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Biomedical subjects

R C Williams

Publications and source records attributed to R C Williams.

At least 217 records · Page 12Linked to original sources
PubMed1987

Monoclonal murine anti-DNA antibody interacts with living mononuclear cells.

A monoclonal mouse antibody, 4-B-5, that reacted with both single-stranded DNA (ssDNA) and double-stranded DNA showed direct reactivity and cell association with mouse thymocytes, as well as peripheral blood mononuclear cells. Absorption of 4-B-5 with peripheral blood mononuclear cells or with mouse thymocytes markedly reduced reactivity with both ssDNA and double-stranded DNA. However, treatment of mouse thymocytes or human T cells with DNase completely eliminated monoclonal anti-DNA reactivity with cells. The cellular reactivity was completely restored when monoclonal anti-DNA was incubated with DNase-treated cells in the presence of normal human serum. Normal human serum was found to contain 574 +/- 639 ng/ml of ssDNA. Coincubation or cell preincubation of mouse thymocytes with ssDNA produced a marked increase in 4-B-5 cell association. These findings indicate that mouse monoclonal anti-DNA can react with and can penetrate both mouse and human mononuclear cells, and that this reactivity may depend on the presence of cell membrane DNA on both types of target cells.

Animals↗
PubMed1987

A family with hereditary angioedema and multiple immunologic disorders.

A 58-year-old man with end-stage renal disease treated with long-term ambulatory peritoneal dialysis gave a history of hereditary angioedema. Results of testing of his serum for antinuclear antibody were strongly positive, and a titer of 1:2,560 for antiribosomal antibody was documented. Three of his six children reported multiple medical problems, including episodic swelling, discoid lupus, and arthritis. An indepth study of this kindred showed a number of associated immunologic disturbances. Three children had decreased C1 esterase inhibitor and decreased C4 levels. T cell enumeration revealed low proportions of suppressor cells in these three children as well as in another unaffected child. Serum IgM level was low in the propositus, and IgG level was also decreased in one of the affected children. HLA typing failed to show a clear association of class I or class II antigens with hereditary angioedema. Family members tested did not demonstrate anti-SS-A, anti-SS-B, antinuclear antibody, anti-double-stranded DNA, anti-Sm, or anti-ribonuclear protein antibodies.

Angioedema↗
PubMed1987

Evidence for the spontaneous formation of disulfide crosslinked aggregates of tubulin during nondenaturing electrophoresis.

Phosphocellulose-purified tubulin has been shown to form a characteristic "ladder" of nonmicrotubular aggregates during nondenaturing gel electrophoresis (J. J. Correia and R. C. Williams, Jr. (1985) Arch. Biochem. Biophys. 239, 120-129). In this paper we describe evidence that the intersubunit bonds responsible for formation of these oligomeric particles are disulfides. Two-dimensional nondenaturing-denaturing gel electrophoresis demonstrates that each aggregate zone is composed of alpha- and beta-subunits of tubulin. Omission of beta-mercaptoethanol during the sodium dodecyl sulfate (SDS)-electrophoresis step causes a pattern of aggregates to appear and implicates disulfide linkages in their stabilization. Molecular weights, estimated from mobilities in the second (SDS) dimension of two-dimensional gels, suggest that the aggregates are crosslinked in units of monomers, not heterodimers. Consistent with this conclusion, alpha- or beta-subunits alone (isolated by isoelectric focusing) will form the same ladder of aggregates. The disulfide crosslinking of tubulin is also achievable in solution. It is favored by high concentrations of alcohol, the presence of oxidizing agents, high pH, and high temperature, conditions that denature tubulin and cause rapid noncovalent aggregation or precipitation. When aggregate formation was monitored as a function of time by SDS-gel electrophoresis in the absence of beta-mercaptoethanol and by quantitative sulfhydryl and disulfide titrations, the most effective conditions for the crosslinking reaction included greater than 75% alcohol, excess H2O2, or excess iodine. These results suggest that proximity of a hydrophobic gel matrix, high pH, the presence of oxidizing agents, high protein concentration, tubulin's propensity to aggregate nonspecifically, and the availability of as many as 20 sulfhydryls in alpha beta-tubulin contribute, during nondenaturing gel electrophoresis, to the spontaneous formation of disulfide-crosslinked tubulin aggregates.

Chromatography, Ion Exchange↗
PubMed1987

Effects of immunoglobulin G from patients with systemic lupus erythematosus on human B cell function.

We examined the effect of systemic lupus erythematous (SLE) sera and Ig fractions on IgG and IgM release by cultured normal peripheral blood mononuclear cells (PBMC) when these cells were preincubated with serum dilutions or Ig fractions. Increases in both IgM and IgG (P less than 0.001 and less than 0.01) in cultured cell supernatants were recorded when PBMC were preincubated with SLE serum dilutions. IgG but not IgM from SLE was found to stimulate PBMC to release IgG (P less than 0.01). Similar results were obtained when SLE IgG was preincubated with adherent cell depleted cells (ADC) or isolated normal B cell fractions. When normal PBMC were preincubated with SLE serum or IgG and subsequently stimulated with pokeweed mitogen (PWM), a relatively blunted IgG release was observed (P less than 0.05); however, IgM release was significantly increased (P less than 0.001). This effect was not observed when PBMC were preincubated with SLE IgM, normal serum dilutions, or normal Ig fractions. Relative blunting of PWM response after PBMC were preincubated with SLE IgG was not reversed in PBMC depleted of adherent cells, OKT8+, or OKT9+ cells. Depletion of PBMC of LeuM1 cells increased IgG release in response to PWM when cells had been preincubated with SLE IgG. SLE serum or Ig fractions did not induce B cell growth factor release by T cells. SLE IgG appeared to act directly on B cell enriched populations to release IgG; this was not associated with significant increase in thymidine uptake, or apparent lysis of cells.

Antibodies, Monoclonal↗
PubMed1987

In vitro inhibition of anti-DNA producing cells from systemic lupus erythematosus patients by autologous anti-F(ab')2 antibodies.

We have previously documented an inverse relationship between serum levels of anti-F(ab')2 antibodies and disease activity in systemic lupus erythematosus (SLE). The present study focused on the specific in vitro inhibition of anti-DNA producing cells from SLE patients by autologous anti-F(ab')2 antibodies. Peripheral blood lymphocytes (PBL) from eleven inactive SLE patients with no apparent disease activity were cultured in vitro to evaluate anti-DNA antibody secretion. Low levels of synthesis of anti-DNA antibody were detected in 3 of 11 SLE patients using unstimulated PBL; on the contrary, pokeweed mitogen stimulation of cultured cells increased production of anti-DNA in all SLE subjects. Parallel cultures were also performed in the presence of heterologous and autologous anti-F(ab')2 antibodies and results on production of anti-DNA evaluated. Lymphocytes from SLE patients in remission showed inhibition of synthesis of anti-DNA antibodies when autologous anti-F(ab')2 antibodies were added to the cultures, whereas production of anti-tetanus toxoid IgG by the same cells was not significantly altered under the same conditions. These data suggest that a functional anti-idiotypic role may be assigned to anti-F(ab')2 antibodies during clinical remission of SLE.

Antibodies, Anti-Idiotypic↗
PubMed1987

Tissue c-myc protein expression and immune response in systemic lupus erythematosus.

Frozen sections of kidney tissue from 12 patients with systemic lupus erythematosus were examined for the expression of oncogene proteins. Polyclonal rabbit antibody to c-myc peptide or to whole 65,000 D c-myc protein, was used to identify c-myc protein within tissue sections. Patterns of nuclear staining in the Hep-2 cell line were similar to those seen with monospecific human SLE sera showing anti-Sm reactivity. In addition c-myc staining was not abolished by prior incubation of tissue sections with human serum containing anti-Sm antibodies. Five of 12 kidney biopsy tissues from patients with systemic lupus erythematosus (SLE) showed positive speckled c-myc protein staining within nuclei of monocyte/macrophage cells of glomerular tufts. Positive staining was in all instances completely abolished by prior absorption of anti-c-myc antibody with c-myc protein. No c-myc protein was identified within SLE immune complex deposits.

Adult↗
PubMed1986

Separation of proteins on a polymeric fluorocarbon high-performance liquid chromatography column packing.

A rigid, inert fluorocarbon resin has been evaluated as an high-performance liquid chromatography packing for the separation of proteins and peptides. Samples are separated with gradient elution of the mobile phase from aqueous buffer to acetonitrile. The selectivity and retention of the fluorocarbon resin packing for proteins are similar to that of aliphatic hydrocarbon-bonded silica packings.

Animals↗
PubMed1986

Fibers of RecA protein and complexes of RecA protein and single-stranded phi X174 DNA as visualized by negative-stain electron microscopy.

Monomers of purified RecA protein polymerize into helical fibers whose pitch is 7.2 nm to 7.5 nm and whose diameter is 11 nm. Either short (approximately 0.2 micron), single fibers, or bundles of aligned, longer fibers, can be formed preferentially, by varying the Mg2+ concentration. When RecA protein is bound to circular, single-stranded phi X174 DNA it forms helical fibers of different classes of contour lengths, ranging from 0.98 micron, depending upon the conditions of assembly. Two different helical pitches are found, one of 9.3 nm when the incubation buffer contains, besides the obligatory Mg2+, either ATP gamma S or ATP accompanied by single-strand binding protein, and one of 5.5 nm when the latter additives are omitted. Preformed fibers of the compact type can be converted to open ones of 9.3 nm pitch upon addition of ATP gamma S, even after the removal of unbound RecA. All signs of helicity are obliterated upon glutaraldehyde cross-linking except in those fibers whose assembly has been mediated by ATP gamma S. RecA protein and single-strand binding protein are competitively bound to single-stranded DNA. Composite complexes, however, are not encountered unless ATP gamma S is present. Otherwise, segments of DNA that are coated by one or the other protein are seen as separate regions. When the assembly of complexes of single-stranded DNA and RecA is mediated by single-strand binding protein and ATP, the axial separation between successive bases is 0 X 42 nm, somewhat greater than the axial distance between bases in one strand of duplex DNA in the B form. It is proposed that the bases of the single-stranded DNA in the complex are located near its inner surface, and that base-pairing with double-stranded DNA takes place following invasion of the central cavity of the complex.

Adenosine Triphosphate↗
PubMed1986

Purification and characterization of actophorin, a new 15,000-dalton actin-binding protein from Acanthamoeba castellanii.

Actophorin is a new actin-binding protein from Acanthamoeba castellanii that consists of a single polypeptide with a molecular weight of 15,000. The isoelectric point is 6.1, and amino acid analysis shows an excess of acidic residues over basic residues. The phosphate content is less than 0.2 mol/mol. There is 0.4 +/- 0.1 mg of actophorin/g of cells, so that the molar ratio of actin to actophorin is about 10:1 in the cell. Unique two-dimensional maps of tryptic and chymotryptic peptides and complete absence of antibody cross-reactivity show that Acanthamoeba actophorin, profilin, capping protein, and actin are separate gene products with minimal homology. Actophorin has features of both an actin monomer-binding protein and an actin filament-severing protein. Actophorin reduces the extent of actin polymerization at steady state in a concentration-dependent fashion and forms a complex with pyrene-labeled actin that has spectral properties of unpolymerized actin. During ultracentrifugation a complex of actophorin and actin sediments more rapidly than either actin monomers or actophorin. Although actophorin inhibits elongation at both ends of actin filaments, it accelerates the late stage of spontaneous polymerization like mechanical shearing and theoretical predictions of polymer fragmentation. Low concentrations of actophorin decrease the length and the low shear viscosity of actin filaments. High concentrations cause preformed filaments to shorten rapidly. Ca2+ is not required for any of these effects. Muscle and amoeba actin are equally sensitive to actophorin.

Actin Depolymerizing Factors↗
PubMed1986

Immunofluorescence studies of florid rheumatic Aschoff lesions.

We studied cardiac tissues of a patient who died of severe rheumatic myocarditis. Multiple Aschoff lesions were present throughout both ventricles and auricles. Immunofluorescence studies showed large monocytoid cells staining with OKM1 and anti-Leu M-3 as well as anti-Ia. Scattered T cells in areas of focal myocarditis stained with OKT3. Parallel staining for cardiac myosin-heavy chain antigens showed patchy dissolution of cardiac muscle fibers and traces of cardiac myosin within large monocytoid Aschoff cells.

Adult↗
PubMed1986

Immunoglobulin from systemic lupus erythematosus serum induces interferon release by normal mononuclear cells.

Ig fractions from patients with systemic lupus erythematosus (SLE) were tested with cultured normal peripheral blood mononuclear cells for induction of interferon release. Lymphocyte eluates, euglobulins containing IgG and IgM, and IgG or IgM from DEAE or sucrose gradients all induced interferon production. Lymphocytotoxic antibody in SLE sera showed a high correlation with capacity of isolated Ig fractions to induce interferon. Most interferon produced was of the gamma type. Monoclonal SLE IgM antilymphocyte antibody induced interferon synthesis.

Antibodies, Monoclonal↗
PubMed1986

Characterization of alpha-actinin from Acanthamoeba.

Characterization of a protein from Acanthamoeba that was originally called gelation protein [T.D. Pollard, J. Biol. Chem. 256:7666-7670, 1981] has shown that it resembles the actin filament cross-linking protein, alpha-actinin, found in other cells. It comprises about 1.5% of the total amoeba protein and can be purified by chromatography with a yield of 13%. The native protein has a molecular weight of 180,000 and consists of two polypeptides of 90,000 Da. The Stokes' radius is 8.5 nm, the intrinsic viscosity is 0.35 dl/dm, and the extinction coefficient at 280 mm is 1.8 X 10(5)M-1 X cm-1. Electron micrographs of shadowed specimens show that the molecule is a rod 48 nm long and 7 nm wide with globular domains at both ends and in the middle of the shaft. On gel electrophoresis in sodium dodecylsulfate the pure protein can run as bands with apparent molecular weights of 60,000, 90,000, 95,000, or 134,000 depending on the method of sample preparation. Rabbit antibodies to electrophoretically purified Acanthamoeba alpha-actinin polypeptides react with all of these electrophoretic variants in samples of purified protein and cell extracts. By indirect fluorescent antibody staining of fixed amoebas, alpha-actinin is distributed throughout the cytoplasmic matrix and concentrated in the hyaline cytoplasm of the cortex. The protein cross-links actin filaments in the presence and absence of Ca++. It inhibits slightly the time course of the spontaneous polymerization of actin monomers but has no effect on the critical concentration for actin polymerization even though it increases the apparent rate of elongation to a small extent. Like some other cross-linking proteins, amoeba alpha-actinin inhibits the actin-activated ATPase of muscle myosin subfragment-1. Although Acanthamoeba alpha-actinin resembles the alpha-actinin from other cells in shape and ability to cross-link actin filaments, antibodies to amoeba and smooth muscle alpha-actinins do not cross react and there are substantial differences in the amino acid compositions and molecular dimensions.

Actinin↗
PubMed1986

Altered function of synovial fluid granulocytes in patients with acute inflammatory arthritis: evidence for activation of neutrophils and its mediation by a factor present in synovial fluid.

In rheumatoid arthritis (RA) a chronic inflammatory state exists in which the synovial fluid is periodically filled with large numbers of polymorphonuclear leukocytes (PMNs). Oxygen radicals produced by these cells have been implicated as mediators of tissue damage and may be directly involved in the pathogenesis of RA. We examined the production of oxygen radicals by synovial fluid PMNs (SF-PMNs) and peripheral blood PMNs (PB-PMNs) by measuring chemiluminescence (CL) as well as superoxide anion (O2-) release. Increased spontaneous CL in the presence of luminol and increased CL in response to phorbol myristate acetate (PMA) was observed in SF-PMNs when compared to PB-PMNs. When zymosan was used as the stimulus in the absence of luminol, a slightly lower CL response was observed in SF-PMNs as compared to PB-PMNs. No significant differences were observed in the generation of O2- generation with any stimulus. Preincubation of normal PB-PMNs in 10% synovial fluid enhanced the luminol-dependent spontaneous and PMA-stimulated CL as well as zymosan-stimulated CL. When O2- release from normal PB-PMNs pretreated with 10% synovial fluid was compared to untreated controls, enhancement of spontaneous O2- release was observed. PMA- and zymosan-stimulated responses did not differ significantly from controls. Increased spontaneous and PMA-stimulated release of myeloperoxidase (MPO) was also observed in normal PB-PMNs pretreated with synovial fluid. These findings may explain the increased luminol-dependent CL since this type of CL requires the presence of MPO. Our findings suggest that the enhanced chemiluminescence observed in normal PMNs treated with synovial fluids may be related to increases in spontaneous O2- generation and myeloperoxidase release. Increased MPO release may account for enhanced CL observed in SF-PMNs.

Acute Disease↗