Insulin-dependent diabetes mellitus (IDDM) is associated with CTLA4 polymorphisms in multiple ethnic groups.

Linkage disequilibrium (association) analysis was used to evaluate a candidate region near the CTLA4/CD28 genes using a multi-ethnic collection of families with one or more children affected by IDDM. In the data set unique to this study (Spanish, French, Mexican-American, Chinese and Korean), the transmission/disequilibrium test (TDT) revealed a highly significant deviation for transmission of alleles at the (AT)n microsatellite marker in the 3' untranslated region (P = 0.002) and the A/G polymorphism in the first exon (P = 0.00002) of the CTLA4 gene. The overall evidence for transmission deviation of the CTLA4 A/G alleles is also highly significant (P = 0.00005) in the combined data set (669 multiplex and 357 simplex families) from this study and a previous report on families from USA, Italy, UK, Spain and Sardinia. Significant heterogeneity was observed in these data sets. The British, Sardinian and Chinese data sets did not show any deviation for the A/G polymorphism, while the Caucasian-American data set showed a weak transmission deviation. Strong deviation for transmission was seen in the three Mediterranean-European populations (Italian, Spanish and French) (P = 10(-5)), the Mexican-American population (P = 0.002) and the Korean population (P = 0.03). These results suggest that a true IDDM susceptibility locus (designated IDDM12) is located near CTLA4.

[The kinetics of cell-mediated cytotoxicity to schistosomula of Schistosoma japonicum with sera from infected mice at different duration].

The tests of antibody-dependent cell-mediated cytotoxicity (ADCC) were performed to observe the kinetics of in vitro killing of Schistosoma japonicum schistosomula mediated by eosinophils, macrophages or neutrophils with infected mouse sera of different duration. The results showed that when complement was not involved, the killing rates of schistosomula mediated by eosinophils or macrophages began to increase significantly at 4 wk post-infection, reached a plateau in 5-7 wk and 6-8 wk respectively, and then declined to the levels of that at 4 wk till 11 wk. In case of neutrophils, there was no significant cytotoxicity detected. When complement was involved, all the three effector cells could mediate cytotoxicity to schistosomula, with the killing rates higher than those without complement at the correspondent time intervals. These indicate that ADCC appears to be an important ingredient in the acquired resistance to S. japonicum, and demonstrate that eosinophil- and macrophage-mediated cytotoxicity to schistosomula is complement-independent, whereas neutrophil-mediated cytotoxicity to schistosomula is complement-dependent. This method may be of some value for choosing optimal time for vaccination and estimating the effectiveness of a candidate vaccine.

[Study on McAbs against protein "target antigen" in Schistosoma japonicum].

In the studies presented here, we demonstrated the feasibility of producing large number of hybridoma cell lines secreting McAbs against S. japonicum including 16 cell lines secreting IgG McAbs (7 for IgG1, 5 for IgG2a, 1 for IgG2b, 3 for IgG3) and 13 McAbs for IgM, on the basis of successfully extracting 24-26kD and 90kD "target antigen" proteins known as important antigens for inducing schistosome protective immunity. All the above cell lines were characterized for localization of the McAbs, mediating in vitro ADCC against schistosomula, epitope recognized by the McAbs on different kinds of schistosome antigens. Studies have shown the evidences that the McAbs can be used to identify the "target antigen" on the surface of schistosome, to isolate and purify "target antigen" of S. japonicum by chromatography column bearing McAbs, as well as to immunize animals as antigens for accumulation of data in the development of vaccine against S. japonicum via anti-idiotype antibodies.

[Screening of Schistosoma japonicum adult worm genomic DNA library by specific antibody].

S. japonicum adult worm genomic DNA libraries were screened by enzymeimmunoassay. The antigens produced by recombinant lambda gt 11 plaques were transferred to nitrocellulose filters. Clones encoding given antigens were detected with infected rabbit sera (IRS) which were preabsorbed with lysate of induced lambda gt 11 in Y1090 cells. Eight putative positives were picked up from 97 plates in the primary screening and were rescreened a second time, and, 2 of them consistently gave good positive signals in subsequent screenings. One of the phage clones was purified to homogeneity and mixed with Y1089 cells. The expressed products still showed positive when rescreened by ELISA, indicating that the clone encoding S. japonicum antigen could be recognized by IRS. The immunoscreening method described here was able to efficiently isolate single clones encoding S. japonicum antigens. The method has the advantages of screening libraries efficiently, reliably and specifically, and it might open the way to screen genes encoding S. japonicum antigens inducing protective immunity (Fig. 1).