Short-term Evaluation of Serum Amyloid A after Exercise in Clinically Healthy Horses

Serum amyloid A (SAA), the major equine acute-phase protein, is often measured after the race to investigate whether poor performances could depend on inflammation. The aim of this study was to assess whether there is an increase in concentration of SAA in serum samples collected from 12 clinically healthy Standardbred horses 1 hour after a standard race. Exercise induced an increase in red blood cells, hematocrit, and total proteins but not in SAA. However, a two- to threefold increase of SAA concentration as compared with prerace values was found in three horses. In conclusion, the concentration of SAA in most of the samples collected 1 hour after the race remains unchanged as compared with prerace samples. However, individual variability in response to exercise exists. The evaluation of SAA immediately after the race is not clinically useful.     

Racing Induces Changes in the Blood Concentration of Serum Amyloid A in Thoroughbred Racehorses

Intensive exercise results in the increased blood concentration of the acute phase proteins in horses competing in some sport disciplines. In this study, the blood level of serum amyloid A (SAA) was analyzed in Thoroughbred racehorses during 5 days after completion of the race. Samples were collected from 25 healthy Thoroughbred horses beginning with 4 hours after the race and repeated daily up to the fifth day after the race. Serum amyloid A analysis was performed using commercial enzyme-linked immunosorbent assay kit, and the results were presented as median, interquartile range (IQR), and range. Data were analyzed using Friedman's nonparametric analysis of variance. The acute phase response (APR) was reflected by an increased SAA level after the race, reaching significantly higher concentrations on days 1 (P < .001) and 2 (P = .005) and falling below the level of the first sample on day 5 (P = .006). The median peak concentration observed on day 1 after the race was 3.84 mg/L (IQR, 2.32 to 8.86). Racing induces minute changes in SAA concentration typical for the exercise-induced APR; however, the significance of this reaction in the context of horse health and fitness remains unclear.     

Evaluation of a Commercially Available Human Serum Amyloid A (SAA) Turbidimetric Immunoassay for Determination of Feline SAA Concentration

Serum amyloid A (SAA) is an acute-phase protein in cats likely to be useful for diagnosing and monitoring inflammatory diseases, especially if rapid, reliable and automated assays can be made available. A commercially available automated human SAA turbidimetric immunoassay (SAA-TIA) was evaluated for determination of SAA in cats. Intra-assay and inter-assay imprecisions were in the ranges 2.1–9.9% and 7.0–12.5%, respectively, and without significant inaccuracy. Eighty-eight cats were divided into groups according to (A) the presence or absence of an acute-phase response (APR) (n = 23 and 65, respectively) and (B) clinical diagnosis (clinically healthy cats, cats diagnosed with inflammatory/infectious diseases, endocrine/metabolic diseases, neoplastic diseases, and miscellaneous disorders (n=43, 13, 8, 4 and 20, respectively)). The observed SAA concentrations were, as expected, different for (A) cats with and without an APR and (B) cats with inflammatory/infectious diseases compared to other diagnostic groups, except neoplastic diseases. In conclusion, the SAA concentration in cats could be measured reliably using the commercially available TIA designed for measuring human SAA, which should facilitate implementation of the parameter for routine diagnostic purposes. Hansen, A.E., Schaap, M.K. and Kjelgaard-Hansen, M., 2006. Evaluation of a commercially available human serum amyloid A (SAA) turbidimetric immunoassay for determination of feline SAA concentration. Veterinary Research Communications, 30(8), 863–872     

Comparison of Serum Amyloid A in Horses With Infectious and Noninfectious Respiratory Diseases

The acute phase protein serum amyloid A (SAA) has been shown to be a useful inflammatory parameter in the horse, but studies showing SAA responses to specific respiratory disease etiologies are limited. The goal of this study was to evaluate SAA responses in horses with infectious and noninfectious respiratory diseases as well as healthy, control horses. Two hundred seven horses were grouped into the following categories: equine influenza virus (EIV), equine herpesvirus-4 (EHV-4), Streptococcus equi subspecies equi (S. equi ss equi), inflammatory airway disease (IAD), and healthy controls. Serum amyloid A concentrations were determined for all horses on serum using a stall-side lateral flow immunoassay test. Serum amyloid A levels were found to be significantly greater for infectious respiratory diseases (EIV, EHV-4, S. equi ss equi) and horses with IAD when compared to control horses. There was a significant difference between viral and bacterial infections and IAD. Although SAA values from horses with S. equi ss equi were significantly greater when compared to horses with viral infections (EIV/EHV-4), the wide range of SAA values precluded accurate classification of the infectious cases. In conclusion, SAA is more reliably elevated with infections of the respiratory tract rather than noninfectious airway conditions. This can facilitate early detection of respiratory infections, help track disease progression, and aid practitioners in making recommendations about proper biosecurity and isolation of potentially contagious horses.     

Serum Amyloid A and Haptoglobin Concentrations and Liver Fat Percentage in Lactating Dairy Cows with Abomasal Displacement

Background: There has been increased interest in measuring the serum concentration of acute phase reactants such as serum amyloid A [SAA] and haptoglobin [haptoglobin] in periparturient cattle in order to provide a method for detecting the presence of inflammation or bacterial infection.
Objectives: To determine whether [SAA] and [haptoglobin] are increased in cows with displaced abomasum as compared with healthy dairy cows.
Animals: Fifty-four adult dairy cows in early lactation that had left displaced abomasum (LDA, n = 34), right displaced abomasum or abomasal volvulus (RDA/AV, n = 11), or were healthy on physical examination (control, n = 9).
Materials and Methods: Inflammatory diseases or bacterial infections such as mastitis, metritis, or pneumonia were not clinically apparent in any animal. Jugular venous blood was obtained from all cows and analyzed. Liver samples were obtained by biopsy in cattle with abomasal displacement.
Results: [SAA] and [haptoglobin] concentrations were increased in cows with LDA or RDA/AV as compared with healthy controls. Cows with displaced abomasum had mild to moderate hepatic lipidosis, based on liver fat percentages of 9.3 ± 5.3% (mean ± SD, LDA) and 10.8 ± 7.7% (RDA/AV). [SAA] and [haptoglobin] were most strongly associated with liver fat percentage, r _s=+0.55 ( P < .0001) and r _s=+0.42 ( P = .0041), respectively.
Conclusions and Clinical Importance: An increase in [SAA] or [haptoglobin] in postparturient dairy cows with LDA or RDA/AV is not specific for inflammation or bacterial infection. An increase in [SAA] or [haptoglobin] may indicate the presence of hepatic lipidosis in cattle with abomasal displacement.     

Serum amyloid A isoforms in serum and synovial fluid in horses with lipopolysaccharide-induced arthritis

The aim of the study was to determine the intraarticular serum amyloid A (SAA) response pattern in horses with inflammatory arthritis. Inflammatory arthritis was induced by injection of lipopolysaccharide (LPS) into the radiocarpal joint of four horses. Serum and synovial fluid (SF) samples were collected before and at 4, 8, 12, 24, 48, 72, 96, and 144 h after injection. Concentrations of SAA were measured by immunoturbidometry, and expression of SAA isoforms was visualized by denaturing isoelectric focusing and Western blotting. The LPS injection caused systemic and local clinical signs of inflammation. Serum amyloid A appeared in serum and SF within 8 h after LPS injection. Isoelectric focusing showed three major SAA bands with apparent isoelectric points (pI) of 7.9, 8.6, and >9.3 in serum and SF. Synovial fluid contained two additional isoforms with highly alkaline apparent pI values (apparent pI value extrapolated from standard curve = 10.0 and 10.2), which were not present in any of the serum samples. In conclusion, intraarticular injection of LPS induced systemic and local inflammatory responses in the horses. By demonstrating SF-specific SAA isoforms the results of the present study suggest that SAA is synthesized locally in the equine inflamed joint, similar to what has been demonstrated in humans previously. The marked local SAA synthesis suggests an important pathophysiological role in inflammatory arthritis.     

Changes in Serum and Urine SAA Concentrations and Qualitative and Quantitative Proteinuria in Abyssinian Cats with Familial Amyloidosis: A Five‐year Longitudinal Study (2009–2014)

Background

Diagnosis of familial amyloidosis (FA) in Abyssinian cats usually is made on postmortem examination.

Hypothesis/Objectives

Sequential analysis of serum SAA (sSAA), urinary SAA (uSAA), urinary protein:creatinine (UPC) ratio, or sodium‐dodecylsulfate agarose gel electrophoresis (SDS‐AGE) may facilitate early identification of cats with FA.

Animals

Twenty‐three Abyssinian cats belonging to cattery A or B (low and high prevalence of FA, respectively).

Methods

Prospective longitudinal study using 109 blood and 100 urine samples collected over 4‐year period every 4 months, if possible, or more frequently in case of illness. Cats that died during study were necropsied. Health status of live cats was checked 5 years after enrollment. Serum amyloid A (sSAA) and urinary SAA (uSAA) were measured using ELISA kit. The UPC ratio and SDS‐AGE also was performed.

Results

Familial amyloidosis was not identified in cattery A, whereas 7/14 cats from cattery B had FA. Serum amyloid A concentrations were not significantly different between cats in catteries A and B or between cats with or without FA, despite frequent peaks in cats from cattery B. Conversely, uSAA was significantly higher in cattery B, especially in the terminal phases of FA. Proteinuria occasionally was found in cats from both catteries, especially in those with FA. Urine protein electrophoresis identified mixed proteinuria only in cats with FA.

Conclusions and Clinical Importance

Serum amyloid A and UPC ratio are not helpful for early identification of Abyssinian cats with FA. Conversely, increases in uSAA with or without mixed proteinuria may be found before onset of clinical signs in cats with FA.     

Investigation of the Usefulness of Serum Amyloid A in Supporting the Diagnosis of Equine Proliferative Enteropathy

The objective of this study was to determine if serum amyloid A (SAA), a major acute-phase protein, could help support the diagnosis of equine proliferative enteropathy (EPE) caused by Lawsonia intracellularis infection in foals. Archived serum samples from 101 foals with enteric signs and hypoproteinemia were available for SAA testing. Based on immunodiagnostics for L. intracellularis, the foals were divided into EPE-suspect (67) and non–EPE-suspect cases (34). Serum amyloid A values ranged from 0 to 2,761 μg/mL (median 466 μg/mL) and from 0 to 2,555 μg/mL (median 192 μg/mL) for the EPE-suspect and the non–EPE-suspect cases, respectively. Although SAA can be measured patient-side and help determine the severity of the underlying inflammatory condition, SAA was unable to consistently support the diagnosis of EPE in hypoproteinemic foals with enteric signs.     

Influence of Disease Process and Duration on Acute Phase Proteins in Serum and Peritoneal Fluid of Horses with Colic

Background

The acute phase proteins (APP) serum amyloid A (SAA), haptoglobin, and fibrinogen are valuable blood biomarkers in equine inflammatory diseases, but knowledge of factors influencing their concentrations in blood and peritoneal fluid (PF) of horses with colic is needed.

Objectives

The objective of this study was to investigate the influence of demographics (age, sex, breed), disease process (simple obstruction, strangulating obstruction, inflammatory), disease location, disease duration, hypovolemia, and admission hospital on concentrations of APP, lactate and white blood cell counts (WBC) in horses with colic admitted to 2 referral hospitals.

Animals

The study included 367 horses with colic admitted at 2 referral hospitals.

Methods

Prospective multicenter observational study of clinical data, as well as blood and PF biomarkers. Associations between biomarker concentrations and clinical variables were analyzed using multivariate linear regression analysis.

Results

Increasing pre‐admission duration of colic was associated with increased concentrations of APP in blood and PF. Blood concentrations of SAA and fibrinogen were associated with disease process (inflammatory, strangulations, simple obstructions) in more colic duration groups (5–12 and >24 hours) than any of the other biomarkers. No relevant associations between demographic factors, hospital, or hydration status and the measured biomarkers were found.

Conclusions and Clinical Importance

In horses with colic, concentrations of APP are associated mainly with disease process and duration of colic and may thus be used for assessment of disease independently of demographic or geographic factors. Serum amyloid A may be a diagnostic marker for use in colic differential diagnosis, but further evaluation is needed.     

Serum amyloid A is not a marker for relapse of multicentric lymphoma in dogs

BACKGROUND: Serum amyloid A (SAA) is an acute phase protein whose concentration increases in inflammatory, infectious, and neoplastic conditions in animals and human beings. Multicentric lymphoma is a common cancer in dogs, and chemotherapy is indicated to attain long-term survival. However, frequent relapses lead to changes in chemotherapeutic protocols. OBJECTIVES: The aims of this study were to evaluate SAA as a marker for relapse of multicentric lymphoma in dogs and to determine whether chemotherapy induces changes in the concentration of SAA during treatment. METHODS: SAA was measured by an ELISA test in healthy control dogs (n=20), in healthy dogs receiving chemotherapy (n=8), and in dogs with lymphoma (n=20). All dogs receiving chemotherapy were randomly assigned to 2 treatment groups, one receiving cyclophosphamide, vincristine, and prednisone (CVP) and the other receiving vincristine, cyclophosphamide, methotrexate, and L-asparaginase (VCMA) protocols. SAA concentration was determined before chemotherapy at weeks 1-4 in healthy dogs receiving chemotherapy and in dogs with lymphoma, then every 3 weeks for 4 months in healthy dogs, and at relapse and in the sample prior to relapse in dogs with lymphoma. SAA was measured only once in the healthy control dogs. Results were analyzed using repeated measures ANOVA followed by Tukey multiple comparison tests to compare groups and weeks of treatment. RESULTS: Mean SAA concentration was significantly higher in dogs with lymphoma before chemotherapy compared with healthy and chemotherapy control dogs. No increase in SAA concentration was found at relapse. No differences were observed in SAA concentration based on type of chemotherapy protocol. CONCLUSIONS: SAA is not a marker of relapse in dogs with multicentric lymphoma, nor does chemotherapy regimen affect SAA concentration.     

Serum amyloid A (SAA) mRNA expression in chicken and quails in response to bacterial stress

Monitoring of acute phase proteins such as serum amyloid A at gene expression level may provide quick information about immune status of the host and its susceptibility towards common infections. Present study was carried out to evaluate and compare the mRNA expression of SAA gene in Rhode Island Red chicken (RIR) and Japanese quails using real time PCR analysis in response to inactivated Salmonella gallinarum culture. The results showed that expression of SAA gene was approximately 17–33 folds higher in case of birds administered with bacterial culture when compared to un-inoculated controls and expression was higher and quicker in case of quails than RIR chicken. The SAA genes from chicken and quail were cloned and upon sequence analysis it was observed that deduced amino acid sequence of SAA from chicken and quails were having approximately seven percent variation which might have significance in function of this protein in these species.     

The use of receiver operating characteristic (ROC) analysis to assess the diagnostic value of serum amyloid A, haptoglobin and fibrinogen in traumatic reticuloperitonitis in cattle

Acute phase proteins (APPs) such as serum amyloid A (SAA), haptoglobin (Hp), and plasma fibrinogen (Fb) are diagnostic and prognostic biomarkers. This study determined the pattern of SAA in traumatic reticuloperitonitis (TRP) in cows, and compared the findings with Hp, and plasma Fb concentrations in the differential diagnosis of TRP using receiver operating characteristic (ROC) analysis. Blood samples were collected from 89 cows: 53 cases of TRP, 16 animals with other internal disorders and 20 healthy controls.SAA and Hp had a similar pattern and magnitude of changes in TRP. Using the ROC method, the optimal cut-off for the diagnosis of TRP was 68 μg/mL for SAA and 0.74 g/L for Hp, with 100% sensitivity and 86.1% specificity; either of these parameters may therefore be used as a sensitive and relatively specific tool for the differential diagnosis of TRP in cattle. The diagnostic accuracy of plasma Fb was significantly lower than either SAA or Hp (P < 0.01). The combined use of these APPs (whilst aware of their conditional interdependence) may not provide any additional diagnostic information than can be obtained using either SAA or Hp alone.     

Concentration of Serum Prealbumin (Pr) Protein in Sick Horses and Its Correlation to Blood Leucocyte Count and Albumin Content in Serum

Studies of Pr protein concentrations in sera of sick horses were carried out using Mancini et al.’s (1965) immunodiffusion technique. Relative values against a chosen standard of 100 % were determined for a total of 102 horses. Horses with acute infections had Pr protein values significantly above the normal. The highest individual Pr protein value recorded in this group was 202. Horses suffering from acute laminitis and malignant tumours also had increased Pr protein values.There was a positive correlation between the Pr protein value and the blood leucocyte count and a negative correlation between the Pr protein value and the albumin content in serum.     

Comparison of paired serum and lithium heparin plasma samples for the measurement of serum amyloid A in horses using an automated turbidimetric immunoassay

This study aimed to evaluate whether equine serum amyloid A (SAA) concentrations could be reliably measured in plasma with a turbidimetric immunoassay previously validated for equine SAA concentrations in serum. Paired serum and lithium-heparin samples obtained from 40 horses were evaluated. No difference was found in SAA concentrations between serum and plasma using a paired t test (P = 0.48). The correlation between paired samples was 0.97 (Spearman’s rank P < 0.0001; 95% confidence interval 0.95–0.99). Passing-Bablok regression analyses revealed no differences between paired samples. Bland–Altman plots revealed a positive bias in plasma compared to serum but the difference was not considered clinically significant. The results indicate that lithium-heparin plasma samples are suitable for measurement of equine SAA using this method. Use of either serum or plasma allows for greater flexibility when it comes to sample collection although care should be taken when comparing data between measurements from different sample types.     

Acute phase proteins in cattle after exposure to complex stress

Stressors such as weaning, mixing and transportation have been shown to lead to increased blood concentrations of acute phase proteins (APP), including serum amyloid A (SAA) and haptoglobin, in calves. This study was therefore undertaken to assess whether SAA and haptoglobin levels in blood mirror stress in adult cattle. Six clinically healthy Holstein cows and two Holstein heifers were transported for four to six hours to a research facility, where each animal was housed in solitary tie stalls. Blood samples for evaluation of leukocyte counts and serum SAA and haptoglobin concentrations were obtained before (0-sample) and at 8, 24 and 48 hours after the start of transportation. Upon arrival the animals gave the impression of being anxious, and they appeared to have difficulty coping with isolation and with being tied on the slippery floors of the research stable. Serum concentrations of SAA and haptoglobin increased significantly in response to the stressors (P < 0.01 and 0.05 at 48 hours, respectively). Additionally, the animals had transient neutrophilia at 8 and 24 hours (P < 0.05). In conclusion, the results of the study suggest that SAA and haptoglobin may serve as markers of stress in adult cattle.     

Giardia and Cryptosporidium infections in neonatal reindeer calves: Relation to the acute phase response

This longitudinal observational study was conducted to investigate the spontaneous effect of Giardia and Cryptosporidium infections on acute phase response (APR) in reindeer calves (Rangifer tarandus tarandus) in Finnish Lapland.Serum (n = 609) and faecal samples (n = 366) were collected from 54 reindeer calves aged zero to 33 days. The samples were analysed for Giardia, Cryptosporidium, acute phase proteins (APP) and γ-globulins.Linear regression models were used to investigate associations of early Giardia infection (before 12 days of life) with the response of APPs and acquiring of passive immunity.Giardia was detected in 100% and Cryptosporidium in 23% of calves. There was a negative association between early Giardia infection and γ-globulin concentrations (p = 0.032) and a positive association with serum amyloid A (SAA) concentrations (p = 0.042). The results suggest a protective effect of colostrum against Giardia infection and that early infection may induce activation of APR.     

Analytical validation of commercially available methods for acute phase proteins quantification in pigs

The aim of this study was to validate commercially available methods for porcine haptoglobin (Hp), C-reactive protein (CRP), serum amyloid A (SAA) and major acute phase protein (Pig-MAP) determinations. Intra and inter assay coefficients of variation (CVs) were lower than 20% in all cases with exception of inter assay CVs for CRP and Pig-MAP assays with samples of low acute phase proteins concentration, and for SAA assay at any acute phase proteins concentration. All methods showed good linearity and detection limits were low enough to detect APPs levels in healthy animals. Hp and SAA were very affected by haemolysis. Lipaemia influenced mainly on SAA determination. Over 15-fold increase was observed in CRP and SAA concentrations after artificially induced inflammation by a single subcutaneous dose of turpentine, whereas Hp and Pig-MAP increased less than 5-fold.     

Serum paraoxonase type-1 activity in pigs: Assay validation and evolution after an induced experimental inflammation

Paraoxonase 1 (PON1) is a serum enzyme synthesised and secreted primarily by the liver. It possesses anti-inflammatory properties limiting the production of pro-inflammatory mediators. The objectives of this study were to validate three spectrophotometric assays for the quantification of PON1 activity in pig serum, and to determine if PON1 activity in porcine behaves as a negative acute phase protein (APP), decreasing in inflammatory conditions. An analytical validation using three different substrates – 5-thiobutil butyrolactone (TBBL), phenylacetate (PA) and 4-(p)-nitrophenyl acetate (pNA) – was performed. In addition, inflammation was experimentally induced in five pigs by subcutaneous injection of turpentine oil, while five control pigs were left untreated. The treated pigs showed significant increases in CRP and decreases in albumin, indicating an inflammatory condition. The three substrates used would be suitable for PON1 activity measurements in serum samples, since they offer adequate precision (coefficients of variation < 10%), sensitivity (0.01, 0.15, 0.02 U/mL for TBBL, pNA and PA respectively) and accuracy (r = 0.99). In addition, PON1 behaves as a negative APP in pigs since a significant decrease (P < 0.05) in its activity after 72 h of the induction of the inflammation was observed with all substrates.