Objective: To observe the effects of electroacupuncture (EA) on the protein and gene expressions of Bax, Caspase-3 and Bcl-2 in cerebral cortex of type 2 diabetic rats with cognitive impairment (CI), and to explore the mechanism of EA in improving the learning and memory abilities. Methods: A total of 100 Sprague-Dawley (SD) rats were divided into a normal group (n=10) and a model group (n=90) by random number table method. Rats in the model group were intraperitoneally injected with a small dose of streptozotocin (STZ) to establish the type 2 diabetic models, after being fed with high-fat and high-sugar diet for 1 month. Twenty CI rats were selected from the 50 successful model rats by the Morris water maze (MWM) test and randomly divided into a model group and an EA group according to the blood glucose level and MWM data (n=10). Rats in the EA group received acupuncture at Zusanli (ST 36), Neiting (ST 44) and Yishu (Extra), of which Zusanli (ST 36) and Neiting (ST 44) were stimulated by EA apparatus, 20 min/time, once a day for 6 d a week and 4 consecutive weeks. The rats in the model and the normal groups were fixed without treatment. After 4-week treatment, the random blood glucose level of the rats was measured; the learning and memory abilities of rats were measured by MWM; terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay was used to detect apoptotic cells; Western blot (WB) and real-time quantitative polymerase chain reaction (RT-qPCR) were used to detect the protein and gene expressions of Bax, Caspase-3 and Bcl-2 in cerebral cortex. Results: After modeling, the random blood glucose level and the escape latency tested by MWM were significantly increased, and the number of crossing the platform tested by the MWM was decreased in the EA and model groups, and were significantly different from those in the normal group (P<0.05 or P<0.01), while the differences between the model group and the EA group were not statistically significant (all P>0.05). After 4-week treatment, the random glucose level and the escape latency tested by MWM were significantly increased (both P<0.05), and the number of crossing the original platform tested by the MWM was significantly reduced (P<0.01), the protein and gene expressions of Bax and Caspase-3 were significantly increased (all P<0.001), the protein and gene expressions of Bcl-2 were significantly reduced (both P<0.001), and the number of neuron apoptosis was significantly increased (P<0.001) in the model group than in the normal group; the random blood glucose level was significantly reduced (P<0.05), the escape latency tested by MWM was significantly shortened (P<0.05), and the number of crossing the original platform tested by MWM was significantly increased (P<0.05), the protein and gene expressions of Bax and Caspase-3 were significantly reduced (all P<0.001), the protein and gene expressions of Bcl-2 were significantly increased (both P<0.001), and the number of neuron apoptosis was significantly reduced (P<0.001) in the EA group than in the model group. Conclusion: EA can improve the learning and memory damages induced by type 2 diabetic model rats with CI; the action mechanism may be achieved via anti-apoptosis.

OBJECTIVETo investigate the cardiac effect of interleukin-2 (IL-2) and to explore the underlying mechanism.

METHODSThe video tracking system and spectrofluorometric method were used to measure the cell contraction and intracellular calcium. Fura-2/AM was used as a calcium fluorescence probe. Langendorff perfusion technique was used to determine the effect of IL-2 on the intact heart.

RESULTSCompared with the control group, IL-2 5 U/ml, 50 U/ml significantly decreased cell contraction amplitude [(74.95+/-4.79) vs (98.09+/-5.02)%, (64.30+/-5.24) vs (97.38+/-4.05)%], peak velocity of cell shortening [(70.23+/-4.85)% vs (98.09+/-5.46)%, (61.15+/-5.20)% vs (97.38+/-6.85)%], peak velocity of cell relengthening [(71.22+/-4.79)% vs (98.32+/-6.08)%, (68.16+/-5.24)% vs (97.55+/-5.00)%] and end- diastolic cell length [(88.28+/-5.84)% vs (97.95+/-5.52)%, (84.18+/-6.52)% vs (98.94+/-6.76)%]. IL-2 (5 U/ml, 50 U/ml) also markedly inhibited intracellular calcium transient [(74.94+/-4.90)% vs (98.09+/-3.74)%,(71.00+/-5.28)% vs (97.38+/-5.52)%], and elevated end-diastolic calcium level of ventricular myocytes [(113.91+/-5.93)% vs (100.10+/-3.02)%, (119.09+/-7.12)% vs (100.52+/-6.00)%], which were attenuated by the opioid receptor antagonist naloxone (Nal,10 nmol/L). In the isolated perfused rat heart,when compared with the control group, IL-2 50 U/ml markedly decreased left ventricular developed pressure [(79.91+/-2.18) vs (93.84+/-2.94)mmHg], maximal rate of rise of left ventricular pressure [(2370.7358.29) vs (2591.50+/-62.81)mmHg] maximal rate of fall of left ventricular [-(1460.95+/-38.6) vs -(1634.24+/-54.05) mmHg/s] and heart rate [(217.35+/-10.56) vs (244.52+/-11.23) beats/min], but increased left ventricular end-diastolic pressure (11.44+/-1.02 vs 9.23+/-0.46). Pretreatment with Nal (10 nmol/L) antagonized the cardiac depression and left ventricular end-diastolic pressure elevation induced by IL-2.

CONCLUSIONThe cardiac effect of IL-2 is mediated by opioid receptors on the membrane of cardiomyocytes.

Animals ; Calcium ; metabolism ; Depression, Chemical ; In Vitro Techniques ; Interleukin-2 ; pharmacology ; Male ; Myocardial Contraction ; drug effects ; Naloxone ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Receptors, Opioid, kappa ; drug effects ; physiology

Objective To investigate the relationship of pathological response of breast cancer after neoadjuvant chemotherapy with the imaging findings in dynamic contrast-enhanced MRI.Methods Forty- five patients with pathologically confirmed breast carcinoma who finished courses of neoadjuvant chemotherapy had breast MRI prior to operation.Dynamic contrast-enhanced MRI scans were performed on a 1.5 T scanner using 3D SPGR sequence before and repeated 6 times after administration of Gd-DTPA. Pathological response was assessed by a pathologist according to Miller & Payne five points classification blinded to breast MRI results.Grade 5 was defined as pCR(pathological complete response).Grade 4 and 5 were defined as major histopathological response(MHR).The type of time signal intensity curve(TIC) (three types),pattern of residual enhancement of each breast cancer were recorded and correlated with pathological findings.Fisher exact test was used for statistical analysis.Results Grade 5 responses were achieved in seven patients;grade 4 in sixteen patients;grade 3 in sixteen patients and grade 1—2 in six patients.70.0%(14/20)of type Ⅰ time signal intensity curve correlated with MHR,while all 6 type Ⅲ curves showed non-MHR response.The type of time signal intensity curve and pathological response grades had statistically significant correlation(P=0.001).18 of the 23 cases with MHR exhibited residual enhancement,while the remaining 5 cases showed no enhancement.Of the 18 MHR cases with residual enhancement,11 showed non-mass-like enhancement and 7 showed mass-like enhancement.The mass(non- mass)morphological pattern in dynamic contrast enhanced-MRI had statistically significant differences in pathological response(P=0.012).Conclusions Pathological response of breast carcinoma after neoadjuvant chemotherapy could be characterized using dynamic contrast-enhanced MRI by identifying patterns of residual contrast enhancement and kinetic curve.Favorable pathological responses correlated with Type Ⅰ TIC,non-enhancement,and non-mass-like residual enhancement.

OBJECTIVETo study the mechanism of gene JWA involved in oxidative stress under hydrogen peroxide (H(2)O(2)) exposure.

METHODSBoth MCF-7 (human breast cancer cell line) and WI-38 (human embryo lung fibroblast cell line) cells were treated with 1 mmol/L of H(2)O(2) with or without pre-incubation of taurine (tau). Malondialdehyde (MDA) and glutathione (GSH) contents in supernatant of cell culture were measured; RT-PCR and Western blotting were carried out for evaluation of the expressions of JWA mRNA and protein respectively. Heat shock proteins (HSP27, HSP70 and HSP90) were also analyzed.

RESULTSThe contents of MDA before and after H(2)O(2) treatment in MCF-7 cells were (0.531 +/- 0.038), (0.674 +/- 0.410) mmol/L respectively, (P < 0.01), while those in WI-38 cells were (0.572 +/- 0.035), (0.683 +/- 0.028) mmol/L respectively, (P < 0.01). The contents of GSH before and after H(2)O(2) treatment in MCF-7 cells were (0.053 +/- 0.002), (0.044 +/- 0.002) g/L respectively, (P < 0.01), while those in WI-38 cells were (0.058 +/- 0.002), (0.050 +/- 0.002) g/L respectively, (P < 0.01). The expression of JWA mRNA was down regulated, at 6 h it decreased by 68.4%, while in WI-38 cells no obvious change found. JWA protein and HSP27 showed markedly increased after H(2)O(2) treatment in both cells but not in similar extent.

CONCLUSIONOxidative stress signal pathways of JWA gene varied between cancer and non-cancer cell lines; JWA protein may have a similar function as HSP27 and act as an important signal molecule in H(2)O(2) induced cell injury.

Blotting, Northern ; Cell Line ; Cell Line, Tumor ; Gene Expression Regulation ; drug effects ; Glutathione ; metabolism ; HSP70 Heat-Shock Proteins ; metabolism ; HSP90 Heat-Shock Proteins ; metabolism ; Heat-Shock Proteins ; genetics ; metabolism ; Humans ; Hydrogen Peroxide ; administration & dosage ; Intracellular Signaling Peptides and Proteins ; Malondialdehyde ; metabolism ; Oxidative Stress ; RNA, Messenger ; drug effects ; genetics ; metabolism

OBJECTIVETo investigate effect of AP-1 and Ets binding site adjacent to matrix metalloproteinase-9 (MMP-9) promoter on activation of MMP-9 transcription of nasopharyngeal carcinoma cells transfected with EBV-encoded latent membrane protein 1 (LMP1), and to ascertain if cross-talk between c-Jun and Ets1 is involved in LMP1-regulating expression of MMP-9.

METHODSSite-directed mutagenesis technique was used to establish a series of mutants, including MMP-9-CAT-Ets(-540)mt, MMP-9-CAT-AP-1(-533)mt and MMP-9-CAT-AP-1(-533)/Ets(-540)mt. After the mutants were transfected into LMP1-expressing NPC HNE2 cells regulated by Tet-on system (pTet-on-LMP1 HNE2), CAT activity of these mutants were assayed with induction of LMP1. With blockade of c-Jun or Ets1 antisense oligonucleotides, the activity of MMP-9 induced by LMP1 was assayed with gelatin zymography.

RESULTSThe CAT activity of MMP-9-Ets(-540)mt-CAT, MMP-9-AP-1(-533)mt-CAT, MMP-9-AP-1(-533)/Ets(-540) mt-CAT decreased significantly compared to MMP-9-CAT wt. After blockade with c-Jun or Ets1 antisense oligonucleotides, activity of MMP-9 induced by LMP1 decreased significantly, especially with combined blockade of c-Jun and Ets1.

CONCLUSIONThe results suggest that transcription factor AP-1 and Ets play an crucial role in activation of MMP-9 transcription induced by LMP1, and cross-talk between c-Jun/Ets1 is involved in expression of MMP-9 mediated by LMP1.

Herpesvirus 4, Human ; genetics ; Humans ; Matrix Metalloproteinase 9 ; biosynthesis ; genetics ; Nasopharyngeal Neoplasms ; metabolism ; virology ; Proto-Oncogene Protein c-ets-1 ; genetics ; Proto-Oncogene Proteins c-jun ; genetics ; Transfection ; Tumor Cells, Cultured ; Viral Matrix Proteins ; genetics

OBJECTIVETo study and explore the relativity of adults HBV vicinal failure and HLA-DR, T cell subset, trace viruses infection. To accumulate date for formulating preventing adult HBV infection prophylactic-therapeutic measures.

METHODSSelect 20 adults randomly who had vaccinated with 10 microgYDV and produced anti-HBS successfully, and another 20 hadn't produced anti-HBs to form two groups-defeated group and contral group. Blood samples from two groups were taken for detecting the level of DR range gene phenotype: T cell subset, white blood cell HLA-DR, HLA-B27, HLA DRB1 * 07, DRB1* 04, DRB1 * 1001, DQB1 * 0401 and so on.

RESULTSThe level of CD4(-)/CD8(-) is lower in the infection group than in healthy group. But the average level of HLA-DR and HLA-B27 is higher in the infection group. The differences of HLA DRB1 * 07 gene frequency between two groups were significant (P <0.05), but the levels of CD3, CD4, CD8, CD7, CD4/CD8 and HLA DRB1 * 04, DRB1 * 1001, DQB1 * 0401 were not significant.

CONCLUSIONThe failure of HBV vaccination on adults may have relation to HLA-DR, HLA-B27, HLA DRB1 * 07, CD4(-)/CD8(-), etc.

Adult ; HLA-DR Antigens ; genetics ; immunology ; Hepatitis Antibodies ; blood ; Hepatitis B ; genetics ; immunology ; prevention & control ; virology ; Hepatitis B Vaccines ; administration & dosage ; immunology ; Hepatitis B virus ; genetics ; immunology ; Humans ; Phenotype ; T-Lymphocyte Subsets ; immunology

OBJECTIVETo develop a double antibody sandwich ELISA assay for quantitative determination of recombinant human interferon alpha1b.

METHODSMouse monoclonal antibodies with different binding site on rIFN-alpha1b were screened to select optimized candidates as coating and HRP-labeled index antibodies respectively. And a double antibodies sandwich ELISA was assembled; the reliable lower detection limit, specificity, accuracy and reproducibility were evaluated and validated.

RESULTSThe quantitative sandwich ELISA had a reliable lower detection limit of 10 ng/ml, with a liner detection range 10-100 ng/ml (R2 = 0.992), variation coefficient inter-plates is less than 10%.

CONCLUSIONThe developed sandwich ELISA was a sensitive and specific, accuracy and reproducibility method for quantitative determination of recombinant human interferon alpha1b in final product.

Animals ; Antibodies, Monoclonal ; analysis ; Enzyme-Linked Immunosorbent Assay ; instrumentation ; methods ; Humans ; Interferon-alpha ; blood ; Mice ; Mice, Inbred BALB C

OBJECTIVESTo observe the role of PPARgamma during the activation process of hepatic stellate cells (HSC).

METHODSBy morphology and RT-PCR, we study the changes of expression of PPARgamma in culture-activated HSC or in vivo activated HSC induced by dimethylnitrosamine (DMN).

RESULTSIn vitro, the expression level of PPARgamma in freshly isolated HSC (0.72+/-0.01) significantly reduced to 0.48+/-0.03 on the third day of culture (t = 19.8372, P<0.01), and reduced 70% on the seventh culture-day and could not be detected after the second passage. In vivo, HSC freshly isolated from normal control rats expressed PPARgamma (0.76+/-0.01). During the development of rat liver fibrosis induced by DMN, the expression level significantly reduced to 0.46+/-0.02 after the third injection of DMN (t = 29.5318, P<0.01), and reduced 66% on the end of first week and could not be detected on the end of second and third week.

CONCLUSIONThe expression of PPARgamma might play an important role on the maintenance of resting-form of HSC, and the reduction of expression of PPARgamma might be an early event during the activation process of HSC.

Animals ; Liver ; cytology ; Liver Cirrhosis ; etiology ; pathology ; Male ; RNA, Messenger ; analysis ; Rats ; Rats, Wistar ; Receptors, Cytoplasmic and Nuclear ; physiology ; Transcription Factors ; physiology

OBJECTIVEThis case-control study was aimed to detect the single nucleotide polymorphisms (SNPs) in JWA promoter region, to assess the effect of SNP on transcriptional activity, and to probe the relationship between SNP and the risk of bladder cancer.

METHODSThe design of one control per case was adopted. The JWA gene promoter region in 155 patients with bladder cancer and in 155 cancer-free controls was amplified by PCR-SSCP technique, and the SNP were confirmed by direct DNA sequencing. The recombinant plasmids of JWA promoter fragment which contain the SNP were constructed as CAT reporter gene and were transfected transiently into NIH 3T3 cells for disclosing whether SNP changes the transcriptional activity of the promoter.

RESULTSA novel SNP -76 G-->C at promoter region of JWA gene was found. The frequencies of the C allele and GC genotype at JWA promoter -76G-->C in bladder cancer group (10.00% and 20.00% respectively) were significantly higher than those in control group (5.16% and 10.32% respectively) (P < 0.05). The transcriptional activity of -76GC allele genotype was significantly down-regulated as compared with that of -76GG allele genotype (P < 0.01). Multivariate logistic regression analysis revealed that JWA polymorphism at promoter -76G-->C is an independent novel risk factor for bladder cancer.

CONCLUSIONThe JWA -76G-->C variant genotype may play an important role in transcription regulation of JWA gene and in the susceptibility to bladder cancer.

Aged ; Animals ; Case-Control Studies ; Female ; Genetic Predisposition to Disease ; genetics ; Heat-Shock Proteins ; genetics ; Humans ; Intracellular Signaling Peptides and Proteins ; genetics ; Male ; Mice ; Middle Aged ; NIH 3T3 Cells ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Polymorphism, Single-Stranded Conformational ; Promoter Regions, Genetic ; genetics ; Transfection ; Urinary Bladder Neoplasms ; genetics

OBJECTIVETo examine the role of Polo-like kinase 1(PLK1) in the migration and invasiveness of human colorectal cancer cells.

METHODSNine colorectal cancer cell lines were cultured. Cell lines with the highest level of PLK1 expression was selected by PCR and Western blot. Three siRNA oligo segments targeting PLK1 were designed and selected cell lines transfected. Successful transfection was confirmed using real-time PCR and Western blot. Changes in migration and invasiveness of the selected cell line were evaluated by Transwell test.

RESULTSColorectal cancer cell line SW1116 was selected with the highest expression of PLK1 at both mRNA level and protein level. The expression of PLK1 in SW1116 was reduced by the three siRNA oligo segments to varying degrees, and the No.1 siRNA oligo segment was the most efficient. In migration test, the number of cells crossing through chambers in PLK1-siRNA group was 44 ± 14, which was lower than that in the negative control group (242 ± 40) and in blank control group(240 ± 38). In invasion test, the number of cells crossing through chambers in PLK1-siRNA group was 62 ± 3, which was lower than that in negative control group (207 ± 12) and in blank control group (211 ± 15). These differences were statistically significant(P<0.01).

CONCLUSIONPLK1 silencing by siRNA may inhibit the migration and invasiveness of colorectal cancer cells, suggesting that PLK1 might play an important role in the infiltration and metastasis of colorectal cancer.

Cell Cycle Proteins ; genetics ; metabolism ; Cell Line, Tumor ; Cell Movement ; Colorectal Neoplasms ; metabolism ; pathology ; Humans ; Neoplasm Invasiveness ; Protein-Serine-Threonine Kinases ; genetics ; metabolism ; Proto-Oncogene Proteins ; genetics ; metabolism ; RNA, Small Interfering ; genetics ; Transfection