αB crystalline upregulates the expression of matrix metalloproteinases in trabecular meshwork cells through TLR1/2

Purpose: Our previous study identified the TLR1/2 complex as the receptor for alpha B crystalline (CRYAB). Here, we aimed to determine whether CRYAB could influence the expression of matrix metalloproteinases (MMPs) in trabecular meshwork (TM) cells through TLR1/2. Methods: The expression of P65, P38, ERK and JNK, which are downstream of TLR1/2, was identified by western blot (WB) with the interference of a TLR1/2 inhibitor or siRNA-TLR1 (siRNA-TLR2). MMP2 and MMP9 were tested by WB analyses using the same interference conditions in TM cells. C57BL/6N mice were used to study the effects in vivo by anterior chamber injection of CRYAB. Finally, immunohistochemistry was performed to evaluate the alterations in MMP2 and MMP9 between the CRYAB injection group and the normal control group. Results: For the NF-kappa B pathway, the expression of P65 was increased in TM cells with the addition of exogenous CRYAB (P<0.01), which was dramatically reduced with inhibition of TLR1/2 by its inhibitor (CU-CPT22) or knockdown of TLR1 (or TLR2) with siRNA-TLRs (P<0.01). For the MAPK pathway, P38, ERK and JNK showed no significant difference when CRYAB or CU-CPT22 was added. Subsequently, MMP2 and MMP9 were upregulated, which was consistent with the increased level of p65 (P<0.001). Finally, elevated expression of MMP2 and MMP9 in mice was demonstrated in trabecular meshwork tissue compared to that of the normal control. Conclusion: CRYAB triggers the activation of MMPs in TM cells in vitro and in vivo, possibly following the TLR1/2-mediated proinflammatory effect on TM cells.