At the recent Elanco Immunity Science Symposium held in Vienna, Professor James Roth from Iowa State University described the latest advances in the understanding of the immune system with reference to dairy cows.1 Professor Roth was joined by five other professors from around Europe to review and collate information on this exciting area of research for an invited audience of vets and scientists from around Europe. Their presentations shed new light on what has become a dynamic and rapidly advancing area of veterinary medicine. Around the time of parturition – referred to as the vital 90 days covering the 60 days pre and 30 days post calving – the immune system faces significant challenges and it is common for these animals to be affected by production related diseases at this time. There is potential to improve the veterinary response to these animals by being aware of the degree to which these animals are compromised and relating this to our understanding of disease processes around the periparturient period.
The immune system is made up of innate (native) and adaptive (acquired) responses. The innate system produces an immediate response after being activated by danger signals. Innate responses are not antigen specific and are required to activate adaptive mechanisms. The adaptive response is acquired, taking place 10-14 days after exposure. Adaptive responses are antigen specific and through ‘memory’ activate the innate system. While adaptive responses are based around antibody and cell mediated responses, the innate system consists of natural defensive barriers, phagocytes and neutrophils, natural killer cells, cytokines, complement and antimicrobial peptides. Veterinary clinical thinking often focuses on the adaptive response but interest in the capacity of the innate system is increasing, as more is understood about its unique qualities. In the face of microbial invasion, sentinel cells, such as macrophages and dendritic cells, secrete cytokines – interleukins (IL1, IL6), tumour necrosis factor alpha (TNFα) and HMGB1 (High-mobility group protein B1, an intracellular cytokine). In the case of a weak response, the action is local at the site of infection. A stronger stimulus leads to systemic effects in the liver, brain and bone marrow. The results include increased white cell production to help fight the infection. Neurological effects such as depression or sleepiness encourage the animal to self-isolate, which in nature would reduce the spread of disease – in food producing animals this protective function cannot naturally occur due to management factors. Synthesis of acute phase proteins is fuelled by amino acid release from muscle, in turn reducing live weight gains. The acute phase proteins perform a number of useful functions including microbial inhibition, coagulation and chemotaxis.
Neutrophil structure and function
Neutrophils are important components of the innate system and are released from bone marrow as short lived cells, surviving only a matter of hours. The cells contain lysozyme granules that can kill bacteria and glycogen granules, the cells being able to be fuelled by anaerobic glycolysis allowing them to enter low oxygen areas of the body where there are dead and dying tissues. Neutrophils are known to roll along blood vessel walls, loosely adhering to endothelial cells and moving as a result of the forces imposed by vascular flow. Selectins mediate this rolling action. Integrins allow adherence and once adhered, neutrophils can digest the adhesions between endothelial cells and squeeze through into infected tissue, sensing the chemotactic gradient. Steroids down-regulate expression of integrins and reduce neutrophil adhesion. This is one of the reasons that stress and steroid administration can be detrimental in infected animals. Morphologically, neutrophils have large membranes that facilitate phagocytosis by wrapping around a pathogen. Most bacteria repel this membrane but antibodies opsonise bacteria, making them easier to engulf. Bacterial destruction involves both oxidation and iodination. Professor Roth noted that the neutrophil worked out long before man that bleaching and formaldehyde can kill bacteria. The close interrelationship between the innate and adaptive immune system is also evidenced by the process occurring during antibody dependent cell mediated cytotoxicity (ADCC) which is mediated by natural killer cells, neutrophils and macrophages. In this process antibody binds antigens on the target cell, receptors on the killer cell recognise bound antigen and cross link, in turn triggering activation of the killer cell to destroy the target. A new mechanism described in recent years is that of neutrophil extracellular traps (or NETs).2 This is thought to result in extracellular killing, as a result of the neutrophil digesting its nuclear material and throwing out a sticky mixture of DNA, histones and granular proteins to engulf the pathogen. NETS offer some prospects to better understand immunity in the periparturient period and Professor Nahum Shpigel elaborated on Professor Roth’s comments, describing how high betahydroxybutyrate levels (BHBA) are associated with reduced NETs functionality.3 Repeated bouts of mastitis and metritis also suggest that immunological ‘memory’ plays little part in the dairy cow’s defences with regard to these conditions.
Neutrophil driven defences
Although neutrophils are a key component of innate defences and have a variety of ways in which they exert their protective effect, the functional capability can be affected by many factors. Physiological states such as the neonatal period, high circulating progesterone levels, stress and the periparturient period can all influence how neutrophils might fulfil their protective function in bovine animals. Up to 5 months of age, calves experience reduced neutrophil function both in terms of lower iodination and lower ADCC responses.4 As has already been noted, stress and steroids can reduce neutrophil adherence by as much as 25 per cent; random migration is increased but the ability to adhere and leave the bloodstream is reduced.5 High progesterone levels produce a very similar picture to high circulating cortisol levels. In contrast, high oestrogen-low progesterone levels are associated with high levels of neutrophil oxidation and iodination and at this point the animal is best positioned to respond effectively to bacterial challenge.6,7 During the periparturient phase, neutrophil function dips at the point of transition before beginning to increase again post parturition. Study data has demonstrated that both IL8 production, chemotaxis of neutrophils to cotyledon supernatant – a process that normally aids expulsion of the placenta – and myeloperoxidase activity of neutrophils, were all lower in cows experiencing retained placenta.8 This suggests that immune function, particularly neutrophil function, is a critical factor in the development of retained placenta. Pathogens can also impact upon neutrophil function. Bovine Viral Diarrhoea (BVD) has been shown to reduce neutrophil iodination and administration of a modified live vaccine of the same agent has the same effect, reducing iodination by as much as 75 per cent of normal function – an effect that can be compounded by also administering cortisol at the same time.9,10 Capsular material from some bacteria, such as Pasturella and Brucella can also reduce neutrophil iodination.11,12 Being able to increase neutrophil function could clearly have significant benefits for the periparturient cow in particular. Cytokines, interferon gamma and granulocyte colony stimulating factor (GCSF) are potential candidates for this role. Interferon can improve some aspects of neutrophil function in immunosuppressed calves.13 Ultrafiltered colostral whey has also been shown to increase neutrophil function in both cows and periparturient cows.14 This may be attributable to passive transfer of cytokines, rather than antibodies (which would be filtered out). GCSF stimulates bone marrow to produce neutrophils and daily administration to periparturient cows has resulted in increased numbers circulating in the blood, reduced random migration and increased ADCC and phagocytosis.15 While antibody-led adaptive immunity has previously been the focus of veterinary intervention, it’s clear that the innate immune system plays an important role in the defences of production animals and is particularly important during the periparturient period. The role of neutrophils is key in responding to infection and reduced function and expression throughout the vital 90 days means that the dairy cow is vulnerable to developing related conditions such as metritis, mastitis and retained placenta. Text: Susan McKay BVM&S, MRCVS, MBA – Illustrations: Elanco
- Elanco Immunity Science Symposium, Vienna 2013, Professor James Roth, Innate vs. adaptive immunity with emphasis on the role of neutrophils
- Lee W.L., Grinstein S., (2004), The Tangled Webs that Neutrophils Weave, Science, 303, 5663, 1477‐1478
- Grinberg N., Elazar S., Rosenshine I., Shpigel N.Y., (2008), β‐Hydroxybutyrate Abrogates Formation of Bovine Neutrophil Extracellular Traps and Bactericidal Activity against Mammary Pathogenic Escherichia coli, Infection and Immunity, 76, 2802‐2807
- Hauser M.A., Koob M.D., & Roth J.A., (1986), Variations of neutrophil function with age in calves, American Journal of Veterinary Research, 47, 152‐153
- Roth J.A., Kaeberle M.L., Hsu W.H., (1982), Effects of ACTH administration on bovine polymorphonuclear leukocyte function and lymphocyte blastogenesis, American Journal of Veterinary Research, 43, 412‐416
- Roth J.A., Kaeberle M.L., Hsu W.H., (1982), Effect of Estradiol and Progesterone on Lymphocyte and Neutrophil Functions in Steers, Infection and Immunopathology, 35, 997‐1002
- Roth J.A., Kaeberle M.L., Appell L.H., Nachreiner R.F., (1983), Association of Increased Estradiol and Progesterone Blood Values with Altered Bovine Polymorphonuclear Leukocyte Function, American Journal of Veterinary Research, 44, 247‐253
- Kimura K., Goff J.P., Kehrli M.E. Jr.,Reinhardt T.A., (2002), Decreased Neutrophil Function as a Cause of Retained Placenta in Dairy Cattle, Journal of Dairy Science, 85, 544‐550
- Roth J A, Kaeberle M L, Griffith Roth J.A., Kaeberle M.L., Griffith R.W., (1981), Effects of Bovine Viral Diarrhea Virus Infection on Bovine Polymorphonuclear Leukocyte Function, American Journal of Veterinary Research, 42, 244‐250
- Roth J.A., Kaeberle M.L., (1983), Suppression of neutrophil and lymphocyte function induced by a vaccinal strain of bovine viral diarrhea virus with and without the administration of ACTH, American Journal of Veterinary Research, 44, 2366‐2372
- Ryu, H., Kaeberle, M.L., Roth, J.A., Griffith, R.W., (1984), Effect of type A Pasteurella multocida fractions on bovine polymorphonuclear leukocyte functions, Infection & Immunity, 43, 66‐71
- Canning, P.C., Roth, J.A., Tabatabai, L.B., Deyoe, B.L., (1985), Isolation of components of Brucella abortus responsible for inhibition of function in bovine neutrophils, Journal of Infectious Diseases, 152, 913‐921
- Roth J.A., Frank D.E., (1989), Recombinant Bovine Interferon‐y as an Immunomodulator in Dexamethasone‐Treated and Non-treated Cattle, Journal of Interferon Research, 9, 143‐151
- Hagiwara K., Kataoka S., Yamamnaka H., Kirisawa R., Iwai, H., (2000), Detection of cytokines in bovine colostrum, Veterinary Immunology and Immunopathology 76, 183‐190
- Kehrli M. E. Jr., Goff J.P., Stevens M.G., Boone T.C., (1991), Effects of Granulocyte Colony‐Stimulating Factor Administration to Periparturient Cows on Neutrophils and Bacterial Shedding, Journal of Dairy Science, 74,2448‐2458