Acute-phase Plasma Protein Research Articles

Regulation of major acute-phase plasma proteins by hepatocyte-stimulating factors of human squamous carcinoma cells.

Human squamous carcinoma (COLO-16) cells release factors which specifically stimulate the synthesis of major acute-phase plasma proteins in human and rodent hepatic cells. Anion exchange, hydroxyapatite, lectin, and gel chromatography of conditioned medium of COLO-16 cells result in separation into three distinct forms of hepatocyte-stimulating factors (designated HSF-I, HSF-II, and HSF-III) with apparent molecular weights of 30,000, 50,000 and 70,000, respectively. None of the preparations contains detectable amounts of thymocyte-stimulating activity. Each of the three HSF forms stimulates the accumulation of mRNA for alpha 1-antichymotrypsin in the human hepatoma cell line, HepG2. When the same factors were added to primary cultures of adult rat hepatocytes, the expression of the same set of plasma proteins was modulated as by nonfractionated medium. The hormonally induced accumulation of mRNA for acute phase proteins is qualitatively comparable to that occurring in the liver of inflamed rats. Unlike in human cells, in rat liver cells dexamethasone acts additively and synergistically with HSFs. The only functional difference between the three HSF forms lies in the level of maximal stimulation. HSF-I represents the predominant form produced by normal human keratinocytes and closely resembles in molecular size and isoelectric point the activity produced by activated peripheral blood monocytes while the larger molecular weight forms are more prevalent in human as well as mouse squamous carcinoma cells. The observation that HSFs from different sources elicit essentially the same pleiotropic response in hepatic cells led to the hypothesis that the species-specific reaction of adult liver cells to inflammatory stimuli is pre-programmed and that the function of any HSF is to trigger and tune the execution of this fixed cellular process.

Biologic Properties of Epidermal Cell Thymocyte-Activating Factor (ETAF)

Recent studies have shown that in addition to Langerhans cells, keratinocytes can play an active role in immunologic events. As detected by the thymocyte co-stimulator assay, keratinocytes produce a factor with interleukin-1 (IL-1)/leukocytic pyrogen (LP)-like activity termed epidermal cell-derived thymocyte-activating factor (ETAF). ETAF like IL-1/LP can mediate fever. By gel filtration and isoelectric focusing the fever-inducing activity and the ETAF activity in the thymocyte assay co-purify. ETAF activity could be eliminated using a heterologous antibody against IL-1/LP. This antibody could also be used to immunoprecipitate ETAF. ETAF and IL-1/LP induce a peripheral neutrophilia and in vitro are chemotactic for neutrophils. Recently, ETAF has been shown to be a potent T-cell chemoattractant. ETAF-containing preparations have also been shown to stimulate hepatocytes in vitro to synthesize acute phase plasma proteins. In the case of rat hepatocytes, this can be explained by elevated levels of mRNA, although recent purifications have suggested that this hepatocyte-stimulating factor may be separable from the co-stimulator activity. ETAF has also been shown to induce muscle proteolysis in vitro. In addition to its effect on inflammatory and immune events, ETAF has growth promoting effects on keratinocytes and fibroblasts. Thus, ETAF has a multiplicity of divergent biologic effects. It remains to be determined whether the multiplicity of these effects are subserved by a single cytokine or by a family of related cytokines. In either case, ETAF may play an important role in both local cutaneous and systemic inflammatory and immune events.

Secretion of C-reactive protein becomes more efficient during the course of the acute phase response.

We studied the kinetics of synthesis and secretion of the acute phase plasma protein, C-reactive protein, in primary hepatocyte cultures prepared from rabbits manifesting differing degrees of the acute phase response to inflammatory stimulus. In cultures prepared from progressively more responsive animals, rate of C-reactive protein secretion increased to a much greater degree than did intracellular C-reactive protein content, resulting in a progressive decrease in the ratio of intracellular content to rate of secretion. This ratio, which represents the time required to secrete the amount of C-reactive protein contained within the intracellular pool, decreased from 18 h in cultures from unstimulated rabbits to 2.5 h in cells from highly responsive animals. In contrast, these ratios for albumin were short and fell within a narrow range (0.8-2.1 h). In pulse-chase labeling experiments, the time required for secretion of 50% of pulse-labeled C-reactive protein varied markedly, ranging from well over 6 h in cells from a minimally responsive animal to about 75 min in cells from a highly responsive rabbit. In contrast, the half-time for secretion of albumin was consistently about 45 min in the same cultures. Taken together, these findings indicate that the process by which C-reactive protein is secreted becomes more efficient during the course of the acute phase response. Recent studies have indicated that secretory proteins pass from the rough endoplasmic reticulum to Golgi at different and characteristic rates, possibly by a receptor-mediated process in which rate of transfer is determined by receptor affinity. We postulate that C-reactive protein secretion is regulated, during the course of the acute phase response, either by alterations in availability of specific receptors or by competition between different secretory proteins for a common receptor.

Circulating hyaluronate in rheumatoid arthritis: relationship to inflammatory activity and the effect of corticosteroid therapy.

The mean serum hyaluronate concentration in a group of patients with rheumatoid arthritis (n = 37) was 232 +/- 182 (SD) microgram/l and significantly greater (p less than 0.001) than that of age and sex matched healthy controls (mean level 42 +/- 25 micrograms/l) as well as that of patients with ankylosing spondylitis (mean level 58 +/- 33 micrograms/l). In rheumatoid arthritis positive correlations were found between serum hyaluronate and acute-phase plasma proteins, while neither rheumatoid factor titre nor the presence of circulating immune complexes were related to the hyaluronate levels. Increased serum hyaluronate could not be explained by impaired renal or liver function or by drug therapy. During treatment with corticosteroids but not with non-steroidal anti-inflammatory drugs the serum hyaluronate concentrations were significantly reduced. The data obtained suggested an increased production of hyaluronate in rheumatoid arthritis, and the increase seems to be related to the activity of the inflammatory process.

Acute Phase Response in Infectious Disease

In considering the pathology associated with infectious diseases, the most common host response to such infection is inflammation. The mechanism(s) whereby inflammation is initiated and the cell types involved will dictate the kinds of acute phase plasma changes that can be seen associated with the infection. Bacteria seem to initiate the classical type of inflammatory response and plasma protein changes similar to those seen in experimental inflammation induced by chemical means. Viruses, on the other hand, in the absence of cytopathology do not appear to induce the same kind of inflammatory changes and avoid the induction of the acute phase protein response since they may not initiate activation of monocytes and/or macrophages. Those viruses that do cause macrophage activation would be expected to have acute phase protein changes associated with that activation. Parasites, however, appear to initiate the acute phase plasma response only when their migration leads to tissue destruction and local inflammation such as caused by parasitemia with Trypanosoma cruzi in the mouse or with migration of Nippostrongylus brasiliensis in the rodent. Human parasitic diseases require much more investigation in order to clarify the role played by acute phase proteins in the subsequent establishment of the host-parasite relationship. We postulate that the macrophage or monocyte on interaction with the infectious pathogen becomes activated and secretes a number of factors, including interleukin 1 and hepatocyte-stimulating factor, which have a marked effect on the total acute phase reaction. In addition to an effect on phagocytic and immune systems, the mediators cause hepatocytes to markedly increase the secretion of plasma acute phase proteins. Some of these proteins return to the site of inflammation and interact with the infectious pathogen and/or cells and proteins of the host, thereby affecting the final outcome of inflammation. We also propose that the initial interaction of an organism such as a parasite and the mammalian host involves early recognition by the macrophage, thereby initiating both the humoral and cellular acute phase reactions and subsequently affects the immune response against the parasite. Variations in the acute phase reaction may help to explain differences in susceptibility to infectious organisms and the presence or lack of host killing mechanisms for the parasite.

Human keratinocytes and monocytes release factors which regulate the synthesis of major acute phase plasma proteins in hepatic cells from man, rat, and mouse.

Human keratinocytes and activated monocytes produces factors which can stimulate the proliferation of thymocytes. The same activity has also been implicated in regulating the expression of plasma proteins in liver cells during the acute phase reaction. To assess whether factors produced by such cells can directly influence liver cells to change the production of acute phase plasma proteins, we studied in tissue culture the response pattern of hepatic cells from three species: human hepatoma cells ( HepG2 cells), and primary cultures of rat and mouse hepatocytes. Conditioned media from the squamous carcinoma COLO-16 cells, normal epidermal cells, and activated peripheral monocytes were able to stimulate the synthesis of specific acute phase plasma proteins: alpha 1-antichymotrypsin in HepG -2 cells, alpha 1-antichymotrypsin, alpha 1-acid glycoprotein, alpha 1-acute phase protein, and alpha 2-macroglobulin in rat hepatocytes, and alpha 1-acid glycoprotein, haptoglobin, and hemopexin in mouse hepatocytes. Only in rat cells, dexamethasone was found to have further enhancing effect. The increased production of plasma proteins could be explained by an elevated level of functional mRNA. Comparing thymocyte-stimulating activities with the effects on plasma protein production, we found some difference both between the conditioned media of epidermal cells and monocytes, and between the responses of the three hepatic cell systems. Furthermore, gel chromatography of conditioned media resulted in partial separation of activities regulating liver cells and thymocytes. Since there is no strict correlation between thymocyte- and hepatocyte-stimulating activities, the presence of different sets of specific factors is assumed.

Synthesis and regulation of acute phase plasma proteins in primary cultures of mouse hepatocytes.

Adult mouse hepatocytes respond in vivo to experimentally induced acute inflammation by an increased synthesis and secretion of alpha 1-acid glycoprotein, haptoglobin, hemopexin, and serum amyloid A. Concurrently, the production of albumin and apolipoprotein A-1 is reduced. To define possible mediators of this response and to study their action in tissue culture, we established primary cultures of hepatocytes. Various hormones and factors that have been proposed to regulate the hepatic acute phase reaction were tested for their ability to modulate the expression of plasma proteins in these cells. Acute phase plasma and conditioned medium from activated monocytes influenced the production of most acute phase plasma proteins, and the regulation appears to occur at the level of functional mRNA. Purified hormones produced a significant anabolic response in only a few cases: dexamethasone was found to be effective in maintaining differentiated expression of the cells; and glucagon produced a specific inhibition of haptoglobin synthesis. When cells were treated with a combination of conditioned monocyte medium and dexamethasone, secretion of proteins was markedly reduced. The carbohydrate moieties of all plasma glycoproteins were incompletely modified, apparently as a result of decreased intracellular transport of newly synthesized plasma proteins. Although primary hepatocytes were not phenotypically stable in tissue culture, the cells nevertheless retained a broad response spectrum to exogenous signals. We propose this as a useful system to study the production of plasma proteins and thereby pinpoint the nature and activity of effectors mediating the hepatic acute phase reaction.

Analysis of the effects of snake venom proteinases on the activity of human plasma C1 esterase inhibitor, α1-antichymotrypsin and α2-antiplasmin

Incubation of C1 esterase inhibitor with Crotalid, Viperid and Colubrid snake venoms resulted in enzymatic inactivation of the inhibitor. Intact inhibitor (104 kDa) was converted into an active intermediate species of 89 kDa and then a further cleavage resulted in formation of an 86-kDa inactive inhibitor. In contrast, C1 esterase inhibitor did not lose activity during incubation with Elapid venoms; however, the intact inhibitor was gradually converted to an active species of 89 kDa during the incubation. Human α 1-antichymotrypsin was inactivated by all venoms tested, including those from the Elapid family. The 67-kDa intact inhibitor was converted by the venom proteinases to an inactive 63-kDa form. The results suggest that this acute-phase plasma protein is readily susceptible to inactivation by venom proteinases. Human α 2-antiplasmin (68 kDa) was cleaved to form a 61-kDa active intermediate, which then underwent a second cleavage to produce an inactive 53-kDa product. Elapid venoms had no effect on α 2-antiplasmin activity and did not cleave this inhibitor. All inhibitors were inactivated with catalytic amounts of venom proteinases. No stable proteinase-proteinase inhibitor complexes were detected, and no random proteolysis of the inhibitors occurred.

The acute-phase response in (NZB X NZW)F1 and MRL/l MICE.

The acute-phase plasma protein response to disease activity in murine models of autoimmune lupus-like disease was investigated by measurement of the concentration of serum amyloid P component (SAP) in NZB X W and MRL/l mice. The levels of SAP, which is a major acute-phase protein in mice, did not rise at all in response to progression of disease in NZB X W mice between the ages of 1 and 9 mo. This resembles the behavior of acute-phase proteins such as C-reactive protein and serum amyloid A protein in human systemic lupus erythematosus, and just as in human lupus, where the occurrence of intercurrent microbial infection can stimulate an acute-phase response, so injection of bacterial lipopolysaccharide or casein into the NZB X W mice stimulated "normal" acute-phase SAP production. In marked contrast, MRL/l mice developed greatly increased levels of SAP, which correlated closely with progression of their pathology as they aged. The disease profile of the MRL/l strain includes rheumatoid factors and spontaneous polyarthritis and their SAP response resembles the behavior of acute phase proteins in human rheumatoid arthritis. Different patterns of acute-phase response in different autoimmune disorders may thus be a reflection of the genetic predisposition to particular diseases and/or contribute to their pathogenesis. The existence of animal counterparts for the various clinical patterns of human acute-phase protein production will assist in experimental investigation of the underlying mechanisms and of the biological role of the acute-phase response.

An in vitro bioassay for leukocytic endogenous mediator(s) using cultured rat hepatocytes.

Primary cultures of adult rat hepatocytes were used to assay for the presence of leukocytic mediator(s) (LEM), a neutrophil derived protein(s) capable of stimulating the synthesis of acute-phase plasma proteins when injected into rats. In the presence of physiological concentrations of dexamethasone (40 mM), the hepatocytes secreted a variety of plasma proteins as demonstrated by crossed immunoelectrophoresis. The addition of LEM to hepatocytes increased the secretion of several acute-phase related plasma proteins, including fibrinogen and hepatoglobin, and decreased albumin secretion. These results mimic the acute-phase response observed in the intact animal. Fibrinogen secretion was used as a quantitative marker for determining LEM activity. The rate of fibrinogen secretion depended upon both the concentration of dexamethasone and LEM present during a given 24-h assay period. One unit of LEM activity is defined as that concentration of LEM capable of producing a 50% maximal stimulation of fibrinogen secretion.

Multiple Systems Organ Failure

The correlations between the acute phase and nutritional plasma proteins and intravenous amino acid dosage have been explored in a group of 25 trauma septic patients of whom 14 survived. The two groups of patients appeared to have equal cardiopulmonary function and exogenous nutritional support. The surviving group showed significant associated changes (p less than or equal to 0.05) between alpha1 acid glycoprotein, alpha2 HS glycoprotein, and ceruloplasmin (acute-phase proteins) and between prealbumin, retinol-binding protein, and transferrin (nutritional proteins). There were no correlations in concentration changes between these two groups of plasma proteins. The surviving group showed significant positive correlations between the nutritional plasma protein and intravenous amino acid dosage (prealbumin, p less than or equal to 0.001; transferrin, p less than or equal to 0.008; retinol-binding protein, p less than or equal to 0.001; and albumin, p less than or equal to 0.004) but no correlations with the acute-phase proteins. The nonsurviving patients showed significant intercorrelations between the acute-phase and nutritional proteins that were not seen in the surviving patients, and showed no relationship between intravenous amino acid dosage and the plasma levels of nutritional proteins. The data are consistent with increased obligatory catabolism of amino acids in the nonsurviving patient which based upon the amino acid behavior documented in the first paper in this series probably involves the branched-chain amino acids.

The Response of the Acute-Phase Plasma Protein α2-Macroglobulin to Vitamin A Deficiency in the Rat

Alpha2-macroglobulin (alpha2-M) was determined in the sera of vitamin A-deficient and pair-fed control rats. Alpha2-macroglobulin levels were assayed by radial immunodiffusion using a purified standard sample from sera from turpentine-stressed rats. There was no significant increase in the serum level of alpha2-M in vitamin A-deficient rats relative to pair-fed controls, indicating that there was no acute response to the stress of the deficiency. When turpentine was injected to elicit an acute response, the serum level of alpha2-M rose equally (by almost 100-fold) in both deficient and normal rats. It was concluded that the response to vitamin A deficiency of serum glycoprotein synthesis shows some specificity in that the synthesis of alpha2-M in response to stress was not affected by vitamin A, whereas we previously found that levels of alpha1-M, which is very similar to alpha2-M, decline with vitamin A deficiency.

Increased plasma protein binding of propranolol and chlorpromazine mediated by disease-induced elevations of plasma alpha1 acid glycoprotein.

To assess the importance of disease-induced increases in plasma concentrations of alpha1 acid glycoprotein (an acute-phase plasma protein that binds cationic drugs), we determined binding of propranolol in plasma from 53 patients and 25 healthy volunteers. Binding was increased in 10 patients with Crohn's disease (P less than 0.002), nine with inflammatory arthritis (P less than 0.002) and eight with chronic renal failure with superimposed inflammatory disease (P less than 0.01) as compared with healthy controls. The plasma binding of control subjects did not differ from that of 12 patients with chronic hepatic disease (P greater than 0.45) or 14 with uncomplicated renal failure (P greater than 0.80). Chlorpromazine binding, determined in 60 subjects, yielded similar results. Percentage of free drug and alpha1 acid glycoprotein concentration were inversely correlated (r = -0.77 with propranolol, P less than 0.001, and r = -0.69 with chlorpromazine, P less than 0.001). Increases in plasma protein binding in patients with inflammatory disease appear mediated by increases in alpha1 acid glycoprotein concentration, which may influence drug kinetics.

The site of synthesis and functions of acute phase plasma proteins: Close relation-ship with the reticulo-endothelial system

The evidence that the acute phase glycoproteins of plasma are synthesized by the liver parenchymal cells is critically examined, and is found to be inconclusive. Some evidence is cited which favors the reticulo-endothelial system (RES) in general, and Kupffer cells in particular, as the site of synthesis of these proteins: 1. The entire RES contains non-glycogenic periodic acid Schiff-positive substances. 2. The diseases which affect glycoprotein levels are also known to affect the function of the RES. 3. When the animal is subjected to stress, the RES function is initially depressed and then stimulated. A similar biphasic behavior is shown by plasma glycoprotein levels. 4. Adrenal cortico-steriods are the major regulators of the RES function and of the synthesis of acute phase proteins. Moreover, both are stimulated at low concentrations, and depressed at high concentrations of the hormone. Some of the glycoproteins of the acute phase (prothrombin, the third component of complement, haptoglobin, transferrin and ceruloplasmin) have defense-related functions. The others seem to participate in phenomena like detoxification, promotion of phagocytosis, wound healing, prevention of tissue injury by lysosomal enzymes, prevention of trauma and recovery from inflammation. It is proposed that the acute phase proteins, together with antibodies, form major components of the defense system, and the RES attempts to deal with injury by mobilization of increased amounts of these substances.

The role of the acute phase reaction in inflammation.

Inflammation and injury to tissue results in a variety of local and systemic events, however although the local events of oedema formation and cellular infiltration have received considerably more attention the systemic response to inflammation is no less profound. The particular systemic event which forms the substance of this communication is the change in the circulating levels of plasma proteins which occurs after inflammatory injury, and the manner in which these changes in plasma concentration are controlled by changes in plasma concentration are controlled by changes in the rate of synthesis. A discussion of the role of the liver in controlling inflammatory events, in relation to the synthesis of an anti-inflammatory protein has been given; the present work is an extension of this and describes the changes in concentration and synthesis rate of albumin, fibrinogen and alpha1 acid glycoprotein during adjuvant arthritis in the rat. The changes which occur are regulated at the liver by alteration of the rate of synthesis of the individual protein. For example albumin at the height of adjuvant arthritis falls to a third of its normal plasma level whereas the level of alpha1 acid glycoprotein increases up to twenty-fold; these changes are reflected by similar changes in their synthesis rate by the liver. The effect of the fall in albumin concentration on the plasma binding of anti-inflammatory drugs (and their toxicity) in relation to these findings will be discussed along with the biological role of the acute phase plasma proteins and hence the influence of the liver in the response to injury.

Effect of ACTH on biosynthesis of fibrinogen in the rabbit.

ARTICLESEffect of ACTH on biosynthesis of fibrinogen in the rabbitAC Atencio, and L LorandAC Atencio, and L LorandPublished Online:01 Nov 1970https://doi.org/10.1152/ajplegacy.1970.219.5.1161MoreSectionsPDF (1 MB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInWeChat Previous Back to Top Next Download PDF FiguresReferencesRelatedInformationCited ByThe Coagulation SystemFatty acids, fibrinogen and blood flow: A general mechanism for hyperfibrinogenemia and its pathologic consequencesMedical Hypotheses, Vol. 6, No. 5Effects of insulin on plasma fibrinogen levels in rats submitted to tissue injury or ACTH administrationExperientia, Vol. 35, No. 5Hematologic and serologic studies in 6‐sulfanilamidoindazole arthritis23 November 2005 | Arthritis & Rheumatism, Vol. 21, No. 6The site of synthesis and functions of acute phase plasma proteins: Close relation-ship with the reticulo-endothelial systemMedical Hypotheses, Vol. 3, No. 2Plasma fibrinogen response in the rat after thyroid stimulating hormone therapyExperientia, Vol. 32, No. 11The effect of leucocytic pyrogen on rabbit fibrinogen. A third viewThrombosis Research, Vol. 9, No. 5The role of sialic acid in determining the life-span of circulating cells and glycoproteinsExperientia, Vol. 32, No. 2Regulation of Net Biosynthesis of Albumin, Fibrinogen, α1-Acid Glycoprotein, α2-(Acute Phase) Globulin, and Haptoglobin by Direct Action of Hormones on the Isolated Perfused LiverDelayed Incorporation of [ 75 Se]Selenomethionine into Fibrinogen: Its Effect upon Kinetic Studies of Fibrinogen with [ 75 Se]Selenomethionine in RabbitsBritish Journal of Haematology, Vol. 26, No. 4Acute-Phase ReactantsEffect of Whole-body X-irradiation on Plasma-protein Synthesis: Role of Adrenals3 July 2009 | International Journal of Radiation Biology and Related Studies in Physics, Chemistry and Medicine, Vol. 23, No. 5Insulin transformation in the liverActa Diabetologica Latina, Vol. 9, No. 1Effect of spironolactone and phenobarbital on the plasma fibrinogen in ratsExperientia, Vol. 28, No. 11Fibrinogen catabolism in rats fed thrombogenic dietsAtherosclerosis, Vol. 16, No. 3 More from this issue > Volume 219Issue 5November 1970Pages 1161-1165 Copyright & PermissionsCopyright © 1970 by American Physiological Societyhttps://doi.org/10.1152/ajplegacy.1970.219.5.1161PubMed4319643History Published online 1 November 1970 Published in print 1 November 1970 Metrics

Mechanism for hormone-induced alterations in serum ceruloplasmin.

Mechanism for hormone-induced alterations in serum ceruloplasmin.