BACKGROUND:Matrix metaloproteinase-1 can degrade extracelular matrix, which is mainly colagen type I, and has the potential to reverse fibrosis tissue. OBJECTIVE:To construct the recombinant adenovirus vector containing human matrix metaloproteinase-1 (hMMP-1) gene with GatewayTM Clone Technology, and observe the capacity of degrading colagen type IIIin vitro. METHODS: The gene hMMP-1 was amplified by using PCR from the pcDNA3.1 plasmid and was cut down by the double endonuclease. The linear gene fragment was connected to the entry vector pENTERTM 1A. Then the entry clone and the destination vectors pJTI? R4 Dest CMV-N-EmGFP pA Vector recombined using the LR reaction to form the expression clone pAd-hMMP-1-eGFP. The linear pAd-hMMP-1-eGFP cut down by endonucleasePac I was transfected into HEK293A cels to packaging the Ad-hMMP-1-eGFP. The transfected situation was observed under a fluorescence microscope, the target protein expression was detected by western-blot assay and RT-PCR. Cels can be divided into three groups: blank control group: HEK293A cels, AD-EGFP group: HEK293A cels were infected by Ad-eGFP, AD-HMMP1-EGF group: HEK293A cels were infected by Ad-hMMP1-eGFP and colagen type III. The content of colagen type III was detected by ELISA kits after 24, 48 and 72 hours. RESULTS AND CONCLUSION: It was confirmed that the entry vector and the destination vector both contained hMMP-1 target gene by restriction analysis and sequencing. The green fluorescent protein was observed in the 293A cels transfected by the Ad-hMMP-1-eGFP at 4 days. The fluorescence intensity was the highest at 10 days. The virus was colected at 12 days, the viral titer was determined as 4.84 × 1010 PFU/mL, the target protein was efficient expressionvia western-blot assay. Blank control group and AD-EGFP group had no obvious change of colagen content with the extension of time. The rate of colagen degradation in AD-HMMP1-EGFP group was 24%, 56% and 81% respectively at 24, 48, 72 hours. AD-HMMP1-EGFP group degraded colagen significantly compared with the other two groups (P < 0.01). The recombinant adenovirus vector containing hMMP-1 was successfuly constructed by using the Gateway technology, this method was more efficient and specific than with the traditional methods. The hMMP1 degraded colagen type III significantlyin vitro.

Two patients underwent liver transplantation of blood type incompatibility were collected from Fuzhou General Hospital of Nanjing Military Area Command of Chinese PLA. Case 1: A male who had primary hepatic carcinoma underwent classic orthotopic liver transplantation; the blood of donor was type A, and the blood of recipient was type O. Case 2: A female having history of type B hepatitis underwent classic orthotopic liver transplantation due to pregnancy combining with severe liver disease and coagulation disorder; the blood of donor was type B, and the blood of recipient was type O. Immunohistochemistry staining was used to observe pathological changes and deposition of various immunoglobulin and complement in two cases following liver transplantation of blood type incompatibility under optic microscope and to investigate diagnostic standard of humoral rejection. The results showed that linear or granular depositions of IgG, IgM, IgA, C4c, C4d, and Clq were found in endothelial cells of hepatic sinusoid, suggesting that IgG and other immunoglobulin exhibited a strongly positively diffused deposition on the endothelial cells of hepatic sinusoid, while expression of C4d and other complements was also found. All those mentioned above could be considered as evidences to prove onset of humoral rejection in transplanted liver tissue.

Objective To detect and explore the expression of ARD1 and its clinical significance in the nasopharyngeal in-flammatory tissue ,nasopharyngeal carcinoma group and its subgroups .Methods Expression of ARD1 in nasopharyngeal carcinoma (56 cases) and nasopharyngeal inflammatory tissue (20 cases) were detected by immunohistochemical staining SP ,the correlation between the expression of ARD1 and age ,gender ,histological grade ,TNM clinical stage and tumor metastasis were analysed . Results The positive expression rate of ARD1 were 10 .00% (2/20) ,55 .35% (31/56) in the nasopharyngeal inflammatory tissue and nasopharyngeal carcinoma ,respectively .The expression level of ARD1 in nasopharyngeal carcinoma was significantly higher than in the nasopharyngeal inflammatory tissue ,the difference was significant (P< 0 .05) ;expression of ARD1 in the nasopharynge-al carcinoma was correlated with the histological grade of nasopharyngeal carcinoma(P< 0 .05) and the expression was increased in poor differenciation tissue .But there was no statistical difference between the expression of ARD1 and the patient′s age ,gender , TNM clinical stage ,tumor metastasis(P> 0 .05) .Conclusion The expression of ARD1 is high in nasopharyngeal carcinoma ,and has a closely correlation with diffferentiation level of tumor ,which suggested that ARD1 may be involved in the the occurrence and development of nasopharyngeal carcinoma .However ,further research needs to be done for its mechanism in the nasopharyngeal car-cinoma .

98%), the peak level of uptake occurred earlier, intracellular fluorescence intensity maintained much more stable. Expressions of CD19+, CD22+, CD20+ increased significantly. A_~570 values of MNCs proliferation and IgG levels in supernatant were all higher. CONCLUSION: CD40 ligand-PLL carrier system may delivery CpG ODN targeting to B lymphocytes, enhancing its immunological efficiency.

BACKGROUND: CpG oligodeoxynucleotide (ODN) is a type of highly effective immune adjuvant with low toxicity, which has an extensive application in gene therapy for many diseases. However, the specificity for species and cells leading to low uptake by cells and degradation by nuclease blocks its clinical application. OBJECTIVE: To explore the specific delivery and its immunologic efficacy of CpG ODN targeting B lymphocytes of umbilical cord blood by CD40 ligand-receptor-mediated carrier system. DESIGN, TIME AND SETTING: An observation and control experiment was performed at the Department of Hematology, and Department of Pediatric, the First Affiliated Hospital of Jinan University from April 2004 to October 2007. MATERIALS: Fresh umbilical cord blood with heparin was obtained from healthy, natal infant. Informed consent was obtained from his parents, and the experiment was approved by the hospital Ethics Committee. METHODS: CD40 ligand (CD40L)-EDC-PLL-CpG ODN conjugated complex was prepared. Mononuclear cells (MNCs) from umbilical cord blood were co-cultured with conjugated complexes. Uptake rate, mean fluorescence intensity of FAM marked CpG ODN, expressions of MNCs, proliferations of lymphocytes and the IgG levels of culture supematants were detected by flow cytometry, fluorescence techniques, MTT assay and ELISA, respectively. MAIN OUTCOME MEASURES: The uptake rate, the mean fluorescence intensity of CpG ODN by MNCs, subgroups and proliferations of lymphocytes, and IgG levels of culture supematants. RESULTS: Compared to the pure CpG ODN group, the uptake rate of the conjugated complexes group was higher (98%), the peak level of up-taking occurred earlier, and intracellular fluorescence intensity maintained much more stable. Expressions of CD19+, CD22+, and CD20+ was increased, A value and IgG levels in supematants were all higher than that of the control group. CONCLUSION: CD40 tigand-receptor-mediated carrier system is helpful for CpG ODN delivery targeting to B lymphocyte, enhancing its immunological efficiency.

The curcumin prodrugs, which could be selectively activated in tumor cells, were prepared to establish a basis for the targeted chemotherapy for cancer. On the basis of the molecular structure of curcumin, the N-maleoyl-L-valine-curcumin (NVC), N-maleoyl- glycine-curcumin (NGC) were chemically synthesized and identified by IR and NMR spectroscopy. After treatment with these two prodrugs for 6 - 24 h, the rates of growth inhibition on human bladder cancer EJ cells and renal tubular epithelial (HKC) cells were detected by MTT colorimetry. Our results showed that after the treatment with 20 micromol/L - 40 micromol/L NVC and NGC for 6 - 24 h, the growth inhibitory effects on EJ cells were 6.71% - 65.13% (P < 0.05), 10.96% - 73.01% (P < 0.05), respectively, in both dose- and time-dependent manners. When compared with the curcumin of same concentrations, the growth inhibitory effects of these two prodrugs on HKC cells were significantly decreased (P < 0.01). It is concluded that activation of curcumin prodrugs via hydrolysis functions of cellular esterase could inhibit the growth activities of tumor cells, and reduce the side effects on normal diploid cells. This provided a novel strategy for further exploration of tumor-targeted chemotherapeutic drugs.
Antineoplastic Agents, Phytogenic/*pharmacology ; Curcumin/*pharmacology ; Dose-Response Relationship, Drug ; Prodrugs/*chemical synthesis ; Prodrugs/*pharmacology ; Tumor Cells, Cultured ; Urinary Bladder Neoplasms/*pathology

The effect of Smac gene on the TRAIL-induced apoptosis of the prostate cancer cell line PC-3 and the molecular mechanism were investigated. The Smac gene was transfected into PC-3 cells under the induction of liposome. The intrinsic Smac gene expression was detected by Western blotting. After treatment with TRAIL as an apoptosis inducer, in vitro cell growth activity was assayed by MTT colorimetry. The apoptosis rate of PC-3 cells was determined by annexin V-FITC and propidium iodide staining flow cytometry. The expression of cellular XIAP and caspase-3 genes was examined by Western blotting. Smac-transfected cells (PC-3/Smac group) had significantly increased Smac protein level as compared with PC-3 controls (P<0.01). After induction with 100-200 ng/mL TRAIL for 12-36 h, cellular proliferation rate in PC-3/Smac group was significantly lower than in PC-3 controls (P<0.05). After induction with 100 ng/mL TRAIL for 24 h, the apoptosis rate in PC-3/Smac group was significantly enhanced as compared with that of PC-3 controls (P<0.05). Accordingly, the XIAP expression level was down-regulated significantly (P<0.05) and caspase-3 subunit P20 was up-regulated significantly (P<0.05). It is suggested that the over-expression of cellular Smac can inhibit inhibitor of apoptosis proteins (IAPs), enhance caspases activity and the apoptosis rate of PC-3 cells induced by TRAIL, which may provide a useful experimental basis for prostate cancer therapy.

Objective To analyze the advantages and disadvantages of minimally invasive video -assisted thyroidectomy(MIVAT)and totally endoscopic thyroidectomy (TET).Methods The study included 70 patients undergoing MIVAT and TET at Qilu Hospital of Shandong University and Yantai Yuhuangding Hospital from Jan . 2012 to Dec.2012.46 patients undergoing MIVAT(MIVAT group)and 24 patients undergoing TET(TET group) were analyzed retrospectively .Data of gender,age,operative data,intraoperative and postoperative complications , length of stay , feelings of patients and cosmetic effect were collected and analyzed .MIVAT and TET were com-pared in terms of safety ,trauma and cosmetic effect .Results ①About safety:no permanent recurrent laryngeal nerve(RLN)palsy and parathyroid injury occurred in either group .One case in MIVAT group and no case in TET group was converted to open surgery (2.2%vs 0%,P>0.05) .②About trauma:the intraoperative blood loss and postoperative drainage amount in the MIVAT group were significantly less than those in the TET group ( P <0.05).Postoperative VAS score at 6 hours was similar,but MIVAT group showed lower postoperative VAS score than TET group at 12 hours and 24 hours ( P<0.05 ) .No significant difference was observed between the 2 groups about C reactive protein (CRP)on the 1st day before operation,the 1st and 3rd day after operation(P>0.05).③About cosmetic effect:the 2 groups had similar patient satisfaction index .Conclusions The study shows similar results in safety and trauma .The 2 approaches are both safe and valid .Patients using the two ap-proaches are equally satisfied with the cosmetic effect .

Objective To establish standard parameters of cervical alignment and cervical range of motion(ROM) in asymptomatic population,and to explore the influential factors such as age,sex and cervical disc degeneration.Methods The cervical standard lateral,flexion and extension plain radiographs of 212 asymptomatic volunteers were analyzed retrospectively.The volunteers,including 128 females and 84 males with ages ranging from 20 to 79 years,were divided into 6 groups from 3rd to 8th decade of life and were divided into 4 groups based on the scoring system of cervical disc degeneration.These plain films were blindly measured by 3 spine surgeons with Mimics software,and each spine surgeon mcasured them for 3 times.Several parameters,including C2-C7 cervical alignment,total ROM,flexion ROM,extension ROM and segmental ROM were measured.The score of disc degeneration were assessed from C2-C3 to C6-C7.Pearson correlation analyses was used to quantify the relation between cervical alignment and total ROM.Multiple linear regression analyses were required to account for influential factors.Inter-and intrarater correlation coefficient was analyzed.Results The C2-C7 cervical alignment was 21.40°± 12.15°,and the total ROM was 63.59°± 15.37°.Sex had a significant impact on the cervical alignment (regression coefficient was-2.472,P ＜ 0.05).Both sex and age had significant impacts on the total ROM (regression coefficient was 3.863 and-6.463 respectively,P ＜ 0.05).Sex had a significant impact on C2,3 and C5,6 segmental ROM; age had a significant impact on all of the five segmental ROM from C2,3 to C6,7; cervical disc degeneration had a significant impact on the C4-5,C5-6 and C6-7 segmental ROM.The cervical alignment had no significant association with both of the extension and total ROM (r=-0.106 and 0.215,respectively,P ＞ 0.05),but had a significantly negative association with flexion angle (r=-0.401,P＜ 0.05).The measurement of cervical alignment,total ROM and segmental ROM showed excellent intra-rater agreement and excellent inter-rater agreement.Conclusion Sex is an influential factor of the cervical alignment.Sex and age are two influential factors of the total ROM.Sex,age and cervical disc degeneration are influential factors of the segmental ROM.The cervical alignments do not have an impact on total ROM.

Objective To investigate the relationship between serum HBeAg level and inflammation grade (G)/fibrosis stage (S) in the liver tissues of chronic hepatitis B (CHB) patients in the immune clearance phase (IC). Methods Both liver biopsy samples and serum samples were consecutively collected from CHB patients in Liver Center,First Affiliated Hospital,Fujian Medical University during March 2007 to June 2010.Electro-chemiluminescence and fluorogenic quantitative polymerase chain reaction (PCR) methods were used to determine HBeAg titer and hepatitis B virus (HBV) DNA level,respectively.The relationships between HBeAg titer and liver pathological stages were analyzed using Spearman rank correlation analysis.Receive operating characteristic (ROC) curve was used to evaluate the diagnostic value of HBeAg for liver pathological stages.Results Totally 249 patients with CHB were enrolled into this study.The serum HBeAg absorbances in patients with liver inflammation G1 to G4 were (2.93±2.85),(2.96±2.74),(2.69±2.67) and (2.30±2.41) lg s/co,respectively,while those in patients with liver fibrosis S1 to S4 were (2.99±2.74),(2.89±2.73),(2.58±2.55) and (2.32±2.44) lg s/co,respectively,which indicated that serum HBeAg titers were significant different in patients with different grading and staging of liver tissues (x2 =47.13,P＜0.01; x2 =74.12,P＜0.01).Spearman rank correlation analysis showed that serum HBeAg titer was negatively correlated with inflammation grades and fibrosis stages of liver tissues (r=-0.418 and-0.532,respectively; both P＜0.01).ROC curve analysis revealed that the areas under the curve (AUC) were 0.74 (G≥≥3) and 0.73 (G≥4),and the HBeAg (s/co) cut-off values were 2.95 and 2.64 lg s/co,respectively.Similarly,ROC curve analysis revealed that the AUC were 0.80 (S≥3) and 0.77 S≥4),and the HBeAg cut-off values were 2.99 and 2.82 lg s/co,respectively.Conclusions The serum HBeAg titer is negatively correlated with the inflammation grades and fibrosis stages m liver tissues of CHB patients in IC phase.The level of HBeAg may be used as an adjunctive noninvasive marker to reflect the inflammation and fibrosis status in the liver.