Three Supernumerary Marker Chromosomes in a Patient with Developmental Delay, Mental Retardation, and Dysmorphic Features

We characterized three supernumerary marker chromosomes (SMCs) simultaneously present in a 2-year- and 10-month-old male patient with mental retardation and dysmorphic features. Peripheral blood chromosome analysis revealed two to three SMCs in 25/26 cells analyzed. The remaining one cell had one SMC. Microarray comparative genomic hybridization (aCGH) showed mosaicism for gains of 5q35.3, 15q11.2q13.3, and 18p11.21q11.1 regions. All three gains contain multiple OMIM genes. FISH studies indicated that one of the SMCs is a dicentric ring 15 with two copies of the 15q11.2q13.3 region including SNRPN/UBE3A and two copies of the 5q35.3 region. One of the der(18)s contains the 18 centromere and 18p11.2 regions, while the other der(18) has a signal for the 18 centromere only. The phenotype of the patient is compared with that of patients with tetrasomy 15q11.2q13.3, trisomy 5q35.3, and trisomy 18p11.2. Our study demonstrates that aCGH and FISH analyses are powerful tools, which complement the conventional cytogenetic analysis for the identification of SMCs

Bilberry Supplementation after Myocardial Infarction Decreases Microvesicles in Blood and Affects Endothelial Vesiculation

Scope: Diet rich in bilberries is considered cardioprotective, but the mechanisms of action are poorly understood. Cardiovascular disease is characterized by increased proatherogenic status and high levels of circulating microvesicles (MVs). In an open-label study patients with myocardial infarction receive an 8 week dietary supplementation with bilberry extract (BE). The effect of BE on patient MV levels and its influence on endothelial vesiculation in vitro is investigated. Methods and results: MVs are captured with acoustic trapping and platelet-derived MVs (PMVs), as well as endothelial-derived MVs (EMVs) are quantified with flow cytometry. The in vitro effect of BE on endothelial extracellular vesicle (EV) release is examined using endothelial cells and calcein staining. The mechanisms of BE influence on vesiculation pathways are studied by Western blot and qRT-PCR. Supplementation with BE decreased both PMVs and EMVs. Furthermore, BE reduced endothelial EV release, Akt phosphorylation, and vesiculation-related gene transcription. It also protects the cells from P2X7-induced EV release and increase in vesiculation-related gene expression. Conclusion: BE supplementation improves the MV profile in patient blood and reduces endothelial vesiculation through several molecular mechanisms related to the P2X7 receptor. The findings provide new insight into the cardioprotective effects of bilberries

CNTN6 copy number variations in 14 patients: A possible candidate gene for neurodevelopmental and neuropsychiatric disorders

Background: Neurodevelopmental disorders are impairments of brain function that affect emotion, learning, and memory. Copy number variations of contactin genes (CNTNs), including CNTN3, CNTN4, CNTN5, and CNTN6, have been suggested to be associated with these disorders. However, phenotypes have been reported in only a handful of patients with copy number variations involving CNTNs. Methods: From January 2009 to January 2013, 3724 patients ascertained through the University of Pittsburgh Medical Center were referred to our laboratory for clinical array comparative genomic hybridization testing. We screened this cohort of patients to identify individuals with the 3p26.3 copy number variations involving the CNTN6 gene, and then retrospectively reviewed the clinical information and family history of these patients to determine the association between the 3p26.3 copy number variations and neurodevelopmental disorders. Results: Fourteen of the 3724 patients had 3p26.3 copy number variations involving the CNTN6 gene. Thirteen of the 14 patients with these CNTN6 copy number variations presented with various neurodevelopmental disorders including developmental delay, autistic spectrum disorders, seizures and attention deficit hyperactivity disorder. Family history was available for 13 of the 14 patients. Twelve of the thirteen families have multiple members with neurodevelopmental and neuropsychiatric disorders including attention deficit hyperactivity disorder, seizures, autism spectrum disorder, intellectual disability, schizophrenia, depression, anxiety, learning disability, and bipolar disorder. Conclusions: Our findings suggest that deletion or duplication of the CNTN6 gene is associated with a wide spectrum of neurodevelopmental behavioral disorders

Mouse and rat BDNF gene structure and expression revisited

Brain-derived neurotrophic factor (BDNF) has important functions in the development of the nervous system and in brain plasticity-related processes such as memory, learning, and drug addiction. Despite the fact that the function and regulation of rodent BDNF gene expression have received close attention during the last decade, knowledge of the structural organization of mouse and rat BDNF gene has remained incomplete. We have identified and characterized several mouse and rat BDNF transcripts containing novel 5′ untranslated exons and introduced a new numbering system for mouse and rat BDNF exons. According to our results both mouse and rat BDNF gene consist of eight 5′ untranslated exons and one protein coding 3′ exon. Transcription of the gene results in BDNF transcripts containing one of the eight 5′ exons spliced to the protein coding exon and in a transcript containing only 5′ extended protein coding exon. We also report the distinct tissue-specific expression profiles of each of the mouse and rat 5′ exon-specific transcripts in different brain regions and nonneural tissues. In addition, we show that kainic acid-induced seizures that lead to changes in cellular Ca2+ levels as well as inhibition of DNA methylation and histone deacetylation contribute to the differential regulation of the expression of BDNF transcripts. Finally, we confirm that mouse and rat BDNF gene loci do not encode antisense mRNA transcripts, suggesting that mechanisms of regulation for rodent and human BDNF genes differ substantially. © 2006 Wiley-Liss, Inc

P2X4 receptor function in the nervous system and current breakthroughs in pharmacology

ATP is a well-known extracellular signalling molecule and neurotransmitter known to activate purinergic P2X receptors. Information has been elucidated about the structure and gating of P2X channels following the determination of the crystal structure of P2X4 (zebrafish), however there is still much to discover regarding the role of this receptor in the central nervous system (CNS). In this review we provide an overview of what is known about P2X4 expression in the CNS and discuss evidence for pathophysiological roles in neuroinflammation and neuropathic pain. Recent advances in the development of pharmacological tools including selective antagonists (5-BDBD, PSB-12062, BX430) and positive modulators (ivermectin, avermectins, divalent cations) of P2X4 will be discussed

Adenosine Triphosphate Release and P2 Receptor Signaling in Piezo1 Channel-Dependent Mechanoregulation

Organs and tissues and their constituent cells are physiologically submitted to diverse types of mechanical forces or stress, one common sequence of which is release of intracellular ATP into extracellular space. Extracellular ATP is a well-established autocrine or paracrine signaling molecule that regulates multiple cell functions and mediates cell-to-cell communications via activating the purinergic P2 receptors, more specifically, ligand-gated ion channel P2X receptors and some of the G-protein-coupled P2Y receptors. The molecular mechanisms that sense mechanical and transduce forces to trigger ATP release are poorly understood. The Piezo1, a newly identified mechanosensing ion channel, shows widespread expression and confers mechanosensitivity in many different types of cells. In this mini-review, we briefly introduce the Piezo1 channel and discuss the evidence that supports its important role in the mechanoregulation of diverse cell functions and, more specifically, critical engagement of ATP release and subsequent P2 receptor activation in Piezo1 channel-dependent mechanoregulation. Such ATP release-mediated coupling of the Piezo1 channel and P2 receptors may serve a signaling mechanism that is more common than we currently understand in transducing mechanical information to regulation of the attendant cell functions in various organs and tissues

A MATTER OF NUTRIENT EXCESS AND SHEAR STRESS

The metabolic syndrome is a clustering of several risk factors associated with diabetes and cardiovascular disease. Diabetes is a global problem with 90% of patients suffering from type 2 diabetes (T2D), which is attributable to various factors including genetic predispositions and lifestyle. In T2D, there is a reduction in β cell mass and function, and impaired glucose homeostasis. It is often associated with insulin resistance in target tissues and has a profound influence on vascular health. This, thesis investigates the novel role of cocaine and amphetamine regulated transcript (CART) in the pancreatic β cells and purinergic signaling mechanisms in the vascular endothelial cells. Using rat pancreatic clonal β cell lines (INS-1 832/13) and rat islets, we provide evidence for a novel role of CART in β cell survival and proliferation. Exogenous addition of CART 55-102 attenuated glucotoxicity-induced β cell death. This effect of CART was likely due to the activation of signaling molecules important for cell survival, namely, CREB, IRS-2, PKB, FoxO1, p44/42-MAPK, and p90RSK. Furthermore, the pharmacological inhibition of the kinases blocked the proliferative effect of CART on β cells. In conclusion, CART 55-102 protects β cells against glucotoxicity and promotes proliferation and may thereby contribute to the preservation of β cell mass. Physical inactivity and metabolic risk factors together form a continuum that over time negatively affects endothelial function. The vascular endothelium is exposed to fluid shear stress from flowing blood and maintains vascular homeostasis. The resident endothelial cells express a repertoire of specialized purinergic receptors that respond to extracellular nucleotides mediating various physiological responses. In human vascular endothelial cells, both ATPγS and shear stress increased KLF2, a transcription factor important for atheroprotection, in part via the P2X4 receptor. Additionally, shear stress-induced endothelial cell alignment and cytoskeletal remodeling, as seen in atheroprotective regions of the vascular tree, was found to be P2Y2 receptor-dependent. Using pharmacological inhibitors of P2X7 and P2X4, we observed differential effects of these receptors in inhibiting inflammatory genes, reactive oxygen species, and leukocyte adhesion in endothelial cells exposed to high glucose and palmitate. In addition, we demonstrate a novel role of UTP and ATP in regulating miR-22 transcription that inhibits ICAM-1 and leukocyte-endothelial adhesion, an effect possibly mediated by P2Y2 receptors. In conclusion, this thesis provides a molecular basis for treatment strategies, albeit at a cellular level, aiming at (i) preserving β cell mass and function and (ii) enhancing vascular health that range from lifestyle interventions to specific pharmacological therapies

The ATP Receptors P2X7 and P2X4 Modulate High Glucose and Palmitate-Induced Inflammatory Responses in Endothelial Cells.

Endothelial cells lining the blood vessels are principal players in vascular inflammatory responses. Dysregulation of endothelial cell function caused by hyperglycemia, dyslipidemia, and hyperinsulinemia often result in impaired vasoregulation, oxidative stress, inflammation, and altered barrier function. Various stressors including high glucose stimulate the release of nucleotides thus initiating signaling via purinergic receptors. However, purinergic modulation of inflammatory responses in endothelial cells caused by high glucose and palmitate remains unclear. In the present study, we investigated whether the effect of high glucose and palmitate is mediated by P2X7 and P2X4 and if they play a role in endothelial cell dysfunction. Transcript and protein levels of inflammatory genes as well as reactive oxygen species production, endothelial-leukocyte adhesion, and cell permeability were investigated in human umbilical vein endothelial cells exposed to high glucose and palmitate. We report high glucose and palmitate to increase levels of extracellular ATP, expression of P2X7 and P2X4, and inflammatory markers. Both P2X7 and P2X4 antagonists inhibited high glucose and palmitate-induced interleukin-6 levels with the former having a significant effect on interleukin-8 and cyclooxygenase-2. The effect of the antagonists was confirmed with siRNA knockdown of the receptors. In addition, P2X7 mediated both high glucose and palmitate-induced increase in reactive oxygen species levels and decrease in endothelial nitric oxide synthase. Blocking P2X7 inhibited high glucose and palmitate-induced expression of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 as well as leukocyte-endothelial cell adhesion. Interestingly, high glucose and palmitate enhanced endothelial cell permeability that was dependent on both P2X7 and P2X4. Furthermore, antagonizing the P2X7 inhibited high glucose and palmitate-mediated activation of p38-mitogen activated protein kinase. These findings support a novel role for P2X7 and P2X4 coupled to induction of inflammatory molecules in modulating high glucose and palmitate-induced endothelial cell activation and dysfunction