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Biomedical subjects

T M Saba

Publications and source records attributed to T M Saba.

At least 19 recordsLinked to original sources
PubMed1992

Liver and spleen phagocytic depression after peripheral ischemia and reperfusion.

Liver and spleen phagocytic clearance of blood-borne microparticulate tissue debris and products of intravascular coagulation after trauma and surgical injury is an important mechanism to limit the deposition of debris in the pulmonary vascular bed. Plasma fibronectin (pFn) modulates this clearance process. We evaluated the effect of a localized peripheral ischemia and reperfusion injury on liver and spleen phagocytic function. Male rats (250 to 350 g) underwent 4 hours of tourniquet-induced bilateral hindlimb ischemia, followed by 18 hours of reperfusion after release of the tourniquet. Rats subjected to ether anesthesia alone or anesthesia followed by groin incision without ischemia were the control and sham groups, respectively. Reticuloendothelial (RE) phagocytic function was assessed at 15 minutes and 18 hours after the start of reperfusion by the in vivo liver and spleen removal of blood-borne iodine 125 (125I)-test microparticles, which were coated with gelatin (denatured collagen) to enhance their interaction with pFn. Liver and spleen particle uptake in control and sham rats was similar. In contrast, after 4 hours of ischemic injury with 15 minutes of reperfusion, we observed a 30% to 40% decrease (p less than 0.05) in liver and spleen particle uptake as compared with sham controls with partial restoration of this removal mechanism by 18 hours. This depression in liver and spleen phagocytic function was associated with a significant (p less than 0.05) increase in the deposition of the 125I-test particles in the lung. RE depression was not due to a deficiency of pFn; indeed, a marked elevation (588 +/- 12 micrograms/mL versus 1,083 +/- 40 micrograms/mL) of pFn was observed by immunoassay over the 18-hour reperfusion interval. Comparative bioassay of humoral (opsonic) versus cellular (Kupffer's cell) activity revealed that Kupffer's cells in livers from controls or ischemia-reperfusion rats exhibited normal phagocytic function when incubated in plasma harvested from either control or 4-hour ischemic rats. The opsonic activity of plasma harvested after ischemia and reperfusion was also more than adequate, consistent with the immunoassay analysis. Thus, the impaired liver and spleen clearance mechanism after peripheral ischemia and reperfusion injury did not appear to be due to either a macrophage cellular deficit or a lack of pFn. This clearance depression may be mediated by splanchnic malperfusion, which is known to develop after peripheral ischemia and reperfusion and associated soft tissue injury.

Animals↗
PubMed1992

Rebound elevation of fibronectin after tissue injury and ischemia: role of fibronectin synthesis.

Plasma fibronectin (pFn) stimulates macrophage phagocytosis of tissue debris; pFn deposition in tissues may influence vascular integrity. Although the acute depletion of pFn after surgery and/or injury has been described, less attention has been given to the rebound hyperfibronectinemia presumably "triggered" by the early pFn depletion. Using a model that compartmentalized the site of tissue injury and thus attenuated the initial pFn depletion, we studied this rebound elevation of pFn in anesthetized rats (250-350 g) after the surgical trauma of groin dissection alone (sham group) or surgery coupled with 4 h of hindlimb ischemia (experimental group). Nonoperated control rats were also anesthetized. Shams had baseline (preoperative) 6-, 8-, and 22-h postoperative pFn levels of 573 +/- 61, 598 +/- 62, 695 +/- 57, and 929 +/- 87 micrograms/ml, respectively. In the surgery-ischemia group, pFn also elevated to 1,117 +/- 40 micrograms/ml at 22 h postsurgery. Nonoperated control rats (only anesthetized) had no elevation of pFn. Intravenous infusion of gelatin-coated lipid particles (50 mg/100 g) depleted pFn by 89.3% but was unable to prevent the rebound elevation of pFn. The blood clearance of 125I-labeled pFn was very similar in control, sham, and experimental rats. In contrast, pFn synthesis over the 22-h period was dramatically altered and equal to 2.12 +/- 0.16, 3.40 +/- 0.56, and 4.49 +/- 0.17 mg pFn synthesized/100 g body wt, in control, sham, and experimental rats respectively. Thus a rapid increase in pFn synthesis contributes to the rebound hyperfibronectinemia after sublethal surgical injury.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Simultaneous measurement of fluid and protein permeability in isolated rabbit lungs during edema.

Fluid conductance and protein permeability have been studied in isolated perfused lung models of pulmonary edema. However, previous studies have not investigated changes of both fluid conductance and protein permeability in the same isolated lung preparation after injury. Arachidonic acid (AA) metabolites are involved in the inflammatory processes that lead to the development of pulmonary edema. The hemodynamic effects of AA have been well established; however, controversy exists concerning the ability of AA to alter the permeability of the pulmonary microvasculature to fluid and protein. The purpose of this study was to simultaneously determine whether transvascular fluid conductance and protein permeability are increased in isolated perfused rabbit lungs with pulmonary edema induced by AA. Indomethacin (80 microM) was added to the perfusate to inhibit the hemodynamic effects of AA and produce a pressure-independent model of pulmonary edema. Fluid conductance was assessed by determination of the capillary filtration coefficient (Kf), and protein permeability was evaluated by measurement of 125I-albumin clearance. The injection of AA (3 mg/200 ml of perfusate) into the pulmonary arterial catheter resulted in an increase in lung weight over the remaining 30-min experimental period. Kf (microliter.s-1 x cmH2O-1 x g dry lung-1) was increased (P < 0.05) in AA-treated lungs at 10 and 30 min post-AA injection when compared with control lungs and baseline values (determined 10 min before AA injection). Albumin clearance was also greater (P < 0.05) in lungs that received AA. 125I-albumin clearance was measured at different rates of fluid flux produced by elevation of venous pressure.(ABSTRACT TRUNCATED AT 250 WORDS)

Albumins↗
PubMed1991

Blood-borne fragments of fibronectin after thermal injury.

Fibronectin is an adhesive protein that can promote phagocytosis and endothelial cell adhesion. Plasma fibronectin declines following burn in animals and patients, potentially due to its complexing with circulating collagenous debris as well as its rapid binding to sites of tissue injury. Such depletion of fibronectin initiates an opsonic deficiency of the plasma. In view of the sensitivity of fibronectin to proteolytic enzymes, an additional factor that could contribute to the decrease of plasma opsonic activity after burn is the proteolytic fragmentation of fibronectin in the blood. In the current study, we determined if fibronectin fragments appear in the blood of anesthetized rats after a sublethal full-thickness skin burn of 15% to 16% of body surface. Plasma fibronectin concentration was quantified by enzyme-linked immunosorbent assay and the presence of fibronectin fragments in plasma was determined by immunoblot analysis. All blood was collected in an antiprotease mixture to yield final plasma concentrations of 0.15% EDTA, 3mmol/L phenylmethylsulfonyl fluoride, and 3 mmol/L iodoacetate to prevent degradation of fibronectin after sampling. Plasma fibronectin decreased 60% to 70% within 30 minutes post-burn, and this low level lasted for at least 4 hours. Within 30 minutes post-burn, two prominent fragments of fibronectin with a molecular weight of 110 +/- 2.2 kd and 122 +/- 3.3 Kd, respectively, were also detected in the plasma. Peak concentration of these fragments was detected at 60 minutes post-burn, but their level declined by 4 hours. By 4 hours, both bands appeared to resolve into doublets. To rule out the possibility that the fragments of fibronectin detected in the plasma were actually generated by coagulation enzymes activated at the site of peripheral blood sampling, rapid direct inferior vena cava sampling was performed, which also yield the presence of the fragments. Thus, fibronectin fragments exist in the plasma following thermal injury. Because fragments of fibronectin can compete with the intact fibronectin molecule with respect to its ability to stimulate macrophage phagocytosis, such fragments may contribute to altered systemic phagocytic host defense following thermal injury. Furthermore, because fibronectin peptides can compete with matrix fibronectin and impair adhesion of cultured endothelial cells, such circulating fragments may also influence the integrity of the vascular barrier.

Animals↗
PubMed1991

Proteolysis of gelatin-bound fibronectin by activated leukocytes: a role for leukocyte elastase.

Fragmentation of subendothelial matrix-bound fibronectin by proteases released from stimulated leukocytes has been implicated in lung vascular injury. We studied the degradation of fibronectin bound to denatured collagen by inflammatory polymorphonuclear leukocytes (PMNL). Tissue culture wells coated with denatured collagen (gelatin) were pretreated with 125I rat plasma fibronectin to allow for fibronectin binding prior to the addition of rat inflammatory PMNL. The release of both intact and fragmented fibronectin from the 125I-labelled artificial matrix was quantified following the addition of PMNL stimulated by the phagocytosis of opsonized zymosan as well as leukocyte elastase. Stimulated PMNL released three times more radiolabelled fibronectin from the denatured collagen surface during a 4 h incubation as compared with unstimulated PMNL. This pattern of 125I-fibronectin release could also be elicited by the addition of purified leukocyte elastase alone, in the absence of PMNL. The release of radiolabelled fibronectin by stimulated PMNL was blocked in a dose-dependent manner by the addition of both methoxysuccinyl-alanine-alanine-valine chloromethyl ketone (AAPVCK), a leukocyte elastase specific inhibitor as well as phenylmethylsulfonylfluoride (PMSF), a non-specific serine protease inhibitor. Western blot analysis coupled with autoradiography confirmed the presence of fibronectin fragments in the medium after addition of PMNL or leukocyte elastase. The large molecular weight fragments (60-200 kD) were not labelled, but the smaller molecular weight fragments (less than 45 kD), derived from the artificial matrix, were labelled. Thus, fibronectin complexed with denatured collagen is susceptible to proteolytic degradation by stimulated inflammatory PMNL. Such a process may have a role in the pathogenesis of acute vascular injury following microvascular margination of activated blood leukocytes.

Amino Acid Chloromethyl Ketones↗
PubMed1991

Leukocyte elastase-independent proteolysis of gelatin-bound fibronectin by inflammatory macrophages.

Fragmentation of lung matrix fibronectin by proteases released from activated phagocytic cells has been implicated in lung vascular injury. We examined whether denatured collagen (gelatin)-bound fibronectin can be degraded by peritoneal exudate mononuclear phagocytes harvested from rats 96 h after intraperitoneal casein injection. Microtiter plates were pretreated with gelatin and then supplemented with purified 125I rat plasma fibronectin, which readily bound to the gelatin. Stimulated inflammatory exudate cells were added and proteolysis of the bound fibronectin was studied by the release of [125I]fibronectin fragments into the media. Following 2 h of incubation, peritoneal exudate mononuclear macrophages stimulated with opsonized zymosan released three times more radiolabeled fibronectin into the medium as compared to background controls, and 1.5 times more radiolabeled fibronectin as compared to cells not stimulated with zymosan. Western blot analysis and autoradiography confirmed the presence of fragments of fibronectin in the culture medium. Some of these fragments were clearly derived from the radiolabeled matrix, but others that were not labeled were potentially released directly from the added stimulated macrophages. The release of radiolabeled fibronectin was inhibited by N-p-tosyl-L-lysine chloromethyl ketone (TLCK), a trypsin specific inhibitor, but not by methoxysuccinyl-alanine-alanine-proline-valine-chloromethyl-ketone (AAPVCK), a leukocyte elastase-specific inhibitor. These results suggest that fibronectin bound to denatured collagen is susceptible to leukocyte elastase-independent enzymatic degradation by stimulated inflammatory exudate mononuclear phagocytic cells. Such proteolysis may mimic a pathological process associated with lung vascular injury during the sequestration of activated macrophages in the lung microcirculation and interstitium.

Animals↗
PubMed1991

Incorporation of circulating fibronectin into various tissues during sepsis: colocalization with endogenous tissue fibronectin.

We studied the plasma clearance and tissue incorporation of intravenously infused purified human plasma fibronectin into various tissues during a period of acute lung vascular injury induced by lethal postoperative bacteremia in sheep. Lung, liver, spleen, and heart tissue were examined for both endogenous sheep tissue fibronectin as well as the experimentally infused human fibronectin using dual-label immunofluorescence. Awake sheep (n = 4) received a postoperative iv infusion of 5 x 10(9) live Pseudomonas over a 60-min infusion interval. Bacterial challenge was started 2 hr after starting the iv fibronectin infusion of purified human plasma fibronectin (100 mg iv bolus; 4 hr iv at 100 mg/hr). Human fibronectin displayed a biphasic rate of clearance from the plasma with entrance into lymph. Human fibronectin readily incorporated in all tissues studied, including the lung which was the focus of vascular injury. Analysis of tissue sections by dual-label immunofluorescence indicated that the exogenous human fibronectin colocalized with the endogenous sheep fibronectin. Thus, the plasma fibronectin concentration may influence the lung vascular barrier due to its incorporation into the tissue pool of fibronectin. Moreover, the plasma may serve as a reservoir for soluble fibronectin which can enter and colocalize with the insoluble tissue pool of fibronectin in various tissues.

Animals↗
PubMed1991

Kinetics of plasma fibronectin: increased lung tissue incorporation after postoperative bacteremia.

Fibronectin is an adhesive glycoprotein that may influence the permeability of the vascular barrier. We compared the plasma disappearance of purified human fibronectin (hFn) and its incorporation into lung tissue in control nonbacteremic and bacteremic sheep after surgery to determine the influence of postoperative bacteremia on the plasma clearance and lung deposition of hFn. Lymph fistulas were surgically prepared 48 h before either saline or bacterial challenge to allow for collection of timed samples of plasma, lung lymph, and peripheral lymph. On the day of the study, the sheep were anesthetized and given either 5 X 10(8) Pseudomonas aeruginosa in saline or saline alone. In parallel, they were infused intravenously with 100 mg of hFn, and the hFn concentration in plasma, lymph, and tissue extracts was subsequently determined by enzyme-linked immunosorbent assay over an 8-h experimental interval. Localization of endogenous sheep fibronectin (sFn) and the injected hFn in serial lung tissue samples was determined by dual-label immunofluorescence. In nonbacteremic control sheep, plasma hFn levels declined with an initial rapid slope (t1/2 = 0.53 +/- 0.19 min), followed by a second, gradual slope (t1/2 = 21.7 +/- 2.5 h), whereas the concentration of hFn in lung lymph and peripheral lymph rose exponentially with time. In bacteremic sheep, the early plasma disappearance of hFn was similar, but the second phase of plasma clearance was faster (t1/2 = 7.4 +/- 0.3 h). Lung tissue from control and bacteremic sheep contained the same level of extractable hFn. However, tissue extraction followed by immunofluorescence microscopy indicated that lung tissue from bacteremic sheep contained more nonextractable hFn than lung tissue from nonbacteremic sheep. Thus postoperative bacteremia that elicits acute lung vascular injury will increase the plasma disappearance of hFn and its incorporation into the lung tissue.

Animals↗
PubMed1991

Fibronectin fragments released from phorbol ester-stimulated pulmonary artery endothelial cell monolayers promote neutrophil chemotaxis.

We have recently shown that monolayer cultures of calf pulmonary artery endothelial (CPAE) cells pretreated with phorbol myristate acetate (PMA) generate a conditioned medium that is chemotactic for human polymorphonuclear leucocytes (PMNL). Fibronectin (Fn) is a multidomain protein found in the plasma and subendothelial extracellular matrix that induces attachment and migration of a variety of cell types. The present study was designed to evaluate the role of Fn or fragments of Fn present in conditioned medium from phorbol ester-stimulated endothelial cells as potential chemotactic factors for human PMNL. A large number of Fn fragments were revealed by Western immunoblotting of serum-free conditioned medium 4 hr after treatment of CPAE monolayers with PMA. Gelatin-Sepharose affinity chromatography of 4-hr conditioned medium demonstrated chemotactic activity for PMNL in both gelatin-binding and non-gelatin-binding fractions. The addition of bovine Fn antiserum to the conditioned medium inhibited PMNL chemotaxis in a dose-dependent manner while having no effect on PMNL chemotaxis generated by zymosan-activated serum. One site on the Fn molecule known to interact with phagocytic cells is the cell-binding domain containing the Arg-Gly-Asp (RGD) sequence. Pretreatment of PMNL with a RGD-containing peptide (1 mM GRGDSPK) for 10 min completely inhibited the expression of chemotactic activity present in conditioned medium and in the gelatin-binding and non-gelatin-binding fractions. PMNL chemotaxis was not stimulated by either intact Fn or the RGD-containing septapeptide tested over a wide concentration range. However, incubation of PMNL with a purified 120,000-MW fragment of Fn containing the cell-binding domain stimulated chemotaxis in a dose-dependent manner. In contrast, a purified 45,000 MW fragment of Fn containing the gelatin-binding domain was not chemotactic for PMNL. When a monoclonal antibody directed against the cell-binding domain of Fn was incubated with conditioned medium, a significant reduction in PMNL chemotaxis was observed. These results demonstrate that phorbol ester-stimulated pulmonary artery endothelial cells release Fn fragments and suggest an important role for Fn fragments containing the cell-binding domain in stimulating the migration of PMNL.

Amino Acid Sequence↗
PubMed1990

Blood-borne collagenous debris complexes with plasma fibronectin after thermal injury.

Plasma fibronectin augments the clearance of blood-borne foreign and effete complexes by mononuclear phagocytes. The release of a "gelatin-like" ligand into plasma after thermal injury has been reported. We quantified the release of this collagenous debris from thermally injured skin, and its potential interaction with soluble fibronectin in plasma using anesthetized rats. Collagen-like material debris in the plasma was detected by assay of hydroxyproline. Fibronectin was measured by a double antibody enzyme-linked immunosorbent assay (ELISA) technique. Over a 24-hour postburn interval, plasma hydroxyproline increased from 6.7 +/- 0.6 micrograms/mL to a maximum of 19.0 +/- 3.3 micrograms/mL at 60 minutes postburn, and normalized by 6 hours. A direct correlation existed between the magnitude of burn injury and the increase in plasma hydroxyproline. In parallel, plasma fibronectin declined over a 15-minute to 2-hour period postburn, and normalized by 3 to 4 hours with rebound hyperfibronectinemia observed at 24 hours. The elevation in total plasma hydroxyproline was not due to an increase in plasma Clq (zero time, 26.2 +/- 1.4 micrograms/mL; 60 minutes, 23.9 +/- 1.1 micrograms/mL). Tracer studies with 125I-fibronectin showed that the acute decline of plasma fibronectin was due to its uptake by the liver and binding to sites of tissue injury. Total hydroxyproline in extracts of burn skin, used as an index of soluble collagenous material, rose from 15 +/- 3.3 micrograms/g skin at zero time to 129.3 +/- 43.7 micrograms/g skin by 5 minutes postburn, with a decline to 38 +/- 22 micrograms/g skin by 24 hours. The formation of circulating fibronectin-gelatin complexes in vivo was documented by cross-immunoelectrophoresis coupled with autoradiography using 125I-gelatin as a model ligand. Thus, collagenous tissue debris from burned skin may enter the plasma after thermal injury and directly complexes with soluble fibronectin before hepatic phagocytic clearance.

Animals↗
PubMed1990

High-affinity binding of fibronectin to cultured Kupffer cells.

Hepatic Kupffer cells are a major component of the reticuloendothelial or macrophage system. They were the first phagocytic cell type whose phagocytosis was shown to be influenced by plasma fibronectin, a dimeric opsonic glycoprotein. In the current study, the binding of soluble radioiodinated fibronectin purified from rat serum to isolated rat hepatic Kupffer cells was investigated using a cultured Kupffer cell monolayer technique. Binding was specific, since unlabeled purified fibronectin competed in a dose-dependent manner with the 125I-fibronectin for binding to the Kupffer cells. Addition of gelatin enhanced the binding of 125I-fibronectin to Kupffer cells. The phagocytosis of gelatinized-coated red cells by Kupffer cells was increased either by preopsonizing the target particles with purified fibronectin or by the addition of purified fibronectin to the culture medium. In contrast, exposure of the Kupffer cells to medium containing purified fibronectin followed by wash-removal of the fibronectin did not increase the uptake of gelatin-coated red blood cells, even though fibronectin was detected on the surface of the Kupffer cells by immunofluorescence. Trypsinized monolayers expressed decreased capacity to bind 125I-fibronectin as well as fibronectin-coated sheep erythrocytes. The binding of 125I-fibronectin-gelatin complexes was inhibited by excess unlabeled fibronectin. We calculated that specific high-affinity (Kd = 7.46 x 10(-9) M) binding sites for fibronectin exist on Kupffer cells. There are approximately 2,800-3,500 binding sites or putative "fibronectin receptors" per Kupffer cell. These sites appear to mediate the enhanced phagocytosis of gelatin-coated particles opsonized by fibronectin.

Animals↗
PubMed1990

Plasma fibronectin levels during cardiopulmonary bypass.

Plasma fibronectin, also called cold-insoluble globulin, is a cryoprecipitable glycoprotein with both opsonic and adhesive activities. It binds to collagen, actin, and heparin and can form soluble as well as cryoprecipitable complexes in the cold. Fibronectin augments particulate phagocytosis by the reticuloendothelial system and can influence lung vascular permeability. Plasma fibronectin deficiency is temporally associated with respiratory failure in septic surgical, trauma, and burn patients. We measured plasma fibronectin and albumin levels in nine adults undergoing elective cardiopulmonary bypass to determine whether dilution alone could account for the changes in plasma fibronectin. Plasma fibronectin concentration decreased 17% with the surgical trauma of opening of the chest and placement of the vascular cannulas. On heparinization and initiation of cardiopulmonary bypass, plasma fibronectin fell an additional 48% (P less than 0.001), whereas albumin concentration (corrected for albumin in the pump prime) fell only 25% (P less than 0.001), emphasizing that dilution was not the only mechanism contributing to the decline in plasma fibronectin. Fibronectin levels began to increase after discontinuation of cardiopulmonary bypass and in association with diuresis, but unexpectedly they remained subnormal until 4 days postoperation. Thus the decline in fibronectin concentration with cardiopulmonary bypass may be due to dilution as well as opsonic consumption and possible complexing with heparin in the cold.

Adult↗
PubMed1989

Effect of repetitive low-dose endotoxin on liver parenchymal and Kupffer cell fibronectin release.

Repetitive low-dose endotoxin, at a dose which will result in endotoxin tolerance, produces a marked but transient 2- to 3-day increase in plasma fibronectin. This elevation of fibronectin appears to contribute to increased hepatic Kupffer cell phagocytic function observed with repetitive low-dose endotoxin administration. Although numerous cell types synthesize fibronectin, hepatocytes are believed to be the major cell source of fibronectin in the plasma. Since Kupffer cells also synthesize fibronectin, we sought to determine the relative contribution of hepatic Kupffer cells, as compared to parenchymal cells, to the elevation of plasma fibronectin following repetitive low-dose endotoxin administration. Kupffer cells isolated from rats previously treated for 3 consecutive days with 100 micrograms Salmonella enteritidis endotoxin released greater (p less than 0.01) amounts of fibronectin over time in culture (3, 6, 12 and 24 hr) as compared to Kupffer cells isolated from normal rats. Experiments in which fibronectin was normalized to DNA content of the cells in culture also showed similar results for fibronectin release by Kupffer cells (normal: 2.9 +/- 0.5 ng per microgram DNA per 24 hr; endotoxin-treated: 53.3 +/- 1.3 ng per microgram DNA per 24 hr). Hepatocytes from endotoxin-treated rats released less (p less than 0.01) fibronectin over time than hepatocytes isolated from normal animals. As with Kupffer cells, results for fibronectin release by hepatocytes were similar when normalized to the DNA content (normal: 190.0 +/- 9.4 ng per microgram DNA per 24 hr; endotoxin-treated: 83.3 +/- 4.2 ng per microgram DNA per 24 hr).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Contribution of hepatic fibronectin synthesis to regulation of plasma fibronectin.

Intravenous injection of gelatinized particles, which are phagocytized by the reticuloendothelial system, elicits an acute depletion of plasma fibronectin followed by restoration to normal concentrations in 6-8 h and a rebound elevation at 24 h. We determined the contribution of hepatic fibronectin synthesis to the restoration of plasma fibronectin in rats after particle infusion by measuring the net incorporation of 75selenomethionine into plasma fibronectin during the recovery period. Rats injected intravenously with gelatinized particles had a greater (P less than 0.01) incorporation of labeled 75selenomethionine into fibronectin but less (P less than 0.05) incorporation of 75selenomethionine into total plasma protein than control rats. Inhibition (86%) of hepatic fibronectin synthesis by pretreatment with cycloheximide limited the recovery of fibronectin levels by only 60%, suggesting a source(s) other than hepatic synthesis may contribute to the restoration of plasma fibronectin. Using purified human plasma fibronectin as a tracer in the plasma pool, we found that 26% of the soluble fibronectin consumed from the plasma during particle clearance was subsequently released back into the plasma over a 4-h interval. Tissue analysis indicated that 125I-labeled fibronectin, which was previously incorporated into the tissues, was not released from the tissue pool after the injection of gelatinized particles. Thus the normalization and regulation of plasma fibronectin levels after its acute depletion due to blood-borne particles is a result of 1) an increase in hepatic synthesis of fibronectin, 2) the release of fibronectin previously consumed as an opsonin during particle clearance, and 3) the release of soluble intact fibronectin from a preformed storage pool.

Animals↗
PubMed1989

Plasma fibronectin synthesis in normal and injured humans as determined by stable isotope incorporation.

In humans, plasma fibronectin decreases early after operative injury, burn, or trauma, followed by a rapid restoration with a secondary decline typically observed if such patients become septic. We determined the rate of plasma fibronectin and plasma fibrinogen synthesis in normal subjects and injured patients using a stable isotope incorporation technique with [15N]glycine. During a constant 14-h infusion of [15N]glycine, the enrichment of [15N]glycine in both the free plasma glycine precursor pool as well as the urinary hippurate pool was determined; the latter used as an estimate of intracellular hepatic precursor enrichment. [15N]Glycine enrichment in both plasma fibronectin and fibrinogen was also quantified. The synthesis rate (Js/V) expressed in micrograms per milliliter of plasma per hour and the fractional synthesis rate (FSR) expressed as percentage of the plasma pool produced per day were determined. In normal subjects, the FSR for plasma fibronectin using 15N enrichment into urinary hippurate was 35.35 +/- 1.46%/d, whereas the Js/V was 4.45 +/- 0.19 micrograms/ml plasma per h. In normal subjects, the FSR for plasma fibronectin using 15N enrichment into free plasma glycine was 14.73 +/- 0.63%/d, whereas the Js/V was 1.98 +/- 0.09 micrograms/ml plasma per h. Early (2-3 d) after burn injury, fibronectin synthesis was increased (Js/V = 5.74 +/- 0.36; P less than 0.05), whereas later after injury, fibronectin synthesis began to decline (Js/V = 3.52 +/- 0.24; P less than 0.05) based on 15N enrichment of urinary hippurate. In contrast, the Js/V and FSR of plasma fibrinogen, a well-documented acute-phase plasma protein, revealed a sustained elevation (P less than 0.05) after injury in both the trauma and burn patients. Thus, plasma fibronectin synthesis is elevated early postinjury, which may contribute to the rapid restoration of its blood level. However, once fibronectin levels have normalized, the synthesis of plasma fibronectin appears to decline.

Adult↗
PubMed1989

Phorbol myristate acetate-treated endothelium stimulates polymorphonuclear leukocyte adhesion and superoxide secretion.

The adherence of polymorphonuclear leukocytes (PMNLs) to the pulmonary vascular endothelium may contribute to acute lung injury. We studied the mechanism whereby treatment of bovine pulmonary artery endothelial cells with the potent inflammatory agent phorbol myristate acetate (PMA) stimulated the adherence and secretion of superoxide anion by human PMNLs. Treatment of endothelial cell monolayers with 100 ng/ml PMA resulted in a time-dependent increase in PMNL adherence to the endothelial cells. Treatment of endothelial cells with PMA or the less-lipophilic active phorbol ester 4-beta-phorbol 12,13-dibutyrate stimulated PMNL adherence in a dose-dependent manner, but the inactive phorbol ester 4-alpha-12,13 dideconoate had no effect on PMNL adherence. When formalin-fixed endothelial cells were treated with PMA, the increase in PMNL adherence was similar to that observed with unfixed endothelial cells. Treatment of fixed endothelial cells with 4-beta-phorbol 12,13 dibutyrate did not promote adherence. PMA-induced stimulation of PMNL adherence to endothelial cells was not eliminated by washing the PMA-treated endothelial cells, and the 4-hour conditioned medium obtained from PMA-treated endothelial cells also stimulated PMNL adherence to untreated endothelial cell monolayers PMNL adherence induced by PMA also stimulated the secretion of superoxide anion. Thus, PMA bound to the vascular endothelium will promote both PMNL adhesion and the secretion of reactive oxygen intermediates. Furthermore bound PMA rereleased from the endothelium can also stimulate PMNL adherence and superoxide anion secretion at a distant site.

Animals↗