OBJECTIVETo investigate the impact of ethanol extracts from Sedum sarmentosum (ESB) on STAT-3 signaling and its probable molecular mechanism in inducing apoptosis.

METHODMTT assay was used to detect the impact of ESB on HepG2 cell proliferation. FITC-Annexin V-FITC /PI double-labeling were used to investigate the impact on hepatoma carcinoma cell apoptosis. Western blot analysis was used to test the expression levels of cell apoptosis-related proteins Caspase-3, Caspase-9, PARP, P-STAT-3 (Tyr705) , STAT-3, Bcl-2, Mcl-1.

RESULTESB could notably inhibit proliferation of HepG2 cells, and induce HepG2 cell apoptosis, with the dose-dependent inhibitory effect. In addition, ESB could inhibit STAT-3 signaling, down-regulate Mcl-1 and Bcl-2 expressions, and induce degradation/activation of apoptosis-related proteins Caspase-3 and Caspase-9 and PARP degradation in a dose-dependent manner.

CONCLUSIONESB inhibits HepG2 cell proliferation and induces apoptosis by inhibiting STAT-3 signaling and Mcl-1 and Bcl-2 expressions.

Apoptosis ; drug effects ; Blotting, Western ; Caspase 3 ; metabolism ; Caspase 9 ; metabolism ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Ethanol ; chemistry ; Flow Cytometry ; Hep G2 Cells ; Humans ; Myeloid Cell Leukemia Sequence 1 Protein ; metabolism ; Plant Extracts ; chemistry ; pharmacology ; Poly(ADP-ribose) Polymerases ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; STAT3 Transcription Factor ; metabolism ; Sedum ; chemistry ; Signal Transduction ; drug effects ; Time Factors

Objective To study the risk factors in patients with trauma accompanied by multiple organ dysfunction syndrome.Method The data of 107 patients with trauma in ICU,Tongji Hospital,Tongji Medical College,Huazhong University of Science and Technology,were retrospectively studied.All patients were divided into 2 groups:MODS group and non-MODS group.The clinical and laboratory,results,ISS score,APACHEⅢscore and GCS score were studied.Results There were no differences on gender,age and mobility of shock between the two groups.There were significant differences on the treatment of shock,ISS score,APACHEⅢscore,CCS score,the levels of blood sugar and platelet between two groups.The Logistic regression analysis showed the main risk factors were treatment of shock,ISS score and APACHEⅢscore.Conclusions The risk factors in patients with trauma accompanied by multiple organ dysfunction syndrome were the treatment of shock, ISS score and APACHEⅢscore.

Objective To investigate the effect of mesenchymal stem cells (MSC) on the activity of nuclear factor (NF)-?B in rats with myocardial infarction.Methods MSC were isolated from SD rats (120—150 g in weight).SD rats (180—200 g in weight) were subjected to MI by left coronary artery occlusion,and were allo- cated into three groups randomly:1)sham group (without ligation of the artery,n=8);2)injection of PBS solu- tion (n=8);3)injection of MSC (n=8).MSC or PBS solution was injected into myocardium from epicardium instantly after MI models were established.Four weeks after transplantation,cardiac function was evaluated u- sing physiological recorder.Western blot were performed to investigate the nuclear factor-? activity.The ex- pressions of tumor necrosis factor (TNF)-? and interleukin (IL)-6 were evaluated by RT-PCR and Western blot. Results 1)Mortality was 20%(2/10) in sham group,33.3%(4/12) in PBS group and 20%(2/10) in MSC group with no statistic differences between them(P=0.646).2) Hemodynamic measurements showed that MSC trans- plantation caused significant improvement in cardiac function,comparing with MI+PBS group.3) MSC inhibi- ted the activities of NF-?B in myocardium and down-regulated the expression of TNF-? and IL-6 in mRNA and protein level.Conclusion Transplantation of MSC improved cardiac function in MI rats,which may partly at- tribute to their immuno-inflammatory regulation mechanism.

The aim of study was to investigate the killing effect of double suicide gene system mediated by retroviral vector on K562 cells in vivo and ex vivo. CDglyTK gene was transfected into PA317 cells by using lipofectamine. K562 cells were infected with viral supernatant. K562/CDglyTK cells were treated with 5-fluorocytosine (5-FC) and/or ganciclovir (GCV). Mice were randomly divided into three groups: tumor formation, tumor inhibition and tumor therapy. Each mouse was implanted with K562/CDglyTK cells or K562 cells. The results indicated that the killing effect of 5-FC in combination with GCV on K562/CDglyTK was more significant than using 5-FC or GCV alone. In vivo study showed that after being injected subcutaneously with K562 cells and K562/CDglyTK cells, there was not obvious difference in tumor formation rate of mice, 5-FC + GCV could suppress tumor formation of the K562/CDglyTK cells. After being treated with 5-FC and GCV, the median tumor volume of mice implanted with K562/CDglyTK cells decreased obviously, compared with the control group. Their median survival was significantly prolonged. It is concluded that double suicide genes are more effective for killing effect on K562 cells in vivo and in ex vivo. It may be applicable to clinical gene therapy.
Cytosine Deaminase ; genetics ; Flucytosine ; pharmacology ; Ganciclovir ; pharmacology ; Genes, Transgenic, Suicide ; genetics ; Genetic Therapy ; Genetic Vectors ; genetics ; Humans ; K562 Cells ; Protein-Tyrosine Kinases ; genetics ; Receptor Protein-Tyrosine Kinases ; biosynthesis ; genetics ; Recombinant Fusion Proteins ; genetics ; Recombination, Genetic ; Retroviridae ; genetics

Chronic myelogenous leukemia (CML) is a clonal myeloproliferative disease of transformed hematopoietic progenitor cells. In order to investigate the role of sphingosine kinase-1 (SphK-1)/sphingosine 1-phosphate (S1P) signal pathway in the expression of CML cells, and to explore whether P210(bcr/abl) involved is activating SphK-1/S1P signal pathwey, the expressions of SphK-1 and S1P receptor mRNA in bcr/abl positive K562 cells and bcr/abl positive primary CML cells were detected by RT-PCR, the imatinib mesylate, the specific inhibitor of P210(bcr/abl) was employed to inhibit the P210(bcr/abl) tyrosine kinases of K562 cells and CML primary cells, and then the intracellular SphK-1 activity was assayed. The results indicated that after being cultured with 2.5 micromol/L imatinib mesylate for 0.5, 2, 6, 24 and 48 hours, the intensions of inhibiting SphK-1 activity were 0.007%, 38.9%, 34.6%, 28.1% and 76.1% resepectively. SphK-1 activity in CML cells also was reduced by 2.5 micromol/L imatinib mesylate (16.8% - 41.9% decrease). It is concluded that the CML cells express SphK-1 and different S1P receptor, and P210(bcr/abl) fusion protein in CML cells can activate SphK-1.
Benzamides ; Fusion Proteins, bcr-abl ; genetics ; metabolism ; Humans ; Imatinib Mesylate ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; genetics ; metabolism ; Lysophospholipids ; genetics ; metabolism ; Phosphotransferases (Alcohol Group Acceptor) ; genetics ; metabolism ; Piperazines ; pharmacology ; Pyrimidines ; pharmacology ; RNA, Messenger ; genetics ; metabolism ; Signal Transduction ; genetics ; Sphingosine ; analogs & derivatives ; genetics ; metabolism

The aim of this research was to understand the influence of rhG-CSF on the sphingosine kinase (SphK) activity of monocytes. The peripheral blood monocytes were collected from 6 peripheral blood progenitor cell donors on the fifth day of mobilization with rhG-CSF and from 5 blood donors' buffy coats. The mRNA expressions of monocyte G-CSF receptor and SphK were tested with RT-PCR. The changes of SphK activity of monocytes were assayed after being treated with rhG-CSF. The results showed that the two kinds monocytes collected from both blood donors and peripheral blood progenitor cell donors mobilized with rhG-CSF expressed mRNA of G-CSF receptor and SphK. The SphK activity of monocytes collected from blood donors was not changed significantly after being treated with rhG-CSF (P > 0.05). The SphK activity of monocytes collected from peripheral blood progenitor cell donors transiently increased by (39.6 - 87.2)% after being treated by means of rhG-CSF (P < 0.05) without obviously dose-dependent effect. It is concluded that the SphK activity of monocytes collected from peripheral blood progenitor cell donors can be activated by rhG-CSF.
Granulocyte Colony-Stimulating Factor ; pharmacology ; Hematopoietic Stem Cell Mobilization ; Humans ; Monocytes ; cytology ; enzymology ; Phosphotransferases (Alcohol Group Acceptor) ; drug effects ; metabolism ; Receptors, Granulocyte Colony-Stimulating Factor ; biosynthesis ; genetics ; Recombinant Proteins

The study was to understand the impact on the proliferation and cytotoxicity of donor's T cells during mobilization with rhG-CSF. The peripheral blood mononuclear cells (PBMNC) were collected from 15 donors before mobilization and on fifth day of mobilization with rhG-CSF. After the PBMNC were activated with 500 ng/ml of CD3 monoclonal antibody and 500 microg/ml of rhIL-2 for 96 hours, the activated T cells were collected for testing proliferation, cytotoxicity, Fas expression, perforin and Fas ligand (FasL) mRNA expression, the IFN-gamma concentration in the culture medium of the activated T cells was determined by radioimmunoassay. The results showed that the proliferation activity of T lymphocytes and the cytotoxicity of T cells activated with CD3 monoclonal antibody and rhIL-2 were reduced markedly after mobilization with rhG-CSF (P < 0.05). The Fas molecule expression in the activated T cells was very high both before and after mobilization with rhG-CSF (P > 0.10). The activated T cells expressed perforin mRNA and didn't express FasL mRNA both before and after mobilization with rhG-CSF. The concentration of IFN-gamma in the culture medium of the activated T cells decreased significantly after mobilization with rhG-CSF (P < 0.01). It is concluded that activity of proliferation and cytotoxicity of donor's T cells is impaired after mobilization with rhG-CSF.
Adolescent ; Adult ; Cell Proliferation ; drug effects ; Cells, Cultured ; Fas Ligand Protein ; biosynthesis ; genetics ; Female ; Granulocyte-Macrophage Colony-Stimulating Factor ; administration & dosage ; pharmacology ; Hematopoietic Stem Cell Mobilization ; methods ; Humans ; Male ; Middle Aged ; RNA, Messenger ; biosynthesis ; genetics ; Recombinant Proteins ; T-Lymphocytes ; cytology ; drug effects ; T-Lymphocytes, Cytotoxic ; drug effects ; immunology ; fas Receptor ; biosynthesis ; genetics

BACKGROUND: Alloying can improve the corrosion resistance and slow the degradation of pure magnesium. In addition, increasing studies have shown that zinc has good antitumor effect. OBJECTIVE: To evaluate the effect of zinc-containing magnesium alloy on the proliferation and apoptosis of human osteosarcoma U2OS cells in vitro.METHODS: The corrosion resistance of pure magnesium and different zinc-containing (2%, 4%, 6%) magnesium alloys were observed and compared by the hydrogen release assay in the Hank's solution. MTT assay was used to examine the effects of different zinc-containing magnesium alloy extractions on the proliferation of U2OS cells (or MC3T3-E1 cells) after co-culture of 1, 3, 5 days. After 24-hour co-culture with pure magnesium, different zinc-containing magnesium alloys and titanium alloy extractions, the apoptotic rates of U2OS cells were analyzed by flow cytometry, and the expression of apoptosis-related proteins were detected by western blot assay.RESULTS AND CONCLUSION: The corrosion resistance of pure magnesium was improved after addition of Zn within the initial 100 hours, and the magnesium alloy containing 4% zinc exhibited the optimal corrosion resistance. Different zinc-containing magnesium alloy extractions obviously inhibited the U2OS proliferation in a zinc level-depended manner, and the cytotoxicity of different zinc-containing magnesium alloy extractions to MC3T3-E1 was graded 0-1. Different zinc-containing magnesium alloy extractions could also induce obvious apoptosis in U2OS cells in a zinc level-depended manner. Different zinc-containing magnesium alloy extractions, especially the magnesium alloy containing 4% zinc, up-regulated the expression of p53 and Bax proteins and down-regulated the expression of Bcl-2 protein in U2OS cells, leading to the disorder of Bcl-2/Bax. These findings suggest that different zinc-containing magnesium alloy extractions can inhibit the proliferation and induce apoptosis of U2OS cells in vitro.

Aim To observe the effect of interventional therapy of 5-fluorouracil(5-Fu), mitomycin C (MMC) and adriamycin (ADM) on solid malignant turmors in association with aterial infusion of NaHCO3.Methods Patients were randomly divided into two groups.With Seldinger technique,through the femoral atery to tumor atery.The patients in the control group were infused by anticarcinoma agents simply ,and patients in the treatment group were initially infused by NaHCO3,and then by NS 30 ml and anticarcinoma agents seperately. Results Partial remission (PR) in the group treated with NaHCO3 and anticarcinoma agents was significantly higher than in the group treated simply with anticarcinoma agents.Conclusion Aterial infusion of NaHCO3 into malignant tumors can increase the efficacy of ADM,MMC and 5-Fu.

OBJECTIVETo investigate the effects of hypoxia induced by cobalt chloride on the expression and the activity of matrix metalloproteinase-2 (MMP-2) in rat hepatic stellate cells (HSC-T6) and to clarify the possible mechanisms.

METHODSHSC-T6 cell line was grown in Dulbecco's modified Eagle medium with 10% fetal calf serum at 37 degrees C and 5% CO2. When reaching confluence, the cells were incubated with serum-free medium in the presence of cobalt chloride (0, 50, 100, 200 micromol/L) for six hours, and then the supernatant and the cells were harvested. The expression of the MMP-2 mRNA and HIF-1alpha protein in HSC-T6 cells was detected using RT-PCR and Western blot respectively. The activity of the MMP-2 in the supernatant was detected by zymography. The binding reaction between HIF-1a protein and MMP-2 gene sequence was investigated by electrophoresis mobility shift assay.

RESULTSWhen the concentration of CoCl2 increased from 0 micromol/L to 200 micromol/L, the expressions of MMP-2 mRNA (the rate of light density) were increased from 0.53+0.12 to 1.57+0.11 and the differences among these four groups were significant (F=34.21). The activity of MMP-2 (the value of light density*band area) decreased gradually from 84.49+5.38 to 53.70+3.42, and the differences among these four groups were also significant (F=29.54). The expressions of HIF-1a were increased gradually with the increase of the CoCl2 concentration. The shift band in the lane of the nuclear protein extraction and the MMP-2 probe containing hypoxia response element showed delays when compared with the lane of the sole probe, and the binding was partially abolished when competing sense oligonucleotides were used.

CONCLUSIONSOur results suggest that chemical hypoxia can up-regulate the expression of MMP-2 mRNA and decrease the activity of the enzyme. HIF-1alpha may play a part in the regulation of MMP-2 transcription under hypoxic conditions.

Animals ; Cell Hypoxia ; Cell Line ; Cobalt ; pharmacology ; Hepatic Stellate Cells ; enzymology ; Hypoxia ; metabolism ; Hypoxia-Inducible Factor 1, alpha Subunit ; metabolism ; Matrix Metalloproteinase 2 ; metabolism ; RNA, Messenger ; genetics ; Rats