Expression Of Serum Amyloid A Research Articles

Serum Amyloid A Facilitates Early Lesion Development in Ldlr-/- Mice.

BackgroundAtherosclerosis is a chronic inflammatory disorder, and several studies have demonstrated a positive association between plasma serum amyloid A (SAA) levels and cardiovascular disease risk. The aim of the study was to examine whether SAA has a role in atherogenesis, the underlying basis of most cardiovascular disease.Methods and ResultsMice globally deficient in acute-phase isoforms Saa1 and Saa2 (Saa−/−) were crossed to Ldlr−/− mice (Saa−/−Ldlr−/−). Saa−/−Ldlr−/− mice demonstrated a 31% reduction in lesional area in the ascending aorta but not in the aortic root or innominate artery after consuming a high-fat, high-cholesterol Western-type diet for 6 weeks. The lesions were predominantly macrophage foam cells. The phenotype was lost in more mature lesions in mice fed a Western-type diet for 12 weeks, suggesting that SAA is involved in early lesion development. The decreased atherosclerosis in the Saa−/−Ldlr−/− mice occurred despite increased levels of blood monocytes and was independent of plasma lipid levels. SAA is produced predominantly by hepatocytes and macrophages. To determine which source of SAA may have a dominant role in lesion development, bone marrow transplantation was performed. Ldlr−/− mice that received bone marrow from Saa−/−Ldlr−/− mice had slightly reduced ascending aorta atherosclerosis compared with Saa−/−Ldlr−/− mice receiving bone marrow from Ldlr−/− mice, indicating that the expression of SAA by macrophages may have an important influence on atherogenesis.ConclusionsThe results indicate that SAA produced by macrophages promotes early lesion formation in the ascending aorta.

MRNA expression of genes involved in inflammation and haemostasis in equine fibroblast-like synoviocytes following exposure to lipopolysaccharide, fibrinogen and thrombin

BackgroundStudies in humans have shown that haemostatic and inflammatory pathways both play important roles in the pathogenesis of joint disease. The aim of this study was to assess mRNA expression of haemostatic and inflammatory factors in cultured equine fibroblast-like synoviocytes exposed to lipopolysaccharide (LPS), fibrinogen and thrombin. Synovial membranes were collected from metacarpo-phalangeal joints of 6 skeletally mature horses euthanized for non-orthopaedic reasons. Passage 4 fibroblast-like synoviocytes were left non-treated or treated with either 0.1 μg/ml LPS, 5 mg/ml fibrinogen or 5 U/ml thrombin and harvested at time points 0, 6, 24 and 48 h. mRNA expression of serum amyloid A (SAA), interleukin-6 (IL-6), monocyte chemotactic protein 1 (MCP-1), tissue factor (TF), plasminogen activator inhibitor 1 (PAI-1), urokinase plasminogen activator (uPA), vascular endothelial growth factor (VEGF) and protease activator receptor 1 (PAR-1) was assessed using quantitative real time reverse transcriptase PCR.ResultsLPS caused a significant increase in mRNA expression of SAA, IL-6, MCP-1 and uPA, and a decrease in TF, PAI-1 and PAR-1 when compared to non-treated cells. Treatment with thrombin resulted in increased mRNA expression of SAA, IL-6, MCP-1 and PAI-1, and a decreased PAR-1 expression compared to non-treated cells. The fibrinogen-treated synoviocytes showed significantly increased mRNA expression of IL-6, MCP-1, TF and PAI-1, and decreased PAR-1 expression compared to non-treated cells.ConclusionLPS, fibrinogen and thrombin induced an increased gene expression of inflammatory markers in isolated equine fibroblast-like synoviocytes. LPS caused changes in gene expression promoting increased fibrinolysis, while fibrinogen and thrombin changed the gene expression resulting potentially in reduced fibrinolysis. Overall, it appeared that both inflammatory and haemostatic stimuli affected expression of genes involved in inflammatory and haemostatic pathways, supporting their importance in equine joint diseases.

Intrinsic Stability, Oligomerization, and Amyloidogenicity of HDL-Free Serum Amyloid A.

Serum amyloid A (SAA) is an acute-phase reactant protein predominantly bound to high-density lipoprotein in serum and presumed to play various biological and pathological roles. Upon tissue trauma or infection, hepatic expression of SAA increases up to 1,000 times the basal levels. Prolonged increased levels of SAA may lead to amyloid A (AA) amyloidosis, a usually fatal systemic disease in which the amyloid deposits are mostly comprised of the N-terminal 1-76 fragment of SAA. SAA isoforms may differ across species in their ability to cause AA amyloidosis, and the mechanism of pathogenicity remains poorly understood. In vitro studies have shown that SAA is a marginally stable protein that folds into various oligomeric species at 4 °C. However, SAA is largely disordered at 37 °C, reminiscent of intrinsically disordered proteins. Non-pathogenic murine (m)SAA2.2 spontaneously forms amyloid fibrils in vitro at 37 °C whereas pathogenic mSAA1.1 has a long lag (nucleation) phase, and eventually forms fibrils of different morphology than mSAA2.2. Remarkably, human SAA1.1 does not form mature fibrils in vitro. Thus, it appears that the intrinsic amyloidogenicity of SAA is not a key determinant of pathogenicity, and that other factors, including fibrillation kinetics, ligand binding effects, fibril stability, nucleation efficiency, and SAA degradation may play key roles. This chapter will focus on the known structural and biophysical properties of SAA and discuss how these properties may help better understand the molecular mechanism of AA amyloidosis.

Divergent effect of glycosaminoglycans on the invitro aggregation of serum amyloid A.

Serum amyloid A (SAA) is an apolipoprotein involved in poorly understood roles in inflammation. Upon trauma, hepatic expression of SAA rises 1000 times the basal levels. In the case of inflammatory diseases like rheumatoid arthritis, there is a risk for deposition of SAA fibrils in various organs leading to Amyloid A (AA) amyloidosis. Although the amyloid deposits in AA amyloidosis accumulate with the glycosaminoglycan (GAG) heparan sulfate, the role GAGs play in the function and pathology of SAA is an enigma. It has been shown that GAG sulfation is a contributing factor in protein fibrillation and for co-aggregating with a plethora of amyloidogenic proteins. Herein, the effects of heparin, heparan sulfate, hyaluronic acid, chondroitin sulfate A, and heparosan on the oligomerization and aggregation properties of pathogenic mouse SAA1.1 were investigated. Delipidated SAA was used to better understand the interactions between SAA and GAGs without the complicating involvement of lipids. The results revealed-to varying degrees-that all GAGs accelerated SAA1.1 aggregation, but had variable effects on its fibrillation. Heparan sulfate, hyaluronic acid, and heparosan did not affect much the fibrillation of SAA1.1. In contrast, chondroitin sulfate A blocked SAA fibril formation and facilitated the formation of spherical aggregates of various sizes. Interestingly, heparin caused formation of spherical SAA1.1 aggregates of various sizes, vast amounts of thin protofibrils, and few long fibrils of various heights. These results suggest that GAGs may have an intrinsic and divergent influence on the aggregation and fibrillation of HDL-free SAA1.1 invivo, with functional and pathological implications.

A link between inflammation and metastasis: serum amyloid A1 and A3 induce metastasis, and are targets of metastasis-inducing S100A4

S100A4 is implicated in metastasis and chronic inflammation, but its function remains uncertain. Here we establish an S100A4-dependent link between inflammation and metastatic tumor progression. We found that the acute-phase response proteins serum amyloid A (SAA) 1 and SAA3 are transcriptional targets of S100A4 via Toll-like receptor 4 (TLR4)/nuclear factor-κB signaling. SAA proteins stimulated the transcription of RANTES (regulated upon activation normal T-cell expressed and presumably secreted), G-CSF (granulocyte-colony-stimulating factor) and MMP2 (matrix metalloproteinase 2), MMP3, MMP9 and MMP13. We have also shown for the first time that SAA stimulate their own transcription as well as that of proinflammatory S100A8 and S100A9 proteins. Moreover, they strongly enhanced tumor cell adhesion to fibronectin, and stimulated migration and invasion of human and mouse tumor cells. Intravenously injected S100A4 protein induced expression of SAA proteins and cytokines in an organ-specific manner. In a breast cancer animal model, ectopic expression of SAA1 or SAA3 in tumor cells potently promoted widespread metastasis formation accompanied by a massive infiltration of immune cells. Furthermore, coordinate expression of S100A4 and SAA in tumor samples from colorectal carcinoma patients significantly correlated with reduced overall survival. These data show that SAA proteins are effectors for the metastasis-promoting functions of S100A4, and serve as a link between inflammation and tumor progression.

Expression of serum amyloid A in uterine cervical cancer

BackgroundAs an acute-phase protein, serum amyloid A (SAA) is expressed primarily in the liver. However, its expression in extrahepatic tissues, especially in tumor tissues, was also demonstrated recently. In our study, we investigated the expression of SAA in uterine cervical carcinomas, and our results suggested its potential as a serum biomarker.MethodsQuantitative real-time polymerase chain reaction (RT-PCR), immunohistochemistry (IHC) and enzyme-linked immunosorbent assay (ELISA) were used to evaluate the SAA gene and protein expression levels in the tissues and sera of patients with non-neoplastic lesions (NNLs), cervical intraepithelial neoplasia (CIN) and cervical carcinoma (CC).ResultsCompared with NNLs, the SAA gene (SAA1 and SAA4) expression levels were significantly higher in uterine CC (mean copy numbers: 138.7 vs. 5.01, P < 0.000; and 1.8 vs. 0.079, P = 0.001, respectively) by real-time PCR. IHC revealed cytoplasmic SAA protein staining in tissues from adenocarcinoma and squamous cell carcinoma of the cervix. The median serum concentrations (μg/ml) of SAA were 6.02 in patients with NNLs and 10.98 in patients with CIN (P = 0.31). In contrast, the median serum SAA concentration was 23.7 μg/ml in uterine CC patients, which was significantly higher than the SAA concentrations of the NNL group (P = 0.002) and the CIN group (P = 0.024).ConclusionsOur data suggested that SAA might be a uterine CC cell product. High SAA concentrations in the serum of CC patients may have a role in monitoring disease occurrence and could have therapeutic applications.Virtual slidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1433263219102962.

Immunomodulatory factors and infectious agents associated with the hepatic gene expression of the IGF system in nursery pigs

Recent findings suggest there is a complex interaction between the IGF system and the inflammatory immune response. The objective of this study was to determine whether gene expression of growth factors (IGF-1, IGF-binding protein-3 (IGFBP-3) and growth hormone receptors (GHR)) in the liver is associated with gene expression of immunomodulators in the liver, including C-reactive protein (CRP), serum amyloid A (SAA), haptoglobin (Hp), interferon-α (IFN-α), IFN-γ, tumour necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, IL-10 and IL-18, as well as with the presence of Salmonella spp., Lawsonia intracellularis, Brachyspira spp., enterotoxigenic Escherichia coli, methicillin-resistant Staphylococcus aureus, swine influenza virus, and porcine reproductive and respiratory syndrome virus (PRRSV) in nursery pigs (n=74) from commercial farms (n=4). Gene expression was quantified using reverse transcription quantitative-PCR (RT-qPCR) and the data were modelled using logistic regression methods. Pigs with elevated IGF-1 expression were less likely to have increased expression of TNF-α (odds ratio (OR)=0.14, P<0.01) and IL-18 (OR=0.19, P<0.05), and less likely to be colonized with PRRSV (OR=0.03, P<0.01). Pigs with increased expression of IGFBP-3 were more likely to have elevated IL-6 expression (OR=8.5, P<0.05). It was also observed that IGFBP-3 and IGF-1 were significantly associated when Hp expression was low (OR=30, P<0.05), but this association was not significant when Hp expression was high (P=0.54). Pigs with increased expression of GHR were less likely to have elevated expression of SAA (OR=0.01, P<0.05) and IL-1β (OR=0.03, P<0.05), but more likely to have increased expression of CRP (OR=290, P<0.01). Overall, there appears to be an inverse association between the hepatic expression of the IGF system (IGF-1, IGFBP-3, GHR) and certain cytokines (IL-1β, IL-18, TNF-α) and acute-phase proteins (SAA, Hp).

Deficits in serum amyloid A contribute to increased neonatal mortality during murine listeriosis

To understand the increased susceptibility of preterm neonates to infection. A murine listeriosis model using immunohistochemistry, microarray technology, and real-time polymerase chain reaction (PCR). We report that recombinant serum amyloid A (SAA) administered prophylactically 18 h before intraperitoneal (i.p.) inoculation with Listeria monocytogenes conferred a dramatic survival benefit compared with administration of only vehicle in neonatal mice. Neonates that received the recombinant SAA protein had significantly fewer Listeria colony counts on plating of infected liver and showed significantly more activated macrophages, but SAA did not affect postnatal growth. Real-time PCR was used to confirm the microarray findings that gene expression levels for the SAA proteins 1 (Saa1) and 2 (Saa2), in addition to that for orosomucoid-2 (Orm2), were strikingly elevated in the adult compared with those in the neonate. Real-time PCR analysis showed that of the acute phase cytokines, tumor necrosis factor (TNF) gene expression increased exponentially with time in the infected adult, whereas neonates did not show similar increases. The increased susceptibility of neonatal mice to listeriosis is in part mediated by a deficiency in the acute phase response, specifically expression of SAA, and that prophylactic SAA protein before neonatal murine listeriosis results in more macrophage activation, lower Listeria counts, and greater survival.

Serum amyloid A, phospholipase A2-IIA and C-reactive protein as inflammatory biomarkers for prostate diseases

Serum amyloid A (SAA), secreted group IIA phospholipase A₂ (sPLA₂-IIA), and C-reactive protein (CRP) are acute-phase proteins whose serum concentrations increase not only during inflammatory disorders, but also in the course of malignant diseases. In this study we analyzed serum levels of these inflammatory markers along with prostate-specific antigens (PSA) in patients with benign prostatic hyperplasia (BPH, n = 55), localized prostate cancers (PCa, n = 55), and metastatic prostate cancers (mPCa, n = 27) using immunological assays. We found that in comparison to healthy individuals (n = 55), patients with BPH, PCa and mPCa have elevated serum levels of SAA, sPLA₂-IIA, and CRP, in addition to elevated levels of PSA. Significant differences with respect to inflammatory biomarkers were found between localized and metastatic PCa (p < 0.001), suggesting a prognostic value of these parameters. In addition, serum concentrations of SAA and sPLA₂-IIA positively correlate with CRP in BPH patients (p < 0.05) and in patients with PCa and mPCa (p < 0.001), but not with PSA levels, Gleason score, or tumor stage, emphasizing a role of SAA and sPLA₂-IIA as circulating biomarkers of inflammation rather than of neoplastic transformation. In contrast to PSA, which differed significantly between BPH and localized PCa patients (p < 0.01), such a difference was not found for SAA, sPLA₂-IIA, and CRP. In order to elucidate whether the elevated levels of SAA and sPLA₂-IIA can be caused by cancer cell-associated synthesis, in vitro studies were performed. These analyses demonstrated the expression of SAA and sPLA₂-IIA in LNCaP and PC-3 prostate cell lines, which can be further upregulated by pro-inflammatory cytokines in a cell type-dependent manner. This might suggest that, in addition to the hepatic origin, SAA and sPLA₂-IIA can also be synthesized and secreted by prostatic cancer tissue itself. The results of the present study emphasize the utility of SAA, sPLA₂-IIA, and CRP as circulating biomarkers of inflammation during BPH development and PCa progression.

Serum Amyloid A: Expression Throughout Human Ovarian Folliculogenesis and Levels in Follicular Fluid of Women Undergoing Controlled Ovarian Stimulation

Serum amyloid A (SAA) is an acute phase protein expressed primarily in the liver in response to various injuries and inflammatory stimuli and is recognized as a modulator of inflammation. Ovarian reproductive functions including folliculogenesis and ovulation use inflammatory processes; thus, studying SAA in this context is of interest. We investigated the expression and localization of SAA in ovarian developing follicles and its levels in follicular fluids. Nonradioactive in situ hybridization and immunohistochemical staining were applied on ovarian paraffin tissue sections. ELISA and RT-PCR were applied on follicular aspirates and blood samples from women undergoing controlled ovarian stimulation for in vitro fertilization. Expression of SAA mRNA and protein was found in follicular cells at all stages of follicular development, from primordial and primary follicles through antral follicles and corpora lutea. Expression was observed in granulosa, theca and luteal cells, and oocytes. Expression of SAA was also found in granulosa cells recovered from follicular aspirates. The SAA protein was detected in follicular fluids. Its levels were somewhat lower than in peripheral blood with strong correlation between the two compartments and with significant correlation with patient's body mass index. High follicular fluid SAA levels were associated with reduced pregnancy rate. SAA is locally produced in ovarian developing follicles and is a constituent of follicular fluids, suggesting its role within the follicular environment. Elevated follicular SAA levels are associated with decreased pregnancy rate and may signify lower reproductive performance.

Hypoxia Increases Serum Amyloid A3 (SAA3) in Differentiated 3T3-L1 Adipocytes

Hypoxia has been implicated as a possible cause of adipose tissue inflammation. Furthermore, the acute phase protein serum amyloid A (SAA) has been associated with the modulation of the adipogenic process, and it is well-known that obese individuals have increased levels of SAA. The effect of hypoxia in the expression and production of SAA was examined in murine 3T3-L1 adipocytes. Hypoxia leads to a substantial increase in SAA3 mRNA and protein level, apparently in a time-dependent manner (threefold in 48 h), in fully differentiated 3T3-L1, followed by reestablishment of gene expression to basal levels after 24 h of reoxygenation. Hypoxia-induced SAA may be one of the key molecules to the development of the inflammatory response in adipose tissue.

Interleukin-17A contributes to the expression of serum amyloid A in chronic rhinosinusitis with nasal polyps

The expression and regulation of serum amyloid A (SAA) in patients with chronic rhinosinusitis with nasal polyps (CRSwNP) have not been well documented. This study enrolled 24 CRSwNP patients and 19 controls to evaluate the expression of SAA in polyp tissues in Chinese adult patients and investigate underlying mechanism. The levels of SAA and interleukin (IL)-17A and myeloperoxidase (MPO) in nasal tissues were detected using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. In addition, the mRNA expression of SAA was examined in cultured polyp epithelial cells (PECs) in the presence of various cytokines (IFN-γ, IL-4, IL-5 and IL-17A) using qRT-PCR, and the role of extracellular signal-related kinase (ERK) signalling in SAA expression was evaluated by western blot analysis. We found the levels of SAA, IL-17A and MPO were significantly upregulated in polyp tissues compared with the controls (p < 0.05), and significant correlations between SAA and IL-17A mRNA levels, as well as between SAA and MPO protein levels, were observed in polyp tissues (p < 0.05). In the in vitro culture system, IL-17A was found to significantly increase SAA mRNA expression in PECs via ERK signaling pathway, in a time- and dose-dependent manner (p < 0.05). Our results suggested a regulatory mechanism underlying excessive SAA production in polyp tissues, which might gain more insights into the pathophysiology of CRSwNP in Chinese adult patients.

Effect of immunomodulatory therapy on the endometrial inflammatory response to induced infectious endometritis in susceptible mares

The objective of the present study was to evaluate the effect of immunomodulatory therapy (glucocorticoids (GC) and mycobacterium cell wall extract (MCWE)) on the endometrial gene expression of inflammatory cytokines in susceptible mares with induced infectious endometritis. Endometrial gene expression of pro- and anti-inflammatory cytokines; interleukin (IL)-1β, IL-6, IL-8, IL-10, tumor necrosis factor (TNF)-α, IL-1 receptor antagonist (ra), acute phase protein (APP) serum amyloid A (SAA) and clinical parameters were evaluated. Five mares were classified as susceptible to persistent endometritis based on their endometrial histopathology and ability to clear an induced uterine inflammation. To investigate the effect of immunomodulatory therapy, the mares were inoculated with 105 colony forming units (CFU) Escherichia coli in three consecutive estrus cycles in a modified cross-over study design. Thus, each mare served as its own control and the treatment type was performed in randomized order. The effect of treatment with MCWE (1.5 mg Settle IV), dexamethasone (0.1 mg per kg IV) or no treatment was investigated. All mares were free from uterine inflammation before each E. coli inoculation. Endometrial biopsies were recovered 3, 24 and 72 h post inoculation. Relative gene-expression analyses were performed by quantitative reverse transcriptase PCR (qRT-PCR). Endometrial gene expression of inflammatory cytokines was modulated by administration of GC. Expression of proinflammatory cytokines (IL-1β, IL-6, IL-8) and SAA was significantly lower in the GC treated group late in the study period (72 h) compared to “no treatment” and MCWE treatment. Increased expression of the anti-inflammatory cytokine IL-10 was observed 3 and 24 h after E. coli infusion and GC treatment. A significant decrease of SAA expression was observed after MCWE treatment compared to “no treatment”. MCWE and GC treatment had a significant effect on the clearance of uterine pathogens and number of mares retaining fluid after E. coli infusion. The results of the current investigation suggest that GC is capable of effectively modulating the innate immune response to induced infectious endometritis in susceptible mares.

Abstract 167: PLTP Acts in Concert with SAA to Reduce Plasma HDL Levels in Mice and Modulates Nascent HDL Formation in Hepatocytes

Phospholipid transfer protein (PLTP), which binds phospholipids and facilitates their transfer between lipoproteins in plasma, plays a key role in lipoprotein remodeling. PLTP levels increase during acute inflammation and increased PLTP activity is associated with the inflammatory markers C-reactive protein (CRP) and serum amyloid A (SAA) as seen in patients with type 2 diabetes and cardiovascular disease. SAA, an HDL apolipoprotein, is highly induced during inflammation. In this study we investigated whether HDL remodeling by PLTP is affected by SAA, and the significance of PLTP on HDL biogenesis. Over-expression of PLTP in mice using an adenoviral vector reduced HDL-C and phospholipid (PL) in a dose dependent manner with the liberation of lipid-poor apoA-I. Co-expression of PLTP and SAA produced a significantly greater reduction in HDL-C and PL than expression of either PLTP or SAA alone. Studies were carried out to examine the lipidation of apoA-I and formation of nascent HDL particles by primary mouse hepatocytes. Over-expression of PLTP in hepatocytes markedly affected the levels as well as the species of nascent HDL particles produced by cells. Although the formation of smaller nascent HDLs was reduced by PLTP, the formation of the larger HDLs was increased. PLTP expression in hepatocytes did not change ABCA1 levels. Co-expression of SAA with PLTP exerted only a modest effect on the levels and types of HDL generated in the presence of PLTP. Our findings suggest that the remodeling of plasma HDL by PLTP and SAA contributes to the reduced HDL-C levels observed during inflammation. PLTP also significantly affects apoA-I lipidation and nascent HDL biogenesis in hepatocytes.

Uropathogenic Escherichia coli Induces Serum Amyloid A in Mice following Urinary Tract and Systemic Inoculation

Serum amyloid A (SAA) is an acute phase protein involved in the homeostasis of inflammatory responses and appears to be a vital host defense component with protective anti-infective properties. SAA expression remains poorly defined in many tissues, including the urinary tract which often faces bacterial challenge. Urinary tract infections (UTIs) are usually caused by strains of uropathogenic Escherichia coli (UPEC) and frequently occur among otherwise healthy individuals, many of whom experience bouts of recurrent and relapsing infections despite the use of antibiotics. To date, whether SAA is present in the infected urothelium and whether or not the induction of SAA can protect the host against UPEC is unclear. Here we show, using mouse models coupled with immunofluorescence microscopy and quantitative RT-PCR, that delivery of UPEC either directly into the urinary tract via catheterization or systemically via intraperitoneal injection triggers the expression of SAA. As measured by ELISA, serum levels of SAA1/2 were also transiently elevated in response to UTI, but circulating SAA3 levels were only up-regulated substantially following intraperitoneal inoculation of UPEC. In in vitro assays, physiological relevant levels of SAA1/2 did not affect the growth or viability of UPEC, but were able to block biofilm formation by the uropathogens. We suggest that SAA functions as a critical host defense against UTIs, preventing the formation of biofilms both upon and within the urothelium and possibly providing clinicians with a sensitive serological marker for UTI.

Investigation of serum amino acid and serum amyloid A concentrations in chickens with amyloid arthropathy

Background: Increased proteolytic cleavage of serum amyloid A (SAA) may potentially contribute to the development of AA amyloid deposition Objective: To study the possible relationship between amyloid artropathy and expression of SAA and some serum amino acids. Animals and methods: Values of 15 serum amino acids and SAA were investigated in chickens with experimentally induced amyloid arthropathy. Thirty-four, 5-week-old chicks were allocated into two groups: one group was injected intra-articularly with 0.25 mL complete Freund's adjuvant at the left tibio-metatarsal joint to induce amyloid arthropathy, whereas the other group served as control. All pullets were necropsied 13 weeks after injection. Collected tissue samples were examined histopathologically. Blood samples were collected and SAA concentrations were measured with enzyme-linked immunosorbent assay. High-performance liquid chromatography was used to assess the amino acid concentrations in serum. Results: Amyloid accumulation in joints occurred only in the experimental group (89%). SAA concentrations of 166 ± 17 and 423 ± 39 (SD) ng/mL were found in the control and experimental groups, respectively (p < 0.001). In the experimental group, an increase was observed in all examined amino acid concentrations except for citrulline. The most significant (p < 0.001) increases were noticed in serine (from 159 ± 15 to 360 ± 29 µmol/L), glycine (from 151 ± 20 to 279 ± 16 µmol/L), isoleucine (from 48 ± 2 to 80 ± 6 µmol/L), and phenylalanine (from 49 ± 2 to 90 ± 3 µmol/L). Conclusion: The results of this study suggest that there is a positive correlation between some serum amino acid values, especially serine, glycine, isoleucine, and phenylalanine, and the high concentrations of SAA in chickens with amyloid arthropathy.

Widespread expression of SAA and Hp RNA in bovine tissues after evaluation of suitable reference genes

The serum amyloid A (SAA) and haptoglobin (Hp) are the most prominent acute phase proteins (APPs) in cow. Liver mainly produces APPs, but extra hepatic expression has also been demonstrated in some tissues. The major aim of the present study was to assess the constitutive SAA and Hp mRNA expression by quantitative PCR (qPCR) in a wide panel of 33 bovine tissues, including gastrointestinal tract, respiratory system, urogenital system, mammary gland, hematopoietic system, central nervous system, eye, thyroid and heart. Normalization of gene expression in different samples requires reference genes, which are stably expressed. Therefore, seven reference genes were investigated (ACTB, GAPDH, HMBS, SDHA, YWHAZ, SF3A1, EEF1A2) and three genes, namely SF3A1, HMBS and ACTB, were selected after assessing their stability with geNorm™ and NormFinder© softwares.The qPCR analysis confirmed liver as the principal source of SAA and Hp, but also identified both APPs’ mRNA in almost all tissues.The highest expression rate of SAA was found in thyroid, followed by pancreas and submandibulary gland. Hp mRNA expression was detected at high concentration in pancreas and submandibulary gland.The present data indicated a widespread expression of SAA and Hp also in non pathological conditions, thus envisaging a possible role as immunomodulatory and protective molecules. To understand where SAA and Hp come from is the prerequisite to their utilization as Acute Phase Reaction biomarkers.

Nascent HDL formation by hepatocytes is reduced by the concerted action of serum amyloid A and endothelial lipase

Inflammation is associated with significant decreases in plasma HDL-cholesterol (HDL-C) and apoA-I levels. Endothelial lipase (EL) is known to be an important determinant of HDL-C in mice and in humans and is upregulated during inflammation. In this study, we investigated whether serum amyloid A (SAA), an HDL apolipoprotein highly induced during inflammation, alters the ability of EL to metabolize HDL. We determined that EL hydrolyzes SAA-enriched HDL in vitro without liberating lipid-free apoA-I. Coexpression of SAA and EL in mice by adenoviral vector produced a significantly greater reduction in HDL-C and apoA-I than a corresponding level of expression of either SAA or EL alone. The loss of HDL occurred without any evidence of HDL remodeling to smaller particles that would be expected to have more rapid turnover. Studies with primary hepatocytes demonstrated that coexpression of SAA and EL markedly impeded ABCA1-mediated lipidation of apoA-I to form nascent HDL. Our findings suggest that a reduction in nascent HDL formation may be partly responsible for reduced HDL-C during inflammation when both EL and SAA are known to be upregulated.

Serum amyloid A protein and hypoxic ischemic encephalopathy in the newborn

Serum amyloid A (SAA) is an acute phase inflammatory marker that is closely associated with ischemic injuries. Its expression in neonatal hypoxic ischemic encephalopathy (HIE) has not been studied. To test the hypothesis that SAA is increased in neonatal HIE and its concentration correlates with the severity of encephalopathy. We conducted a prospective case-control study on 54 full-term neonates; 27 cases with evidence of perinatal compromise and 27 healthy controls. Blood samples were collected from cases and controls at postnatal day 1 and day 7, and SAA was measured by ELISA. SAA concentrations (μg ml(-1)) were significantly increased in cases when compared with controls at day 1 and at day 7 (P<0.001). SAA concentrations at day 1 were greater in cases who died when compared with those who survived, and correlated significantly with the severity of HIE (146.9 ± 56.4 vs 79.8 ± 24.7 vs 58.1 ± 21.5) in severe, moderate and mild HIE, respectively (P=0.001). The expression of SAA is increased in response to hypoxia ischemia of the neonate. The increased concentration correlates with the severity of encephalopathy and is associated with mortality. This is the first study of SAA in neonatal HIE.

Validation of a liver adenoma classification system in a tertiary referral centre: Implications for clinical practice

A molecular and pathological classification system for hepatocellular adenomas (HCA) was recently introduced and four major subgroups were identified. We aimed to validate this adenoma classification system and to determine the clinical relevance of the subtypes for surgical management. Paraffin fixed liver tissue slides and resection specimens of patients radiologically diagnosed as HCA were retrieved from the department of pathology. Immunostainings included liver-fatty acid binding protein (L-FABP), serum amyloid A (SAA), C-reactive protein (CRP), glutamine synthetase (GS) and β-catenin. From 2000 to 2010, 58 cases (71 lesions) were surgically resected. Fourteen lesions were diagnosed as focal nodular hyperplasia with a characteristic map-like staining pattern of GS. Inflammatory HCA expressing CRP and SAA was documented in 36 of 57 adenomas (63%). Three of these inflammatory adenomas were also β-catenin positive as well as GS positive and only one was CRP and SAA and GS positive. We identified eleven L-FABP-negative HCA (19%) and four β-catenin positive HCA (7%), without expression of CRP and SAA and with normal L-FABP staining, one of which was also GS positive. Six HCA were unclassifiable (11%). In three patients multiple adenomas of different subtypes were found. Morphology and additional immunohistochemical markers can discriminate between different types of HCA in>90% of cases and this classification, including the identification of β-catenin positive adenomas may have important implications in the decision for surveillance or treatment. Interpretation of nuclear staining for β-catenin can be difficult due to uneven staining distribution or focal nuclear staining and additional molecular biology may be required.