ACS DIVISION OF BIOLOGICAL CHEMISTRY

October 6th, 2014 by Maher, K., Jerič–Kokelȷ, B., Butinar, M., Mikhaylov, G., Manček–Keber, M., Stoka, V., Vasilȷeva, O., Turk, B., Grigoryev, S. A., Kopitar–Jerala, N.

Stefin B (cystatin B) is an endogenous cysteine cathepsin inhibitor and the loss of function mutations in the stefin B gene were reported in patients with Unverricht-Lundborg disease (EPM1). In the present study we demonstrated that stefin B deficient (StB KO) mice were significantly more sensitive to the lethal LPS-induced sepsis and secreted higher amounts of pro-inflammatory cytokines IL-1β and IL-18 in the serum. We further showed that increased caspase-11 gene expression and better pro-inflammatory caspase-1 and -11 activation determined in StB KO bone marrow-derived macrophages (BMDMs) resulted in enchanced IL-1β processing. Pretreatment of macrophages with a cathepsin inhibitor E-64d did not affect secretion of IL-1β, suggesting that the increased cathepsin activity determined in StB KO BMDMs is not essential for the inflammasome activation. Upon LPS stimulation stefin B was targeted into mitochondria and the lack of stefin B resulted in the increased destabilization of mitochondrial membrane potential and mitochondrial superoxide generation. Collectively, our study demonstrates that the LPS-induced sepsis in StB KO mice is dependent on caspase-11 and mitochondrial reactive oxygene species (mtROS), but is not associated with the lysosomal destabilization and increased cathepsin activity in the cytosol.