Function

a) Providing help when CD4⁺ T cells are limited/ MHCI dependent manner:

Another hint regarding the role of CD40L⁺ CD8⁺ T cells in immunity comes from CD40L itself. Proper B cell activation and respective humoral immunity is dependent on T cell mediated help. The actual concept relies on MHCII presentation of antigens by B cells that are detected by CD4⁺ T cells, leading to T cell activation and CD40L upregulation. Interaction of CD40L with CD40 on the B cell provides the necessary “help” signal to induce B cell maturation, class switching and somatic hypermutation, which can be further supported by T cell secreted cytokines (IL-4, IFN-). Helper-type Tc2, Tc17 and Tc22 CD8⁺ T cells express CD40L with an intensity comparable to CD4⁺ T cells. CD40L^high IL-2 co-secreting clones were shown to be superior to CD40L^int clones (IFN- co-producers) in B cell activation and induction of Ig secretion (Hermann et al., 1995). Furthermore, in HIV patients with reduced CD4⁺ T cell levels, the majority of CD8⁺ T cells exhibit an IL-4 secreting Tc2 phenotype and is capable of providing B cell help (Maggi et al., 1994, 1997). The increased frequency of CD40L⁺ CD8⁺ T cells when CD4⁺ T cell-mediated help is limited such as in ThPOK^-/- and MHCII^-/- mice models and CD4⁺ depletion experiments supports the ability of helper-type CD8⁺ T cells to provide MHCI dependent B cell help. As humoral immunity is ineffective against intracellular infections that are classically presented on MHCI, this CD8⁺ T cell mediated help might rather play a role in the response to cross-presented antigens.

Help was furthermore shown to be an important aspect of cell mediated immune responses (Bennett et al., 1998; Janssen et al., 2003; Schoenberger et al., 1998; Shedlock and Shen, 2003). CD4⁺ T cells activate and license DC by CD40L-CD40 interaction, which leads to MHC upregulation and enables the DC to provide sufficient secondary (co-stimulatory receptors) and tertiary (cytokines) signals for optimal CD8⁺ T cell activation and induction of an effector response.

Huber and Lohoff showed that a reverse variant of mediating help also exists.

Non-cytotoxic, IL-17 producing CD8⁺ T cells were necessary for the accumulation of strongly pathogenic CD4⁺ T cells in the CNS of murine EAE model (Huber and Lohoff, 2015; Huber et al., 2009, 2013). Accordingly, IL-17 producing CD8⁺ T cells were identified in active lesions of MS patients (Tzartos et al., 2008).

Therefore, CD40L⁺ CD8⁺ T cells might not only contribute to efficient effector responses of CD8⁺ T cells but also the recruitment and activation of antigen specific CD4⁺ T cells.

b) Cytokine secretion:

Helper-type CD8⁺ T cells secrete a specific repertoire of cytokines (IL-4, IL-13, IL-17, IL-22, IL-10) that differ from classical CD8⁺ T cell mediated IFN- secretion (Figure 26). The functions of these cytokines were mostly identified by analysis of CD4⁺ T cells. However, the effect of the cytokines on their target cell is independent of their origin and therefore should also be induced by CD8⁺ T cell derived cytokines. IL-4, IL-5 and IL-13 secretion by Th2 cells is critical for the recruitment of eosinophils, which mediate the expulsion of parasites (Sallusto, 2016). CD4⁺ T cell derived IL-4 and IL-5 moreover support B cell maturation and class switch in the presence CD40L-CD40 interaction. Th17 derived IL-17 in turn supports the clearance of extracellular bacteria and fungi by the mobilization of neutrophils (McDermott and Klein, 2018). And IL-22 produced by Th22 cells is assumed to play a role in skin regeneration as well as skin protection by inducing the production of antimicrobial peptides (Duhen et al., 2009; Sigmundsdottir et al., 2007). As helper-type CD8⁺ T cells are also capable to express CD40L in combination with IL-4 and IL-5, they can mediate according B cell class switching in a MHCI dependent manner (Hermann et al., 1995). IL-13, a feature of not only Th2 cells but also of all helper-type memory CD8⁺ T cells Tc2, Tc17 and Tc22 has a protective function in skin regulating basal cell activity and subsequently it contributes to wound healing (Dalessandri et al., 2016). The different cytokines also can induce the release of chemokines by other cells, which modulate the recruitment of further cells. Accordingly, helper-type CD8⁺ T cells are capable of inducing CCL26 in keratinocytes, which in turn mediate the recruitment of CCR4⁺ cellswhereas cytotoxic CD8⁺ T cells were demonstrated to recruit further CXCR3⁺ cells by the induction of CXCL11, instead (Figure 41).

There are multiple conditions where Tc2, Tc17 and Tc22 cells were detected. Tumors at barrier sites such as skin or lung (melanoma, lung cancer) often contain Tc2 cells (Minkis et al., 2008; Roberts et al., 2009; Sheu et al., 2001). IL-13 producing Tc2 (most likely together with Tc17, Tc22) cells furthermore are widely associated with several autoimmune diseases in tissues.

They are enriched in patients with systemic sclerosis (Cascio et al., 2017), alopecia areata (Czarnowicki et al., 2018), rheumatoid arthritis (Cho et al., 2012) and best described in asthma. Despite the vast majority of lung CD8⁺ T cells exhibit a cytotoxic memory (CXCR3⁺CCR5⁺) phenotype in patients with mild asthma only the small fraction expressing CCR4⁺ and producing IL-13 has been associated with disease progress (Campbell et al., 2001). Adoptive transfer of Tc2 cells that favored low antigen concentrations during priming but not Tc1 primed OT-1 cells led to lung eosinophila and bronchial hyperresponsiveness, classical characteristics of asthma (Sawicka et al., 2004). Additionally, increased frequencies of IL-13 producing CD8⁺ T cells were found among bronchoaveolar lavage cells of asthmatic patients compared to healthy (Dakhama et al., 2013) and IFN-knockout led to a Tc2/Tc17 biased phenotype of lung CD8⁺ T cells with reduced cytotoxicity, paralleled by an aggravated disease (Tang et al., 2012).

Hondowicz and his team showed that in mice sensitized with house dust CD4⁺ TRM in lung arise together with circulatory TCM in secondary lymphoid organs, a mechanism we propose to underlie the appearance of helper-type CD8⁺ T cell in the circulation (Hondowicz et al., 2016). A severe form of asthma is associated with high levels of Ig-E, which can mediate eosinophil and neutrophil activation.

Secretion of Ig-E requires CD40L dependent activation of B cells in the presence of IL-4 and Tc2 cells could provide both as potential contributor to the worsening of asthma.

But how are these helper-type cells activated in autoimmune diseases? IL-17 and IL-22 producing CD8⁺ T cells in the epidermis were identified as major driver of psoriasis induction (Di Meglio et al., 2016). Triggers such as tissue traumata can result into a complex formation of keratinocyte derived antimicrobial peptide LL37 with self-DNA/RNA, which activates dendritic cells in skin to secrete IL-23. Those DC activate autoreactive CD8⁺ T cells which in turn release IL-17 and contribute to skin lesions – a process also known as Koebner phenomenon (Lande et al., 2014). Similarly, Mrp8 and Mrp14 can induce autoimmunity. They

belong to the group of DAMPS (damage-associated molecular pattern molecules) that are constitutively expressed endogenous proteins and released upon tissue damage. The Mrp8 and Mrp14 protein load is increased in several autoimmune disorders such as multiple sclerosis, rheumatoid arthritis and psoriasis. When presented on DC paralleled by CD40-CD40L interaction with CD8⁺ T cells Mrp8 and Mrp14 might mediate TLR4 ligation and IL-17 induction in the CD8⁺ T cells (Loser et al., 2010). Therefore, atypical structures that become accessible by cell disruption and/or complex formation possibly mimic high affine peptides for CD40L⁺ CD8⁺ T cells and activate them in a peptide independent mechanims.

While CD40L expressing CD8⁺ T cells might usually contribute to tissue maintenance and/or tolerance of commensal bacteria, their CD40L expression is a disadvantage when untypical structures become accessible for example by tissue disruption.