ABSTRACT:Objective To prepare polymeric nanomicelles capable of simultaneously loading doxorubicin (DOX) and Bcl‐2 small interfering RNA (Bcl‐2 siRNA ) , and to explore their in vitro cytotoxicity and cellular uptake in MCF‐7 human breast cancer cells .Methods Copolymer poly (ethylene glycol )‐g‐polyethylenimine‐g‐poly(γ‐benzyl‐L‐glutamate) was synthesized by the combination of reductive amination and carbodiimide methods , and its chemical structure was verified by 1 H NMR .Empty and drug‐loaded copolymeric nanomicelles were prepared by dialysis method and characterized by transmission electron microscope and dynamic light scattering .The ability of the nanomicelles to compress Bcl‐2 siRNA was measured by by agarose gel electrophoresis method . The release profiles of DOX and Bcl‐2 siRNA from the nanomicelles were explored by means of fluorescence spectrometry and dialysis method .The in vitro cytotoxicity and cellular uptake of DOX and Bcl‐2 siRNA co‐loaded nanomicelles in MCF‐7 human breast cancer cells were characterized by MTT assay and confocal laser scanning microscopy , respectively .Results The critical micelle concentration of the copolymer was about 4 mg/L ,and the sizes of self‐assembled empty and drug‐loaded nanomicelles were smaller than 200 nm .The drug‐loading efficiency and drug‐loading content of DOX in the nanomicelles were 88 .7% and 15 .1% ,respectively .The DOX‐loaded nanomicelles could efficiently compress Bcl‐2 siRNA when an N/P ratio was ≥64 .The zeta potential of DOX and Bcl‐2 siRNA co‐loaded nanomicelles was +30 mV .The release behavior of the cargoes from the nanomicells was pH‐sensitive , and the release of Bcl‐2 siRNA was more sensitive to acidic pH than that of DOX . The nanomicelles could simultaneously deliver DOX and Bcl‐2 siRNA into MCF‐7 cells , and the co‐delivered DOX and Bcl‐2 siRNA significantly increased the cytotoxicity of DOX (P<0 .05) .Conclusion The polymeric nanomicelles can co‐load DOX and Bcl‐2 siRNA and deliver them into MCF‐7 cells , and DOX in combination with Bcl‐2 siRNA can synergistically inhibit the growth of MCF‐7 cells and promote cell apoptosis ,suggesting that the nanomicells may be a promising carrier for the co‐delivery for chemotherapeutics and genes .

Objective To investigate the effects and mechanisms of rosiglitazone on the expressions of nuclear factor-κB and matrix metalloprotease (MMP-9) in peripheral blood monocyte-derived macrophages (MDMs) in patients with coronary heart disease. Method This was a clinical case-control study. Forty-eight actue coronary symdrome (ACS) patients (ACS group), and 20 patients with stable angina (SA) (control group) were collected. They were performed coronary arteriography in the Department of Cardiology of the Second Xiangya Hospital from March to April in 2007. Exclusion criteria included acute infection, trauma or surgery patients within four weeks, cerebral vascular accident, liver and kidney dysfunction, cancer, and so on. The peripheral blood mononuclear cells were isolated and transformed into MDMs with macrophage colony-stimulating factor treatment. The transformed MDMs were randomly assigned into subgrougs and incubated with 0 /μmol/L, 1 μmol/L, 10 μmol/L, 20 μmol/L of rosiglitazone respectively. The expressions of PPAR-γ mRNA, MMP-9 mRNA were determined by RT-PCR and nuclear factor-κB P65 (NF-KB P65) expression by immunohistochemistry. Multiple comparisons were examined for significant differences using analysis of variance (ANOVA). Results The basal expression of PPAR-y mRNA was lower, in contrast, the levels of NF-KB P65 and MMP-9 mRNA were higher in ACS group than control group. PPAR-γ mRNA expression were significantly upregulated in both ACS and control groups with rosiglitazone treatment. PPAR-γ mRNA expression was positive correlation, while the expressions of MMP-9 mRNA were negative correlation with the rosiglitazone concentration in the ACS group. Rosiglitazone inhibited the expression of NF-KB in a concentration-independent manner in ACS and control groups. Conclusions The expression of PPAR-y mRNA is inhibited, while the activity of NF-KB and expression of MMP-9 mRNA are enhanced in MDMs of ACS cases. Rosiglitazone intervention may inhibit NF-KB activity and MMP-9 expression by upregulation of PPAR-y expression in MDMS of patiens with ACS.

Objective To observe the effect of Bushen Gutai mixture on uterine blood supply of rat model of abortion induced by hydroxyurea tablets combined with mifepristone through PKA-CREB signal pathway and its mechanism of calming fetus.Methods 60 pregnant rats of SPF grade SD rats were prepared by closing cages at 2∶1.According to the order of pregnancy,60 pregnant rats were randomly divided into 6 groups with 10 rats in each group:Bushen Gutai mixture group(low,middle and high dose),normal pregnancy group,model group and Di qu progesterone group.On the 1st to 9th day of pregnancy,except the normal group,the pregnant rats in each group were gavaged with hydroxyurea tablets at 5∶00 pm every day(450 mg·kg-1),and at 10∶00 am on the 10th day of pregnancy.Mifepristone tablets were given by intragastric administration(4.0 mg·kg-1).At 9∶00 am every day from the 1st to 9th day of pregnancy,Bushen Gutai mixture was given to the low,middle and high dose groups(0.5,1.0,2.0 g·kg-1),didrone group(3.02 mg·kg-1),model group and normal pregnancy group with 0.9%normal saline.24 hours after the last administration of pentobarbital sodium(50 mg kg-1),all pregnant rats were killed by intraperitoneal injection of pentobarbital sodium,and the uterine decidual tissue of pregnant rats was bluntly isolated in sterile environment.Hematoxylin-eosin(HE)staining was used to observe the lumen diameter and wall thickness of spiral artery in uterine decidual tissue.Immunohistochemical staining(IHC)was used to detect the expression of vascular endothelial growth factor(VEGF)in decidual tissues.Western blot was used to detect the protein expression of phosphorylated protein kinase A,protein kinase A,phosphorylated cyclic adenosine monophosphate response element binding protein,Cyclic Adenosine monophosphate response element binding protein and aquaporin 5 in the decidua of pregnant rats.The apoptosis of decidual cells was detected by in situ end labeling(TUNEL)of DNA fragmentation.Results Compared with the model group,the wall thickness of spiral artery was higher than that in other groups(P<0.05),the lumen diameter was lower than that in other groups(P<0.05)and the expression of VEGF protein was lower than that of other groups(P<0.01).Compared with the model group,the apoptosis level of decidual cells in uterine decidua of abortive rats in high,middle and low dose groups of Bushen Gutai mixture and diqu progesterone group decreased in varying degrees.Bushen Gutai mixture can up-regulate the levels of p-PKA/PKA and p-CREB/CREB in uterine decidua(P<0.01)and promote the expression of AQP5 protein in uterine decidua of abortion rats(P<0.01).Conclusion Bushen Gutai mixture can improve uterine blood supply of aborted rats by activating PKA and CREB phosphorylation,up-regulating AQP5 expression,promoting physiological recasting of spiral artery and high expression of Vascular endothelial growth factor.

Based on the resutls of literature review and interviews of experts, two rounds of Delphi surveys were conducted. The mean, importance ratio, coefficient of variation and coordination coefficient were used for assessment of survey from multiple perspectives, and finally form a framework model of factors affecting the efficacy of Tuina therapy. A total of 37 experts were selected for questionnaire surveys, the positive coefficients of experts' participatation in the first round and second round were 92.5% and 80.0%, respectively. The overall coordination coefficient in the second round is 0.68. The items were included into the consensus meeting if the importance ratio of items were equal to and more than 80%. After the expert consensus meeting, 22 items were included to form a framework model of factors affecting the efficacy of Tuina therapy, and summarized as 5 major influencing factors, including diagnostic factors, treatment factors, prognostic factors, patient factors, and doctor-patient communication. This framework can guide and help young Tuina practitioners to improve clinical efficacy. It is also clearly pointed out that the effect of Tuina for pain is not only related to disease diagnosis or manipulation, but also related to home exercise, health care, and doctor-patient communication.