Kinetics of local and systemic isoforms of serum amyloid A in bovine mastitic milk

doi:10.1016/j.vetimm.2004.09.031

Veterinary Immunology and Immunopathology

Volume 104, Issues 1–2, 10 March 2005, Pages 21-31

https://doi.org/10.1016/j.vetimm.2004.09.031 Get rights and content

Abstract

The aim of the present study was to characterise the serum amyloid A (SAA) response to intramammary inoculation of Escherichia coli and to examine the distribution of hepatically and extrahepatically produced SAA isoforms in plasma and milk from cows with mastitis.

Milk and plasma SAA concentrations were determined before and after experimental induction of E. coli mastitis in six dairy cows. The milk SAA response was characterised by low or undetectable levels before inoculation, very rapid and large increases in concentration after inoculation, and rapid decline towards baseline levels after resolution of disease.

In plasma from cows with experimentally induced E. coli mastitis, four hepatically derived SAA isoforms with apparent isoelectric point (pI) values of 5.8, 6.2, 6.8 and 7.4 were demonstrated by denaturing isoelectric focusing. In milk three highly alkaline isoforms with apparent pI values above 9.3 appeared 12 h post-inoculation. These isoforms were not present in any of the plasma samples, and it therefore seems likely that they were locally produced, tissue-specific isoforms. At 24–36 h post-inoculation one or more acidic isoforms corresponding to those found in plasma appeared in the milk samples.

The isoforms demonstrated in plasma from cows with E. coli mastitis were also present in serum obtained from three cows with clinical Streptococcus uberis mastitis.

In conclusion, experimentally induced E. coli mastitis is accompanied by a prominent SAA response. The results of the present study indicate that SAA accumulation in mastitic milk is the result of both local synthesis of SAA and of hepatically derived SAA gaining access to the milk due to increased permeability of the blood–milk barrier.

Introduction

In veterinary medicine the acute phase protein serum amyloid A (SAA) is usually mentioned in relation to its hepatic production and its rapidly increasing serum concentrations during infections or inflammatory diseases. However, SAA has recently been shown also to be present in normal bovine colostrum (McDonald et al., 2001) and in milk from cows suffering from mastitis (Eckersall et al., 2001, Nielsen et al., 2004). Extrahepatic production of SAA in the healthy bovine mammary gland has been demonstrated by cloning of cDNA from mammary epithelial cells (Kho et al., 2000, McDonald et al., 2001), but the exact nature of the milk SAA protein and the regulation of its synthesis during episodes of mastitis have not yet been elucidated. In laboratory rodents, synthesis of one member of the SAA protein family, SAA3, has been shown to take place in a variety of cell types other than hepatocytes during acute phase states (Rokita et al., 1987, Meek et al., 1992, Marhaug et al., 1997, Vallon et al., 2001). In contrast with SAA1 and SAA2, which are primarily produced in the liver and released into the systemic circulation, murine SAA3 is found in only minute amounts in the high-density lipoprotein fraction of serum (Husby et al., 1994), which has lead to the suggestion that this isoform possesses a predominantly local function. Analogously, local production of SAA in the bovine mammary gland may play a role in the innate protection against mastitis pathogens. Insight into factors related to innate immunity in the bovine udder may promote the understanding of the pathogenesis of mastitis and improve knowledge about factors important for individual susceptibility towards mastitis pathogens.

The aims of the present study were (1) to examine the SAA response to intramammary inoculation of Escherichia coli; (2) to examine the presence of hepatically and extrahepatically produced SAA isoforms in plasma/serum and milk during episodes of mastitis in dairy cows; (3) to describe the time-sequence of production of the isoforms; and (4) to study the variation in synthesis of the different SAA isoforms between individual cows.

Section snippets

Animals and samples

Plasma and milk samples were obtained from six dairy cows (cows A–F) before and after intramammary inoculation with 50 CFU of a non-verotoxic, encapsulated strain of E. coli 0:157 isolated from a case of clinical mastitis (originally described by Lohuis et al., 1990a, Lohuis et al., 1990b) as described by Kornalijnslijper et al. (2003). The cows were in their second or third lactation, and they were 100% Holstein–Friesian, except cow D, which was a 50% Holstein–Friesian crossbreed. The cows

Signs of mastitis after inoculation with E. coli

All cows developed clinical mastitis with local signs (swelling, increased firmness of the inoculated quarter, increased SCC, and changes in milk appearance) and systemic signs (fever, anorexia, and depression). Severity of mastitis varied between cows: cows A and B developed mild mastitis, cows C and D developed moderate mastitis, and cows E and F developed severe mastitis (Table 1). The infection was self-limiting in all cows, and bacteria were cleared from the udder within 42 to 240 h without

Serum amyloid A response to intramammary inoculation with E. coli

Prior to inoculation with E. coli plasma and milk contained no or very low amounts of SAA. Low concentrations of SAA and other acute phase proteins have often been demonstrated in plasma and milk of healthy cattle (Alsemgeest et al., 1994, Werling et al., 1996, Heegaard et al., 2000, Eckersall et al., 2001). This has generally been considered to be within physiological limits (Saini et al., 1998).

Milk SAA concentrations increased approximately 80 times within 12 h post-inoculation, whereas the

Conclusions

In conclusion, the present study demonstrates several SAA isoforms in both plasma/serum and milk from cows with mastitis. Some SAA isoforms were only present in mastitic milk and never appeared in plasma/serum, thus suggesting that they might be a result of intramammary SAA synthesis. Serum amyloid A accumulation in milk commenced within few hours of inoculation and SAA levels seemed to reflect severity of infection. These characteristics of milk SAA suggest that this protein holds a potential

Acknowledgements

This research was supported through a European Community Marie Curie Research Fellowship. The authors are solely responsible for the information published. It does not represent the opinion of the Community, and the Community is not responsible for any use that might be made of data appearing therein. Bodil H. Nielsen, D.V.M., is thanked for her kind donation of the serum samples from cows with S. uberis mastitis, and Tridelta Development Ltd., Greystones Co., Wicklow, Ireland is thanked for

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Several quantitative trait loci (QTL) affecting mastitis incidence and mastitis-related traits such as somatic cell score exist in dairy cows. Previously, QTL haplotypes associated with susceptibility to Escherichia coli mastitis in Nordic Holstein-Friesian (HF) cows were identified on Bos taurus autosome 9. In the present study, we induced experimental E. coli mastitis in Danish HF cows to investigate the effect of 2 E. coli mastitis-associated QTL haplotypes on the cows’ disease phenotypes and recovery in early lactation. Thirty-two cows were divided in 2 groups bearing haplotypes with either low (HL) or high (HH) susceptibility to E. coli. In addition, biopsies (liver and udder) were collected from half of the cows (n = 16), resulting in a 2 × 2 factorial design, with haplotype being one factor (HL vs. HH) and biopsy being the other factor (biopsies vs. no biopsies). Each cow was inoculated with a low E. coli dose (20 to 40 cfu) in one front quarter at time 0 h. Liver biopsies were collected at −144, 12, 24, and 192 h; udder biopsies were collected at 24 h and 192 h post-E. coli inoculation. The clinical parameters: feed intake, milk yield, body temperature, heart rate, respiration rate, rumen motility; and the paraclinical parameters: bacterial counts, somatic cell count (SCC), and milk amyloid A levels in milk; and white blood cell count, polymorphonuclear neutrophilic leukocyte (PMNL) count, and serum amyloid A levels in blood were recorded at different time points post-E. coli inoculation. Escherichia coli inoculation changed the clinical and paraclinical parameters in all cows except one that was not infected. Clinically, the HH group tended to have higher body temperature and heart rate than the HL group did. Paraclinically, the HL group had faster PMNL recruitment and SCC recovery than the HH group did. However, we also found interactions between the effects of haplotype and biopsy for body temperature, heart rate, and PMNL. In conclusion, when challenged with E. coli mastitis, HF cows with the specific Bos taurus autosome 9-located QTL haplotypes were associated with differences in leukocyte kinetics, with low-susceptibility cows having faster blood PMNL recruitment and SCC recovery and a tendency for a milder clinical response than the high-susceptibility cows did.
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Kinetics of local and systemic isoforms of serum amyloid A in bovine mastitic milk

Abstract

Introduction

Section snippets

Animals and samples

Signs of mastitis after inoculation with E. coli

Serum amyloid A response to intramammary inoculation with E. coli

Conclusions

Acknowledgements

Res. Vet. Sci.

Res. Vet. Sci.

Res. Vet. Sci.

Vet. Immunol. Immunopathol.

J. Biol. Chem.

Biochem. Biophys. Acta

Biochem. Biophys. Res. Commun.

J. Dairy Sci.

J. Dairy Sci.

FEBS Lett.

J. Immunol. Methods

Vet. Immunol. Immunopathol.

J. Comp. Path.

Respir. Physiol.

Res. Vet. Sci.

Concentrations of serum amyloid A (SAA) and haptoglobin (Hp) as parameters of inflammatory diseases in cattle

Vet. Q.

First evidence for the existence of multiple isoforms of bovine serum amyloid-A (apoSAA)

Scand. J. Immunol.

Serum amyloid A is a chemoattractant: induction of migration, adhesion, and tissue infiltration of monocytes and polymorhonuclear leukocytes

J. Exp. Med.

Serum amyloid A is an activator of PMN antimicrobial functions: induction of degranulation, phagocytosis, and enhancement of anti-Candida activity

J. Leukoc. Biol.

Serum amyloid A concentrations in cows given endotoxin as an acute-phase stimulant

Am. J. Vet. Res.

Syrian and armenian hamsters differ in serum amyloid A gene expression. Identification of novel syrian hamster serum amyloid A subtypres

J. Immunol.

Over-expression of the muring pIgR gene in the mammary gland of transgenic mice influences the milk composition and reduces its nutritional value

Transgenic Res.

Acute phase proteins in serum and milk from dairy cows with clinical mastitis

Vet. Rec.

Acute phase protein values in blood of cattle as indicator of animals with pathological proceses

Arch. Lebensmittelhygiene