Knockout of TLR4 and TLR2 impair the nerve regeneration by delayed demyelination but not remyelination

Background: Knockout of either toll-like receptor 4 (TLR4) or 2 (TLR2) had been reported to delay the Wallerian degeneration after peripheral nerve injury by deterring the recruitment of the macrophages and clearance of myelin debris. However, the impact on the remyelination process is poorly understood. In this study, the effect of TLR2 and TLR4 knockout on the nerve regeneration and on the remyelination process was studied in a mouse model of sciatic nerve crush injury. Results: A standard sciatic nerve crush injury by a No. 5 Jeweler forcep for consistent 30 seconds was performed in Tlr4(-/-) (B6.B10ScN-Tlr4(lps-del)/JthJ), Tlr2(-/-)(B6.129-Tlr2(tm1Kir)/J) and C57BL/6 mice. One centimeter of nerve segment distal to the crushed site was harvested for western blot analysis of the myelin structure protein myelin protein zero (Mpz) and the remyelination transcription factors Oct6 and Sox10 at day 0, 3, 7, 10, 14, 17, 21, 28. Nerve segment 5-mm distal to injured site from additional groups of mice at day 10 after crush injury were subjected to semi-thin section and toluidine blue stain for a quantitative histomorphometric analysis. With less remyelinated nerves and more nerve debris, the histomorphometric analysis revealed a worse nerve regeneration following the sciatic nerve crush injury in both Tlr4(-/-) and Tlr2(-/-) mice than the C57BL/6 mice. Although there was a delayed expression of Sox10 but not Oct6 during remyelination, with an average 4-day delay in the demyelination process, the subsequent complete formation of Mpz during remyelination was also delayed for 4 days, implying that the impaired nerve regeneration was mainly attributed to the delayed demyelination process. Conclusions: Both TLR4 and TLR2 are crucial for nerve regeneration after nerve crush injury mainly by delaying the demyelination but not the remyelination process.