Studies using SERPINB deficient mice established the

Studies using SERPINB1-deficient mice established the significant role for SERPINB1 in protecting lung antimicrobial proteins from proteolysis during microbe infection and its regulatory role in sustaining the balance of neutrophil reserve [15]. Neutrophil serine proteases (NSPs) (elastase, cathepsin G, and proteinase-3) kill invading microbes [18]. However, excess NSPs in the lungs play a central role in inflammatory pulmonary disease. On infection, wild-type (WT) and SERPINB1-KO mice show similar early responses, production of cytokines and chemokines, recruitment of neutrophils. However, SERPINB1 mice fail to efficiently clear bacteria due to a deficient recruitment of neutrophils to the lung because these Fmoc-Gln(Trt)-OPfp show a shorter survival and increased release of neutrophil proteases in the extracellular space generating sustained pro-inflammatory cytokine production. The negative impact of overproduction of pro-inflammatory cytokines has been established by its association with fatal outcome of human influenza [15], [20]. The administration of a recombinant SERPINB1 protein to SERPIN KO mice normalized bacterial clearance, indicating that the regulation of pulmonary innate immunity by SERPINB1 is a non redundant key process during the host response to infection. In line with this, other results suggest that SERPINB1 may be a novel marker of active ulcerative colitis and may play an important role in the pathogenesis of inflammatory bowel disease, since it is seen accumulated in the intestinal lumen and in intestinal epithelial cells of patients [21]. Taken together these results indicate that SERPIN B1 has an important role in the resolution of inflammatory diseases.
LEI derived-DNase II: another function of SERPINB1 The eye lens is a useful model for the examination of many fundamental processes. Cellular differentiation of the lens is accompanied by nuclear degeneration morphologically related to nuclear modifications seen during apoptosis [22]. This includes the degradation of DNA in oligonucleosomes. We examined in detail this phenomenon in chicken lens and obtained results indicating that single strand and double strand cleavage of DNA cumulate in lens fiber nuclei [23]. However, the absence of 3′OH ends directed our attention towards the involvement of a DNase II (an endonuclease that releases 3′ phosphate ends) in this process. By using polyclonal antibodies, done in our laboratory, directed against DNase II, purified from porcine spleen, we showed that the enzyme responsible for lens differentiating cell nuclei has a cytoplasmic localization in chick lens epithelial cells (that is a not differentiated cell), but it concentrates in the nuclei of fibers cells [24]. Moreover, only this specific anti-DNase II antibody was able to inhibit the degradation of DNA from fibers cells in vitro. The crucial role played by DNase II in these processes drove our interest to its molecular structure, that was totally unknown by this time. We have then undertaken the cloning and molecular analysis of this protein. To do this, we first sequenced the protein by Edman degradation. Search for similarities of the obtained sequences within the databases showed 100% identity of our peptides with a protein of the serpin superfamily, the Leukocyte Elastase Inhibitor (LEI), afterwards called SERPINB1 [25]. This was very intriguing: we sequenced an endonuclease of 27kDa and obtained the sequence of an anti-protease of 42kDa. Many experiments were necessary in order to prove that this was not a technical error. Explaining these results was a major challenge. Many hypotheses were investigated [26]. Finally, the hypothesis of a post-translational modification was retained as explained below. Actually, after cloning the cDNA in bacteria we produced a protein of 42kDa with an anti-protease activity, as expected. The protein, purified using its His-tag had not DNase activity. However, if we purified the recombinant protein not by using its His-tag but by applying the same protocol used to purify DNase II from pig spleen, we generated a 35kDa protein that did have an endonuclease activity [25]. The same effect was obtained if we submitted the recombinant protein purified by the His-tag to an acidic pH, a mandatory step in the purification of DNase II from tissues. These results allowed us to conclude that the DNase activity was derived from LEI by post-translational modification. We therefore called it L-DNase II for LEI-derived DNase II. Later on, we showed that the acidic treatment could be replaced by elastase cleavage and that the elastase cleaved form of LEI had also a DNase II activity.