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S Zucker

Publications and source records attributed to S Zucker.

At least 91 records · Page 5Linked to original sources
PubMed1986

Analysis of the cell membrane proteolytic enzymes of the B16, F1, F10, and BL6 melanoma and their role in target cell destruction.

The tumor-induced red blood cell (RBC) cytolysis assay has been used to demonstrate that three B16 melanoma sublines, the F1, F10, and BL6, cause the cytolysis of normal red blood cells in vitro. RBC cytolysis was inhibited for all three sublines by metalloprotease inhibitors. Cell membrane preparations have been prepared for all three sublines and tumor cell membrane-induced RBC cytolysis was also shown to be inhibited by metalloprotease inhibitors. The F10 and BL6 sublines were shown to have cell membrane-bound proteases. The BL6 subline has a cell membrane enriched in an enzyme with a trypsin-like arginine specificity. The trypsin-like protease may have a metal dependence. The BL6 subline has a collagenolytic cell membrane enzymes and a chymotrypsin-like cell membrane enzyme. B16 cell membrane enzymes may be responsible for RBC cytolysis in vitro in a process requiring divalent cations.

Animals↗
PubMed1986

Reticuloendothelial system Fc receptor function in the drug induced lupus erythematosus syndrome.

In view of reported delayed clearance of 51Cr labelled IgG sensitized autologous erythrocytes in systemic lupus erythematosus, we measured clearance half-time (t1/2) in 16 patients with the drug induced lupus erythematosus syndrome. The mean t1/2 for the patients with drug induced lupus was 52 min, and for 16 normals mean was 48 min, a difference without statistical significance. Nine of the patients with drug induced lupus had laboratory abnormalities only, and mean t1/2 in this group was not significantly different from that in the 7 symptomatic patients. T1/2 was not significantly associated with Clq or DNA binding levels in symptomatic or asymptomatic patients. Normal mean clearance of model immune complexes was thus present in patients with drug induced lupus, with and without symptoms, and may protect against renal and central nervous system disease in drug induced lupus erythematosus.

Adult↗
PubMed1985

The proteolytic activities of chymopapain, papain, and papaya proteinase III.

The three proteinases present in papaya latex: papain (EC 3.4.22.2) chymopapain and papaya proteinase III (EC 3.4.22.6), were standardized by active-site titration, and compared in proteolytic activity against azocasein, serum albumin and cartilage proteoglycan. The activities were all of the same order, although there were differences in pH dependence. SDS-polyacrylamide gel electrophoresis of the early products of digestion of albumin and phosphorylase a showed very similar patterns for the three papaya proteinases. Kinetic parameters for hydrolysis of benzyloxycarbonyl-phenylalanyl-arginyl-7(4-methyl)coumarylamide were determined for the three enzymes. Values for kcat/Km varied only within a factor of 2, but the individual constants were much higher for papain than for chymopapain and papaya proteinase III. In contrast to the results obtained with the synthetic substrate, the kinetic parameters for the initial hydrolysis of succinyl-albumin were very similar for the three papaya proteinases. This was consistent with their similar proteolytic activities in other assays.

Albumins↗
PubMed1985

Plasminogen-dependent fibrinolytic activity in normal human lymphocytes: diminished lymphocyte plasminogen activator in chronic lymphocytic leukemia.

Discrepancies in correlations between fibrinolytic activity and metastatic potential of malignant cells has resulted in speculation on the putative role of plasminogen activators (PA) in cancer. In this report we have compared lymphocyte PA from 40 patients with chronic lymphocytic leukemia (CLL) to normal human B- and T-lymphocytes. Lymphocytes were isolated from peripheral blood by Ficoll-Hypaque centrifugation. The B- and T-cells were further separated on nylon wool columns. Cell PA activity and cell membrane PA were determined using 3H-fibrin-coated plates with added human plasminogen. Lymphocytes did not lyse 3H-fibrin in the absence of plasminogen. Plasminogen-dependent fibrinolytic activities of normal B- and T-lymphocytes were comparable. The addition of protease inhibitors with trypsin or plasmin specificity to lymphocytes significantly inhibited normal PA, thus substantiating the serine protease spectrum of lymphocyte PA. Examination of lymphocytes from greater than 95% of patients with chronic lymphocytic leukemia revealed a marked decrease in lymphocyte and cell membrane PA as compared to normals. No correlation between Stage of CLL and lymphocyte PA was observed. Likewise, an inhibitor of PA in CLL lymphocytes was not detected. The function of PA in normal B-lymphocyte physiology and the potential pathogenetic role of diminished PA in CLL lymphocytes remain to be explored.

Fibrinolysis↗
PubMed1985

Role for different cell proteinases in cancer invasion and cytolysis.

The crucial role of non-plasminogen dependent serine proteinases is tissue invasive and cytolytic functions of Walker 256 cancer cells has been documented using a rat urinary bladder invasion and a 125I-labelled fibroblast cytolysis assay. The invasive capacity of these cancer cells was abrogated by non toxic concentrations of the serine proteinase inhibitors, diisopropylfluorophosphate and phenylmethylsulfonylfluoride, but not by metallo or cysteine proteinase inhibitors. Although tumour cell collagenase activity and plasminogen activator were demonstrated, these proteolytic enzymes were not essential in these in vitro assays. These results suggest that different categories of proteinases play specific roles in the complicated process of cancer invasion.

Animals↗
PubMed1985

Anemia in cancer.

In summary, anemia developing in a patient with cancer can be due to several different factors. A relative failure of erythropoiesis, in conjunction with a modestly shortened erythrocyte survival, is the most likely explanation for the anemia and can occur in patients with or without bone marrow invasion. Several theories have been proposed to explain the mechanism of limited red cell production in cancer. Internal iron starvation and cancer toxic factors have been widely implicated. Immunoglobulin inhibitors of erythropoiesis occur in the rare entity, pure red cell aplasia, which is sometimes associated with thymomas. Autoimmune hemolytic anemia and microangiopathic hemolytic anemia can also occur in patients with solid cancers, pointing out the need for a complete evaluation of anemia in any patient with recent-onset anemia. Successful treatment and prognostic implications of anemia in cancer is dependent on proper diagnosis.

Anemia↗
PubMed1985

Diversity of melanoma plasma membrane proteinases: inhibition of collagenolytic and cytolytic activities by minocycline.

The tissue-destructive proteinases of B16-BL6 melanoma cells from C57BL/6 mice and subcellular fractions were examined. Cancer cell organelles were isolated following nitrogen cavitation with the use of sucrose density gradient centrifugation. Serine, cysteine, and metalloproteinases were assayed with the use of radiolabeled proteins and synthetic substrates. Tumor-induced red blood cell lysis was quantitated by measurement of the release of isotope from 59Fe-labeled red blood cells (RBC) cocultivated with melanoma cells; the RBC were from Wistar rats. Enzyme inhibitors with specificity toward different classes of proteinases were used in the above assays to categorize the enzymes responsible for substrate degradation. Results indicated that intact melanoma cells, cell organelles, and cytosol contain proteinases that can degrade collagen and gelatin and lyse normal RBC. Melanoma plasma membranes are highly enriched in collagenase, gelatinase, cysteine proteinase, plasminogen activator, and cytolytic activity. The inhibition of tumor collagenolytic, gelatinolytic, and cytolytic activities by EDTA and 1,10-phenanthroline but not by diisopropyl fluorophosphate and N alpha-p-tosyl-L-lysine chloromethyl ketone indicates that metalloproteinases are the active enzymes in these assays. Minocycline, a synthetic tetracycline with demonstrable inhibitory activity with other mammalian collagenases, also inhibited melanoma collagenolytic and cytolytic activities.

Animals↗
PubMed1985

Diversity of human pancreatic cancer cell proteinases: role of cell membrane metalloproteinases in collagenolysis and cytolysis.

In this study we have examined the tissue-destructive proteinases of human pancreatic ductal cancer cell lines derived initially from xenogenic transplants. Cancer cell organelles were isolated following nitrogen cavitation using sucrose density gradient centrifugation. Serine, cysteine, and metalloproteinases were assayed using radiolabeled protein and synthetic substrates. Tumor-induced RBC lysis was quantitated by measuring the release of isotope from 59Fe-labeled RBCs co-cultivated with tumor cells or subcellular fractions. Enzyme inhibitors with specificity toward different classes of proteinases were used in the above assays to categorize the enzymes responsible for substrate degradation. Results indicated that intact pancreatic cancer cells (RWP-1 and RWP-2 cell lines), cell homogenate, and cytosol contain proteinases which were able to degrade [3H]collagen (type I) and [3H]gelatin and lyse normal RBCs. Cancer cell membrane fractions were enriched in collagenolytic, gelatinolytic, and cytolytic activities which could be abrogated by EDTA but not by inhibitors of serine or cysteine proteinases, which indicates that metalloproteinases are the active enzymes in these assays. Although plasminogen activator and cysteine proteinases were also enriched in the tumor cell membranes, these activities were not required for collagen degradation or cytolysis. We conclude that human cancer cell membrane proteinases are advantageously situated to facilitate damage to surrounding normal tissues.

Cell Line↗
PubMed1983

Isolation and characterization of a trypsin-like serine proteinase from the membranes of Walker 256 carcino-sarcoma cells.

A serine proteinase was isolated from Walker-256-carcino-sarcoma plasma-membrane-enriched preparations by affinity chromatography employing soya-bean trypsin inhibitor as the ligand. This enzyme was termed 'memsin' owing to its membrane location and trypsin-like substrate specificity. Analysis of this preparation by steric-exclusion high-pressure liquid chromatography (h.p.l.c.) resulted in a single peak of enzyme activity. Calculations of the rates of inactivation of memsin by peptidyl-chloromethanes and comparison with rate constants obtained with other serine proteinases indicated that memsin closely resembled trypsin and acrosin. Digestion of oxidized ribonuclease by memsin and analysis of the resulting peptides by h.p.l.c. yielded a chromatogram that was very similar to one generated by a tryptic digest of oxidized ribonuclease. This enzyme could possibly play a role in tumour-cell invasion.

Animals↗
PubMed1983

Apolipoprotein E synthesis in human kidney, adrenal gland, and liver.

Human tissues were incubated in vitro with radiolabeled amino acids to determine whether plasma apolipoproteins are synthesized in human kidney. Subsequently, tissue extracts were screened with antisera directed against apolipoprotein E (apo E), apolipoprotein B (apo B), apolipoprotein AI (apo AI), and bulk apolipoproteins of high density lipoprotein (HDL). Newly synthesized apo E, but not apo AI or apo B, was identified in kidney and adrenal cortex. Estimates of relative rates of apo E synthesis in vitro suggest that a substantial portion of adrenal and kidney protein synthesis is committed to apo E synthesis. The relative rate of apo E synthesis was 4-6 times greater in kidney cortex than in kidney medulla. Analysis of immunoreactive apo E showed that kidney and adrenal apo E species have the same electrophoretic mobility in NaDodSO4/polyacrylamide gels as does plasma apo E. Further characterization by high resolution two-dimensional gel analysis indicated that the isoforms of newly synthesized kidney and adrenal apo E correspond to specific isoforms of plasma apo E. These findings suggest that apolipoproteins arising from peripheral tissues may play an important role in lipid transport and metabolism.

Adrenal Glands↗
PubMed1983

Mechanism of BCG-activated macrophage-induced tumor cell cytotoxicity: evidence for both oxygen-dependent and independent mechanisms.

Activated peritoneal macrophages have been shown to be cytotoxic to cancer cells. Bacillus Calmette-Guerin (BCG)-activated rat peritoneal macrophages have a basal cytolytic potential for 3H-thymidine-labelled Walker 256 cancer cells in vitro that can be markedly enhanced by digitonin. This stimulation of cytotoxicity can be partially inhibited by catalase and the combination of superoxide dismutase plus catalase. This suggests that digitonin stimulates activated macrophages to produce superoxide, hydrogen peroxide and possibly other free radicals which can augment macrophage-induced tumor cell cytotoxicity. After a 2-hour incubation with digitonin, macrophages are no longer stimulated by digitonin. However, after a 2-hour drug preincubation period, inhibitors of serine protease activity (DFP, TLCK, SBTI) and inhibitors of protein synthesis (cycloheximide) are potent inhibitors of basal macrophage-induced tumor cytotoxicity. We suggest that BCG-activated macrophages have two mechanisms for destroying cancer cells: one mediated by proteolytic activity, and a second mechanism dependent on the generation of oxygen-derived free radicals.

Animals↗
PubMed1983

Characterization of tumor cell membrane serine proteases by polyacrylamide gel electrophoresis.

Studies in our laboratory have indicated that tumor cell membrane-bound proteases are responsible for the ability of tumor cells to lyse normal cells in vitro. In order to evaluate the tumor cell membrane enzymes, a purified tumor cell membrane preparation was prepared and the nonionic detergent Triton X-100 was used to extract active enzymes from the cell membranes. The solubilized membrane enzymes were then studied by Triton X-100 polyacrylamide gel electrophoresis under non-denaturing conditions. Using this technique the tumor cell membranes were shown to contain esterproteases that reacted with the substrates alpha-naphthyl acetate and naphthol-AS-aminocaproate. These esterproteases were inhibited by diisopropyl fluorphosphate and tosyl lysine chloromethyl ketone but not by tosylamide phenylethyl chloromethyl ketone, soybean trypsin inhibitor p-chloromercuribenzene sulfonic acid; N-ethylmaleimide choline iodide, alpha-1-anti-trypsin. NaF, epsilon-aminocaproic acid, ethylenediamine tetraacetic acid, or eserine. SBTI affinity chromatography of the tumor cell membrane extract revealed that some of the serine esterproteases bound to the SBTI column. The proteolytic activity of the tumor cell membrane extract and a fraction eluted from the SBTI affinity column was demonstrated using casein. We conclude that the tumor cell membranes contain previously undescribed serine proteases that are identifiable by their esterase activity and inhibitor profiles in polyacrylamide gels.

Animals↗
PubMed1983

Organ distribution of circulating very low density lipoproteins (VLDL): fate of hematopoietic growth inhibitory VLDL in the rat.

The organ distribution of very low density lipoproteins, with known hematopoietic cell growth inhibitory effects, was studied in the rat. Animals received intravenous injections of 125I-labelled VLDL and tissue uptake was monitored over 4 days. Uptake into 8 organs was studied using a technique that excluded blood associated radioactivity. While we demonstrated that the liver predominated in tissue uptake of radioactivity on a total organ basis, bone marrow had the greatest 125I-VLDL uptake per gram of tissue, six times greater for bone marrow than liver. Thus we have demonstrated that the bone marrow is a very active organ in terms of uptake of VLDL. This suggests that the in vitro inhibitory effect of rat VLDL on bone marrow cell proliferation may have physiologic significance in the in vivo regulation of marrow cell proliferation.

Animals↗
PubMed1982

Role of tumor cell membrane-bound serine proteases in tumor-induced target cytolysis.

The tumor-induced marrow and red blood cell cytolysis assays have been used to explore the mechanism of cancer cell destruction of normal cells. Previously, we suggested that tumor-induced cytolysis was caused by tumor cell membrane-bound serine proteases. In this study, we have shown that concentrations of the broad-spectrum serine protease inhibitor diisopropylfluorophosphate that did not inhibit tumor cell DNA and protein synthesis completely abrogated tumor-induced red blood cell cytolysis. In addition, tumor cell membranes isolated by differential and sucrose density gradient centrifugation and characterized by electron microscopy and enzyme marker analysis were cytolytic for rat 59Fe-labeled red blood cells. The specific activity expressed as release index (%) per microgram of protein was 1.620 for the tumor cell membrane preparations as compared to 0.002 for intact Walker 256 tumor cells. Tumor cell membranes solubilized in Triton X-100 had activity in the p-toluenesulfonyl-L-arginine methyl ester assay for trypsin-like enzymes and the N-benzoyl-L-tyrosine ethyl ester assay for chymotrypsin-like enzymes. The enzyme activities demonstrated in these assays could be inhibited by N-alpha-p-tosyl-L-lysine chloromethyl ketone HCl and L-1-tosylamide-alpha-phenyl-ethyl chloromethyl ketone, respectively. Using [3H]diisopropylfluorophosphate affinity labeling of the tumor cell membrane proteins followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, we have identified membrane-bound serine protease(s) that appear to be responsible for tumor-induced marrow and red blood cell cytolysis.

Animals↗
PubMed1981

Rat plasma lipoprotein inhibitors of lymphocyte proliferation: specific membrane receptor for very low density lipoproteins.

We have compared the inhibitory effects of plasma lipoproteins on DNA synthesis by phytohemagglutinin-stimulated rat lymphocytes. The order of suppressor potency was: very low density lipoproteins (VLDL) greater than low density lipoproteins much greater than high density lipoproteins. We have further demonstrated the presence of specific receptor sites for VLDL on rat spleen lymphocytes. The number of VLDL receptors was estimated at 31,000 by a modified Scatchard analysis. Our results suggest that in the normal physiologic state, the proliferation of circulating lymphocytes may be suppressed by the combined effects of plasma lipoproteins. Furthermore, it would appear that the binding of plasma VLDL to specific receptors on rat lymphocytes would be the initial physiological event in this inhibitory process.

Animals↗
PubMed1981

Membrane receptors for very low density lipoprotein (VLDL) inhibitor of lymphocyte proliferation.

Physiologic concentrations of human plasma very low density lipoproteins inhibit the DNA synthesis of lymphocytes stimulated by allogeneic cells or lectins. In this report we have compared the effects of isolated lipoproteins [very low density lipoproteins (VLDL), low density lipoproteins (LDL), and high density lipoproteins (HDL)] and lipoprotein-depleted plasma (LDP) on DNA synthesis by phytohemagglutinin-stimulated human lymphocytes. The relative potency for the inhibition of lymphocyte proliferation was VLDL greater than LDL greater than HDL greater than LDP. Fifty percent inhibition of DNA synthesis was observed at a VLDL protein concentration of 1.5--2.0 microgram/ml. We have further demonstrated the presence of specific receptors for VLDL on human lymphocytes. Native VLDL was more effective than LDL in competing for 125I-VLDL binding sites. Subsequent to binding to lymphocytes, 125I-VLDL was internalized and degraded to acid-soluble products. Based on a Scatchard analysis of VLDL binding at 4 degrees C, the number of VLDL receptors per lymphocyte was estimated at 28,000 +/- 1300. Based on an estimated mean binding affinity for the VLDL receptor complex at half saturation of approximately 8.8 X 10(7) liter/mole, it is estimated that 91% of lymphocyte VLDL receptors are occupied at physiologic VLDL concentrations in blood. Although the immune regulatory role of plasma lipoproteins is uncertain, we suggest tha VLDL and LDL-In may maintain circulating blood lymphocytes in a nonproliferative state via their respective cell receptor mechanisms.

Cholesterol↗
PubMed1981

Rat hematopoietic cell receptors for very low density lipoprotein (VLDL) growth inhibitor.

Physiologic concentrations of rat plasma very low density lipoproteins (VLDL) have profound inhibitory effects in tissue culture on the proliferation of hormonally stimulated bone marrow granulocytic and erythrocytic cells and mitogen-stimulated spleen cells. In this study we have evaluated the in vitro uptake and binding of 125I-VLDL by rat marrow cells and spleen lymphocytes. To this end, VLDL were isolated from rat plasma by sequential ultracentrifugation flotation and radioiodinated using iodine monochloride. Biological activity of 125I-VLDL was ascertained by demonstrating that 125I-VLDL and native VLDL had comparable proliferative inhibitory effects when added to erythropoietin-stimulated marrow cells and PHA-stimulated lymphocytes. After 1 h exposure of marrow to VLDL, exhaustive cell washing did not reverse the lipoprotein growth inhibitory effect. The cell uptake of VLDL was evaluated by adding 125I-VLDL to marrow or spleen cells. Uptake of 125I-VLDL by rat cells showed a preference for binding of VLDL as compared to chylomicrons, LDL, or HDL. Based on Scatchard plot analysis of 125I-VLDL binding at 37 degrees C, the approximate number of saturable VLDL receptors available per marrow or spleen cell during a 3 h incubation was 34,000 and 63,000, respectively. We conclude that rat marrow cells and lymphocytes have specific receptors for plasma VLDL and that receptor binding of VLDL is an initial step in its growth inhibitory effect. The physiologic role of plasma lipoprotein cell growth inhibitors will remain speculative, however, until the in vivo distribution of the biologically active lipoprotein moiety to extravascular sites of hematopoiesis has been determined.

Animals↗