Anti-inflammatory Effect And Regulation On Associated Signal Transduction Pathway Of Doramectin

Doramectin, an ivermectin derivative, is a new generation of macrolideantiparasite drug. It has been used in the treatment of parasites such asgastrointestinal roundworms, lungworms, earworms, grubs, sucking lice and mangemites. In addition to their antibacterial and anti-parasitic activity, some macrolidesagents have been known to regulate inflammatory responses. In this study, the invitro anti-inflammatory effect and molecular mechanism of Doramectin onLPS-stimulated murine RAW264.7macrophages were investigated. Based on the invitro results, the effects of doramectin on LPS-induced toxicemia, acute lung injury,asthma and atopic dermatitis were studied.First, in vitro inflammatory model was performed by LPS-stimulated RAW264.7murine macrophages to investigate the effect of different concentrations ofdoramectin on LPS-induced TNF-α, IL-6and IL-1β, as well as inflammatorymediator production such as NO and PGE2. As result, doramectin inhibited not onlyTNF-α, IL-6and IL-1β secretion but also NO and PGE2synthesis. These resultsindicated that doramectin may have in vitro anti-inflammatory effect throughregulating the secretion of inflammatory cytokines and mediators. Additionally, themechanisms by which doramectin regulates these pro-inflammatory mediators andcytokines were assessed. Among these, Toll-like receptor (TLR4), mitogen-activatedprotein kinases (MAPKs) and nuclear factor-κB (NF-κB) pathways may play anessential role in the pathogenesis of inflammation. TLR4is the most importantreceptor in LPS-induced inflammatory response while NF-κB and MAPKs areknown as important molecular targets for anti-inflammatory therapy. In order toascertain the molecular mechanisms involved in anti-inflammatory effects ofdoramectin, the effect of doramectin on TLR4receptor, NF-κB and MAPKs signaltransduction pathways in LPS-stimulated RAW264.7murine macrophages werefurther investigated. The results showed that doramectin inhibited TLR4expression,NF-κB activity, p38and ERK1/2MAPKs phosphorylation. This observation suggested that doramectin inhibited cytokine TNF-α, IL-6, IL-1β, NO and PGE2secretion by inhibiting TLR4expression and regulating both NF-κB and p38,ERK1/2MAPKs pathways.To evaluate the clinical therapeutic effect of the drug, an investigation on invivo anti-inflammatory effect of doramectin was done. In order to choose the bestanti-inflammatory concentration, the effect of different concentrations on mousemortality and levels of inflammatory cytokines was monitored. The results showedthat doramectin significantly improved murine survival rate, decreased TNF-α, IL-6and IL-8, increased IL-10level in serum, but had little effect on IL-1β. It suggestedthat doramectin improved murine survival rate through regulating cytokineproduction.To investigate the effect of doramectin on acute lung injury (ALI), a mousemodel of LPS-induced ALI was established, and as result doramectin significantlyinhibited the W/D ratio of lungs, protein concentrations, number of inflammatorycells in BALF and lung MPO activity. Doramectin also significantly decreasedTNF-α, IL-6and IL-1β levels of BALF in LPS-induced ALI mice. Histopathology oflung showed that doramectin had certain protection on inflammatory infiltration ofALI. The results showed that doramectin had a protective effect on LPS-induced ALImouse model.Next, a mouse model of OVA-induced allergy asthma was established in theaim to investigate the effect of doramectin (3mg/kg) on asthmatic mice. The findingssuggested that doramectin significantly inhibited OVA-IgE, OVA-IgG, IL-4, IL-5,IL-13and CCL11levels in serum, and increased IFN-γ level in BALF, but had littleeffect on IL-10. Furthermore, doramectin dramatically reduced RL and lunghistopathology indicated that doramectin restored Cdyn in OVA-challenged mice inresponse to methacholine and markedly reduced the degree of inflammatory cellinfiltration in peribronchial and perivascular areas, mucus overproduction and gobletcell hyperplasia. In this study, doramectin showed a protective effect against OVA-induced asthmatic mouse model.Finally, the effect of doramectin (3mg/kg) on atopic dermatitis was examinedand the data proved that doramectin significantly decreased the ear thickness, IgElevel in serum, IL-4, IL-6, IL-1β and IL-18level and MPO activity in ear;doramectin was also found to significantly inhibit total cell and lymphocyte countsin cervical lymph nodes. The levels of IL-4and IL-13were decreased while the level of IFN-γ in submandibular lymph node increased. The inflammatory changes inepidermal thickness and dermal thickness, the degree of inflammatory cellinfiltration, as well as edema in ear tissue were significantly reduced.In this study, it has been demonstrated that doramectin has anti-inflammatoryeffect in vitro and in vivo by regulating TLR4-NF-κB, ERK1/2and p38MAPKssignal pathway. In addition to its anti-parasitic activity, doramectin proved to play animportant anti-inflammatory effect. In conclusion, doramectin is effective in thetreatment of inflammatory diseases and may be an innovative model for improvingclinical practice.