Thomas M. Fasy

Structure and binding properties of monoclonal antibodies to core histones from autoimmune mice.

Histones are frequent targets of self-reactive antibodies during autoimmune syndromes. We report the specificities and V region genes of three IgG anti-histone MAbs obtained from autoimmune mice. Each of the MAbs, named LG2-1, LG2-2 and BWA3, is directed against a different determinant located in the basic amino-terminal domain of core histones. LG2-1 reacts with a peptide from histone H3 (residues 30-45), LG2-2 recognizes the amino-terminus of H2B (residues 1-13) and BWA3 binds an epitope corresponding to a region of high sequence similarity between H2A and H4 (residues 1-20 and 1-29, respectively). The analysis of their V region sequences indicates that the H chain CDRs of these MAbs are remarkable for the presence of negatively charged amino acid residues that may play a role in the binding to cationic histones. The H chain importance in conferring reactivity to histones is corroborated by the observation that each of the VH gene segments of these MAbs is very similar to VH genes of previously described murine anti-histone antibodies.

Lymphomatoid papulosis with antigen deletion and clonal rearrangement in a 4-year-old boy

Lymphomatoid papulosis (LyP) is rarely seen in children. We report a case of LyP in a 4‐year‐old boy in whom immunopathologic studies demonstrated T cell antigen deletions. In contrast to all but two previous reports, a T suppressor (CD‐8) phenotype was predominant. Southern blot analysis of DNA isolated from a typical skin lesion indicated a clonal rearrangement of the T cell receptor β gene. Because of a 10% frequency of malignant lymphomas in patients with LyP, long‐term observation is cruciai, especially in chiidren. We recommend routine clonal rearrangement studies for aid in diagnosis and follow‐up, and as possible prognostic indicators in children with this condition.

Chrysotile asbestos fibers mediate homologous recombination in Rat2λ fibroblasts: Implications for carcinogenesis

Asbestos fibers are widespread environmental carcinogens whose mutagenicity is now established. Nonetheless, the molecular nature of these mutations and the mechanisms by which they accelerate carcinogenesis remain poorly understood. We have assessed the ability of asbestos fibers to promote homologous recombination, a potent mechanism for generating intrachromosomal rearrangements, such as deletions, and mitotic recombination. For this, we have developed a new assay which determines the extent to which a marker gene present in DNA introduced by asbestos can recombine with homologous genes residing in a transfected cell. We have demonstrated that Calidria chrysotile fibers are mutagenic and are able to mediate transfection of molecularly marked mutant lacI genes in a manner that results in their preferential recombination with homologous wild-type genes in the transfected cell. Asbestos induced recombination events may play a significant role in asbestos mutagenesis and carcinogenesis, and promotion of recombination may underlie the well-recognized synergy of asbestos with other carcinogens.

ras and c-myc protein expression in colorectal carcinoma

This study was performed to determine the correlation of tumorrasand c-myconcogene expression with clinical and prognostic variables in patients prone to develop colorectal cancer. One hundred eighteen patients with colorectal cancer were studied; mean age was 40 years. Fifty-three were young patients (age 40 or less), 49 had ulcerative colitis, and 16 had multiple polyposis coli. Immunoperoxidase stains of paraffin-embedded cancer sections were performed for the c-mycandrasproteins.rasstaining was found to correlate with Dukes stage and prognosis. Patients with tumors negative forrasprotein stain had an actuarial five-year survival of 61 percent versus 44 percent for those tumors with a positive stain (P<0.05). This correlation was not seen with the c-mycstain. Positiverasoncogene stain appears to be a useful indicator of advanced stage and poor prognosis in colorectal cancer occurring in cancer-prone patients.

Deoxyribonuclease I treatment of histones for the detection of anti-histone antibodies in solid-phase immunoassays : effect of protease contamination in commercial deoxyribonuclease I preparations

In this report, we tested two commercial preparations of DNase I for their effect on histone H1 and its recognition by a monoclonal anti-H1 antibody. Our results indicate that digestion with the DNase I preparation of a lesser grade results in the complete proteolysis and loss of antigenicity of histone H1 even though this preparation is adversited as containing less than 0.01% chymotrypsin and less than 0.005% protease

Effects of H1 histones and a monoclonal autoantibody to H1 histones on clot formation In vitro: Possible implications in the antiphospholipid syndrome

Histones are known to bind anionic phospholipids (PLs). Binding of procoagulant PLs by histones released during cell injury/death may interfere with coagulation and may serve a local regulatory anticoagulant function. Histone H1 prolonged the PT and APTT of normal pooled plasma (NPP). These increased clotting times disappeared when anti-H1 monoclonal antibody (mAb) was added to the incubation. Dilute Russell Viper Venom Time was also prolonged with the addition of histone H1. When H1 was added to plasma from a patient with the antiphospholipid syndrome (APL plasma), there was a further prolongation of the abnormal APL clotting time which was partially corrected by anti-H1 mAb. Platelet neutralization times were increased with added H1 and were further increased using APL plasma. when disrupted endothelial cells were incubated with plasma with and without anti-H1 antibodies, the addition of anti-H1 antibodies decreased clotting times. These data support the theory that histones released during cell injury may have a regulatory anticoagulant role in clot formation and the anti-H1 effect of some APL plasmas may inhibit this, thereby contributing to thrombosis seen in APL patients.

Anti-histone antibodies in subacute sensory neuropathy

SummaryWe investigated the levels of anti-histone antibodies in the sera of 7 patients with subacute sense neuropathy. IgG antibodies to histones HI and H3 were significantly elevated in 4 of these patients. The ai H1 antibodies reacted mainly with determinants located in the central globular and the carboxy-termi domain of the H1 molecule. We also observed reactivity of these sera with histone H1°, a variant founc terminally-differentiated cells such as neurons. This study suggests a potential for histones to serve autoantigens in humorally-mediated paraneoplastic diseases.

Histone-Reactive IgA Antibodies in Adult IgA Nephropathy and Other Primary Glomerulonephritis

The levels of histone-reactive IgA antibodies in the sera of adult patients with IgA mesangial glomerulonephritis, membranous glomerulonephritis, membranoproliferative glomerulonephritis and idiopathic nephrotic syndrome (minimal change disease+segmental glomerulosclerosis+IgM nephropathy) were evaluated by an enzyme-linked immunosorbent assay. Increased levels of IgA antibodies to all five major histones (H1, H2A, H2B, H3, H4) were found in all four disease groups when compared to normal controls. These histone-reactive IgA antibodies were restricted to the IgA1 subclass and their levels did not correlate with the levels of total serum IgA, nor with serum creatinine, creatinine clearance, and 24-hour proteinuria. Increasing ionic strength resulted in only partial inhibition of the binding to histones and, in individual patients, levels of reactivity with individual histones were usually correlated. This study shows that elevated levels of IgA antibodies reactive with self antigens are present in primary glomerulonephritis and extends previous observations indicating that anomalies of the IgA system occur in various forms of primary glomerulonephritis and are not limited to IgA nephropathy.

DNA ploidy of colorectal cancer and synchronous polyps in polyposis coli

Sixteen patients with polyposis coli and cancer were studied retrospectively to determine the incidence of DNA ploidy abnormalities in the tumors and synchronous polyps. Six patients (37 percent) had nondiploid tumors. Nondiploid tumors were more likely to be advanced and had a significantly worse prognosis (17 percentvs.76 percent 5-year survival;P<0.01). Only 4 of 20 polyps studied were nondiploid. There was no association between tumor and polyp ploidy. All nondiploid polyps were found in patients with synchronous diploid cancers. Patients with nondiploid polyps were more likely to be older and have more advanced tumors than those with diploid polyps. DNA ploidy abnormalities seem to occur with the same frequency in polyposis coli as in the nonpolyposis population, and tumor ploidy correlates with prognosis