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Biomedical subjects

S Zucker

Publications and source records attributed to S Zucker.

At least 19 recordsLinked to original sources
PubMed1992

Secretion of gelatinases and tissue inhibitors of metalloproteinases by human lung cancer cell lines and revertant cell lines: not an invariant correlation with metastasis.

Numerous studies have reported a correlation between production of 72-kDa (MMP-2) and 92-kDa (MMP-9) type-IV collagenases/gelatinases and the metastatic potential of cancer cells. An abrogating effect of tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2) on metastases has also been noted. In this report we have used sensitive enzyme-linked immunoassays to measure MMP-2, MMP-9, TIMP-1 and TIMP-2 levels in eight human lung-cancer cell lines which were characterized for biological behavior in nude mice. We demonstrated that the Calu-6 and A549 cell lines with the highest metastatic, invasive and tumorigenic potential secreted the highest levels of MMP-2. MMP-9 and TIMP-1 secretions were comparatively low in all cell lines. TIMP-2 secretion, which exceeded MMP-2 secretion for all cell lines, did not correlate with metastatic potential. To further explore these correlations, the metastatic Calu-6 cell line was transfected with a K-rev-1 cDNA expression construct. The K-rev revertant cell lines demonstrated a more differentiated phenotype and were less tumorigenic, invasive and metastatic in nude mice. Nonetheless, the Calu-6 revertant cell lines secreted higher levels of MMP-2 than the parent cell line. In conclusion, invasion and metastasis by lung-cancer cells requires not only enhanced MMP production, but also other less well-understood tumorigenic characteristics. The multiplicity of factors required by cancer cells for dissemination helps to explain the minute fraction of cancer cells from a primary tumor that ever develop into a metastasis.

Animals↗
PubMed1992

Immunoassay of type IV collagenase/gelatinase (MMP-2) in human plasma.

We have developed a sensitive and specific sandwich type enzyme-linked immunosorbent assay (ELISA) to detect type IV collagenase (MMP-2) in human plasma which employs the combination of affinity purified rabbit polyclonal antibodies and mouse monoclonal antibodies to human MMP-2. The MMP-2 ELISA detects latent and activated MMP-2, MMP-2 complexed with TIMP and MMP-2 complexed with alpha 2 macroglobulin (65% efficiency). To determine whether physiologic conditions associated with increased connective tissue turnover are accompanied by increased MMP-2 levels in plasma, we compared enzyme levels in pregnant and nonpregnant women. Plasma MMP-2 (mean +/- standard deviation) was significantly increased (p less than 0.05) in the second half of pregnancy (650 +/- 312) as compared to early pregnancy (356 +/- 139) or the nonpregnant state (387 +/- 193). As a result of the linkage between type IV collagenase production by cancer cells and the metastatic phenotype, the assay of MMP-2 in plasma is of potential clinical value in cancer.

Antibodies, Monoclonal↗
PubMed1992

Modulation of the antigenic phenotype of human melanoma cells by differentiation-inducing and growth-suppressing agents.

Tumor cells often display alterations in their normal program of cellular differentiation. A promising approach for the treatment of cancer involves the induction of terminal differentiation and a loss of proliferative capacity in cancer cells. In human melanoma cells, the combination of mezerein (MEZ) and fibroblast interferon (IFN-beta), results in a rapid and irreversible suppression of cell growth with a concomitant increase in the synthesis of melanin. The induction of terminal differentiation is associated with alterations in the expression of several cellular genes, including fibronectin, ISG-15 and ISG-54, and changes in the expression of specific cell surface antigens, including intercellular adhesion molecule-1 (ICAM-1) and HLA Class I antigens. In the HO-1 human melanoma cell line, induction of terminal differentiation by MEZ plus IFN-beta results in an induction and/or increased expression of ICAM-1, HLA Class I antigens and HLA Class II antigens. IFN-beta and MEZ alone can modulate expression of these antigens to a lower extent than does the combination of compounds. Induction of terminal differentiation and the irreversible suppression of cell growth is not a prerequisite for antigenic modulation in HO-1 cells. This is indicated by the inability of immune interferon (IFN-gamma), a strong inducer of ICAM-1, HLA Class I antigens and HLA Class II antigens synthesis, or the combination of IFN-beta plus IFN-gamma which synergistically but reversibly suppresses HO-1 growth, to induce melanin synthesis or terminal differentiation in HO-1 cells.(ABSTRACT TRUNCATED AT 250 WORDS)

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1992

Human pleural effusions are rich in matrix metalloproteinases.

We identified and characterized type IV collagenase and gelatinase activity in pleural fluid from 32 patients. The capacity to substantially degrade type IV collagen was demonstrated in every pleural sample. Comparable results were also noted for the degradation of a radiolabeled gelatin substrate. Gelatin gel zymography of the pleural fluids revealed two prominent zones of lysis at 66 kDa and 92 kDa. These were identified by specific polyclonal antibodies as human matrix metalloproteinases MMP-2 and MMP-9. The concentration of MMP-2 in pleural fluid, as measured by enzyme-linked immunoassay, averaged 1,622 ng/ml whereas those of MMP-9 were 210 ng/ml. Substrate degradation activity was compared in both serum and pleural fluid from three patients and found to be similar. In serum this enzymatic activity was primarily due to MMP-9 whereas in pleural fluid, the predominant gelatinase was MMP-2. This was confirmed by immunoassay that showed that MMP-2 levels were two to five times higher in pleural fluid than in serum. We conclude that substantial amounts of MMP-2 and, to a lesser degree, MMP-9 are present in pleural effusions. The bioactivity and the immunoactivity of these enzymes did not help to distinguish among pleural fluids characterized as transudates, nonmalignant exudates, or malignant exudates. The differences in the distribution of these enzymes in pleural fluid and blood suggest that their presence is not due simply to the ultrafiltration of plasma, but rather to synthesis by the resident cells at the pleural surfaces.

Biomarkers↗
PubMed1992

Characterization of metalloproteinases and tissue inhibitors of metalloproteinases in human plasma.

In this study, we have identified and characterized metalloproteinases and tissue inhibitors of metalloproteinases (TIMPs) in human plasma. Treatment of plasma with trypsin or aminophenylmercuric acetate resulted in activation of latent gelatinolytic activity. Fractionation of plasma by gelatin Sepharose chromatography resulted in the isolation of 72 kDa and 92 kDa gelatinases/type IV collagenases. The 72 kDa gelatinase was purified by gel filtration chromatography. Stromelysin-1 was isolated from plasma by Matrex green A affinity chromatography. Immunoblotting of plasma fractions with antibodies to unique peptide regions of human gelatinases differentiated the 72 kDa gelatinase from the 92 kDa gelatinase. Antibodies to the amino terminal peptides of each enzyme were used to determine that plasma gelatinases circulate as latent proenzymes. Immunoblotting with antibodies directed against human stromelysin identified a 57 kDa stromelysin. TIMP-1 (28 kDa) and TIMP-2 (21 kDa) were also identified by immunoblotting of gelatin Sepharose bound plasma proteins using non-crossreacting antibodies to each protein.

Amino Acid Sequence↗
PubMed1992

Growth inhibition and modulation of antigenic phenotype in human melanoma and glioblastoma multiforme cells by caffeic acid phenethyl ester (CAPE)

The active component of the honeybee hive product propolis, caffeic acid phenethyl ester (CAPE), has been shown to display increased toxicity toward various oncogene-transformed cell lines in comparison with their untransformed counterparts (Su et al., 4: 231-242, 1991). This observation provides support for the concept that it is the transformed phenotype which is specifically sensitive to CAPE. In the present study, we have determined the effect of CAPE on the growth and antigenic phenotype of a human melanoma cell line, HO-1, and a human glioblastoma multiforme cell line, GBM-18. For comparison, we have also tested the effects of mezerein (MEZ), mycophenolic acid (MPA) and retinoic acid (RA), which can differentially modulate growth, differentiation and the antigenic phenotype in these human tumor cell lines. Growth of both cell lines was suppressed by CAPE in a dose-dependent fashion, with HO-1 cells being more sensitive than GBM-18 cells. The antiproliferative effect of CAPE was enhanced in both cell types if CAPE and MEZ were used in combination. Growth suppression was associated with morphological changes in H0-1 cells, suggesting induction of a more differentiated phenotype. CAPE also differentially modulated the expression of several antigens on the surface of the two tumor cell lines. These results suggest a potential role for CAPE as an antitumor agent, an antigenic modulating agent and possibly a differentiation inducing agent.

Antigens, Neoplasm↗
PubMed1991

Proteinase-alpha 2 macroglobulin complexes are not increased in plasma of patients with cancer.

Alpha 2-macroglobulin, a major glycoprotein component of plasma, is unique in its capacity to bind and inhibit the proteolytic activities of all classes of proteinases. Since proteinases implicated in cancer dissemination (type-IV collagenase, plasminogen activator, cathepsins B) are normal constitutents of blood, we have explored the hypothesis that elevated tissue levels of activated proteinases bound to alpha 2M might be detected in plasma of patients with cancer. To test this premise, blood was collected from 149 subjects (33 healthy controls, 31 patients with infections and non-malignant diseases, 16 with myeloproliferative disease, 10 with gastrointestinal cancer, 7 with genito-urinary cancer, 16 with lung cancer, 14 with lymphoma, 11 with miscellaneous cancers and 11 with chronic lymphocytic leukemia and myeloma). Plasma was assayed for alpha 2M-proteinase complexes using a sandwich ELISA which employs a mouse monoclonal antibody (MAb) that binds to a neo-antigenic determinant on complexed alpha 2M and a rabbit polyclonal anti-native human alpha 2M antibody. The concentration of complexed alpha 2M in healthy controls was 14.2 +/- 9.8 micrograms/ml (mean +/- standard deviation). No significant differences in complexed alpha 2M were noted between normal and cancer groups (range 7.4-14.6 micrograms/ml). On the basis of these data, we propose that, in patients with cancer, activated proteinases are bound locally to inhibitors in the tissues and are not available to form complexes with plasma alpha 2M. An alternative explanation is that proteinases are not secreted in excess by cancer cells in vivo.

Adult↗
PubMed1991

Detergent extraction and characterization of tumor hemolytic factor from plasma membranes of oncogene transformed fibroblasts.

Cancer cells have the capacity to lyse erythrocytes by a cell-contact-requiring phenomenon. Subcellular fractionation procedures have revealed that the hemolytic principle resides in the cancer cell plasma membrane. In this study we report the detergent extraction of a potent hemolytic factor from the plasma membranes of ras-oncogene-transformed fibroblasts. Ammonium-sulfate partitioning (60-100%) of detergent-extracted proteins was used to enrich hemolytic activity. Tumor membrane Hemolytic Factor (mTHF) was inactivated by treatment with papain, suggesting that it is a protein. mTHF was inhibited by serum, but was unaffected by extremes of temperature and pH, also by metal chelation with EDTA. Surface radio-iodination of tumor cells and isolation of cell organelles was used to characterize the outer plasma membrane localization of mTHF. mTHF retained hemolytic activity when reconstituted into stable phospholipid vesicles. Pre-incubation of mTHF with red cell ghosts led to an abrogation of hemolytic activity. mTHF-induced hemolysis consists of a 2-stage phenomenon: an early binding step, followed by hemolysis after 4 hr.

Animals↗
PubMed1990

Localization of collagenase at the basal plasma membrane of a human pancreatic carcinoma cell line.

We have recently presented biochemical evidence for collagen and gelatin degrading activities associated with plasma membranes of various human cancer cell lines. In this report we describe the localization of interstitial collagenase at the basal plasma membrane of the human pancreatic cancer cell line RWP-I, using immunofluorescence and ultrastructural immunogold labeling techniques. Collagenase was expressed on the extracellular face of the plasma membrane. Furthermore, the immunogold labeling was concentrated on the long, finger-like microvillous projections typically seen on the basal cell surface, while the short, brush-like projections characteristic of the apical cell surface were unlabeled. When the cytoplasmic face of the membrane was made accessible, the number of reactive sites increased markedly, indicating a high concentration of enzyme at the inner surface of the plasma membrane. When plasma membrane fractions of RWP-I cells were prepared by differential centrifugation, high salt washes virtually failed to extract collagenase activity from the membrane, while detergent extraction with n-octyl glucoside, a detergent used in the purification of integral membrane proteins, yielded soluble collagenase activity. When detergent extracted membrane fractions were passed over an anticollagenase immunoaffinity column, collagenase was specifically bound, as demonstrated by the TCA and TCB degradation of type I collagen by the bound material. Gelatinolytic activity did not bind to the column. Furthermore, immunoprecipitation of 125I-labeled detergent extracts of tumor membranes yielded a single Mr 55,000 band consistent with the zymogen form of the connective tissue collagenase. These morphological and biochemical findings suggest that collagenase is a tightly associated component of the basal plasma membrane, where it occupies a strategic location for directional proteolysis during cell migration and invasion.

Cell Membrane↗
PubMed1990

Extraction of type-IV collagenase/gelatinase from plasma membranes of human cancer cells.

Tumor proteinases are considered to be important in the process of cancer invasion and metastasis. We have proposed that the surface membrane localization of these proteinases places them in an optimal site to facilitate the invasion of surrounding extracellular matrix. In this study, we have used the organic solvent, n-butanol, and the detergent, n-octyl-glucoside, to sequentially extract metalloproteinases from crude plasma membranes of human RWP-I pancreatic cancer cells. Anion exchange chromatography and gel permeation chromatography were employed to further purify enzymes with the capacity to degrade gelatin, type-IV collagen, and carboxymethylated transferrin. Gelatin zymography was used to demonstrate proteinase bands of 92, 70 and 62-kDa. Immunoblotting of solubilized, partially purified pancreatic cancer plasma membrane proteins using polyclonal rabbit antibodies, which have specificity for type-IV collagenase/gelatinase, resulted in the recognition of a 70-kDa protein, but not the 92-kDa gelatinase. A type-IV collagenase/gelatinase of 68-kDa was similarly identified in A2058 human melanoma cancer cell plasma membranes.

Cell Line↗
PubMed1990

Purification and characterisation of soluble tumour haemolytic factor isolated from oncogene transformed fibroblasts.

Numerous studies have shown that intact cancer cells and cell extracts have the capacity to lyse erythrocytes in vitro. The transformation of NIH-3T3 fibroblasts by ras oncogenes has recently been demonstrated to result in tumour cells releasing a haemolytic factor. The purpose of this study has been to purify and further characterise the soluble tumour haemolytic factor (sTHF) produced by mouse fibroblasts transformed by T24 human bladder cancer DNA and by the cloned Harvey murine sarcoma viral oncogene. To this end, transformed fibroblasts were cultivated in serum-free medium. The cell-free supernatant was treated with ammonium sulphate and the precipitate achieved at 60-100% saturation was dialysed and applied to a gel filtration column. A haemolytic factor was eluted with an Mr between 65,000 and 75,000. Zinc chelate and strong anion exchange column chromatography resulted in greater than 3,000-fold enrichment of sTHF. SDS-PAGE of sTHF resulted in a single protein band of 66,000 Da. Soluble THF had no immunological cross-reactivity with known cytokines produced by lymphocytes and macrophages. The pathophysiological role of sTHF in cancer remains to be determined.

Animals↗
PubMed1990

Chlorpromazine-induced immunopathy: progressive increase in serum IgM.

Long-term chlorpromazine therapy has been associated with the asymptomatic development of a high incidence of antinuclear antibodies, coagulation inhibitors, and increased serum levels of IgM. The purpose of this study has been to characterize the natural history of this chlorpromazine-induced (CPZ) immunopathy. To this end we carried out a prospective study of schizophrenic patients with the immunopathy to compare the effect of continuing CPZ versus switching to haloperidol therapy. Although no marked differences were noted between the 2 groups at the end of 5 years, 6 of 29 patients who continued to receive CPZ, as compared to none of 14 patients on haloperidol, had progressive elevations of serum IgM. In spite of a high incidence of antinuclear antibodies, none of the patients developed a lupus-like syndrome. One patient, however, who had been maintained on CPZ for more than 15 years, developed Waldenström macroglobulinemia, as characterized by an IgM monoclonal gammopathy and a lymphocyte immunoglobulin heavy and kappa light chain gene rearrangement. Another CPZ-treated patient developed immune thrombocytopenia. Based on the potential serious sequelae of prolonged stimulation of the immune system by CPZ, we recommend that patients who develop an increase in serum IgM while on CPZ be switched to other types of anti-psychotic medications.

Antibodies, Antinuclear↗
PubMed1989

Identification of tissue factor in two human pancreatic cancer cell lines.

We have studied the effects of two human pancreatic cancer and two human small cell lung cancer cell lines on clotting and platelet aggregation. Both pancreatic lines markedly shortened recalcification times and induced platelet aggregation. The lung cancer lines produced little shortening of recalcification times and no platelet aggregation. The clotting and aggregation activities of the pancreatic lines were further characterized. Recalcification times following the addition of cancer cell line material to plasmas deficient in factors VII and X were markedly prolonged, suggesting that the activity is due to tissue factor. Hirudin, an inhibitor of thrombin from the saliva of leeches, and rabbit polyclonal immunoglobulin G anti-bovine brain tissue factor inhibited both procoagulant and aggregation activities. Apyrase (an enzyme degrading ADP), diisopropylfluorophosphate (a serine protease inhibitor) and L-trans-epoxysuccinylleucylamido(4-guanidino)butane (a cysteine protease inhibitor) failed to inhibit these activities. Increasing concentrations of heparin inhibited platelet aggregation. Subcellular fractionation studies showed these activities to be localized to the plasma membrane. The association between mucin and the acceleration of clotting has been well described. The absence of mucin in electron micrographs of these pancreatic whole cells, membrane fractions, and shed microvesicles, as well as the failure of chaotropic agents (i.e., agents stripping material extrinsic to the cell membrane such as mucin) to abrogate this activity support these activities being intrinsic to the plasma membrane. These data strongly suggest that these activities are due to tissue factor which appears to be released as microvesicles in vitro. The release of tissue factor via microvesicles in vivo is one possible mechanism for the coagulopathy sometimes seen in patients with pancreatic carcinoma.

Adenocarcinoma↗