Faculty Bibliography

Genome-wide association studies have implicated PLEXIN D1 (PLXND1) in body fat distribution and type 2 diabetes. However, a role for PLXND1 in regional adiposity and insulin resistance is unknown. Here we use in vivo imaging and genetic analysis in zebrafish to show that Plxnd1 regulates body fat distribution and insulin sensitivity. Plxnd1 deficiency in zebrafish induced hyperplastic morphology in visceral adipose tissue (VAT) and reduced lipid storage. In contrast, subcutaneous adipose tissue (SAT) growth and morphology were unaffected, resulting in altered body fat distribution and a reduced VAT:SAT ratio in zebrafish. A VAT-specific role for Plxnd1 appeared conserved in humans, as PLXND1 mRNA was positively associated with hypertrophic morphology in VAT, but not SAT. In zebrafish plxnd1 mutants, the effect on VAT morphology and body fat distribution was dependent on induction of the extracellular matrix protein collagen type V alpha 1 (col5a1). Furthermore, after high-fat feeding, zebrafish plxnd1 mutant VAT was resistant to expansion, and excess lipid was disproportionately deposited in SAT, leading to an even greater exacerbation of altered body fat distribution. Plxnd1-deficient zebrafish were protected from high-fat-diet-induced insulin resistance, and human VAT PLXND1 mRNA was positively associated with type 2 diabetes, suggesting a conserved role for PLXND1 in insulin sensitivity. Together, our findings identify Plxnd1 as a novel regulator of VAT growth, body fat distribution, and insulin sensitivity in both zebrafish and humans.

BACKGROUND: Dalbavancin is a lipoglycopepetide antibiotic with activity against gram positive pathogens recently approved for treatment of acute bacterial skin and skin structure infections. Pending the introduction of antimicrobial susceptibility tests, we examined the utility of vancomycin inhibitory concentrations to predict dalbavancin susceptibility in a panel of isolates obtained from phase 3 registration studies. FINDINGS: 99.6% of Staphylococcus aureus and 99.0% of beta-hemolytic streptococci which are susceptible to vancomycin will have an MIC at or below the US FDA susceptibility breakpoint for dalbavancin. CONCLUSION: Vancomycin should be considered as a surrogate for in vitro dalbavancin susceptibility testing.

Structural and functional studies of ribosomal subunits require the dissociation of intact ribosomes into individual small and large ribosomal subunits. The dissociation of the prokaryotic 70S ribosomes into the 50S and 30S subunits is achieved by dialysis against a buffer containing a lower Mg(2+) concentration. Eukaryotic 80S ribosomes are dissociated into 60S and 40S subunits by incubation in a buffer containing puromycin and higher KCl and Mg(2+) concentrations.

The prostate gland plays an important role in male reproduction, and is also an organ prone to diseases such as benign prostatic hyperplasia (BPH) and prostate cancer. The prostate consists of ducts with an inner layer of epithelium surrounded by stroma. Reciprocal signaling between these two cell compartments is instrumental to normal prostatic development, homeostasis, regeneration, as well as tumor formation. Hedgehog (HH) signaling is a master regulator in numerous developmental processes. In many organs, HH plays a key role in epithelial-mesenchymal signaling that regulates organ growth and tissue differentiation, and abnormal HH signaling has been implicated in the progression of various epithelial carcinomas. In this review, we focus on recent studies exploring the multipotency of endogenous postnatal and adult epithelial and stromal stem cells and studies addressing the role of HH in prostate development and cancer. We discuss the implications of the results for a new understanding of prostate development and disease. Insight into the cellular and molecular mechanisms underlying epithelial-mesenchymal growth regulation should provide a basis for devising innovative therapies to combat diseases of the prostate.

The synthesis of proteins in the endoplasmic reticulum (ER) that exceeds the protein folding capacity of this organelle is a frequent cause of cellular dysfunction and disease. An example of such a disease is alpha-1-antitrypsin (A1AT) deficiency, caused by destabilizing mutations in this glycoprotein. It is considered that the mutant proteins are recognized in the ER by lectins and are subsequently degraded through the proteasome, leading to a deficiency in this enzyme in the afflicted patients. We previously established a Drosophila model of this disease by overexpressing the null Hong Kong (NHK) allele of this gene and found that the Drosophila lectin, ER degradation-enhancing alpha-mannosidase-like protein 2 (EDEM2), can accelerate the degradation of A1AT when overexpressed. NHK is a rare allele, and in this study, we investigated in depth the mechanisms through which Drosophila EDEMs affect the degradation of the Z variant, which is the predominant disease allele. Specifically, we report that the Z allele does not activate ER stress signaling as prominently as the NHK allele, but similarly requires both Drosophila EDEM1 and EDEM2 for the degradation of the protein. We demonstrate that EDEMs are required for their ubiquitination, and without EDEMs, glycosylated A1AT mutants accumulate in cells. These results support the role of the EDEM-mediated ubiquitination of the alpha-1-antitrypsin Z (ATZ) allele, and establish a Drosophila model for the study of this protein and disease.

Inflammation drives atherosclerotic plaque progression and rupture, and is a compelling therapeutic target. Consequently, attenuating inflammation by reducing local macrophage accumulation is an appealing approach. This can potentially be accomplished by either blocking blood monocyte recruitment to the plaque or increasing macrophage apoptosis and emigration. Because macrophage proliferation was recently shown to dominate macrophage accumulation in advanced plaques, locally inhibiting macrophage proliferation may reduce plaque inflammation and produce long-term therapeutic benefits. To test this hypothesis, we used nanoparticle-based delivery of simvastatin to inhibit plaque macrophage proliferation in apolipoprotein E deficient mice (Apoe-/- ) with advanced atherosclerotic plaques. This resulted in rapid reduction of plaque inflammation and favorable phenotype remodeling. We then combined this short-term nanoparticle intervention with an eight-week oral statin treatment, and this regimen rapidly reduced and continuously suppressed plaque inflammation. Our results demonstrate that pharmacologically inhibiting local macrophage proliferation can effectively treat inflammation in atherosclerosis.

OBJECTIVE: The purpose of this study was to determine the role of the endothelial glucocorticoid receptor in the pathogenesis of atherosclerosis. APPROACH AND RESULTS: Control mice and mice lacking the endothelial glucocorticoid receptor were bred onto an Apoe knockout background and subjected to high-fat diet feeding for 12 weeks. Assessment of body weight and total cholesterol and triglycerides before and after the diet revealed no differences between the 2 groups of mice. However, mice lacking the endothelial glucocorticoid receptor developed more severe atherosclerotic lesions in the aorta, brachiocephalic artery, and aortic sinus, as well as a heightened inflammatory milieu as evidenced by increased macrophage recruitment in the lesions. CONCLUSIONS: These data suggest that the endothelial glucocorticoid receptor is important for tonic inhibition of inflammation and limitation of atherosclerosis progression in this model.

Latent transforming growth factor-beta binding protein-1 (LTBP-1) is an extracellular protein that is structurally similar to fibrillin and has an important role in controlling transforming growth factor-beta (TGF-beta) signaling by storing the cytokine in the extracellular matrix and by being involved in the conversion of the latent growth factor to its active form. LTBP-1 is found as both short (LTBP-1S) and long (LTBP-1L) forms, which are derived through the use of separate promoters. There is controversy regarding the importance of LTBP-1L, as Ltbp1L knockout mice showed multiple cardiovascular defects but the complete null mice did not. Here, we describe a third line of Ltbp1 knockout mice generated utilizing a conditional knockout strategy that ablated expression of both L and S forms of LTBP-1. These mice show severe developmental cardiovascular abnormalities and die perinatally; thus these animals display a phenotype similar to previously reported Ltbp1L knockout mice. We reinvestigated the other "complete" knockout line and found that these mice express a splice variant of LTBP-1L and, therefore, are not complete Ltbp1 knockouts. Our results clarify the phenotypes of Ltbp1 null mice and re-emphasize the importance of LTBP-1 in vivo.

Protein folding homeostasis in the lumen of the endoplasmic reticulum is defended by signal transduction pathways that are activated by an imbalance between unfolded proteins and chaperones (so called ER stress). Collectively referred to as the unfolded protein response (UPR) this homeostatic response is initiated by three known ER stress transducers: IRE1, PERK and ATF6. These ER-localised transmembrane (TM) proteins posses lumenal stress sensing domains and cytosolic effector domains that collectively activate a gene expression programme regulating the production of proteins involved in the processing and maturation of secreted proteins that enter the ER. However, beyond limiting unfolded protein stress in the ER the UPR has important connections to lipid metabolism that are the subject of this review.