Integration of Continuous Glucose Monitoring Data into an Electronic Health Record System: Single-Center Implementation.

Managing data from continuous glucose monitoring (CGM) systems presents challenges to health care provider teams that rely on the electronic health record (EHR) during patient visits. A method of integrating CGM data with the EHR that relies on the Dexcom API was developed by Northwestern Medicine and Dexcom to address these challenges. Here, we describe the data management steps and user interface of the integrated system.

Post-sequelae symptoms and comorbidities after COVID-19.

The frequency, severity, and forms of symptoms months after coronavirus 2019 (COVID-19) are poorly understood, especially in community settings. To better understand and characterize symptoms months after community-based COVID-19, a retrospective cohort analysis was conducted. Three hundred and twenty-eight consecutive persons with a positive test for SARS-CoV-2 in the Johns Hopkins Health System, Maryland, March-May 2020, were selected for the study.

Single-molecule analysis of fluorescently labeled G-protein-coupled receptors reveals complexes with distinct dynamics and organization.

G-protein-coupled receptors (GPCRs) constitute the largest family of receptors and major pharmacological targets. Whereas many GPCRs have been shown to form di-/oligomers, the size and stability of such complexes under physiological conditions are largely unknown. Here, we used direct receptor labeling with SNAP-tags and total internal reflection fluorescence microscopy to dynamically monitor single receptors on intact cells and thus compare the spatial arrangement, mobility, and supramolecular organization of three prototypical GPCRs: the β(1)-adrenergic receptor (β(1)AR), the β(2)-adrenergic receptor (β(2)AR), and the γ-aminobutyric acid (GABA(B)) receptor.

Rotational diffusion of the α(2a) adrenergic receptor revealed by FlAsH labeling in living cells.

The fluorescein arsenical hairpin binder (FlAsH) shows much promise to determine the relative orientations of protein regions and structures even in living cells and in the plasma membrane. In this study, we characterized FlAsH's photophysical properties by steady-state anisotropy and time-resolved single photon counting for further applications with G-protein coupled receptors. We find that FlAsH has a relatively high initial anisotropy of 0.

Fluorescent labeling of tetracysteine-tagged proteins in intact cells.

In this paper, we provide a general protocol for labeling proteins with the membrane-permeant fluorogenic biarsenical dye fluorescein arsenical hairpin binder-ethanedithiol (FlAsH-EDT₂). Generation of the tetracysteine-tagged protein construct by itself is not described, as this is a protein-specific process. This method allows site-selective labeling of proteins in living cells and has been applied to a wide variety of proteins and biological problems.

Sensing G protein-coupled receptor activation.

G protein-coupled receptors (GPCRs) are the key elements of a highly regulated transduction machinery that generates different signaling outcomes to hormones and neurotransmitters. Until recently, it was assumed that diverse ligands of a given GPCR differ only in their ability to alter the balance between the OFF and the ON state of the receptor. However, it has now become evident that their activation mechanisms are more complex and that receptors presumably display distinguishable active conformational states, which are induced by different agonists and correlate to specific signaling outputs.

Formation of a ternary complex among NHERF1, beta-arrestin, and parathyroid hormone receptor.

β-Arrestins are crucial regulators of G-protein coupled receptor (GPCR) signaling, desensitization, and internalization. Despite the long-standing paradigm that agonist-promoted receptor phosphorylation is required for β-arrestin2 recruitment, emerging evidence suggests that phosphorylation-independent mechanisms play a role in β-arrestin2 recruitment by GPCRs. Several PDZ proteins are known to interact with GPCRs and serve as cytosolic adaptors to modulate receptor signaling and trafficking.

Site-specific, orthogonal labeling of proteins in intact cells with two small biarsenical fluorophores.

The fusion of fluorescent proteins to proteins of interest has greatly advanced fluorescence microscopy, but is often limited by their large size. Here, we report site-specific, orthogonal labeling of two cellular proteins in intact cells with two small fluorescent dyes: fluorescein arsenical hairpin binder, FlAsH, and its red analogue, ReAsH, which bind to tetracysteine motifs. Development of a sequential labeling method to two different motifs, CCPGCC and FLNCCPGCCMEP, allowed site-specific labeling with FlAsH and ReAsH, respectively.

Regulation of the glutamate transporter EAAT4 by PIKfyve.

The excitatory amino-acid transporter EAAT4 (SLC1A6), a Na(+),glutamate cotransporter expressed mainly in Purkinje cells, serves to clear glutamate from the synaptic cleft. EAAT4 activity is stimulated by the serum and glucocorticoid inducible kinase SGK1. SGK1-dependent regulation of the Na(+),glucose transporter SGLT1 (SLC5A1) and the creatine transporter CreaT (SLC6A8) has recently been shown to involve the mammalian phosphatidylinositol-3-phosphate-5-kinase PIKfyve (PIP5K3).

[Chromatographic determination of radiochemical purity - replacement of ITLC SG].

Unlabelled: Thin layer chromatography is well established for quality control of radiopharmaceuticals. A convenient and widely used stationary phase are ITLC SG strips. However, the Pall Corporation stopped manufacturing of the silica gel impregnated glass fibre strips (ITLC SG).

Regulation of the glutamate transporter EAAT2 by PIKfyve.

The Na(+),glutamate cotransporter EAAT2 is expressed in astrocytes and clears glutamate from the synaptic cleft. EAAT2 dependent currrent is stimulated by the serum and glucocorticoid inducible kinase SGK1. Phosphorylation targets of SGK1 include the human phosphatidylinositol-3-phosphate-5-kinase PIKfyve (PIP5K3).

Differential proteoglycan expression in two spinal cord regions after dorsal root injury.

Dorsal root injury leads to reactive gliosis in the spinal cord dorsal root entry zone and dorsal column, two regions that undergo Wallerian degeneration, but have distinct growth-inhibitory properties. This disparity could in part be due to differences in the number of degenerating sensory fibers, differences in glial cell activation, and/or to differential expression of growth-inhibitory molecules such as chondroitin sulfate proteoglycans. Laser capture microdissection of these two spinal cord white matter regions, followed by quantitative analysis of mRNA expression by real-time PCR, revealed that glial marker transcripts were differentially expressed post-injury and that the chondroitin sulfate proteoglycans Brevican and Versican V1 and V2 were preferentially up-regulated in the dorsal root entry zone, but not the dorsal column.

Regulation of the Na(+)-coupled glutamate transporter EAAT3 by PIKfyve.

The Na(+), glutamate cotransporter EAAT3 is expressed in a wide variety of tissues. It accomplishes transepithelial transport and the cellular uptake of acidic amino acids. Regulation of EAAT3 activity involves a signaling cascade including the phosphatidylinositol-3 (PI3)-kinase, the phosphoinositide dependent kinase PDK1, and the serum and glucocorticoid inducible kinase SGK1.

Local delivery of glial cell line-derived neurotrophic factor improves facial nerve regeneration after late repair.

Objectives/hypothesis: Facial nerve regeneration is limited in some clinical situations: in long grafts, by aged patients, and when the delay between nerve lesion and repair is prolonged. This deficient regeneration is due to the limited number of regenerating nerve fibers, their immaturity and the unresponsiveness of Schwann cells after a long period of denervation. This study proposes to apply glial cell line-derived neurotrophic factor (GDNF) on facial nerve grafts via nerve guidance channels to improve the regeneration.

Embryo development, stress protein (Hsp70) responses, and histopathology in zebrafish (Danio rerio) following exposure to nickel chloride, chlorpyrifos, and binary mixtures of them.

Two different classes of chemicals were tested in a multilevel approach in this study: NiCl(2) as a representative for heavy metals and chlorpyrifos, a pesticide. Both, the single substances and mixtures of them were investigated for their effects on embryonic development, histological alterations, and the stress protein (Hsp70) response in the zebrafish Danio rerio. Fishes were exposed from fertilization of eggs up to a maximum of 168 h post fertilization, depending on the investigated endpoint.

Fluorescence resonance energy transfer analysis of alpha 2a-adrenergic receptor activation reveals distinct agonist-specific conformational changes.

Several lines of evidence suggest that G-protein-coupled receptors can adopt different active conformations, but their direct demonstration in intact cells is still missing. Using a fluorescence resonance energy transfer (FRET)-based approach we studied conformational changes in alpha(2A)-adrenergic receptors in intact cells. The receptors were C-terminally labeled with cyan fluorescent protein and with fluorescein arsenical hairpin binder at different sites in the third intracellular loop: N-terminally close to transmembrane domain V (I3-N), in the middle of the loop (I3-M), or C-terminally close to transmembrane domain VI (I3-C).

Persisting atypical and cystic forms of Borrelia burgdorferi and local inflammation in Lyme neuroborreliosis.

Background: The long latent stage seen in syphilis, followed by chronic central nervous system infection and inflammation, can be explained by the persistence of atypical cystic and granular forms of Treponema pallidum. We investigated whether a similar situation may occur in Lyme neuroborreliosis.

Method: Atypical forms of Borrelia burgdorferi spirochetes were induced exposing cultures of Borrelia burgdorferi (strains B31 and ADB1) to such unfavorable conditions as osmotic and heat shock, and exposure to the binding agents Thioflavin S and Congo red.

Delayed priming promotes CNS regeneration post-rhizotomy in Neurocan and Brevican-deficient mice.

A wealth of literature has provided evidence that reactive tissue at the site of CNS injury is rich in chondroitin sulfate proteoglycans which may contribute to the non-permissive nature of the CNS. We have recently demonstrated using a murine model of human brachial plexus injury that the chondroitin sulfate proteoglycans Neurocan and Brevican are differentially expressed by two subsets of astrocytes in the spinal cord dorsal root entry zone (DREZ) following dorsal root lesion (Beggah et al., Neuroscience 133: 749-762, 2005).

Conformational changes in G-protein-coupled receptors-the quest for functionally selective conformations is open.

The G-protein-coupled receptors (GPCRs) represent one the largest families of drug targets. Upon agonist binding a receptor undergoes conformational rearrangements that lead to a novel protein conformation which in turn can interact with effector proteins. During the last decade significant progress has been made to prove that different conformational changes occur.

Lentivirus-mediated expression of glutathione peroxidase: neuroprotection in murine models of Parkinson's disease.

Reactive oxygen species are considered to contribute to the pathogenesis of Parkinson's disease (PD). In order to study viral vector-mediated overexpression of the antioxidant enzyme glutathione peroxidase (GPX) as a potential neuroprotective approach in both an in vitro and in vivo model of PD, we have developed a lentiviral vector carrying the human GPX1 gene. Neuroblastoma cells infected with this vector showed a 2-fold increase in GPX activity compared to cells infected with a control vector.

Progressive and selective striatal degeneration in primary neuronal cultures using lentiviral vector coding for a mutant huntingtin fragment.

A lentiviral vector expressing a mutant huntingtin protein (htt171-82Q) was used to generate a chronic model of Huntington's disease (HD) in rat primary striatal cultures. In this model, the majority of neurons expressed the transgene so that Western blot analysis and flow cytometry measurement could complement immunohistological evaluation. Mutant huntingtin produced a slowly progressing pathology characterized after 1 month by the appearance of neuritic aggregates followed by intranuclear inclusions, morphological anomalies of neurites, loss of neurofilament 160, increased expression in stress response protein Hsp70, and later loss of neuronal markers such as NeuN and MAP-2.

Increased fiber outgrowth from xeno-transplanted human embryonic dopaminergic neurons with co-implants of polymer-encapsulated genetically modified cells releasing glial cell line-derived neurotrophic factor.

We investigated whether a continuous supply of glial cell line-derived neurotrophic factor (GDNF) via encapsulated genetically modified cells can promote survival and fiber outgrowth from xenotransplanted human dopaminergic neurons. Cells genetically engineered to continuously secrete GDNF were confined in hollow fiber-based macrocapsules. Each hemiparkinsonian rat received either a single C2C12-hGDNF capsule (n=8) or a C2C12-control capsule (n=8) concomitantly with human embryonic ventral mesencephalic cell suspension transplants.

Lesion-induced differential expression and cell association of Neurocan, Brevican, Versican V1 and V2 in the mouse dorsal root entry zone.

Lack of regeneration in the CNS has been attributed to many causes, including the presence of inhibitory molecules such as chondroitin sulfate proteoglycans (CSPGs). However, little is known about the contribution of CSPGs to regeneration failure in vivo, in particular at the dorsal root entry zone (DREZ), a unique CNS region that blocks regeneration of sensory fibers following dorsal root injury without glial scar formation. The goal of the present study was to evaluate the presence, regulation, and cellular identity of the proteoglycans Brevican, Neurocan, Versican V1 and Versican V2 in the DREZ using CSPG-specific antibodies and nucleic acid probes.

Local GDNF expression mediated by lentiviral vector protects facial nerve motoneurons but not spinal motoneurons in SOD1(G93A) transgenic mice.

Approximately 2% of amyotrophic lateral sclerosis (ALS) cases are associated with mutations in the cytosolic Cu/Zn superoxide dismutase 1 (SOD1) gene. Transgenic SOD1 mice constitute useful models of ALS to screen therapeutical approaches. Glial cell line-derived neurotrophic factor (GDNF) holds promises for the treatment of motoneuron disease.