Faculty Bibliography

OBJECTIVE: This study examined the effect of adjunctive intranasal insulin therapy on body metabolism in patients with schizophrenia. METHOD: Each subject had a DSM-IV diagnosis of schizophrenia or schizoaffective disorder and had been on stable dose of antipsychotic agent for at least one month. In an 8-week randomized, double-blind, placebo-controlled study, subjects received either intranasal insulin (40IU 4 times per day) or placebo. The whole body dual-energy X-ray absorptiometry (DXA) was used to assess body composition. Lipid particles were assessed using nuclear magnetic resonance (NMR) spectroscopy. All assessments were conducted at baseline, and repeated at week 8. RESULTS: A total number of 39 subjects completed the study (18 in the insulin group, 21 in the placebo group). There were no significant differences between the two groups in week 8 changes for body weight, body mass index, waist circumference, as well as various measures of lipid particles (p's>0.100). The DXA assessment showed no significant differences between the two groups in week 8 changes for fat mass, lean mass or total mass (p's>0.100). CONCLUSION: In the present study, adjunctive therapy of intranasal insulin did not seem to improve body metabolism in patients with schizophrenia. The implications for future studies were discussed.

Introduction: Most cases of familial arrhythmogenic right ventricular cardiomyopathy (ARVC) associate with mutations in desmosomal proteins, most commonly plakophilin-2 (PKP2). A crosstalk between PKP2 and connexin43 (Cx43) has been proposed as a pathogenic mechanism. We speculate that a) Cx43 and PKP2 are in close physical proximity, allowing for direct intermolecular interaction and b) the structure of the Cx43- PKP2 complex depends on expression of the scaffolding protein ankyrin-G (AnkG). To test these hypotheses, we implemented a novel method (direct stochastic reconstruction microscopy; dSTORM) that allows for spatial resolution of fluorescence microscopy images in the nanoscale. Methods: Neonatal rat ventricular myocytes were labeled with antibodies to Cx43 and PKP2 and imaged using a custom- made microscopy system. On-off cycles of light emission were recorded in 2000 frames, and the image reconstructed by custom-made software. Cells were treated with siRNAfor AnkG, or non-targeted constructs, and the characteristics of Cx43 and PKP2 clusters compared to control. Results: Optical resolution of dSTORM images was 20 nm. Cx43 was found in circular clusters of two predominant sizes: 13313+/-328 and 25035+226 nm^2. PKP2 clusters were of various shapes and widespread size distribution, but consistently found less than 40 nm away from a Cx43 plaque, with signals overlapping on the edges of the plaques. Loss of AnkG expression drastically altered Cx43 cluster morphology becoming less circular and of a larger dimension. Close proximity to PKP2 was maintained. Yet, the total number of PKP2 clusters was significantly decreased. Conclusion: We implemented a method that breaks the optical resolution barrier imposed by the diffraction properties of light (~300 nm), to reach a range previously reserved to electron microscopy (~20 nm). We demonstrate that PKP2 populates the edge of Cx43 plaques (the perinexus). Cx43 cluster architecture depends on AnkG expression and likely, Cx43-cytoskeletal interacti!

The cardiac intercalated disc harbors mechanical and electrical junctions as well as ion channel complexes mediating propagation of electrical impulses. Cardiac connexin43 (Cx43) co-localizes and interacts with several of the proteins located at intercalated discs in the ventricular myocardium. We have generated conditional Cx43D378stop mice lacking the last five C-terminal amino acid residues, representing a binding motif for zonula occludens protein-1 (ZO-1), and investigated the functional consequences of this mutation on cardiac physiology and morphology. Newborn and adult homozygous Cx43D378stop mice displayed markedly impaired and heterogeneous cardiac electrical activation properties and died from severe ventricular arrhythmias. Cx43 and ZO-1 were co-localized at intercalated discs in Cx43D378stop hearts, and the Cx43D378stop gap junction channels showed normal coupling properties. Patch clamp analyses of isolated adult Cx43D378stop cardiomyocytes revealed a significant decrease in sodium and potassium current densities. Furthermore, we also observed a significant loss of Nav1.5 protein from intercalated discs in Cx43D378stop hearts. The phenotypic lethality of the Cx43D378stop mutation was very similar to the one previously reported for adult Cx43 deficient (Cx43KO) mice. Yet, in contrast to Cx43KO mice, the Cx43 gap junction channel was still functional in the Cx43D378stop mutant. We conclude that the lethality of Cx43D378stop mice is independent of the loss of gap junctional intercellular communication, but most likely results from impaired cardiac sodium and potassium currents. The Cx43D378stop mice reveal for the first time that Cx43 dependent arrhythmias can develop by mechanisms other than impairment of gap junction channel function.

Inflammation of human bronchial epithelia (HBE) activates the endoplasmic reticulum (ER) stress transducer inositol-requiring enzyme 1 (IRE1)alpha, resulting in IRE1alpha-mediated cytokine production. Previous studies demonstrated ubiquitous expression of IRE1alpha and gut-restricted expression of IRE1beta. We found that IRE1beta is also expressed in HBE, is absent in human alveolar cells, and is upregulated in cystic fibrosis and asthmatic HBE. Studies with Ire1beta(-/-) mice and Calu-3 airway epithelia exhibiting IRE1beta knockdown or overexpression revealed that IRE1beta is expressed in airway mucous cells, is functionally required for airway mucin production, and this function is specific for IRE1beta vs. IRE1alpha. IRE1beta-dependent mucin production is mediated, at least in part, by activation of the transcription factor X-box binding protein-1 (XBP-1) and the resulting XBP-1-dependent transcription of anterior gradient homolog 2, a gene implicated in airway and intestinal epithelial mucin production. These novel findings suggest that IRE1beta is a potential mucous cell-specific therapeutic target for airway diseases characterized by mucin overproduction.

MicroRNAs (miRNAs) are small, ~22 nucleotide (nt) sequences of RNA that regulate gene expression at posttranscriptional level. These endogenous gene expression inhibitors were primarily described in cancer but recent exciting findings have also demonstrated a key role in cardiovascular diseases (CVDs), including atherosclerosis. MiRNAs control endothelial cell (EC), vascular smooth muscle cell (VSMC), and macrophage functions, and thereby regulate the progression of atherosclerosis. MiRNA expression is modulated by different stimuli involved in every stage of atherosclerosis, and conversely miRNAs modulates several pathways implicated in plaque development such as cholesterol metabolism. In the present review, we focus on the importance of miRNAs in atherosclerosis, and we further discuss their potential use as biomarkers and therapeutic targets in CVDs.

INTRODUCTION: Acne vulgaris is one of the most common skin conditions in children and adolescents. The presentation, differential diagnosis, and association of acne with systemic pathology differs by age of presentation. Current acknowledged guidelines for the diagnosis and management of pediatric acne are lacking, and there are variations in management across the spectrum of primary and specialty care. The American Acne and Rosacea Society convened a panel of pediatric dermatologists, pediatricians, and dermatologists with expertise in acne to develop recommendations for the management of pediatric acne and evidence-based treatment algorithms. METHODS: Ten major topic areas in the diagnosis and treatment of pediatric acne were identified. A thorough literature search was performed and articles identified, reviewed, and assessed for evidence grading. Each topic area was assigned to 2 expert reviewers who developed and presented summaries and recommendations for critique and editing. Furthermore, the Strength of Recommendation Taxonomy, including ratings for the strength of recommendation for a body of evidence, was used throughout for the consensus recommendations for the evaluation and management of pediatric acne. Practical evidence-based treatment algorithms also were developed. RESULTS: Recommendations were put forth regarding the classification, diagnosis, evaluation, and management of pediatric acne, based on age and pubertal status. Treatment considerations include the use of over-the-counter products, topical benzoyl peroxide, topical retinoids, topical antibiotics, oral antibiotics, hormonal therapy, and isotretinoin. Simplified treatment algorithms and recommendations are presented in detail for adolescent, preadolescent, infantile, and neonatal acne. Other considerations, including psychosocial effects of acne, adherence to treatment regimens, and the role of diet and acne, also are discussed. CONCLUSIONS: These expert recommendations by the American Acne and Rosacea Society as reviewed and endorsed by the American Academy of Pediatrics constitute the first detailed, evidence-based clinical guidelines for the management of pediatric acne including issues of special concern when treating pediatric patients.

Eukaryotic cells respond to stress caused by the accumulation of unfolded/misfolded proteins in the endoplasmic reticulum by activating the intracellular signaling pathways referred to as the unfolded protein response (UPR). In metazoans, UPR consists of three parallel branches, each characterized by its stress sensor protein, IRE1, ATF6, and PERK, respectively. In Drosophila, IRE1/XBP1 pathway is considered to function as a major branch of UPR; however, its physiological roles during the normal development and homeostasis remain poorly understood. To visualize IRE1/XBP1 activity in fly tissues under normal physiological conditions, we modified previously reported XBP1 stress sensing systems (Souid et al., Dev Genes Evol 217: 159-167, 2007; Ryoo et al., EMBO J 26: 242-252, 2007), based on the recent reports regarding the unconventional splicing of XBP1/HAC1 mRNA (Aragon et al., Nature 457: 736-740, 2009; Yanagitani et al., Mol Cell 34: 191-200, 2009; Science 331: 586-589, 2011). The improved XBP1 stress sensing system allowed us to detect new IRE1/XBP1 activities in the brain, gut, Malpighian tubules, and trachea of third instar larvae and in the adult male reproductive organ. Specifically, in the larval brain, IRE1/XBP1 activity was detected exclusively in glia, although previous reports have largely focused on IRE1/XBP1 activity in neurons. Unexpected glial IRE1/XBP1 activity may provide us with novel insights into the brain homeostasis regulated by the UPR.

OBJECTIVE: Emerging evidence suggests that neuronal guidance cues, typically expressed during development, are involved in both physiological and pathological immune responses. We hypothesized that endothelial expression of such guidance cues may regulate leukocyte trafficking into the vascular wall during atherogenesis. Approach and Results- We demonstrate that members of the netrin, semaphorin, and ephrin family of guidance molecules are differentially regulated under conditions that promote or protect from atherosclerosis. Netrin-1 and semaphorin3A are expressed by coronary artery endothelial cells and potently inhibit chemokine-directed migration of human monocytes. Endothelial expression of these negative guidance cues is downregulated by proatherogenic factors, including oscillatory shear stress and proinflammatory cytokines associated with monocyte entry into the vessel wall. Furthermore, we show using intravital microscopy that inhibition of netrin-1 or semaphorin3A using blocking peptides increases leukocyte adhesion to the endothelium. Unlike netrin-1 and semaphorin3A, the guidance cue ephrinB2 is upregulated under proatherosclerotic flow conditions and functions as a chemoattractant, increasing leukocyte migration in the absence of additional chemokines. CONCLUSIONS: The concurrent regulation of negative and positive guidance cues may facilitate leukocyte infiltration of the endothelium through a balance between chemoattraction and chemorepulsion. These data indicate a previously unappreciated role for axonal guidance cues in maintaining the endothelial barrier and regulating leukocyte trafficking during atherogenesis.

OBJECTIVE: The persistence of myeloid-derived cells in the artery wall is a characteristic of advanced atherosclerotic plaques. However, the mechanisms by which these cells are retained are poorly understood. Semaphorins, a class of neuronal guidance molecules, play a critical role in vascular patterning and development, and recent studies suggest that they may also have immunomodulatory functions. The present study evaluates the expression of Semaphorin 3E (Sema3E) in settings relevant to atherosclerosis and its contribution to macrophage accumulation in plaques. Approach and Results- Immunofluorescence staining of Sema3E, and its receptor PlexinD1, demonstrated their expression in macrophages of advanced atherosclerotic lesions of Apoe(-/-) mice. Notably, in 2 different mouse models of atherosclerosis regression, Sema3E mRNA was highly downregulated in plaque macrophages, coincident with a reduction in plaque macrophage content and an enrichment in markers of reparative M2 macrophages. In vitro, Sema3E mRNA was highly expressed in inflammatory M1 macrophages and in macrophages treated with physiological drivers of plaque progression and inflammation, such as oxidized low-density lipoprotein and hypoxia. To explore mechanistically how Sema3E affects macrophage behavior, we treated macrophages with recombinant protein in the presence/absence of chemokines, including CCL19, a chemokine implicated in the egress of macrophages from atherosclerotic plaques. Sema3E blocked actin polymerization and macrophage migration stimulated by the chemokines, suggesting that it may immobilize these cells in the plaque. CONCLUSIONS: Sema3E is upregulated in macrophages of advanced plaques, is dynamically regulated by multiple atherosclerosis-relevant factors, and acts as a negative regulator of macrophage migration, which may promote macrophage retention and chronic inflammation in vivo.