Euphoesulatin A prevents osteoclast differentiation and bone loss via inhibiting RANKL-induced ROS production and NF-κB and MAPK signal pathways.

Euphoesulatin A (Eup A), a new jatrophane diterpenoid isolated from the Euphorbia esula L. (Euphorbiaceae), was reported to inhibit RANKL-induced osteoclastogenesis. However, the underlying mechanism and the effect in osteoporosis mouse model are still unclear. This study is the first to demonstrate that Eup A inhibits osteoclastogenesis in vitro and in vivo. Mechanistic analysis suggested that Eup A (3, 6, 12 µM) dose-dependently inhibited osteoclastogenesis by down-regulating the activation of NFATc1 and NF-κB and MAPKs signal pathways. Moreover, Eup A (10 mg/kg) significantly prevented bone loss in ovariectomized mice. This work provides in vitro and in vivo evidence that Eup A could be a potential candidate for the development of anti-osteoporosis agents.

Molecular Characterization of bla IMP - 4 -Carrying Enterobacterales in Henan Province of China.

Carbapenem-resistant Enterobacterales (CRE) pose a serious threat to clinical management and public health. We investigated the molecular characteristics of 12 IMP-4 metallo-ß-lactamase-producing strains, namely, 5 Enterobacter cloacae, 3 Escherichia coli, 2 Klebsiella pneumoniae, and 2 Citrobacter freundii. These strains were collected from a tertiary teaching hospital in Zhengzhou from 2013 to 2015. The minimum inhibitory concentration (MIC) results showed that each bla IMP - 4-positive isolate was multidrug-resistant (MDR) but susceptible to colistin. All of the E. coli belonged to ST167, two C. freundii isolates belonged to ST396, and diverse ST types were identified in E. cloacae and K. pneumoniae. S1-PFGE, Southern blotting, and PCR-based replicon typing assays showed that the bla IMP - 4-carrying plasmids ranged from ∼52 to ∼360 kb and belonged to FII, FIB, HI2/HI2A, and N types. N plasmids were the predominant type (8/12, 66.7%). Plasmid stability testing indicated that the bla IMP - 4-carrying N-type plasmid is more stable than the other types of plasmids. Conjugative assays revealed that three of the bla IMP - 4-carrying N plasmids were transferrable. Complete sequence analysis of a representative N type (pIMP-ECL14-57) revealed that it was nearly identical to pIMP-FJ1503 (KU051710) (99% nucleotide identity and query coverage), an N-type bla IMP - 4-carrying epidemic plasmid in a C. freundii strain. PCR mapping indicated that a transposon-like structure [IS6100-mobC-intron (K1.pn.I3)-bla IMP - 4 -IntI1-IS26] was highly conserved in all of the N plasmids. IS26 involved recombination events that resulted in variable structures of this transposon-like module in FII and FIB plasmids. The bla IMP - 4 gene was captured by a sul1-type integron In1589 on HI2/HI2A plasmid pIMP-ECL-13-46.

Inhibitory mechanism of reveromycin A at the tRNA binding site of a class I synthetase.

The polyketide natural product reveromycin A (RM-A) exhibits antifungal, anticancer, anti-bone metastasis, anti-periodontitis and anti-osteoporosis activities by selectively inhibiting eukaryotic cytoplasmic isoleucyl-tRNA synthetase (IleRS). Herein, a co-crystal structure suggests that the RM-A molecule occupies the substrate tRNAIle binding site of Saccharomyces cerevisiae IleRS (ScIleRS), by partially mimicking the binding of tRNAIle. RM-A binding is facilitated by the copurified intermediate product isoleucyl-adenylate (Ile-AMP). The binding assays confirm that RM-A competes with tRNAIle while binding synergistically with L-isoleucine or intermediate analogue Ile-AMS to the aminoacylation pocket of ScIleRS. This study highlights that the vast tRNA binding site of the Rossmann-fold catalytic domain of class I aminoacyl-tRNA synthetases could be targeted by a small molecule. This finding will inform future rational drug design.

Synthesis and evaluation of andrographolide derivatives as potent anti-osteoporosis agents in vitro and in vivo.

In this work, we found that 14-deoxy-11,12-didehydroandrographolide (2), a derivative of andrographolide (AP, 1), had greatly reduced cytotoxicity compared with AP and exhibited moderate anti-osteoclastogenesis activity. Thirty compounds were synthesized by introducing anti-osteoporosis chemotypes at C-19 of 2. Six of them exhibited stronger inhibition of osteoclastogenesis than AP. Of note, compound 12g displayed the most potent activity with IC50 value of 0.35 µM. The expression levels of osteoclast-specific genes such as TRAcP, CTSK, NFATc1, and MMP-9 were also decreased by 12g treatment. Furthermore, Western blot and immunofluorescence analyses demonstrated that compound 12g inhibited osteoclast differentiation through downregulation of RANKL-induced NF-κB signaling pathway. In an ovariectomized (OVX) female mice model, compound 12g significantly ameliorated bone loss. Therefore, compound 12g exhibited promising in vivo efficacy and low toxicity, indicating its therapeutic potential for the treatment of osteoporosis.

Discovery of novel liver X receptor inverse agonists as lipogenesis inhibitors.

Based on the co-crystal structures of LXRß and its agonists (spiro [pyrrolidine-3,3'-oxindole] derivatives) discovered by us previously, we designed and synthesized a compound library to explore the agonistic activities. The library was screened with luciferase reporter assays, interestingly, it resulted in the discovery of 10 LXR inverse agonists besides 5 LXR agonists. To clarify the mechanism of the actions, we conducted molecular dynamics (MD) simulations on the LXR and inverse agonists complexes, and revealed that H3, H11 and H12 configurations are the key to turn on agonism or inverse agonism status for LXR. Binding tightly with H3, pushing H11 out and destabilizing H12 could form a bigger hydrophobic groove to accommodate NCOR1 to turn on LXR inverse agonism. The inverse agonist 10rr was further studied, and found as a lipogenesis inhibitor through down-regulating LXR target genes SREBP-1c, ACC, FAS and SCD-1, and demonstrated lipid-lowering effects in 3T3-L1 cells, HepG2 cells and mice with Triton WR-1339-induced hyperlipidemia. Therefore, we have proved that LXR inverse agonists can be promising agents for hyperlipidemia treatment.