Therapeutic treatment with Ibrutinib attenuates imiquimod-induced psoriasis-like inflammation in mice through downregulation of oxidative and inflammatory mediators in neutrophils and dendritic cells

Abstract

Psoriasis is clinically characterized by well-demarcated silvery plaques which may appear on the extremities, scalp, and sacral area. The multidimensional interactions among innate immune cells [neutrophils and dendritic cells (DCs)], adaptive immune cells and skin resident cells result in characteristic features of psoriatic inflammation such as acanthosis, hyperkeratosis, and parakeratosis. Tec family kinases are involved in the pathogenesis of several inflammatory diseases. One of them is Bruton’s tyrosine kinase (BTK) which is reported to carry out inflammatory and oxidative signaling in neutrophils and DCs. Effect of BTK inhibitor with regard to psoriatic inflammation has not been explored previously especially in a therapeutic setting. In the current investigation, effect of BTK inhibitor, Ibrutinib on oxidative/inflammatory signaling in dermal/splenic neutrophils [phosphorylated BTK (p-BTK), inducible nitric oxide synthase (iNOS), nitrotyrosine], CD11c + DCs (p-BTK, iNOS, nitrotyrosine, MCP-1, TNF-α) and enzymatic antioxidants [superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR)] in imiquimod (IMQ)-induced psoriatic inflammation was evaluated using therapeutic mode. Our results show that IMQ treatment led to induction of p-BTK expression along with concomitant increase in oxidative stress in neutrophils, and CD11c + DCs in skin/periphery. Therapeutic treatment with Ibrutinib caused attenuation of IMQ-induced oxidative stress in CD11c + DCs and neutrophils. Further there were dysregulations in antioxidants enzymes (SOD/GPx/GR) in the skin of IMQ-treated mice, which were corrected by Ibrutinib. In short, our study reveals that BTK signaling in neutrophils and CD11c + DCs upregulates oxidative stress which is concomitant with psoriatic inflammation in mice. Ibrutinib attenuates psoriasis inflammation through downregulation of oxidative stress in these innate immune cells.

Introduction

Psoriasis is a very prevalent inflammatory disease which manifests itself in the form of well-demarcated silvery plaques resulting from an interplay between the innate and adaptive immune system (Chiricozzi et al., 2018; Schön, 2019). There is a multidirectional communication between innate immune cells and resident skin cells such as neutrophils/dendritic cells (DCs), and keratinocytes respectively. These interactions impact differentiation of adaptive immune cells ultimately resulting in histopathological features of psoriasis such as acanthosis, hyperkeratosis and parakeratosis (Schön, 2019; Lowes et al., 2014; Boehncke, 2015).

Neutrophils are reported to be present in psoriatic lesions where they can secrete different inflammatory mediators including oxidants and cytokines (Lin et al., 2011; Schon et al., 2017; Havnaer et al., 2019; Katayama, 2018). Out of these, oxidants play a significant function in amplification of dermal inflammation by causing widespread damage to biological macromolecules. This is corroborated by the findings which indicate redox imbalance in psoriasis (Zhou et al., 2009; Bacchetti et al., 2013). For example, increased lipid peroxidation products and reduced antioxidants in psoriatic patients have been reported (Nemati et al., 2014; Péter et al., 2016). Bruton’s tyrosine kinase (BTK) signaling has also been shown to be involved in neutrophilic inflammation (Volmering et al., 2016; Ito et al., 2015; Krupa et al., 2014). However, its context in psoriatic inflammation is largely undiscovered.

Dermal inflammation in psoriasis is also characterized by presence of large number of DCs in the epidermis and dermis (Schön, 2019; Lowes et al., 2014). DCs are important contributors in amplification of dermal inflammation as they are the key cells which are involved in secretion of various cytokines, chemokines and oxidants (Guttman-Yassky et al., 2007; Lowes et al., 2014). Mediators released by dermal DCs act on keratinocytes and help them proliferate and secrete other chemokines/cytokines and antimicrobial peptides which further enhance psoriatic inflammation (Schön, 2019; Lowes et al., 2014; Nadeem et al., 2017). Inflammatory effects of BTK activation in neutrophils and DCs have been investigated in earlier studies as well, however whether BTK signaling is involved in regulation of inflammatory and oxidative mediators in DCs and neutrophils remains largely unexplored especially with regard to psoriasis-like inflammation in mice (Stadler et al., 2017; Ye et al., 2019; Li et al., 2014; Weber et al., 2017).

Several immune cells or their mediators have been the target of immunotherapy with regards to psoriatic inflammation. This includes antagonism of IL-12/IL-23, IL-17A and TNF-α signaling on various immune cells. However, there is still lacunae in the current therapies which needs to be overcome such as lack of clinical response even after use of multiple immunomodulators (Greb et al., 2016). In this regard, BTK inhibition could be beneficial as this strategy has shown ameliorative effects in multiple inflammatory diseases in the past (Stadler et al., 2017; Weber et al., 2017; Rip et al., 2018; Di Paolo et al., 2011; de Porto et al., 2019). However, this strategy has not been investigated in a therapeutic setting in the context of psoriatic inflammation. BTK is a crucial member of Tec kinase family and mediates multiple roles in neutrophils and DCs which include regulation of several downstream transcription factors or other protein kinases. Activation of toll-like receptors (TLRs) is also well connected with BTK signaling in innate immune cells such as neutrophils and DCs (Weber et al., 2017; Rip et al., 2018; Li et al., 2014; Ye et al., 2019). All of these observations strongly suggest that BTK signaling could be therapeutically targeted to ameliorate psoriasis-like inflammation.

Therefore, we utilized BTK inhibitor, Ibrutinib therapeutically in imiquimod (IMQ) mouse model of psoriatic inflammation. Our data disclose that BTK signaling in neutrophils and CD11c + DCs is related with upregulation of oxidative and inflammatory mediators in IMQ model. Therapeutic treatment with BTK inhibitor, Ibrutinib causes marked reduction of inflammatory and oxidative mediators in the skin and periphery of IMQ-treated mice.