0.70 ± 0.04), and Bax (0.48 ± 0.03vs.0.67 ± 0.04) were significantly decreased in 16μg/ml quercetin + inhibitor group(P<0.05). while the expression levels of Bcl-2 (0.54 ± 0.03vs. 0.44 ± 0.02) was significantly increased (P<0.05).Conclusions Quercetin can induce the apoptosis of human colon cancer HT-29 cells and the effective mechanism may relate to the activation of Traf6/TAK1 signaling pathway.

Induced apoptosis by quercetin in human colon cancer HT-29 cells through Traf6/TAK1 signaling pathway / 国际中医中药杂志

Objective To study the effect of quercetin on the apoptosis of human colon cancer HT-29 cells and observe the possible mechanism.Methods Human colon cancer HT-29 cells were cultured in vitro and divided into four groups, including the control group, the 16μg/ml quercetin group, the Traf6 inhibitor group and the 16μg/ml quercetin + inhibitor group. The cells in control group were cultured with complete medium and other groups were treated with quercetin or/and Traf6 inhibitor. Flow cytometry was used to observe the impact of quercetin on the apoptosis of HT-29 cells; Western Blot technology was used to detect the expression levels of Traf6, TAK1, p-TAK1, caspase-3, Bax and Bcl-2; RT-PCR was used to investigate the expression level of Traf6 mRNA after treating for 24 h.Results Compared with the 16μg/ml quercetin group, the expression levels of Traf6 (0.59 ± 0.03 vs. 0.96 ± 0.04), p-TAK1 (0.43 ± 0.02vs. 0.72 ± 0.04), caspase-3 (0.59 ± 0.03vs. 0.70 ± 0.04), and Bax (0.48 ± 0.03vs.0.67 ± 0.04) were significantly decreased in 16μg/ml quercetin + inhibitor group(P<0.05). while the expression levels of Bcl-2 (0.54 ± 0.03vs. 0.44 ± 0.02) was significantly increased (P<0.05).Conclusions Quercetin can induce the apoptosis of human colon cancer HT-29 cells and the effective mechanism may relate to the activation of Traf6/TAK1 signaling pathway.

Effects of SphK1 and FAK on epithelial-mesenchymal transition in colon cancer HCT116 cells / 中国病理生理杂志

[ ABSTRACT] AIM:To investigate the effects of sphingosine kinase l ( SphK1) and focal adhesion kinase ( FAK) on the epithelial-mesenchymal transition ( EMT) of human colon cancer HCT 116 cells.METHODS:Human colon cancer HCT116 cells were divided into 3 groups.N, N-dimethylsphingosine (DMS) was used to suppress the activity of SphK1. PF573228 was used to suppress the activation of FAK .The cells treated with equal volume of culture medium severed as control group.The cell viability was measured by MTT assay .The protein expression of SphK1, FAK and the EMT relative protein E-cadherin, N-cadherin, vimentin and matrix metalloproteinase (MMP) 2 was analyzed by Western blot.The mR-NA expression of SphK1, sphingosine-1-phosphate (S1P), FAK, E-cadherin and vimentin was detected by real-time PCR. The ability of tumor cell migration was measured by wound-healing assay.RESULTS:The cell viability of HCT116 cells was suppressed by DMS and PF 573228 in dose and time dependent manners .DMS significantly suppressed the expression of SphK1, FAK, N-cadherin, vimentin and MMP2, meanwhile enhanced the expression of E-cadherin.PF573228 reduced the expression of FAK , SphK1, N-cadherin, vimentin and MMP2, meanwhile increased the expression of E-cadherin (P<0.01).In addition, the migration ability of HCT116 cells was significantly decreased by treating with DMS and PF573228 (P<0.01).Compared with control group , the mRNA expression of FAK, SphK1, S1P and vimentin was de-creased, while the expression of E-cadherin was increased significantly in PF573228 group and DMS group (P<0.05). CONCLUSION:SphK1 and FAK signaling pathways may play an important role in the occurrence of EMT in the colon cancer HCT116 cells.

Interaction between MMS2 and P53 on proliferation and apoptosis of colon cancer cells / 中国免疫学杂志

Objective:To explore the regulatory effects of proliferation and apoptosis on THC-8307 by MMS2 siRNA and P53 siRNA.Methods:We experimentally suppressed the MMS2 and P53 expression in human colon cancer cells by the interference RNA technology ( RNAi) as monitored by Real-time qRT-PCR and Western blot.THC-8307 cells that express rate significantly reduced were collected as case group , while using untreated cells as the blank control group , and mock-treated cells as the negative control group.After separately interfering the target genes in each group ,test the relationship and expression level of the two genes.Utilizing flow cytometry techniques to test cells proliferation and apoptosis rate of each group.Results: Compared to the control group , colon cancer cells in which MMS2 and P53 were silenced displayed significant increase of P53,MMS2 mRNA and protein levels(P<0.05).MMS2-depleted cells displayed increase in apoptosis rates ,for both early and later stages ( P<0.05 ).Conclusion: MMS2 and P53 negatively regulate each other in colon cancer cells proliferation and apoptosis .

Effect of angiogenesis on Solanine and VEGF antibody in chicken embryo transplantation model of human colon cancer cells / 重庆医学

Objective To establish model of the chicken embryo transplantation of human colon cancer cells ,and investigate the effect of Solanine、VEGF antibody and Solanine combined with VEGF antibody on human colon cancer cells induce tumor angio‐genesis and tumor proliferation .Methods The model of the chicken embryo transplantation of human colon cancer HT‐29 cells were divided into three experimental group and control group .We added to the chick embryo chorioallantoic membrane with Sola‐nine、VEGF antibody and Solanine+ VEGF antibody mixture ,PBS was added to the control group .Then we analysed picture through the stereomicroscope and IPP 6 .0 image analysis software ,using immunohistochemistry envision method to detect of CD34 antigen and ki‐67 antigen ,and observing effect of Solanine group ,VEGF antibody group ,Solanine+ VEGF antibody group and the effect on the tumor angiogenesis and tumor proliferation .Results The tumor angiogenesis ,CD34 antigen and ki‐67 antigen of Sola‐nine+VEGF antibody group were significantly better than those of VEGF antibody group and Solanine group(P<0 .01);VEGF antibody group had statistical significant difference with Solanine group(P<0 .01);the effect of other three groups were better than that of the control group(P<0 .01) .Conclusion Solanine、VEGF antibody and Solanine combined with VEGF antibody could in‐hibit tumor angiogenesis and tumor proliferation of human colon cancer cell line HT‐29 to induce .It provides a new way for anti‐an‐giogenes .

Anti-proliferation effect of zoledronic acid on human colon cancer line SW480

Objective: To investigate the anti-proliferation effect and mechanism of zoledronic acid (ZOL) on human colon cancer line SW480. Methods: SW480 cells were treated with 0, 12.5, 25, 50, 100 and 200 μmoL/L of ZOL for 48 h, and CCK-8 assay was employed to obtain the survival rate of SW480 cells. SW480 cells were treated with 25 μmoL/L of ZOL for 0, 12, 24, 48 and 72 h, and then the survival rate was obtained. SW480 cells of the ZOL group were treated with 25 μmoL/L of ZOL for 48 h, while cells of the CsA + ZOL group were pretreated with 10 μmoL/L of CsA for 0.5 h and then treated with 25 μmoL/L of ZOL for 48 h. Then the survival rates of SW480 cells of the control group, ZOL group and CsA + ZOL group were determined. Flow cytometry was employed to detect the apoptosis rate and the mitochondrial transmembrane potential (▵Ψm) of the three groups and Western blot was used to detect the expressions of cyt C in the cytosol of the three groups. Results: ZOL inhibited the proliferation of SW480 cells, and the inhibition rate positively correlated with the concentration of ZOL and the action time (P < 0.01). The cell survival rate and the ▵Ψm of the ZOL group were greatly lower than those of the control group, while the apoptosis rate and the expression of cyt C in the cytosol were obviously higher than those of the control group. All the differences showed distinctly statistical significances (P < 0.01). The cell survival rate and the ▵Ψm of the CsA + ZOL group were all lower than those of the control group, but substantially higher than those of the ZOL group; while the apoptosis rate and the expression of cyt C in the cytosol were higher than those of the control group, but distinctly lower than those of the ZOL group. All the differences were statistically significant (P < 0.01). Conclusions: ZOL can induce the apoptosis in human colon cancer line SW480 and then inhibit the proliferation of SW480 cells directly by opening the mitochondrial permeability transition pore abnormally, decreasing ▵Ψm, and releasing the cyt C into the cytosol. And the effect enhances with the increases of the concentration of ZOL and the action time.

Naringenin-Mediated ATF3 Expression Contributes to Apoptosis in Human Colon Cancer

Naringenin (NAR) as one of the flavonoids observed in grapefruit has been reported to exhibit an anti-cancer activity. Activating transcription factor 3 (ATF3) is associated with apoptosis in human colon cancer cells. This study was performed to investigate the molecular mechanism by which NAR stimulates ATF3 expression and apoptosis in human colon cancer cells. NAR reduced the cell viability and induced an apoptosis in human colon cancer cells. ATF3 overexpression increased NAR-mediated cleaved PARP, while ATF3 knockdown attenuated the cleavage of PARP by NAR. NAR increased ATF3 expression in both protein and mRNA level, and increased the luciferase activity of ATF3 promoter in a dose-dependent manner. The responsible region for ATF3 transcriptional activation by NAR is located between -317 and -148 of ATF3 promoter. p38 inhibition blocked NAR-mediated ATF3 expression, its promoter activation and apoptosis. The results suggest that NAR induces apoptosis through p38-dependent ATF3 activation in human colon cancer cells.

Anti-proliferation effect of zoledronic acid on human colon cancer line SW480

OBJECTIVE@#To investigate the anti-proliferation effect and mechanism of zoledronic acid (ZOL) on human colon cancer line SW480.@*METHODS@#SW480 cells were treated with 0, 12.5, 25, 50, 100 and 200 μmoL/L of ZOL for 48 h, and CCK-8 assay was employed to obtain the survival rate of SW480 cells. SW480 cells were treated with 25 μmoL/L of ZOL for 0, 12, 24, 48 and 72 h, and then the survival rate was obtained. SW480 cells of the ZOL group were treated with 25 μmoL/L of ZOL for 48 h, while cells of the CsA + ZOL group were pretreated with 10 μmoL/L of CsA for 0.5 h and then treated with 25 μmoL/L of ZOL for 48 h. Then the survival rates of SW480 cells of the control group, ZOL group and CsA + ZOL group were determined. Flow cytometry was employed to detect the apoptosis rate and the mitochondrial transmembrane potential (△Ψm) of the three groups and Western blot was used to detect the expressions of cyt C in the cytosol of the three groups.@*RESULTS@#ZOL inhibited the proliferation of SW480 cells, and the inhibition rate positively correlated with the concentration of ZOL and the action time (P < 0.01). The cell survival rate and the △Ψm of the ZOL group were greatly lower than those of the control group, while the apoptosis rate and the expression of cyt C in the cytosol were obviously higher than those of the control group. All the differences showed distinctly statistical significances (P < 0.01). The cell survival rate and the △Ψm of the CsA + ZOL group were all lower than those of the control group, but substantially higher than those of the ZOL group; while the apoptosis rate and the expression of cyt C in the cytosol were higher than those of the control group, but distinctly lower than those of the ZOL group. All the differences were statistically significant (P < 0.01).@*CONCLUSIONS@#ZOL can induce the apoptosis in human colon cancer line SW480 and then inhibit the proliferation of SW480 cells directly by opening the mitochondrial permeability transition pore abnormally, decreasing △Ψm, and releasing the cyt C into the cytosol. And the effect enhances with the increases of the concentration of ZOL and the action time.

Bacillus spp. or Bacillus spp.-Derived Membrane Vesicles Induce the Intrinsic Pathways of Apoptosis of Human Colon Cancer Cell Lines

The striking increase in colorectal cancer (CRC) has shown the great fatality in Korea for more than 15 years. The leading edge of this rising incidence rate is mainly due to the people's dietary changes in Korea. Some studies have reported that the dietary fiber does not have significant cytotoxic effects on CRC cells, which contrasts to the effects of probiotics. It gives a positive evaluation that the nonpathogenic spore-forming Bacillus species among the probiotics including fermented bacteria might have optimistic effects on CRC incidence rate. Recently, we isolated Bacillus lentus (BL) from Korean soybean fermented food. BL showed the cytotoxic effect on human colon carcinoma cell lines HCT116 and SW480. Interestingly, BL did not have effect on human dermal fibroblast cells and human hepatoma cell line HepG2. It suggested that BL has the target cell-specific cytotoxicity toward human colon carcinoma cells. To clarify the death signaling pathway underlying the BL-induced apoptosis in cancer cells, we analyzed the expression of caspases, Bax and Bcl-2 by western blotting. The apoptotic effects by cytotoxic elements were executed by direct BL contact or membrane-derived vesicles isolated from BL. Treatment of HCT116 with BL activated caspase-9, -3 and increased cleavage form of poly (ADP-ribose) polymerase (PARP). However, caspase-8 activity was not increased by BL. BL-activated intrinsic pathway increased the pro-apoptotic Bax, decreased the anti-apoptotic Bcl-2 proteins on mitochondria, disrupted the mitochondrial membrane potential, and then released the cytochrome c from mitochondria. The membrane-derived vesicles (MVs) from BL induced apoptosis of the HCT116. Here, we propose that BL as a strong candidate for the development of apoptosis-specific anti-tumor agent will give great contribution to the understandings of the tumor-microbe interdisciplinary areas.