Objective To investigate the significance of human cyomeg Movirus(HCMV)pp65 IgG antibody avidity index(AI)for the clinical diagnosis of HCMV primary infection through the experimental model of HCMV primaly infection in BALB/c mice.Methods 6～8 weeks,female,specific-pathogen-free BALB/c mice were divided into 5 groups.6 mice in each group. And he mice were injected with 2×106 PFU/m1,2×105 PFU/mi,2×104 PFU/ml,2×103 PFU/ml and 2×102 PFU/ml of HCMV intraperitoneally respectively. Another 6 mice were injected intraperitoneally with the maximum dose of HCMV kept at 56℃ for 30 min as inactivated virus group.And HF negative control group was established at same time. All the mice ere sacrificed to obtain brain and lung tissues for the following experiments after 1 montll.(1)Tissue samples obtained from mice were inoculated in human embryo fibroblasts(HF)monolayers after routine treatment for virus isolation.HCMV specific eytopathie effect(CPE) was observed bv inverted phase-contrast microscopy.HCMV UL83 DNA in the ultures as tested by PCR and pp65 antigen was detected by indirect immunofluorescence.(2)Extracted mRNA from tissue samples and HCMV pp67 mRNA were analyzed by reverse transcriptase PCR(RT-PcR).(3)Immunoglobulin M(IgM)antibody and immunoglobulin G(1gG)antibody avidity Was investigated for their usefulness in distinguishing primary genital HCMV infections rom nonprimary infections with ELISA kit using truncated pp65 protein.ResultsHCMV can be isolated in the tissues from the mice injected with 2×106 PFU/ml and 2×105 PFU/m1.RT- PCR and ELISA showed positive results in the same groups.The infective rates were 100%.The analysis of the low doses groups,inactivated group and HF negative ontrol group all showed negative results.Conclusions BALB/c mice can be infected with HCMV and appeared as primary infection after1 month.Determination of HCMV pp65 IgM and HCMV p065 IgG-AI by ELISA incorporated with virus isolation and RT-PCR are helpful for distinguishing primary infections from nonpfimary infections.The detection of HCMV p65 IgM and HCMV pp65 IgG-AI by ELISA utilizing recombinant protein pp5 as antigens can be used for preliminary screening.

Objective To study the value of the liver and spleen interventional therapy in primary carcinoma of the liver with hyperspleenism. Methods The treatment was given to 25 patients. Using the method of Seldinger,the 5FRH into artery hepatica propria to infuse 2/3 of the overall dose of chemotherapy medicine and embolism was inserted into the liver segment. Then 1/3 of it was infused into artery lienalis and gelatin sponge was infused into artery lienalis Rr. lienales. Combined regimens were used in chemotherapy: 5-Fu 750mg, EADM 40mg, MMC 10mmg,and 10 of the 25 added DDP 40mg. LUF and gelatin sponge were used for embolisation. Results Of the 25 patients,the WBC and PLT increased in 25 patients 4 weeks after the operation. The difference was significant. The survival rate at 6 and 12 months was 100% and 60% respectively,with 1 case of CR,8 cases of PR and 6 cases of SD. Patients had fever after the treatment but few with abnormal liver function tests. Conclusion Liver and spleen interventional therapy may relieve hyperspleenism and suggests that the treatment of primary carcinoma of the liver is effective.

Objective To establish a new ELISA method by use of shorted recombinant antigen pp65 for detection of IgM antibodies to human cytomegalovirus (HCMV).Method According to HCMV pp65 sequence of nucleotide and amino acid, the shorted pp65 was decided by computer soft ware of protein and attained through gene engineering technique. The by computer soft ware of protein and attained through gene engineering technique. The recombinant enzyme- linked immunosorbent assay (REC-ELISA ) method was developed using pp65 recombinant protein as antigen and was applied to detecting anti-HCMV-IgM in sera. It was compared to ELISA by use of whole virus as antigen and Biocheck CMV IgM enzyme immunoassay test kit.Results To detect HCMV IgM by REC-ELISA, the best quantity of pp65 was 3.5?g percent cavity, the best dilution of HRP~*anti-HCMV IgM was 1∶[KG-*2]800 and that of serum was 1∶[KG-*2]100. The positive coefficient of variation (CV) was 9.5% in stability assay. The average CV of Inter- assay and Intra-assay was 4.5% and 9.6% respectively. The positive value of REC- ELISA was 44%, that of ELISA using whole virus was 50% and BioCheck was 45%. REC-ELISA using most suitable condition was concordant with BioCheck, which was 97.0%. Its youden′s index was 92.8% and its specificity (98.2%) was higher than that of ELISA using whole virus as antigen (90.9%), too.Conclusion REC-RLISA had good sensitivity, specificity and reproducibility. The method was easy and quick. Recombinant protein was harmless and easy gained compared with the whole virus. It can be large-scale production and standardization. Therefore, the application value and potential of REC-ELISA was very large.

Objective To define the nuclear agrresome formation is determinated by which part of the UL76 of
HCMV. Full-length, conserved N terminal and unconserved C terminal of pUL76 were constructed to eukaryotic ex-pression plasmid pEGFP-N1 . Methods Primers were designed to amplify full-length and different part of pUL76 according to standard sequence of HCMV AD169 which had been submitted to GenBank(FJ527563. 1). These frag-ments were constructed to eukaryotic expression plasmid pEGFP-N1 . The recombinant plasmids were designated pEGFP-UL76,pEGFP-UL76N, pEGFP-UL76C respectively. Double digestion and sequencing were performed to verify the accuracy of recombinant plasmids construction. Empty vector and three recombinant plasmids were transi-ent transfected to HELF and HepG-2 cells respectively. Reverse transcriptation PCR and Western blot were per-formed to detect the RNA and protein expression level respectively. Different nuclear aggresome formations were visualized with an Olympus fluorescence microscopy. Results pEGFP-N1 and pEGFP-UL76N were unable to in-duce nuclear aggresome formation, whereas pEGFP-UL76 and pEGFP-UL76C were able to elicit nuclear aggresome formation. Conclusion The unconserved C terminal of pUL76 is sufficient to induce nuclear aggresome formation.

Objective To explore the value of automatic photography in the observation of results of Schistosoma japoni?cum miracidium hatching experiments. Methods Some fresh S. japonicum eggs were added into cow feces,and the samples of feces were divided into a low infested experimental group and a high infested group(40 samples each group). In addition,there was a negative control group with 40 samples of cow feces without S. japonicum eggs. The conventional nylon bag S. japonicum miracidium hatching experiments were performed. The process was observed with the method of flashlight and magnifying glass combined with automatic video(automatic photography method),and,at the same time,with the naked eye observation meth?od. The results were compared. Results In the low infested group,the miracidium positive detection rates were 57.5% and 85.0%by the naked eye observation method and automatic photography method,respectively(χ2=11.723,P<0.05). In the high infested group,the positive detection rates were 97.5%and 100%by the naked eye observation method and automatic pho?tography method,respectively(χ2= 1.253,P > 0.05). In the two infested groups,the average positive detection rates were 77.5% and 92.5% by the naked eye observation method and automatic photography method,respectively(χ2 = 6.894,P <0.05). Conclusion The automatic photography can effectively improve the positive detection rate in the S. japonicum miracidi?um hatching experiments.

Objective: To realize the requirement of adjacent area accuracy,axial surface physical contour and smoothness in the process of inlay computer-aided design(CAD).Methods:A compromised method was brought forward that some assistant 3-D constraint lines from occlusion to gingiva were used to lead the inlay contour on axial corner and other characterized areas,and two perpendicular 3-D constructing lines at adjacent area were used to construct accurate 3-D configuration of this area.The axial surface was completely defined by the network of specified curves which were two constructing lines and four borders.Results:Inlay axial surface was smooth,the contour was consistent with the residual tooth body and the 3-D configuration in adjacent area was accurate.Conclusion:It is feasible to realize the inlay axial surface accuracy,smoothness,and consistency using the method of special distributed 3-D B-Spline curve network.

Objective To detect CK-19 mRNA expression by quantitative real-time RT-PCR in axillary drainage fluid of rectal cancer and investigate its clinical significance.Methods Axillary drainage fluids were collected from 59 patients with rectal cancer and 15 patients with benign abdominal lesion from Sep.2010 to Dec.2010.Level of CK-19 mRNA in axillary drainage fluid was detected using specific primers by real-time RTPCR.The data were statistically analyzed to investigate the relationships between CK-19 mRNA level and tumor invasion,lymph node status,tumor stage and tumor differentiation level.Results The positive rate of CK-19 mRNA expression in patients with rectal cancer was 67.8％,which was significantly higher than that in patients with benign abdominal lesion.The expression of CK-19 mRNA was significantly correlated with the depth of tumor invasion,lymphnode status,tumor stage and histopathological differentiation( P ＜ 0.05 or P ＜ 0.01 ).Ck129 mRNA expression was associated with the pathological level,the higher of the lymph node translation level,the higher expression in the axillary drainage fluid after rectal cancer surgery (r =0.674,P =0.021 ).The lower of the lymph node differentiation level,the higher expression in the axillary drainage fluid after rectal cancer surgery (r =-0.741,P =0.014).Conclusion Quantitative detection of CK-19 mRNA in axillary drainage fluid of rectal cancer by RT-PCR could enhance the diagnostic sensibility of colorectal cancer micrometastases.RT-PCR assay is suitable for predicting peritoneal micrometastasis of rectal cancer,which is a reference for postoperative treatment and prognosis prediction.

Objective To explore the change of opioid peptide content such as ?-endorphin(?-EP), dynorphinA_ 1-13(DynA_ 1-13) in plasma and cerebrospinal fluid(CSF) of patients with cerebral hemorrhage, and the interference effect of Naloxone on them.Methods 60 patients with cerebral hemorrhage were randomly divided into Naloxone-treated group and control group. Conventional treatment was used in both groups, then Naloxone-treated group intravenous drip of Naloxone, each 3.0 mg, once a day, for 14 d. The concentrations of ?-EP, DynA_ 1-13 of the patients were measured with radioimmunoassay (RIA) during the acute stage before treatment and at 7th, 14th day after treatment compared with those of normal group (the normal people and the patients were treated by operations). Changes of ?-EP and DynA_ 1-13 in bleeding part and amount of bleeding were analyzed. The scores of GCS and NDS before and after treatment were examined at the same time.Results (1) The content of ?-EP, DynA_ 1-13 in plasma and CSF of patients with cerebral hemorrhage were obviously higher since acute stage than those of the normal group(all P

Objective To define that human cytomegalovirus (HCMV) can cross the placenta of the Balb/c mice and induce liver damage in developing fetus. Methods HCMV AD169 (6.0 logTCID 50 , 3.0 logTCID 50 , 1.5 logTCID 50 per mouse respectively) was injected into the peritoneum of mice (half of mice were female) when they were about 10 weeks old (weight 25 30g). Then, these mice were paired to mate. Fetus on day about to give birth was removed from the uterus and liver was obtained for virus isolation, pathological studies, examination of the viral DNA positive cells by in situ hybridization using digoxigenin labelled HCMV DNA oligonucleotide probe. Results HCMV could be isolated from the supernatant of tissues; HCMV DNA was found by PCR in supernatant of cell culture with CPE; the presence of viral DNA sequence in hepatocytes was confirmed by in situ hybridization; pathological changes in liver consist of swollen cytoplasm and destroyed nuclei of hepatocytes and distinct intranuclear inclusion in hepatocytes with a cellular infiltrates of predominantly phagocytic cells. All above were found more obviously in fetal mouse liver tissues from the group inoculated with 6.0 log TCID 50 HCMV AD169 as compared with 3.0 log TCID 50 group. In contrast, these positive results couldn't be well found in 1.5 log TCID 50 group. Nothing could be found in normal controls. Conclusions Our research suggests that primary maternal HCMV infection during pregnancy could induce congenital infection in fetus by transplacental transmission and induce fetal liver damage. The mouse model will provide the basis for the study on pathogenesis of congenital HCMV infection in liver and the development of prevention, diagnosis and antiviral agents for congenital HCMV infection in human being.