Real-World Assessments of mySugr Mobile Health App.

Mobile health (mHealth) solutions such as diabetes self-management apps improve glycated hemoglobin, particularly those that provide a feedback loop between patient and health care provider. mHealth apps that incorporate behaviorally designed interventions can improve patient access to diabetes self-management education and ongoing support. The mySugr mobile app was designed to support patients in their diabetes self-management. Most studies of mHealth apps were conducted under controlled conditions and did not elucidate the nuances of patient perceptions and utilization of these apps in everyday life. In this article, we discuss findings from real-world observations of changes in glycemic control and patient satisfaction associated with the use of the mySugr mHealth app.

Identification of a novel gene cluster in the upstream region of the S-layer gene sbpA involved in cell wall metabolism of Lysinibacillus sphaericus CCM 2177 and characterization of the recombinantly produced autolysin and pyruvyl transferase.

The S-layer protein SbpA of Lysinibacillus sphaericus CCM 2177 assembles into a square (p4) lattice structure and recognizes a pyruvylated secondary cell wall polymer (SCWP) as the proper anchoring structure to the rigid cell wall layer. Sequencing of 8,004 bp in the 5'-upstream region of the S-layer gene sbpA led to five ORFs-encoding proteins involved in cell wall metabolism. After cloning and heterologous expression of ORF1 and ORF5 in Escherichia coli, the recombinant autolysin rAbpA and the recombinant pyruvyl transferase rCsaB were isolated, purified, and correct folding was confirmed by circular dichroism. Although rAbpA encoded by ORF1 showed amidase activity, it could attack whole cells of Ly. sphaericus CCM 2177 only after complete extraction of the S-layer lattice. Despite the presence of three S-layer-homology motifs on the N-terminal part, rAbpA did not show detectable affinity to peptidoglycan-containing sacculi, nor to isolated SCWP. As the molecular mass of the autolysin lies above the molecular exclusion limit of the S-layer, AbpA is obviously trapped within the rigid cell wall layer by the isoporous protein lattice. Immunogold-labeling of ultrathin-sectioned whole cells of Ly. sphaericus CCM 2177 with a polyclonal rabbit antiserum raised against rCsaB encoded by ORF5, and cell fractionation experiments demonstrated that the pyruvyl transferase was located in the cytoplasm, but not associated with cell envelope components including the plasma membrane. In enzymatic assays, rCsaB clearly showed pyruvyl transferase activity. By using RT-PCR, specific transcripts for each ORF could be detected. Cotranscription could be confirmed for ORF2 and ORF3.

Analysis of the cell surface layer ultrastructure of the oral pathogen Tannerella forsythia.

The Gram-negative oral pathogen Tannerella forsythia is decorated with a 2D crystalline surface (S-) layer, with two different S-layer glycoprotein species being present. Prompted by the predicted virulence potential of the S-layer, this study focused on the analysis of the arrangement of the individual S-layer glycoproteins by a combination of microscopic, genetic, and biochemical analyses. The two S-layer genes are transcribed into mRNA and expressed into protein in equal amounts. The S-layer was investigated on intact bacterial cells by transmission electron microscopy, by immune fluorescence microscopy, and by atomic force microscopy. The analyses of wild-type cells revealed a distinct square S-layer lattice with an overall lattice constant of 10.1 ± 0.7 nm. In contrast, a blurred lattice with a lattice constant of 9.0 nm was found on S-layer single-mutant cells. This together with in vitro self-assembly studies using purified (glyco)protein species indicated their increased structural flexibility after self-assembly and/or impaired self-assembly capability. In conjunction with TEM analyses of thin-sectioned cells, this study demonstrates the unusual case that two S-layer glycoproteins are co-assembled into a single S-layer. Additionally, flagella and pilus-like structures were observed on T. forsythia cells, which might impact the pathogenicity of this bacterium.

Differential functions of ApoER2 and very low density lipoprotein receptor in Reelin signaling depend on differential sorting of the receptors.

ApoER2 and very low density lipoprotein (VLDL) receptor transmit the Reelin signal into target cells of the central nervous system. To a certain extent, both receptors can compensate for each other, and only the loss of both receptors results in the reeler phenotype, which is characterized by a gross defect in the architecture of laminated brain structures. Nevertheless, both receptors also have specific distinct functions, as corroborated by analyses of the subtle phenotypes displayed in mice lacking either ApoER2 or VLDL receptor. The differences in their function(s), however, have not been defined at the cellular level. Here, using a panel of chimeric receptors, we demonstrate that endocytosis of Reelin and the fate of the individual receptors upon stimulation are linked to their specific sorting to raft versus non-raft domains of the plasma membrane. VLDL receptor residing in the non-raft domain endocytoses and destines Reelin for degradation via the clathrin-coated pit/clathrin-coated vesicle/endosome pathway without being degraded to a significant extent. Binding of Reelin to ApoER2, a resident of rafts, leads to the production of specific receptor fragments with specific functions of their own and to degradation of ApoER2 via lysosomes. These features contribute to a receptor-specific fine tuning of the Reelin signal, leading to a novel model that emphasizes negative feedback loops specifically mediated by ApoER2 and VLDL receptor, respectively.

Sympathetic reserve, serum potassium, and orthostatic intolerance after endurance exercise and implications for neurocardiogenic syncope.

AIMS: To elucidate the mechanisms of orthostatic intolerance (OI) after endurance exercise which are incompletely understood. METHODS AND RESULTS: We investigated beat-to-beat haemodynamic and autonomic parameters in 51 male athletes during supine rest and after active standing the day before and 2 h after a marathon run. None of the subjects before the marathon [non-orthostatic intolerance (Non-OI)], but 14 after the marathon [orthostatic intolerance (OI)] exhibited with pre-syncope. There were no differences between OI and Non-OI before the marathon. After the marathon, only Non-OI was able to increase sympathetic modulation to resistance vessels from already increased basal levels in response to standing; OI could not. OI instead exhibited a decrease in total peripheral resistance and a paradoxical increase in parasympathetic sinus node modulation. We observed a significant correlation between serum potassium before the race and the maximally achieved sympathetic drive after the marathon (r = 0.55, P = 0.001). CONCLUSION: Post-exercise OI is associated with a 'high basal sympathetic modulation of vasomotor tone in combination with a diminished orthostatic sympathetic response' to resistance vessels. This situation may mimic the OI in some clinical conditions, which are also known to be associated with increased 'basal' sympathetic tone. The role of serum potassium deserves further study.

Structural characterization of the acid-degraded secondary cell wall polymer of Geobacillus stearothermophilus PV72/p2.

The secondary cell wall polymer (SCWP) from Geobacillus stearothermophilus PV72/p2, which is involved in the anchoring of the surface-layer protein to the bacterial cell wall layer, is composed of 2-amino-2-deoxy- and 2-acetamido-2-deoxy-D-glucose, 2-acetamido-2-deoxy-D-mannose, and 2-acetamido-2-deoxy-D-mannuronic acid. The primary structure of the acid-degraded polysaccharide--liberated by HF-treatment from the cell wall--was determined by high-field NMR spectroscopy and mass spectrometry using N-acetylated and hydrolyzed polysaccharide derivatives as well as Smith-degradation. The polysaccharide was shown to consist of a tetrasaccharide repeating unit containing a pyruvic acid acetal at a side-chain 2-acetamido-2-deoxy-alpha-D-mannopyranosyl residue. Substoichiometric substitutions of the repeating unit were observed concerning the degree of N-acetylation of glucosamine residues and the presence of side-chain linked 2-acetamido-2-deoxy-beta-D-glucopyranosyl units: [Formula: see text].

The proprotein convertase PCSK9 induces the degradation of low density lipoprotein receptor (LDLR) and its closest family members VLDLR and ApoER2.

The proprotein convertase PCSK9 gene is the third locus implicated in familial hypercholesterolemia, emphasizing its role in cardiovascular diseases. Loss of function mutations and gene disruption of PCSK9 resulted in a higher clearance of plasma low density lipoprotein cholesterol, likely due to a reduced degradation of the liver low density lipoprotein receptor (LDLR). In this study, we show that two of the closest family members to LDLR are also PCSK9 targets. These include the very low density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2) implicated in neuronal development and lipid metabolism. Our results show that wild type PCSK9 and more so its natural gain of function mutant D374Y can efficiently degrade the LDLR, VLDLR, and ApoER2 either following cellular co-expression or re-internalization of secreted human PCSK9. Such PCSK9-induced degradation does not require its catalytic activity. Membrane-bound PCSK9 chimeras enhanced the intracellular targeting of PCSK9 to late endosomes/lysosomes and resulted in a much more efficient degradation of the three receptors. We also demonstrate that the activity of PCSK9 and its binding affinity on VLDLR and ApoER2 does not depend on the presence of LDLR. Finally, in situ hybridization show close localization of PCSK9 mRNA expression to that of VLDLR in mouse postnatal day 1 cerebellum. Thus, this study demonstrates a more general effect of PCSK9 on the degradation of the LDLR family that emphasizes its major role in cholesterol and lipid homeostasis as well as brain development.

Reconstitution of the Reelin signaling pathway in fibroblasts demonstrates that Dab1 phosphorylation is independent of receptor localization in lipid rafts.

The Reelin signaling pathway operates in migrating neurons and is indispensable for their correct positioning during embryonic brain development. Many biochemical and cell biological studies to dissect the Reelin pathway at the molecular level are hampered by the lack of a cell line harboring a functional Reelin signaling pathway. Here we present fibroblast cell lines in which all required functional components of the pathway have been reconstituted. These cells react upon Reelin treatment in the same way as primary neurons. We have subsequently used these cell lines to study the subcellular localization of ApoER2 and the VLDL receptor and could demonstrate that receptor-mediated Dab1 phosphorylation does not depend on lipid rafts and that phosphorylated Dab1 remains bound to the receptor tail when the pathway is activated by Reelin.

Multi-site-frequency electromechanocardiography for the prediction of ejection fraction and stroke volume in heart failure.

BACKGROUND: Methods for stroke volume (SV) and ejection fraction (EF) measurements require the presence of qualified physicians and are not suited for continuous monitoring. AIM: To develop an automated non-invasive method for the measurement and continuous monitoring of SV and EF. METHODS: We have designed a method for the measurement of EF and SV using multiple-site-impedance (z0) measurements, applying multiple frequencies of 5, 40 and 200 kHz whereby various segments of the human body, including volume changes within these segments, could be defined electrically. The obtained variables were used to train neuronal nets and related by multiple regression analyses to cardiac output (CO) as measured by a partial rebreathing Fick method (CO(r-fick)) or EF as measured by echocardiography (EF(echo)), respectively. A total of 129 subjects (48 with normal heart function and 81 with CHF, NYHA I-IV) were investigated. RESULTS: The multiply derived values of z0 and of change of impedance (dz/dt) were shown, by multiple regression analysis, to be significantly related to CO(r-fick) and to EF(echo), (total r=0.77, n=35, p<0.001, and r=0.81, n=47, p<0.001, respectively.). By training a neuronal net with the electrical data of 67 (out of 94) subjects, EF(echo) could be predicted in the remaining 27 subjects which were unknown to the neuronal net with a combined r=0.71 (p<0.001,n=27). In contrast, conventional impedance cardiography (ICG) was unable to predict either CO(r-fick) or EF(echo). CONCLUSION: The new method, which we call multi-site-frequency electromechanocardiography (msf-ELMC) appears promising for the automated electrical measurement of the mechanical heart action in patients with normal and reduced cardiac function.

Hemodynamic and autonomic changes induced by Ironman: prediction of competition time by blood pressure variability.

We hypothesized that the extreme endurance exercise of an Ironman competition would lead to long-standing hemodynamic and autonomic changes. We investigated also the possibility of predicting competition performance from baseline hemodynamic and autonomic parameters. We have investigated 27 male athletes before competition, 1 h after, and then for the following week after the competition. The Task Force monitor was used to measure beat-to-beat hemodynamic and autonomic parameters during supine rest and active standing. Heart rate (P < 0.001) was increased, and stroke index (P = 0.011), systolic blood pressure (P = 0.004), diastolic blood pressure (P < 0.001), total peripheral resistance index (P < 0.001), and baroreceptor reflex sensitivity (P < 0.001) were decreased after the competition. The 0.05- to 0.17-Hz band of heart rate and blood pressure variability was increased (P < 0.001 and P < 0.001, respectively), the 0.17- to 0.40-Hz band of heart rate interval variability was decreased after the competition (P < 0.001). All parameters returned to baseline values 3 days after the competition. After the competition, the autonomic response to orthostasis was significantly impaired. The 0.05- to 0.17-Hz band of diastolic blood pressure variability before competition and weekly net exercise training, but not the other hemodynamic and autonomic parameters, were related to competition time in multivariate regression analysis (multiple r = 0.70, P < 0.001). The marked hemodynamic and autonomic changes after an ultraendurance race, which are compatible with myocardial depression in the face of sympathetic activation and reduction of afterload, return to baseline after only 1-3 days. Because the 0.05- to 0.17-Hz band of diastolic blood pressure variability contributes to the prediction of competition time, the analysis of blood pressure variability in the frequency domain deserves further study for the prediction of endurance capacity.

Receptor clustering is involved in Reelin signaling.

The Reelin signaling cascade plays a crucial role in the correct positioning of neurons during embryonic brain development. Reelin binding to apolipoprotein E receptor 2 (ApoER2) and very-low-density-lipoprotein receptor (VLDLR) leads to phosphorylation of disabled 1 (Dab1), an adaptor protein which associates with the intracellular domains of both receptors. Coreceptors for Reelin have been postulated to be necessary for Dab1 phosphorylation. We show that bivalent agents specifically binding to ApoER2 or VLDLR are sufficient to mimic the Reelin signal. These agents induce Dab1 phosphorylation, activate members of the Src family of nonreceptor tyrosine kinases, modulate protein kinase B/Akt phosphorylation, and increase long-term potentiation in hippocampal slices. Induced dimerization of Dab1 in HEK293 cells leads to its phosphorylation even in the absence of Reelin receptors. The mechanism for and the sites of these phosphorylations are identical to those effected by Reelin in primary neurons. These results suggest that binding of Reelin, which exists as a homodimer in vivo, to ApoER2 and VLDLR induces clustering of ApoER2 and VLDLR. As a consequence, Dab1 becomes dimerized or oligomerized on the cytosolic side of the plasma membrane, constituting the active substrate for the kinase; this process seems to be sufficient to transmit the signal and does not appear to require any coreceptor.