**Abstract**

Helper CD4+ T cells are essential in shaping effective antibody response and cytotoxic T cell response against pathogen invasion. There are two subtypes of pathogen-specific helper T cells in mice and humans; type 1 (Th1) and type 2 (Th2), with Th1 producing interferon-gamma (IFNγ) and Th2 producing interleukin-4 (IL-4). While effective Th1 controls intracellular pathogens like viruses, efficient Th2 controls extracellular pathogens like most parasites. However, the most predominant CD4+ T cell subtype in cattle is Th0, which produces both IFNγ and IL-4, and only exists in small amounts in mice and humans. Moreover, in many bovine infections, both IFNγ and IL-4 were detected in the blood and both antigenspecific IgG2 (Th1 associated bovine antibody) and antigen-specific IgG1 (Th2 associated bovine antibody) were upregulated in the serum, suggesting bovine CD4+ T cell responses may vary from those in mice and humans. How bovine CD4+ T cell differentiation differs from that in mice and humans and how some critical bovine pathogens regulate immunity to establish chronic infections are largely unknown. This chapter summarizes current literature and identifies the knowledge gaps to provide insights into future research in the field.

**Keywords:** bovine, CD4+ T cell differentiation, antigen-specific clones, Th0 responses, pathogens, chronic infections

## **1. Introduction**

CD4+ T cells, also called helper T cells, are important regulators of adaptive immune responses, which are antigen-specific and critical in protecting animals from pathogen infections. The control of intracellular pathogens, such as viruses, primarily depends on antigen-specific CD8+ T cell response, whereas antibodies (produced by B cells) or humoral immune responses are mostly responsible for the control of extracellular pathogens such as most bacteria and parasites. CD4+ T cells are the lynchpin in shaping both CD8+ T cell and antibody responses [1, 2].

Common lymphoid progenitor cells migrate from the bone marrow into the thymus for further development and maturation into T cells. Inside the thymus, these progenitor cells proliferate into a large pool of T cells, with each expressing a unique T cell receptor (TCR) through a genetic recombination. After TCR recombination, T cells must go through two selection processes, and only a fraction of them pass through these selections and become either CD4+ or CD8+ T cells [3]. Surviving CD4+ T cells then exit the thymus as naïve CD4+ T cells but without the ability to help CD8+ T cells and B cells. To become fully functional, naïve CD4+ T cells need to become activated and differentiated into specialized effector subtypes; helper

type 1 (Th1) to facilitate CD8+ T cell responses, and helper type 2 (Th2) to facilitate antibody responses [4]. Naïve CD4+ T cells constantly survey secondary lymphoid tissues to detect pathogens through their antigen-specific TCRs [5]. As opposed to antibodies, which bind directly to pathogens or their derivatives, TCRs can only recognize short chains of amino acids (derived from pathogens) that are presented by major histocompatibility-II (MHC-II) expressed on antigen presenting cells (APCs) [2]. This recognition process provides the 1st signal required to activate naïve CD4+ T cells. Along with the 1st signal, APCs also offer co-stimulation as the 2nd signal and cytokine signaling, as the 3rd signal, to the naïve CD4+ T cell. Combined, these three signals coordinate CD4+ T cell differentiation into distinct effector subtypes with different helper functions [2].

Studies in humans and mice have identified numerous helper subtypes, including: Th1, Th2, Th3, Th9, Th17, Treg, and Tr1 [2, 6]. Among these, Th1 and Th2 are considered to play major roles in defending the host from pathogen invasion [7–9]. Th1 cells help CD8+ T cells to gain killing functions, which leads to apoptosis of infected cells and induces Interferon gamma (IFNγ) mediated immunity [10–13]. On the other hand, Th2 cells help B cells differentiate into plasma cells, which produce pathogen-specific antibodies [14]. Antibodies or humoral immunity contribute to the control of extracellular pathogens by mechanisms like neutralizing toxins, preventing bacterial attachment to the host cell, and stimulating basophil and mast cells to release toxic chemicals that induce the expulsion of large gastrointestinal parasites [15, 16]. Although antibodies are mostly responsible for controlling extracellular pathogens, they can also play important roles in cell-mediated killing of intracellular pathogens [17]. For instance, during intracellular infections in mice, Th1 cells help B cells become plasma cells that secrete antigen- specific immunoglobulin subtype G2a (IgG2a), which in turn can help killing infected cells through antibody dependent cytotoxicity (ADCC) [18, 19]. In short, Th1 is responsible for control of intracellular pathogens mostly through shaping CD8+ T cell responses and Th2 is for control of extracellular pathogens through antibody responses. In addition, antibodies can be involved in both Th1 and Th2 responses, but with unique subtypes, such as IgG2 for Th1, and IgG1 for Th2 in cattle. This will be discussed further in Section 2.

There are many similarities in the immune system across species. Therefore, knowledge generated from the research in mice and humans has been extensively applicable to study immune responses in cattle [20–23]. In the past several decades, however, unique features have been discovered in the bovine immune system that are not shared with that of mice and humans, such as high prevalence of circulating γδ T cells [24], production of IL-10 by γδ T cells [25], regulation of CD4+ T cell activation by neutrophils [26], which are able to secrete IL-10, and high prevalence of hybrid helper T cells (*i.e.,* co-express both Th1 and Th2 cytokines), which is relatively low in humans and mice [22, 27, 28].

Cattle industry suffers billions of dollar's losses annually due to infections, and many of the commercially available vaccines for cattle are not fully effective [29–32]. Understanding the mechanisms underlying bovine CD4+ T cell differentiation, which seems to be partially different from that of mice and humans, is critical to identify novel strategies to achieve more effective immunity after vaccinations, such as through generating strong Th1 responses against intracellular pathogens and Th2 responses against extracellular pathogens. In this chapter, we will summarize the current knowledge and key findings on bovine CD4+ T cell responses, highlight the existing knowledge gaps, and provide some insights on future directions.
