Objective To investigate the expression of cyclooxygenase-2(COX-2)in breast cancers, and understand its clinical significance.Methods The immunohistochemistry was used to determine the ex- pression of COX-2 in the 40 tissues of breast cancer and 18 adjacent noncancerous mammary tissues.Results The positive expression of COX-2 was detected in 52.5% of breast cancer tissues and in 11.1% of adjacent noncancerous mammary tissues.There was a significant difference in the expression of COX-2 between breast cancer tissues and adjacent noncancerous mammary tissues(P0.05).Conclusion COX-2 was over expressed in breast cancer and correlated with metastatic lymph nodes and HER-2/neu.The expression of COX-2 may play an important role in carcinogen- esis,progression and prognosis of breast cancer.

Objective To investigate the different impacts of the components of metabolic syndrome (MS) on cardiovascular disease (CVD) risk factors and Framingham risk value (FRV) in newly-diagnosed type 2 diabetes mellitus (T2DM) patients without coronary heart disease (CHD).Methods A total of 212 newly-diagnosed T2DM patients was divided into three groups based on the components of MS,including body mass index (BMI),triglyceride (TG),high density lipoproteins cholesterol (HDL-C),and mean arterial pressure (MAP) tertile,respectively.The general CVD risk factors [smoking,BMI,TG,HDL-C,total cholesterol (TC),low density lipoproteins cholesterol (LDL-C),HbA1 C,homeostasis model assessment of insulin resistance (HOMA-IR),and FRV] were compared among the groups.Logistic regression analysis was used to observe the different impact of MS components on CVD risk.Results With the increasing of BMI,TG and MAP tertiles,all the CVD risk factors and the ratio of FRV middle-risk group or high-risk group had the tendency of augmentation.With the increasing of HDL-C tertiles,all the CVD risk factors and the ratio of FRV middle-risk group or high-risk group was in downtrend.Binary logistic regression analysis indicted that BMI,systolic pressure,diastolic pressure,HbA1 c and TG were risk factors of FRV non-low-risk group (middle-risk and high-risk groups),HDL-C was the protective factor.The odds ratios (ORs) of BMI,systolic pressure,diastolic pressure,HbA1 c,TG and HDL-C were 2.794 (95％ CI:2.390-2.408),2.601 (95 ％ CI:1.974-3.701),1.476 (95 ％ CI:1.231-2.048),2.964 (95 ％ CI:1.925-3.282),1.464(95％ CI:0.934-2.294),and 0.732(95％ CI:0.023-0.962),respectively.Logistic stepwise regression analysis indicated that systolic pressure,BMI,HbA1 C,and HDL-C were entered into the regression equation,and the partial regression coefficient was 0.784,1.213,1.679,and-0.854,respectively (P ＜ 0.05 or P ＜ 0.01).Conclusions All the components of MS in newly-diagnosed T2DM were correlated with CVD risk factors.However,they should be weighed differently.

Objective To construct the human IL-24 recombined adenovirus(Ad-IL24)and to observe the effect of Ad-IL24 on ex vivo culture of HL-60 cells.Methods pAdTrack-CMV-IL24 was constructed by PCR with pcDNA3.0-IL24 recombined plasmid as a template,enzyme digestion and ligation.The pAdTrack-CMV-IL24 lineared by Pme Ⅰ was co-transformed into BJ5183 with pAdEasy-1.The pAdEasy-1-pTrack-CMV-IL24 recombined adenovirus vector was lineared with Pac Ⅰ and then transfected in to QBI-293A cells.The Ad-IL24 was obtained and used to infect HL-60 ceils,and the effect on HL-60 cells was tested by LSCM,Mrrr,FCM,ELISA and immunohistochemistry staining.Results The pAdEasy-1-pTrack-CMV-IL24 vector was constructed and the Ad-IL24 was successfully obtained.The effect of inhibiting growth of HL-60 cells and inducing cell apoptosis by IL-24 was proved by LSCM,MTT and FCM.IL-24 down-regulated the expression of anti-apoptosis factor Bcl-2 and increased the secretion of IFN-γ,TNF-α of HL-60 cells.Conclusion Ad-IL24 can inhibit the growth of HL-60 cells and induce cell apoptosis through down-regulating expression of anti-apoptosis factor and increasing the secretion of IFN-γ,TNF-α of HL-60 cells.The human Ad-IL24 may provide a basic study for the future target therapy to tumors.

Objective:To construct the cells which can express LIF protein steadily and study the effect of rhLIF and IL-24 on HL-60 cells.Methods:ECV-304 cells were infected by eukaryotic expression plasmid pcDNA3-LIF,then the positive cells were selected by G418.The positive cells were collected and their culture supernatant was stored for further use.Expression of LIF mRNA was detected by RT-PCR.The pAdeasy-1-pAdTrack-CMV-IL-24 was extracted from DH5a.The recombined adenovirus vector was lineared with PacI and transfected into 293 cells.The IL-24 recombined adenovirus(Ad-IL-24) was obtained and used to infect HL-60 cells.At the same time,the culture supernatant containing rhLIF was added into the HL-60 cells which was identified as positive expression of IL-24 by RT-PCR.Synergistic effect of the cytokines on HL-60 cells was tested by LSCM, FCM and immunohistochemistry stain assay.Results:The cells expressing LIF protein steadily were constructed successfully, the high titer of the recombined adenovirus(Ad-IL-24) was obtained. Expression of IL-24 in infected HL-60 cells was identified by RT-PCR.The apoptosis of HL-60 cells induced by rhLIF and IL-24 was proved by LSCM ,FCM and immunohistochemistry stain assay. They had synergistic effect.Conclusion:rhLIF and IL-24 can inhibit growth of HL-60 cells and induce apoptosis of the cells.They have synergistic effect.

Thirty SHRs were obtained randomly to hypertension, model group, captopril group and Qingre jiangya capsule group. Ten Wistar rats were used as control group. The hippocampus tissue was removed to explore the damage of spontaneously hypertensive rats (SHR) and the protective effect of Qingre jiangya capsule after continuously administered for 14 days. And then the data were processed by principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA). The research results revealed captopril group was significantly different from the other three groups. The classification of other three groups is also very clear after captopril group removed. This suggested that Qingre jiangya capsule could improve the overall metabolism compared with captopril. Four metabolites were identified: dimethylglycine, glycerophosphocholine, aldosterone and noradrenaline. Hypertension hippocampus damage may mainly be expressed in tyrosine metabolism, aldosterone-regulated sodium, vascular smooth muscle contraction reabsorption, and Qingre jiangya capsule could reverse the hippocampus tissue damage of SHR.
Animals ; Capsules ; pharmacology ; Drugs, Chinese Herbal ; pharmacology ; Hippocampus ; drug effects ; Hypertension ; drug therapy ; Male ; Rats ; Rats, Inbred SHR ; Rats, Wistar

Objective To establish Ad-LIF-OSM transgenic feeder cells for the expansion of CD34+ hematopoietic stem/progenitor cell and tentatively study its effect in expansion and differentiation of cord blood hematopoietic stem cell(HSC) in vitro.Methods In the foundation of pAdTrack-CMV-LIF-polyA+promoterΔ,the OSM gene was inserted to the vector plasmid.Then we structure the transfer plasmid pAdTrack-CMV-LIF-polyA+promoterΔ-OSM.The transfer vector and backbone vector were further cotransfected for homologous recombination. The result pAdEasy-1-pAdTrack-CMV-LIF-polyA + promoterΔ-OSM homologous recombination plasmid were transfected into the human embryonic kidney 293 (QBI-293A) cells,leading to formation of the recombinant adenviruses Ad-LIF-OSM which co-expressing LIF and OSM.Infect the feeder layer cells with groups of Adenovirus,detection the expressing of LIF and OSM in WI-38 cells by RT-PCR and ELISA.Compares the stem cells differentiation and proliferation of the different experimental groups in vitro by transwell and cell counting.Results The sequencing results show that the OSM genes were anastomotic in Ad-LIF-OSM.LIF and OSM gene could be detected in feeder layer cells which infected by Ad-LIF-OSM.Exogenetic LIF and OSM have special effect in culturing HSC in vitro.Conclusion The adenoviral vector co-expressing LIF and OSM (Ad-LIF-OSM) were successfully constructed.Ad-LIF-OSM transgenic feeder cells can effectively proliferate umbilical cord blood CD34+ HSPC in vitro and delay it differentiate.

AIM:To investigate the effects of C1q/TNF related protein 3 (CTRP3) on the insulin sensitivity of insulin resistant 3T3-L1 adipocytes.METHODS: The insulin resistance model of 3T3-L1 adipocytes was induced by palmic acid cultivation.The adipocytes were treated with different concentrations of recombinant CTRP3 protein (10, 50, 250,1 250 μg/L) for 12 h, and for different times (2, 6, 12, 24 h) at the concentration of 250μg/L.The glucose con-sumption was detected by the glucose oxidase method.The glucose transport ratio was measured by 2-deoxidation-[3H]-glucose intake method.The contents of TNF-αand IL-6 in the supernatant were detected by ELISA.The mRNA expression of TNF-α, IL-6 and glucose transporter-4 (GLUT-4) was measured by real-time PCR.The protein expression of GLUT-4 was detected by Western blotting.RESULTS:Compared with normal control ( NC) group, the glucose consumption and glucose intake ratio of insulin resistance ( IR) group was decreased by 50.6%and 57.9%, respectively.Compared with IR group, with the increase in CTRP3 (10, 50, 250,1 250 μg/L) in intervention groups, the glucose consumptions were in-creased by 22.1%, 42.9%, 76.6% and 80.5%, respectively, and the glucose intake ratios were increased by 39.0%, 68.0%, 108.0%and 111.0%, respectively.With the increased duration (2, 6, 12 and 24 h) of CTRP3 treatment at the concentration of 250 μg/L, the glucose intake ratio was increased by 23.0%, 79.0%, 109.0%and 114.0%, respectively. The contents of TNF-αand IL-6 in the supernatant were decreased by 17.4%and 17.1%respectively as treated with CTRP3 at the concentration of 250 μg/L for 12 h, and the mRNA expression of TNF-αand IL-6 was decreased by 26.0% and 18.9%respectively, while the mRNA and protein expression of GLUT-4 was increased by 61.5%and 55.6%respectively. CONCLUSION:CTRP3 may increase the insulin sensitivity of insulin resistant 3T3-L1 adipocytes by down-regulating the expression of inflammatory factors, improving the insulin signal transduction and increasing the expression of GLUT-4.

Objective To study the antitumor effect and mechanism of co-cultured cytokine-induced killer(CIK) cells and autologous DC modified with IL-24 gene on A549 cells in vitro. Methods DC and CIK cells were prepared routinely from human peripheral blood mononuclear cells(PBMC). Recombinant adenovirus vector pAdEasy-1-pTrack-CMV-IL-24 was extracted from DH5α, it was lineared with Pac I and transfected into A293 cells, and then the IL-24 recombined adenovirus(Ad-IL-24) was obtained. Ad-IL-24 was used to infect DC. The cells obtained were named DC-IL-24. RT-PCR and ELISA were used to evaluate the expression of IL-24 gene in transfected DC. The phenotypes change of DC were identified by flow cytometry analysis, the concen-tration of IL-12 and TNF-α in supernatant of DC were determined by EIJSA. The ability of CIK producing per-forin was measured by homolysis method. FCM was used to determine the cytotoxicity of cocultured CIK cells and autologous DC modified with IL-24 gene to A549 cells. Results We obtained the high titre of Ad-IL-24.IL-24 gene was transfered into DC successfully via Ad-IL-24. The green fluorescence was observed on DC by fluorescence microscope. The expression rate of CD80, CD83, HI.A-DR, CD40, CXCR4 on DC-IL-24 was sig-nificantly increased compared with that of the control group. DC-IL-24 produced markedly higher levels of IL-12 and TNF-α as compared with DC. DC-IL-24 can enhance the ability of CIK cells producing perforin. On com-parison with non-transfected DC co-cultured with CIK cells, transfected DC co-cultured with CIK cells had a sig-nificantly higher lytic activity against A549 cells. Conclusion IL-24 gene modification can enhance the anti-tu-moral immunity of DC. The mechanism of which might be related to the increased secretion of IL-12 and TNF-α, up-regulation expression of co-stimulatory molecules and MHC Ⅱ class molecules on DC, promoting the acti-vation and maturation of DC, and then enhancing CIK cells to generate specific anti-tumoral immunity.

AIM: To determine the role of Fas antigen and caspase-8 in modulating apoptosis of osteosarcoma cells induced by bacterial redox protein azurin. METHODS: The human osteosarcoma cell line U2OS was treated with bacterial redox protein azurin (150 mg/L) for 6, 12, 24 and 48 h, respectively. Cell immunohistochemistry and quantitative image pattern analysis were applied for detecting the expression of Fas antigen. Caspase-8 activity was detected using caspase-8 fluorescent assay kit. The apoptotic rate was measured by FCM. RESULTS: Compared with the control group, the expression of Fas antigen and activity of caspase-8 significantly increased in U2OS cells treated with 150 mg/L azurin (P

Objective To investigate the impacts of low-grade inflammation on the correlation of serum testosterone (T) and carotid intima-media thickness (CIMT) in type 2 diabetes mellitus (T2DM) men.Methods Based on the concentration of C-reactive protein (CRP) and T,a total of 247 patients was divided into low-grade inflammation with low T group (LI-LT,CRP ≥ 2.0 mg/L,T ＜ 12.0 nmol/L,n =65),low-grade inflammation with normal T group (LI-NT,n =67),non-low-grade inflammation with low T group (NLI-LT,n =56),and non-low-grade inflammation with normal T group (NLI-NT,n =59).General information,medical history,and anthropometry data were collected.Glycosylated hemoglobin AI c (HbA1c),blood fat,and CIMT were detected.Results Compared to NLI-NT group,CIMT in NLI-LT group was increased without statistical significance [(0.87 ±0.09) vs (0.90 ±0.10)mm,t =1.693,P =0.090].CIMT in LI-LT group was increased significantly compared to that of LI-NT group [(0.99 ± 0.10) vs (1.07 ±0.12)mm,t =5.208,P =0.000].Correlation analysis indicated that serum T correlated negatively with CIMT (n =247,r =-0.368,P ＜0.01) in whole.The correlation coefficient of T and CIMT was-0.582 (P =0.000),and-0.098 (P =0.087) in patients with (n =132) and without (n =115) low-grade inflammation,respectively.To make CIMT dependent coefficient and serum T independent coefficient in multiple regression analysis,the partial regression coefficient was-0.062 (95％ CI:-0.094 ～-0.029,P =0.008),and-0.045 (95％ CI:-0.087 ～-0.002,P =0.036),respectively,before and after the adjustment of age,smoking,family history,T2DM course,body mass,blood pressure,HbAlc,and blood fat.After the additional adjustment of CRP,the partial regression coefficient was-0.019 (95％ CI:-0.120 ～ 0.042,P =0.287).Conclusions The negative relationship between serum T and CIMT in T2DM men might be modulated by low-grade inflammation.