Role Of TLR4in Idiopathic Inflammatory Myopathies

The idiopathic infammatory myopathies (IIM) encompass a heterogeneousgroup of rare muscle disorders related to adaptive immunity which is also calledautoimmune myositis. The main subtypes of IIM include polymyositis (PM),dermatomyositis (DM）, inclusion body myositis (IBM), non-specific myositis(NSM) and immune-mediated necrotizing myopathy (IMNM). The commonclinical symptoms are wield or serious weakness, Endomysia infammatory cellinfltrate, creatine kinase (CK) elevation and abnormal electromyogramdemonstrating. Currently the therapeutic efficacy of IIM is limit and manymedicine have lots of side effect, that means clinic needs more effective medicine.So it is essential and important to study the mechanism of IIM.Toll-like receptor4is one important member of Toll-like receptor familywhich can trigger an appropriate effective immune response by recognizingPAMPs and in turn leads to many inflammatory and autoimmune diseases. Themain two signaling pathways are activated by TLR4: MyD88-dependent andMyD88-independent. The former finally activates the transcription factor NF-κB, which promotes the expression of proinfammatory cytokines. Some studiessuggest TLR4play an important role in autoimmune diseases, and some showedTLR4-knockout mice could avoid some autoimmune diseases.However, there are rare studies related to the role and signaling of TLR4inIIM at home and broad, that are investigated in our study by detecting theexpression of TLR4in mice with experimental autoimmune myositis (EAM) andmuscle biopsy samples from PM patients. We hope the study may supply a neworientation for therapy of IIM.AIMS:The aim of our study was to investigate TLR4, MyD88and NF-кB mRNAlevels in mouse muscle and lymph node with EAM and muscle biopsy samplesfrom PM patients to determine the role of TLR4in IIM.METHODS:Experiment one: Thirty femal BALB/c mice were randomly divided into fivegroups (n=6): mice of group1were normally controlling; mice with EAM inother four groups were killed at different time point: group2in the first week，group3in the second week，group4in the third week and group5in the fourthweek since they had been given myosin for preparing EAM. The expression ofTLR4in muscle of each mouse was detected with immunohistochemistry (IHC).The expressions of TLR4, MyD88and NF-κB mRNA in muscle and lymph nodeof each mouse were measured with real-time fluorescent quantitative polymerasechain reaction (RT-PCR).Experiment two: muscle biopsy samples from patients newly diagnosed withactive PM (n=17) From April2009to May2011in neurology department ofXijing Hospital. Muscle tissue sections from seven patients with non-specific muscle manifestations but normal histological findings were used as the controls(n=7). The expression of TLR4in muscle tissue was detected withimmunohistochemistry (IHC). The expressions of TLR4, MyD88and NF-κBmRNA in muscle tissue were measured with real-time fluorescent quantitativepolymerase chain reaction (RT-PCR).RESULTS:Experiment one:（1）The expressions of TLR4in mouse muscle of eachEAM group were significantly high compared with those in the normal controlgroup by IHC（P<0.01）.（2）The expressions of TLR4, MyD88and NF-κBmRNA in mouse muscle and lymph node of each EAM group were significantlyhigh compared with those in the normal control group by RT-PCR, which wassignificantly highest in group3of all（P<0.01）.（3）In muscle and lymph nodeof mice with EAM, the expression level of TLR4mRNA had significant positivecorrelations with the expressions of MyD88mRNA and NF-κB mRNA(P<0.01),and the latter two also had significant positive correlations(P<0.01).（4）Theexpressions of TLR4mRNA in mouse spleen of each EAM group weresignificantly low compared with that in the normal control group byRT-PCR(P<0.01). The expressions of MyD88mRNA and NF-κB mRNA inmouse spleen of the third EAM group were significantly low compared with thatin the normal control group by RT-PCR (P<0.01). The expressions of MyD88mRNA had positive correlations with the expressions of NF-κBmRNA (P<0.01).Experiment two:（1）The expressions of TLR4in muscle biopsy samplesfrom patients with PM were significantly high compared with those in the normalcontrol group by IHC（P<0.01）.（2）The expressions of TLR4, MyD88andNF-κB mRNA in muscle biopsy samples from patients with PM weresignificantly high compared with those in the normal control group by RT-PCR （P<0.01）（.3）In muscle biopsysamples from patients withPM, the expressionlevel of TLR4mRNA had significant positive correlations with the expressions ofMyD88mRNA and NF-κB mRNA(P<0.01), and the latter two also hadsignificant positive correlations(P<0.01).CONCLUSION:（1） The expression of TLR4in muscle and lymph node of each EAMgroup were higher compared with those in the normal control group,which had some relation to inflammation and up regulated withinflammatory aggravation.（2） The results of the two experiments suggest TLR4took part in thedevelopment of IIM.（3） TLR4run its function mainly by MyD88-dependent pathway in IIMthat could activate NF-κB for promoting the release of inflammatoryfactors.