MicroRNA miR-199a-5p regulates smooth muscle cell proliferation and morphology by targeting Wnt2 signaling pathway

MicroRNA miR-199a-5p impairs tight junction formation leading to increased urothelial permeability in bladder pain syndrome. Now using transcriptome analysis in urothelial TEU-2 cells we implicate it in the regulation of cell cycle, cytoskeleton remodeling, TGF and Wnt signaling pathways. MiR-199a-5p is highly expressed in the smooth muscle layer of the bladder and we altered its levels in bladder smooth muscle cells (SMC) to validate the pathway analysis. Inhibition of miR-199a-5p with antimiR increased SMC proliferation, reduced cell size and up-regulated miR-199a-5p targets, including Wnt2. Overexpression of Wnt2 protein or treating SMCs with recombinant Wnt2 closely mimicked the miR-199a-5p inhibition, whereas down-regulation of Wnt2 in antimiR-expressing SMCs with shRNA restored cell phenotype and proliferation rates. Overexpression of miR-199a-5p in the bladder SMCs significantly increased cell size and up-regulated SM22, SM alpha-actin and SM myosin heavy chain mRNA and protein levels. These changes, as well as increased expression of ACTG2, TGFB1I1, and CDKN1A were mediated by up-regulation of smooth muscle-specific transcriptional activator myocardin at mRNA and protein levels. Myocardin-related transcription factor (MRTF-A) downstream targets Id3 and MYL9 were also induced. Up-regulation of myocardin was accompanied by down-regulation of Wnt-dependent inhibitory Kruppel-like transcription factor 4 (KLF4) in miR-199a-5p overexpressing cells. In contrast, KLF4 was induced in antimiR-expressing cells following the activation of Wnt2 signaling, leading to repression of myocardin-dependent genes. MiR-199a-5p plays a critical role in the Wnt2-mediated regulation of proliferative and differentiation processes in the smooth muscle and may behave as a key modulator of smooth muscle hypertrophy, relevant for organ remodeling.

Dendritic spine morphology determines membrane-associated protein exchange between dendritic shafts and spine heads.

The purpose of this study was to examine whether variability in the shape of dendritic spines affects protein movement within the plasma membrane. Using a combination of confocal microscopy and the fluorescence loss in photobleaching technique in living hippocampal CA1 pyramidal neurons expressing membrane-linked GFP, we observed a clear correlation between spine shape parameters and the diffusion and compartmentalization of membrane-associated proteins. The kinetics of membrane-linked GFP exchange between the dendritic shaft and the spine head compartment were slower in dendritic spines with long necks and/or large heads than in those with short necks and/or small heads. Furthermore, when the spine area was reduced by eliciting epileptiform activity, the kinetics of protein exchange between the spine compartments exhibited a concomitant decrease. As synaptic plasticity is considered to involve the dynamic flux by lateral diffusion of membrane-bound proteins into and out of the synapse, our data suggest that spine shape represents an important parameter in the susceptibility of synapses to undergo plastic change.

Investigation of retinal morphology alterations using spectral domain optical coherence tomography in a mouse model of retinal branch and central retinal vein occlusion.

Retinal vein occlusion is a leading cause of visual impairment. Experimental models of this condition based on laser photocoagulation of retinal veins have been described and extensively exploited in mammals and larger rodents such as the rat. However, few reports exist on the use of this paradigm in the mouse. The objective of this study was to investigate a model of branch and central retinal vein occlusion in the mouse and characterize in vivo longitudinal retinal morphology alterations using spectral domain optical coherence tomography. Retinal veins were experimentally occluded using laser photocoagulation after intravenous application of Rose Bengal, a photo-activator dye enhancing thrombus formation. Depending on the number of veins occluded, variable amounts of capillary dropout were seen on fluorescein angiography. Vascular endothelial growth factor levels were markedly elevated early and peaked at day one. Retinal thickness measurements with spectral domain optical coherence tomography showed significant swelling (p<0.001) compared to baseline, followed by gradual thinning plateauing two weeks after the experimental intervention (p<0.001). Histological findings at day seven correlated with spectral domain optical coherence tomography imaging. The inner layers were predominantly affected by degeneration with the outer nuclear layer and the photoreceptor outer segments largely preserved. The application of this retinal vein occlusion model in the mouse carries several advantages over its use in other larger species, such as access to a vast range of genetically modified animals. Retinal changes after experimental retinal vein occlusion in this mouse model can be non-invasively quantified by spectral domain optical coherence tomography, and may be used to monitor effects of potential therapeutic interventions.

A numerical model for inert gas transport in the lung based on fractal airway morphology

Patients suffering from cystic fibrosis (CF) show thick secretions, mucus plugging and bronchiectasis in bronchial and alveolar ducts. This results in substantial structural changes of the airway morphology and heterogeneous ventilation. Disease progression and treatment effects are monitored by so-called gas washout tests, where the change in concentration of an inert gas is measured over a single or multiple breaths. The result of the tests based on the profile of the measured concentration is a marker for the severity of the ventilation inhomogeneity strongly affected by the airway morphology. However, it is hard to localize underlying obstructions to specific parts of the airways, especially if occurring in the lung periphery. In order to support the analysis of lung function tests (e.g. multi-breath washout), we developed a numerical model of the entire airway tree, coupling a lumped parameter model for the lung ventilation with a 4th-order accurate finite difference model of a 1D advection-diffusion equation for the transport of an inert gas. The boundary conditions for the flow problem comprise the pressure and flow profile at the mouth, which is typically known from clinical washout tests. The natural asymmetry of the lung morphology is approximated by a generic, fractal, asymmetric branching scheme which we applied for the conducting airways. A conducting airway ends when its dimension falls below a predefined limit. A model acinus is then connected to each terminal airway. The morphology of an acinus unit comprises a network of expandable cells. A regional, linear constitutive law describes the pressure-volume relation between the pleural gap and the acinus. The cyclic expansion (breathing) of each acinus unit depends on the resistance of the feeding airway and on the flow resistance and stiffness of the cells themselves. Special care was taken in the development of a conservative numerical scheme for the gas transport across bifurcations, handling spatially and temporally varying advective and diffusive fluxes over a wide range of scales. Implicit time integration was applied to account for the numerical stiffness resulting from the discretized transport equation. Local or regional modification of the airway dimension, resistance or tissue stiffness are introduced to mimic pathological airway restrictions typical for CF. This leads to a more heterogeneous ventilation of the model lung. As a result the concentration in some distal parts of the lung model remains increased for a longer duration. The inert gas concentration at the mouth towards the end of the expirations is composed of gas from regions with very different washout efficiency. This results in a steeper slope of the corresponding part of the washout profile.

A stereotaxic, population-averaged T1w ovine brain atlas including cerebral morphology and tissue volumes

Standard stereotaxic reference systems play a key role in human brain studies. Stereotaxic coordinate systems have also been developed for experimental animals including non-human primates, dogs, and rodents. However, they are lacking for other species being relevant in experimental neuroscience including sheep. Here, we present a spatial, unbiased ovine brain template with tissue probability maps (TPM) that offer a detailed stereotaxic reference frame for anatomical features and localization of brain areas, thereby enabling inter-individual and cross-study comparability. Three-dimensional data sets from healthy adult Merino sheep (Ovis orientalis aries, 12 ewes and 26 neutered rams) were acquired on a 1.5 T Philips MRI using a T1w sequence. Data were averaged by linear and non-linear registration algorithms. Moreover, animals were subjected to detailed brain volume analysis including examinations with respect to body weight (BW), age, and sex. The created T1w brain template provides an appropriate population-averaged ovine brain anatomy in a spatial standard coordinate system. Additionally, TPM for gray (GM) and white (WM) matter as well as cerebrospinal fluid (CSF) classification enabled automatic prior-based tissue segmentation using statistical parametric mapping (SPM). Overall, a positive correlation of GM volume and BW explained about 15% of the variance of GM while a positive correlation between WM and age was found. Absolute tissue volume differences were not detected, indeed ewes showed significantly more GM per bodyweight as compared to neutered rams. The created framework including spatial brain template and TPM represent a useful tool for unbiased automatic image preprocessing and morphological characterization in sheep. Therefore, the reported results may serve as a starting point for further experimental and/or translational research aiming at in vivo analysis in this species.

The genetics of reproductive organ morphology in two Petunia species with contrasting pollination syndromes

Main conclusion Switches between pollination syndromes have happened frequently during angiosperm evolution. Using QTL mapping and reciprocal introgressions, we show that changes in reproductive organ morphology have a simple genetic basis. In animal-pollinated plants, flowers have evolved to optimize pollination efficiency by different pollinator guilds and hence reproductive success. The two Petunia species, P. axillaris and P. exserta, display pollination syndromes adapted to moth or hummingbird pollination. For the floral traits color and scent, genetic loci of large phenotypic effect have been well documented. However, such large-effect loci may be typical for shifts in simple biochemical traits, whereas the evolution of morphological traits may involve multiple mutations of small phenotypic effect. Here, we performed a quantitative trait locus (QTL) analysis of floral morphology, followed by an in-depth study of pistil and stamen morphology and the introgression of individual QTL into reciprocal parental backgrounds. Two QTLs, on chromosomes II and V, are sufficient to explain the interspecific difference in pistil and stamen length. Since most of the difference in organ length is caused by differences in cell number, genes underlying these QTLs are likely to be involved in cell cycle regulation. Interestingly, conservation of the locus on chromosome II in a different P. axillaris subspecies suggests that the evolution of organ elongation was initiated on chromosome II in adaptation to different pollinators. We recently showed that QTLs for pistil and stamen length on chromosome II are tightly linked to QTLs for petal color and volatile emission. Linkage of multiple traits will enable major phenotypic change within a few generations in hybridizing populations. Thus, the genomic architecture of pollination syndromes in Petunia allows for rapid responses to changing pollinator availability.

From supramolecular sheets to fibers and back: establishing the critical parameters that govern the morphology of pyrene-based self-assembled materials

The precise arraying of functional entities in morphologically well-defined shapes remains one of the key challenges in the processing of organic molecules1. Among various π-conjugated species, pyrene exhibits a set of unique properties, which make it an attractive compound for the utilization in materials science2. In this contribution we report on properties of self-assembled structures prepared from amphiphilic pyrene trimers (Py3) consisting of phosphodiester-linked pyrenes. Depending on the geometry of a pyrene core substitution (1.6-, 1.8-, or 2.7- type, see Scheme), the thermally-controlled self-assembly allows the preparation of supramolecular architectures of different morphologies in a bottom-up approach: two-dimensional (2D) nanosheets3 are formed in case of 1.6- and 2.7-substitution4 whereas one-dimensional (1D) fibers are built from 1.8- substituted isomers. The morphologies of the assemblies are established by AFM and TEM, and the results are further correlated with spectroscopic and scattering data. Two-dimensional assemblies consist of an inner layer of hydrophobic pyrenes, sandwiched between a net of phosphates. Due to the repulsion of the negative charges, the 2D assemblies exist mostly as free-standing sheets. An internal alignment of pyrenes leads to strong exciton coupling with an unprecedented observation (simultaneous development of J- and H-bands from two different electronic transitions). Despite the similarity in spectroscopic properties, the structural parameters of the 2D aggregates drastically depend on the preparation procedure. Under certain conditions extra-large sheets (thickness of 2 nm, aspect ratio area/thickness ~107) in aqueous solution are formed4B. Finally, one-dimensional assemblies are formed as micrometer-long and nanometer-thick fibers. Both, planar and linear structures are intriguing objects for the creation of conductive nanowires that may find interest for applications in supramolecular electronics.

From supramolecular sheets to fibers and back: establishing the critical parameters that govern the morphology of pyrene-based self-assembled materials

The precise arraying of functional entities in morphologically well-defined shapes remains one of the key challenges in the processing of organic molecules1. Among various π-conjugated species, pyrene exhibits a set of unique properties, which make it an attractive compound for the utilization in materials science2. In this contribution we report on properties of self-assembled structures prepared from amphiphilic pyrene trimers (Py3) consisting of phosphodiester-linked pyrenes. Depending on the geometry of a pyrene core substitution (1.6-, 1.8-, or 2.7- type, see Scheme), the thermally-controlled self-assembly allows the preparation of supramolecular architectures of different morphologies in a bottom-up approach: two-dimensional (2D) nanosheets3 are formed in case of 1.6- and 2.7-substitution4 whereas one-dimensional (1D) fibers are built from 1.8- substituted isomers. The morphologies of the assemblies are established by AFM and TEM, and the results are further correlated with spectroscopic and scattering data. Two-dimensional assemblies consist of an inner layer of hydrophobic pyrenes, sandwiched between a net of phosphates. Due to the repulsion of the negative charges, the 2D assemblies exist mostly as free-standing sheets. An internal alignment of pyrenes leads to strong exciton coupling with an unprecedented observation (simultaneous development of J- and H-bands from two different electronic transitions). Despite the similarity in spectroscopic properties, the structural parameters of the 2D aggregates drastically depend on the preparation procedure. Under certain conditions extra-large sheets (thickness of 2 nm, aspect ratio area/thickness ~107) in aqueous solution are formed4B. Finally, one-dimensional assemblies are formed as micrometer-long and nanometer-thick fibers. Both, planar and linear structures are intriguing objects for the creation of conductive nanowires that may find interest for applications in supramolecular electronics.

Trends in incidence of oesophageal and gastric cancer according to morphology and anatomical location, in Switzerland 1982-2011.

QUESTION UNDER STUDY/PRINCIPLES This study aimed to evaluate trends in the incidence of oesophageal and gastric cancer by anatomical location and histology using nationally representative Swiss data. METHODS We included all oesophageal and gastric cancers recorded in 10 Swiss population-based cancer registries 1982-2011. We calculated age-standardised incidence rates (ASIRs) per 100 000 person-years (PY) (European standard) for both cancer sites stratified by sex, language region (German, French-Italian), morphology and anatomical location. To assess time trends, we estimated annual percentage changes (APCs) with 95% confidence intervals (95% CIs). RESULTS ASIR of oesophageal adenocarcinoma increased in both sexes and language regions (p <0.001). The steepest increase occurred in males of the German-speaking region (APC 6.8%, 95% CI 5.8-7.8) with ASIRs of 0.8 per 100,000 PY in 1982-1987 and 3.9 per 100.000 PY in 2007-2011. Incidence of oesophageal squamous cell carcinoma decreased significantly in males of both language regions by around -1.5% per year. In contrast, a slight but significant increase (APC 1.4%, 95% CI 0.3-2.4]) of oesophageal squamous cell carcinoma was observed in females of the German-speaking region. We observed stable rates for cancer of the gastric cardia. The incidence of noncardia gastric cancer decreased substantially in both sexes and language regions (p <0.001). CONCLUSION In Switzerland, the incidence of oesophageal adenocarcinoma has risen whereas incidence of noncardia gastric cancer has decreased substantially as observed in other developed countries.

Paleohydraulic reconstruction of a 40 kyr-old terrace sequence implies that water discharge was larger than today

The evolution of landscapes crucially depends on the climate history. This is particularly evident in South America where landscape responses to orbital climate shifts have been well documented. However, while most studies have focused on inferring temperature variations from paleoclimate proxy data, estimates of water budget changes have been complicated because of a lack of adequate physical information. Here, we present a methodology and related results, which allowed us to extract water discharge values from the sedimentary record of the 40 Ka-old fluvial terrace deposits in the Pisco valley, western Peru. In particular, this valley hosts a Quaternary cut-and-fill succession that we used, in combination with beryllium-10 (10Be)-based sediment flux, gauging records, channel geometries and grain size measurements, to quantitatively assess sediment and water discharge values c. 40 Ka ago in relation to present-day conditions. We compare these discharge estimates to the discharge regime of the modern Pisco River and find that the water discharge of the paleo-Pisco River, during the Minchin pluvial period c. 40 Ka ago, was c. 7–8 times greater than the modern Pisco River if considering the mean and the maximum water discharge. In addition, the calculations show that inferred water discharge estimates are mainly dependent on channel gradients and grain size values, and to a lesser extent on channel width measures. Finally, we found that the c. 40 Ka-old Minchin terrace material was poorer sorted than the modern deposits, which might reflect that sediment transport during the past period was characterized by a larger divergence from equal mobility compared to the modern situation. In summary, the differences in grain size distribution and inferred water discharge estimates between the modern and the paleo-Pisco River suggests that the 40 Ka-old Minchin period was characterized by a wetter climate and more powerful flood events.

Twelve percent of hips with a primary cam deformity exhibit a slip-like morphology resembling sequelae of slipped capital femoral epiphysis

BACKGROUND In some hips with cam-type femoroacetabular impingement (FAI), we observed a morphology resembling a more subtle form of slipped capital femoral epiphysis (SCFE). Theoretically, the morphology in these hips should differ from hips with a primary cam-type deformity. QUESTIONS/PURPOSES We asked if (1) head-neck offset; (2) epiphyseal angle; and (3) tilt angle differ among hips with a slip-like morphology, idiopathic cam, hips after in situ pinning of SCFE, and normal hips; and (4) what is the prevalence of a slip-like morphology among cam-type hips? METHODS We retrospectively compared the three-dimensional anatomy of hips with a slip-like morphology (29 hips), in situ pinning for SCFE (eight hips), idiopathic cam deformity (171 hips), and 30 normal hips using radial MRI arthrography. Normal hips were derived from 17 asymptomatic volunteers. All other hips were recruited from a series of 277 hips (243 patients) seen at a specialized academic hip center between 2006 and 2010. Forty-one hips with isolated pincer deformity were excluded. Thirty-six of 236 hips had a known cause of cam impingement (secondary cam), including eight hips after in situ pinning of SCFE (postslip group). The 200 hips with a primary cam were separated in hips with a slip-like morphology (combination of positive fovea sign [if the neck axis did not intersect with the fovea capitis] and a tilt angle [between the neck axis and perpendicular to the basis of the epiphysis] exceeding 4°) and hips with an idiopathic cam. We evaluated offset ratio, epiphyseal angle (angle between the neck axis and line connecting the center of the femoral head and the point where the physis meets the articular surface), and tilt angle circumferentially around the femoral head-neck axis. Prevalence of slip-like morphology was determined based on the total of 236 hips with cam deformities. RESULTS Offset ratio was decreased anterosuperiorly in idiopathic cam, slip-like, and postslip (eg, 1 o'clock position with a mean offset ranging from 0.00 to 0.14; p < 0.001 for all groups) compared with normal hips (0.25 ± 0.06 [95% confidence interval, 0.13-0.37]) and increased posteroinferiorly in slip-like (eg, 8 o'clock position, 0.5 ± 0.09 [0.32-0.68]; p < 0.001) and postslip groups (0.55 ± 0.12 [0.32-0.78]; p < 0.001) and did not differ in idiopathic cam (0.32 ± 0.09 [0.15-0.49]; p = 0.323) compared with normal (0.31 ± 0.07 [0.18-0.44]) groups. Epiphyseal angle was increased anterosuperiorly in the slip-like (eg, 1 o'clock position, 70° ± 9° [51°-88°]; p < 0.001) and postslip groups (75° ± 13° [49°-100°]; p = 0.008) and decreased in idiopathic cam (50° ± 8° [35°-65°]; p < 0.001) compared with normal hips (58° ± 8° [43°-74°]). Posteroinferiorly, epiphyseal angle was decreased in slip-like (eg, 8 o'clock position, 54° ± 10° [34°-74°]; p < 0.001) and postslip (44° ± 11° [23°-65°]; p < 0.001) groups and did not differ in idiopathic cam (76° ± 8° [61°-91°]; p = 0.099) compared with normal (73° ± 7° [59°-88°]) groups. Tilt angle increased in slip-like (eg, 2/8 o'clock position, 14° ± 8° [-1° to 30°]; p < 0.001) and postslip hips (29° ± 10° [9°-48°]; p < 0.001) and decreased in hips with idiopathic cam (-7° ± 5° [-17° to 4°]; p < 0.001) compared with normal (-1° ± 5° [-10° to 8°]) hips. The prevalence of a slip-like morphology was 12%. CONCLUSIONS The slip-like morphology is the second most frequent pathomorphology in hips with primary cam deformity. MRI arthrography of the hip allows identifying a slip-like morphology, which resembles hips after in situ pinning of SCFE and distinctly differs from hips with idiopathic cam. These results support previous studies reporting that SCFE might be a risk factor for cam-type FAI.

Effects of Antiseptic Solutions Commonly Used in Dentistry on Bone Viability, Bone Morphology, and Release of Growth Factors

PURPOSE Antiseptic solutions are commonly used in dentistry for a number of sterilization procedures, including harvesting of bone chips, irrigation of extraction sockets, and sterilization of osteonecrotic bone. Despite its widespread use, little information is available regarding the effects of various antiseptic solutions on bone cell viability, morphology, and the release of growth factors. MATERIALS AND METHODS The antiseptic solutions included 1) 0.5% povidone iodine (PI), 2) 0.2% chlorhexidine diguluconate (CHX), 3) 1% hydrogen peroxide (H2O2), and 4) 0.25% sodium hypochlorite (HYP). Bone samples collected from porcine mandibular cortical bone were rinsed in the antiseptic solutions for 10 minutes and assessed for cell viability using an MTS assay and protein release of transforming growth factor (TGF-β1), bone morphogenetic protein 2 (BMP2), vascular endothelial growth factor (VEGF), interleukin (IL)-1β, and receptor activator of nuclear factor κB ligand (RANKL) using an enzyme-linked immunosorbent assay at 15 minutes and 4 hours after rinsing. RESULTS After antiseptic rinsing, changes to the surface protein content showed marked alterations, with an abundant protein layer remaining on CHX-rinsed bone samples. The amount of surface protein content gradually decreased in the following order: CHX, H2O2, PI, and HYP. A similar trend was also observed for the relative cell viability from within bone samples after rinsing, with up to 6 times more viable cells found in the CHX-rinsed bone samples than in the HYP- and PI-rinsed samples. An analysis of the growth factors found that both HYP and PI had significantly lower VEGF and TGF-β1 protein release from bone samples at 15 minutes and 4 hours after rinsing compared with CHX and H2O2. A similar trend was observed for RANKL and IL-1β protein release, although no change was observed for BMP2. CONCLUSIONS The results from the present study have demonstrated that antiseptic solutions present with very different effects on bone samples after 10 minutes of rinsing. Rinsing with CHX maintained significantly higher cell viability and protein release of growth factors potent to the bone remodeling cycle.