Objective To investigate the role of hydroxyapatite nanoparticle (nHAP) in the gene transfection of human colorectal cancer cell line SW480/M5 and the possible mechanisms.Methods The combination and protection of nHAP-Mg2+ to DNA were analyzed by gelose gelatin electrophoresis.Liposome and nHAP modified by magnesium chloride was combined,and the PEGFP-N1 plasmids were transfected into SW480/M5 cells.The gene transfection rate and the mean fluorescence intensity were observed by flow cytometry.The effect of nHAP-Mg2+ on the growth of the cells were studied by MTT.Results At appropriate proportion,nHAP-Mg2+ could combine the plasmids compeletly and protected the DNA.The gene could not be transferred by nHAP-Mg2+ alone.Combining the nanoparticles and liposome,the gene could be transferred very efficiently and the transfection rates were significantly higher than the liposome (P ＜ 0.05).The inhibition of cell growth was increased along with the concentration of nHAP-Mg2+ wether it was used alone or with the combination of liposome (P ＜ 0.05).Conclusions nHAP-Mg2+ has the ability to combining and protecting DNA and can be used to transfer gene as the adjunct carrier of liposome for the gene therapy of tumor cells to elevate the gene tansfection and expression rate and also enhance the anti-tumor effection.

Objective To investigate the pathological change in articular cartilages after firearm injury.Methods Four rabbits from 28 New Zealand healthy rabbits were chosen as control group and subjected to joint capsule incision only. Another 24 rabbits were equally divided into 6 experimental groups( groups B to G) and subjected to medial femoral condyle cartilage surface damage by the nail gun.After the operation, their specimens were collected after 6 h,3 d,7 d,14 d,28 d and 56 d, respectively.Tissue sections were observed and stained by HE staining and toluidine blue staining.The histolopathological changes in articular cartilage after firearm injury were detected.Results The color of articular cartilages in experimental groups became lighter, the cell number increased but then decreased, the articular cartilage layer disappeared, the cell shape became uneven, cells began to cluster and the Mankin score increased, and the statistical differences between experimental groups and control group were significant.Conclusion The histological pathological changes in articular cartilages after fiream injury seem to follow some pattern.The degeneration seems obvious after 7 days and then becomes heavier.

Objective Cerebral microbleeds (CMBs) are important indicators of cerebral small vessel disease .However, it is still unclear whether endothelial dysfunction is involved in CMBs .The aim of this study is to investigate the correlation between CMBs and soluble E-selectin (sE-selectin) in patients with acute ischemic stroke . Methods Based on the results of MRI (3.0 T) susceptibility weighted imaging , we divided patients with first acute ischemic stroke into a CMBs group ( n=63 ) and a non-CMBs group (n=63), and recruited another 45 volunteers with normal MRI findings as controls .We collected and conducted comparative a-nalysis on the demographic data , biochemical variables ( including the sE-selectin level ) , vascular risk factors , and the number of CMBs of the patients . Results Ordinal logistic regression analysis showed a significantly positive correlation between sE -selectin and the number of CMBs (OR=1.062, 95%CI:1.023-1.103, P=0.002), higher systolic blood pressure associated with more CMBs (OR=1.014, 95%CI:1.002-1.025, P=0.021). Conclusion Serum sE-selectin is significantly positively correlated with and can be used as a biological marker for the severity of CMBs .

Objective To investigate the effects of olfactory bulb(OB) lesion on neural stem cells proliferation and expression of NMDA receptor subunit 2B in subventricular zone(SVZ) of rats.Methods Sixty adult female SD rats were randomly divided into normal group,saline group and OB lesion group.OB lesion was induced by N-methyl-D-aspartic acid (NMDA) injection.Each group was respectively divided into four time points including 3 d,7 d,14 d and 28 d.Immunohistochemistry staining was used to detect the number of Nestin,Ki67 and NR2B-positive cells in the SVZ.Results (1) Nestin positive cells in the SVZ were shown at the different time of three groups.Seven days after OB lesion,IOD value of nestin-positive cells began to increase((29601± 1788)/0.01 mm2,P＜0.05),reached the maximum at 14 d((49800±3701)/0.01 mm2,P＜0.05) and still sustained a high level at 28 day((27600±3209)/0.01 mm2,P＜0.05).(2)Ki67 positive cells in the SVZ were shown at the different time of three groups.The number of Ki67-positive cells was increased significantly at 7 d,14 d and 28 d after OB lesion compared to normal group and saline group (P＜0.05).(3)NR2B immune expression in the SVZ was shown at the different time of three groups.The NR2B-positive cells increased at 3 d after OB lesion(58.80±2.95,P＜0.05),reached the maximum at 14 d(68.40±4.04,P＜0.05).At 28 d of OB lesion,the number of positive cells was reduced,but still sustained a high level(62.20±3.56,P＜0.05).(4)The positive cells of NR2B and Ki67 were highly positive correlation at different time after OB lesion(r=0.968,P＜0.05).Conclusions OB lesion can stimulate neural stem cell proliferation and increases the expression of NR2B.The increased mode of NR2B is in accordance with the schedule of the neural stem cells increase induced by OB lesion.Therefore,it indicates that the NMDA receptor subunit 2B may be involved in neural stem cell proliferation.

Background and purpose:We investigated whether lfuorine-18 lfuorodeoxyglucose (18F-FDG) maximal standard uptake value (SUVmax) of the primary tumor (SUV-T), SUVmax of the regional lymph nodes (SUV-N) or the overall loco-regional lesion SUVmax (SUV-TOTAL) was related to survival of patients with stage Ⅲ non-small cell lung cancer (NSCLC) who received Cetuximab and combined definitive chemoradiotherpay. Methods:From September 2009 to July 2012, seventeen patients with unresectable stageⅢNSCLC receiving cetuximab with cisplatin/vinorelbine (NP) followed by concomitant NP and intensity-modulated radiotherapy (IMRT) at the Fudan University Shanghai Cancer Center were enrolled onto a prospectively study. All patients received positron emission tomography/computerized tomography (PET/CT) scans within 2 weeks before enrolment. Univariate analysis were used to assess the correlation between SUV-T, SUV-N, SUV-TOTAL, gender, age, histology, tumour-node-metastasis (TNM) stage, performance status (PS) as well as smoking status and survival. The factors which showed statistical signiifcance entered into multivariate Cox-regression model. Survival functions of different populations were estimated by Kaplan-Meier method and compared by Log-rank test. Results:In the univariate analysis, SUV-T, SUV-N, SUV-TOTAL, PS and smoking status were prognostic factors. The best cut-off values for SUV-T, SUV-N and SUV-TOTAL were 11, 11 and 20, respectively. Multivariate analysis revealed that SUV-TOTAL (P=0.012), SUV-T (P=0.025), and SUV-N (P=0.033) were independent predictors of survival with hazard ratio (HR) of 14.7, 11.2, and 6.2, respectively. Conclusion:Local, regional and locoregional maximal SUVs deifned by 18F-FDG PET-CT scanning may have a strong correlation with survival in this patients setting, which merits further study.

Objective:To study the therapeutic effect of a novel double-target system,in which human umbilical cord-derived MSCs were used as vehicles to deliver fusion protein scFvCD20:sTRAIL to non-Hodgkin ’ s lymphoma. Methods: The traditional methods in molecular biology were used to construct lentivirus expression vectors pLenR. scFvCD20: sTRAIL and contrast vectors. Human umbilical cord-derived MSCs ( HUMSCs ) were labeled with the copGFP by transducing with pseudo viral particles which had been packaged in 293T cells with four plasmid-lentivirus packaging system. Fusion protein scFvCD20:sTRAIL were secreted from MSC. scFvCD20:sTRAIL after that HUMSCs were infected by pseudo viral particles. CCK8 assay was applied to detect the antigen-restricted cell death induced by scFvCD20:sTRAIL in CD20-positive BJAB and Raji cells as well as CD20-negtive Jurkat cells and human normal peripheral blood mononuclear cells (PBMCs). To evaluate the therapeutic effect of MSC. scFvCD20:sTRAIL in vivo,ge-netically modified HUMSCs were intravenously injected into tumor-bearing mice with BJAB cells. The volume of tumor was measured every three days, and the inhibition ratio of tumor was calculated according to tumor volume. Results: Lentivirus expression vectors pLenR. scFvCD20:sTRAIL, pLenR. ISZ:sTRAIL, pLenR. scFvCD20 and pLenR. CopGFP were successfully constructed and these constructs could be expressed stably in HUMSCs by lentivirus transduction. scFvCD20:sTRAIL fusion protein produced a potent inhibition of cell proliferation in CD20-positive BJAB cells,moderate inhibition of the growth of Raji cells,and weak inhibition in CD20-negtive Jurkat cells when compared with ISZ-sTRAIL treatment,and it had no effect on normal human peripheral blood mononuclear cells (PBMCs). The MSC. scFvCD20:sTRAIL treatment significantly inhibited the tumor growth when compared with those treated with MSC. ISZ-sTRAIL. Conclusion: A double-target therapeutic system is well established, in which HUMSCs migrated to tumor site, secreted a novel fusion protein scFvCD20:sTRAIL,and thus locally concentrated scFvCD20:sTRAIL extended antigen-restricted anti-tumor activity. The engineered HUMSCs secreting scFvCD20:sTRAIL showed potent effect on inhibiting tumor growth in BJAB lymphoma malignancy,which may play an essential role in the clinical research .

AIM:To investigate whether oxidative stress is able to induce autophagy in mesenchymal stem cells (MSCs), and to explore the effects of autophagy on MSC proliferation and apoptosis under oxidative stress circumstance as well as the underlying mechanism for promoting the therapeutic effects of transplanted MSCs on treating diabetes mellitus e -rectile dysfunction ( DMED) .METHODS: Hydrogen peroxide ( H2 O2 ) was applied to simulate the oxidative stress cir-cumstance.The effects of H2 O2 at concentration of 0, 50, 100, 200, 400μmol/L on the viability of MSCs were tested by the method of Trypan blue exclusion and MTT assay respectively .The methods of MTT assay , Western blot and transmis-sion electron microscope ( TEM) were used to explore the effects of H 2 O2 on MSC apoptosis and autophagy .RESULTS:The proliferation of MSCs was obviously inhibited by H 2 O2 in a dose-dependent manner ( P<0.01) and the 50%inhibiting concentration (IC50) was (384.58 ±16.89) μmol/L.H2O2 induced apoptosis and autophay of MSCs .The proliferation rate of MSCs was suppressed by H 2 O2 significantly ( P<0.05 ) , with a further decline by blockade of autophagy ( P<0.05) whereas increased by blockade of apoptosis (P<0.05).H2O2 induced MSCs apoptosis obviously (P<0.05), with an augment of apoptosis ( P<0.05) by blockade of autophagy .Furthermore, the H2 O2 increased expression of cleaved caspase-3 and cleavage of poly ADP-ribose polymerase 1 (PARP1), Which were decreased by apoptosis blockade whereas were enhanced by blockade of autopahgy .CONCLUSION:Oxidative stress plays a dual role in MSC survival , which in-duces MSC apoptosis and autophagy .Moreover , blockade of autophagy intensifies MSC apoptosis .Therefore , it is a promis-ing method to ameliorate the effects of stem-cell based therapy on DMED by enhancing protective autophagy to increase the survival rate of transplanted MSCs against oxidative stress circumstance caused by diabetes mellitus .

Objective To investigate the effects of Guiqi polysaccharide (GQP) on the telomerase activity in premature senescence of WI-38 cells;To discuss its mechanism of action.Methods MTT assay was used to observe the effects of different concentrations of H2O2 on the proliferation of WI-38 cells and screened the best concentration of H2O2 to induce premature senescence cellular model in WI-38 cells. ELISA and chemical staining method were used to evaluate the protection of GQP in telomerase activity and senescence-associated β-galactosidase activity in premature cellular senescence.Results 200μmol/L H2O2 treatment could induce premature senescence in WI-38 cells. There were no significant differences in senescence-associated β-galactosidase and telomerase activity between GQP group and normal control group (P>0.05), but had significant difference with model group (P<0.01).Conclusion GQP can increase the telomerase activity and inhibit the senescence-associated β-galactosidase activity in premature senescence of WI-38 cells.

Objective To explore the feasibility of establishing a swine model of liver cirrhosis with portal hypertension by portal infusion of 80％ alcohol.Methods A total of 13 Guizhou miniature pigs were randomly divided into three groups,experiment group 1 (n=5),experiment group 2 (n=5) and control group (n=3).Experiment groups of pigs received portal infusion of 80％ alcohol in volumes of 5 ml in group 1,and 10 ml in group 2,and the pigs in control group received portal perfusion of saline in volumes of 10 ml.All animals were performed direct portal angiography,the portal vein pressures and diameter were also detected before,immediately and 6 weeks after the infusion.All animals underwent liver biopsies before and 6 hours,1-6 weeks after operation.And contrast-enhanced abdominal CT was performed before and 6 weeks after operation.All animals were dissected 6 weeks after operation,aud each leaf of liver specimens were performed histological examination.Results There was no statistically significant difference of the portal venous pressure and diameter before infusion and 6 weeks after infusion in the experiment group 1 and control group (all P＞0.05).In the experiment group 2,compared with pre infusion,the portal vein pressure and diameter were higher than those of immediately and 6 weeks after infusion (all P＜0.05).In both experiment group 1 and group 2,all pigs had developed into liver fibrosis,the METAVIR score of 2 pigs in group 1 and 5 pigs in group 2 respectively were up to grade 4.Conclusion Portal infusion of 80％ alcohol is more suitable for establishing a swine model of liver cirrhosis with portal hypertension.

Aim To investigate the expression of AB-CB5 and MDR1 in the cell line KG1 a and samples from acute myeloid leukemia ( AML) and their effects on multidrug resistance. Methods The expression of ABCB5 and P-gp ( the expressed product of MDR1 ) in KG1 a cells were detected by flow cytometry as well as Western blot analysis; KG1 a cells were transfected with the specific siRNA of ABCB5 using lipo2000 to reduce the expression of ABCB5; intracellular rhoda-mine123 was measured by flow cytometry;cell viability was detected by MTT; the expressions of ABCB5 and MDR1 in samples from AML were detected by real time PCR. Results ABCB5 and P-gp were overexpressed in KG1 a;the specific siRNA of ABCB5 transiently in-hibited the expression of ABCB5 in KG1 a; the siAB-CB5-KG1 a cells increased the intracellular rhodamine 123 and have been more sensitive to adriamycin com-pared with the parent KG1a. ABCB5 gene expression in samples from AML was higher than healthy people. Further, the expression of ABCB5 in 38 relapse or re-fractory AML significantly exceeded the 33 drug sensi-tive. And we found a significant positive correlation between ABCB5 expression and MDR1 gene expression in the 38 patients with relapse or refractory AML. Conclusion ABCB5 , as well as P-gp contributes to mediate multidrug resistance of AML, which provides a novel target for the therapy of relapse or refractory AML.