Transferrin receptor 1-mediated iron uptake regulates bone mass in mice via osteoclast mitochondria and cytoskeleton.

Increased intracellular iron spurs mitochondrial biogenesis and respiration to satisfy high-energy demand during osteoclast differentiation and bone-resorbing activities. Transferrin receptor 1 (Tfr1) mediates cellular iron uptake through endocytosis of iron-loaded transferrin, and its expression increases during osteoclast differentiation. Nonetheless, the precise functions of Tfr1 and Tfr1-mediated iron uptake in osteoclast biology and skeletal homeostasis remain incompletely understood. To investigate the role of Tfr1 in osteoclast lineage cells in vivo and in vitro, we crossed Tfrc (encoding Tfr1)-floxed mice with Lyz2 (LysM)-Cre and Cathepsin K (Ctsk)-Cre mice to generate Tfrc conditional knockout mice in myeloid osteoclast precursors (Tfr1ΔLysM) or differentiated osteoclasts (Tfr1ΔCtsk), respectively. Skeletal phenotyping by µCT and histology unveiled a significant increase in trabecular bone mass with normal osteoclast number in long bones of 10-week-old young and 6-month-old adult female but not male Tfr1ΔLysM mice. Although high trabecular bone volume in long bones was observed in both male and female Tfr1ΔCtsk mice, this phenotype was more pronounced in female knockout mice. Consistent with this gender-dependent phenomena, estrogen deficiency induced by ovariectomy decreased trabecular bone mass in Tfr1ΔLysM mice. Mechanistically, disruption of Tfr1 expression attenuated mitochondrial metabolism and cytoskeletal organization in mature osteoclasts in vitro by attenuating mitochondrial respiration and activation of the Src-Rac1-WAVE regulatory complex axis, respectively, leading to decreased bone resorption with little impact on osteoclast differentiation. These results indicate that Tfr1-mediated iron uptake is specifically required for osteoclast function and is indispensable for bone remodeling in a gender-dependent manner.

The Cytoplasmic Dynein Associated Protein NDE1 Regulates Osteoclastogenesis by Modulating M-CSF and RANKL Signaling Pathways.

Cytoskeleton organization and lysosome secretion play an essential role in osteoclastogenesis and bone resorption. The cytoplasmic dynein is a molecular motor complex that regulates microtubule dynamics and transportation of cargos/organelles, including lysosomes along the microtubules. LIS1, NDE1, and NDEL1 belong to an evolutionary conserved pathway that regulates dynein functions. Disruption of the cytoplasmic dynein complex and deletion of LIS1 in osteoclast precursors arrest osteoclastogenesis. Nonetheless, the role of NDE1 and NDEL1 in osteoclast biology remains elusive. In this study, we found that knocking-down Nde1 expression by lentiviral transduction of specific shRNAs markedly inhibited osteoclastogenesis in vitro by attenuating the proliferation, survival, and differentiation of osteoclast precursor cells via suppression of signaling pathways downstream of M-CSF and RANKL as well as osteoclast differentiation transcription factor NFATc1. To dissect how NDEL1 regulates osteoclasts and bone homeostasis, we generated Ndel1 conditional knockout mice in myeloid osteoclast precursors (Ndel1ΔlysM) by crossing Ndel1-floxed mice with LysM-Cre mice on C57BL/6J background. The Ndel1ΔlysM mice developed normally. The µCT analysis of distal femurs and in vitro osteoclast differentiation and functional assays in cultures unveiled the similar bone mass in both trabecular and cortical bone compartments as well as intact osteoclastogenesis, cytoskeleton organization, and bone resorption in Ndel1ΔlysM mice and cultures. Therefore, our results reveal a novel role of NDE1 in regulation of osteoclastogenesis and demonstrate that NDEL1 is dispensable for osteoclast differentiation and function.

Selective Inhibition of Aurora Kinase A by AK-01/LY3295668 Attenuates MCC Tumor Growth by Inducing MCC Cell Cycle Arrest and Apoptosis.

Merkel cell carcinoma (MCC) is an often-lethal skin cancer with increasing incidence and limited treatment options. Although immune checkpoint inhibitors (ICI) have become the standard of care in advanced MCC, 50% of all MCC patients are ineligible for ICIs, and amongst those treated, many patients develop resistance. There is no therapeutic alternative for these patients, highlighting the urgent clinical need for alternative therapeutic strategies. Using patient-derived genetic insights and data generated in our lab, we identified aurora kinase as a promising therapeutic target for MCC. In this study, we examined the efficacy of the recently developed and highly selective AURKA inhibitor, AK-01 (LY3295668), in six patient-derived MCC cell lines and two MCC cell-line-derived xenograft mouse models. We found that AK-01 potently suppresses MCC survival through apoptosis and cell cycle arrest, particularly in MCPyV-negative MCC cells without RB expression. Despite the challenge posed by its short in vivo durability upon discontinuation, the swift and substantial tumor suppression with low toxicity makes AK-01 a strong potential candidate for MCC management, particularly in combination with existing regimens.

Metabolism, CB1 cannabinoid receptor binding and in vivo activity of synthetic cannabinoid 5F-AKB48: Implications for toxicity.

AKB48 and its fluorinated derivative 5F-AKB48 are synthetic cannabinoids (SCs) which have caused hospitalizations and deaths in human users. Abuse of SCs is dangerous because users may mistake them for natural cannabis, which is generally considered to be unlikely to elicit adverse effects. The present studies were designed to investigate the in vitro oxidative metabolism of 5F-AKB48 by human microsomal fractions from different organs and sexes as well as recombinant human cytochrome P450s (P450s). Mass spectrometry data tentatively provides evidence for the existence of mono-, di-, and trihydroxylated metabolites in a successive metabolism. Experiments utilizing P450s revealed that the most active enzymes (CYP2D6, CYP2J2, CYP3A4, and CYP3A5) effectively produced mono- and dihydroxylated metabolites, while CYP3A4/5 also produced significant amounts of the trihydroxylated metabolite. Moreover, although the affinity and potency of Phase I metabolite 4OH-5F-AKB48 is reduced when compared to that of the parent drug, this metabolite nevertheless retains similar high affinity for CB1 receptors, and greater efficacy for G protein activation, when compared to THC. Finally, 5F-AKB48 produced time- and dose-dependent cannabimimetic effects in mice which were more potent, but shorter acting, than those of Δ9-THC, and were attenuated by prior treatment with the CB1 antagonist rimonabant. Based on our data, we hypothesize that while many cases of toxicity result from genetic mutations, which can lead to a decrease or even absence of activity for Phase I drug-metabolizing enzymes, other P450s could potentially increase their role in the metabolism of these SCs. Because many metabolites of SCs remain biologically active, they could contribute to the deleterious effects of these substances.

Synthesis, optimization and applications of ZnO/polymer nanocomposites.

Polymer composites have established an excellent position among the technologically essential materials because of their wide range of applications. An enormous research interest has been devoted to zinc oxide (ZnO) based polymer nanocomposites, due to their exceptional electrical, optical, thermal, mechanical, catalytic, and biomedical properties. This article provides a review of various polymer composites consisting of ZnO nanoparticles (NPs) as reinforcements, exhibiting excellent properties for applications such as the dielectric, sensing, piezoelectric, electromagnetic shielding, thermal conductivity and energy storage. The preparation methods of such composites including solution blending, in situ polymerization, and melt intercalation are also explained. The current challenges and potential applications of these composites are provided in order to guide future progress on the development of more promising materials. Finally, a detailed summary of the current trends in the field is presented to progressively show the future prospects for the development of ZnO containing polymer nanocomposite materials.

Pharmacological characterizationof synthetic cannabinoid MAM-2201:radioligand binding and abuse-related effects / 中国药理学与毒理学杂志

OBJECTIVE Over 30% of all new psychoactive substances identified by the UN Office on Drugs and Crime in 2016 were synthetic cannabinoids. The recent emergence of MAM-2201 on the illicit market is troubling because this drug has no precedent in either the scientific or patent literature, and appears to be a novel compound developed specifically as a ″graymarket″ drug of abuse bystruc?turally combining the known synthetic cannabinoids JWH- 122 and AM- 2201. There is currently no published information regarding the pharmacology of MAM-2201. METHODS The present studies characterized cannabinoid-like effects of MAM-2201 in vitro (interactions with cannabinoid type 1 receptors [CB1Rs]) and in vivo (in mice and rats). RESULTS In a radioligand binding assay using [3H]CP55,940 in HEK cell membranes transfected with the CB1R, MAM-2201 (Ki=5.4 nmol·L- 1), had higher binding affinity than WIN 55,212-2 (Ki=80 nmol·L-1), and D9-THC (Ki=8.3 nmol·L-1). The Emax values for MAM-2201 and WIN 55,212-2 in an assay of agonist inhibition of forskolin-stimulated cAMP were 85% (EC50=0.45 nmol·L-1) and 95%, respectively, as compared with the D9-THC Emax of 74%. In mice, MAM-2201 (0.003-1.0 mg·kg-1, IP) produced dose-dependent cannabimimetic effects which were both more potent and more effective than those of D9-THC. MAM-2201 and D9-THC dose-dependently produced hypothermia:ED50=0.287 and 25.4 mg·kg-1, analgesia: ED50=0.125 and 29.4 mg·kg-1, and catalepsy: ED50=0.301 and 18.9 mg·kg-1 in adult male CD1 mice. Importantly, MAM-2201 also elicited convulsant effects at a dose of 1.0 mg·kg-1 in 8/8 murine subjects. In rats, MAM-2201 produced dose-dependent D9-THC-like intero?ceptive effects in subjects trained to discriminate 3.0 mg·kg-1 (IP) D9-THC from saline. CONCLUSION MAM-2201 binds CB1Rs with high affinity and agonist efficacy, and functions as a potent cannabinoid agonist in vivo across several complementary measures of cannabinoid activity in two rodent species.