Background and Purpose: Osteoclasts are unique cells to absorb bone. Targeting osteoclast differentiation is a therapeutic strategy for osteolytic diseases. Natural marine products have already become important sources of new drugs. The naturally occurring nitrobenzoyl sesquiterpenoids first identified from marine fungi in 1998 are bioactive compounds with a special structure, but their pharmacological functions are largely unknown. Here, we investigated six marine fungus-derived nitrobenzoyl sesquiterpenoids on osteoclastogenesis and elucidated the mechanisms. Experimental Approach: Compounds were first tested by RANKL-induced NF-κB luciferase activity and osteoclastic TRAP assay, followed by molecular docking to characterize the structure-activity relationship. The effects and mechanisms of the most potent nitrobenzoyl sesquiterpenoid on RANKL-induced osteoclastogenesis and bone resorption were further evaluated in vitro. Micro-CT and histology analysis were used to assess the prevention of bone destruction by nitrobenzoyl sesquiterpenoids in vivo. Key Results: Nitrobenzoyl sesquiterpenoid 4, with a nitrobenzoyl moiety at C-14 and a hydroxyl group at C-9, was the most active compound on NF-κB activity and osteoclastogenesis. Consequently, nitrobenzoyl sesquiterpenoid 4 exhibited suppression of RANKL-induced osteoclastogenesis and bone resorption from 0.5 μM. It blocked RANKL-induced IκBa phosphorylation, NF-κB p65 and RelB nuclear translocation, NFATc1 activation, reduced DC-STAMP but not c-Fos expression during osteoclastogenesis in vitro. Nitrobenzoyl sesquiterpenoid 4 also ameliorated LPSinduced osteolysis in vivo. Conclusion and Implications: These results highlighted nitrobenzoyl sesquiterpenoid 4 as a novel inhibitor of osteoclast differentiation. This marine-derived sesquiterpenoid is a promising lead compound for the treatment of osteolytic diseases.
Diabetic nephropathy (DN) is a diabetic complication that can cause renal failure. β‐amyrin has been identified to possess anti‐diabetic property. This study was designed to evaluate the potential role of β‐amyrin in DN and its underlying mechanism. Streptozotocin‐induced diabetic mice were used as the in vivo model, and high glucose (HG)‐stimulated human proximal tubular HK‐2 cells were utilized as the in vitro model. Renal histological changes in mice were assessed by hematoxylin–eosin and periodic acid‐Schiff staining. HK‐2 cell viability and apoptosis were detected by Cell Counting Kit‐8 assay and flow cytometry analysis, respectively. β‐amyrin was found to ameliorate kidney injury in DN mice and suppressed inflammatory response as well as apoptosis of HG‐stimulated HK‐2 cells. miR‐181‐5p expression in murine renal tissues and HK‐2 cells was detected by in situ hybridization (ISH) and fluorescence in situ hybridization (FISH). MiR‐181b‐5p, a previously identified target for diabetic kidney disease, was downregulated in renal tissues and HG stimulated HK‐2 cells, and β‐amyrin induced the upregulation of miR‐181b‐5p. Binding relationship between miR‐181b‐5p and high mobility group box 2 (HMGB2) was confirmed by luciferase reporter assay. MiR‐181b‐5p bound to 3′ untranslated region of HMGB2 to suppress its expression. As shown by immunohistochemical staining and immunofluorescence staining, HMGB2 was upregulated in the in vivo and in vitro models of DN, and β‐amyrin induced the downregulation of HMGB2. Moreover, HMGB2 overexpression neutralized the suppressive effects of miR‐181b‐5p elevation on the inflammatory response and apoptosis of HG‐treated HK‐2 cells. Overall, β‐amyrin ameliorates DN in mice and suppresses inflammatory response and apoptosis of HG‐stimulated HK‐2 cells via the miR‐181b‐5p/HMGB2 axis.
Nuclear receptor peroxisome proliferator-activated receptor c (PPAR-c) activation can prevent immunoinflammatory disorders and diabetes. B cells play protective roles during inflammation as well. However, the roles of endogenous PPAR-c in the regulatory properties of B cells to relieve inflammation remain unknown. Here, we developed B-cell-specific PPARc knockout (B-PPAR-c À/À ) mice and found that the conditional deletion of PPAR-c in B cells resulted in exaggerated contact hypersensitivity (CHS). Meanwhile, interferon-c (IFN-c) of CD4 + CD8 + T cells was up-regulated in B-PPAR-c À/À mice in CHS. This showed that the regulatory function of B cells in B-PPAR-c À/À mice declined in vivo. Whereas splenic CD5 + CD1d hi regulatory B-cell numbers and peripheral regulatory T-cell numbers were not changed in naive B-PPAR-c À/À mice. Loss of PPAR-c in B cells also did not affect either CD86 or FasL expression in splenic CD5 + CD1d hi regulatory B cells after activation. Notably, interleukin-10 (IL-10) production in CD5 + CD1d hi regulatory B cells reduced in B-PPAR-c-deficient mice. In addition, functional IL-10-producing CD5 + CD1d hi regulatory B cells decreased in B-PPAR-c À/À mice in the CHS model. These findings were in accordance with augmented CHS. The current work indicated the involvement of endogenous PPAR-c in the regulatory function of B cells by disturbing the expansion of IL-10-positive regulatory B cells.
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