Objective To produce the recombinant NS3 protease of the hepatitis C virus in insect cells. Methods The gene of HCV serine proteinase domain which encodes 181 amino acids was inserted into pFastBacHTc and the recombinant plasmid pFBCNS3N was transformed into DH10Bac competent cells for transposition. After the recombinant bacmid had been determined to be correct by both blue-white colonies and PCR analysis, the isolated bacmid DNAs were transfected into Sf9 insect cells. The bacmid DNA was verified to replicate in insect cells and package into baculovirus particles via PCR and electronic microscope analysis. The insect cells infected by recombinant baculovirus were determined by SDS-PAGE and Western-blot assays. The recombinant protein was soluted in N-Lauryl Sarcosine Sodium (NLS) and purifed by metal-chelated-affinity chromatography, then the enzymatic activity was measured with the protein substrate NS5ab and the antigenicity of recombinant protease was determined by enzyme-linked immunoabsorbent assay. Results The HCV NS3 protease domain was expressed in insect cells at high level and it was partially resolved in NLS;Totally 0.2 mg recombinant serine proteinase domain with high purity was obtained by metal-chelated-affinity chromatography from 5?10 7 cells and both enzymatic activity and antigenicity of the protein were proven to be good. Conclusion The recombinant HCV NS3 protease expressed in insect cells is a membrane-binding protein with good enzymatic activity and antigenicity.

Objective To evaluate the reliability and validity of Bowel Function Index applied to assess the constipation induced by strong opioid drugs for patients with cancer pain.Methods 126 patients with constipation induced by strong opioid drugs for pain caused by cancer were selected.BFI,Patient Assessment of Constipation Symptoms(PAC-SYM) and clinical observations were used to collect data which were conducted for reliability analysis,correlation analysis and tests for several independent samples to evaluate the reliability and validity.Results The internal consistency determined by Cronbach αt was 0.86 for the total BFI score.Significant correlations were found among BFI,PAC-SYM and clinical observations.The BFI total score and each item scores of three groups all showed significant differences.Conclusions The Chinese version of BFI is a simple,valid and reliable,clinically relevant tool to assess the constipation induced by strong opioid drugs for pain caused by cancer.

Hypoxia/ischemia is a common clinical pathophysiological process and cause of death,and it is a common problem in extreme conditions such as high altitude,astronautics and diving.Hamartin is a kind of effective endogenous neuroprotectant and could increase cell tolerance to acute hypoxia or ischemia, thus,is of significance to research.The role of hamartin in hypoxia/ischemia has been a research focus of many scientists.Elucidating the related protective effect and its DNA methylation on hypoxia/ischemia can not only reduce injury,but also lay a basic for further studying the role of hamartin and its DNA methylation in other pathophysiological processes and provide theoretical guidance for the following clinical study.In this paper,we review the structure,mechanism and role of hamartin and the effect of its DNA methylation on hypoxia/ischemia.

Objective To investigate X‐ray ,CT and MRI features of synovial chondromatosis of the temporomandibular joint (TMJ) .Methods X‐ray ,CT and MRI features of eight patients of synovial chondromatosis of TMJ with histo‐pathologically con‐firmed were analyzed retrospectively .X‐ray examination and CT scanning were performed in all eight patients .Routine MRI scanning was performed in six patients and contrast‐enhanced MRI scanning was performed in two patients synchronously .Results Tumors occured unilaterally in all eight cases ,which occured on the right TMJ in six cases and on the left side in two cases .On X‐ray films , widen joint space and calcificated loose bodies occured in all eight cases .On CT scanning ,cystic‐solid mixed mass around the joint and calcificated loose bodies occured in all eight cases .On MR scanning ,multiple nodular long T1 and short T2 signal occured in six cases . Arthroedema and synovial hyperplasia with iso T1 and iso or slightly long T2 signal in six cases .On contrast‐enhanced MR ,homoge‐neous enhancement occurred in svnovial tissue and the edge of loose bodies in two cases .Conclusion The synovial chondromatosis of TMJ owns typical imaging features .The imaging findings can serve as a reference to improve diagnosis of synovial chondromatosis of TMJ .

Objective To investigate the clinical application of interventional combination embolization of spleen and kidney in treating traumatic splenic and renal hemorrhage. Methods Since Jan. 2010 to early 2014, a total of 24 patients with traumatic splenic and renal hemorrhage were admitted to authors’ hospital. After angiography, super-selective embolization with Gelfoam particles was employed to occlude the bleeding arteries. The results were analyzed. Results Technical success with single procedure was achieved in all the 24 patients. All 24 patients survived the trauma. The complication was mild. Follow-up examination showed that the embolization results were perfect. Conclusion For the treatment of traumatic splenic and renal hemorrhage, interventional combination embolization of spleen and kidney is safe and effective with reliable clinical results, and this technique can reduce visceral function damage to the greatest possible advantage.

Purpose To investigate the diameter change of dorsal root ganglion (DRG) in lumbar disc herniation using three-dimensional MR neurography. Materials and Methods Sixty-ifve patients with lumbar disc herniation and 30 healthy volunteers were selected. Bilateral DRG diameter was measured using MR three-dimensional constructive interference steady state (3D-CISS) sequence at the level of L3-S1 in the control group and at the level of herniation disc in patient group including central and lateral subgroups. The relationship between the sagittal index and DRG diameter at the level of herniation disc was analyzed. Results In the control group, the DRG diameters increased from the level of L3 to S1. The DRG diameters of the central subgroup were bigger than those of the control group (t=-2.485--2.253, P<0.05). The DRG diameters of the lateral subgroup were bigger on the diseased side than the contralateral side (t=1.956-2.432, P<0.05). The DRG diameters of the contralateral side in lateral subgroup were slightly bigger than those of control group without statistical signiifcance (t=-1.248--0.981, P>0.05). The sagittal index was not correlated with DRG diameter. Conclusion 3D-CISS sequence clearly demonstrates morphological changes of lumbosacral nerve root and measures its diameter.

Objective To establish a mouse model of prostate cancer expressing human PSCA for the development of new anti-tumor drugs or vaccines. Methods The total RNA of DU145 cells,a human prostate cancer cell line,was isolated by using TRIzol reagent according to the (RT-PCR),the first-strand cDNA was synthesized using the SuperScript First-Strand synthesis system. The human PSCA gene was amplified with the primers and cloned into the plasmid pcDNA3.1 to generate pcDNA-PSCA. DNA sequencing was used to confirm the constructs. The mouse prostate tumor cell line RM-1 cells,syngeneic to C57BL/6,were transfected with pcDNA-PSCA plasmids followed by selection using G418. RT-PCR analysis was performed to examine the validity of the constructs. Expression of PSCA on the cell surface was determined by staining with anti-PSCA antibody,and the anti-PSCA antibody was detected using an FITC-conjugated goat anti-rabbit IgG antibody,and analyzed by flow cytometry. 4-6-week-old male C57BL/6 mice purchased from the Laboratory Animals Center were inoculated with different amounts of RM-PSCA cells to search for suitable cell population which can form tumor in mouse,and the mice were monitored twice a week. The growth and the survival time of mice were measured,respectively. The tumor volume was measured by vernier caliper according to the formula:V=0.5a×b~2,where a and b are the long and short diameters of the tumor,respectively. Results The plasmid pcDNA-PSCA was successfully constructed and the PSCA was successfully expressed in RM-PSCA 7~# and RM-PSCA 28~# cells by RT-PCR and confirmed by flow cytometry. 1×10~5 RM-PSCA cells were sufficient to get tumor growth in 100% of inoculated mice. The tumor grew quickly and the volume of the tumor reached 12000 mm~3 within 34 days. All the mice died within 40 days and their mean survival time was 37 days. Conclusion A PSCA-expressing tumor model in mice has been successfully established. It can be used to evaluate the activities of drugs or vaccines.

Objective To explore the effect of auricular plaster therapy on associated symptoms and quality of life in patients with constipation induced by strong opioid drugs for pain.Methods A total of 60 patients selected were randomly divided into the intervention group and the control group with 30 cases in each group.The intervention group was treated with auriculotherapy based on the pattern/syndrome differentiation after detecting the auricular sensitive acupoints' distribution in patients with opioid-induced constipation after receiving opioid therapy.BFI,self-made scale on symptoms accompanied by constipation and PAC-QOL were used to collect data about constipation symptoms and the level of quality of life before or after the intervention,and still with a subset of the intervention group followed for more than two weeks.Results The BFI and PAC-QOL scores of the intervention and control group were lower after intervention,and the decline score of the two groups showed significant difference.But for the self-made scale,the score of each item was slightly lower compared with that before treatment,which showed no statistically significant difference between groups.A follow-up of 2 weeks showed that the long-term efficacy of auricular plaster therapy was significantly lower than before,but was less desirable than short-term.Conclusions Scientific and reasonable choice of acupoint prescription according to syndrome differentiation can effectively improve symptoms accompanied by constipation and patients' quality of life.

This paper reports a study regarding the verity of the hypocholestero-lemic effect of konjac-polysaccharide. The konnyaku powder (KP) used in this study was prepared and refined from the tubers of Amorphophallus ko-njac K. Koch and contained 84.8% of glucomannan. Male and female Spra-ue-Dawley rats aged 5 weeks were divided into 5 groups and fed on normal basal diet, hypercholesterolemic diet (control diet) and 3 test diets (i.e. KP was added to the control diet at a dosage of 2.5%, 5% or 10%) respe- ctively, for 12 weeks.The results obtained from this study showed that KP could markedly lower the level of the cholesterol in sera and livers of rats feeding hyper-cholesterolemic diets. At the end of the 4th week of the feeding experiment, the serum cholesterol level of the 5% and the 10% KP groups, and the liver cholesterol level of the 10% KP group were shown to be significantly lower than those of the control group. At the end of the 12th week, serum cholesterol levels of all the 3 KP groups were found to be lowered to the level of the normal group and so did the liver cholesterol level of the 10% KP group. The lipotropic (anticholesteatosis) effect of KP was also confirmed by the hepatic histopathological examination. Besides the hypocholest-eroletmic eftect, KP diets can also increase the bulk of stool. Finally, there were not any harmful effects on the absorption and utilization of Ca, Fe, Zn and Cu being found.

Objective To express human-mouse chimeric antibody against anthrax protective anti-gen and to analyze its biological activities. Methods A new mammalian bipromoter expression vector was constructed with dihydrofolate reduetase(DHFR) gene as the selection and complication marker. First, the light and heavy chain variable region gene of the monoclonal antibody 5E1 were cloned by RT-PCR, at the same time the human IgG1 heavy chain constant region gene and kappa type constant region gene were cloned. Next, the human-mouse chimeric antibody genes were synthesized by fusion PCR. Then, the hu-man-mouse chimeric antibody gene were inserted into MCS of pSecTag and B1 to construct pSecTag-5E1L and B1-5E1H, respectively. Finally, heavy chain expression cassette excised from the B1-5E1H with Bgl Ⅱ/BamH Ⅰ was further cloned into the Bgl Ⅱ site of the pSecTag-5E1L to construct pSecTag-5E1. Plasmid pSecTag-5E1 was transfected into CHO(dhfr) engineering cells and high production cell clones that were screened by enhancing MTX concentration. After collecting medium and purifying chimeric antibody with af-finity chromatogram, purified chimeric antibody was analyzed by SDS-PAGE, Western blot. Results A sta-ble and high production cell line was acquired at MTX concentration 5×10~(-8) mol/L. Conclusion The hu-man-mouse chimeric antibodies were successfully expressed in CHO cells.