Veterinary Antibodies and Reagents

Bovine Immunity: Cytokines – Interleukin-2 (IL-2)

This article gives an overview of Interleukin 2 (IL-2) focusing on its biology and the function it carries out in the immune response with comparisons to human and mouse. Finally, the role IL-2 plays in various bacterial and viral diseases is briefly examined.

Introduction

IL-2 is a 15 kD four-alpha-helix bundle cytokine. It is a soluble molecule secreted mainly by CD4+ T helper (Th) cells. CD8+ T cells, natural killer (NK) cells, and natural killer T (NKT) cells also produce IL-2 but at lower levels. Dendritic cells (DC) activated by microbial triggers produce small amounts of IL-2.


IL-2 biology

IL-2 binds to the IL receptor (IL-2R) subunit, CD25 (IL-2Rα), inducing a conformational change in IL-2 which increases its affinity for CD122 (IL-2Rβ). This IL-2-CD25 dimer first binds the CD122 subunit followed by binding to the cytokine receptor γ chain (γc, CD132) culminating in a quaternary IL-2-IL-2R complex (a trimeric receptor of CD25-CD122-γc binding the soluble IL-2 molecule). Cells that do not express CD25 can bind soluble IL-2 directly, via the dimeric IL-2R (CD122-γc). Also, IL-2 can bind to CD25 expressed on DCs; this IL-2-CD25 dimer can then be presented to dimeric IL-2R on neighboring T cells, inducing their activation pathways. It is thought that CD25 increases the binding affinity of IL-2 to the dimeric IL-2R which leads to signal transduction. In the absence of CD25, binding affinity is lower and signal transduction is dependent on the cell expressing sufficient levels of dimeric IL-2R.


IL-2 in immune response

During steady-state conditions, the antigen major histocompatibility complex (MHC) class II complexes on DC’s stimulate CD4+ T cells to produce IL-2, which is taken up by CD4+ T cells and CD25+ effector cells (CD8+ T cells and CD4+CD25+ T regulatory (Treg) cells) that are in close proximity. During an immune response, activated DCs expressing CD25 can bind to T cells (CD4+, CD8+ and Treg) or capture DC derived IL-2 to present it to CD25low effector cells or CD8+ T cells (Driesen et al. 2008, Boyman and Sprent 2012).

IL-2 is predominantly secreted by activated T cells in the secondary lymphoid tissues; there it is taken up by these cells and other CD25+ cells, including Treg cells.

The rate of IL-2 turnover in the steady state influences T cell homeostasis. The highest levels of IL-2 producing cells are CD4+ T cells expressing intermediate levels of CD25 followed by CD25lowCD4+ T cells which produce more than CD25hi CD4+ T cells. NKT cells produce less than CD25hi CD4+ T cells but more than NK and CD8+ T cells. During an immune response, antigen-activated CD4+ and CD8+ T cells rapidly produce IL-2. These elevated levels of IL-2 in the secondary lymphoid organs are consumed by CD4+, CD8+, and CD25+ Treg cells. CD25+ Treg cells are important in maintaining systemic IL-2 homeostasis, as depletion of these cells causes an increase in serum IL-2 levels.

Mice deficient in IL-2 (IL-2-/-) have enlarged lymph nodes due to activation and expansion of lymphocytes (lymphadenopathy). Furthermore, IL-2 deficient mice develop the same phenotype and clinical signs (accumulation of lymphocytes in the intestines and ulcerative colitis) as cattle with malignant catarrhal fever (MCF).


IL-2 role in bovine disease


Malignant catarrhal fever (MCF)

MCF is a lymphoproliferative disease of cattle (and other hoofed animals) caused by the gamma-herpes viruses of the genus Macavirus. This disease is characterized by vasculitis and necrosis in a variety of tissues brought about by the infiltration and accumulation of large numbers of CD8+ T cells. Compared with healthy cattle, MCF infected cattle have a significant decrease in IL-2 transcripts (Meier-Trummer et al. 2009, Russel et al. 2012). Administration of low doses of IL-2 can result in clinical recovery of cows from MCF (Braun et al. 2015).

Bovine tuberculosis (TB)

Similar to human and mouse, where multifunctional T cells co-expressing IFN-γ, TNFα and IL-2 provide a protective immunity against infectious diseases (e.g. HIV or Mycobacterium tuberculosis), CD4+ T cells in cattle infected with bovine tuberculosis are also multifunctional and predominately exhibit IFN-γ+IL-2+TNFα+ and IFN-γ+IL-2-TNFα+CD4+ T cells (Whelan et al. 2011).

Cattle infected with bovine TB have measurable levels of antigen specific IL-2, whereas in cattle vaccinated with Mycobacterium bovis (M.bovis) BCG, the IL-2 levels are not detectable. However, when BCG vaccinated cattle are infected with a virulent (pathogenic) form of M.bovis IL-2, production is induced. Therefore, IL-2 levels could be used to differentiate between cattle that are uninfected, infected or vaccinated with BCG (Rhodes et al. 2014).

In mouse and human there are two functionally distinct subpopulations of memory T cells of the CD45RA-CD45RO+ phenotype important in protecting against pathogens; these can be subdivided by their expression of CD62L and CCR7. They are:

  • T cell memory cells (Tcm) (CD62L+CCR7+) which predominantly reside in lymphoid tissue
  • Effector memory T (Tem) cells (CD62L-CCR7-) found in peripheral tissue and blood

Compared with Tem cells, Tcm cells show elevated levels of proliferation and IL-2 production and are able to generate Tem (Sallusto et al. 1999). Assaying the Tcm (CCR7+CD62LhiCD45RO+), Tem (CCR7-CD62Llow/int CD45RO+) and T effector cells (CCR7-CD62L-/lowCD45RO-) response to M.bovis in cattle also shows an elevated proliferative response by the Tcm to the antigen and a reversion to the effector phenotypes (mainly Tem T cells) (Maggioli et al. 2015).

Bovine leukemia virus (BLV)

BLV is a delta-retrovirus that infects cattle’s lymphocytes and mammary epithelial cells and is transmitted by blood and milk. Major difference between BLV and other delta-retroviruses is that BLV mainly targets B cells. Around 30% of infected cattle develop persistent lymphocytosis (PL) caused by polyclonal expansion of CD5+ B cells. CD4+T cells in BLV infected cows with PL have a reduced expression of IL-2 (and IL-4) mRNA (Amills et al. 2002).

It has recently been shown that exposure to bovine leukemia virus is associated with breast cancer in humans. The frequency of BLV DNA in mammary epithelium was found to be significantly higher than in normal controls (Buehring et al. 2015).


IL-2 controls polymorphonuclear neutrophils  functions

Neutrophils are critical in combating bacterial infections (e.g. Staphylococcus aureus), their accumulation at the site of tissue infection (e.g. mastitis), and their production of cytokines enables them to recruit more neutrophils. In periparturient cows a higher incidence of mastitis occurs during the first 2 weeks of calving. Suppression of the immune system (innate and adaptive) in the periparturient cow means that Th1 cytokines (IL-2) are not available to activate neutrophils that are critical in protecting mammary tissue from infection.


Post-partum mammary gland infection

The expression levels of IL-2, IL-4 and IFN-γ mRNA in peripheral blood monocytes is lower in buffaloes that develop postpartum reproductive disorder than those that don’t. Neutrophil function is also impaired, manifested as lower levels of superoxide and hydrogen peroxide production before parturition and during early postpartum. At calving, lower expression levels of IL-2, IFNγ, and IL-4 mRNA (Patra et al. 2013) are seen.


IL-2 as a biomarker

In addition to IFN-γ being used as a biomarker to diagnose bovine TB, IL-2 and IL-17 and in some cases IL-10, have been identified as potential biomarkers to study the progression of TB in cattle (Blanco et al. 2014).


IL-2 Immunotherapy

One of the first experiments to study the use of IL-2 in domestic food animals (pigs and cattle) was carried out by Anderson et al. in 1987. Animals vaccinated and injected with recombinant human IL-2 (rHuIL-2) were less severely affected when challenged with bacteria or viruses. Further studies revealed that rHuIL-2 increased NK cell activity (Bhagyam et al. 1988). Recombinant bovine IL-2 (rBoIL-2) is three times more effective than rHuIL-2.  The lower doses of rBoIL-2 needed cause fewer side effects, but deliver enhanced immunity and protection against bovine herpesvirus-1 .

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References:

  • Amills M et al. (2002). Reduced IL-2 and IL-4 mRNA expression in CD4+ T cells from bovine leukemia virus-infected cows with persistent lymphocytosis. Virology 304, 1-9.
  • Bhagyam RC et al. (1988). Activation of swine peripheral lymphocytes with human recombinant interleukin-2. Immunology 64, 607-613.
  • Blanco FC et al. (2014). Identification of potential biomarkers of disease progression in bovine tuberculosis. Vet Immunol Immunopathol 160, 177-183.  
  • Boyman O and Sprent J (2012). The role of interleukin‑2 during homeostasis and activation of the immune system. Nat Rev Immunol 12, 180-190.
  • Braun U et al. (2015). Interleukin-2 for the treatment of cows with malignant catarrhal fever. Schweiz Arch Tierheilkd 157, 31-38.
  • Buehring GC et al. (2015). Exposure to Bovine Leukemia virus is associated with breast cancer: A case-control study. PloS One 10.
  • Driesen J et al. (2008). CD25 as an immune regulatory molecule expressed on myeloid dendritic cells. Immunobiology 213, 849-858.
  • Maggioli MF et al. (2015). Characterization of effector and memory T cell subsets in the immune response to bovine tuberculosis in cattle. PLoS One 10.
  • Meier-Trummer CS et al. (2009). Malignant Catarrhal Fever of Cattle Is Associated with Low Abundance of IL-2 Transcript and a Predominantly Latent Profile of Ovine Herpesvirus 2 Gene Expression. PLoS One 4.
  • Patra MK et al. (2013). Neutrophil functions and cytokines expression profile in buffaloes with impending postpartum reproductive disorders. AJAS 26.
  • Rhodes SG et al. (2014). Use of Antigen-Specific Interleukin-2 to Differentiate between Cattle Vaccinated with Mycobacterium bovis BCG and Cattle Infected with M. bovis. Clin Vaccine Immunol 21, 39–45.
  • Russell GC et al. (2012). Host gene expression changes in cattle infected with Alcelaphine herpesvirus 1. Virus Res 169, 246–254.
  • Sallusto F et al. (1999). Two subsets of memory T lymphocytes with distinct homing potentials and effector functions. Nature 401, 708–712.
  • Whelan AO et al. (2011). Development of an Antibody to Bovine IL-2 Reveals Multifunctional CD4 TEM Cells in Cattle Naturally Infected with Bovine Tuberculosis. PLOS One 6.

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