Sujay K. Singh

Passive Transfer of Arthritis to Mice by Injection of Human Anti-Type II Collagen Antibody

The serum IgG fraction from a patient with seronegative rheumatoid-like arthritis which contained a high anti-type II collagen antibody titer was injected intravenously into mice susceptible to type II collagen-induced arthritis. A mild, transient, inflammatory arthritis was observed in 20 to 25% of the animals, whereas histologic signs of disease were evident in most of the injected mice. Purified human anti-type II collagen immunoglobulin injected into the knee joints of mice was also shown to induce a transient, inflammatory arthritis. Radiolabeled human anti-type II collagen IgG was shown to accumulate in the peripheral joints of mice, and the specificity of the antibody was shown to be similar to the specificity of anticollagen antibody eluted from the joints of mice with collagen-induced arthritis.

TRANSCRIPTIONAL CONTROL OF MHC CLASS II ANTIGEN EXPRESSION ON MOUSE T CELL LINES

We have analysed the factors which regulate MHC class II expression in mouse T cell lines. Two such lines, BW 5147 and PLT‐24.2, were used in this study. Using 5‐azacytidine (5 AzaC) we have shown that hypomethylation of DNA can induce class II antigen synthesis in BW 5147. The expression of class II in PLT‐24.2 cells seems to be under a different control mechanism. Southern blot analysis of I‐Aβ gene in PLT‐24.2 suggests that the expression of class II in this cell line is probably the outcome of a gene rearrangement. We hypothesise that insertion of viral long terminal repeats (LTR) next to the class II genes in transformed T cell lines can act as a promoter for the expression of class II antigens.

DNA restriction fragment analysis of Eαgenes in E-negative H-2 recombinant mouse strains

In the mouse, there are two classes of immune response (Ia) molecules, A and E, each with a distinct ~ chain with a molecular mass of 34 000 and a/3 chain with a molecular mass of 28 000. All the strains of mice have been found to express A molecules, while some strains do not express an E molecule on the cell surface (Jones et al. 1981). Mice With the b and s haplotypes synthesize only E;~ polypeptide chains, which can be detected in the cytoplasm but not on the lymphocyte cell surface. These mice have a deletion in the E~ gene that includes the promoter and part of the first exon; this prevents expression of the gene (Mathis et al. 1983). Similar deletions of the same size and location have been found in E-negative wild mouse strains (Dembid et al. 1984). Mice with q and fhaplotypes do not express an E molecule on the cell surface, and both the polypeptide chains are absent (Jones et al. 1981). Molecular genetic studies by Mathis and co-workers (1983) have shown that thefhaplotype synthesizes a normal amount of E~ mRNA which is predominantly of an aberrant size, while the q haplotype expresses very low levels of E~ mRNA. Recombinant mouse strains with crossovers between two E-negative haplotypes do not express an E molecule, as expected. In such recombinant mouse strains, the origin of the E~ allele cannot be determined by serology. These E~ genes can be identified by DNA restriction fragment analysis. In this study, we analyzed five E-negative recombinant mouse strains by DNA restriction fragment analysis (Table 1). They were typed by microcytotoxicity analysis for H-2K, H-2D, A, and E antigens. Immunoprecipitation analysis with anti-E serum confirmed the absence of E molecules. To determine if the B10.RBF, B10.RBQ1, B10. RKQ1, and B10. RKQ2 recombinant mouse strains have functional E~ genes, they were crossed to the E-negative f k strain A.TFR5 (E~E~) which has a functional E~ gene and a nonfunctional E f gene. If the recombinant has a functional E¢ gene, then transcomplementation will occur in F 1 between this E¢ polypeptide chain and the E~ gene from A.TFR5. These recombinants had functional E¢ genes derived from k or b haplotypes (Fig. 1). The recombinants were also analyzed by Ouchterlony immunodiffusion for S region-encoded C4 complement components Ss and Slp. The typing of B10.RBF and A.TFR4 showed that in these recombinants, the C4 genes are derived from thefhaplotype, which places the crossover point between the Ee gene and the C4 genes. The aUelic origin of the C4 genes of B10.RBQ1, B10.RKQ1, and B10.RKQ2 could not be determined by serology, since Ss and Sip antigens are not distinguishable between the b and q haplotypes. Thus, these studies placed the crossover in these recombinants between the E¢ gene and the H-2D gene. The five recombinant mouse strains were examined by restriction fragment analysis with two restriction endonucleases which cut within the E~ gene (Dembid et al. 1984). Recombinants B10.RBQ1, B10.RKQ1, and B10.RKQ2 have either the E~ or E q alleles, B10.RBF either

Mutation in the Aβ gene of B6.C-H-2bm12 generates unique T-cell recognition sites

Four functional Ia genes are coded within the I (immune response) region of mouse, e. g., A~, A~, E~, and E~ (Jones et al. 1978, Klein et al. 1981). In recent years using congenic inbred and recombinant strains of mice, several investigators have demonstrated a relationship between Ia molecules and immune response. The findings on the only known Ia mutant, B6.C-H-2 b.a2, have helped in further characterization of fine specificities on an la molecule. Tryptic peptide maps revealed a limited number of differences between B6 and bm12, mapping the mutation to the A beta polypeptide chain (McKean et al. 1981). The tryptic peptide studies showed that this mutation is not due to a single-point mutation, but involves a minimum of three different amino acid substitutions. Recently, McIntyre and Seidman (1984) have cloned and sequenced a gene coding the bml2 A-beta polypeptide chain. Three nucleotide substitutions result in three amino acid substitutions within a span of five amino acids; isoleucine at position 67, arginine at position 70, and threonine at position 71 are being substituted by phenylalanine, glutamine, and lysine respectively. Two of these amino acid substitutions (arginine-glutamine and threonine-lysine) are nonconservative changes in the tertiary structure of the beta polypeptide chain resulting in the observed alterations in the serology and function of the bml2 A molecule. Amino acid sequence comparison also shows that two of the amino acid substitutions found in the bml2 A;~ chain are found at the same positions in several human class II beta chains (McIntyre and Seidman 1984). These comparisons suggest that the bml2 mutation in the A beta gene arose from a gene-conversion-type event in which another class II beta chain gene acted as a donor sequence. Comparison of the bml2 nucleotide substitutions to the sequence of a cosmid clone for the E~ gene has suggested that the E~ served as this donor sequence (Widera and Flavell 1984), where a minimum of 14 nucleotides of the A;~ gene are

Structural Characterization of I-Ab Subsets Using Monoclonal Antibodies

The murine Ia antigens are cell surface glycoproteins composed of two noncovalently linked polypeptide chains with apparent molecular weights of 35,000 (α) and 29,000 (β) respectively (1). Recently, a third chain, an invariant chain of 31,000 MW, has been detected by Jones et al. (2). The expression of Ia antigens are controlled by two distinct loci, called I-A and I-E, that code for distinct subsets of Ia molecules. Previous reports from our laboratory have suggested the existence of more than one Ia molecule encoded by I-A and I-E loci in H-2b and H-2k haplotypes (3,4).