Loss of Nmp4 optimizes osteogenic metabolism and secretion to enhance bone quality

A goal of osteoporosis therapy is to restore lost bone with structurally sound tissue. Mice lacking the transcription factor Nuclear Matrix Protein 4 (Nmp4, Zfp384, Ciz, ZNF384) respond to several classes of osteoporosis drugs with enhanced bone formation compared to wild type (WT) animals. Nmp4-/- mesenchymal stem/progenitor cells (MSPCs) exhibit an accelerated and enhanced mineralization during osteoblast differentiation. To address the mechanisms underlying this hyper-anabolic phenotype, we carried out RNA-sequencing and molecular and cellular analyses of WT and Nmp4-/- MSPCs during osteogenesis to define pathways and mechanisms associated with elevated matrix production. We determined that Nmp4 has a broad impact on the transcriptome during osteogenic differentiation, contributing to the expression of over 5,000 genes. Phenotypic anchoring of transcriptional data was performed for the hypothesis-testing arm through analysis of cell metabolism, protein synthesis and secretion, and bone material properties. Mechanistic studies confirmed that Nmp4-/- MSPCs exhibited an enhanced capacity for glycolytic conversion- a key step in bone anabolism. Nmp4-/- cells showed elevated collagen translation and secretion. Expression of matrix genes that contribute to bone material-level mechanical properties were elevated in Nmp4-/- cells, an observation that was supported by biomechanical testing of bone samples from Nmp4-/- and WT mice. We conclude that loss of Nmp4 increases the magnitude of glycolysis upon the metabolic switch, which fuels the conversion of the osteoblast into a super-secretor of matrix resulting in more bone with improvements in intrinsic quality

Transcriptomic Analysis of Healthy and Atopic Dermatitis Samples Reveals the Role of IL-37 in Human Skin

Atopic dermatitis (AD) is a chronic inflammatory skin disease that affects up to one in five children and millions of adults in developed countries. Clinically, AD skin lesions manifest as subacute and/or chronic lichenified eczematous plaques, which are often intensely pruritic and prone to secondary bacterial and viral infections. Despite the emergence of novel therapeutic agents, treatment options and outcomes for AD remain suboptimal. An improved understanding of AD pathogenesis may help improve patient outcomes. Dysregulated Th2-polarized skin inflammation and impaired skin barrier function interact to drive AD pathogenesis; however, much remains to be understood about the molecular mechanisms underlying this interplay. The current study used published clinical trial datasets to define a skin-related AD gene signature. This meta-analysis revealed significant reductions in IL1F7 transcripts (encodes IL-37) in AD patient samples. Reduced IL1F7 correlated with lower transcripts for key skin barrier function genes in the epidermal differentiation complex. Immunohistochemical analysis of normal (healthy) human skin specimens and an in vitro three-dimensional human skin model localized IL-37 protein to the epidermis. In comparison with normal human skin, IL-37 levels were decreased in AD patient skin. Addition of Th2 cytokines to the aforementioned in vitro three-dimensional skin model recapitulates key aspects of AD skin and was sufficient to reduce epidermal IL-37 levels. Image analysis also indicated close relationship between epidermal IL-37 and skin epidermal differentiation complex proteins. These findings suggest IL-37 is intimately linked to normal keratinocyte differentiation and barrier function and implicates IL-37 as a potential biomarker and therapeutic target for AD

Identification of heme as the ligand for the orphan nuclear receptors REV-ERBalpha and REV-ERBbeta

The nuclear receptors REV-ERBalpha (encoded by NR1D1) and REV-ERBbeta (NR1D2) have remained orphans owing to the lack of identified physiological ligands. Here we show that heme is a physiological ligand of both receptors. Heme associates with the ligand-binding domains of the REV-ERB receptors with a 1:1 stoichiometry and enhances the thermal stability of the proteins. Results from experiments of heme depletion in mammalian cells indicate that heme binding to REV-ERB causes the recruitment of the co-repressor NCoR, leading to repression of target genes including BMAL1 (official symbol ARNTL), an essential component of the circadian oscillator. Heme extends the known types of ligands used by the human nuclear receptor family beyond the endocrine hormones and dietary lipids described so far. Our results further indicate that heme regulation of REV-ERBs may link the control of metabolism and the mammalian clock.status: publishe

TGFβ-Mediated induction of SphK1 as a potential determinant in human MDA-MB-231 breast cancer cell bone metastasis

Mechanistic understanding of the preferential homing of circulating tumor cells to bone and their perturbation on bone metabolism within the tumor-bone microenvironment remains poorly understood. Alteration in both transforming growth factor β (TGFβ) signaling and sphingolipid metabolism results in the promotion of tumor growth and metastasis. Previous studies using MDA-MB-231 human breast cancer-derived cell lines of variable metastatic potential were queried for changes in sphingolipid metabolism genes to explore correlations between TGFβ dependence and bone metastatic behavior. Of these genes, only sphingosine kinase-1 (SPHK1) was identified to be significantly increased following TGFβ treatment. Induction of SPHK1 expression correlated to the degree of metastatic capacity in these MDA-MB-231-derived cell lines. We demonstrate that TGFβ mediates the regulation of SPHK1 gene expression, protein kinase activity and is critical to MDA-MB-231 cell viability. Furthermore, a bioinformatic analysis of human breast cancer gene expression supports SPHK1 as a hallmark TGFβ target gene that also bears the genetic fingerprint of the basal-like/triple-negative breast cancer molecular subtype. These data suggest a potential new signaling axis between TGFβ/SphK1 that may have a role in the development, prognosis or the clinical phenotype associated with tumor-bone metastasis

Improving Combination Osteoporosis Therapy in a Preclinical Model of Heightened Osteoanabolism

Combining anticatabolic agents with parathyroid hormone (PTH) to enhance bone mass has yielded mixed results in osteoporosis patients. Toward the goal of enhancing the efficacy of these regimens, we tested their utility in combination with loss of the transcription factor Nmp4 because disabling this gene amplifies PTH-induced increases in trabecular bone in mice by boosting osteoblast secretory activity. We addressed whether combining a sustained anabolic response with an anticatabolic results in superior bone acquisition compared with PTH monotherapy. Additionally, we inquired whether Nmp4 interferes with anticatabolic efficacy. Wild-type and Nmp4-/- mice were ovariectomized at 12 weeks of age, followed by therapy regimens, administered from 16 to 24 weeks, and included individually or combined PTH, alendronate (ALN), zoledronate (ZOL), and raloxifene (RAL). Anabolic therapeutic efficacy generally corresponded with PTH + RAL = PTH + ZOL > PTH + ALN = PTH > vehicle control. Loss of Nmp4 enhanced femoral trabecular bone increases under PTH + RAL and PTH + ZOL. RAL and ZOL promoted bone restoration, but unexpectedly, loss of Nmp4 boosted RAL-induced increases in femoral trabecular bone. The combination of PTH, RAL, and loss of Nmp4 significantly increased bone marrow osteoprogenitor number, but did not affect adipogenesis or osteoclastogenesis. RAL, but not ZOL, increased osteoprogenitors in both genotypes. Nmp4 status did not influence bone serum marker responses to treatments, but Nmp4-/- mice as a group showed elevated levels of the bone formation marker osteocalcin. We conclude that the heightened osteoanabolism of the Nmp4-/- skeleton enhances the effectiveness of diverse osteoporosis treatments, in part by increasing hyperanabolic osteoprogenitors. Nmp4 provides a promising target pathway for identifying barriers to pharmacologically induced bone formation