In the present study, 13 clinical cases of canine mammary adenocarcinoma were evaluated in order to understand the effect of Tarantula cubensis extract (TCE) on tumor tissue. Punch biopsies were taken from the tumors before treatment with TCE. Subcutaneous injections of TCE were administered three times at weekly intervals (3 mL per dog). Between days 7 and 10 after the third injection, the tumor masses were extirpated by complete unilateral mastectomy. Pre- and post-treatment tumor tissues were immunohistochemically assessed. The expression of B-cell lymphoma 2 (Bcl-2) was found to be higher in pre-treatment compared to post-treatment tissues (p < 0.01) whereas Ki-67 expression was lower in post-treatment tissues (p < 0.01). No significant differences in fibroblast growth factor or vascular endothelial growth factor expression were observed between pre- and post-treatment tissues (p > 0.05). The apoptotic index was determined to be low before treatment and increased during treatment. These results suggest that TCE may be effective for controlling the local growth of canine mammary adenocarcinoma by regulating apoptosis.
Adenocarcinoma/*drug therapy/physiopathology ; Animals ; Apoptosis/drug effects ; Dog Diseases/*drug therapy/physiopathology ; Dogs ; Female ; Mammary Neoplasms, Animal/*drug therapy/physiopathology ; Mammary Neoplasms, Experimental/*drug therapy/physiopathology ; Mitosis/drug effects ; Spiders/*chemistry

AIM: To investigate the active constituents of Lignum Sappan (Caesalpinia sappan L.) on growth-related signaling and cell mitosis. METHOD: The influence of the ethyl acetate (EtOAc) extract of Lignum Sappan and its constituents on growth-related signaling were evaluated by a luciferase assay in cells stably-transfected with NF-κB, STAT1, or STAT3 responsive luciferase reporter plasmid. The inhibitory effect on the cell cycle was determined by flow cytometric analysis. The anti-tumor activities were assessed in vitro and in vivo. RESULTS: The EtOAc extract of Lignum Sappan had inhibitory activities on growth-related signaling and cell mitosis. Three major active compounds were sappanchalcone, brazilin, and butein. Sappanchalcone blocked cell cycle progression in the G2/M phase, brazilin inhibited TNFα/NF-κB signaling, while butein inhibited IL-6/STAT3 signaling, as well as TNFα/NF-κB signaling. The three compounds all demonstrated cytotoxic activities against human tumor cells in vitro. In a S180 tumor cell-bearing mice model, the anti-tumor efficacy of the EtOAc extract was better than the individual compounds acting alone. CONCLUSION These results indicate that Lignum Sappan contains multiple active compounds with different antitumor activities, which act synergistically to enhance their anti-tumor effects. The EtOAc extract of Lignum Sappan may be better than individual active constituent as a novel medicine for the treatment of cancer.
Animals ; Antineoplastic Agents, Phytogenic ; pharmacology ; therapeutic use ; Benzopyrans ; pharmacology ; therapeutic use ; Caesalpinia ; Cell Cycle Checkpoints ; drug effects ; Chalcones ; pharmacology ; therapeutic use ; Hep G2 Cells ; Humans ; Interleukin-6 ; metabolism ; Male ; Mice, Inbred BALB C ; Mitosis ; drug effects ; NF-kappa B ; metabolism ; Phytotherapy ; Plant Extracts ; pharmacology ; therapeutic use ; STAT3 Transcription Factor ; metabolism ; Sarcoma ; drug therapy ; metabolism ; Signal Transduction ; Tumor Necrosis Factor-alpha ; metabolism

To further test whether protein kinase A (PKA) can affect the mitotic cell cycle, one-cell stage mouse embryos at S phase (22 h after hCG injection) were incubated in M16 medium containing various concentrations of H-89, a PKA inhibitor. With increasing concentrations of H-89 (0-50 μmol/L), the G(2) phase of eggs was decreased and the cleavage rate was accelerated. A concentration of 40 μmol/L H-89 led to all of the mouse eggs entering the M phase of mitosis. Furthermore, to study the role of PKA in regulating the phosphorylation status of S149 and S321 sites of cell division cycle 25B (CDC25B) on one-cell stage fertilized mouse eggs, pBSK-CDC25B-WT, pBSK-CDC25B-S149A, pBSK-CDC25B-S321A and pBSK-CDC25B-S149A/S321A were transcribed into mRNAs in vitro, then mRNAs were microinjected into S phase of mouse fertilized eggs and cultured in M16 medium pretreated with H-89. Then, the cleavage of fertilized eggs, maturation promoting factor (MPF) activity and phosphorylation status of CDC2-Tyr15 were observed. In the presence of 40 μmol/L H-89, the cleavage rate of fertilized eggs in CDC25B-S/A-mRNAs and CDC25B-WT-mRNA injected groups was significantly higher than that in the control groups, and the peak of MPF activity appeared in the CDC25B-S/A-mRNAs and CDC25B-WT-mRNA injected groups earlier than that in the control groups. CDC2-Tyr15 phosphorylation state was consistent with MPF activity. In conclusion, the present study suggests that PKA regulates the early development of mouse embryos by phosphorylation of S149 and S321 of CDC25B, which plays an important role in the regulation of G(2)/M transition in the mitotic cell cycle of fertilized mouse eggs.
Animals ; Cyclic AMP-Dependent Protein Kinases ; genetics ; physiology ; Embryonic Development ; physiology ; Female ; Male ; Mice ; Microinjections ; Mitosis ; drug effects ; Phosphorylation ; Serine ; genetics ; metabolism ; Zygote ; cytology ; growth & development ; cdc25 Phosphatases ; genetics ; metabolism

Microtubule is one of the key components of the cytoskeleton and plays an important role in the maintenance of cell shape and the process of signal transduction and mitosis. Due to the extreme importance of microtubule in the process of mitosis, tubulin becomes one of the most important targets for development of new anticancer drugs and tubulin inhibitors are used for the treatment of cancer nowadays. These inhibitors have antitumor activity by inhibiting or promoting the assembly of tubulin to microtubules and interfering the process of cell mitosis. This review summarized the research progress of the tubulin inhibitors, especially the introduction of the tubulin inhibitors of pharmacological activities and the progress of clinical research. Also, the development trend of these inhibitors is discussed.
Antineoplastic Agents ; chemical synthesis ; chemistry ; pharmacology ; Humans ; Microtubules ; drug effects ; metabolism ; Mitosis ; drug effects ; Molecular Structure ; Neoplasms ; drug therapy ; Stilbenes ; chemical synthesis ; chemistry ; pharmacology ; Structure-Activity Relationship ; Tubulin ; metabolism ; Tubulin Modulators ; chemical synthesis ; chemistry ; pharmacology

Liver cancer is one of the most common neoplastic diseases with high mortality in China. Currently, antimicrotubule drugs such as paclitaxel (PTX) and vincristine (VCR), are used as the common agents in the clinical chemotherapy for liver cancer. However, the responses of patients to these drugs vary markedly. Successful identification of intracellular factors influencing liver cancer's sensitivity to antimicrotubule drugs would be of great clinical importance. In this study, by engineering human hepatoma cell HepG2 to overexpress synuclein-gamma (SNCG), we investigated if SNCG is a molecular factor associated with the sensitivity to antimicrotubule drug treatment. Real-time RT-PCR and Western blotting assays showed SNCG was successfully overexpressed in HepG2/ SNCG cells compared with HepG2/Neo cells. The overexpressed SNCG altered the proliferation activity in HepG2 cells, which was 66% higher than that of HepG2/Neo cells through MTT method. The overexpressed SNCG also reduced sensitivity of HepG2 cells to antimicrotubule drugs: after PTX or VCR treatment, the proportion of HepG2/SNCG cells in G2/M arrest was significantly lower than that in HepG2/Neo cells. Correspondingly, HepG2/SNCG cells showed significantly lower mitotic index than HepG2/Neo cells. Meanwhile, HepG2/SNCG cells showed higher resistance to PTX and VCR than HepG2/Neo cells, with resistance index 21 and 15 respectively. Our studies suggested that the overexpression of SNCG could confer resistance to antimicrotubule drugs in hepatoma cells; and it indicated that SNCG may be as a potential response marker for antimicrotubule drugs in liver cancer chemotherapy.
Antineoplastic Agents, Phytogenic ; pharmacology ; Cell Cycle ; Cell Proliferation ; Drug Resistance, Neoplasm ; Gene Expression Regulation, Neoplastic ; Genetic Vectors ; Hep G2 Cells ; drug effects ; metabolism ; Humans ; Microtubules ; drug effects ; Mitosis ; drug effects ; Mitotic Index ; Paclitaxel ; pharmacology ; Plasmids ; RNA, Messenger ; metabolism ; Transfection ; Vincristine ; pharmacology ; gamma-Synuclein ; biosynthesis ; genetics ; physiology

Pin1 is a phosphorylation-dependent peptidyl-prolyl cis/trans isomerase, which specifically catalyzes the amide bond isomerization of phosphoserine-proline or phosphothreonine-proline in mitotic phosphoproteins. Pin1 induces the conformational changes to control the function of phosphoproteins. Depletion of Pinl on various human cancer cell lines cause mitotic arrest and apoptosis. Pin1 is an attracting therapeutic target for anticancer and its inhibitors might be potential anticancer drug. In this review, Pin1 inhibitors and the catalytic mechanism, the biological function of Pin1 and its role in oncogenesis are summarized.
Apoptosis ; Enzyme Inhibitors ; chemical synthesis ; pharmacology ; Humans ; Mitosis ; drug effects ; NIMA-Interacting Peptidylprolyl Isomerase ; Neoplasms ; enzymology ; Peptidylprolyl Isomerase ; antagonists & inhibitors ; metabolism ; Phosphoproteins ; chemistry ; metabolism ; Phosphorylation ; drug effects ; Signal Transduction ; drug effects

OBJECTIVETo investigate the effect of lidamycin (LDM) on telomerase activity in human hepatoma BEL-7402 cells under the condition of LDM inducing mitotic cell death and senescence.

METHODSChromatin condensation was detected by co-staining with Hoechst 33342 and PI. Cell multinucleation was observed by Giemsa staining and genomic DNA was separated by agarose gel electrophoresis. Fluorescent intensity of Rho123 was determined for mitochondrial membrane potential. MTT assay and SA-beta-gal staining were employed to analyze the senescence-like phenotype. The expression of proteins was analyzed by Western blot. Telomerase activity was assayed by telomerase PCR-ELISA.

RESULTSMitotic cell death occurred in LDM-treated cells characterized by unique and atypical chromatin condensation, multinucleation and increased mitochondrial membrane potential. However, no apoptotic bodies or DNA ladders were found. In addition, apoptosis-related proteins remained nearly unaltered. Senescence-like phenotype was identified by increased and elongated size of cells, growth retardation, enhanced SA-beta-gal activity and the changes of senescence-related protein expression. Telomerase activity markedly decreased (P<0.01) in LDM-treated hepatoma BEL-7402 cells.

CONCLUSIONMitotic cell death and senescence could be triggered simultaneously or sequentially after exposure of hepatoma BEL-7402 cells to LDM. The decrease in telomerase activity may play a key role in the defective mitosis and aging morphology. Further investigation of detailed mechanism is needed.

Aminoglycosides ; pharmacology ; Antibiotics, Antineoplastic ; pharmacology ; Apoptosis ; drug effects ; Azure Stains ; Benzimidazoles ; Carcinoma, Hepatocellular ; enzymology ; pathology ; Cell Nucleus ; drug effects ; metabolism ; Cellular Senescence ; drug effects ; Chromatin ; metabolism ; DNA, Neoplasm ; analysis ; Dose-Response Relationship, Drug ; Enediynes ; pharmacology ; Genome, Human ; genetics ; Humans ; Liver Neoplasms ; enzymology ; pathology ; Membrane Potential, Mitochondrial ; drug effects ; Mitosis ; drug effects ; Phenotype ; Propidium ; Telomerase ; metabolism ; Time Factors ; beta-Galactosidase ; metabolism

OBJECTIVETo observe the effect of polo-like kinase 1 (PLK1) gene depletion on mitosis phenotype and elucidate its vital role in gastric cancer cell line (MKN45) mitosis.

METHODSThe PLK1 expression in MKN45 cells was blocked by RNA interference (RNAi), the expression level of PLK1 mRNA and protein were measured by real-time quantitative PCR and Western blot, respectively. The morphological change of microtubules and mitosis phenotype in MKN45 cells were observed by immunofluorescence staining and laser confocal microscopy, the morphological changes of cells were observed by reverse microscopy, the variation of cell cycle distribution was detected by flow-cytometry.

RESULTSAfter RNAi targeting PLK1, PLK1 mRNA and protein level decreased obviously, the cell microtubules became obscure and lost cohesiveness, the mitosis phenotype also varied substantially (P < 0.05), more gastric cancer cells became rounded and showed G(2) phase cell DNA content (P < 0.05).

CONCLUSIONPLK1 gene plays a key role in mitosis and its inhibition can lead to mitosis arrest in MKN45 cells.

Adenocarcinoma ; enzymology ; metabolism ; pathology ; Cell Cycle Proteins ; biosynthesis ; genetics ; Cell Line, Tumor ; G2 Phase ; drug effects ; Humans ; Mitosis ; drug effects ; Protein-Serine-Threonine Kinases ; biosynthesis ; genetics ; Proto-Oncogene Proteins ; biosynthesis ; genetics ; RNA Interference ; RNA, Messenger ; biosynthesis ; genetics ; RNA, Small Interfering ; genetics ; pharmacology ; Stomach Neoplasms ; enzymology ; metabolism ; pathology ; Transfection

AIMTo compare the effects of the non-mitogenetic human acidic fibroblast growth factor (nmhaFGF) and the human acidic fibroblast growth factor (haFGF) on the proliferation and MAPK signal transduction pathway of the malignant tumor cell and to study the clinical safety of nmhaFGF.

METHODSThe mammary tumor cells (MCF-7) were treated with haFGF and nmhaFGF separately. The mitogenic activities of both haFGF and nmhaFGF were detected by MTT method and the cell cycle was analyzed by flow cytometer (FCM). The expression levels of the signal proteins, Grb2 (growth factor receptor bound 2) and ERK1/2 (extracellular signal-regulated kinase 1/2), were detected by semi-quantitative Western blotting method.

RESULTSThe mitogenic activity of nmhaFGF was obviously lower than that of haFGF. The activity of nmhaFGF was weaker than that of the haFGF. The ratio of G1/G0, G2/M of haFGF was markedly lower than that of nmhaFGF and control group, and was reverse in S phase. The expression levels of both Grb2 and ERK1/2 of the nmhaFGF treated group were lower than that of the haFGF treated group and approaching the control group.

CONCLUSIONThe mitogenic activity of the nmhaFGF decreased remarkably. Its mechanism probably via down-regulation of the expression of the signal moleculars, MAPK-ERK1/2 and Grb2.

Breast Neoplasms ; metabolism ; pathology ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Down-Regulation ; Female ; Fibroblast Growth Factor 1 ; genetics ; pharmacology ; GRB2 Adaptor Protein ; metabolism ; Humans ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Mitosis ; drug effects ; Mutation

OBJECTIVETo investigate the role of STK15 in regulating mitosis of gastric cancer cells (MKN45) by gene silencing through RNA interference mechanism.

METHODSRNA interference technique was used to inhibit STK15 expression in MKN45 cells. The expression levels of STK15 mRNA and protein were measured by real-time quantitative RT-PCR and Western blot respectively and cell morphological changes were investigated by reverse microscopy. In addition, cell cycle distribution and cellular proliferation were determined by flow-cytometry and MTT assay respectively. Finally, the mitotic phenotype of MKN45 cells was studied by immunofluorescence staining and confocal microscopy.

RESULTSSilencing of STK15 gene by RNA interference was confirmed by marked decrease of STK15 mRNA and protein levels in the treated MKN45 cells. This silencing correlated with rounding of the cells, decreasing of DNA content in G(2) phase (P < 0.05) and a lowered proliferation index (P < 0.05), along with alterations of mitotic phenotype of MKN45 (P < 0.05).

CONCLUSIONSTK15 gene may play a key role in regulating cellular mitosis and its inhibition by RNA interference leading to mitosis arrest in MKN45 cells.

Adenocarcinoma ; metabolism ; pathology ; Aurora Kinase A ; Aurora Kinases ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; DNA, Neoplasm ; metabolism ; Gene Silencing ; Humans ; Mitosis ; drug effects ; Protein-Serine-Threonine Kinases ; biosynthesis ; genetics ; RNA Interference ; RNA, Messenger ; biosynthesis ; genetics ; RNA, Small Interfering ; pharmacology ; Stomach Neoplasms ; metabolism ; pathology