OBJECTIVE: To investigate the effect of danusertib (Danu), an inhibitor of Aurora kinase, on the proliferation, cell cycle, apoptosis, and autophagy of hepatocellular carcinoma HepG2 cells and explore the underlying mechanisms. METHODS: MTT assay was used to examine the effect of Danu on the viability of HepG2 cells to determine the IC50 of Danu. The effect of Danu on cell cycle distribution, apoptosis and autophagy were determined using flow cytometry. Western blotting was used to detect the expressions of the proteins related to cell cycle, apoptosis and autophagy. Chloroquine was used to suppress Danuinduced autophagy to test the apoptosis-inducing effect of Danu. RESULTS: Danu significantly inhibited the proliferation of HepG2 cells with IC of 39.4 μmol and 14.4 μmol at 24 h and 48 h, respectively. Danu caused cell cycle arrest in G/M phase in HepG2 cells and led to polyploidy accumulation via up-regulating the expressions of p53 and p21 and down-regulating the expressions of cyclin B1 and DC2. Danu also caused apoptosis of HepG2 cells through up-regulating the expressions of Bax, Puma, cleaved caspase-3, cleaved caspase-9, cleaved PARP and cytochrome C and down-regulating the expressions of Bcl-xl and Bcl-2. Danu induced autophagy via activating AMPK signaling and inhibiting PI3K/PTEN/AKT/mTOR axis, and inhibition of Danu-induced autophagy with chloroquine enhanced the pro-apoptotic effect of Danu. CONCLUSIONS Danu inhibits cell proliferation and induces cell cycle arrest in G/M phase, apoptosis and cytoprotective autophagy in HepG2 cells.
Apoptosis ; drug effects ; Autophagy ; drug effects ; Benzamides ; pharmacology ; Carcinoma, Hepatocellular ; pathology ; Cell Cycle ; drug effects ; Cell Division ; drug effects ; Cell Proliferation ; drug effects ; Hep G2 Cells ; Humans ; Liver Neoplasms ; pathology ; Neoplasm Proteins ; metabolism ; Protein Kinase Inhibitors ; pharmacology ; Pyrazoles ; pharmacology

Objective: To explore the apoptosis-inducing effect of the Chinese medicinal compound CFF-1 on prostate cancer cells and its related molecular mechanisms. METHODS: Normal prostate WPMY-1 cells and prostate cancer LNCaP, CWR22Rv1, PC3 and DU145 cells were treated in dehydrated alcohol with CFF-1 at 0, 2, 5, or 10 mg/ml for 24 hours. Then the viability of the prostate cells was detected by morphological observation, MTT and CCK-8 assay, nuclear condensation and disruption measured by DAPI staining, the cell cycle and apoptosis calculated by flow cytometry, the activity of the PI3K/AKT/FOXO1 signaling pathway and the expressions of its downstream apoptosis- and cycle-related proteins determined by Western blot. RESULTS: CFF-1 significantly arrested the cell cycle in the G1 phase, decreased the cell viability and increased the nuclear condensation and disruption in a dose-dependent manner, and elevated the apoptosis rate of prostate cancer cells. At the molecular level, CFF-1 dose-dependently reduced the activity of the PI3K/AKT signaling pathway and phosphorylation of the FOXO1 protein, increased the transcription activity of FOXO1, and eventually regulated the expressions of cell apoptosis- and cycle-related genes. CONCLUSIONS The Chinese medicinal compound CFF-1 can significantly inhibit the growth, arrest the cycle, and induce the apoptosis of prostate cancer cells by decreasing the activity of the PI3K/AKT/FOXO1 signaling pathway, which suggests its potential clinical application value in the treatment of prostate cancer.
Antineoplastic Agents, Phytogenic ; pharmacology ; Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Division ; Cell Line, Tumor ; Cell Proliferation ; Cell Survival ; Drugs, Chinese Herbal ; pharmacology ; Forkhead Box Protein O1 ; metabolism ; Humans ; Male ; Neoplasm Proteins ; metabolism ; Phosphatidylinositol 3-Kinases ; metabolism ; Phosphorylation ; Prostatic Neoplasms ; drug therapy ; metabolism ; pathology ; Proto-Oncogene Proteins c-akt ; metabolism ; Signal Transduction ; drug effects

OBJECTIVETo determine the effect of aspirin on cell proliferation, alkaline phosphatase (ALP) activity, cell cycle and apoptosis in murine bone marrow stromal cells, so as to explore an appropriate dose range to improve bone regeneration in periodontal treatment.

METHODSST2 cells were stimulated with aspirin (concentrations of 1, 10, 100 and 1 000 μmol/L) for 1, 2, 3, 5 and 7 d. Cell proliferation was measured by methyl thiazolyl tetrazolium (MTT) assay. After ST2 cells were treated for 1, 3 and 7 d, ALP activity was measured by ALP kit, cell cycle and apoptosis were measured by flow cytometry (FCM) after treated for 48 h.

RESULTSMTT assays showed that various doses of aspirin have different effects on the cell growth. Briefly, lower concentrations (1, 10 μmol/L) of aspirin promoted the cell growth, the A value of 0, 1 and 10 μmol/L aspirin 7-day-treated cells were 0.313±0.012, 0.413±0.010 and 0.387±0.017 respectively (P <0.01 vs control), and so did the ALP level ([4.3±0.9], [6.0±0.3] and [7.7±0.4] μmol·min(-1)·g(-1), P <0.05 vs control), while higher concentrations, especially 1000 μmol/L of aspirin might inhibit the cell growth with time going, A value and ALP level were 0.267±0.016, (4.3±1.3) μmol·min(-1)·g(-1) respectively (P <0.05 vs control). Cell cycle analysis revealed no changes in comparison to control cells after treatment with 1 or 10 μmol/L aspirin, but it was observed that cell mitosis from S phase to G2/M phase proceeded at higher concentrations of 100 μmol/L aspirin, and the cell cycle in phase G0/G1 arrested at 1000 μmol/L. Parallel apoptosis/necrosis studies showed that the percentage of cells in apoptosis decreased dramatically at all doses of aspirin, the apoptosis rates of ST2 cells responded to 0, 1, 10, 100 and 1000 μmol/L aspirin were (11.50±0.90)%, (5.30±0.10)%, (5.50±0.10)%, (4.90±0.90)% and (7.95±0.25)% respectively (P<0.05 vs control).

CONCLUSIONSThis study demonstrated that lower dosage of aspirin can promote ST2 cells growth, osteogenic activity and inhibit its apoptosis. Aspirin maybe used for the bone reconstruction with a proper concentration.

Alkaline Phosphatase ; metabolism ; Animals ; Apoptosis ; drug effects ; Aspirin ; administration & dosage ; pharmacology ; Bone Regeneration ; Cell Cycle ; drug effects ; Cell Division ; Cell Line, Tumor ; Cell Proliferation ; Flow Cytometry ; Formazans ; Mesenchymal Stromal Cells ; cytology ; drug effects ; enzymology ; Mice ; Periodontics ; Tetrazolium Salts ; Time Factors

OBJECTIVETo investigate the inhibitory effect of HSP90 inhibitory 17-AAG on proliferation of multiple myeloma cells and its main mechanism.

METHODSThe multiple myeloma cells U266 were treated with 17-AAG of different concentrations (200, 400, 600 and 800 nmol/L) for 24, 48, and 72 hours respectively, then the proliferation rate, expression levels of β-catenin and C-MYC protein, as well as cell cycle of U266 cells were treated with 17-AAG and were detected by MTT method, Western blot and flow cytometry, respectively.

RESULTSThe 17-AAG showed inhibitory effect on the proliferation of U266 cells in dose- and time-depetent manners (r = -0.518, P < 0.05 and r = -0.473, P < 0.05), while the culture medium without 17-AAG displayed no inhibitory effect on proliferation of U266 cells (P > 0.05). The result of culturing U266 cells for 72 hours by 17-AAG of different concentrations showed that the more high of 17-AAG concentration, the more low level of β-catenin and C-MYC proteins (P < 0.05); At same time of culture, the more high of 17-AAG concentration, the more high of cell ratio in G1 phase (P < 0.05), at same concentration of 17-AAG, the more long time of culture, the more high of cell ratio in G1 phase (P < 0.05).

CONCLUSIONThe HSP90 inhibitory 17-AAG can inhibit the proliferation of multiple myeloma cells, the down-regulation of Wnt/β-catenin signaling pathway and inhibition of HSP90 expression may be the main mechnisms of 17-AAG effect.

Apoptosis ; Benzoquinones ; pharmacology ; Cell Cycle ; Cell Division ; Cell Line, Tumor ; drug effects ; Cell Proliferation ; drug effects ; Down-Regulation ; HSP90 Heat-Shock Proteins ; antagonists & inhibitors ; Humans ; Lactams, Macrocyclic ; pharmacology ; Multiple Myeloma ; metabolism ; pathology ; Proto-Oncogene Proteins c-myc ; metabolism ; Wnt Signaling Pathway ; drug effects ; beta Catenin ; metabolism

To investigate the effect of jianpi-jiedu (JPJD) prescription-contained serum on colorectal cancer SW48 cell proliferation and the underlying mechanisms.
 Methods: Crude extract from JPJD was made by water extract method and the main components of crude extract from JPJD were analyzed by ultra-performance liquid phase high resolution time of flight mass spectrometry (UPLC-Q-TOF/MS). The low, medium, and high-concentration of JPJD-contained serum were prepared by the serum pharmacological method. The effect of serum containing JPJD on SW48 cell proliferation was determined by MTT assay. The cell cycle was detected by flow cytometric method. The protein levels of mammalian target of rapamycin (mTOR), phospho-mTOR, P-P53, and -P21, and the mRNA level of mTOR were examined by Western blot and RT-PCR, respectively.
 Results: Seven compounds including calycosin-7-glucoside, astragaloside, ginsenoside-Re, ginsenoside-Rb1, glycyrrhizinic acid, apigenin, atractylenolide-II were identified. MTT assays demonstrated that the SW48 cell proliferation was inhibited by medium and high concentration of JPJD-contained serum and the percentages of cells at G1 phase in SW48 cell cultured in the medium and high concentration of JPJD serum group were significantly higher than those in the control group (P<0.05). Meanwhile, the levels of mTOR mRNA and phospho-mTOR protein in the medium and high concentration of JPJD serum groups were substantially lower than those in the control group (P<0.05). Conversely, the expressions of phospho-P53 and P21 protein were significantly increased in the medium and high concentration of JPJD serum group compared with those in the control group.
 Conclusion: JPJD prescription-contained serum can inhibit SW48 cell proliferation, which may be related to mTOR-P53-P21 signaling pathways.
Animals ; Apigenin ; Blotting, Western ; Cell Cycle ; Cell Division ; Cell Proliferation ; drug effects ; genetics ; Colorectal Neoplasms ; Cyclin-Dependent Kinase Inhibitor p21 ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Flow Cytometry ; Ginsenosides ; Glycyrrhizic Acid ; Humans ; Lactones ; Phosphorylation ; genetics ; RNA, Messenger ; Saponins ; Sesquiterpenes ; Signal Transduction ; TOR Serine-Threonine Kinases ; drug effects ; Triterpenes ; Tumor Suppressor Protein p53 ; drug effects

OBJECTIVETo investigate the effects of β-elemene in suppressing the proliferation and apoptosis of SGC7901 gastric cancer cells in vitro and explore the underlying mechanisms.

METHODSUsing MTT assay, flow cytometry, and clonogenic survival assay, we assessed the effects of β-elemene on the viability, apoptosis, cell cycle distribution, and clonogenic survival of gastric cancer SGC7901 cells and gastric mucosal epithelial GES-1 cells. Western blotting was employed to determine the changes in the protein expression profiles in SGC7901 cells in response to β-elemene treatment.

RESULTSβ-elemene significantly suppressed the cell viability and increased the apoptosis of SGC7901 cells, and these effects were less obvious in GES-1 cells. β-elemene decreased clonogenic survival of SGC7901 cells, increased the proportion of G2/M phase cells, decreased the expression of Bcl-2, and increased the expression of Bax and cleaved caspase-3. β-elemene did not obviously affect the expression of total p21-activated protein kinase 1 (Pak1) but decreased the level of phospho-Pak1 (Thr423) and phospho-ERK1/2 (Thr202/Tyr204) in SGC7901 cells.

CONCLUSIONβ-elemene inhibits the proliferation and induces apoptosis of gastric cancer cells possibly by inhibiting Pak1/ERK signaling and regulating apoptosis-associated proteins such as Bcl-2 and Bax.

Apoptosis ; Apoptosis Regulatory Proteins ; metabolism ; Cell Cycle ; Cell Division ; Cell Line, Tumor ; drug effects ; Cell Proliferation ; Cell Survival ; Humans ; Sesquiterpenes ; pharmacology ; Signal Transduction ; Stomach Neoplasms ; pathology

OBJECTIVETo investigate whether CYC116 can potentiate matrine-dependent growth inhibition and apoptosis in multiple myeloma (MM) cells.

METHODSThe dose response relationship of matrine to dexamethasone-resistant and dexamethasone-sensitive MM cells was first established. Myeloma RPMI8226 cells were treated with matrine alone or combined with CYC116 for 24 h. Cell proliferation was measured using an MTT assay and apoptosis induction was evaluated by flow cytometry. Activation of the caspase pathway and expression of apoptosis regulator proteins were detected by Western blotting.

RESULTSMatrine significantly induced growth arrest and apoptosis in both drug-resistant and drug-sensitive MM cells. Treatment with the combination of matrine and CYC116 had a stronger cytotoxic effect on MM cells than did single drug treatments. Enhanced apoptosis observed following the combined treatment of matrine and CYC116 was associated with higher levels of activation of caspase-9, caspase-3, and poly adenosine diphosphate ribose polymerase (PARP) and down-regulation of the anti-apoptotic proteins Bcl-2 and Mcl-1 and the signaling proteins p-Akt and nuclear factor κB (NF-κB).

CONCLUSIONCYC116 enhances the growth inhibitory and apoptotic effects of matrine on MM cells.

Alkaloids ; pharmacology ; Apoptosis ; drug effects ; Cell Division ; drug effects ; Cell Line, Tumor ; Humans ; Multiple Myeloma ; pathology ; Pyrimidines ; pharmacology ; Quinolizines ; pharmacology ; Thiazoles ; pharmacology

OBJECTIVEThis aimed to investigate the effect of specific sequence oligodeoxynucleotide MT01 on the biological properties of osteoblasts invaded by Porphyromonas gingivalis (P. gingivalis ) by evaluating proliferation, cell cycle, and apoptosis.

METHODSMG63 osteoblasts were recovered and incubated with MT01, CpG ODN, metronidazole (MNZ), and gentamicin (GEN) for 3 h. P. gingivalis (the multiplicity of infection was 100:1) was added subsequently and cocultured for another 24 and 48 h. Cells with PBS comprised the blank group, whereas cells with P. gingivalis comprised the negative controls. Six experimental groups were established: PBS group, P. gingivalis group, MT01+P. gingivalis group, CpG ODN+ P. gingivalis group, MNZ+P. gingivalis group, and GEN+P. gingivalis group. The proliferative ability was measured by methyl thiazolyl tetrazolium assay, and the percentages of apoptosis and cell cycle were examined by flow cytometry.

RESULTSCompared with the blank group, proliferation increased significantly in the MT01+P. gingivalis group (P < 0.05). The ratio of cells was lower at the G₁ phase and higher at the S phase in the MT01+P. gingivalis group compared with the results in the P. gingivalis group (P < 0.05). Early cell apoptosis in the MT01+P. gingivalis group was significantly lower than that in the P. gingivalis group (P < 0.05).

CONCLUSIONMT01 can promote the proliferation, reduce the ratio of the G₁phase, increase the ratio of the S phase, and inhibit the early apoptosis of osteoblasts invaded by P. gingivalis.

Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Division ; Cell Proliferation ; drug effects ; Flow Cytometry ; Gentamicins ; pharmacology ; Humans ; Metronidazole ; pharmacology ; Oligodeoxyribonucleotides ; pharmacology ; Osteoblasts ; cytology ; drug effects ; Porphyromonas gingivalis ; pathogenicity

OBJECTIVETo explore apoptosis of multiple myeloma (MM) cells and its mechanism by the combined inhibition of mTORC2 signaling pathway and heat shock protein 90.

METHODSThe effects of Rapamycin, 17-AAG and the combination on proliferation of MM cell lines U266 and KM3 were assessed using MTT at different time points (0, 8, 24, 48 hour). Cell apoptosis and cell cycle distribution were measured by flow cytometry. The specific proteins p-AKT (ser473), p-AKT (thr450), p-S6 (S235/236) and AKT were detected by Western blotting.

RESULTSRapamycin, 17- AAG and the combination suppressed the proliferation of MM cell lines U266 and KM3, especially the combination of Rapamycin and 17-AAG synergistically inhibited the proliferation (P<0.05); Rapamycin induced G1 arrest both at 24 and 48 hours, 17-AAG also induced G1 arrest, especially at 48 hours (P<0.01); Rapamycin, 17-AAG alone decreased the expression of AKT and induced MM cell apoptosis to some extent (P<0.01); Chronic rapamycin treatment inhibited mTORC2; Inhibition of both mTORC2 and chaper on pathways degraded AKT and induced MM cell apoptosis, which was significantly higher than that of any single agent (P<0.01).

CONCLUSIONInhibition of both mTORC2 and chaper on pathways decreased the expression of AKT to induce apoptosis of MM cells in vitro.

Apoptosis ; Benzoquinones ; pharmacology ; Cell Cycle ; Cell Division ; Cell Line, Tumor ; drug effects ; HSP90 Heat-Shock Proteins ; metabolism ; Humans ; Lactams, Macrocyclic ; pharmacology ; Mechanistic Target of Rapamycin Complex 2 ; Multiple Myeloma ; pathology ; Multiprotein Complexes ; antagonists & inhibitors ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Signal Transduction ; Sirolimus ; pharmacology ; TOR Serine-Threonine Kinases ; antagonists & inhibitors ; metabolism

This study was purposed to explore the mechanisms of cladribine (2-CdA)-inducing apoptosis of multiple mycloma RPMI 8226 cells. The MTT method was used to determine cell proliferation after being treated with 2-CdA. Apoptosis and cell cycle progression were examined by flow cytometry. Transmission electron microscopy was used to observe ultrastructural changes of RPMI 8226 cells. RT-PCR and Western blot were used to analyze the mRNA and protein expression levels of BCL-2, MCL-2 and caspase-3 respectively. The results showed that the 2-CdA inhibited proliferation of RPMI 8226 cells in time and dose-dependent manner. Typical apoptotic morphological and ultrastructure changes could be observed by electron microscopy. Flow cytometry showed that 2-CdA induced myeloma cell apoptosis and arrested myeloma cells in the G2/M phase. The mRNA expression of BCL-2 and MCL-1 decreased but that of caspase-3 not apparently changed. Western blot results suggested that the change trend of BCL-2 MCL-1 and caspase-3 was the same as result of RT-PCR. It is concluded that 2-CdA exhibits inhibitory effects on RPMI 8226 cells in vitro. Activating the mitochondrial and death receptor pathways of apoptosia may be the potential mechanism, meanwhile, the cell cycle arrest may also play a critical role in apoptosis.
Apoptosis ; drug effects ; Caspase 3 ; Cell Division ; Cell Line, Tumor ; Cell Proliferation ; Cladribine ; pharmacology ; Humans ; Multiple Myeloma ; pathology ; Proto-Oncogene Proteins c-bcl-2