Mechanical stress induces tumor necrosis factor-α production through Ca2+ release-dependent TLR2 signaling

We studied centrifugation-mediated mechanical stress-induced tumor necrosis factor-alpha (TNF-alpha) production in the monocyte-like cell line THP-1. The induction of TNF-alpha by mechanical stress was dependent on the centrifugation speed and produced the highest level of TNF-alpha after 1 h of stimulation. TNF-alpha production returned to normal levels after 24 h of stimulation. Mechanical stress also induced Toll-like receptor-2 (TLR2) mRNA in proportion to the expression of TNF-alpha. The inhibition of TLR2 signaling by dominant negative myeloid differentiation factor 88 (MyD88) blocked TNF-alpha expression response to mechanical stress. After transient overexpression of TLR2 in HEK-293 cells, mechanical stress induced TNF-alpha mRNA production. Interestingly, mechanical stress activated the c-Src-dependent TLR2 phosphorylation, which is necessary to induce Ca2+ fluxes. When THP-1 cells were pretreated with BAPTA-AM, thapsigargin, and NiCl2 center dot 6H(2)O, followed by mechanical stimulation, both TLR2 and TNF-alpha production were inhibited, indicating that centrifugation-mediated mechanical stress induces both TLR2 and TNF-alpha production through Ca2+ releases from intracellular Ca2+ stores following TLR2 phosphorylation. In addition, TNF-alpha treatment in THP-1 cells induced TLR2 production in response to mechanical stress, whereas the preincubation of anti-TNF-alpha antibody scarcely induced the mechanical stress-mediated production of TLR2, indicating that TNF-alpha produced by mechanically stimulated THP-1 cells affected TLR2 production. We concluded that TNF-alpha production induced by centrifugation-mediated mechanical stress is dependent on MyD88-dependent TLR2 signaling that is associated with Ca2+ release and that TNF-alpha production induced by mechanical stress affects TLR2 production.