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:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Methods for the isolation and culture of macrophages from normal human intestine are described. After disaggregation using sequential treatments of dithiothreitol, ethylenediaminetetraacetate, collagenase, and deoxyribonuclease, Percoll density gradients (1.064 SG) produced single cell suspensions from which macrophages were readily purified by adherence to plastic. Macrophages were characterized by morphology, phagocytosis, cytoplasmic staining for nonspecific esterase, presence of membrane Fc receptors, Ia-like antigens, and lysozyme synthesis and secretion. In 10 separate experiments, recovery of viable mononuclear cells was 2.9 +/- 0.6 x 10(6) cells/g of mucosa. Thirty percent of these cells were phagocytic. After adherence to plastic, the macrophage recovery was 1.05 +/- 0.2 x 10(6) cells/g mucosa and 90% +/- 0.4% of the adherent cells in the monolayer were phagocytic. Fifty-five percent of the adherent cells showed Fc receptors for immunoglobulin G, while 94% expressed the Ia-like antigen on their membrane. The successful isolation and culture of human intestinal macrophages in large numbers will allow detailed study of their role in the mucosal immune response in health and disease.
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PMID:Isolation and preliminary characterization of human intestinal macrophages. 657 64

Advanced human periodontitis is considered to be a B-cell lesion, but the cellular infiltrate contains several cell types, the distribution of which has not been determined. This experiment was designed to characterize and identify the immunocompetent cells on histological sections and in eluates from diseased human gingiva. Immunoglobulin-bearing cells were detected on histological sections by direct immunofluorescence with F(ab')2 antisera monospecific for human immunoglobulin G (IgG), IgA, or IgM. Plasma cells predominated in the central portion of the lamina propria, with the proportions positive for IgG, IgA, and IgM accounting for 65.2 +/- 9.5, 11.2 +/- 1.1, and 1.3 +/- 1.1% of the total infiltrating cells, respectively. T lymphocytes, identified by indirect immunofluorescence with monoclonal antibody (Leu-1) against human T cells, accounted for 29.3 +/- 10.0% of the total infiltrated cells. Most of the T cells were located subjacent to the pocket epithelium, but there were a few in the central lamina propria. Similarly, Fc receptor-bearing cells detected by EA rosetting and macrophages and monocytes detected by nonspecific esterase staining with alpha-naphthylbutyrate esterase were also localized to the region immediately subjacent to the pocket epithelium. Infiltrated cells were harvested from minced gingival tissue after digestion with collagenase, hyaluronidase, and DNase. The eluates contained 35.3 +/- 6.0% T lymphocytes, 30.0 +/- 14.9% Fc receptor-bearing cells, and 12.9 +/- 4.4% monocytes and macrophages. Whereas T gamma cells comprised 13.3 +/- 1.4% of peripheral blood T cells, they accounted for only 6.0 +/- 2.0% of the eluate T cells. In contrast, T mu cells accounted for 44.7 +/- 4.9% of the T cells in the eluates and 51.6 +/- 4.4% in the peripheral blood. The decreased proportion of T gamma cells in the gingiva may indicate a form of abnormal immune regulation concerned with T suppression of B-cell proliferation.
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PMID:Identification and distribution of immunocompetent cells in inflamed gingiva of human chronic periodontitis. 660 70

A study was made of mycobacterial-induced granulomas in guinea-pig lymph nodes. Live BCG (Pasteur) induced a granuloma containing epithelioid cells while Cobalt irradiated Mycobacterium leprae induced a granuloma comprised of phagocytic macrophages. The granulomas were quantitated by measurement of lymph node weight and the areas of infiltration in histological sections. The time course of granuloma formation induced by Co-irradiated M. leprae was veary different from the time course of the granuloma formation induced by BCG. Collagen synthesis assessed by incorporation of 14C-proline into collagenase sensitive protein was greater in lymph nodes draining the site of injection of Co-irradiated BCG than those draining the site of injection of Co-irradiated M. leprae during the first 10 weeks. Collagen synthesis was delayed in the nodes from animals injected with live BCG for at least 10 weeks. Single cell suspensions of draining lymph nodes containing granulomas consisted of lymphocytes and large cells (epithelioid cells and macrophages). A high proportion of the large cells were found to be non-adherent in the live BCG-induced epithelioid cell granuloma. In contrast, M. leprae-induced granulomas contained a high percentage of adherent large cells. In both the granulomas, the majority of large cells were esterase positive and showed the presence of fibronectin. Most of the large cells in the granulomas did not carry receptors for the Fc component of IgG or the C3 component of complement and did not exhibit peroxidase activity.
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PMID:Comparison of mycobacterial granulomas in guinea-pig lymph nodes. 675 60

Fluorescent conjugates of hydroxyethyl (OEt) starch of Ficoll are selectively ingested and retained in vivo by spleen marginal-zone (MZ) and lymph node marginal-sinus macrophages of mice, whereas similar conjugates of type 3 pneumococcal capsular polysaccharide (SIII) are generally retained by macrophages (Kupffer cells, histiocytes, macrophages of spleen, lymph nodes, bone marrow and peritoneal cavity). MZ and other macrophages are readily identifiable by fluorescence after injection in vivo of OEt starch and SIII labeled with tetraethylrhodamine isothiocyanate and fluorescein isothiocyanate. Collagenase digestion was required for recovery of intact MZ macrophages from spleen in single cell suspensions and for maximum yields of other macrophages. MZ macrophages are larger and morphologically distinct from other macrophages, but resemble them in respect of EA, EAC receptors and acid phosphates and nonspecific esterase content and are equally radio-resistant. The appear normal in CBA/N nude and in beige mice. Freshly isolated MZ macrophages in suspension have adherent lymphocytes, dispersible by EDTA treatment, with B but not T cell markers. It is suggested that selective adherence to MZ macrophages is a factor in determining B cell traffic. MZ macrophages did not have demonstrable surface I-A or I-EC antigens. Only 4--8% of other spleen macrophages freshly isolated by collagenase treatment expressed I-A in the same preparation, whereas 35% of other cells (lymphocytes and blasts) reacted with monoclonal anti-I-A and anti-I-EC. After adherence to glass or plastic, 40% or more red-pulp, but not MZ macrophages, became I-A-positive. When taken from mice recently restimulated with sheep erythrocytes, half the red-pulp macrophages expressed I-A even before adherence. The relation of MZ to other macrophages is not known. However, their properties are consistent with the demonstrated ability of thymus-independent antigens selectively taken up by these cells to elicit long-lasting IgM antibody responses by direct interaction with B cells. The unexpected observation that only a small proportion of spleen macrophages freshly isolated from unstimulated mice had detectable surface I-A, but that this proportion was much increased after attachment to plastic, is discussed in relation to the possibility that macrophages do not express surface Ia antigens unless they have been stimulated.
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PMID:Different macrophage populations distinguished by means of fluorescent polysaccharides. Recognition and properties of marginal-zone macrophages. 694 Jul 55

Lymphocyte subpopulations in peripheral blood (PBL) and intestinal mucosa (IML) of 10 patients with inflammatory bowel disease (IBD) were compared with those of 11 non-IBD controls. PBL were separated on Ficoll/hypaque gradients, and IML were isolated by incubation in dithiothreitol, EDTA, and collagenase. These methods yielded cells of good viability and with intact HLA A and B-antigens. T-cells, identified by neuraminidase-treated sheep RBC rosettes and non-specific esterase staining, comprised approximately 91% of the IML from normal mucosa of all groups. B-cells, identified by erythrocyte-antibody-complement rosettes and surface immunoglobulins, were only 7% of these IML populations. Cell yields were two-fold or more greater from abnormal IBD mucosa, with T-cells ranging from 55 to 95% and B-cells from 2 to 36%. The percentage of Fc receptor bearing cells was low in all specimens. By these methods, T-lymphocytes predominated in intestinal mucosa of both IBD and non-IBD patients, but there is marked increase in the percentage of B-cells isolated from abnormal mucosa in IBD.
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PMID:Lymphocyte subpopulations of intestinal mucosa in inflammatory bowel disease. 696 6

Thymic adherent cells were isolated after their enrichment on density gradients. The predominant cell type found was the macrophage, as determined by morphology, surface receptors for Fc and C3, phagocytosis and esterase activity. There was, in addition, a minor fraction of cells with a distinctive dendritic morphology. These dendritic cells had surface properties similar to macrophages but limited phagocytic capacity. Approximately 50% of thymic adherent cells bear Ia antigens detected by immunofluorescence by using either A.TH anti-A.TL or monoclonal anti-I-A antibodies. These cells were also found to be an extremely effective antigen-presenting source for macrophage-depleted immune T cells, supporting the idea that the Ia antigens detected are of functional significance. Our data indicate that the macrophage is the predominant adherent cell type in general, and the principal Ia-bearing cell in particular, isolated by either physical disruption of the thymus or by collagenase dissociation of thymis stroma.
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PMID:IA antigens and antigen-presenting function of thymic macrophages. 735 86

Differentiated mammalian cell lines can be established by introducing viral oncogenes into primary cells. Such lines can retain their original specialised functions while being adapted to prolonged life in culture; but most transformed cell lines obtained in this way characteristically show altered properties compared with the primary cells. The result of these changes is that transformed cell lines no longer provide a good model of the original tissue, and indeed often resemble other transformed lines more than the initial cell type. In our laboratory three murine peritoneal macrophage-like cell lines have been isolated by transforming primary cells with SV40 origin-deleted DNA. These lines have been in continuous culture for approximately 1 year and have been shown to express many macrophage-specific properties throughout this time, including Fc receptors and staining for non-specific esterase. The cell lines phagocytosed IgG-coated particles, they were positive for the murine macrophage-specific marker F4/80 and they showed antigen-presentation function. Lysozyme, acid phosphatase, plasminogen activator, collagenase, prostaglandin E2 and 5'-nucleotidase activities have also been detected in these lines. In this paper the method of DNA transformation will be described as well as some of the assays used for the characterization of the three immortalized cell lines.
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PMID:Establishment and characterization of murine macrophage-like cell lines following transformation with simian virus 40 DNA deleted at the origin of replication. 808 36

The time course of inflammatory cell infiltration into guinea pig lungs following a single 4 h exposure to 2 ppm O3 was established by measuring the changing cell populations recovered by both bronchoalveolar lavage (BAL) and collagenase tissue digestion. Analysis of BAL-recovered albumin was used as an indicator of permeability damage and demonstrated an increase immediately following ozone exposure, reaching a maximum within 24 h, but returning to air-control levels by 7 days post-ozone exposure. A twofold enhancement in macrophages was observed in the lavage-recovered cell population after 2 days, returning to air-control numbers by 7 days. Collagenase digest-recovered monocytes and macrophages, identified by nonspecific esterase staining, were found to be elevated between 2 and 14 days following O3 exposure. Immediately following O3 exposure, a 4.5-fold increase in collagenase digest-recovered neutrophils was observed, with a subsequent decline to air-exposed lung levels during the next 12 h. In contrast, BAL-recovered neutrophils were observed to be increased immediately following O3 exposure at a level that was sustained for up to 3 days. The tissue accumulation of neutrophils was not associated with their subsequent appearance in the lavageable spaces. Although significant increases in collagenase digest-recovered eosinophils could not be detected, lavage-recovered eosinophil numbers were transiently increased by threefold after 3 days. By employing both BAL and collagenase digestion to evaluate this model of reversible lung injury, this study demonstrated that the use of BAL-recovered cell measurements alone does not adequately reflect the early inflammatory cell changes taking place within oxidant-exposed lungs.
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PMID:Guinea pig lung inflammatory cell changes following acute ozone exposure. 820 31

The contribution of interstitial macrophages (IM) to lung defense, homeostasis, and pathophysiology is for the most part unknown. Studies on this cell type are difficult because they are not readily accessible in large numbers or in high purity. In the present work, various nonenzymatic and enzymatic methods were compared with the aim of isolating and characterizing pure populations of lung IM. The results of our studies demonstrate that most procedures currently used to isolate IM yield subpopulations of alveolar macrophages (AM) or IM highly contaminated by AM and granulocytes. We found that lavage of the lung yielded only one half of the total AM present in the tissue. The remainder of the AM could only be obtained by extensive washing of cut and disaggregated lung tissue, which is considered by some investigators to be an effective procedure for IM isolation. According to our results, cells recovered by lavage and washing of cut and disaggregated lung tissue were morphologically and histochemically identical, were strongly positive for nonspecific esterase, highly phagocytic, and appeared to represent subpopulations of AM with apparently varying degrees of adherence to the alveolar walls. We also found that IM could be obtained in high purity by sequential digestion of the remaining lung tissue with 60 and 175 IU/ml of collagenase followed by selective adherence. Digestion of the tissue with 60 IU/ml of collagenase resulted in a highly enriched population of granulocytes and also reduced their contamination in the IM population. The resulting IM were distinct from AM by morphology and histochemistry. Like AM, these cells displayed Fc receptor-mediated phagocytosis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Isolation and partial characterization of subpopulations of alveolar macrophages, granulocytes, and highly enriched interstitial macrophages from rat lung. 847 32

The identification and purification of human osteoclast precursors is essential to further our understanding of the mechanisms that control human osteoclast differentiation. Osteoclastoma tissue potentially provides a rich source of human osteoclast precursors, and in previous studies we have demonstrated the existence of a population of mononuclear cells within this tissue that is reactive with osteoclast-selective vitronectin receptor monoclonal antibodies. In this study, mononuclear cells expressing the vitronectin receptor, as defined by their ability to react with a murine monoclonal antibody to the beta 3 chain of the vitronectin receptor (87MEM1), were isolated from collagenase digests of osteoclastoma tissue using a fluorescence activated cell sorter. Based on their fluorescence signal and size, approximately 2-3% of the viable cells (typically 2 x 10(5)) were obtained and prepared for further phenotyping. The isolated cells demonstrated a number of phenotypic characteristics of osteoclasts: positive tartrate-resistant acid phosphatase (TRAP) activity, reactivity with human osteoclast-selective antibodies, expression of calcitonin receptors, cathepsin K (a novel osteoclast-selective cysteine proteinase) mRNA, and osteopontin mRNA and protein. These phenotypic characteristics were also detected in mononuclear cells within cryostat sections of the native osteoclastoma tissue as well as in resorption lacunae of sections of human bone. In contrast, isolated peripheral blood monocytes were negative for TRAP activity and osteopontin expression and, unlike the osteoclastoma-derived cells, demonstrated strong nonspecific esterase activity. Significantly, when the osteoclastoma-derived 87MEM1 positive cells were cocultured on whale dentine for 1-3 weeks with stromal cells, extensive resorption of the dentine surface was observed. This is the first demonstration of the purification of human osteoclast precursors. These cells provide an homogeneous cell population for studying cellular events that occur during human osteoclast differentiation.
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PMID:Purification and characterization of fully functional human osteoclast precursors. 891 68


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