Objective:To investigate the mechanism of miR-128 on the expression of AK2 protein through the STAT3 signaling pathway on the biological behavior of cervical cancer cells .Methods: The expression of AK2 and miR-128 in cervical cancer tissues and cells was detected by qPCR and Western blot .Double luciferase assay was used to detect the interaction between miR-128 and AK2.CCK-8 proliferation assay was used to detect the effect of miR-128 on the enhancement of cervical cancer cells .The effect of miR-128 on the tumorigenesis of cervical cancer cells was exa mined by tumor formation test in nude mice .Western blot was used to detect the effect of miR-128 on STAT3 signal pathway protein level .Western blot was used to detect the inhibitory effect of miR-128 on p-STAT3 by overexpressing AK2 protein.Results:The expression level of AK2 in cervical cancer tissues was higher than that in normal cervical tissues,and the expression level of miR-128 in cervical cancer C33a cells was lower.Double luciferase assay confirmed that miR-128 could directly target the expression of AK 2.CCK-8 proliferation test showed that miR-128 could inhibit the proliferation of cervical cancer cell lines .In vivo tumorigenesis test showed that the increase of miR-128 could inhibit the tumorigenesis ability of cervical cancer cells[volume(3.05±0.35)cm3 vs (0.86±0.11)cm3,P=0.031;weight(3.26±0.39)g vs (0.89±0.15)g,P=0.016 ];Western blot showed that miR-128 could inhibit the activation of p-STAT [ ( 42.12 ±6.28 )% vs ( 91.25 ±9.29 )%, P<0.05 ] ,while the overexpression of AK 2 could reverse the inhibitory effect of miR-128 on p-STAT.Conclusion: miR-128 is used to regulate the expression of AK 2 and regulate the biological behavior of cervical cancer cells through the activation of STAT 3 pathway.

OBJECTIVEThe purpose of the present study was to investigate the in vitro effects of baicalin on induction of apoptosis in human prostate cancer cell line DU145.

METHODHuman prostate cancer cell line DU145 was treated with different concentration of baicalin in vitro. The apoptosis rate was determined by FACS analysis, cell cycle distribution was detected by flow cytometry, morphological changes and protein analysis were determined by means of electron microscope techniqueand immunohistochemical techniquerespectively.

RESULT50micromol x L(-1) and 125 micromol x L(-1) of baicalin dose-dependently induced apoptosis and inhibited the proliferation of prostate cancer cell DU145 in a dose and time-dependent manner. DNA flow cytometric analysis indicated that baicalin induced a arrest in G1 phase, showing a typical apoptosis peak. Electron microscopy detected a characteristic appearance of the apoptotic cells morphology. Immunohistochemical analysis revealed that induction of apoptosis by ways of inhibition of the bcl-2, loss of the Bax, and upregulation of Fas.

CONCLUSIONThe results indicate that baicalin may induce apoptosis and inhibit proliferation of prostate cancer cells, and has direct anti-tumor effects on human prostate cancer cells.

Antineoplastic Agents, Phytogenic ; pharmacology ; Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Flavonoids ; isolation & purification ; pharmacology ; G1 Phase ; Humans ; Male ; Plants, Medicinal ; chemistry ; Prostatic Neoplasms ; metabolism ; pathology ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Scutellaria ; chemistry ; bcl-2-Associated X Protein ; fas Receptor ; metabolism

The purpose of this study was to investigate the effect of the overexpression of β(1)-adrenoceptor (β(1)-AR) on the contractile function and cell survival of rat cardiomyocytes injured by isoprenaline (ISO). The rat cardiomyocytes were isolated using the collagenase perfusion method and then transfected with β(1)-AR gene using adenoviruses vector. Four hours after the infection, the rat cardiomyocytes were treated with ISO for 24 h to imitate the high catecholamine levels of chronic heart failure. Western blot was performed to measure the protein expression of β(1)-AR. The percentages of rod cells were measured to test cell survival. Video-based edge-detection system was used to measure the contractile function of the cardiomyocytes. The results indicated that the expression of β(1)-AR in β(1)-AR-transfected cardiomyocytes was significantly increased compared with that in control group (P<0.01). Meanwhile, β(1)-AR transfection also increased β(1)-AR protein levels in ISO-injured cardiomyocytes. The cardiomyocyte survival was significantly decreased in ISO group compared with that in control group. β(1)-AR-transfection alone had no effect on cardiomyocyte survival in β(1)-AR group, but it further decreased cardiomyocyte survival in β(1)-AR+ISO group. Contractile amplitudes of ISO-injured cardiomyocytes were significantly decreased regardless of whether they were transfected with β(1)-AR or not, although β(1)-AR-transfected cardiomyocytes showed significantly increased contractile function compared with control group (P<0.05). These results suggest that the overexpression of β(1)-AR has no significant protective effect on rat cardiomyocytes injured by ISO.
Animals ; Cell Survival ; Cells, Cultured ; Female ; Heart Failure ; metabolism ; Isoproterenol ; pharmacology ; Male ; Myocardial Contraction ; drug effects ; Myocytes, Cardiac ; cytology ; metabolism ; pathology ; Rats ; Rats, Sprague-Dawley ; Receptors, Adrenergic, beta-1 ; genetics ; metabolism ; Transfection

OBJECTIVETo investigate the therapeutic effect, long term survival and side effect on NSCLC patients treated with nadaplatin combined with paclitaxol and cisplatin combined with paclitaxol.

METHODSNSCLC patients with stage IIIB or IV were randomized into two groups in this prospective clinical study. TN group: nadaplatin 30 mg/m2 dl-3, paclitaxol 175 mg/m2 dl, repeated every 4 weeks. TP group: DDP 30 mg/m2 dl-3, paclitaxol 175 mg/m2 dl, repeated every 4 weeks.

RESULTSSixty patients were enrolled and 57 were evaluable with 30 in TN group and 27 in TP group. The overall response rate were 43.3% vs. 48.1% (P = 0.716), and the disease control rate were 86.7% vs. 88.8% in TN and TP group (P = 0.799), respectively. The median survival time was 14.3 vs. 13.0 months, and the 1- and 2-year survival rate was 62.5% vs. 59.1%, 0% vs. 5.8% in TN and TP group (P = 0.839), respectively. The rates of neutropenia and thrombocytopenia were similar in TN and TP groups whereas more patients in TP group than in TN group suffered from anemia (38.5% vs. 17.5%, P = 0.001), nausea and vomiting (82.6% vs. 35.6%, P = 0.000), fatigue (35.9% vs. 14.1%, P = 0.000) and peripheral neurotoxicity (50.0% vs. 21.9%, calculated by case, P = 0.023).

CONCLUSIONNadaplatin combined with paclitaxol is an effective treatment regimen for NSCLC patients. When compared with similar regimen with cisplatin, the response rate and survival were similar; however, nadaplatin regimen shows some superiority as regards some treatment side effect.

Adult ; Aged ; Anemia ; chemically induced ; Antineoplastic Combined Chemotherapy Protocols ; adverse effects ; therapeutic use ; Carcinoma, Non-Small-Cell Lung ; drug therapy ; pathology ; Cisplatin ; administration & dosage ; adverse effects ; Female ; Humans ; Lung Neoplasms ; drug therapy ; pathology ; Male ; Middle Aged ; Neoplasm Staging ; Neutropenia ; chemically induced ; Organoplatinum Compounds ; administration & dosage ; adverse effects ; Paclitaxel ; administration & dosage ; adverse effects ; Prospective Studies ; Remission Induction ; Survival Analysis ; Thrombocytopenia ; chemically induced ; Treatment Outcome

OBJECTIVETo investigate the effects of ARNT2 on invasion and migration of HCCLM6 cells.

METHODSFour short hairpin oligos targeting to ARNT2 were s cloned into the pLVTHM vector. Lentiviral vectors shRNA-ARNT2i, pCMV-dR8.74 and pMD2G were cotransfected into 293T cells using Lipofectamine 2000. HCCLM6 was infected with virus supernatant. ARNT2 mRNA and protein expressions were detected using quantitative Real time-PCR and Western blot, respectively. The invasion and migration of HCCLM6 cells were evaluated using wound healing assay and cell invasion assay in vitro. Statistical analysis was performed with SPSS 16.0.

RESULTSThe relative mRNA levels of ARNT2 were 0.154+/-0.024, 0.860+/-0.145, 1.004+/-0.009 in shRNA-ARNT2i virus infected HCCLM6 cells, mock-infected cells and control vector virus infected cells (F = 113.14, P more than 0.01). The expression of ARNT2 at protein level was 16.45+/-1.6, 44.56+/-2.07 in the HCCLM6 cells infected with shRNA-ARNT2i virus and negative control vector virus, respectively (t = 18.58, P less than 0.01). The scrape wound of HCCLM6 cells infected with shRNA-ARNT2i virus healed faster than cells infected with control vector virus or mock-infected cells. The number of cells invading through Matrigel was higher in the HCCLM6 cells infected with shRNA-ARNT2i virus (13.25+/-1.04) than that in mock-infected HCCLM6 cells and the HCCLM6 cells infected with negative control vector virus (6.50+/-2.56, 6.75+/-2.05) (F = 29.645, P less than 0.01).

CONCLUSIONInhibition of ARNT2 gene promotes the invasion and migration of HCCLM6 cells.

Aryl Hydrocarbon Receptor Nuclear Translocator ; genetics ; metabolism ; Basic Helix-Loop-Helix Transcription Factors ; genetics ; metabolism ; Blotting, Western ; Carcinoma, Hepatocellular ; genetics ; metabolism ; pathology ; Cell Line, Tumor ; Cell Movement ; Gene Expression Regulation, Neoplastic ; Genetic Vectors ; genetics ; Humans ; Lentivirus ; genetics ; Liver Neoplasms ; genetics ; metabolism ; pathology ; Neoplasm Invasiveness ; Polymerase Chain Reaction ; methods ; RNA, Messenger ; genetics ; metabolism ; RNA, Small Interfering ; genetics ; Transfection

OBJECTIVETo understand and master the situation in which enterovirus caused hand-foot-and-mouth disease (HFMD) in Xuzhou district in 2009 so as to provide scientific basis for the control and prevention of hand-foot-and-mouth disease.

METHODSThe researchers adopted fluorescence RT-PCR method to detect EV and EV71 as well as the CA16 specificity RNA from 222 samples of anal swabs and oropharyngeal swabs from the 240 cases who were diagnosed clinically as hand-foot-mouth disease infected by enterovirus. Also, the researchers conducted EV71-IgM antibody detection on 114 samples of acute phase serum with ELA method.

RESULTSAmong the 240 enterovirus infected patients, the total EV infection rate is 72.50% (174/240), among which EV71 infection rate is 57.92% (139/240), CoxA16 infection rate is 9.17% (22/240), and other EV infection rate is 5.42% (13/240). The EV71-RNA positive rate of the samples of 222 anus swabs among the 240 suspected enterovirus infected patients is 45.94% (102/222), the samples of swallow swab EV71-RNA positive rate is 25.68% (57/222) and the EV71-IgM antibody positive rate of 114 samples of acute phase serum is 86.84% (99/114). The EV71-RNA positive rate of oropharyngeal swabs of 254 healthy children is 1.57% (4/254) , and no CoxA16-RNA was detected. In the oropharyngeal swabs of 54 close contacts (medical personnel), the EV-RNA detected is negative. The positive rate of EV71-IgM antibody of the 258 healthy children's serum samples is 2.71% (7/258).

CONCLUSIONThe widespreading of hand-foot-mouth disease in Xuzhou district is caused mainly by type 71 enterovirus. Inapparent infection of type 71 enterovirus exists among children under the age of 3 during the time of widespreading period and IgM antibody develops in them. It is difficult for adults to be infected by EV71 even if they contact the contagion source closely. The positive rate of EV71-IgM antibody in the samples of acute phase serum of suspected cases is the highest (86.84%), and the second highest is the positive rate of RNA of EV71 of anal swabs (45.94%) and of the EV71 of oropharyngeal swabs (25.68%). ELA reagent kit is used in the early diagnosis of EV71 infection for it is easy to operate, fast and economic, so, it is worth popularizing in the grass-root medical units.

Child, Preschool ; China ; Enterovirus A, Human ; genetics ; isolation & purification ; Female ; Hand, Foot and Mouth Disease ; virology ; Humans ; Infant ; Male

To develop a fed-batch fementation process of E. coli TOP10 containing a recombinant plasmid pBAD/HBs Fab. Cells were grown in semi-defined medium at 37 degrees C, and the feed operation using glycerol as carbon source was performed when dissolved oxygen increased. When the target cell concentration reached to 64g/L, arabinose was added to a final concentration of 0.02%. Cells were grown for another 5h with the culture temperature decreased from 37 degrees C to 30 degrees C. In the whole process, cell growth was monitored by measuring OD600 of samples taken at 1/2h intervals and the dissolved oxygen was kept above 30%. After the fementation, E. coli pellets were collected for purification of Fab protein. The specificity of Fab protein was confirmed by Western blot, and binding activity to HBsAg was verified by Dot blot. Cell concentration we got is 96g wet bacteria per liter, the Fab protein is about 6% of total protein of the host, that is 80mg per liter. Stable fermentation parameters were obtained for fermentation to improve productivity of the Fab protein. The Fab protein was produced in the form of soluble biologically active protein, it's better than inclusion bodies from which biologically active protein can only be recovered by complicated and costly denaturation and refolding process.
Blotting, Western ; Escherichia coli ; genetics ; metabolism ; Fermentation ; physiology ; Hepatitis B Surface Antigens ; immunology ; Immunoglobulin Fab Fragments ; biosynthesis ; genetics ; immunology ; Temperature

OBJECTIVETo investigate the effects of adenovirus-mediated PTEN and P27 on the invasion of PC-3 in vitro and angiogenesis, along with their synergy in the treatment of prostate cancer.

METHODSRecombinant adenovirus vectors of the human tumor suppressor genes PTEN and P27 were constructed. The replication-incompetent recombinant adenovirus was packaged and propagated in HEK293 cells. The viral titer was examined by plaque assay and the mRNA and protein expressions of PTEN and P27 in human prostate cancer cell line PC-3 infected with Ad-PTEN and Ad-P27 were determined by RT-PCR and Western blot respectively. The invasion of PC-3 cells in vitro was examined by Boyden chamber assay. MTT assay was used to testify the effect of supernatant from PC-3 infected with Ad-PTEN and Ad-P27 on the proliferation of endothelial cells ECV-304 and the CAM test was used to testify the effect of PTEN and P27 on angiogenesis. The difference between the combined therapy group and the single gene therapy group was also examined.

RESULTSThe viral titers of Ad-PTEN and Ad-P27 were 1.8 x 10(7) pfu/ml and 1.2 x 10(9) pfu/ml respectively. Adenovirus infection verified that the mRNA and protein expression of PTEN and P27 were steady in human PC-3 cells. The invasion in vitro of PC-3 cells was significantly inhibited by infection with Ad-PTEN or/and Ad-P27. CAM and MTT assays of ECV-304 confirmed that the supernatant from PC-3 cells infected with Ad-PTEN or/and Ad-P27 could inhibit the angiogenesis effectively. There was a significant difference between the combined therapy group and the single gene therapy group.

CONCLUSIONThe combined gene therapy of Ad-PTEN and Ad-P27 plays a synergistic role in inhibiting the invasiveness of PC-3 cells and angiogenesis.

Adenoviridae ; genetics ; Cell Line, Tumor ; Cyclin-Dependent Kinase Inhibitor p27 ; genetics ; Humans ; Male ; Neoplasm Invasiveness ; PTEN Phosphohydrolase ; genetics ; Prostatic Neoplasms ; blood supply ; pathology ; Transfection

This study is to determine age-specific prostate-specific antigen (PSA) distributions in Chinese men without prostate cancer (PC) and to recommend reference ranges for this population after comparison with other studies. From September 2003 to December 2006, 9 374 adult men aged from 18 to 96 years agreed to participate in the study. After all cases of PC were excluded, 8 422 adult men participated in statistical analysis and were divided into five age groups. Simple descriptive statistical analyses were carried out and quartiles and 95th percentiles were calculated for each age group. The age-specific PSA reference ranges are as follows: 40-49 years, 2.15 ng mL(-1); 50-59 years, 3.20 ng mL(-1); 60-69 years, 4.10 ng mL(-1); 70-79 years, 5.37 ng mL(-1). The results indicate that the ethnic differences in PSA levels are obvious. The currently adopted Oesterling's age-specific PSA reference ranges are not appropriate for Chinese men. The reference ranges of this study should be more suitable to Chinese men.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Aging ; blood ; ethnology ; China ; Humans ; Male ; Middle Aged ; Prostate-Specific Antigen ; blood ; Prostatic Neoplasms ; blood ; diagnosis ; ethnology ; Reference Values ; Retrospective Studies ; Young Adult

OBJECTIVESTo investigate whether the human PC-3 cell infected with recombinant Ad-PTEN and Ad-p27Kip1 can steadily produce PTEN and p27Kip1 protein and change the biologic behaviors such as cell proliferation, cell cycle and apoptosis. The synergistic effect of PTEN and p27Kip1 on the therapy for prostate cancer has also been investigated.

METHODSWe constructed recombinant adenovirus vector of human tumor suppressor gene PTEN and p27Kip1. The viral titer was examined by plaque assay and the mRNA and protein expressions of PTEN and p27Kip1 in human prostate cancer cell line PC-3 infected with Ad-PTEN and Ad-p27Kip1 were determined by RT-PCR and Western blot respectively. MTT assay was used to determine the effect of PTEN and p27Kip1 on growth and proliferation of PC-3 cell; the change of cell cycle and apoptosis was examined by flow cytometry, and to compare between the combined therapy group and single gene therapy group.

RESULTSThe viral titers of Ad-PTEN and Ad-p27Kip1 were 1.8 x 10(7) pfu/ml and 1.2 x 10(9) pfu/ml respectively. After infected by adenovirus, it had been verified that the mRNA and protein expression of PTEN and p27Kip1 were steady in human PC-3 cell. Ad-PTEN and Ad-p27 Kip1 inhibited the growth and proliferation of PC-3 cells. The progression of cell cycle of PC-3 cell was arrested in G(0)-G(1) phase, meanwhile the apoptosis rate of PC-3 was also affected after Ad-PTEN or/and Ad-p27 Kip1 infected. There was significant difference between combined therapy group and single gene therapy group.

CONCLUSIONThe recombinant Ad-PTEN and Ad-p27Kip1 vector were constructed successfully and the expression of specific PTEN and p27Kip1 was high, steadily in PC-3 cell line. These results suggested that combination of PTEN with p27Kip1 has an application value in treatment of prostate cancer in future.

Adenoviridae ; genetics ; Apoptosis ; drug effects ; Cell Division ; drug effects ; Cell Line, Tumor ; Chromosomes, Human, Pair 10 ; genetics ; Cyclin-Dependent Kinase Inhibitor p27 ; Gene Deletion ; Genetic Therapy ; Genetic Vectors ; Humans ; Intracellular Signaling Peptides and Proteins ; genetics ; pharmacology ; Male ; PTEN Phosphohydrolase ; genetics ; pharmacology ; Prostatic Neoplasms ; genetics ; physiopathology ; therapy ; Transfection