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)

Fundamental examination was carried out on the liberation of single cells from hepatoma cells in culture. dRLa-74 cells derived from rat hepatoma, which hardly disperse as single cell suspensions with several proteolytic enzymes or EDTA alone, dispersed with a high yield of single cells by the combination of trypsin and EDTA. HUH-6 cells derived from human hepatoblastoma also showed similar results. The degree of cell dissociation by the combination was dependent on the incubation temperature or pH.
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PMID:Preparation of single cell suspensions from hepatoma cells in culture. 16 72

Human alpha 2-macroglobulin (alpha 2M) is a unique 720-kDa proteinase inhibitor with a broad specificity. Unlike most other proteinase inhibitors, it does not inhibit proteolytic activity by blocking the active site of the proteinase. During complex formation with a proteinase, alpha 2M entraps the proteinase molecule in a reaction that involves large conformational changes in alpha 2M. We describe the molecular cloning of alpha 2M cDNA from the human hepatoblastoma cell line HepG2. The cDNA was subcloned under control of the adenovirus major late promoter in a mammalian expression vector and introduced into the baby hamster kidney (BHK) cell line. Transformed clones were isolated and tested for production of human alpha 2M with a specific enzyme-linked immunosorbent assay. Human recombinant alpha 2M (r alpha 2M), secreted and purified from isolated transfected BHK cell lines, was structurally and functionally compared to alpha 2M purified from human serum. The results show that r alpha 2M was secreted from the BHK cells as an active proteinase-binding tetramer with functional thiol esters. Cleavage reactions of r alpha 2M with methylamine and trypsin showed that the recombinant product, which was correctly processed at the N-terminus, exhibited molecular characteristics similar to those of the human serum derived reference. Moreover, r alpha 2M-trypsin complex bound to purified human placental alpha 2M receptor with an affinity indistinguishable from that of a complex formed from serum-derived alpha 2M and trypsin.
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PMID:Expression of human alpha 2-macroglobulin cDNA in baby hamster kidney fibroblasts: secretion of high levels of active alpha 2-macroglobulin. 169 56

DNA synthesis of human hepatoblastoma HepG2 cells is reversibly inhibited by butyrate. When butyrate is removed from the culture medium, cells re-enter the cell cycle, synthesizing DNA with a time lag of about 12 h. HepG2 cells, growth-inhibited for 30 h with butyrate, synthesize and accumulate a nuclear protein, called D. Protein D synthesis is inhibited in cells which, released from the butyrate block, have resumed DNA synthesis as well as in growing cells never exposed to butyrate. Protein D has been purified from growth-arrested cells and partially sequenced. The amino acid sequences of five internal trypsin peptides indicate that protein D is a novel nuclear protein.
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PMID:Identification of a novel nuclear protein synthesized in growth-arrested human hepatoblastoma HepG2 cells. 184 69

Using the trypsin-Giemsa staining technique, cytogenetic analysis was done to determine the karyotypic characteristics in cells from the early nontumorigenic passage 12 and the late tumorigenic passage 147 of the human breast milk-derived cell line HBL-100. Acquisition of exclusive marker chromosomes in the late passage of HBL-100 may be responsible for the tumorigenic behavior of these cells in athymic nude mice.
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PMID:Cytogenetic characterization of two human milk-derived cell line (HBL-100) passages differing in tumorigenicity. 233 16

Monoclonal antibody 7B10, raised against the human breast cancer cell line T47D, identifies an antigen found in human breast carcinomas and in normal breast. Western blot and immunoprecipitation studies detected a Mr 76,000 antigen in cytosol, cell membrane, and cell culture supernatants of T47D cells. 7B10 binding to T47D cell extracts was affected by proteolytic digestion with protease type VI, trypsin, and subtilisin while it was not altered by neuraminidase digestion. Adsorption of breast cancer cell line extracts with concanavalin A reduced 7B10 immunoreactivity more than 70%. These results suggest that the antigen is a glycoprotein and that the epitope does not contain sialic acid. 7B10 was reactive with neither human milk fat globule membrane, nor skimmed milk, nor the milk-derived HBL 100 cell line. Conversely binding was detected in more than 50% of normal breast epithelial cells in culture. 7B10 immunostaining was positive on frozen sections of normal breast and nonmalignant mastopathies in 30 to 90% cells. In frozen sections of other normal tissues, 7B10 immunoreactivity was detected only in colon, apocrine glands of skin, parotid ducts, and luteal phase endometrium, confirming previous data on paraffin sections. Strong, homogeneous immunostaining was observed on frozen sections of intraductal and invasive lobular breast carcinomas (100% of cases), while more heterogeneous staining was found on invasive ductal carcinomas. Colon and rectal carcinomas, one carcinoma of the esophagus, and some cells in serous ovarian carcinomas also showed 7B10 reactivity. Immunoblotting of the 7B10-immunoreactive fraction isolated by Sepharose CL-6B chromatography of a breast carcinoma tissue sample extract identified the Mr 76,000 antigen, which was also detected in several breast cancer specimens, in colon adenocarcinomas, and in serous ovarian carcinoma fresh tumor extracts. The Mr 76,000 glycoprotein described here represents a breast cancer-associated antigen previously undescribed, mainly expressed in normal breast and breast tumors.
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PMID:Characterization and distribution in normal and tumoral human tissues of breast cancer-associated antigen defined by monoclonal antibody 7B10. 255 58

This study reports the purification and characterization of a high molecular weight human breast cancer-associated antigen identified by a previously described (1,2) murine monoclonal antibody, BCD-B4. Immunohistochemical analysis indicated that BCD-B4 recognizes an antigen expressed in an altered form on the human breast carcinoma cell line, BT-20, compared to the non-malignant human mammary epithelial cell line, HBL-100. Chemical treatments and enzymatic digestions suggested that the recognized moiety was a protein. The antigenic determinant was resistant to neuraminidase and periodate treatments but was sensitive to trypsin and proteinase K. The antigen was purified by affinity chromatography and its molecular weight, determined by SDS-PAGE analysis under non-reducing conditions, was proven to be 250 Kd. Under reducing conditions, the molecule dissociated into two polypeptides of 125 and 45 Kd, respectively. Both subunits could be isolated from normal HBL-100 and neoplastic BT-20 cellular protein extracts by affinity chromatography. The higher molecular weight subunit showed; however, qualitative and quantitative differences between the two cell lines: it was expressed in greater quantity on BT-20 cells and its molecular weight was 15 Kd higher. Both subunits could also be identified by immunoblots of BT-20 cells.
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PMID:Affinity purification of a high molecular weight human breast cancer-associated antigen identified by the BCD-B4 monoclonal antibody. 367 57

Insulin-like growth factors (IGF's) circulate in blood complexed to specific carrier proteins. The BRL 3A and BRL 3A2 rat liver cell lines secrete a 30,000-50,000 mol wt IGF carrier protein. Since the liver parenchymal cell is a likely source of IGF carrier protein synthesis, we have evaluated medium conditioned by three human hepatocellular carcinoma- or hepatoblastoma-derived cell lines for the presence of IGF carrier protein. The HEP G2 and the HEP 3B, but not the PLC/PRF/5, cell lines secrete a specific IGF carrier protein(s) into serum-free medium. This carrier protein is specific for IGF molecules. Multiplication-stimulating activity, IGF I, and IGF II were equipotent in competing for the binding of [125I]multiplication-stimulating activity to HEP G2 medium. The HEP G2 IGF carrier protein is trypsin sensitive, acid stabile, and does not contain a glycoprotein moiety. It has a molecular size of 30,000-50,000, as assessed by Sephadex G-200 gel filtration. These studies suggest that the HEP G2 cell line is a useful model to study the synthesis and secretion of human IGF carrier protein.
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PMID:Demonstration that a human hepatoma cell line produces a specific insulin-like growth factor carrier protein. 618 61

Serum-free medium conditioned by activated cells of the acute monocytic leukemia line, THP-1, was examined for growth-inhibitory activity with several established human cell lines. Free-floating clusters of THP-1 cells were activated into adherent nonproliferating cells by a 24-hr exposure to 10(-7) M mezerein in Roswell Park Memorial Institute 1640 medium containing 1% fetal bovine serum. Adherent cells were incubated for an additional 24 hr in serum-free medium containing insulin (5 micrograms/ml). Dose-response studies revealed that a cervical carcinoma (HeLa), a melanoma (A375Ag5), and several mammary carcinoma cell lines (MCF-7, BT474, MDA-MB415, and T47D) were growth inhibited by this conditioned medium. We concluded, from the results of thymidine release assays and from experiments on reversibility, that inhibition was a cytostatic and not a cytolytic response. In contrast, THP-1 conditioned medium stimulated the growth of two mammary lines (ZR75-1 and HBL-100), a lung type II carcinoma (549), and a colon adenocarcinoma (SW48). Preliminary characterization showed that the inhibitory activity was stable to acid and urea treatment but was destroyed by trypsin and sodium dodecyl sulfate. Molecular sieve chromatography of acetic acid-extracted material separated the inhibitory and stimulatory components.
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PMID:Production of growth-inhibitory activity in serum-free medium by human monocytic leukemia cells. 634 88

CD6, a type I cell surface glycoprotein expressed predominantly by thymocytes and mature T lymphocytes, becomes phosphorylated on tyrosine residues following T cell activation and has been implicated as an accessory molecule in T cell activation. The purpose of this study was to identify cell lines and tissues which express CD6 ligand(s), determine the requirements for CD6 binding, and biochemically characterize the putative CD6 ligand(s). Binding studies with a CD6 immunoglobulin fusion protein, CD6-Rg, allowed the identification of a number of human cell lines which express a CD6 ligand(s). The binding to these cell lines was trypsin sensitive, in part required divalent cations, was blocked by an anti-CD6 mAb, and could be downregulated by tumor necrosis factor alpha (TNF alpha), interleukin-1 beta (IL-1 beta) and interferon-gamma (IFN-gamma). Among the cell lines tested, the human breast carcinoma-derived cell line HBL-100 expressed the highest levels of CD6 ligand(s) and was used for immunoprecipitation studies. Following metabolic labeling, CD6-Rg immunoprecipitated glycoproteins of approximately 100, approximately 90, and approximately 45 kDa from HBL-100 cells. Using CD6-Rg we were able to show that murine thymus, lymph nodes, and skin express high levels of CD6 ligand(s) and that CD6-Rg bound to a murine thymic epithelial cell line and to cultured human epidermal keratinocytes.
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PMID:Characterization of a CD6 ligand(s) expressed on human- and murine-derived cell lines and murine lymphoid tissues. 792 88

Hepatic fibrosis often occurs in alcoholic liver diseases without accompanying tissue necrosis or inflammation. However, the precise mechanism of this fibrosis has not been fully clarified. In the present study, using the hepatoblastoma cell line HepG2 as a model for hepatocytes, we identified a factor that stimulates collagen synthesis of fibroblasts in a conditioned medium of HepG2 cells after treatment with ethanol. Type 1 procollagen peptide (PIC) in a culture of human fibroblast IMR-90 markedly increased after incubation with the conditioned medium of ethanol-treated HepG2 cells. The stimulating activity on the production of PIC by IMR-90 remained after the dialysis and evaporation of the conditioned medium of HepG2 cells, indicating this factor was not as volatile from low molecular substances such as acetaldehyde, acetate, or lactate. The activity of this factor diminished with heat or trypsin treatment. A gel chromatographic analysis disclosed that the molecular weight of this factor was approximately 8000 Da. These results suggest that a polypeptide factor secreted from HepG2 cells by treatment with ethanol stimulates collagen synthesis of fibroblasts.
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PMID:Detection of activity in the conditioned medium of ethanol-treated HepG2 cells which stimulates collagen synthesis in IMR-90 cells. 865 93


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