Background:The maturation process of IL-33 (IL-1F11) remains unclear. Results: IL-33 ligand affinity column isolates neutrophil proteinase 3. Conclusion: PR3 is an IL-33-processing enzyme. Significance: PR3 has a dual function in IL-33 biological activity.
α 1 -Antitrypsin (AAT) is a member of the serine proteinase inhibitor family that impedes the enzymatic activity of serine proteinases, including human neutrophil elastase, cathepsin G and neutrophil proteinase 3. Here, we expressed recombinant AAT by fusing the intact AAT gene to the constant region of IgG1 to generate soluble recombinant AAT-Fc protein. The recombinant AATFc protein was produced in Chinese hamster ovary (CHO) cells and purified using mini-protein A affinity chromatography. Recombinant AAT-Fc protein was tested for antiinflammatory function and AAT-Fc sufficiently suppressed tumor necrosis factor (TNF)-α-induced interleukin (IL)-6 in human peripheral blood mononuclear cells (PBMCs) and inhibited cytokine-induced TNFα by different cytokines in mouse macrophage Raw 264.7 cells. However, AAT-Fc failed to suppress lipopolysaccharide-induced cytokine production in both PBMCs and macrophages. In addition, our data showed that AAT-Fc blocks the development of hyperglycemia in a streptozotocin-induced mouse model of diabetes. Interestingly, we also found that plasma-derived AAT specifically inhibited the enzymatic activity of elastase but that AAT-Fc had no inhibitory effect on elastase activity.
Proteolysis targeting chimeras (PROTACs) are an emerging strategy for promoting targeted protein degradation by inducing the proximity between targeted proteins and E3 ubiquitin ligases. Although successful degradation of numerous proteins by PROTACs has been demonstrated, the elements that determine the degradability of PROTAC-targeted proteins have not yet been explored. In this study, we developed von Hippel-Lindau-Cereblon (VHL-CRBN) heterodimerizing PROTACs that induce the degradation of CRBN, but not VHL. A quantitative proteomic analysis further revealed that VHL-CRBN heterodimerizing PROTACs induced the degradation of CRBN, but not the well-known immunomodulatory drug (IMiD) neo-substrates, IKAROS family zinc finger 1 (IKZF1) and −3 (IZKF3). Moreover, truncation of disordered regions of CRBN and the androgen receptor (AR) attenuated their PROTAC-induced degradation, and attachment of the disordered region to stable CRBN or AR facilitated PROTAC-induced degradation. Thus, these results suggest that the intrinsically disordered region of targeted proteins is essential for efficient proteolysis, providing a novel criterion for choosing degradable protein targets.
Avascular necrosis of the femoral head (ANFH) is commonly observed in patients treated with excessive glucocorticoid (GC). Single administration of lipopolysaccharide (LPS) has shown to induce immune stimulatory factors. However, the effect of repeated administration of LPS on GC-induced ANFH has not been studied. Thus, the purpose of this study was (i) to examine the cytokine profile induced by repeated LPS administrations and (ii) to test the effect of repeated LPS treatments on GC-induced ANFH. A mouse necrosis model of ANFH was designed by chronic GC administration with co-treatment of LPS. Mice body weights in the LPS/prednisolone (PDN) co-treated group were lower than that of the untreated control group, but spleen weights were greater than the control group. The levels of IL-6, TNFα, and IL-33 in the liver and spleen of the LPS/PDN group were lower than the untreated control group, whereas TNFα level in the femoral head of the LPS/PDN group increased. Collectively, the effect of repeated LPS on the pathogenesis of GC-induced ANFH was associated with the TNFα level in the femoral head, but the pathogenesis did not correspond to cytokine levels in immune tissues.
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