PubMed Health⌕ Search

Biomedical subjects

H F Upchurch

Publications and source records attributed to H F Upchurch.

At least 19 recordsLinked to original sources
PubMed1991

Localization of cellular transglutaminase on the extracellular matrix after wounding: characteristics of the matrix bound enzyme.

Extending our previous observation that tissue transglutaminase (TGase) binds to extracellular matrix (ECM) fibronectin, we report here that endogenous tissue TGase is localized on the adjacent ECM after puncture wounding embryonic human lung fibroblasts (WI-38). The bound TGase persisted at the wound site for many hours, demonstrated by immunofluorescence and by catalytic activity using an overlay assay. The binding characteristics of TGase with ECM were studied further by the addition of exogenous TGase to cell monolayers and monitoring by immunofluorescence or overlay catalytic activity assays. Binding occurred equally well at 4 degrees C or 37 degrees C. Prior incubation of exogenous TGase with guanosine 5'-triphosphate (GTP), guanosine 5'-diphosphate (GDP), or adenosine triphosphate (ATP) had little effect on the amount bound to matrix, but prior treatment with calcium, magnesium, strontium, or manganese ions enhanced binding 2- to 3-fold. The Ca(++)-dependent change was a concentration-dependent effect on soluble exogenous TGase, rather than an effect on ECM. Immunofluorescent techniques showed that binding of exogenous TGase to ECM was prevented by prior mixing with fibronectin or collagen, but not with several other ECM components, including laminin, elastin, chondroitin sulfate, heparan sulfate, and hyaluronic acid. ECM-bound TGase was released by 2 M potassium thiocyanate (KSCN) treatment but was not released by treatment with a variety of amino acids, salts, reducing agents, glycerol, or other chaotropic agents.

Adenosine Triphosphate↗
PubMed1987

Cellular transglutaminase has affinity for extracellular matrix.

Cellular transglutaminase (TGase) was demonstrated as an intracellular enzyme by immunofluorescence in WI-38 cells. Following cell membrane perturbation by Triton X-100 treatment, TGase was bound to the extracellular matrix and was found to coexist with fibronectin as visualized by immunofluorescence microscopy. The binding of TGase to the cell matrix was blocked by anti-fibronectin antibody. Exogenous sources of soluble TGase were transferred to the extracellular matrix of an untreated or methanol fixed cell. The experimental data indicated that "particulate bound" TGase is a consequence of soluble TGase binding to the extracellular matrix following cell rupture.

Extracellular Matrix↗
PubMed1985

A monoclonal antibody to cellular transglutaminase.

A cellular enzyme-linked immunosorbent assay was developed for estimating cellular transglutaminase in situ using a monoclonal antibody produced to tissue transglutaminase. The minimum level of detection of TGase was 2-5 ng. The enzyme was present in greater amounts in WI-38 and IMR90 cells than in their simian virus-transformed counterparts. The levels of TGase in the virus-transformed cells increased significantly when the cells were grown in the presence of sodium butyrate to induce enzyme activity. Staining of confluent WI-38 cells by indirect immunofluorescence using the monoclonal antibody showed microscopic fibers suggesting that the enzyme may be associated with detergent-insoluble components.

Acyltransferases↗
PubMed1983

Further comparisons of endogenous pyrogens and leukocytic endogenous mediators.

It was recently shown (Murphy et al., Infect. Immun. 34:177-183), that rabbit macrophages produce two biochemically and immunologically distinct endogenous pyrogens. One of these has or copurifies with substances having a molecular weight of 13,000 and a pI of 7.3. This protein was produced by blood monocytes or inflammatory cells elicited in 16-h rabbit peritoneal exudates. These acute peritoneal exudates were produced by the intraperitoneal injection of large volumes of saline containing shellfish glycogen. When the leukocytes in these exudates were washed and incubated at 37 degrees C in saline, they released an endogenous pyrogen. The injection of this pyrogen into rabbits, rats, or mice caused the biological manifestations which have been attributed to leukocytic endogenous mediator. These effects were increases in blood neutrophils, the lowering of plasma iron and zinc levels, and the increased synthesis of the acute-phase proteins. The other rabbit endogenous pyrogen seems to be a family of proteins with isoelectric points between 4.5 and 5.0. These proteins are produced by macrophages in the lung, liver, or in chronic peritoneal exudates. In these experiments, the lower-isoelectric-point endogenous pyrogens were produced by macrophages from the peritoneal cavity of rabbits that had been injected 4 days earlier with 50 ml of light mineral oil. These rabbit pyrogens were found to have leukocytic endogenous mediator activity in mice but to be completely inactive in rats. When injected into rabbits, these proteins produced fever, lowered plasma iron, increased blood neutrophils, but failed to elevate plasma fibrinogen.

Animals↗
PubMed1982

Characterization of a leukocyte-derived endogenous mediator responsible for increased plasma fibrinogen.

Fibrinogen has been the plasma protein most frequently studied after tissue injury. This report presents evidence that leukocytic endogenous mediator (LEM) from macrophages promotes fibrinogen synthesis. LEM has a molecular weight of 13,000-16,000, an isoelectric point (pI) at pH 7.3, is heat labile, and is inactivated by trypsin or sulfhydryl reactive agents. LEM not only promotes increased synthesis of acute phase proteins, but also causes increased neutrophilia and alterations in metal metabolism. There is considerable evidence that LEM may be the same protein as endogenous pyrogen and Interleukin 1 (IL-1). There was no increase in plasma fibrinogen when endotoxin was injected in C3H/HeJ mice; however, this strain of mice responded the same as normal mice to injections of LEM. This provides further evidence that LEM is the endogenous mediator for acute phase protein synthesis during tissue injury. The half-life of LEM is still circulation following its iv injection into rats was less than 10 minutes. There is still considerable doubt about the mechanism LEM uses in promoting increased hepatocyte synthesis of fibrinogen. Some evidence indicates a direct action of LEM upon the hepatocyte, whereas other data suggest an indirect role through other mediators or the central nervous system. In addition to LEM with pI of 7.3, there are proteins with a pI near 5 that will increase plasma fibrinogen. These proteins also have a molecular weight between 13,000 to 16,000 but do not have essential sulfhydryl groups. These proteins also have pyrogenic and IL-1 activities. LEM shows a limited amount of species specificity. For example, the pI 7 LEM prepared from human monocytes or rabbit peritoneal leukocytes will increase plasma fibrinogen in rats, mice, and rabbits; but the pI 5 LEM from rabbits is inactive in rats.

Animals↗
PubMed1980

The activity of partially purified leukocytic endogenous mediator in endotoxin-resistant C3H/HeJ mice.

Partially purified LEM was injected into both the endotoxin-resistant C3H/HeJ and the closely related, but endotoxin-responsive, C3Heb/FeJ strains of mice. Both strains of mice responded to LEM with significant decreases in plasma iron concentration, increased number of peripheral blood neutrophils, increased plasma levels of CSA, and elevations of plasma fibrinogen concentration. Injections of endotoxin produced significant responses in these four determinations in C3Heb/FeJ mice but had no effect in the C3H/HeJ strain. Changes in plasma iron, fibrinogen, CSA, and neutrophils are believed to occur when LEM is produced by an interaction of endotoxin with phagocytic cells.

Animals↗