Biomedical subjects

N Otting

Publications and source records attributed to N Otting.

43 records · Page 3Linked to original sources

PubMed1991

Polymorphism of C4 and CYP21 genes in various primate species.

To study the genetic heterogeneity of the C4 and CYP21 genes in selected primate species we used the technique of restriction fragment length polymorphism (RFLP). Genomic DNA was digested using several restriction endonucleases and filters were hybridized with a 500 bp BamHI/KpnI fragment derived from the 5' section of a human C4-cDNA and with a 1700 bp BamHI obtained from a human CYP21 gene. Abundant RFLP heterogeneity was observed for the C4 genes within a rhesus monkey population but not for the chimpanzee colony analyzed. Duplicated C4-CYP21 clusters can be traced back in humans, but also in chimpanzees, orang-utans and rhesus monkeys. Thus, duplication of the basic C4-CYP21 cluster in primates may have happened more than 30 million years ago. Non-duplicated C4-CYP21 regions were found for the gorilla and orang-utan. Apart from this, shortened C4A and C4B genes were observed in chimpanzees, orang-utans and rhesus monkeys, whereas the so-called long variety of the C4A gene appears to be present in humans and orang-utans. This ancestral modification, resulting from an insertion of a 6.5 kb intron in the C4A gene, therefore predates at least speciation of human and orang-utan which is estimated to have taken place more than 12 million years ago.

Animals↗

PubMed1990

Major histocompatibility complex class II-restricted antigen presentation across a species barrier: conservation of restriction determinants in evolution.

The existence of at least three alleles of the HLA-DRB3 gene within the human population is evident. These alleles express DRw52 determinants and react with monoclonal antibody (mAb) 7.3.19.1. The polymorphic epitope recognized by 7.3.19.1 is not only present on human cells but is also expressed on chimpanzee (Pan troglodytes) class II-positive cells. The 7.3.19.1 determinant already existed before speciation of man and chimpanzee, and is at least 5,000,000 yr old. Two-dimensional gel electrophoresis demonstrated that the various HLA- and Patr-DRw52 molecules that are reactive with 7.3.19.1 exhibit isoelectric point differences due to primary amino acid heterogeneity, as was confirmed by sequencing data. Sequence comparison allowed us to map the binding site of mAb 7.3.19.1 to the alpha helix of the major histocompatibility complex (MHC) class II DRB1 domain surrounding the antigen-binding cleft. Despite MHC sequence variation, chimpanzee antigen-presenting cells can present antigen (purified protein derivative) to human T cell lines and vice versa. Only the HLA- and Patr-DRw52 molecules were shown to function as restriction elements for antigen presentation across this species barrier. It is concluded that these particular restriction determinants probably have been conserved in evolution. The HLA- and Patr-DRw52 molecules represent alleles displaying polymorphism that has been selected for in evolution. Such "biomutants" may thus be more useful to study the biological significance of MHC molecules than MHC variants that have been generated by in vitro mutagenesis experiments.

Amino Acid Sequence↗

PubMed1990

The chimpanzee major histocompatibility complex class II DR subregion contains an unexpectedly high number of beta-chain genes.

The major histocompatibility complex (MHC) class II DR subregion of the chimpanzee was studied by restriction fragment length polymorphism (RFLP) analysis. Genomic DNA obtained from a panel of 94 chimpanzees was digested with the restriction enzyme Taq I and hybridized with an HLA-DR beta probe specific for the 3' untranslated (UT) region. Such a screening revealed the existence of 14 distinct DRB/Taq I gene-associated fragments allowing the definition of 11 haplotypes. Segregation studies proved that the number of chimpanzee class II DRB/Taq I fragments is not constant and varies from three to six depending on the haplotype. Comparison of these data with a human reference panel manifested that some MHC DRB/Taq I fragments are shared by man and chimpanzee. Moreover, the number of HLA-DRB/Taq I gene-associated fragments detected in a panel of homozygous typing cells varies from one to three and corresponds with the number of HLA-DRB genes present for most haplotypes. However, a discrepancy is observed for the HLA-DR4, -DR7, and -DR9 haplotypes since a fourth HLA-DRB pseudogene present within these haplotypes lacks its 3' UT region and thus is not detected with the probe used. These results suggest that chimpanzees have a higher maximum number of DRB genes per haplotype than man. As a consequence, some chimpanzee haplotypes must show a dissimilar organization of the MHC DR subregion compared to their human equivalents. The implications of these findings are discussed in the context of the trans-species theory of MHC polymorphism.

Animals↗

PubMed1990

Phenotypic and functional changes of tumour cells from patients treated with monoclonal anti-idiotypic antibodies.

In this paper data are presented indicating that immunotherapy with monoclonal anti-idiotypic antibodies (MoAb anti-id) can provoke different responses in the B-cell tumour concerned. With respect to the course of disease during and after immunotherapy, the in vitro findings may very well explain the in vivo observations in the two patients (D.E.F., B.O.R.) with B-cell chronic lymphocytic leukaemia (B-CLL) who were treated with MoAb anti-id. After initial tumour reduction, there was a recurrence of tumour cells with altered functional and phenotypic properties. In both cases the recurring tumour cells still expressed the same idiotype. In one patient (D.E.F.) the phenotypic changes (a surface Ig change from IgM, IgG, IgA, and IgD to weakly positive IgM and IgD) and functional changes (a 10-fold increase in [3H]thymidine uptake and a decreased idiotype secretion in vitro), together with the in vivo findings with respect to the course of disease--at relapse an impressive tumour regrowth rate with constant serum idiotype level--suggest that immunoselection might have taken place favouring the survival and relapse of a less mature, more aggressive tumour cell population with a lower idiotype expression. In the second patient (B.O.R.), the phenotypic changes (an isotype change from IgM and IgD to IgM with the loss of IgD, and a gradual decrease in expression of CD19 and CD24) and functional changes (a 10-fold increase of idiotype secretion in vitro), together with the in vivo finding that the serum idiotype level had increased 25-fold compared with the preimmunotherapy serum level with comparable tumour load, strongly suggest an immunotherapy-induced differentiation of the malignant B cell. We also describe an increased expression of CD74, detected by MoAb BoM22, on the recurring tumour cells of patient B.O.R., whereas the expression of HLA-DP, -DQ and -DR did not change. The significance of this finding is unclear.

Antibodies, Anti-Idiotypic↗

PubMed1989

RFLP analysis of the HLA-, ChLA-, and RhLA-DQ alpha chain gene regions: conservation of restriction sites during evolution.

Genomic DNA samples, derived from a panel of 60 chimpanzees and 45 rhesus monkeys, were digested with the restriction enzymes Taq I and Bgl II and hybridized with an HLA-DQ alpha chain cDNA probe. The results were compared with the data available on a human reference panel. Use of the restriction enzyme Taq I and the DQ alpha chain probe allows the detection of five HLA-DQA1 and two HLA-DQA2 gene-associated fragments within the human population. For the ChLA and RhLA systems, 3 and 7 different DQA1-associated restriction patterns were detected, respectively, while for the chimpanzee a nonpolymorphic DQA2 (DX alpha) gene-associated fragment was also observed. The equivalent of the HLA- and ChLA-DQA2 genes appears to be absent in the rhesus monkey. The ChLA-DQA1 and -DQA2 gene-associated RFLP patterns are identical in man and chimpanzee, whereas such restriction site conservation is not seen in the rhesus monkey. The conclusion drawn is that the genetic organization of the HLA-DQA and ChLA-DQA gene regions, and possibly some of their allelic variabilities, already existed before man and chimpanzee separated in evolution. Moreover, the particular duplication which led to the generation of the HLA- and ChLA-DQA2 genes must have happened before speciation of members belonging to the superfamily Hominoidea (man, chimpanzee, etc), but probably after the separation of superfamily Cercopitecoidea (rhesus monkeys, baboons, etc.) from Hominoidea.

Animals↗

PubMed1988

Molecular analysis of HLA-DP specificities HLA-DPw1, -DPw2 and -DPw4: DP beta chain heterogeneity correlates with PLT subtyping.

HLA-DP molecules were isolated from Epstein-Barr virus transformed B cell lines by immunoprecipitation with monoclonal antibody B7/21.2 and subsequently analysed by two-dimensional gel electrophoresis. The results obtained demonstrate that the HLA-DP molecules that can be isolated from cells positive for the HLA-DP specificities HLA-DPw1, -DPw2 and -DPw4 display DP beta chain isoelectric point differences, whereas no DP alpha chain polymorphism was observed. These results suggest that the PLT defined HLA-DP specificities (HLA-DPw1, -DPw2 and -DPw4) are probably DP beta chain structures.

Cell Line↗

PubMed1987

Molecular diversity of HLA-DQ. DQ alpha and beta chain isoelectric point differences and their relation to serologically defined HLA-DQ allospecificities.

HLA-DQ molecules were isolated from a panel of HLA-DR-DQ homozygous cell lines, partially of consanguineous origin, derived by the use of monoclonal antibody SPV-L3, and subsequently analyzed by gel electrophoretic techniques. It is demonstrated that both the DQ alpha and beta chain exhibit an extensive isoelectric point polymorphism. Within a panel of 29 B-cell lines tested, at least 5 distinct alpha and 6 distinct DQ beta chain gene products were observed. Of the 30 theoretically possible DQ alpha-beta dimers, only 10 could be identified within the panel: 5 different dimers are associated with the DQw1 allospecificity; HLA-DQw2 and -DQw3 are associated with 2 types of dimers, whereas another DQ alpha-beta combination was expressed by a cell line with a so-called DQ-blank specificity. The relation between the specificities 2B3 and TA10 appeared to be complicated as far as DQ beta chain isoelectric point differences are concerned: monoclonal antibody IIB3 seems to be reactive with four distinct DQ beta chain alleles whereas monoclonal antibody TA10 only reacted with one type of DQ beta chain. These results suggest that the polymorphic DQ alpha and beta chains may both contribute to the definition of the HLA-DQ allospecificity. A particular DQ beta chain was present in two types of HLA-DQw1 molecules, as well as in one type of HLA-DQw2 and -DQw3 (2B3 positive) molecule, and formed dimers with electrophoretic distinct DQ alpha chains. On the other hand, HLA-DQw2 molecules isolated from HLA-DR3-positive cells and one type of HLA-DQw3 (TA10 positive) molecule were found to be constructed of identical alpha chains but appeared to differ in the composition of their DQ beta chain gene products. The implications of these findings will be discussed.

Alleles↗