Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two rat anti-B16 melanoma monoclonal antibodies (MoAb), designated IB16-6 and IB16-8, recognize an epitope expressed with high density on the surface of B16 parental cells and B16-F1, F10, F10FLR, and BL6 sublines. The purpose of this study was to define by means of cytolytic and clonogenic assays whether these MoAbs reacted with the same or distinct determinants as those recognized on B16 targets by lymphokine-activated killer (LAK) cells. Using 125I-labeled antibody and Scatchard analysis, the affinity constant (KA) of IB16-6 was determined to range from 5.6 to 9.4 x 10(8) liter/M and the number of receptor sites per B16 cell was 4.8 x 10(4) to 2.5 x 10(5). The effects of anti-B16 MoAb on LAK activity were determined by either preincubating 51Cr-labeled B16 target cells with varying concentrations of MoAb, followed by the cytolytic assay, or exposing unlabeled B16 cells to MoAb, and then carrying out a 10-day clonogenic assay. Over a wide range of antibody concentrations, IB16-6 and IB16-8 had minimal effects on LAK activity, and even at MoAb concentrations up to 1 mg there were no changes in target cell sensitivity or colony-forming ability. Enzymatic treatment of B16 melanoma cells with either trypsin or pronase completely removed the epitope recognized by MoAb IB16-6 but did not alter B16 sensitivity to LAK cells. These observations indicate that the LAK recognition unit was distinct from the epitope reactive with MoAb IB16-6 and that the B16 determinant(s) recognized by LAK cells is resistant to proteolytic enzymes. The molecular structure of each of these remains to be determined.
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PMID:Distinct and non-cross-reactive epitopes are recognized on B16 melanoma by LAK cells and anti-B16 monoclonal antibodies. 169 Aug 93

A panel of monoclonal antibodies (MoAbs), produced against the murine B16 melanoma, has been used to characterize its phenotypic diversity. Six MoAbs that did not bind to primary cultures of kidney, brain or liver, spleen cells, thymocytes, 3T3 fibroblasts, melanin, or transferrin receptors were selected for further evaluation. Five MoAbs, which recognized surface antigens expressed on parental B16 cells and the B16-F1, B16-F10, B16-F10 FLR, and B16-BL6 sublines, did not appear to cross-react with each other, suggesting that they identified antigenically distinct epitopes. Four MoAbs, designated as IB16-2, IB16-4, IB16-8, and IB16-10, recognized B16 surface antigens that were variably expressed over short periods of time. This variable expression was independent of the cell cycle and was characteristic of four B16 sublines. Two of these MoAbs, both of the IgG2b isotype, fixed rabbit and guinea pig complement and were cytolytic in the presence of rabbit complement. One MoAb, designated IB16-6, recognized a surface antigen consistently expressed on greater than 90% of cells of both the parental tumor and the sublines. This MoAb bound to several murine and one human melanoma cell line, but not to other histopathological types of tumors or normal tissues. The cellular antigen that this antibody recognized was not detected in the cytoplasm, did not modulate in the presence of IB16-6, and was sensitive to trypsin, pronase, alcohols, acetone, and detergents, thereby suggesting that it was a protein. Our data are among the first that directly show the extent of phenotypic diversity of the B16 melanoma and sublines that have been derived from it.
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PMID:Phenotypic diversity of murine B16 melanoma detected by anti-B16 monoclonal antibodies. 243 32

Studies with four different transplantable murine tumors demonstrated that surgical instruments contaminated by contact with a tumor mass could produce tumors in a surgical wound. Eighty-seven per cent of mice with wounds made by invisibly contaminated scissors developed tumors. Irrigation with water did not prevent tumor growth. Before spilled tumor cells can invade and grow into a recurrence in the wound site, they must first attach to underlying extracellular matrix. We have devised a simple in vitro assay to identify inhibitors of tumor-cell attachment to develop therapeutic compounds that can prevent tumor-cell reimplantation. Various test compounds, including proteases (trypsin and Dispase), known modulators of matrix metabolism (proline analogues, cycloheximide, heparin, cortisone, cortexolone, and heparin-steroid combinations), large molecular weight polymers (agarose, dextran, polyethylene oxide), and synthetic fibronectin peptides were tested for their ability to inhibit mouse melanoma (B16-F10) cell attachment to gelatinized dishes. Most of these compounds had little or no effect on tumor-cell adhesion when cells were plated in serum-containing medium. However we identified three compounds that inhibited tumor-cell attachment in a reversible fashion: (1) a specific inhibitor of collagen deposition (L-azetidine-2-carboxylic acid); (2) a bacterial neutral protease (Dispase); and (3) synthetic fibronectin peptides that contained the arginine-glycine-asparate (RGD) sequence that is responsible for cell binding. Dispase and the RGD-containing peptides also inhibited cell implantation and prevented tumor formation in a surgical wound. We propose that inhibitors of attachment might be used either alone or with other biologic modifiers to prohibit implantation of free tumor cells at the time of surgery and thus, to prevent local tumor recurrence.
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PMID:Inhibition of tumor-cell attachment to extracellular matrix as a method for preventing tumor recurrence in a surgical wound. 268 68

In Exp. 1, 5-8-cell embryos from superovulated cattle were co-cultured with oviducal tissue suspended in Ham's F10 + 10% fetal calf serum (F10FCS) or in F10FCS alone. After 4 days, the proportion of embryos developing into compact morulae or blastocysts was greater (P less than 0.005) in co-culture (38/82; 46%) than in F10FCS (1/27; 4%). In Exp. 2, a solution of collagenase, trypsin, DNAse and EDTA was used to disperse oviducal tissue, which was then cultured in TCM199 + 10% fetal calf serum (M199FCS) to obtain monolayers. Embryos (1-8 cells) were then co-cultured with monolayers or in M199FCS alone. The proportion of embryos developing into compact morulae and blastocysts after 4-5 days was higher (P less than 0.005) in co-culture (15/34; 43%) than in M199FCS (1/37; 3%); mean numbers of cells/embryo were also higher (P less than 0.001) (27.70; range 2-82 in co-culture; 8.83; range 2-18 in M199FCS). In Exp. 3, embryos obtained from in-vitro maturation and fertilization were used to compare development between co-culture and medium conditioned by oviducal tissue. Initial cleavage rate (no. embryos greater than 1 cell/total) was 76% (611/807) and did not differ among treatments. After 5 days, the proportion cleaving to greater than 16 cells was higher (P less than 0.005) in co-culture (71/203; 35%) and conditioned medium (48/205; 23%) compared to M199FCS (14/203; 7%). Similarly, the proportion developing into compact morulae and blastocysts was greater (P less than 0.005) in co-culture (44/203; 22%) and conditioned medium (46/205; 22%) than in M199FCS (7/203; 3%).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Co-culture of early cattle embryos to the blastocyst stage with oviducal tissue or in conditioned medium. 270 4

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.
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PMID:Analysis of the cell membrane proteolytic enzymes of the B16, F1, F10, and BL6 melanoma and their role in target cell destruction. 354 41

The objective of this study was to establish a method by which trophectodermal cells originating from individual preimplantation bovine embryos could be perpetuated in monolayer culture. A single, Day-11 bovine embryo collected nonsurgically from a mixed-breed beef cow was cultured in Ham's F10 medium supplemented with fetal bovine serum, sodium pyruvate, insulin, and epidermal growth factor. After 13 d in culture the embryo had adhered to the surface of the plastic culture vessel and a monolayer covering 0.3 cm2 had developed in the manner of a tissue explant. The monolayer was successfully dispersed using trypsin-EDTA and the cells were passaged. Expansion to a 25-cm2 flask was achieved by the 4th passage. By passaging cultures at a dilution ratio of 1:2, cells were maintained for 38 passages before growth slowed. Transfers beyond the 44th passage were unsuccessful. The cell line, designated BE-13, was successfully frozen and thawed at the 9th, 12th, 15th, and 20th passages. The cell line contains both mono- and binucleate cells with a prominent rough endoplasmic reticulum characteristic of ruminant trophoblast cells. Susceptibility to eight bovine viruses was demonstrated. Such cell lines may provide inexpensive systems for the study of trophoblast metabolism and for investigation of the role of the trophoblast in the pathogenesis of selected bovine abortifacient diseases. Because of their range of viral susceptibility, these cells might also be useful for diagnostic purposes.
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PMID:Monolayer culture of cells originating from a preimplantation bovine embryo. 368 Jan 2

The aggregating properties of murine melanoma cell lines with low metastatic potential (B16-F1) and high metastatic potential (B16-F10 and B16-BL6) were compared. All three types of cells were found to possess Ca2+-dependent and Ca2+-independent intrinsic mechanisms for cell adhesion, though the extent of reaggregation varied in each mechanism. After trypsin treatment at around 1 microgram/ml, F10 and BL6 cells reaggregated in the presence of 1mM Ca2+ to a greater degree than F1 cells. F10 and BL6 cells were also more aggregative than F1 cells after dissociation with collagenase. The apparent adhesiveness of the cells was found to be dependent on both the manner of cell preparation for reaggregation and on the presence of external Ca2+ or serum factors. The results are discussed in relation to the mechanisms of tumor cell arrest with emphasis on the effect of extracellular factors on cell adhesiveness.
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PMID:Adhesive properties of weakly and highly metastatic melanoma cell lines. 608 47

Changes in glycosaminoglycan composition occurring during the cell cycle were determined in B16-F10 cells sorted flow cytometrically with respect to DNA content. Incorporation of 35S-sulfate into heparan sulfate and chondroitin sulfate of unsorted and G1,S, and G2 +M sorted cells was determined following chondroitinase ABC or nitrous acid treatment; the incorporation into surface material was measured as the difference between the radioactivity of control and trypsin-treated cells. Incorporation of 35S-sulfate and 3H-glucosamine into cetyl pyridinium chloride (CPC)-precipitable material was characterized before and after chondroitinase or nitrous acid treatment by Sephadex G50 chromatography. Long-term (48 h) and short-term (1 h) labeling studies demonstrate that (a) the amount of total cellular chondroitin sulfate is greater than that of heparan sulfate, with larger amounts of unsulfated heparan than chondroitin being present; (b) the rate of turnover of heparan sulfate is greater than that of chondroitin sulfate; (c) greatest short-term incorporation of 3H-glucosamine into CPC-precipitable material occurs during S phase; and (d) the rate of turnover of both heparan sulfate and chondroitin sulfate is decreased in S phase relative to G1 and G2 + M.
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PMID:Incorporation of 35S-sulfate and 3H-glucosamine into heparan and chondroitin sulfates during the cell cycle of B16-F10 cells. 623 52

Oesophageal biopsies from endoscopically normal patients were incubated in the following for up to 30 min-Ham's F10 medium:isotonic saline:0.1 N HCl:20, 2 and 0.2 mM bile acids:trypsin:pepsin:lipase; autologous gastric and duodenal juices. The biopsies were then examined by electron microscopy. No morphological change was produced by Ham's F10 or saline. HCl caused little damage. Gastric juice produced widespread severe damage, as did pepsin. Duodenal juice and the enzymes tested caused lysis and internalisation of desmosomes and peripheral cytoplasmic vacuolation. Bile acids split desmosomes and induced microvesiculation of cell membranes. Similar microvesiculation was also induced by duodenal juice. All media except hydrochloric acid eventually produced organelle damage and leaky and disrupted cells. The functional and superficial cells appeared to be able to withstand attack from these experimental media better then did prickle and basal cells. Membrane coating granules were secreted in the presence of hydrochloric acid, but not with enzymes.
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PMID:Oesophageal epithelial ultrastructure after incubation with gastrointestinal fluids and their components. 678 15

The incorporation of [35S]sulfate and [3H] glucosamine into cetyl pyridinium chloride (CPC) precipitable glycosaminoglycans was determined in B16-F10 cultured cells sorted with respect to DNA content. Incorporation into surface material was measured indirectly as the difference between the radioactivity of control and trypsin treated cells. Approximately 80% of the total cellular [35S] sulfate labeled CPC precipitable material, but only 5% of that labeled by [3H]glucosamine, was removed by this mild trypsin treatment. Incorporation of [35S]sulfate into the trypsin removable surface material increased progressively from G1 to S to F2 + M in both long-term (48 hours) and short-term (1 hour) labeled cells, while the ratio of surface to total incorporated [35S]sulfate remained relatively constant. Incorporation of [35S]sulfate into total cellular glycosaminoglycans in long- and short-term labeled cells increased as cells progressed from G1 to S to G2 + M; the incorporation of [3H]glucosamine into CPC precipitable material also increased progressively from G1 to S to G2 + M in long-term labeled cells but was greater during S phase relative to G1 or G2 + M in short-term labeled cells. The degree of sulfation of glycosaminoglycans as represented by the ratio of [35S]sulfate to [3H]glucosamine of double labeled cells was relatively constant in long-term labeled cells but was increased during the G1 and G2 + M phases of short-term labeled cells. Comparison of the degree of sulfation of short-term with long-term labeled cells suggests that highly sulfated glycosaminoglycans may be turned over more rapidly during G1 and G2 + M phases of the cell cycle.
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PMID:Analysis of glycosaminoglycans of flow sorted cells: incorporation of [35S]sulfate and [3H]glucosamine into glycosaminoglycans of B16-F10 cells during the cell cycle. 717 38


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