Potential Efficacy of Proteasome Inhibitor, Delanzomib, for the Treatment of Renal Fibrosis.

Renal fibrosis is scarring and tissue hardening caused by the excess deposition of extracellular matrix proteins in response to chronic inflammation. Renal fibrosis is the primary cause of a progressive loss of renal function, and is an important therapeutic target because it ultimately leads to end-stage renal failure, which can be treated only by either dialysis or kidney transplantation. There is no effective treatment that specifically targets renal fibrosis. Myofibroblasts are known to evade apoptosis by activating molecular mechanisms in response to pro-survival biomechanical and growth factor signals from the fibrotic microenvironment. In this study, we screened and selected compounds that selectively cause cell death in myofibroblasts in vitro and studied their possible potency against renal fibrosis in a mouse model. Several proteasome inhibitors induced selective cell death in myofibroblasts differentiated from the human fibroblast cell line (MRC5). The in vivo antifibrotic effect of Delanzomib (Dz), one of the proteasome inhibitors most sensitive to myofibroblasts in vitro, was investigated in a Unilateral Ureteric Obstruction (UUO) mouse model. Treatment with Dz decreased the expression levels of the actin-alpha-2 (ACTA2) and collagen-type-1-alpha-1 (COL1A1) genes in the kidney, which are common fibrosis markers. These results suggest that Dz might be a compound that suppresses renal fibrosis by inducing selective cell death of myofibroblasts, although further investigation is required.

Role of lipid rafts in innate immunity and phagocytosis of polystyrene latex microspheres.

Understanding of the association of phagocytosis of polymers with signaling of innate immunity of macrophages is the major purpose of this study. Polymer conjugates have been utilized for clinical therapy of cancer and infections, such as Mycobacterium tuberculosis, as effective vectors of drug-delivery systems. They are incorporated through phagocytosis into macrophages and activate innate immunity signaling, which plays a crucial role in its therapeutic and side effects. Macrophage phagocytosis of polystyrene latex microspheres was examined and assayed by treatment of macrophages with the cholesterol depletor methyl-ß-cyclodextrin (MßCD) or the sphingolipid depletor n-octyl-ß-D-glucopyranoside (OGP). Expressions of various mRNAs during phagocytosis were quantified by real-time PCR. Phagocytosis of polystyrene latex microspheres by various macrophages, such as murine monocyte-derived macrophage J774, rat alveolar macrophage NR8383, and murine Kupffer cell KC13-2, was suppressed by treatment with MßCD or OGP in a concentration-dependent manner. The expression of mRNAs of TNFα, IL-1ß, IL-6 and CXCL10 genes induced by lipopolysaccharide (LPS) was not suppressed by treatment with MßCD in J774 cells. Moreover, genes that were induced by LPS were up-regulated even in the absence of LPS by the phagocytosis of polymer conjugates, but such up-regulations were not suppressed by the treatment with MßCD. It was shown that lipid rafts play a significant role in incorporation of polymer conjugates through phagocytosis of macrophages, but their association with signal transduction in innate immunity is very limited.

Role of lipid rafts in phagocytic uptake of polystyrene latex microspheres by macrophages.

BACKGROUND: Micro- and nanospheres have been used as carrier materials for effective delivery of drugs into macrophages via endocytosis including phagocytosis for the treatment of cancer and infections. MATERIALS AND METHODS: To analyze the role of lipid rafts in the endocytic uptake of microspheres by macrophages, we studied the incorporation of polystyrene latex microspheres (PSL MS) by treating macrophages with reagents that affect the physicochemical and biochemical functions of their lipid rafts, such as the cholesterol depletor methyl-ß-cyclodextrin (MßCD) and the inhibitor of scavenger receptors polyinosinic acid (poly I). For this study, we used 3-µm fluorescent polystyrene latex microspheres (F-PSL MS) and J774 murine macrophage. RESULTS: We found that the endocytosis of J774 cells was remarkably reduced in a concentration-dependent manner by treatment of the cells with MßCD and that endocytosis was suppressed by approximately 50% by treatment with 100 µg/ml poly I. These results suggest that scavenger receptors are associated with phagocytosis of F-PSL MS and that their functions are regulated by lipid rafts. CONCLUSION: PSL MS are phagocytosed via a raft-dependent pathway.