Objective To develop a PCR-SSP method for detection of HLA-DRB1 alleles in the patients who were hypersensitive to Platanus Acerifolia pollen allergen,and to probe into the association between the atopic subjects to Platanus Acerifolia pollen allergen and HLA-DRB1 alleles.Methods DNA in whole blood was extracted by phenol-chloroform method.Eight pairs of specific primers for alleles were synthesized,and HLA-DRB1*0401,*0402,*0403,*0404,*0405,*0406,*0407,*0408 alleles in 20 atopic patients and 36 healthy individuals of Jiangsu Province with Han nationality were detected by PCR-SSP(polymerase chain reaction-sequence specific primer).Results By optimizing the experimental conditions PCR-SSP methods for detection of the 8 alleles were established and the distributing data of above-mentioned HLA DRB1 were obtained.The frequency of HLA DRB1*0405 and *0406 in the patients group was higher than that of in healthy controls group,while the frequency of HLA DRB1*0402 in the patients group was lower than that in controls.No significant deference for the other 5 alleles was found between the 2 groups.Conclusion HLA-DRB1*0406和*0405 seems to be the likely suspected candidate alleles responsible for susceptibility to Platanus Acerifolia pollen allergen in the atopic patients,while DRB1*0402 might be contribute to the related resistance to the allergen.

Objective To evaluate if the assay of platelet-associated immunoglobulin G (PAIgG) can improve the sensitivity of monoclonal antibody immobilization of platelet antigens assay (MAIPA) in immune thrombocytopenic purpura (ITP).Methods Anti-GP Ⅱb/Ⅲa and anti-GP Ⅰb/Ⅸ autoantibodies were detected by a modified MAIPA;PAIgG was detected by competitive ELISA.MAIPA and PAIgG were done simultaneously in 190 patients with thrombocytopenia,in which 132 were diagnosed as ITP according to the diagnostic criteria Kappa test was used to analyze the concordance between MAIPA and PAIgG.Results The sensitivities of PAIgG alone or in parallel with MAIPA to ITP were 32.6% and 37.2% higher than MAIPA alone respectively,while their specifieities were 51.7% and 56.9% lower respectively.The sensitivity,specificity,positive predictive value,negative predictive value of PAIgG in series with MAIPA were all not superior to MAIPA alone.The kappa values between MAIPA and PAIgG in ITP and non-immune thrombocytopenia were 0.129 and 0.012 respectively,which meant the concordance between them was poor.Conclusion The detection of PAIgG should not be used as screening before MAIPA in distinguishing immune from non-immune thrombocytopenia.

To study the contribution of T cell subsets in the pathogenesis of Murine hepatitis virus Type3 (MHV-3) induced chronic viral hepatitis in C3H/Hej mice, ninety C3H/Hej mice were chosen to individually receive 10 plaque forming units (PFU) of MHV-3 intraperitoneally. The changes of virus titer and pathology in liver tissue were examined by standard plaque assay and by the hematoxylin/eosin (HE) staining method from 2 days post MHV-3 infection. The ratios of T cell subsets including CD3+CD4+CD8-, CD3+CD4-CD8+, CD3+CD4-CD8-, CD3+CD4+CD25+, CD3+CD4+CD25- and CD3+CD4-CD25+ T lymphocyte of total T lymphocytes in blood, spleen and liver were examined at 0, 2, 4, 6,8, 10, 12, 15, 20, 25, 30, 40 days post MHV-3 infection by flow cytosorting. We observed that the virus titer raised and showed persistent virus duplications and inflammatory changes in the livers of C3H/Hej mice from 2 days post MHV-3 infection. The double negative T cell (DN Treg cell) and CD4+CD25+ T cell ratios increased significantly from 2 days post MHV-3 infection in C3H/Hej mice, and CD3+CD4+CD8-, CD3+CD4-CD8+, CD3+CD4+CD25- and CD3+CD4-CD25+ T cell ratios decreased accordingly. In conclusion, the changes of virus titer and pathology in the livers of C3H/Hej mice post MHV-3 suggest their contribution to viral persistence. Further characterizations of DN Treg cells are that infection indicates that MHV-3 could induce the chronic inflammation in livers of C3H/Hej mice.The increase of the DN Treg cell and CD4+CD25+ T cell ratios in C3H/Hej mice post MHV-3 infection suggests that DN Treg cells and CD4+CD25+ T cells may both have important suppressive immunomodulation functions in the development of chronic viral hepatitis and have important roles in the virus persistent infection. Further characterizations of DNT cell and CD4+CD25+ T cell are under investigation.

Objective To establish a method for rapidly detecting the G-protein ?3 subunit (GNB3) 825 site single nucleotide polymorphism (SNP) and to analyse the relationship between GNB3 825 site gene SNP and obesity. Methods The real-time fluorescent PCR was employed to analyse the GNB3 825 site gene SNP of 420 samples from 21 provinces and the the frequencies of genotypes were compared with those detected by gene sequencing. GNB3 825 site genotype, body weight, body mass index (BMI) and fat content were examined from 207 subjects and the correlation between GNB3 825 site gene SNP and obesity was analysed. Results The result by real-time fluorescent PCR showed that the frequencies of 825T and 825C haploid were 46.90% and 53.10%, respectively, and the frequencies of 825TT, 825TC and 825CC genotype were 22.38%, 51.42% and 28.10%, respectively, with no other genotype detected, which was consistent with the result by gene sequencing. BMI and fat content were significantly higher in subjects with GNB3 825TT than in subjects with other genotypes. Body weight was much higher in subjects with GNB3 825TT genotype than in subjects with 825CC genotype, but not significantly different with 825CT genotype. Conclusion A new rapid method for the detection of GNB3 825 site SNP has been successfully established. There existed significant correlation between GNB3 825TT genotype and obesity.

The ability to induce hypersensity on leaves of tomato and the stability of double-plasmid of an harpin-producing, nitrogen-fixing engineered strain E4 were tested. Hypelsensitivity-inducing experiment indicated that the time and density of hypersensitivity-induction of E4 was similar to those of DH5, the positive control of pCPP430. Although E4 took the same time to induce hypersensitivity as 308R, another positive control of pCPP430, it induced weaker hypersen- sitivity on tobacco leaves. On tomato leaves, there was no difference in time and density of hypersensitivity between E4 and 308R (pCPP430). Results revealed that the two plasmids, pCPP430 and pMC73A, were unstable in host bacteria, with the losing rate of 100% at the 48th generation. The emergence probability of bacteria with either pCPP430 or pMC73A was almost the same.

Objective To study the survival,migration and differentiation of the neural stem cells which transplanted into ventral horn of spinal cord after brachial plexus avulsion.Methods Neural stem ceils isolated from spinal cord of neogenetic rats and cultured,expanded,labeled by BrdU before transplanted. Twenty adult healthy SD rats preformed as the model of brochial plexus avulsion(Roots C_(5～7)),then transplan- rod stem ceils into the C_6 ventral horn of spinal cord.On 1,2,4,8,12 weeks postoperatively,immunohisto- chemistry assay were carried out in the spinal cord.Results Transplanted into ventral horn of spinal cord after brachial plexus avulsion.Neural stem cells can survive,migrate for at least one segment of spinal cord and differentiate to neurons and astrocytes.The differentiation of stem cells were time-depends.Conclusion Neural stem cells can survive,migrate and differentiate after transplanted into ventral horn of spinal cord in the rats which suffered from brachial plexus avulsion.

Postoperative pancreatic leakage remains a persistent problem after pancreaticoduodenectomy.For patients with a soft and nonfibrotic pancreas,double binding continuous hemstitch suture is an optimal method for anastomosis.From January 2011 to June 2012,92 cases of periampullary carcinoma with a soft pancreas underwent pancreaticoduodenectomy,and then a modified technique of pancreaticogastrostomy was performed with 2 continuous hemstitch sutures placed in the mucosal and seromuscular layers of the posterior gastric wall,respectively.The median time for pancreaticogastrostomy was 12 minutes,and only 15 patients had postoperative complications.Two patients developed pancreatic leakage(1 grade A and 1 grade B)postoperatively.Pancreaticogastrostomy with double binding continuous hemstitch sutures is a simple and safe reconstruction procedure for patients with a soft and fragile pancreas who received pancreaticoduodenectomy.

OBJECTIVE: To establish a hepatocyte immunotoxicity model for screening of liver protective medications. METHODS: Cytotoxicity was induced by coincubating BCG-pretreated rat hepatocytes in vivo and with 10 mg/L LPS in vitro. Biphenyldimethylesterate (DDB), malotilate(MLT), silybin(SB) and glycyrrhizin (GRZ) were coincubated along with LPS to prevent the hepatocyte injury and verify the applicability and reliability of the model. AST, LDH and nitric oxide (NO) were measured in both the serum and supernatant. The liver and spleen index were calculated and the liver histopathologic changes were examined microscopically. RESULTS: Supernatant AST, LDH and NO in the BCG combined LPS group were increased in comparison with the control group (P<0.01). This increase was attenuated by the addition of DDB, MLT, SB and GRZ (P<0.05). The serum AST, NO and liver and spleen index were also increased significantly compared with the control group (P<0.01). Microscopic exam revealed serious histopathologic changes in the BCG combined LPS group. Hepatoxicity with associated liver enzyme elevation but histopathologic changes were attenuated by DDB, MLT, SB and GRZ. CONCLUSION: BCG combined LPS-induced hepatocyte immunotoxicity in an in vitro rat model may be a useful technique to assess the effectiveness of liver protective medications.

The release kinetics research of sustained-release formulations of traditional Chinese medicines (TCM) is an inalienable part of the chain of TCM modernization, which plays an important role in the development of modern compound TCM preparation. However, the research method or pattern in line with the specific characteristics of TCM, i.e., multi-component and multi-target, is still lacking. On the basis of material rough set theory, this paper reviewed the advantages and disadvantages of the existing evaluation patterns and methods, a tentative idea about the "total amount" release characteristics evaluation on TCM compound sustained-release preparation has suggested so as to evaluate the release kinetics and to promote the development of evaluation methodology on TCM sustained-release preparations.

Objective To search a new,safe and effective method to obtain semen using rectal Drobe electroejaculation(RPE).Methods RPE procedures were performed in 8 men with psychological ejaculatory failure,used the electrical stimulate instrument(CGS model,made in Italy).The average age was 26 years.Seminal vesicle was normal by B Uhrasonography or CT examination.Except for 1 case of diabetes,the other 7 cases found no hormones,blood,biochemical and nervous system abnormalities.Three cases had nocturnal emission,but no sexual intercourse ejaculation.Results Adequate sperm was successfully taken out from the 8 patients.Mean seminal fluid volume was 2.3 ml,mean total sperm count was 67×106/ml and mean total motile sperm was 21.3%.Three patients felt abdominal distension during the RPE course.Conclusion RPE is a safe,reliable,non-in-vasive,repeatable method to obtain semen.