1. Tissue Engineering Research Center, Academy of Military Medical Sciences and Department of Neural Engineering and Biological Interdisciplinary Studies, Institute of Military Cognition and Brain Sciences, Academy of Military Medical Sciences, Beijing, China.
2. Rice University, Department of Bioengineering, Houston, Texas, USA.
3. Collage of Life Science and Technology, Huazhong Agricultural University, Wuhan, Hubei, China.
After myocardial infarction (MI), the scar tissue contributes to ventricular dysfunction by electrically uncoupling viable cardiomyocytes in the infarct region. Injection of a conductive hydrogel could not only provide mechanical support to the infarcted region, but also synchronize contraction and restore ventricular function by electrically connecting isolated cardiomyocytes to intact tissue.
Methods: We created a conductive hydrogel by introducing graphene oxide (GO) nanoparticles into oligo(poly(ethylene glycol) fumarate) (OPF) hydrogels. The hydrogels were characterized by AFM and electrochemistry workstation. A rat model of myocardial infarction was used to investigate the ability of OPF/GO to improve cardiac electrical propagation in the injured heart in vivo. Echocardiography (ECHO) was used to evaluate heart function 4 weeks after MI. Ca2+ imaging was used to visualize beating cardiomyocytes (CMs). Immunofluorescence staining was used to visualize the expression of cardiac-specific markers.
Results: OPF/GO hydrogels had semiconductive properties that were lacking in pure OPF. In addition, the incorporation of GO into OPF hydrogels could improve cell attachment in vitro. Injection of OPF/GO 4 weeks after myocardial infarction in rats enhanced the Ca2+ signal conduction of cardiomyocytes in the infarcted region in comparison with PBS or OPF alone. Moreover, the injection of OPF/GO hydrogel into the infarct region enhanced the generation of cytoskeletal structure and intercalated disc assembly. Echocardiography analysis showed improvement in load-dependent ejection fraction/fractional shortening of heart function 4 weeks after injection.
Conclusions: We prepared a conductive hydrogel (OPF/GO) that provide mechanical support and biological conduction in vitro and in vivo. We found that injected OPF/GO hydrogels can provide mechanical support and electric connection between healthy myocardium and the cardiomyocytes in the scar via activating the canonical Wnt signal pathway, thus upregulating the generation of Cx43 and gap junction associated proteins. Injection of OPF/GO hydrogel maintained better heart function after myocardial infarction than the injection of a nonconductive polymer.
Coronary heart disease is the leading cause of the rising incidence of heart failure worldwide, especially myocardial infarction (MI) [1-3]. The limited regenerative potential of the heart causes scar formation along the infarcted region following MI, leading to abnormal electric signal propagation and desynchronized cardiac contraction . Injectable hydrogels have demonstrated great potential for cardiac tissue repair and regeneration after MI. The injectable hydrogels can be injected alone or used as the delivery vehicle for therapeutic agents, including cells or growth factors, and can potentially provide mechanical and functional support to the infarcted region [5, 6].
Oligo(poly(ethylene glycol) fumarate) (OPF) is a macromer with alternating poly(ethylene glycol) (PEG) chains and fumarate groups, which allow for crosslinking and degradation through hydrolysis . OPF hydrogels have been employed for the controlled delivery of growth factors and encapsulated cells for a variety of tissue engineering applications , especially in the field of osteochondral tissue regeneration. Previous findings have demonstrated that OPF hydrogels support the proliferation of encapsulated articular chondrocytes . Additionally, previous in vivo investigations have shown the promise of OPF hydrogels as MSC delivery vehicles for osteochondral tissue regeneration . These reports suggested that OPF hydrogel could provide mechanical support to the injured tissue and can be used to support tissue repair after MI. Previously, we have used OPF as a carrier of mouse embryonic stem cells (mESCs) for the treatment of MI. Four weeks after transplantation, OPF hydrogel significantly reduced the infarct size and collagen deposition and improved the cardiac function . However, the implantation of OPF hydrogel alone, which is nonconductive, cannot restore impulse propagation to infarcted regions by electrically connecting isolated cardiomyocytes to intact tissue.
Graphene oxide (GO) is one of the widely investigated nanomaterials that has good electrical conductivity and appropriate mechanical properties. Previous studies have used GO as a scaffold material in tissue engineering applications, and have demonstrated the potential of GO to support cell adhesion, proliferation and differentiation [11-15]. For cardiac tissue engineering, previous studies have reported that GelMA, when blended with carbon-based nanoparticles such as multiwalled carbon nanotubes (CNTs) or GO, generates mechanically strong, electroconductive hydrogels that could support microvascular networks in vivo and support cardiomyocytes in a three-dimensional (3D) microenvironment [16-20].
In the present study, GO was incorporated into OPF hydrogels to generate an electrically conductive OPF/GO hydrogel. We found that electrically conductive OPF/GO composite hydrogels can enhance electric signal propagation in and around the infarcted region and improve cardiac repair when delivered to the heart tissue after MI.
All animal care and experimental protocols complied with the Animal Management Rule of the Ministry of Health, People's Republic of China (Documentation No. 55, 2001). All procedures were approved by the Institutional Animal Care and Use Committee of Academy of Military Medical Sciences, Beijing, China.
OPF and OPF/GO hydrogels were fabricated as described previously [9, 10]. Briefly, 0.16 g of OPF 10 K and 0.08 g of PEGDA were dissolved in 380 µL of phosphate buffered saline (PBS) or the GO dispersion (1.0 mg/mL) and incubated at 23 ℃ for 45 min to eliminate air bubbles. Thermal radical initiators, 74.9 µL of 300 mM N, N, N', N'-tetramethylethylenediamine (TEMED) solution (Sigma) and 74.9 µL of 300 mM ammonium persulfate (APS) solution (Sigma), were then added and incubated at 37 ℃ for 8 min for crosslinking. The electrical resistance of the hydrogel was detected by an electrochemistry workstation (CHI660D) (n=3/group).
Degradability was monitored as follows: OPF and OPF/GO hydrogels were prepared as described above [9, 10], except the polymer solutions were injected in Teflon molds (6 mm diameter by 0.5 mm thick) after mixing, and placed at 37 °C for 20 min afterwards. The resulting hydrogels were subject to degradation in PBS in 12-well tissue culture dishes at 37 °C on a shaker table shaking at 80 rpm. The PBS (2.5 mL/well) was completely changed every other day for the first week and the mass of the hydrogels was monitored every week. The mass remaining ratio of hydrogels (n=3) at each time point were represented as wd/wi × 100%, where wi and wd are the weight of the dried composite after fabrication prior to swelling, and the weight of the dried composite after swelling at each time point, respectively (n=3/group).
After euthanizing adult rats, rat leg skeletal muscle segments were explanted from the hind limbs. The muscles were cut in strips with a width of 10 mm and placed into a cell dish leaving a gap of 5 mm between two muscle pieces. This gap was filled with OPF or OPF/GO hydrogel, which acted as a bridge structure to connect two pieces of muscle tissue together. Voltage was then applied between two gel-connected muscle strips until contraction of both pieces could be observed to determine the voltage threshold (n=3).
Rat cardiac fibroblasts cells were isolated from 1-day-old neonatal Sprague-Dawley rats and seeded onto uncoated polystyrene dishes or those coated with OPF or OPF/GO at a concentration of 250 cells/mm2 to assess cell attachment [13, 30, 37]. After 48 h, cells were stained with live & dead kit to analyze the cytocompatibility of OPF/GO.
To do immunocytochemistry, cultured cardiac fibroblasts were fixed in 4% paraformaldehyde for 30 min at room temperature. After fixation, cells were treated with 0.3% Triton X-100 for 30 min. Cells were then stained with vimentin primary antibody (Abcam) at 1: 200 dilution in blocking buffer for 24 h at 4 ℃. Next, the sample was treated with secondary antibody (1: 200 dilution) in goat serum for 40 min. After washing with PBS, the sample was stained with DAPI (1: 200) diluted in PBS for 40 min at room temperature. The stained samples were then imaged with an inverted laser scanning confocal microscope (Leica SP5X MP, Germany). To assess the viability of cardiac fibroblasts on different materials, an MTT assay was performed according to the manufacturer's instructions. Briefly, a total of 1×104 cardiac fibroblasts were seeded on polystyrene, OPF or OPF/GO-coated 96 well surface, and grown for 48 h. Cardiac fibroblasts were incubated with MTT for 3 h at 37 °C, and cell viability was determined by measuring a test wavelength of OD 570 nm and a reference wavelength of 630 nm.
MI was induced in 27 male Sprague Dawley rats using an established model . Briefly, the rats were anesthetized by injecting with 2% pentobarbital sodium into cavum abdominis. A left-sided open thoracotomy was performed, the heart was exposed, and the proximal left anterior descending artery (LAD) was ligated with 6-0 polypropylene. The rats were randomized into 3 groups and received separate peri-infarct intramyocardial injections of PBS (100 µL; n=7), OPF hydrogel (100 µL; n=10) or OPF/GO hydrogel (100 µL; n=10). Then, the thoracotomy was closed in multiple layers.
Echocardiography (ECHO) was performed 4 weeks after hydrogel or PBS injection. Rats were anesthetized with 2% pentobarbital sodium. Standard transthoracic echocardiography was performed using the echocardiography system equipped with a 13 MHz linear ultrasonic transducer (15L8; Acuson Corporation, Mountain View, CA) in a phased array format. All analyses were performed by a single investigator who was blinded to the treatment groups (n=6/group).
After 24 h, 1, 2, 3, and 4 weeks of injections, rats in all surgical groups were anesthetized, and the heart was exposed and arrested by apical injection of 1 mL 15% KCl. The hearts were explanted and fixed in 2% paraformaldehyde overnight and then were paraffin-embedded. The embedded tissues were serially sectioned at 3 µm in the LV transverse direction. Masson's trichrome staining was performed to measure infarct size. Hematoxylin and eosin staining was performed to measure the maintenance and distribution of OPF or OPF/GO hydrogel in the infarct region. Immunohistochemical staining was performed with antibodies against CD68 (Abcam), F-actin (Abcam), N-cadherin (Abcam), plakoglobin (Abcam), desmoplakin, alpha-smooth muscle actin (α-SMA, Abcam), connexin 43 (Cx43, Abcam) and α-actinin (Abcam). Moreover, the number of CD68+ cells was counted from 6 randomly selected fields at the infarcted zone under a magnification of 60×. Nuclei were stained with 4', 6-diamidino-2-phenyindole (DAPI, Sigma). Morphometric analysis was performed to assess infarct and myocardial wall thickness. Infarct thickness was computed at each time point from the average spoke length. Infarct size was calculated as percentage of length of the infarcted endocardial circumference as previously described . Capillaries were recognized as tubular structures positively stained for α-SMA as described in the literature .
4 weeks after injections, total RNA of the infarcted region was extracted using Trizol (Invitrogen). Total RNA was treated with DNase I (Invitrogen, CA, USA) to eliminate the genomic DNA carry over. DNase-treated RNA was reverse transcribed using high capacity cDNA kit (Toyobo). Real-time quantitative PCR (qPCR) was performed with Fast SYBR Green Master Mix (Toyobo). The corresponding primers were presented in Table S1.
Western blot analysis was carried out using lysates from infarct heart tissues 4 weeks after MI. The infarcted tissue and protein extract (TIANDZ, Beijing, China) were homogenized on ice. The supernatant was collected after centrifugation at 12000×g for 10-15 min. The extracted proteins were determined by using BCA Protein Assay Kit (Thermo Scientific, USA). Equal amounts (50 µg) of denatured proteins were separated on 12% sodium dodecyl sulfate-polyacrylamide gels and transferred to nitrocellulose membrane. The membranes were blocked with 5% defatted milk for 1 h and were incubated with these primary antibodies overnight at 4 ℃. After being washed with Tris-base buffer, they were further incubated using appropriate secondary antibodies (goat anti-mouse or anti-rabbit IgG (Boster, Wuhan, China)) for 1 h at room temperature. The labeled proteins were visualized by enhanced chemiluminescent reagent (Applygen, Beijing, China). GAPDH was used as an internal control to correct the variations of different samples.
Adult CMs were isolated as described in literature , with minor modifications. Briefly, 4 weeks after injections, rats in all surgical groups were anesthetized. Hearts were removed and perfused via aortic cannulation with a constant flow of 3 mL/min in a Langendorff apparatus. Hearts were firstly perfused at 37 °C for 5 min with Tyrode's Solution containing 137 mM NaCl, 5.4 mM KCl, 0.5 mM MgCl2, 5.0 mM D-glucose, 11.6 mM HEPES, and 1.8 mM CaCl2 (pH 7.4). Then, the hearts were perfused for 3 min with the Tyrode's solution containing 130 mM NaCl, 5.4 mM KCl, 1.2 mM KH2PO4, 6.0 mM HEPES, and 10.0 mM D-glucose (pH 7.4), followed by digestion solution (0.8 mg/mL colleganse II) for 16 min. Hearts were then removed from the Langendorff apparatus while the tissues were gray and loosely connected. Desired areas (infarct zone) were then dissected in KB solution containing 50 mM L-glutamic acid, 30 mM KCl, 80 mM KOH, 30 mM KH2PO4, 20 mM taurine, 10 mM HEPES, 10 mM D-glucose, 3 mM MgSO4, 0.5 mM EGTA (pH 7.4), mechanically dissociated, triturated, and resuspended in Tyrode's solution, and calcium was gradually reintroduced through a series of washes. For calcium transient measurements, cells were used on the same day as isolation.
Calcium transients were assessed as previously described . Briefly, isolated cardiomyocytes were loaded with Fluo-4 for 30 min at room temperature before measurements. The loading solution contained a 1:10 mixture of 5 mM Fluo-4 AM in dry DMSO, which was diluted 100-fold into extracellular Tyrode's solution containing suspended myocytes. After 30 min, the loading solution was extracted and added to the fresh Tyrode's solution. Then Fluo-4 fluorescence transients were recorded via live cell station.
Statistical analyses were performed with Origin Pro 8.5 software. *p < 0.05 or **p < 0.01 was considered statistically significant.
To create an electrically conductive hydrogel, GO nanoparticles were incorporated into OPF hydrogel. A schematic of this study is shown in the graphical abstract. GO dispersion was introduced into OPF at 3 different concentrations (0.3 mg/mL, 0.6 mg/mL and 1.0 mg/mL GO/OPF) to form OPF/GO hydrogels (Figure 1A). To investigate the effects of GO nanoparticles on the local conductivity and morphology of the hybrid hydrogels, thin layers of OPF and OPF/GO hydrogels were coated on glass slides. AFM analysis was carried out to determine the morphology of the samples. As shown in Figure 1C, OPF sample showed a uniform topography, while the samples containing GO showed relatively rough surfaces, which was due to the presence of GO nanoparticles on the surfaces of the hydrogels. Additionally, the electrical conductivity of OPF/GO substrates was found to be significantly greater compared to that of OPF substrates (Figure 1D). The OPF/GO with the composite ratio of 1.0 mg/mL showed significantly higher conductivity (4.235×10-3 S/cm) than either 0.6 mg/mL OPF/GO, 0.3 mg/mL OPF/GO or pure OPF (0.905×10-3 S/cm).
To ascertain whether OPF/GO could support conduction between tissues more effectively than pure OPF ex vivo, we developed an assay for bioconductivity between isolated rat muscles. As shown in Figure 1E, pure OPF or OPF/GO hybrid hydrogels were injected into the gap between the tissues and allowed to crosslink. The hydrogels connected two pieces of muscle tissues together and acted as a bridge structure. One side of the muscle piece was then stimulated by applying an electrical voltage, while the beating or contraction of the other side muscle was checked and the applied voltage was increased until the muscles contracted. We observed a lower threshold for muscle tissue connected with OPF/GO (1.0 mg/mL) hybrid hydrogels (4.5±1.5 V, Movie S1) compared to the tissues linked with pure OPF hydrogel (Figure 1F, 10.5±1.5 V, Movie S2).
The prepared hydrogel degraded over time and the degradation ratio increased as culture progressed in all tested groups (Figure 1B). The mass remaining of OPF or OPF/GO was monitored every week until week 4. OPF had lost approximately 73% of its original weight whereas OPF/GO hydrogel with the composite ratio of 1.0 mg/mL lost only about 25% of its original weight, indicating degradation of both types of hydrogels and a higher degradation rate of OPF (n=3/group). Moreover, hydrogel degradation did not have significant effects on its electrophysiological properties (Figure S1).
Synthesis and characterization of OPF/GO conductive hydrogel. (A) Photographs of OPF and OPF/GO hydrogels with various concentrations. (B) Degradation behavior of OPF and OPF/GO hydrogels (n=3/group). (C) Spatial topography of OPF and OPF/GO hydrogels measured by AFM. (D) Conductivity of OPF and OPF/GO with various concentrations (n=3/group). (E) A schematic illustration of the ex vivo setup used for measuring excitation threshold of muscle tissue. One muscle was electrically stimulated while the beating or contraction of the other muscle was monitored. (F) Excitation threshold of muscle tissue connected by pure OPF and OPF/GO hybrid gels, demonstrating that OPF/GO hybrid gels gave rise to the lowest excitation threshold, which was due to the conductivity of the OPF/GO hybrid gels (± standard deviation, n=3/group, *p < 0.05, or **p < 0.01).
We investigated the biocompatibility of OPF/GO on cardiac fibroblasts, an important cell type in the heart after MI (Figure S2A). To demonstrate cell morphology and biomaterial attachment, rat fibroblasts were seeded on the surface of OPF- or OPF/GO hydrogel-coated wells and grown for 48 h (Figure 2A and Figure S2B). The effects of OPF/GO hydrogel on cardiac fibroblasts were investigated via live & dead staining. Compared with the cardiac fibroblasts seeded on OPF-coated surfaces, cells seeded on OPF/GO-coated surfaces maintained excellent viability and few dead cells (red) could be observed on all OPF/GO hydrogels, indicating that the introduction of GO did not induce cytotoxicity even at GO concentrations up to 1.0 mg/mL (Figure 2B). In addition, the introduction of GO nanoparticles improved the poor cell adhesion on the OPF surface, which correlated with increasing GO concentration (Figure 2C). A similar effect was observed for the MTT assay (Figure 2D): the introduction of GO in the OPF hydrogel greatly enhanced cell adhesion, and the adhesion effects were correlated with GO concentration. No significant differences were observed in terms of cytotoxicity between OPF with 0.6 mg/mL GO and 1.0 mg/mL GO. On the other hand, the results in Figure 1D suggested that the electrical conductivity of OPF/GO hydrogel with the composite ratio of 1.0 mg/mL was significantly higher than the rest. Therefore, OPF/GO with the composite ratio of 1.0 mg/mL was used in implantation research after MI.
MI was induced in a rat model by occlusion of the left anterior descending coronary artery according to established laboratory procedures. Subsequently, 100 μL of injectable hybrid hydrogel was injected at three different sites in the peri-infarct regions of the heart. The rat models were divided into three groups and received PBS, OPF and OPF/GO injections, respectively. The rats were sacrificed at different time points (24 h, 1, 2, and 4 weeks) after injection to observe the histological responses to scaffolds using light microscopy.
As shown in Figure S3A, OPF/GO was still visible in H&E-stained heart sections of OPF/GO-treated hearts. The implant experiments suggested that the OPF/GO hydrogels could provide mechanical support to the infarcted region and interact directly with the infarcted tissue. 24 h after injection post MI, OPF or OPF/GO hydrogels were observed in cross sections of left ventricular tissue, which indicates direct interaction with the infarcted tissue at the early stage of MI. Along with partial degradation of OPF hydrogel with time after implantation, the biomaterial was gradually replaced by connective tissue and myofibroblasts. At 2 weeks and 4 weeks, only rare, weak pink-stained isolated islands of biomaterial were detected in the infarct area after OPF injection. However, in the OPF/GO-injected group, OPF/GO hydrogel was still visible and distributed continuously along the infarcted region at 4 weeks post MI (Figure S3B).
Considering the role of OPF and OPF/GO hydrogels in promoting macrophage recruitment and activation, we analyzed the infiltration of macrophages in the infarcted hearts. CD68+ cells were evaluated by immunofluorescence staining in the infarcted area of myocardial sections at 1 week after MI. No significant differences were observed in macrophage infiltration between PBS, OPF and OPF/GO-injected infarcts (Figure 3A). Compared to the PBS group, the ratio of CD68+ cells in the OPF- and OPF/GO-injected hearts was slightly increased (Figure 3B). Moreover, the recruitment of CD68+ cells was not significantly enhanced in or around the OPF/GO hydrogels in the infarcted region (Figure 3C).
OPF/GO hydrogel supports cell growth without toxicity. (A) Representative morphological images of cardiac fibroblast cells cultured for 48 h on OPF- or OPF/GO-coated substrates. (B) Rat cardiac fibroblast cells were plated on the surfaces of wells coated with OPF or OPF/GO, grown for 48 h, and then stained with live & dead kit. (C) Immunofluorescence images of cardiac fibroblasts seeded on OPF- or OPF/GO-coated substrates (vimentin, red). (D) An MTT assay of cardiac fibroblasts cultured on the surface of pure OPF and different composite OPF/GO (0.3 mg/mL, 0.6 mg/mL, 1.0 mg/mL) (n=3, *P<0.05). MTT indicates 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazoliumbromide.
Effects of OPF and OPF/GO hydrogels on macrophage infiltration in infarcted hearts. (A) Representative fluorescence images show CD68-positive cells (left column, merged channel; middle column, CD68 (red); right column, DAPI (blue)) in PBS-, OPF-, and OPF/GO-injected hearts at 1 week post MI. (B) Quantification of CD68-positive cells of the corresponding images of (A). (C) CD68-positive signal is co-localized with the presence of OPF/GO hydrogel (CD68 (green), DAPI (blue)).
As shown in Figure 4A, in the OPF/GO- and OPF-treated group, a cross-striated pattern can be obviously observed in the F-actin-positive area at 4 weeks post MI. Whereas in the PBS injected group, the pattern of the F-actin-positive area was weak and ambiguous (Figure 4A-B). In order to confirm if the enhanced generation of cytoskeletal structures was induced by the mechanical support of OPF and OPF/GO, we further explored the mechanism of the enhanced cytoskeletal structures and examined intercalated disc (ID) formation.
ID is important for cardiac integrity and function . To determine whether the injection of OPF/GO had any influence on the assembly and formation of ID, ID-related proteins in the infarcted regions were characterized by immunofluorescence staining. Marker proteins including N-cadherin (NC) for adherens junctions, and plakoglobin (PG) and desmoplakin (DP) for desmosomes were examined 4 weeks post injection. As shown in Figure 5A, increases in localized structures were seen in the cell contact region, including the plaque pattern of N-C protein, stair-like distribution of PG protein (Figure 5B), and strand-like appearance of DP (Figure 5C). Meanwhile, NC, PG and DP showed larger linear regions in the OPF/GO groups compared to those in the OPF and control groups, indicating the formation of better developed mechanical and gap junctions. To further quantify the ID-associated proteins, we quantified the ID-associated proteins by western blotting. The results suggested that ID-related protein expression in the OPF/GO group had significantly higher levels relative to those in the groups injected with OPF and PBS after 4 weeks (Figure 5D).
Immunohistochemistry and histology of tissue sections revealed higher gap junction remodeling in the infarcted region of the OPF/GO-treated group than those in the OPF and PBS groups. Cx43 red fluorescence could be observed at the gap connexin region of the tissue by immunohistochemistry staining. As shown in Figure S4A, no significant differences were observed in the formation of gap junction-associated proteins between PBS-, OPF- and OPF/GO-injected infarcts 2 weeks after MI. The higher gap junction remodeling was observed obviously in the infarcted region of the OPF/GO hydrogel-treated group compared to those of the OPF- and PBS-treated groups at 3 weeks and 4 weeks (Figure S4B and Figure 6A). The expression levels of gap junction-associated markers Cx43, Cx40, Cx37 and Cx45 were analyzed by qPCR (Figure 6B). The results were consistent with those of the immunohistochemistry analysis, where the mRNA expression level of gap connexin-associated genes in OPF/GO hydrogel was higher than that of OPF and PBS injection at 4 weeks post injection.
To further clarify the possible mechanism of the enhanced generation of connexin-associated proteins, we quantified the downstream signaling molecules of canonical Wnt signaling pathway by western blot (Figure 6C). Our data suggests that the introduction of GO improved the expression of total Akt. Although there were no significant differences in the expression of total GSK-3β among all experimental groups, the introduction of GO could improve the phosphorylation levels of GSK-3β (pGSK-3β) at 14 days post MI. Similarly, pGSK-3β could then induce β-catenin signaling and subsequently cause an increase in Cx43 expression (Figure 6D).
OPF/GO enhanced cytoskeletal structure 4 weeks after MI. (A) F-actin (green) in the infarcted areas of PBS-, OPF- and OPF/GO-treated groups at 4 weeks post MI (DAPI (blue)). (B) α-Tubulin (green) in the border zone cardiomyocytes (marked by actinin staining in red, DAPI (blue)).
Immunofluorescence staining and quantification of ID-related proteins in the hearts of PBS-, OPF-, OPF/GO-injected groups at 4 weeks post MI. Marker proteins including N-cadherin (NC) for adherens junctions (A), and plakoglobin (PG) (B) and desmoplakin (DP) (C) for desmosomes. (D) Western blot revealed the OPF/GO-injected group had significantly higher expression levels of NC, PG, and DP compared to the PBS- and OPF-injected groups.
Masson's Trichrome assay showed that there was extensive fibrous tissue formation in the border area of the PBS group, whereas little fibrosis was observed in the OPF/GO group. The control group receiving PBS injection had collagen zones occupying a large portion of the LV wall of the infarct site (Figure 7A), indicating the presence of widely spread scar tissue. In contrast, the animals injected with OPF/GO had visually more myocardial tissue at the infarct site. Consistent with this observation, LV wall thickness was significantly preserved (from 0.37±0.096 mm to 0.77±0.079 mm) in the OPF/GO injected group (Figure 7B); the infarct size was drastically decreased (from 50.7% to 31.7%; Figure 7C). These results suggest that the structural basis of the infarcted heart was effectively restored after OPF/GO injection.
The MI-damaged left ventricle undergoes progressive remodeling, leading to the deterioration of cardiac function . Cardiac function was evaluated using echocardiography (ECHO) at 4 weeks post MI (Figure 7D). The results of echocardiography showed that LV ejection fraction (EF; Figure 7E) and fractional shortening (FS; Figure 7F), two important indicators for blood pumping function, were improved in both OPF and OPF/GO groups compared to PBS-injected rats. In particular, the hearts treated by OPF/GO show a significantly higher recovery of cardiac function than the OPF-treated group. Consistently, the OPF/GO group had a decrease in left ventricular end-diastolic dimension (LVEDD; Figure 7G) and left ventricular end-diastolic volume (LVESD; Figure 7H), indicating a functional improvement in ventricular filling. Together, these data proved that OPF/GO injection effectively promotes myocardial functional recovery after MI.
Angiogenesis restores blood supply to myocardium, facilitating cardiac function recovery . We next investigated the effects of OPF/GO injection on neovascularization after MI. The OPF/GO injection significantly increased the mRNA level of α-SMA (Figure S5A), vWF (Figure S5B) and VEGF (Figure S5C) at 4 weeks after LAD ligation, suggesting that OPF/GO injection may have an angiogenic effect. 4 weeks post injection, rats that received OPF/GO injections had a higher density of α-SMA-positive capillaries in the infarcted myocardium compared to both PBS- and OPF-treated rats (Figure S5D). The undegraded OPF/GO hydrogel appeared to be distributed in the anterior wall and cell ingrowth was found inside or around the hydrogel area, with some regions of this tissue structure staining positively for α-SMA in higher magnification images (Figure S5E). These data suggested angiogenesis can be an outcome of gap junction enhancement.
The formation of gap junction-associated proteins in the infarcted region. (A) Immunofluorescence staining revealed higher gap junction remodeling in the infarcted region of the OPF/GO hydrogel-treated group compared to the OPF- and PBS-treated groups. The highlighted areas are presented in the bottom images. (B) qPCR analysis of the expression levels of gap junction-associated markers Cx43, Cx40, Cx37 and Cx45. (C) Western blot revealed the downstream signaling molecules of canonical Wnt signaling pathway. (D) A schematic of the possible mechanisms.
Masson's trichrome staining and echocardiographic measurements represent the effects of OPF/GO hydrogel on the morphology and myocardial functional recovery of infarcted hearts. (A) Masson trichrome staining of PBS-, OPF- or OPF/GO-injected heart 4 weeks post injection. Four weeks after injection, hearts were excised and left vetricle wall thickness (B) and infarct size (C) were measured. (D) Systolic ventricular function was improved after OPF/GO hydrogel injection into the border zone. Echocardiography (ECHO) was performed at 4 weeks post injection. Quantification of the parameters reflecting blood pumping function, including ejection fraction (EF) (E) and fractional shortening (FS) (F) 6 weeks after MI. Quantification of the parameters reflecting ventricular filling function, including left ventricular end-diastolic dimension (LVEDD) (G) and left ventricular end-systolic dimension (LVESD) (H). Data is presented as mean ± standard deviation; *p < 0.05, or **p < 0.01 (n=6/group).
To further investigate whether OPF/GO or OPF alone can promote electric conduction between cardiomyocytes and host tissue in the infarcted region, we isolated cardiomyocytes from the left ventricle of OPF/GO-, OPF- or PBS-injected rats 4 weeks after MI and labeled them with the Fluo-4 calcium dye. The Ca2+ fluctuations of cardiomyocytes can be observed (Figure 8A-C). In the OPF/GO-treated group, the isolated cardiomyocytes displayed rhythmic calcium transients and exhibited markedly high amplitudes of calcium transients (Movie S3 and Figure 8A). While the cardiomyocytes isolated from the left ventricle of OPF-treated (Movie S4 and Figure 8B) and PBS-treated (Movie S5 and Figure 8C) rat MI models exhibited a uniform beating frequency, the corresponding calcium transients amplitudes were lower than that of the OPF/GO-treated group. The results suggested that OPF/GO mediated the electrical conduction between cardiomyocytes and host tissue, thus promoting the intrinsic Ca2+ propagation of cardiomyocytes in the infarcted region.
During the past 5 years, investigations into intramyocardial injection of bioengineered, chemically modified materials after MI have shown promising results . For cardiac tissue engineering, electrical stimulation is important in assisting in the development of the tissue construct [2, 24]. Although traditional injectable hydrogels benefit cardiac function by providing structural support to the injured LV and enhancing myocardial regeneration, the contraction of the heart depends on the conduction of electric impulses through cardiac tissue, and conduction velocity is reduced in the infarct region following an MI [25, 26]. A conductive injectable hydrogel would potentially be able to synchronize ventricular contraction and substantially improve heart function post MI.
Ca2+ transients evaluation at 4 week after MI. Spontaneous Ca2+ transients of cardiomyocytes isolated from the left ventricle of OPF/GO- (A), OPF- (B) or PBS- (C) injected rats four weeks after MI.
Conductive hydrogels, such as OPF/GO composite hydrogel, are biocompatible and have semiconductive properties that are capable of transmitting electric currents [27, 28]. The introduction of GO nanoparticles improved the poor cell adhesion of OPF, and is therefore suitable for tissue engineering applications. Our results demonstrated that OPF/GO injection after MI improved cardiac function in comparison with OPF- or PBS-treated groups. Although the mechanism of how OPF/GO improves cardiac function has not been fully investigated, the electric conductive property of GO is believed to contribute to the beneficial effects. GO is a single layer of sp2-hybridized carbon atoms tightly packed into a two-dimensional (2D) honeycomb lattice, and has emerged as a new class of nanomaterial for its mechanical strength and conductive properties . Previous studies have confirmed GelMA/GO hydrogel can be microengineered to support microvascular networks in vivo and support cardiomyocytes in a three-dimensional (3D) microenvironment due to its strong mechanical and electroconductive properties . In this study, we explored the therapeutic efficacy of OPF/GO hydrogel for tissue repair following MI. Calcium transients evaluation showed that the cardiomyocytes in the left ventricle in the OPF/GO-injected group displayed rhythmic calcium transients and exhibited markedly high amplitudes of Ca2+. The results suggested that OPF/GO hydrogel could not only provide mechanical support to the infarcted region, but also enhance electrophysiological properties by electrically connecting isolated cardiomyocytes to intact tissue. The precise mechanisms for the observed effects of OPF/GO hydrogels in this study are not fully understood and deserve further investigations. There could be several reasons that account for the improvement of cardiac function by the OPF/GO injection post MI. Our H&E and immunofluorescence images suggest that GO developed close contacts with the cardiac tissue in or around the infarcted region, and one mechanism of GO/myocyte interaction may rely on the detection of irregular tight contacts between GO and membranes. Such contacts are similar to that seen in cardiomyocytes cultured on carbon nanotubes, which promote growth and trigger spontaneous electrical activity [17, 31]. Furthermore, the electrically conductive networks formed by GO within OPF hydrogel are the key characteristics of GO/OPF that lead to improved cardiac cell adhesion, organization, and cell-cell interactions. GO, bridging the OPF hydrogel, provided additional pathways for direct electrical current flow and action potential propagation. A similar mechanism explained the electric signal propagation between cardiomyocytes through CNT networks .
Overall, graphene-based nanomaterials hold great promise for applications in cardiac tissue engineering. Most synthetic hydrogels used for tissue engineering are mechanically weak and lack conductive properties [33-36]. The addition of graphene-based nanoparticles imparted the required conductivity and surface properties to these hydrogel materials in several ways and further improved the microenvironment of the infarcted region [13-15, 32]. Therefore, these materials could be used in cardiac tissue engineering and injectable hydrogel-based therapeutic strategies.
APS: ammonium persulfate; CMs: cardiomyocytes; CNTs: carbon nanotubes; Cx43: Connexin 43; DP: desmoplakin; ECHO: echocardiography; EF: ejection fraction; FS: fractional shortening; GO: graphene oxide; ID: intercalated disc; LAD: left anterior descending artery; LV: left ventricular; LVEDD: left ventricular end-diastolic dimension; LVESD: left ventricular end-diastolic volume; mESCs: mouse embryonic stem cells; MI: myocardial infarction; MTT: 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazoliumbromide; NC: N-cadherin; OPF: oligo(poly(ethylene glycol) fumarate); PBS: phosphate buffered saline; PEG: poly(ethylene glycol); PG: plakoglobin; TEMED: N, N, N', N'-tetramethylethylene- -diamine.
The authors thank Antonios G. Mikos (Rice University) for providing OPF raw materials. This work was supported by Projects of International Cooperation and Exchanges NSFC (No. 31320103914), National Natural Science Funds for Outstanding Young Scholar (No. 81622027), the Key Program of the National Key Research and Development Program of China (No. 2016YFY1101303, No. 2017YFA0106100), National Natural Science Foundation of China (No. 31370987, 31470940, 81371696), Joint Funds for National Natural Science Foundation of China (No. U1601221), Beijing NOVA Program of China (No. 2016B615).


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redpajama_set_name: RedPajamaGithub


Unfortunately, many of the protections in place for "unaccompanied alien children" (UAC) are at risk of being rescinded by new administrative policies and proposed legislation. Both Executive Order (EO) 13767 on border security and EO 13768 on interior immigration enforcement negatively impact UAC and their families. In addition, bills such as the "Asylum Reform and Border Protection Act of 2017" (H.R. 391) and the "Protection of Children Act of 2017" (H.R. 495) include provisions stripping away essential protections for UAC.
Why should we oppose these rollbacks in protections?
They are against basic child protection principles. Unaccompanied children should be viewed as children first. One of the foundations of the U.S. domestic child welfare system is the best interest of the child principle. The term "best interest of the child" generally refers to the deliberation that courts undertake when deciding what type of services, actions, and orders will best serve a child as well as who is best suited to take care of a child. Our government should operate a UAC system that more fully takes best interest of children into account. Proposals that make it more difficult for children to seek relief, strip them of necessary protections, and target their families for removal and criminal prosecution upend child welfare and do not have best interest principles as their basis.
We have a moral obligation to ensure that UAC receive proper care, are not separated from their families if reunified, and have an opportunity to seek protection. While the Catholic Church recognizes governments' right to control their borders and enforce immigration laws, we hold a strong and pervasive pastoral interest in the welfare and humane treatment of unaccompanied children. The majority of children arriving at our borders present international protection concerns. And, as Pope Francis has said: "Among migrants, children constitute the most vulnerable group, because as they face the life ahead of them, they are invisible and voiceless." Our faith calls on us to oppose proposals which would lead to the inhumane and unjust treatment of these children. It further requires us to firmly reject proposals which would lead to family separation. We have seen firsthand in our churches and parishes the devastation caused by family separation and the family breakdown that often results from such separation.
Releasing children to family or sponsors pending their removal proceedings is a fiscally sound policy. In addition to the moral and humanitarian concerns, detention of children is incredibly costly and an unnecessary use of government resources. The Department of Homeland Security (DHS) spends an estimated $161 per person per day for those in family detention; in contrast, it costs a mere $14 a day to provide a UAC released to a sponsor with follow-up services. Similarly, keeping children in the care of the Department of Health and Human Services is costly and would often be an unnecessary use of limited federal funds. Many UAC present no safety or flight risks relating to their release, and reunification with family is frequently the best option from both a child welfare and a fiscal policy perspective.
How can we support UAC and voice our opposition to these proposals?
Contacting your Lawmakers. Contact your Members of Congress to express your support for unaccompanied children, urge them to support legislation to nullify the recent Executive Orders negatively impacting UAC, and ask them to reject bills which would roll back protections for these children. You can do this by participating in the Justice for Immigrants' Action Alerts.
Providing Transportation to Immigration Court. It is crucial that unaccompanied children who are involved in court proceedings consistently attend any legal hearings for which they are scheduled to appear. Failure to appear at scheduled hearings will likely lead the judge to issue a deportation order in absentia and make them eligible for deportation. To learn more, contact jfi@usccb.org.
Helping Families Enroll UAC in School. For immigrant families, a challenge in the U.S. can be enrolling their children into the local public school system. They may have trouble gathering the requested documentation, may be discouraged from enrolling due to language barriers or their child's age, or may be denied enrollment if they are a sponsor who is not the child's parent or legal guardian. Reach out to local schools to see what resources they offer for immigrant families. Plan an open house event where parents, children, teachers, and the administration can meet and discuss options as community members. Offer to accompany sponsors of UAC to schools to assist with enrollment.
Fostering an Unaccompanied Child. Become a foster parent to an unaccompanied child! USCCB/MRS currently provides foster care to unaccompanied children in the following locations: Texas, Michigan, Arizona, Virginia, New York, California, Washington, Mississippi, Florida and Utah. Those interested in becoming a foster parent must meet state and agency guidelines for training and licensing. If you live in one of the aforementioned states, and are interested in becoming a foster parent, contact migratingchildren@usccb.org for more information. If you reside in Colorado Springs, CO, Denver, CO, Fargo, ND, Fullerton, CA, Lansing, MI, Newton, PA, Philadelphia, PA, Seattle, WA, Washington, DC, Worcester, MA please contact Lutheran Immigrant and Refugee Services at fosterparentinfo@lirs.org.
If there are no USCCB or LIRS programs in your area, please direct your inquiries to DCS_ProjectOfficers@acf.hhs.gov.
 Determining the Best Interest of the Child, Child Welfare Information Gateway (March 2016), https://www.childwelfare.gov/pubPDFs/best_interest.pdf.
 The United Nations High Commissioner for Refugees (UNHCR), Children on the Run: Unaccompanied Children Leaving Central America and Mexico and the Need for International Protection 6 (UNHCR, ed. 2014).
 Pope Francis, 2017 World Day of Migrants and Refugees Message (September 8, 2016), https://w2.vatican.va/content/francesco/en/messages/migration/documents/papa-francesco_20160908_world-migrants-day-2017.html.
 American Academy of Pediatrics, Letter to Secretary Johnson (July 24, 2015), available at http://endfamilydetention.com/wp-content/uploads/2015/07/AAP-Letter-to-Secretary-Johnson-Family-Detention-Final.pdf (finding that detention "is associated with poorer health outcomes, higher rates of psychological distress, and suicidality making the situation for already vulnerable women and children even worse. For children, exposure to early adverse experiences, often referred to as toxic stress, has long-term consequences.").
 Justice for Immigrants, Action Alerts, https://justiceforimmigrants.org/category/action-alerts/ (last visited March 10, 2017).


Depending on the power they can supply, they incorporate a series of power sockets to be able to connect one or more vibrating needles. They can be connected to single-phase or three-phase networks and with input frequencies of 50 or 60 Hz. de 50 o 60 Hz.


군산출장안마 -24시출장샵 クM4군산출장안마u1군산출장안마Sk군산콜걸만남2Z군산마사지Mx군산콜걸λエζ군산마사지 군산출장마사지 Wozencraft, W.C. (2005). 창원릉콜걸샵 . Wilson, D.E.; Reeder, D.M. 함평모텔 찌라시 (영어) 3판. 존스 홉킨스 대학교 출판사. ISBN 영월출장최고시 . 동두천모텔 다방 가격 완진출장안마 .
연천출장안마 Lindblad-Toh, Kerstin; Wade, Claire M; Mikkelsen, Tarjei S.; Karlsson, Elinor K.; Jaffe, David B.; Kamal, Michael; Clamp, Michele; Chang, Jean L.; Kulbokas, Edward J.; Zody, Michael C.; Mauceli, Evan; Xie, Xiaohui; Breen, Matthew; Wayne, Robert K.; Ostrander, Elaine A.; Ponting, Chris P.; Galibert, Francis; Smith, Douglas R.; Dejong, Pieter J.; Kirkness, Ewen; Alvarez, Pablo; Biagi, Tara; Brockman, William; Butler, Jonathan; Chin, Chee-Wye; Cook, April; Cuff, James; Daly, Mark J.; Decaprio, David; 외. (2005). "Genome sequence, comparative analysis and haplotype structure of the domestic dog". 《Nature》 438 (7069): 803 in 803–19. PMID 무안군산 터미널 모텔 . doi : 10.1038/nature04338 .
↑ Nyakatura, K.; 외. (2012). "Updating the evolutionary history of Carnivora (Mammalia): a new species-level supertree complete with divergence time estimates" . 《BMC Biology》 10 (12). doi : 김제서울 조건 카톡 .
↑ Koepfli, Klaus-Peter; Pollinger, John; Godinho, Raquel; Robinson, Jacqueline; Lea, Amanda; Hendricks, Sarah; Schweizer, Rena M.; Thalmann, Olaf; Silva, Pedro; Fan, Zhenxin; Yurchenko, Andrey A.; Dobrynin, Pavel; Makunin, Alexey; Cahill, James A.; Shapiro, Beth; Álvares, Francisco; Brito, José C.; Geffen, Eli; Leonard, Jennifer A.; Helgen, Kristofer M.; Johnson, Warren E.; o'Brien, Stephen J.; Van Valkenburgh, Blaire; Wayne, Robert K. (2015). "Genome-wide Evidence Reveals that African and Eurasian Golden Jackals Are Distinct Species". 《Current Biology》 25 (16): 2158–65. PMID 26234211 . doi : 울진출장안마 .
모든 문서는 부안부산 해운대 출장 에 따라 사용할 수 있으며, 추가적인 조건이 적용될 수 있습니다. 자세한 내용은 봉화삼산동 출장 을 참고하십시오.
Wikipedia®는 미국 및 다른 국가에 등록되어 있는 상주광주 대딸방 소유의 등록 상표입니다.


text:
Safety Profile of Ustekinumab Favorable in IBD, Psoriatic Arthritis, Psoriasis
Kenny Walter
Conference | American College of Gastroenterology
The rates of adverse events was similar across all 4 indications in comparison to the placebo group.
While ustekinumab is prescribed across different indications, including Crohn's disease, ulcerative colitis, psoriatic arthritis, and psoriasis, new research presented at the 2021 American College of Gastroenterology (ACG) Annual Meeting shows the treatment is safe in all 4 indications.
A team, led by Millie D. Long, MD, MPH, FACG, University of North Carolina, presented an integrated analysis of pooled safety data of phase 2 or 3 long-term safety data of inflammatory bowel disease (IBD) studies through up to 5 years in Crohn's disease and 2 years in ulcerative colitis and psoriatic disease studies through up to 5 years in psoriasis and 1 year in psoriatic arthritis.
Four Indications
There are a number of US Food and Drug Administration (FDA) approved indications for ustekinumab, including psoriasis (2009), psoriatic arthritis (2013), Crohn's disease (2016), and ulcerative colitis (2019).
During this time there has been a number of safety studies showing the treatment is in fact safe, including an integrated safety data analysis through year 1 and the 2020 IBD pooled Safety Dataset.
Overall, the investigators examined data from 13 studies for the 4 approved indications, including studies involving concomitant immunomodulators and corticosteroids for IBD and psoriatic arthritis, but not psoriasis patients. Every patient received at least 1 dose of ustekinumab.
Safety outcomes were classified as events per 100 patient-years of follow-up. Also, the placebo-controlled period was 12 weeks in the majority of psoriasis studies and 16 weeks in most of the psoriatic arthritis.
Ustekinumab Safety
Overall, there were 6710 patients treated with ustekinumab, accounting for 13,807 follow-up years, compared to 2501 patients treated with placebo accounting 1244 of person-years of follow-up.
The rates of events per 100 person-years were similar between both ustekinumab treated patients and placebo groups, with the most frequent adverse events being nasopharyngitis (20.83 vs 20.87), upper respiratory tract infection (13.51 vs 14.53), arthralgia (13.99 vs 7.01), abdominal pain (10.77 vs 4.01), and diarrhea (6.67 vs 3.88).
The most serious infections reported in the placebo group compared to the ustekinumab group were anal abscess (0.72 vs 0.17), pneumonia (0.32 vs 0.14), and cellulitis (0.16 vs 0.11). However, events of malignancy and death were not frequently report, especially in the placebo group.
"The safety profile of ustekinumab across pooled indications was favorable through up to 5 years, confirming the well-established safety profile of ustekinumab in psoriatic and IBD indications," the authors wrote.
The study, "Safety of Ustekinumab Across Approved Adult Indications: Pooled Safety Analysis in Crohn's Disease, Ulcerative Colitis, Psoriasis, and Psoriatic Arthritis Through up to 5 Years," was published online by ACG.
American College of GastroenterologyConferenceGastroenterologyUlcerative ColitisFamily MedicineInternal MedicinePsoriatic ArthritisPlaque PsoriasisRheumatologyDermatology

redpajama_set_name: RedPajamaCommonCrawl


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Discussion in 'Misc. Oakley Exchange' started by Anonymous, 8/2/11.
At least that's what I call them, but I don't know what they are really called, so if anyone does, please let me know.
They are 6 sided and roughly 6 inches cubed. Pretty cool, but I really don't have room for them.
nice score . . .
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text:
Michael Rapino is a Canadian-American business executive and the Chief Executive Officer and President of Live Nation Entertainment, Inc, which was formed in 2010 following the merger of Live Nation and Ticketmaster.

In the late 1990s, Rapino co-founded Core Audience Entertainment, a concert promoting company, and later held several roles at Clear Channel Entertainment. He was named CEO of Live Nation in 2005.

Early life and education
Michael Rapino was born in Thunder Bay, Ontario, Canada, and earned a bachelor of administration degree from Lakehead University in 1989. While attending Lakehead, Rapino promoted bands at local bars and signed his first artist, Jeff Healey.

Career
While at Lakehead, Rapino landed a job as a beer rep for Labatt Breweries and started promoting bands at local bars, arranging a weekly ad in The Argus (Thunder Bay) called "The Blue Zone" that featured live music events happening in Thunder Bay that upcoming weekend. After graduation, he moved to Toronto and continued his career as a sales representative for Labatt. He remained at the company for 10 years in different marketing and entertainment roles. In the late 1990s Rapino founded Core Audience Entertainment, a concert promotion company that operated in Canada. Core Audience Entertainment was acquired by SFX Entertainment in 1999, which was in turn acquired by Clear Channel Communications in 2000. Rapino held several roles at Clear Channel Entertainment. He was later named CEO of Live Nation when it was spun off from Clear Channel in 2005.

Live Nation
As the CEO of Live Nation, Rapino initially focused on expanding the business outside of tour management, including artists' recording and marketing. Between 2007 and 2010, Rapino announced deals with artists such as Madonna, U2, and Jay-Z. In 2016, Rapino announced a deal between Lady Gaga and Live Nation's management division.

In 2010, Live Nation merged with Ticketmaster, a live entertainment ticket seller. Rapino was named CEO of the new parent company, Live Nation Entertainment. Prior to the merger's approval by the U.S. Justice Department, Rapino and then-Ticketmaster CEO Irving Azoff testified in front of a Senate antitrust panel. They argued that the merger was necessary to reduce inefficiencies and help the music industry, which was struggling financially at the time.

Rapino has led the company in acquiring several festivals and companies, including House of Blues Entertainment in 2006, Bonnaroo Festival in 2015, and Blue Note Entertainment in 2017. Rapino earned $70.6 million in 2017 and was reported to have one of the highest discrepancies in salary between CEO and his employees who earned an average of $24,000. Also in 2017, he extended his contract with Live Nation through 2022. Rapino was appointed to the SiriusXM Radio board of directors in January 2018. Also in 2018, he was listed as #1 on the Billboard Power 100 list of influential people in the music industry.

Rapino began producing films in 2015. In 2018, he served as an executive producer on the HBO documentary "Believer,"  and "A Star is Born".

Personal life
Rapino met his wife, American model and actress Jolene Blalock, when he cast her in a commercial he was shooting. Rapino and Blalock married in Negril, Jamaica, on April 22, 2003 and have three sons. Rapino and Blalock founded the Rapino Foundation, an organization that assists populations in the developing world.

Honours
In 2015, Rapino was given an honorary doctorate of commerce from Lakehead University.
He also received the Clara Lionel Foundation Diamond Honors Award for his commitment to philanthropy.

References

Living people
American business executives
Canadian business executives
Lakehead University alumni

Year of birth missing (living people)

redpajama_set_name: RedPajamaWikipedia


Following on from his previous release 'Join Us' on Miami based House label Deeper Heaven (Heavenly Bodies Recordings) P.Nichol brings us another fresh piece of dancefloor EDM.
Verging on the Progressive Trance side of things 'Waterworld' is a melodic track featuring a beautiful piano part and laced with synths & FX that will fit in any Trance & Electro House sets.
Juno has it for 1 week exclusive before all other stores.
MP3:192/320 & WAV all same price.
P.Nichol - Waterworld released today!
Should start appearing on all other stores later today.


text:
Panasonic completes Sanyo acquisition
By Associated Press |
TOKYO — Panasonic said Monday that it had taken majority control of Sanyo Electric, officially forming one of the world's largest electronics makers.
Panasonic said it had converted preferred shares to give it a 50.27 percent stake in Sanyo. It acquired the shares in a previously announced $4.6 billion tender offer that ended Dec. 9.
The acquisition gives Panasonic access to Sanyo's advanced solar panel and rechargeable battery technology. But Panasonic must also find a way to bring profitability back to Sanyo, which had a net loss of 30.6 billion yen ($338 million) in the April-September half.
Panasonic has said it will retain the well-known Sanyo brand and keep its shares listed on the Tokyo Stock Exchange.
Panasonic had reportedly hoped to take a larger stake in Sanyo, but the company had to settle with a minimum controlling stake, with some shareholders likely unwilling to sell at a tender offer that was lower than the market price.
The deal was delayed for months while the companies awaited clearance from worldwide anti-monopoly authorities.
WASHINGTON (AP) — President Donald Trump on Thursday chided his supporters who chanted "send her back" when he questioned the loyalty of a Somali-born congresswoman, joining widespread criticism of the campaign crowd's cry and his Republican allies worried about political blowback from the angry scene.
A Sacramento woman was convicted of pimping in a plea deal.
Lack of resolution in nearly half of cases, both locally and nationally, led to closer look at why.


The closed loop system of drying, unique to Whirlpool Dishwashers facilitates steam absorption inside the cavity which turns into water and is drained away. This avoids damage to your kitchen furniture as there is no humidity emission.
*Result based on tests comparing the performance of the 1 hour program of the A+++ comparable models. Models in A++ or less may have different results.
The revolutionary PowerClean Pro Technology uses high-pressure spray nozzles that ensures outstanding cleaning results without the need of scrubbing or hand-prewashing.
Designed to ensure maximum coverage & perfect performance, these powerful water jets remove the toughest stains & oil from the utensils resulting in supreme cleaning results.
The intelligent 6TH SENSE® Technology in Whirlpool Dishwashers intuitively detects the level of soil present on the dishes and optimizes the cleaning resources accordingly. This delivers outstanding cleaning results with up to 50% savings* of time & water.
Thanks to the innovative power clean loading system with 12 foldable racks, Whirlpool Dishwashers provide you with up to 30%* extra space & various loading options to clean all your utensils at once.
*Percentage calculated vs. the loading space on models of the same Whirlpool range without power clean solution.
Thanks to the innovative power clean loading system with 4 foldable racks, Whirlpool Dishwashers provide you with up to 30% extra space & various loading options to clean all your utensils at once.
Whirlpool dishwashers are amongst the silent-most dishwashers presently available in the market. With the noise level as low as 43db*, there is absolutely no noise disturbance, even during the night.
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Whirlpool Dishwashers come with flexible loading solutions of up to 14 place settings. This gives you great usage flexibility, customized loading and a modular third basket, so that even the taller & wider items can be placed in the upper rack.
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Jon Bjornson is an artist, graphic designer and visual thinker living and working in Philadelphia. His design work is about balancing opportunity with constraint, strategy with visual expression. By thinking deeply about how design can leverage a client's brand value, he translates that thinking into creative action. His philosophy is to work with people, not for businesses, and he truly cares about the people who trust him to visually and verbally express who they are.
Jon's recent artwork combines simple geometry and ambiguous spatial relationships with his artistic sense of color and composition; he also creates colorful and dramatic montages that incorporate photography from his various travels.
So Bjornson, what do you do? Mostly, I tell people I'm a graphic designer; my wife refers to me as an artist. To potential clients, I say I'm a brand consultant. Frankly, I like the universality of the term "creative professional." I'm also a dad, husband, and ice hockey nut.
Where do you work? I work in paradise—a 600-square-foot studio in my backyard that I collaboratively designed with an architect friend of mine. Short commute, flexible hours, great view.
Nice. How'd you swing that? With help from family and friends, I decided to simplify life so I could be around to watch my kids grow up. With that in mind, I built the studio in 2010.
What was so complicated? I had run a small firm in Center City Philadelphia for 15 years that employed as many as 10 people; the business was very successful, but the 80-hour weeks were killing me. In 2001, following the dot-com-bubble, I scaled back my business and moved my office out of Center City. In 2005, through another series of cosmic events, I was offered a job with one of my clients and went "in-house" to rebrand the company from the inside. It was an invaluable experience, and I am proud of what I accomplished, but it was pretty stressful. When I finished what I had started, I went back out on my own in 2008. I have been a white-collar fugitive ever since.


text:
Improving health and social care statistics: lessons learned from the COVID-19 pandemic
Transparent and trustworthy
Responsive and proactive
Clear and insightful
The pandemic reinforced the need for statistics to inform society about public health and provide an understanding of how public health programmes are working.
Statistics producers should continue to develop outputs which go beyond operational data in order to support policy evaluation and a better understanding of public health.
Official statistics on health and social care are often a by-product of the delivery of services. This means that they often focus on operational delivery, such as waiting times or hospital activity, rather than emerging population health issues and how they relate to other societal and economic issues. The pandemic demonstrated that good quality and timely data on operational delivery are essential. They helped governments and the public understand the ability of hospitals and care homes to cope with the increased pressures they were facing. It is also important that the public understand how government money is spent on delivering health services. But the pandemic showed that as a society we need to have a good understanding of population health issues – for example, in the case of the pandemic, to answer questions about the spread of coronavirus infections or the long-term impact of coronavirus infection on an individual's health and wellbeing. It is important that producers continue to collect, publish, and develop information which goes beyond operational data and supports a broad understanding of public health. This may be as part of public health surveillance carried out by public health bodies, or as producers have done through the Office for National Statistics (ONS) COVID-19 Infection Survey, or Public Health England's analyses of vaccine effectiveness. These data contribute to the evidence base required to understand public health, which includes academic research and other types of government analysis, such as social research.
The pandemic highlighted the importance of evaluating public health programmes. When we reviewed NHS England Test and Trace statistics in July 2020, we were clear that there were key questions which the publication was not able to answer, such as what proportion of people asked go on to complete self-isolation. We asked the UK Government to find ways to better understand the programme and its impact on the pandemic. We noted that this was unlikely to be answered solely through management information from the Test and Trace programme. We saw many significant improvements in Test and Trace statistics over the following year, as well as the publication of some information about the effectiveness of the programme based on modelling. We welcome the Department of Health and Social Care's recent publication on the impact of NHS Test and Trace on transmission. However, many important questions about the programme have remained unanswered – both as the UK went into the winter 2020 wave of the pandemic, and when we carried out a second review of the statistics prior to changes to self-isolation rules in England in August 2021. These include questions about the vaccination status of those who test positive, and how many close contacts identified through the tracing system go on to test positive. In future, governments must ensure the early collection and publication of measures which allow public health programmes to be scrutinised and evaluated.
Governments must evaluate public health programmes and be transparent in sharing information about them with the public.
ONS should consider how the COVID-19 Infection Survey can be adapted to play a role in understanding public health in future. This should be done in collaboration with other producers, including those in devolved administrations, and should consider the sustainability of representative population sampling.
For the statistical system to be able to respond quickly to future health and social care issues, horizon scanning and sharing plans with each other should be focus areas for collaboration between producers. It is also important that analytical resource and data infrastructures are flexible enough to adapt to new demands.
The pandemic exposed gaps in available data.
To ensure that statistics best serve the public good, these gaps must now be filled. Statistics producers should be proactive in meeting user needs to minimise gaps in future.
The pandemic exposed and reinforced existing data gaps, for example in social care, ethnicity, and mental health. This made it challenging for producers to meet user needs. For example, prior to the pandemic there was not a clear understanding of the number of people and the personal characteristics of people in care homes. As a result, in the early days of the pandemic, there was a lack of understanding about the impact of COVID-19 on people in care homes and initial data focused only on deaths in hospitals.
It is positive to see producers working hard to address these gaps. The recent development of statistics for adult social care in England now provide monthly information about vaccinations, infections, testing and availability of personal protective equipment in care homes. Collaboration between NHS Digital, ONS, NatCen Social Research and academics in England resulted in the large-scale Mental Health of Children and Young People survey. This survey provides a unique insight into the mental health of children and young people during the pandemic. Our recent review of statistics about children and young people found that key statistics about COVID-19 have made this part of the population more visible. However, there are notable gaps in the information available, such as separate analyses for vulnerable children and data on social outcomes which give children themselves a voice. Overall, data gaps have often taken considerable time to address, not least because of the pressures of the pandemic which producers were working under, and in some cases gaps remain.
It is tragic that it has taken the pandemic to focus attention on issues such as gaps in social care data. However, we are now encouraged to see greater attention to this issue from governments across the UK and strong commitments to making improvements. In Scotland, data on social care is a focus of the Independent Review of Adult Social Care in Scotland and 'The Future of Social Care and Support in Scotland' Parliamentary Inquiry. The Chief Statistician in Scotland also recently outlined the ambition to improve the completeness of data on protected characteristics. In Wales, there is a focus on improving the way social care data are used, and we are pleased to see a commitment to create an Equality Data Unit in the Welsh Government's Programme for 2021 to 2026. There is also the ongoing programme of work by the Race Disparity Unit to improve the quality of ethnicity data on health records in England.
Producers must work with users to understand and address existing data gaps, such as those on social care, mental health, and ethnicity and other protected characteristics. The long-term and indirect impacts of the pandemic on society's health and wellbeing will also need to be understood.
Data infrastructure impacted the ability of some statistics producers to respond to the demands of the pandemic.
Flexible and joined-up data infrastructures are needed so producers can respond quickly to new data needs.
The speed of response of producers during the pandemic was highly impressive. As we highlighted in our State of the UK Statistical System report earlier this year, producers were quick to set up new data collections, data sharing processes and to publish new statistical releases. The sense that this was a contribution to a national effort, less bureaucracy, and a focus on 'good enough', rather than waiting for perfection, helped producers to be responsive and agile.
Where good data infrastructures were already in place, producers found it easier to adapt them and respond quickly. However, new operational systems created in response to the pandemic did not always consider future data and publishing needs – such as the issue of understanding the NHS Test and Trace programme in England discussed in lesson 3 above. We recognise that challenges with new operational systems will sometimes result in delays to publishing data. When this happens, producers should provide a clear explanation to users.
Existing challenges relating to data infrastructure also presented problems for producers – for example, fragmented data owned by multiple bodies, legal barriers, inconsistent formats, legacy software, and non-digital data. These issues resulted in challenges for data sharing and linking, and an over-reliance on burdensome manual processes – though we welcome the fact that some producers have since automated these processes. The UK coronavirus dashboard and the NHS COVID-19 Data Store are two examples of outputs produced using automated pipelines. We encourage producers to build on these developments, automating processes using code where possible to reduce the risk of error and create processes which are sustainable and flexible – as recommended in our report on Reproducible Analytical Pipelines. Producers should continue to consider what analytical code can be published or shared with other organisations to enhance transparency, reduce duplication of effort, and improve consistency – for all analysis, not just that relating to COVID-19.
We welcome the focus on improving technical infrastructure and data architecture in the draft data strategy for NHS England. The Department of Health and Social Care should ensure that those working on official statistics are included as key stakeholders in these efforts. Producers will also benefit from collaboration in this area – to learn from each other's successes, such as the National Clinical Data Store in Scotland and the National Data Resource in Wales.
There is a need for improved, system-wide data infrastructures within each country. This will be challenging, but producers should learn from each other's successes to make progress.
Governments must involve statisticians and analysts when setting up new operational systems so that analytical requirements and publishing plans are built-in from the start, and so that there is a good understanding of the expectations of the Code of Practice for Statistics.
Flexible use of analytical resource supported the impressive work by statistics producers.
Sufficient investments in recruitment and retention of skilled statisticians are required so statistics continue to be sustainable and responsive.
The impressive response by producers required a large amount of resource and dedicated effort. Producers were quick to review existing work, deciding what could be paused to allow extra resource to be dedicated to the pandemic response. This included moving analytical staff between departments and organisations through a programme of loans and secondments, often supported by the Analysis Function's analytical resourcing hub. Directors of Analysis played an important role in quickly redeploying staff to work on the pandemic response. However, in hindsight some producers reflected that this flexible sharing of resource could have been done more, and more quickly. The Analysis Function should consider the lessons from the approach taken during the pandemic and develop a model of resource-sharing which most effectively supports new demands in future.
Many producers feel that the past 18 months have been relentless, with staff working round the clock and through weekends and holidays – this level of work has continued for far longer than anticipated at the start of the pandemic. Maintaining staff wellbeing and morale has been challenging. Many producers feel they are still responding to the pandemic while trying to reinstate previous work which was paused and meet new demands. A significant challenge for some producers is recruiting and retaining people with the right skills. In the past we have heard anecdotally that this was a challenge for the whole statistical system. The increased burden on producers during the pandemic appears to have exacerbated it further. However, relatively little is known or published about new starters, leavers and vacancies in the Government Statistical Service (GSS) – a data gap we highlighted in our Statistical Leadership report earlier this year. It is important to collect and share this information so that the scale of the problem can be assessed and effectively resolved.
To respond effectively to new priorities, analytical resource should continue to be shared flexibly across governments and non-government bodies. The Analysis Function should consider a longer-term resourcing approach across departments and professions, and also consider how the model used during the pandemic can be even more agile in future.
Producers should continue to review their existing statistical releases. They should decide, based on balancing user needs with resource and the ongoing burden on staff, which statistics should be continued, restarted, or stopped, and how or with what frequency.
The GSS People Committee should regularly monitor the membership of the Government Statistician Group, including information about central GSS recruitment and professional membership by organisation. The Analysis Function should regularly collate and publish data on the Analysis Function, including breakdowns by profession to better understand similarities and differences across analytical professions.


We're back! After quite some time we are finally back within the German Counterstrike scene with the former roster of Team SkinBaron!
After a prolonged search in which we have talked to great teams and people, we have finally found a team that perfectly fits Wolves eSports in all areas. After the first meeting with our new team we have seen that it would make sense for both sides longterm and we can't wait for them to represent Wolves from now on.
While the team did not make it into Division 2 of the ESL Meisterschaft after a loss against EnRo Griffins, they still have the chance to make it into Division 2 of the 99Damage Liga in the upcoming playoffs. Apart from that, the team will represent Wolves eSports at the SUPREME MASTERS LAN in Amriswil, Switzerland from August 31st – September 2nd. As one of 32 teams, they will fight for their share of the 6475€ prize pool against teams such as Berzerk, mYinsanity, eEriness, and eSuba just to name a few.
In the upcoming months, the team is going to give it their all to achieve the goals they have set themselves – Division 2 of the ESLM and the 99Liga. We believe that these goals are certainly possible and hope that both Wolves and the team will benefit from the combined efforts we are putting in.
With the announcement of a German CSGO team, we are still on the look-out for a European team but stay tuned for more great news coming in the next few days!


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Q: How many lines can pass through a point with the same distance from two other points? Consider three points $A$,$B$ and $C$ in the space which are not on the same line. how many line we can pass through $A$ so that it has the same distance from both points $B$ and $C$ ?
$1)\text{infinity}\qquad2)\text{at least two lines}\qquad3)\text{two lines}\qquad4)\text{one line}$
I know that in the space the points on the perpendicular bisector plane of a segment have the same distance from both endpoint of that segment. So I think if $A$ place on the perpendicular bisector of $BC$ then any line on that plane which passes through $A$ has the same distance from $B$ and $C$.
But I don't know how to solve the problem in general.

A: You reasoning regarding perpendicular bisector plane is valid.
Now, consider vectors $\vec{AB}$ and $\vec{AC}$. The problem boils down to finding vector $\vec{AX}$ such that $\left\lVert\vec{AB} \times \vec{AX}\right\rVert=\left\lVert\vec{AC} \times \vec{AX} \right\rVert$ or $\left\lVert\vec{AB}  \right\rVert \left\lVert\vec{AX}  \right\rVert \sin \alpha=\left\lVert\vec{AC}  \right\rVert \left\lVert\vec{AX}  \right\rVert \sin \beta$ where $\alpha$ and $\beta$ are angles between the corresponding vectors. Thus, we have the equation  $\left\lVert\vec{AB}  \right\rVert \sin \alpha=\left\lVert\vec{AC}  \right\rVert \sin \beta$ which has infinite number of solutions as all we need to do is to pick angle $\alpha$ such as $-1 \le \frac{||\vec{AB}||}{||\vec{AC}||}\sin \alpha \le 1$ and we can find angle $\beta$ to satisfy our equation above.
P.S. Try to solve this problem in 2D to see that two solutions will always exist in 2D.

redpajama_set_name: RedPajamaStackExchange


Devon Ditto is this week's PTO Student of the Week at Powell County High School. Devon was nominated by Mrs. Susan Watson. According to Mrs. Watson, Devon has been very dedicated to his classes and has worked diligently to complete assignments in order to become a PCHS 2018 graduate. Devon enjoys his English class with Mrs. Laura Davis and his classes with Mrs. Susan Watson. He says that both teachers are the best and are always helpful.
Devon played football while at PCHS and enjoys playing all sports. He is athletic and very much a "team player". Devon is very outgoing with a friendly personality. Mrs. Martina Skidmore and her husband Rick are happy having Devon spend time with their active family.
After graduation, Devon's plans to attend a community college and major in psychology. Devon is very excited and looking forward to his future.


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Home › Fact Sheets › Planning - Planning Applications
This Fact sheet is about complaints about planning applications. It should be read together with our general information booklet about our service.
The Council is the local planning authority for its area (in some areas this is a function of a National Park Authority). Planning authorities are responsible for making decisions and taking action about various planning matters. They have to work within the law, government guidance and the authority's own policy. The Ombudsman may be able to help you with your complaint against a planning authority.
He can look at whether there is something wrong in the way in which the Council went about deciding an application for a neighbouring development. This can include the failure to:
Notify you of an application, which may affect you, so that you were unable to object;
Notify other bodies which should have been consulted;
Consult you on amended plans;
Give reasons for its decision to approve an application;
Take your objections into account (this is not the same as not agreeing with your objections);
Take into account relevant planning considerations.
Incorrect /misleading /incomplete planning advice given, before you made an application which can be shown to have caused significant problems for you;
The Council did not have a system in place for ensuring that advice it gave was properly recorded;
The Council failed to make clear that the advice was informal and did not commit the Council to make a particular decision in the future;
The Council failed to follow the proper procedures or to take into account relevant planning considerations.
He cannot look at:
A planning decision which has been properly taken where there is no evidence of administrative fault;
Complaints where there is or was a right of appeal against the Council's decision to the Planning Inspectorate. This can apply where there is delay in deciding an application or where an application has been refused;
A complaint about an application until a decision has been taken and he can consider the effect on you.
He is unlikely to look at your complaint if it relates to private matters between you and your neighbour – e.g. boundary disputes.
The Ombudsman cannot overturn the decision or demolish the development even if you are affected, but, if your complaint is upheld, he will consider other ways of reducing any effect on you.
Planning Aid Wales can give helpful information on planning matters. You can contact them via phone on 029 2062 5000 or via the internet at www.planningaidwales.org.uk
Planning information can also be obtained at http://www.planningportal.gov.uk/wales/public
The Council's own website may also contain information about some planning matters.
The Ombudsman is independent and impartial; he cannot order public bodies to do what he recommends – but, in practice, they almost always do. Examples of cases that the Ombudsman has looked at can be found on our website. Please see www.ombudsman.wales.


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Which countries are part of Scandinavia?
Scandinavia is in northern Europe. But where exactly are its borders and which countries are regarded as Scandinavian countries? What about Scandinavian languages and the flags with the Scandinavian Cross? Learn more about Scandiavia and the countries of northern Europe here.
In most definitions "Scandinavia" is defined as the area of the Scandinavian Peninsula in Northern Europe, which is dominated by the Scandinavian Mountains (see for example Wikipedia or Encyclopædia Britannica).
This means that Norway and Sweden are clearly Scandinavian countries. The northeastern border of the Scandinavian Peninsula is not clearly defined. Northern Finland and most of Finnish Lapland do undoubtedly reach into the Scandinavian Peninsula and Mountains.
The most northern point of Scandinavia is the Nordkap in Norway. In the south of the country is Scandinavia's highest peak, the Galdhøpiggen with an elevation of more than 2,400 meters.
left: Scandinavian Peninsula with Norway and Sweden.
right: The "usual" Scandinavian countries Norway, Sweden, and Denmark.
The issue with a proper definition for "Scandinavia" is clearly that geographical and cultural elements are usually combined. Depending on which aspects you stress, this leads to different results, when determining which countries should be regarded as part of Scandiavia.
Denmark is mostly regarded as one of the Scandiavian countries, which has foremost linguistic, historic, and overall cultural reasons. The Danish autnomous areas of the Faroe Islands and Greenland are viewed separately, though. In pure geographical terms there would be a lot of ground to claim Denmark to be Central Europe's most northern part and perhaps a transition area to Scandinavia. Depending on your point of view regarding Denmark, Scandinavia's most southern point is either at Smygehuk in Trelleborg in southern Sweden, or on the Danish island Falster in the Baltic Sea.
Finland is less frequently counted as part of Scandinavia than Denmark, even though, as stated above, geographically there is a lot of reason to count at least the country's north as part of it. The Finnish history is closely intertwined with that of Sweden in particular, since the country was under Swedish rule for centuries.
Iceland also has close linguistic and historic ties with Scandiavia and is therefore sometimes also seen as part of a Scandinavian or Nordic cultural community of values, even though the island on Europe's outmost margin is situated almost 1,000 kilometers away from the Norwegian coast.
Fenno-Scandinavia with Norway, Sweden, Finland,
as well as Karelia and Kola Peninsula in Russia
Northern Europe and Fenno-Scandinavia
As stated above the Scandinavian Peninsula's northeastern border is not clearly defined. Sometimes a larger area named Fenno-Scandinavia, or Fennoscandia is regarded. Besides the Scandinavian Peninsula it also contains all of Finland ("Fenno") and the Russian territories of the Kola Peninsula and Karelia.
Besides the Scandinavian languages
Faroese and Icelandic are also part of the family of Nordic languages.
Scandinavian and Nordic languages
Denmark, Sweden, and Norway are not only geographically close to each other and have a rich common history. Also their languages are quite similar and usually Danes, Swedes, and Norwegians can easily communicate with each other with everyone still speaking their mother tongue.
Icelandic and Faroese also belong to the North Germanic, or Nordic languages. Even though they are still quite similar, communication with continental Scandinavians is significantly more difficult. That's why Icelandic and Faroese are also referred to as the Insular Nordic languages.
Geographically and historically Finland has close ties with the rest of Scandinavia. The Finnish language, however, sets the country clearly apart. Where the Nordic languages, as English, Dutch, or German, are part of the family of Germanic and Indo-European languages, Finnish is an Uralic language, and thus distantly related to Estonian and Hungarian. About 300,000 Finnish native speakers live in Sweden and Finnish has the status of an official minority language there. More than 5 % of the Finnish population speak Swedish as their first language. In Finland Swedish is one of two official languages and therefore equal to Finnish by law.
Countries and autonomous regions with a Scandinavian Cross-flag
Scandinavian Cross
Another commonality of the countries of Northern Europe, which immediately catches the eye, is the flag design. The flags of Denmark, Sweden, Norway, Iceland, and Finland are all dominated by the Nordic or Scandinavian Cross. There are also some autonomous regions, as the Finnish island Åland, the Faroe Islands and also Shetland und Orkney in the United Kingdom which have a Nordic Cross on their flag.
The Christian symbol was first used on the Danish flag "Dannebrog". According to an old legend the flag fell from the sky during the Battle of Lindanise (near today's Tallin) in 1219. Thereupon the Danes could yet win a battle which before had already appeared to be lost. Historic proof for the use of the flag is confirmed for the 14th century. Later the Danes also introduced similar flags for Norway and Iceland which were both mostly under Danish rule up until the 19th, respectively 20th century.
Sweden introduced its flag in the 16th century. Finland uses its flag since the country's independence in 1917.
Member states (incl. associated members) and observers of the Nordic Council
The Nordic Council
Scandinavia's history was for a long time dominated by the rivalry between Denmark and Sweden, resulting in countless battles and wars up until the 19th century. Nowadays the countries of northern Europe are close allies who work together trustfully in many areas. There are, however, very different view regarding the European Union among the Nordic countries. At first Denmark, Norway, and Sweden were founding members of the free trade organisation EFTA, with Finland joining shortly after and Iceland becoming a member in 1970. In 1973 Denmark left the EFTA and, together with Ireland and the United Kingdom, joined the European Economic Community (which transformed into the EU in 1992). In 1995 Finland and Sweden also became EU-members. Today Iceland and Norway, together with Switzerland and Liechtenstein, are the remaining member states of the EFTA.
One of the most important forums for the political and cultural coordination of the countries of Northern Europe is the Nordic Council, which was established in 1952 and has its headquarters in Copenhagen. It consists of the five souvereign states Denmark, Norway, Sweden, Finland, and Iceland, as well as the autonomous areas Åland, the Faroe Islands, and Greenland as associated members. The Baltic countries Estonia, Latvia, and Lithuania, and the northernmost German state Schleswig-Holstein have an official status as observers in the council.
Even though the decisions of the Nordic Council are not binding for the member states it is valued as an instrument of exchange and cooperation between the countries.
Ski Resorts of Scandinavia
Our first Marmota Map of northern Europe is SKI RESORTS OF SCANDINAVIA with all downhill ski areas in Norway, Sweden, and Finland (with data from skiresort.info). With everything above this leads to two questions:
1) Shouldn't Denmark be on a map with this title?
This is relatively easy to answer. Denmark is a great holiday destination, but unfortunately not the best one if you want to go skiing. Since October 2019 technically there is now a Danish outdoor ski area - CopenHill in Copenhagen on the roof of the incineration plant Amager Bakke, having about 400 slope meters. Since it is a purely artificial area, which can also operate without any snow, we still decided not to include it on our list of Scandinavian ski areas (as we also did with indoor ski resorts). That's why, in the end, we left out Denmark completely on our ski map.
2) Is Finland part of Scandiavia?
We can see that our decision to put Finland on the map as a whole is controversial, considering the map title. For us the crucial point is that Finland's northern part is clearly part of Scandinavia geographically. Defining a border through Finnish territory would also be somehow arbitrary. Even though most Finnish ski areas are in the south of the country, where also the vast majority of the people live there are also strong cultural ties with the rest of Scandinavia. That's why we finally decided to include the Finnish south on our map of the Scandinavian ski areas.
We hope we will be able to produce more maps of northern Europe in the future. We may not have convinced everyone but we still hope to have brought up some valid arguments for our decision to publish our first map with this area and title.
Unskyld dear Danish and all Denmark-fans and ski ahoy!
Area of the Marmota Map SKI RESORTS OF SCANDINAVIA without Denmark and with all of Finland.
Galdhøpiggen
(By now) Scandinavia's undisputed highest mountain – Galdhøpiggen in southern Norway.
All 299 ski resorts in Norway, Sweden and Finland on one map.
The World of Animals
Define Scandinavia's borders just as you wish - on our world map of animals – illustratet by Dieter Braun


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Inflight wifi: the dream, the dismal reality and the future
I was three hours into a four-and-a-half-hour flight across Europe this summer when I called it. In-flight wifi doesn't work. Not in a real, impactful way, anyway – not to the point where you can ethically slap a price tag on it and suggest people can get a bit of work done between the seatbelt sign switch-off and the tea trolley.
In my case that price was £7.99 for an "entire flight" wifi pass. That's £1.77 an hour spent on next to nothing – this in-flight wifi creaked through about 40 minutes of the flight (while we were over land) before disappearing into nothingness and refusing to reconnect. Cabin crew were shruggy at best – and, arguably, it's not their job to troubleshoot my connectivity issues. They're busy making sure we're all alive – or at least caffeinated – and who could argue with that?
Inflight wifi has been a "reality" since around 2006. British Airways, Virgin Atlantic, Emirates, Norwegian and Qatar Airways are just some of the airlines to launch a version of it since then. Some offer free access to business class customers, most charge anything from £2.99 to £20 for a wifi pass. Delta is the next carrier to announce it will be introducing internet at altitude, with plans to launch its free Delta Sync package from as early as 1 February. But 16 years on it remains a hit-and-miss experience.
The problem, says business travel expert John Walton, is that getting wifi on planes is so much more complex than wifi in hotel lobbies, or even on trains. Explaining it to me as though I were a five-year-old (my instructions), Walton tells me how aircraft pick up wifi from two places: from cell towers on the ground – when at lower altitude and closer to land – or from satellites in outer space.
The former is known as air-to-ground (ATG) and is picked up by an aerial on the plane, creating a hotspot to which passengers can connect. For the latter method to work, the aircraft antenna has to be able to reach an orbiting satellite some 20,000 miles from earth. So the signal has to travel much further – hence in-flight wifi often vanishing into thin air as we fly over long stretches of sea, far from the ground towers.
But there are layers of reasons why sky-high wifi seems like a dinosaur – something that should be soundtracked by the bleating of a 1990s modem struggling to dial up. For one,…
Rail passengers face fourth day of massive travel disruption as train strikes rumble on
Why my New Year's resolution is to travel without flying – for the fourth year in a row