Major Acute-phase Plasma Protein Research Articles

Levels of C-reactive protein (CRP) in elderly patients with unipolar depression – case control analysis

Aim: C-reactive protein (CRP) is the major acute-phase plasma protein. Studies show that patients with depression have elevated levels of CRP. The aim of the study was to determine differences in CRP serum level in elderly patients with unipolar depression (DEP) compared with non-depressed elderly patients (nonDEP) using case-control analysis.Methods: Serum level of CRP was measured in 404 (DEP: n = 202, nonDEP: n = 202) Caucasian inpatients aged ≥60 (350 women, 86.7%; mean age = 76.7 years).Results: Mean CRP level in the study groups was: DEP 2.67 ± 2.56 mg/dL, nonDEP 2.41 ± 2.19 mg/dL, the difference was not significant (p = 0.96). The overall rate of being above the high level of CRP (set at 3.0 mg/L) was 33.2% for DEP and 29.2% for nonDEP groups (p = 0.39). It was also found that, in the whole study group, CRP level was not correlated with age (p = 0.10).Conclusions: Elderly patients with depression have no increased CRP levels. A high percentage (∼30%) of all subjects had a CRP level >3 mg/L, which is the cut-off point for increased cardiovascular risk.

Structure of serum amyloid A suggests a mechanism for selective lipoprotein binding and functions: SAA as a hub in macromolecular interaction networks.

Serum amyloid A is a major acute-phase plasma protein that modulates innate immunity and cholesterol homeostasis. We combine sequence analysis with x-ray crystal structures to postulate that SAA acts as an intrinsically disordered hub mediating interactions among proteins, lipids and proteoglycans. A structural model of lipoprotein-bound SAA monomer is proposed wherein two α-helices from the N-domain form a concave hydrophobic surface that binds lipoproteins. A C-domain, connected to the N-domain via a flexible linker, binds polar/charged ligands including cell receptors, bridging them with lipoproteins and rerouting cholesterol transport. Our model is supported by the SAA cleavage in the interdomain linker to generate the 1-76 fragment deposited in reactive amyloidosis. This model sheds new light on functions of this enigmatic protein.

Levels of C-reactive protein (CRP) in patients with schizophrenia, unipolar depression and bipolar disorder

Objective: C-reactive protein (CRP) is the major acute-phase plasma protein. Studies show that patients with various mental disorders have elevated levels of CRP. The aim of the study was to determine differences in CRP serum level in patients with acute schizophrenia, unipolar depression, bipolar depression and bipolar mania. Method: Serum level of CRP was measured in 950 Caucasian inpatients (589 women, 62.0%; mean age 50.3 years). Results: Mean concentration of CRP in study groups was: schizophrenia (n = 485) 5.30 mg/l, unipolar depression (n = 319) 5.61 mg/l, bipolar disorder (n = 146) 4.65 mg/l, bipolar depression (n = 114) 3.82 mg/l and bipolar mania (n = 32) 7.36 mg/l. There was no difference for CRP levels between patients with schizophrenia, unipolar depression, bipolar depression and bipolar mania (P = 0.58). The overall rate of being above the high level of CRP (set at 3.0 mg/l) was 35.7% for schizophrenia, 38.6% for unipolar depression, 40.4% for bipolar disorder, 40.4% for bipolar depression and 40.6% for bipolar mania. There were no significant differences in the risk of having high level of CRP between the clinical groups. The rate of patients being above high level was higher in women. We also found that in whole study group CRP level was positively correlated with age (P = 0.002). Conclusions: Although there is no statistically significant difference in CRP serum level between patients with schizophrenia, unipolar depression, bipolar depression and bipolar mania, our results show that more than one-third (37.4%) of all subjects had CRP level > 3 mg/l, which is the cut-off point for high cardiovascular risk.

Distal Promoter Regions Are Responsible for Differential Regulation of Human Orosomucoid-1 and -2 Gene Expression and Acute Phase Responses

Human orosomucoid (ORM) is a major acute-phase plasma protein, encoded by 2 highly homologous genes, ORM1 and ORM2. Human ORM induction is assumed to be regulated by each proximal promoter region, where putative glucocorticoid responsive elements and CCAAT/enhancer binding protein (C/EBP)β binding sites are located. However, the details of the differential regulation of these genes remain unknown. To explore this, we assessed the role of the distal promoter region of each ORM in HeLa cells. Luciferase-reporter activities of full constructs, containing approximately 1.1 kbp (FULL), and those of deletion constructs, containing up to 188 bp region (DEL) upstream of the transcription start sites of ORM1 and ORM2 were compared under both basal and inducer-treated conditions. For ORM1 and ORM2 DEL constructs, significantly increased activities after dexamethasone (DEX) treatments (alone and combined with interleukin (IL)-1β) were observed. Significantly higher FULL construct activities than DEL construct activities were observed for ORM1 after IL-1β treatment, while those for ORM2 were significantly lower at basal level and after DEX treatments. Upon C/EBPβ overexpression, FULL construct activities were significantly higher than those of DEL constructs at basal level and after IL-1β treatment for ORM1, and at basal level and after inducer-treatments for ORM2. Higher transcription-induction activity in the distal promoter region was evident for ORM1 in the absence of C/EBPβ overexpression, and for ORM2 under C/EBPβ overexpression conditions. These findings suggest that the ORM distal promoter region differentially regulates expression of ORM genes at basal level and in acute phase responses.

Identification of a specific haptoglobin C‐terminal fragment in arthritic synovial fluid and its effect on interleukin‐6 expression

Haptoglobin (Hp), a major acute-phase plasma protein, has been found in arthritic synovial fluid (SF). However, the function and structural modifications of Hp in arthritic SF are unknown. To investigate in vivo generation of modified Hp associated with inflammatory disease, we examined a new Hp isoform in SF from patients with rheumatoid arthritis (RA). Specific Hp fragments of 28000 and 15000 molecular weight were identified in SF of patients with RA, and the two polypeptides were presumed to be fragments of the Hp β-chain (43000 MW) produced by cleavage with plasmin. The 15000 MW fragment, which is a C-terminal region of Hp, was observed at higher frequency and levels in RA than in osteoarthritis. Plasmin activity was also higher in SF of RA patients. A recombinant 15000 MW Hp fragment up-regulated interlukin-6 expression in monocytic cells. These findings indicate that the C-terminal Hp fragment is generated by plasmin in local inflammatory environments and acts as an inflammatory mediator. They further suggest that a specific Hp fragment might be applied as a novel biomarker for the diagnosis and prognosis of inflammatory diseases such as RA.

Pyrrolidine Dithiocarbamate Inhibits Interleukin-6 Signaling through Impaired STAT3 Activation and Association with Transcriptional Coactivators in Hepatocytes

Pyrrolidine Dithiocarbamate Inhibits Interleukin-6 Signaling through Impaired STAT3 Activation and Association with Transcriptional Coactivators in Hepatocytes

Pyrrolidine Dithiocarbamate Inhibits Interleukin-6 Signaling through Impaired STAT3 Activation and Association with Transcriptional Coactivators in Hepatocytes

Interleukin (IL)-6 is a proinflammatory cytokine that has been implicated in the expression of acute phase plasma proteins and hepatic insulin resistance through activation of the JAK/STAT3 pathway. Although previous studies have demonstrated that pyrrolidine dithiocarbamate (PDTC) exerts protection against inflammatory responses, its role in the regulation of IL-6 receptor signaling remains unclear. Here we show that treatment of cultured HepG2 hepatoma cells with PDTC inhibits IL-6-stimulated tyrosine phosphorylation and subsequent nuclear translocation of STAT3 in a dose- and time-dependent fashion. No inhibition of JAK-1 activity was observed. To provide insight into PDTC signaling, we constructed a conditionally active STAT3 by fusing it with the ligand binding domain of the estrogen receptor (STAT3-ER). In the presence of 4-hydroxytamoxifen STAT3-ER was translocated in the nucleus of HepG2 cells in a phosphorylation-independent manner, and treatment with PDTC mitigated the response. Although STAT3 coprecipitated with heat-shock protein 90 (Hsp90) in control cells, coprecipitation of the two proteins was greatly reduced after PDTC treatment or after exposure to geldanamycin, an Hsp90 inhibitor. As a result there was a decrease in IL-6-induced association of STAT3 with the transcriptional coactivators FOXO1a and C/EBPbeta together with significant reduction in the expression of SOCS-3 protein and that of two major acute phase plasma proteins. Importantly, treatment of HepG2 cells and a primary culture of rat hepatocytes with PDTC restored insulin responsiveness that was abrogated by IL-6. These studies are consistent with the ability of PDTC to down-regulate IL-6-induced STAT3 activation by altering the stability of STAT3-Hsp90 complex.

185 C-Reactive Protein in The Tracheobronchial Secretion of A Preterm Infant

Background: C reactive protein (CRP) is a major acute-phase plasma protein displaying rapid and pronounced rise of its serum concentration in response to infection or tissue injury. CRP is usually synthesized by hepatocytes and induced by cytokines. However, extrahepatic synthesis has been recently documented. In particular, CRP has been detected in adult human respiratory tract and it has been shown that its gene is expressed in epithelial cells. Methods: Tracheobronchial secretions (TBS) were collected longitudinally in an infant delivered at 24±3 weeks of gestation who developed a middle right lobe pneumonia 5 weeks after delivery. Cultures of TBS were positive for Staphylococcus aureus. Antibiotic therapy with 100 mg/kg/day of amoxicillin/clavulanic acid and 7.5 mg/kg/day of amikacin was administered intravenously for 10 and 5 days, respectively. Nephelometry was used for the determination of CRP in neonatal serum, while a commercially available ELISA test was utilized for the measurement of both blood and TBS CRP concentrations. The sensitivity of the CRP assay was 0.39 ng/mL (Inter-assay coefficient of variation: 6.5%). To provide evidence that CRP is produced locally in the tracheobronchial system a ratio between CRP and IgG both in serum and in the TBS were calculated. The reason for this is twofold: first, IgG and CRP have a similar molecular size, and second, it is known that IgG are present in extremely low concentrations on the mucosal surface. To allow comparison, CRP and IgG concentrations in TBS were normalized to the total protein content. Results: CRP was present in TBS. A marked increase of CRP in serum and in the TBS was noted during the course of pneumonia with a sharp decrease after antibiotic therapy. The highest CRP concentration in serum (160 mg/L) and TBS (345.9 ng/ml) was found 2 and 3 days after the diagnosis was made, respectively. The CRP – IgG ratio was 21340 times higher in the TBS than in serum. Serum CRP and TBS CRP decrease was observed 1 and 2 days after the initiation of antibiotic therapy, respectively. Conclusion: CRP is detectable in the TBS of preterm infants during pneumonia. It can be speculated that CRP is not only involved in a systemic response against pathogens but also acts as a mediator in the local defense. It remains to be explored whether TBS CRP measurement is of value in clinical practice.

The Carboxyl-terminal Region of STAT3 Controls Gene Induction by the Mouse Haptoglobin Promoter

Haptoglobin (HP) is one of the major acute phase plasma proteins in the mouse, and its synthesis is additively induced by interleukin (IL)-6 and glucocorticoids. STAT3 serves as the mediator of the IL-6 receptor signal and appears to contribute to the transcriptional induction of acute phase protein genes. The carboxyl-terminal region of STAT3, consisting of an acidic domain and containing a serine phosphorylation site, has been proposed to contribute to the induction process. To assess the role of STAT3 in the transcriptional control of the HP promoter, we applied two mutant forms of STAT3: one with a deletion of the carboxyl-terminal 55 amino acid residues, STAT3Delta55C, and the other with a substitution of serine 727 to alanine, STAT3SA. Like the wild-type STAT3, both mutant STAT3 forms are activated by the signal-transducing subunit of the IL-6 receptor, gp130, or by co-transfected IL-3 receptor. Ectopic expression and activation of wild-type STAT3 or STAT3SA in HepG2 hepatoma cells similarly enhance transcription through the IL-6-response element of the HP promoter. This enhancement is specific for STAT3 and cannot be reproduced by STAT1 or STAT5. In contrast, STAT3Delta55C inhibits IL-6-induced transcriptional activation. Interestingly, whereas receptor-activated STAT3 also enhances stimulation of the haptoglobin promoter by dexamethasone through the glucocorticoid receptor, activated STAT3Delta55C reduces the regulation below the level achieved by the glucocorticoid receptor alone. This transdominant action by STAT3Delta55C is dependent on a functional IL-6-responsive element. The data suggest that the carboxyl-terminal domain, but not its serine phosphorylation site of STAT3, is required for transcription as part of the hematopoietin receptor signaling as well as for cooperation with other transcription factors such as the glucocorticoid receptor.

Nonexpression of the Human Serum Amyloid A Three (SAA3) Gene

Serum amyloid A (SAA) is a major acute-phase plasma protein synthesized by the liver. In addition to the two major plasma isoforms described in humans (SAA1 and SAA2), a third form (SAA3) has been demonstrated in several other species and is distinguished by predominant extrahepatic expression. Two clones, Ch11g5-1-1 and HDg1-1, containing the human SAA3 gene are described in this report. The human SAA3 gene is comparable in organization to the SAA1 and SAA2 genes and shares with them 87% nucleotide identity in the region spanning exon 3 through exon 4. Sequences 5' to exon 3, however, are strikingly different from those in the SAA1 and SAA2 genes. For instance, the sequence deduced for amino acids 1-12 (exon 2) has only 25% identity with human SAA1 and SAA2; it most closely resembles that of rabbit SAA3 isolated from synovial fibroblast cultures (75% identity). Although rabbit SAA3 induces collagenase production in an autocrine fashion the human SAA3 gene is not expressed. This is shown by: (i) a single base insertion in the sequence corresponding to codon 31, (ii) the inability of a 918-bp fragment immediately upstream from SAA3 exon sequences to direct transcription of a chloramphenicol acetyltransferase reporter gene, and (iii) the absence of detectable human SAA3 in mRNA.

Interleukin-11 regulates the hepatic expression of the same plasma protein genes as interleukin-6.

Treatment of rat hepatoma H-35 cells with purified human recombinant interleukin-11 (IL-11) resulted in the stimulated production of several major acute phase plasma proteins. The qualitative and quantitative changes were comparable to those mediated by IL-6 or leukemia inhibitor factor (LIF). Like IL-6, IL-11 acted synergistically with IL-1 on type 1 acute phase proteins. The combination of IL-11 and dexamethasone yielded a magnitude of stimulation which was more similar to the combination of LIF and dexamethasone than IL-6 and dexamethasone. IL-11 elicited in treatment of primary cultures of rat hepatocytes a qualitative change of plasma protein production which was similar to that in H-35 cells. Comparison of rat and human hepatoma cells indicated that the IL-11 response did not correlate with that of IL-6 or LIF, suggesting that the action of IL-11 was mediated by an IL-11-specific receptor system. However, the intracellular transduction of the IL-11, IL-6, and LIF signals to the acute phase protein genes seems to rely, in part, on common elements as judged from their stimulatory effects on the transfected expression vector containing the IL-6 response element of the rat beta-fibrinogen gene. The finding that IL-11 shares liver-regulating properties with IL-6 and LIF suggests that IL-11 has the potential of contributing to the control of systemic homeostasis and hepatic acute phase response.

Transcriptional regulation through cytokine and glucocorticoid response elements of rat acute phase plasma protein genes by C/EBP and JunB.

Independent of de novo protein synthesis, interleukin-1, interleukin-6, and dexamethasone caused immediate stimulation of transcriptional activity of most major acute phase plasma protein genes in the rat hepatoma H-35 cells. However, activation of alpha 2-macroglobulin and alpha 1-acid glycoprotein genes were delayed by 2-4 h and required ongoing protein synthesis. The hormones also increased transiently the transcription of the junB gene and the amounts of JunB, C/EBP, and C/EBP-like mRNA. To identify whether JunB and C/EBP have the ability to control both the early and late acute phase reactants, expression vectors for mouse C/EBP and JunB together with reporter gene constructs containing recognized hormone-specific regulatory elements were introduced into hepatoma cells. C/EBP displayed prominent transactivation activity with the interleukin-1 and glucocorticoid regulatory elements of alpha 1-acid glycoprotein, the interleukin-1 regulatory element of haptoglobin gene, and the interleukin-6 regulatory element of beta-fibrinogen. The interleukin-6 regulatory elements of the first two genes and the glucocorticoid response element of the third gene were not affected by C/EBP. These data suggest that normal hormone activation of these three acute phase reactant genes might involve, in part, C/EBP-related factors which have a broad range of specificity. H-35 cells stably transformed with a mouse C/EBP expression vector showed an elevated basal level as well as cytokine inducible expression of some but not all acute phase reactants. Cotransfected JunB resulted in reduced activity of cytokine-responsive constructs and in lower transactivation by C/EBP. JunB appears to function as a modulator of plasma protein expression during the course of acute phase response.

Structure and characterisation of a duplicated human α 1 acid glycoprotein gene

Human α 1-acid glycoprotein (AGP), also known as orosomucoid, is a major acute-phase plasma protein. The amino acid sequence of AGP, which was determined by sequencing from protein isolated from pooled plasma, contained amino acid substitutions in 21 different positions. Genomic and cDNA clones which correspond to one of the possible amino acid sequences have been previously reported. In this paper we present the complete nucleotide sequence of a second gene, AGP2 which is located approx. 3.3 kb downstream from AGP1. The derived amino acid sequence of AGP2 contains 19 of the possible alternative amino acid substitutions as well as two additional differences. It is clear from the results presented here that the AGP in human plasma is the product of two separate gene loci.

Synthesis of hemopexin and cysteine protease inhibitor is coordinately regulated by HSF-II and interferon-β2 in rat hepatoma cells

Rat hepatoma (H-35) cells respond to hepatocyte-stimulating factors by increased expression of major acute phase plasma proteins. The synthesis of hemopexin is stimulated 10-fold by either hepatocyte-stimulating factor-II of human squamous carcinoma cells or hepatocyte-stimulating factor/interferon-β2 of activated human blood monocytes. The hormone specificity, time course and dose-dependence of hemopexin regulation is closely correlated with that of cysteine protease inhibitor. The coordinate expression of hemopexin and other type II acute phase proteins suggests the existence of common molecular regulatory mechanisms.

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.

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.