Temperature- and time-dependent changes in TLR2-activated microglial NF-κB activity and concentrations of inflammatory and anti-inflammatory factors

Therapeutic hypothermia protects neurons following injury to the central nervous system (CNS). Microglia express toll-like receptors (TLRs) that play significant roles in pathological processes in sterile CNS injury. We have examined the effects of culture temperature on the TLR2-activated microglial production of cytokines and nitric oxide (NO), which are known to be associated with CNS damage, and the possible involvement of nuclear factor-kappa B (NF-kappa B) activation underlying such effects. Rat microglia were cultured with a selective TLR2 agonist, Pam(3)CSK(4), under hypothermic, normothermic, and hyperthermic conditions, and with Pam(3)CSK(4) in the presence of a NF-kappa B activation inhibitor at 37 A degrees C. Cytokine and NO levels and NF-kappa B p65 activation were measured. The production of tumor necrosis factor-alpha (TNF-alpha), interleukin-10 (IL-10), and NO and the activation of NF-kappa B p65 were reduced by hypothermia, but augmented by hyperthermia at 3-6, 24-48, 48, and 0.5 h, post-treatment initiation, respectively. Pharmacological inhibition of NF-kappa B activation impaired the Pam(3)CSK(4)-induced TNF-alpha, IL-10, and NO production. In TLR2-activated microglia, hypothermia reduced, while hyperthermia increased, the early activation of NF-kappa B and the subsequent NF-kappa B-mediated production of TNF-alpha, IL-10, and NO in a time-dependent manner, suggesting that attenuation of these factors via suppression of NF-kappa B in microglia is one possible neuroprotective mechanism of therapeutic hypothermia. Moreover, temperature-dependent changes in microglial TNF-alpha production during the early phase and IL-10 and NO production during the late phase indicate that these factors might be useful as clinical markers to monitor hypothermia-related neuronal protection and hyperthermia-related neuronal injury.