Objective To explore the protocol of tissue matching and anti-rejection therapy in highly sensitized patients (HSP). Method The panel reactive antibody (PRA), human leukocyte antigen (HLA) matching and renal transplantation outcomes of 45 HSPs were retrospectively analyzed. Results Hyperacute rejection occurred in 2 patients. Acute rejection occurred in 9 patients and reversed by anti-rejection therapy. One year patient/graft survival rate was 95.6% / 91.1% respectively. Conclusions To avoid specific antibody through HLA matching is the key point for successful renal transplantation of HSP. Antithymocyte globulin (ATG) induction therapy combined with tacrolimus, mycophenolate mofetil therapy can decrease the rate of acute rejection and prolong graft survival.

Objective To obtain the recombinant urease B subunit (ureB) of Helicobacter pylori (Hp) and apply it in serological detection of Hp infected patients. Methods Urease B subunit gene was amplified from the complete genome of Helicobacter pylori by PCR, and cloned into the PinPoint TM Xa Ⅲ fusion expression vector, then was sequenced. The protein of urease B subunit was expressed in E.coli JM109 and purified with affinity chromatography. 113 serum samples of peptic ulcer patients were detected by ELISA combined with purified UreB protein. Results DNA sequence analysis showed the nucleic acid sequence homology of ureB gene was 96.44% and the putative amino acid sequence homology was 99.65%. The recombinant UreB protein was composed of 571 amino acid residues and kept original immunologic reaction with corresponding antibody, and its purity was over 90% after affinity chromatography. The results of ELISA associated with recombination UreB antigen showed the sensitivity and specificity was 92%, 98.5%. Conclusion The recombinant UreB protein will be of value for clinical serodiagnosis and epidemiological study of Helicobacter pylori.

Objective To explore the feasibility and efficacy of desensitization protocol for highly sensitized renal transplant patients (HSP). Methods Thirty-five HSPs ( HLA class-Ⅰ panel reactive antibody ＞50 %), including 27 patients with a positive T and/or B cell cytotoxicity crossmatch (XM) and 8 patients with a negative T and B cell XM, received plasmapheresis plus intravenous immunoglobulin (PP-IVIG)treatment. Results The positive XM was rendered negative by PP-IVIG treatment in 25 of 27 (92.6 %)HSPs, and subsequent transplantation was performed. Two patients did not receive renal transplants due to persistent positive XM. In 25 patients receiving renal transplants, no hyperacute rejection occurred. There were 8 cases of acute rejection, including 5 cases of acute humoral rejection (AHR). All rejection episodes were reversed. During a follow-up period of 52 ± 26 months, the serum creatinine levels at 12th and 24th month were 112± 18 and 130 ± 38 mol/L respectively. The 1- and 3-year graft survival rate was 96. 0 %and 80. 0 % respectively. Conclusion The desensitization therapy by PP-IVIG is effective for HSP. High rate of AHR is the major defect of this protocol. The short-term graft survival rate after this protocol is acceptable but the long-term survival rate needs to be defined.

OBJECTIVE: To explore the effect of artesunate (ART) on cell differentiation and cell cycle distribution of the prostate cancer cell line PC-3 in vitro. METHODS: PC-3 cells were cultivated with ART from logarithmic growth phase. After 48-hour treatment, the cell cycles were detected by flow cytometry (FCM), and enzyme linked immunosorbent assay was used to detect the level of prostate specific antigen (PSA) in cell culture supernatant. The change of cellular morphology was observed under a transmission electron microscope (TEM). RESULTS: In comparison with the blank control group, the rate of G(0)/G(1) plus S stages of PC-3 cells was significantly decreased in the high-dose ART group. The PC-3 cell was arrested in G(2)/M by ART. The rates of G(2)/M of the high-dose ART group and the medium-dose ART group were obviously higher than those of the blank control group and the cisplatin group (P<0.05). The levels of PSA in the three ART groups were significantly lower than that of the normal control group (P<0.05, P<0.01). In the ART groups, TEM showed that some vacuoles appeared in endochylema, cell polarity was enhanced, cell nucleus leaned to one side of the cell, and microvilli increased on the other side of the cell. CONCLUSION: ART can induce PC-3 cell cycle arrest and differentiation in vitro.

Objective To evaluate the analytical performance of Mindray BS‐820 automatic biochemical analysis system used in primary hospitals in Guangzhou area for verifying whether its performance meeting the clinical requirements .Methods Accord‐ing to the EP5‐A2 ,EP6‐A ,EP7‐A2 and EP‐17A files recommended by CLSI and the Pharmaceutical Industry Standard of the Peo‐ple′s Republic of China YY/T0654‐2008 ,the precision ,linear rang ,anti‐interference capability ,sensitivity and carry‐over rate of the Mindray BS‐820 automatic biochemical analysis system were evaluated by adopting 16 routine biochemical items .Results The pre‐cision results of all 16 analyzing items conducted by the Mindray BS‐820 automatic chemical analysis system conformed to the re‐quirements .The results of 15 items showed good linear relation during the testing range (r≥0 .997 9) .In 16 biochemical items ,the anti‐interference capability of TBIL ,TP ,TC ,TG ,LDL ,HDL ,UREA and Ca to bilirubin ,triglyceride and hemoglobin conformed to or were higher than the anti‐interference capability declared by manufacturer ,but among other 8 items ,the anti‐interference capabil‐ity of 3 items was lower than that declared by manufacturer .The limit of blank(LOB) of all 16 analyzing items was less than LOB in the kit instruction .The carry‐over rate of Glu was 0 .02% ,less than 0 .50% .Conclusion This instrument has good precision , sensitivity and low carry‐over rate(0 .02% ) ,the linear range is ideal ,r≥0 .997 9 ,the anti‐interference capability basically satisfies the clinical needs .Therefore this instrument is suitable for the use in the middle‐small scale laboratory department .

Objective To investigate the epitope of HLA class I specific antibody in the serum of highly sensitized patients (HSP). Methods The specificities and epitopes of HLA class I specific antibody from 54 HSP were monitored continually by enzyme linked immunosorbent assay (ELISA). Results 90.7 % of HSP had public epitopes of HLA class I antibody. The common public epitopes were 163E, 142 145TTKH, 83R, 71 74AQTD, 127K, 163R, 163L, 41T, 62 66RNTRN and 41 45ASPRT. The public epitopes were stable in the course of continual monitoring, and it could reflect antibody spectrum more accurately than antibody specificity analysis. Conclusion Most HSP produce HLA class I antibodies in a fairly predictable fashion. These antibodies direct against a small number of high frequency public amino acid residues with high immunogenicity. Analysis of public epitopes of HLA antibodies is very useful to predict acceptable mismatches.

Objective:To analyze the polymorphism of urease B gene(ureB) from different Hp strains. Methods:The sequences of ureB gene from 14 different Hp strains were amplified by PCR to analyze PCR-RFLP with HaeⅢ digestion.At the same time,the sequence of ureB gene was analyzed with biochemical software to compare the homology of nucleotide and amino acid sequence and to profile the phylogenetic tree. Results: The results of 1.7 kb ureB gene digested with HaeⅢ showed there were 2-5 band types in 14 different Hp strains and formed five distinct RFLP types.The two reference strains had the same RFLP type as 3 clinical isolates while the other clinical isolates and 3 isolates from animal model belong to 4 different RFLP types respectively.The nucleotide sequences and putative amino acide sequences of Hp ureB were compared.The various ureB sequences had high homologies(more than 96.6% and 98% in nucleotide sequences and amino acide sequences respectively) among 14 Hp strains.Particularly,there was the highest homologies(100%) between CCS9801、CCS9806、M3 and M10.Phylogenetic tree analysis showed that two reference strains and other isolates from clinical patients and Hp-infected mice model were located in two different lineage respectively in phylogenetic tree of nucleotide sequences while there were some variance in phylogenetic tree of amino acide sequences. Conclusion: Hp ureB was high conserved and homologous in the gene level as well as in the protein level.

Objective To investigate the effects of injecting allogeneic mesenchymal stem cells(MSCs) on the cellular immunity in rat in vivo.Methods Bone marrow-derived MSCs were isolated from Wistar rats.The purity of MSCs was identified by morphological examination with microphotography,and the phenotypes were identified with flow cytometry.Twenty SD rats were randomly divided into 4 groups.Different concentrations of MSCs(5?106/ml for group A,5?105/ml for group B,and 5?104/ml for group C,respectively) and PBS(for group D) were given to allogeneic SD rats via intravenous infusions.The suppressive effect of MSCs on lymphocytes proliferation in recipient rat was analyzed using mixed lymphocyte cultures(MLR) 10 days after cultivation.At the same time,proportions of CD4+,CD8+ and CD4+CD25+/CD4+ T-lymphocytes in peripheral blood and spleen were analyzed with flow cytometry.Results Proliferation rate of splenic lymphocytes in group A(5?106/ml MSCs,8.58%?0.27%) was markedly lowered compared with that in group D(PBS,24.40%?5.21%,P

[Objective]This study was designed to investigate capability of the conditioned media that originated from Renca cells to convert CIM~+ CD25~- T cells into CD4~+ CD25~+ T cells,which can exert immunosuppressive effect on effector T cells in vitro and in vivo.[Methods]The common media were mixed with the conditioned media at different ratios,and fresh enriched CD4~+ CD25~- T cells with MACS were cultured in mixed media for 7 days.At end-point of culture,the cells were collected and detected phenotypes in flow cytometer.Moreover,we detected immunosuppressive effect of converted CD4~+ CD25~+ T cells on effeetor T cells proliferation in one-way mixed lymphocytes reaction by using CCK-8,and we observed survival time and histology of grafts.The delayed type hypersensitivity was determined 14 days after transplantation.[Results]The mixed media could increase ratio of CD4~+ CD25~+ Foxp3~+ T cells in conditioned media ratio-dependent(P＜0.05),compared with control groups,when the mixed media contained no mote than 75% of conditioned media.The converted CD4~+ CD25~+ T cells significantly suppress proliferation of effector T cells in vitro,and prolong survival time of grafts,which were(29.6±1.4)d in converted CD4~+ CD25~+ T cells treated groups(P＜0.05),compared with that in untreated groups(9.8±0.6 d)or PBS treated groups(10.9±0.6 d).Moreover,delayed-type hypersensitivity reaction were conducted at day 14 after transplantation in the recipients,and the results showed that less pad swelling in the group treated with converted CD4~+ CD25~+ T cells than other control groups was found,according to measurement of pad swelling.In addition,progressed to complete necrosis of grafts were exhibited in the mice treated with PBS and untreated mice,whereas better healing of grafts and less lymphocytes infiltration were displayed in the mice treated with converted CD4~+ CD25~+ T cells,which were similar to the mice treated with natural regulatory T cells.[Conclusion]The converted CD4~+ CD25~+ T cells with Renca conditioned media play suppressive role in vitro and in vivo.

Objective To explore the feasibility of establishing mini-pig model of type 1 diabetes by single intrave-nous injection of high dose streptozotocin .Methods Eight male Zhonghua mini-pigs (22.6 ±1.8 kg) were administrated with high-dose streptozotocin (150 mg/kg) into the ear vein .Before and 10, 30, 90 minutes, 1, 3 and 7 days after adminis-tration of streptozotocin , blood samples were obtained respectively , and used to dynamically monitor the fasting blood-glucose. C peptide and insulin levels were evaluated by IVGTT test .Results Since 24 hours after STZ administration , the fasting blood-glucose level was increased significantly compared with that of pre-administration and maintained at 16.7-20.6 mmol/L, while the C peptide and insulin levels were decreased significantly .IVGTT results showed that blood sugar levels at 1 h af-ter intravenous injection of 50%glucose were much higher than 11.1 mmol/L and failed to restore to fasting glucose levels until 2 h, insulin and c-peptide did no response after injection of glucose , always kept at a trace level .Conclusion A sin-gle high-dose streptozotocin injection can be used to establish a mini-pig model of type 1 diabetes successfully .