Introduction
Inflammatory Bowel Disease (IBD) is a group of inflammatory conditions of the colon and small intestine. Crohn’s disease (CD) and ulcerative colitis (UC) are the principal types of inflammatory bowel diseases. The etiology is multifactorial, and many genetic loci have been found to serve as susceptibility genes for different subgroups of patients with IBD [1, 2], likewise, environmental influences are also considered as factors involved in IBD onset [3]. In addition, to its complicated risk factors, the pathogenesis of IBD is not entirely clear, making it difficult to develop new drugs with therapeutic value and acceptable side effects [2].
For more than two decades, animal models of intestinal inflammation have provided a wealth of information about mucosal immunology and the intestinal inflammation encountered in the inflammatory bowel diseases. While no single model has proven to sufficiently represent the complexity of the clinical characteristics of the human disease, the combined data obtained from these various animal models have provided a more detailed understanding of the underlying principles of human IBD and delineate pathways that might help predict the successful outcome of novel therapeutics in the clinic [4, 5].
The most widely used murine model of colitis employs dextran sodium sulfate (DSS), a chemical colitogen with anticoagulant properties, to induce intestinal epithelial damage [6]. The DSS colitis model lends itself to IBD research due to its rapidity, simplicity, reproducibility and controllability. Acute, chronic and relapsing models of intestinal inflammation can be achieved by modifying the concentration of DSS and the frequency of administration [6].
Numerous inflammatory mediators have been implicated in the pathogenesis of IBD. These include cytokines, eicosanoids, reactive oxygen species, nitric oxide, and complement activation products [7]. Similarly, increased expression of different inflammatory mediators (TNF-α, IL-1β, IFN-ɣ, IL-10 and IL-12) are observed as early as the first day of DSS treatment. The production of these inflammatory mediators increases progressively during DSS treatment [8].
Recently, we reported identification of an immunosuppressive peptide (ENV59-GP3) from a specific human endogenous retroviral envelope protein, ENV59, which has been adapted by the immune system and is involved in ameliorating autoimmunity [9, 10]. The ENV59-GP3 peptide showed strong immune modulating activity both in vitro andin vivo in SKG mouse model for rheumatoid arthritis and EAE model for Multiple Sclerosis [9, 10].
In this study we follow up on our previous findings and demonstrate that the ENV59-GP3 peptide treatment, reduces inflammation in the DSS-induced mice model of colitis. Furthermore, we show that in vitro ENV59-GP3 inhibits the KCa3.1 potassium channel, an ion channel responsible for hyperpolarization of the plasma membrane and regulation of calcium influx upon lymphocyte activation.
Potassium channels might be regarded as a new group of therapeutic targets for IBD [11, 12]. The intermediate-conductance calcium/calmodulin-gated potassium channel KCa3.1 also known as the IK or the Gardos channel (encoded by the KCNN4 gene) is expressed in a variety of tissues and cell types including T cells [13, 14], macrophages as well as DCs [14]. In T cells, sustained Ca2+ influx via the CRAC channel is important for sufficient cytokine production and proliferation. Influx of Ca2+ is dependent on a negative membrane potential. Activation of KCa3.1 by elevated intracellular Ca2+ maintains the negative membrane potential through efflux of K+, which helps to sustain Ca2+ entry into the cell. Upon channel opening, the efflux of potassium is driven by its concentration gradient and results in hyperpolarization of the membrane providing the electrochemical gradient necessary for influx of calcium [15].
KCa3.1 channel is known to be involved in regulating T cell function, proliferation, and cytokine production [16], thus suggesting that they could be involved in the pro-inflammatory responses in IBD further up-stream of inflammatory cytokines such as IFN-ɣ and TNF-α. Furthermore, studies have revealed a prominent role of KCNN4 in Ca2+-dependent ion transport and intestinal restitution [17]. In a T cell-mediated mouse model of colitis, inhibition of KCa3.1 ameliorated the disease [11]. Likewise, the KCa3.1 channel blocker NS6180 prevents T-cell activation and inflammation in a rat model of inflammatory bowel disease [18]. Although loss of KCa3.1 did not interfere with CD4 T cell differentiation, both Ca2+ influx and cytokine production were impaired in KCNN4−/− Th1 and Th2 CD4 T cells, whereas T-regulatory and Th17 function were normal. In animal models for other autoimmune diseases, the KCa3.1 inhibitor TRAM-34 or ICA-17043 (Senicapoc) have been reported to prevent experimental autoimmune encephalomyelitis in mice [19] or to inhibit/attenuate inflammatory responses in mice with collagen antibody induced rheumatoid arthritis. Clinical studies with Clotrimazole (form which Tram-34 has been derived) already provided evidence that blockers of this channel may be useful in treatment of chronic diseases such as RA, however, non-target related adverse effects have prevented further development in some cases [20].