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Biomedical subjects

E M Tan

Publications and source records attributed to E M Tan.

At least 217 records · Page 12Linked to original sources
PubMed1984

Antibodies from patients with mixed connective tissue disease react with heterogeneous nuclear ribonucleoprotein or ribonucleic acid (hnRNP/RNA) of the nuclear matrix.

The sera of patients with mixed connective tissue disease (MCTD) have high titers of antibodies directed against nuclear U1-ribonucleoprotein (U1-RNP). This antigen is easily extracted from nuclear preparations with physiologic saline and from tissue sections with 0.1 HCl, leaving the nucleic acids and nuclear matrix behind. When U1-RNP is extracted from HEp-2 cells with 0.1 N HCl, the sera of 32/32 patients with MCTD react with another antigen that is exposed by the extraction procedure. This antigen is not destroyed by trypsin and deoxyribonuclease 1 treatment but is sensitive to both purified ribonuclease A and purified micrococcal nuclease. Absorption studies showed that the antibody reacting with this antigen cannot be absorbed by sheep red blood cells coated with extracts of rabbit thymus that contain U1-RNP. Radioimmunoassay showed that the reaction of the unadsorbed antibody was with heterogeneous nuclear ribonucleoprotein or ribonucleic acid (hnRNP/RNA) and not with transfer RNA or ribosomal RNA. The hnRNP/RNA antigen is demonstrated as discrete particles in the internucleolar chromatin of interphase cells, but in metaphase cells the antigen is diffusely dispersed. The distribution, solubility, and biochemical characteristics suggest that the antigenic moiety is part of the nuclear matrix. Therefore, MCTD sera contain antibodies that react with at least two species of nuclear RNP: small nuclear RNP (snRNP), as described by others, and a high m.w. hnRNP/RNA bound to the nuclear matrix.

Animals↗
PubMed1984

Monoclonal anti-histone autoantibodies derived from murine models of lupus.

We characterized three monoclonal antibodies with histone reactivity which were derived from spleen cells obtained from unmanipulated NZB/NZW or MRL/1 mice. By using an enzyme-linked immunosorbent assay, we noted that all three antibodies reacted with chromatin histones as well as with total histones extracted from chromatin. None of the antibodies appeared to require DNA as part of the antigen. One antibody (BWH-1) recognized a determinant present in the nucleosome core H2A-H2B complex but showed little reactivity with any of the individual histones (H1, H2A, H2B, H3, or H4). In contrast, the other two monoclonal antibodies each recognized multiple individual histones in a unique pattern. Antibody MH-1 reacted with H2A, H2B, and H3; antibody MH-2 reacted with H2A, H3, and H4. MH-1 demonstrated cross-reactivity with poly-1-lysine but not poly-1-arginine or protamine sulfate; the opposite pattern of cross-reactivity was observed with MH-2. The antigenic determinants recognized by MH-2 were all trypsin-sensitive, suggesting that these determinants were present on the N-terminal regions of the respective individual histones. These studies revealed markedly different specificities of anti-histone monoclonal antibodies derived from murine models of systemic lupus erythematosus. These and other similarly derived antibodies may provide interesting tools to understand the specificity and biologic importance of anti-histone autoantibodies in different diseases.

Animals↗
PubMed1984

Anti-native DNA antibodies from autoimmune sera also bind to DNA in mitochondria.

Sera from 20 patients with systemic lupus erythematosus (SLE), selected for elevated titers of antibody to native DNA (nDNA), were examined by indirect immunofluorescence (IF) on tissue culture Hep-2 and rabbit kidney cells. Twelve sera showed a particulate cytoplasmic staining, in addition to nuclear IF. Double IF staining by using a mouse monoclonal anti-nDNA and a human serum containing anti-mitochondrial antibody as probes showed that the cytoplasmic structures recognized by these 12 SLE sera were mitochondria. SLE sera showing mitochondrial staining had high anti-nDNA levels, as assessed by ELISA (3.5 +/- 1.9 O.D.), compared with those not showing this staining pattern (0.8 +/- 0.4 O.D.). Mitochondrial staining was abolished by DNase I pretreatment of the substrates. Liquid phase absorption of serum anti-nDNA with S1 nuclease-treated calf thymus DNA or purified mitochondrial DNA also removed staining. These findings demonstrate that anti-nDNA antibodies from patients with SLE bind to DNA in intact mitochondria. Therefore, mitochondrial IF staining on tissue culture cells in the presence of nuclear staining should be interpreted with caution, because the phenomenon could be entirely related to anti-native DNA. These observations might also provide new insights concerning the nature of immunogenic cellular components stimulating anti-DNA production.

Animals↗
PubMed1984

Pharmacological inhibition of excessive collagen deposition in fibrotic diseases.

Excessive accumulation of collagen is a major pathological feature in diseases characterized by tissue fibrosis. Although several therapeutic approaches have been attempted in such patients, currently no treatment modality would specifically reduce collagen deposition in tissues. In this paper we discuss the mode of action of compounds that interfere with collagen production on the posttranslational level. First, structural analogs of proline, cis-4-hydroxy-L-proline and L-azetidine-2-carboxylic acid, which are incorporated into the newly synthesized polypeptides of procollagen during translation, prevent the polypeptides from folding into a stable triple-helical conformation. As a consequence, the nonhelical polypeptides are subject to degradation by proteases, thus leading to reduced deposition of collagen fibers. Second, several naturally occurring amino acids, polyamines, and their structural analogs prevent the removal of the carboxy-terminal extensions during the extracellular conversion of procollagen to collagen. Because the precursor molecules that contain the carboxy-terminal extensions are unable to assemble into functional fibers, collagen deposition is again reduced. Further development of these and related compounds, with appropriate tissue targeting, could potentially provide us with novel means to reduce the excessive deposition of collagen in fibrotic processes.

Animals↗
PubMed1983

An improved ELISA for anti-native DNA by elimination of interference by anti-histone antibodies.

We evaluated an enzyme-linked immunosorbent assay (ELISA) for antibodies to native DNA (nDNA) in which protamine was used to link DNA to polystyrene. Elevated anti-nDNA was largely restricted to patients with systemic lupus erythematosus (SLE), and within this group good correlation between ELISA and the ammonium sulfate assay was obtained. However, substantial background immunoglobulin binding to protamine coated wells was commonly observed, and it was necessary to subtract this activity from each anti-DNA determination. Many of the SLE sera also contained anti-histone antibodies, and this antibody activity showed significant correlation with the binding to protamine. In contrast, methylated bovine serum albumin (mBSA) did not bind anti-histone antibodies and provided a substrate for coupling nDNA to polystyrene. This modified ELISA allowed the quantitation of antibodies to native DNA without the simultaneous binding of anti-histone antibodies.

Ammonium Sulfate↗
PubMed1983

Detection of a nucleolar 7-2 ribonucleoprotein and a cytoplasmic 8-2 ribonucleoprotein with autoantibodies from patients with scleroderma.

In studies on antinucleolar antibodies in sera from 24 patients with scleroderma, an autoimmune disease, one serum, designated "anti-To", contained antibodies against a nucleolar 7-2 ribonucleoprotein and a novel cytoplasmic 8-2 ribonucleoprotein. The 7-2 and 8-2 RNAs are distinct RNAs with a pppG terminus. They are partially conserved between rat and human species and are present in distinct ribonucleoprotein particles. Eight sera contained antibodies that precipitated particles containing nucleolar U3 RNA; these antibodies appear to be directed against preribosomal particles containing U3 ribonucleoprotein, rather than the U3 ribonucleoprotein particles alone. All these ribonucleoproteins required proteins for antigenicity. These antibodies will be of use in studies on the structure and function of these novel small ribonucleoproteins.

Animals↗
PubMed1983

Autoantibodies to nonhistone nuclear antigens and their clinical significance.

Identification of the immunologic specificity of antinuclear antibodies (ANAs) in the various systemic rheumatic diseases has become increasingly important. The standard immunofluorescence technique may enable detection of antibodies to nuclear antigens present in abundance in the nucleus, such as DNA, histones, Sm, nuclear ribonucleoprotein (nRNP), and SS-B/La. The nuclear antigens present in low concentrations, such as SS-A, proliferating cell nuclear antigen (PCNA), rheumatoid arthritis nuclear antigen (RANA), and Ku antigens, are unique to cell types, and their detection requires special substrates or reagent systems. Anti-Sm, anti-Scl-70, anticentromere, and anti-PM-1 are characteristic serologic markers for systemic lupus erythematosus, scleroderma, the CREST syndrome of scleroderma, and polymyositis, respectively. Distinct profiles of ANA characterize different rheumatic diseases. A number of ANAs are found in SLE, whereas other diseases are characterized by the presence or absence of a certain ANA or by differences in mean ANA titers. Specific ANAs have been used to isolate and characterize nuclear antigens at molecular and functional levels.

Antibodies, Antinuclear↗
PubMed1983

Antinuclear antibodies in diagnosis and management.

Antinuclear antibodies are known to be present in a variety of autoimmune diseases, and they appear to play a primary role in the pathogenesis of a subgroup of "ANA diseases." It has been shown that each presents a distinct ANA profile, characterized by the presence and titer of certain antibodies and the absence of others. These profiles are of practical value in diagnosis.

Adolescent↗
PubMed1983

Proline analogues inhibit human skin fibroblast growth and collagen production in culture.

Several structural analogues of proline have been shown to be incorporated into proteins in place of proline. As a consequence, the proliferation of cells in culture and the extracellular deposition of collagen in animal systems are reduced. In this study, the effects of two proline analogues, cis-4-hydroxy-L-proline and L-azetidine-2-carboxylic acid, on the growth parameters and procollagen production by cultured normal human skin fibroblasts were examined. The results indicated that incubation of the cells with the analogues reduced the rate of fibroblast proliferation and lowered the plating efficiency. Further experiments demonstrated that fibroblasts in the presence of L-azetidine-2-carboxylic acid synthesized procollagen polypeptides which were not in a triple-helical conformation, as judged by limited pepsin proteolysis. Also, a significantly increased fraction of the newly synthesized collagenous peptides was in a dialyzable form, suggesting increased degradation of the nonhelical chains. The rate of translation of collagenous polypeptides and the preprocollagen messenger RNA activity in the cells were not affected by the analogues. The proline analogues thus appear to inhibit the production of procollagen on the posttranslational level by preventing the polypeptides from folding into a stable triple-helical conformation. The nonhelical polypeptides are then readily susceptible to proteolysis leading to reduced deposition of extracellular collagen fibers. Similar experiments were also performed with fibroblasts cultured from patients with active progressive systemic sclerosis. Quantitatively and qualitatively comparable inhibition of procollagen production by L-azetidine-2-carboxylic acid was noted with scleroderma cells as with control fibroblast cultures. The results suggest, therefore, that proline analogues may, in the future, prove useful in limiting excessive collagen deposition in scleroderma and other forms of dermal fibrosis.

Azetidinecarboxylic Acid↗
PubMed1983

Characterization of a phosphoprotein associated with the SS-B/La nuclear antigen in adenovirus-infected and uninfected KB cells.

We have employed sera from patients with autoimmune disease to characterize the nuclear SS-B/La antigen in uninfected and adenovirus-infected KB cells. A 45,000-dalton phosphorylated polypeptide was specifically precipitated with anti-SS-B sera, and the level of phosphorylation was increased after infection. The increased phosphorylation appears to occur at the same amino acid residues phosphorylated in uninfected cells and results from increased phosphorylating activity rather than from altered enzyme specificity. A competition experiment between infected and uninfected cell extracts indicates that the antigen in the infected cell binds more strongly to SS-B/La antibodies. Fragments of adenovirus-induced virus-associated RNA as well as intact molecules complex with SS-B/La antigen and are immune precipitated with autoimmune sera.

Adenoviridae↗
PubMed1983

Frequency of transforming Epstein-Barr virus in oropharyngeal secretions of rheumatoid arthritis patients.

To determine if rheumatoid arthritis (RA) patients have an increased frequency of Epstein-Barr virus (EBV) shedding into saliva, throat washings were collected from 59 patients with RA and 64 healthy adult controls. EBV in filtered samples was detected by transformation of human umbilical cord lymphocytes. EBV was detected in 27% (16/59) of throat washings of RA patients compared to 11% (7/64) of control samples. This difference is significant (p less than 0.05). RA patients on steroids had a frequency of positivity of 43% (10/23) compared to 17% (6/36) in patients not on steroids. No correlation was demonstrated between steroid dose received by the patient and detection of EBV in throat washings. Parotid fluid samples were collected from 18 RA patients and 11 healthy controls and were tested for EBV by the transformation bioassay. EBV was detected in 18% (2/11) of samples from normal controls, but samples from RA patients were uniformly negative.

Adrenal Cortex Hormones↗
PubMed1983

Indirect immunofluorescence studies of proliferating cell nuclear antigen in nucleoli of human tumor and normal tissues.

A proliferating cell nuclear antigen (PCNA) was identified with autoantibodies from a patient with systemic lupus erythematosus. Specific antibodies were purified by affinity chromatography in which Novikoff hepatoma nucleolar proteins were conjugated to Sepharose-4B. The purified anti-PCNA antibodies produced bright nucleolar fluorescence in tumor cells as shown by indirect immunofluorescence. PCNA was found in nucleoli of human cell lines, HeLa, Hep-2, and Namalwa, and a solid human renal and a prostate carcinoma. Both strong and weak nucleolar fluorescence areas were found in the renal and prostate carcinoma indicating that there are varying degrees of proliferation among tumor cells. Two human colon carcinoma cell lines, omega (an aggressive, fast-growing clone of human colon carcinoma cell line HCT 116) and CBS [a slow-growing human colon carcinoma cell line (group 3)], with different growth rates were compared. The fast-growing colon carcinoma cells, omega, exhibited a higher percentage of nucleolar fluorescence (28.5%) than that of the slow growing colon cells (13.6%). By enzyme-linked immunosorbent assays, the omega cell extract had a higher PCNA antigen content (2.8-fold) than that of the CBS cell extract which, in turn, was higher than that of human liver extract. PCNA was also found in a human fetal lung fibroblast cell line (IMR-90). Very weak or negative nucleolar fluorescence was observed in several normal human tissues including liver, kidney, prostate, and cheek cells. Nucleolar fluorescence was also observed in rat Novikoff hepatoma cells. Although normal rat livers do not have PCNA nucleolar fluorescence, nuclear and nucleolar fluorescence were observed at 18 hr after partial hepatectomy.

Antigens↗
PubMed1983

Antinuclear, antinucleolar, and anticytoplasmic antibodies in patients with malignant melanoma.

Immunofluorescence was used to examine antibodies to cellular antigens in the sera of patients with malignant melanoma. Sera from 60 melanoma patients and from 33 control individuals (13 normal subjects and 20 disease controls) were studied. Ninety % of the melanoma sera were found to have antinuclear antibodies when epithelial cell lines or melanoma lines were used as substrates for their detection, compared to 18% in the control group. Antinucleolar antibodies and anticytoplasmic antibodies were present in 32 and 17%, respectively, in malignant melanoma but none in the controls. Antinuclear and antinucleolar antibodies could be classified into different types according to different patterns of staining and susceptibility of antigens to digestion with DNase, RNase, and trypsin. Certain types of antibodies, such as those showing granular nuclear staining, appeared to be associated with less advanced stages of malignant melanoma, whereas those showing nucleolar and large speckled nuclear staining were associated with more advanced stages of the disease.

Antibodies, Antinuclear↗
PubMed1983

Urinary excretion of antinuclear antibodies.

The frequency of antinuclear antibodies (ANA), the immunoglobulin class of ANA and their specificity for known nuclear antigens were determined in 24-h urine collections from patients with systemic lupus erythematosus (SLE) and progressive systemic sclerosis (PSS). Sixteen % of SLE patients had detectable urine ANA by indirect immunofluorescence using mouse kidney substrate. A higher incidence, 32% of SLE and 28% of PSS patients, had detectable ANA in a titer greater than or equal to 1:16 using HEp-2 cell substrate. IgG ANA was the most frequent immunoglobulin class of antibodies present in the urine; 56% of SLE and 29% of PSS patients with urine ANA had more than one immunoglobulin class of antibodies. Antibodies to Sm, nRNP, SS-A and dsDNA were detected in SLE urine; antibodies to SS-A and centromere were detected in PSS urine. Urine ANA detected on mouse kidney substrate and urine dsDNA antibodies correlated with diffuse proliferative glomerulonephritis in patients with SLE. Sixty-two% of SLE patients with urine ANA had proteinuria. In the remaining SLE patients and in all the PSS patients with urine ANA however, protein excretion was normal. SDS-PAGE revealed heavy and light immunoglobulin molecules in both SLE and PSS patients with urine ANA. The intact immunoglobulin was shown to have ANA activity. ANA present in the urine of SLE and PSS patients with apparently normal renal function may be an early sign of altered glomerular capillary membrane permeability.

Adult↗
PubMed1983

Characterization of DNA in polyethylene glycol precipitated immune complexes from sera of patients with systemic lupus erythematosus.

The nature and the quantity of DNA present in the circulating immune complexes (ICs) from 30 patients with systemic lupus erythematosus (SLE) was characterized. DNA was extracted from IC enriched material prepared by polyethylene glycol precipitation of serum and the extracted DNA was labelled with 32P-phosphate. The size and the nature of DNA was determined by polyacrylamide gel electrophoresis and autoradiography. The quantity of DNA in the PEG precipitates from sera of 10 clinically active SLE was found to be significantly higher (mean 159 X 10(4) ct/min, range 49.9-807 X 10(4) ct/min) than 10 normal controls (mean 24.7 X 10(4) ct/min, range 8.7-47.8 X 10(4) ct/min). Four different sizes of DNA fragments were detected: 370-470, 150-240, 30-40 and 20 base pairs (bp). DNA of 30-40 bp and 20 bp were frequently present in both SLE and normals, but the other two large sized DNA fragments were particularly prominent in SLE patients. In the majority of samples, DNA fragments appeared double stranded.

Antigen-Antibody Complex↗
PubMed1983

In vivo and in vitro production of anti-histone antibodies in NZB/NZW mice.

Anti-histone antibodies have been demonstrated in the sera of patients with both idiopathic and drug-induced lupus. We measured anti-histone antibodies in female NZB/NZW (F1) mice, which are considered to be a model of human SLE. Using a sensitive and quantitative enzyme-linked immunosorbent assay (ELISA), we detected minimal serum antibody activity in NZB/NZW mice younger than 4 mo of age and in nonautoimmune mice at all ages tested. Serum anti-histone antibodies progressively increased in NZB/NZW mice from 4 to 8 mo of age and showed an age-related maturation from IgM to IgG. The predominant antibody activity in the older mice was to the individual histones H2B and H3, and the pattern of reactivity to the histone proteins was similar to that seen in human SLE. We also studied the spontaneous in vitro production of anti-histone antibodies using spleen cells from NZB/NZW mice of different ages. Culture supernatants were analyzed for antibody activity by an ELISA with total histones as the antigen. Spleen cells from older NZB/NZW mice, with elevated serum levels, produced 10-fold higher levels of antibody activity compared to age-matched nonautoimmune mice. Antibody production was maximal at 4 days of culture and was inhibited by the addition of puromycin to the culture. Surprisingly, spleen cells from 1 to 3-mo-old NZB/NZW mice, with normal serum levels, also demonstrated significantly elevated production. The antibodies produced by these young mice were mostly IgM, whereas spleen cells from older mice produced mostly IgG anti-histone antibodies. The present results provide the basis for using the anti-histone antibody system to study further the immune abnormalities that allow for autoantibody production.

Aging↗
PubMed1982

Sjögren's syndrome and polymyositis or dermatomyositis.

Of four patients with Sjögren's syndrome, three had polymyositis and one had dermatomyositis. In all, deposition of IgG, IgA, IgM, and C3 was observed in muscle by immunofluorescent techniques. Serologic studies revealed elevated levels of serum IgG and IgM, rheumatoid factor, and antinuclear antibody with specificity for SS-A and SS-B antigens. In muscle there was a mononuclear cell infiltrate with plasma cell predominance around small vessels and capillaries. Ultrastructural changes in the vessels included reduplication of the basement membrane, endothelial thickening, and numerous tubuloreticular and dense inclusions. In two patients, electrondense deposits were noted in the microvasculature. This combination of immunoglobulin deposition in muscle, prominent microvascular changes, and characteristic serology suggests that the myositis in Sjögren's syndrome may result from small-vessel injury by autoantibodies or circulating immune complexes.

Adult↗