Design-functionality relationships for adhesion/growth-regulatory galectins.

Glycan-lectin recognition is assumed to elicit its broad range of (patho)physiological functions via a combination of specific contact formation with generation of complexes of distinct signal-triggering topology on biomembranes. Faced with the challenge to understand why evolution has led to three particular modes of modular architecture for adhesion/growth-regulatory galectins in vertebrates, here we introduce protein engineering to enable design switches. The impact of changes is measured in assays on cell growth and on bridging fully synthetic nanovesicles (glycodendrimersomes) with a chemically programmable surface. Using the example of homodimeric galectin-1 and monomeric galectin-3, the mutual design conversion caused qualitative differences, i.e., from bridging effector to antagonist/from antagonist to growth inhibitor and vice versa. In addition to attaining proof-of-principle evidence for the hypothesis that chimera-type galectin-3 design makes functional antagonism possible, we underscore the value of versatile surface programming with a derivative of the pan-galectin ligand lactose. Aggregation assays with N,N'-diacetyllactosamine establishing a parasite-like surface signature revealed marked selectivity among the family of galectins and bridging potency of homodimers. These findings provide fundamental insights into design-functionality relationships of galectins. Moreover, our strategy generates the tools to identify biofunctional lattice formation on biomembranes and galectin-reagents with therapeutic potential.

Calorimetric Analysis of the Interplay between Synthetic Tn Antigen-Presenting MUC1 Glycopeptides and Human Macrophage Galactose-Type Lectin.

Human macrophage galactose-type lectin (hMGL, HML, CD301, CLEC10A), a C-type lectin expressed by dendritic cells and macrophages, is a receptor for N-acetylgalactosamine α-linked to serine/threonine residues (Tn antigen, CD175) and its α2,6-sialylated derivative (sTn, CD175s). Because these two epitopes are among malignant cell glycan displays, particularly when presented by mucin-1 (MUC1), assessing the influence of the site and frequency of glycosylation on lectin recognition will identify determinants governing this interplay. Thus, chemical synthesis of the tandem-repeat O-glycan acceptor region of MUC1 and site-specific threonine glycosylation in all permutations were carried out. Isothermal titration calorimetry (ITC) analysis of the binding of hMGL to this library of MUC1 glycopeptides revealed an enthalpy-driven process and an affinity enhancement of an order of magnitude with an increasing glycan count from 6-8 µM for monoglycosylated peptides to 0.6 µM for triglycosylated peptide. ITC measurements performed in D2O permitted further exploration of the solvation dynamics during binding. A shift in enthalpy-entropy compensation and contact position-specific effects with the likely involvement of the peptide surroundings were detected. KinITC analysis revealed a prolonged lifetime of the lectin-glycan complex with increasing glycan valency and with a change in the solvent to D2O.

Imitating evolution's tinkering by protein engineering reveals extension of human galectin-7 activity.

Wild-type lectins have distinct types of modular design. As a step to explain the physiological importance of their special status, hypothesis-driven protein engineering is used to generate variants. Concerning adhesion/growth-regulatory galectins, non-covalently associated homodimers are commonly encountered in vertebrates. The homodimeric galectin-7 (Gal-7) is a multifunctional context-dependent modulator. Since the possibility of conversion from the homodimer to hybrids with other galectin domains, i.e. from Gal-1 and Gal-3, has recently been discovered, we designed Gal-7-based constructs, i.e. stable (covalently linked) homo- and heterodimers. They were produced and purified by affinity chromatography, and the sugar-binding activity of each lectin unit proven by calorimetry. Inspection of profiles of binding of labeled galectins to an array-like platform with various cell types, i.e. sections of murine epididymis and jejunum, and impact on neuroblastoma cell proliferation revealed no major difference between natural and artificial (stable) homodimers. When analyzing heterodimers, acquisition of altered properties was seen. Remarkably, binding properties and activity as effector can depend on the order of arrangement of lectin domains (from N- to C-termini) and on the linker length. After dissociation of the homodimer, the Gal-7 domain can build new functionally active hybrids with other partners. This study provides a clear direction for research on defining the full range of Gal-7 functionality and offers the perspective of testing applications for engineered heterodimers.

Galectin-Glycan Interactions: Guidelines for Monitoring by 77 Se NMR Spectroscopy, and Solvent (H2 O/D2 O) Impact on Binding.

Functional pairing between cellular glycoconjugates and tissue lectins like galectins has wide (patho)physiological significance. Their study is facilitated by nonhydrolysable derivatives of the natural O-glycans, such as S- and Se-glycosides. The latter enable extensive analyses by specific 77 Se NMR spectroscopy, but still remain underexplored. By using the example of selenodigalactoside (SeDG) and the human galectin-1 and -3, we have evaluated diverse 77 Se NMR detection methods and propose selective 1 H,77 Se heteronuclear Hartmann-Hahn transfer for efficient use in competitive NMR screening against a selenoglycoside spy ligand. By fluorescence anisotropy, circular dichroism, and isothermal titration calorimetry (ITC), we show that the affinity and thermodynamics of SeDG binding by galectins are similar to thiodigalactoside (TDG) and N-acetyllactosamine (LacNAc), confirming that Se substitution has no major impact. ITC data in D2 O versus H2 O are similar for TDG and LacNAc binding by both galectins, but a solvent effect, indicating solvent rearrangement at the binding site, is hinted at for SeDG and clearly observed for LacNAc dimers with extended chain length.

TF-containing MUC1 glycopeptides fail to entice Galectin-1 recognition of tumor-associated Thomsen-Freidenreich (TF) antigen (CD176) in solution.

Aberrant Mucin-1 (MUC1) glycosylation with the Thomsen-Friedenreich (TF) tumor-associated antigen (CD176) is a hallmark of epithelial carcinoma progression and poor patient prognosis. Recognition of TF by glycan-binding proteins, such as galectins, enables the pathological repercussions of this glycan presentation, yet the underlying binding specificities of different members of the galectin family is a matter of continual investigation. While Galectin-3 (Gal-3) recognition of TF has been well-documented at both the cellular and molecular level, Galectin-1 (Gal-1) recognition of TF has only truly been alluded to in cell-based platforms. Immunohistochemical analyses have purported Gal-1 binding to TF on MUC1 at the cell surface, however binding at the molecular level was inconclusive. We hypothesize that glycan scaffold (MUC1's tandem repeat peptide sequence) and/or multivalency play a role in the binding recognition of TF antigen by Gal-1. In this study we have developed a method for large-scale expression of Gal-1 and its histidine-tagged analog for use in binding studies by isothermal titration calorimetry (ITC) and development of an analytical method based on AlphaScreen technology to screen for Gal-1 inhibitors. Surprisingly, neither glycan scaffold or multivalent presentation of TF antigen on the scaffold was able to entice Gal-1 recognition to the level of affinity expected for functional significance. Future evaluations of the Gal-1/TF binding interaction in order to draw connections between immunohistochemical data and analytical measurements are warranted.

Transient climate and ambient health impacts due to national solid fuel cookstove emissions.

Residential solid fuel use contributes to degraded indoor and ambient air quality and may affect global surface temperature. However, the potential for national-scale cookstove intervention programs to mitigate the latter issues is not yet well known, owing to the spatial heterogeneity of aerosol emissions and impacts, along with coemitted species. Here we use a combination of atmospheric modeling, remote sensing, and adjoint sensitivity analysis to individually evaluate consequences of a 20-y linear phase-out of cookstove emissions in each country with greater than 5% of the population using solid fuel for cooking. Emissions reductions in China, India, and Ethiopia contribute to the largest global surface temperature change in 2050 [combined impact of -37 mK (11 mK to -85 mK)], whereas interventions in countries less commonly targeted for cookstove mitigation such as Azerbaijan, Ukraine, and Kazakhstan have the largest per cookstove climate benefits. Abatement in China, India, and Bangladesh contributes to the largest reduction of premature deaths from ambient air pollution, preventing 198,000 (102,000-204,000) of the 260,000 (137,000-268,000) global annual avoided deaths in 2050, whereas again emissions in Ukraine and Azerbaijan have the largest per cookstove impacts, along with Romania. Global cookstove emissions abatement results in an average surface temperature cooling of -77 mK (20 mK to -278 mK) in 2050, which increases to -118 mK (-11 mK to -335 mK) by 2100 due to delayed CO2 response. Health impacts owing to changes in ambient particulate matter with an aerodynamic diameter of 2.5 µm or less (PM2.5) amount to ∼22.5 million premature deaths prevented between 2000 and 2100.

How altering the modular architecture affects aspects of lectin activity: case study on human galectin-1.

Discoveries on involvement of glycan-protein recognition in many (patho)physiological processes are directing attention to exploring the significance of a fundamental structural aspect of sugar receptors beyond glycan specificity, i.e., occurrence of distinct types of modular architecture. In order to trace clues for defining design-functionality relationships in human lectins, a lectin's structural unit has been used as source material for engineering custom-made variants of the wild-type protein. Their availability facilitates comparative analysis toward the stated aim. With adhesion/growth-regulatory human galectin-1 as example, the strategy of evaluating how changes of its design (here, from the homodimer of non-covalently associated domains to (i) linker-connected di- and tetramers and (ii) a galectin-3-like protein) affect activity is illustrated by using three assay systems of increasing degree of glycan complexity. Whereas calorimetry with two cognate disaccharides and array testing with 647 (glyco)compounds disclosed no major changes, galectin histochemical staining profiles of tissue sections that present natural glycome complexity revealed differences between wild-type and linker-connected homo-oligomers as well as between the galectin-3-like variant and wild-type galectin-3 for cell-type positivity, level of intensity at the same site and susceptibility for inhibition by a bivalent glycocompound. These results underscore the strength of the documented approach. Moreover, they give direction to proceed to (i) extending its application to other members of this lectin family, especially galectin-3 and (ii) then analyzing impact of architectural alterations on cell surface lattice formation and ensuing biosignaling systematically, considering the variants' potential for translational medicine.

Bone loss at the distal femur and proximal tibia in persons with spinal cord injury: imaging approaches, risk of fracture, and potential treatment options.

Persons with spinal cord injury (SCI) undergo immediate unloading of the skeleton and, as a result, have severe bone loss below the level of lesion associated with increased risk of long-bone fractures. The pattern of bone loss in individuals with SCI differs from other forms of secondary osteoporosis because the skeleton above the level of lesion remains unaffected, while marked bone loss occurs in the regions of neurological impairment. Striking demineralization of the trabecular epiphyses of the distal femur (supracondylar) and proximal tibia occurs, with the knee region being highly vulnerable to fracture because many accidents occur while sitting in a wheelchair, making the knee region the first point of contact to any applied force. To quantify bone mineral density (BMD) at the knee, dual energy x-ray absorptiometry (DXA) and/or computed tomography (CT) bone densitometry are routinely employed in the clinical and research settings. A detailed review of imaging methods to acquire and quantify BMD at the distal femur and proximal tibia has not been performed to date but, if available, would serve as a reference for clinicians and researchers. This article will discuss the risk of fracture at the knee in persons with SCI, imaging methods to acquire and quantify BMD at the distal femur and proximal tibia, and treatment options available for prophylaxis against or reversal of osteoporosis in individuals with SCI.

Improving present day and future estimates of anthropogenic sectoral emissions and the resulting air quality impacts in Africa.

The African continent is undergoing immense social and economic change, particularly regarding population growth and urbanization, where the urban population in Africa is anticipated to increase by a factor of 3 over the next 40 years. To understand the potential health impacts from this demographical shift and design efficient emission mitigation strategies, we used improved Africa-specific emissions that account for inefficient combustion sources for a number of sectors such as transportation, household energy generation, waste burning, and home heating and cooking. When these underrepresented emissions sources are combined with the current estimates of emissions in Africa, ambient particulate matter concentrations from present-day anthropogenic activity contribute to 13 210 annual premature deaths, with the largest contributions (38%) coming from residential emissions. By scaling both the population and the emissions for projected national-scale levels of growth, the predicted health impact grows to approximately 78 986 annual premature deaths by 2030 with 45% now resulting from emissions related to energy combustion. In order to mitigate this resulting increase in premature deaths, three scenarios have been developed which reduce sector-specific future emissions based on prior targets for technological improvements and emission controls in transportation, energy production and residential activities. These targeted potential mitigation strategies can avoid up to 37% of the estimated annual premature deaths by 2030 with the largest opportunity being a reduction of 10 868 annual deaths from switching half of the energy generation in South Africa to renewable technologies.

Vegetation dynamics and rainfall sensitivity of the Amazon.

We show that the vegetation canopy of the Amazon rainforest is highly sensitive to changes in precipitation patterns and that reduction in rainfall since 2000 has diminished vegetation greenness across large parts of Amazonia. Large-scale directional declines in vegetation greenness may indicate decreases in carbon uptake and substantial changes in the energy balance of the Amazon. We use improved estimates of surface reflectance from satellite data to show a close link between reductions in annual precipitation, El Niño southern oscillation events, and photosynthetic activity across tropical and subtropical Amazonia. We report that, since the year 2000, precipitation has declined across 69% of the tropical evergreen forest (5.4 million km(2)) and across 80% of the subtropical grasslands (3.3 million km(2)). These reductions, which coincided with a decline in terrestrial water storage, account for about 55% of a satellite-observed widespread decline in the normalized difference vegetation index (NDVI). During El Niño events, NDVI was reduced about 16.6% across an area of up to 1.6 million km(2) compared with average conditions. Several global circulation models suggest that a rise in equatorial sea surface temperature and related displacement of the intertropical convergence zone could lead to considerable drying of tropical forests in the 21st century. Our results provide evidence that persistent drying could degrade Amazonian forest canopies, which would have cascading effects on global carbon and climate dynamics.

Cryptococcus gattii infection in solid organ transplant recipients: description of Oregon outbreak cases.

Cryptococcus gattii was recognized as an emerging infection in the Pacific Northwest in 2004. Out of 62 total infections in Oregon since the outbreak, 11 were in solid organ transplant (SOT) recipients. SOT recipients were more likely to have disseminated disease and higher mortality than normal hosts, who mostly had isolated mass lesions. The median time from transplantation to C. gattii diagnosis was 17.8 months. The primary sites of infection were lung (n = 4), central nervous system (n = 3), or both (n = 4). The Oregon-endemic strain, VGII (subtypes IIa and IIc) was present in 10 of 11 patients; the median fluconazole minimum inhibitory concentration (MIC) was 12 µg/mL (range 2-32 µg/mL) for this strain. We found C. gattii infection among organ transplant recipients was disseminated at diagnosis, had low cerebrospinal fluid cryptococcal antigen titers, and was associated with an elevated fluconazole MIC and high attributable mortality.

Reliability of surface EMG as an assessment tool for trunk activity and potential to determine neurorecovery in SCI.

STUDY DESIGN: Reliability and validity study. OBJECTIVE: This study investigates the responsiveness and reliability of the brain motor control assessment (BMCA) as a standardized neurophysiological assessment tool to: (i) characterize trunk neural activity in neurologically-intact controls; (ii) measure and quantify neurorecovery of trunk after spinal cord injury (SCI). SETTING: Kessler Foundation Research Center, West Orange, NJ. METHODS: A standardized BMCA protocol was performed to measure surface electromyography (sEMG) recordings for seven bilateral trunk muscles on 15 able-bodied controls during six maneuvers (inhalation, exhalation, neck flexion, jendrassik, unilateral grip). Additionally, sEMG recordings were analyzed for one chronic SCI individual before electrical stimulation (ES), after ES of the lower extremities while supine, and after active stand training using body-weight support with bilateral ES. sEMG recordings were collected on bilateral erector spinae, internal and external obliques, upper and middle trapezius, biceps and triceps. For each maneuver a voluntary response index was calculated: incorporating the magnitude of sEMG signal and a similarity index (SI), which quantifies the distribution of activity across all muscles. RESULTS: Among all maneuvers, the SI presented reproducible assessment of trunk-motor function within (ICC: 0.860-0.997) and among (P⩾0.22) able-bodied individuals. In addition, potential changes were measured in a chronic SCI individual after undergoing two intensive ES protocols. CONCLUSION: The BMCA provides reproducible characterization of trunk activity in able-bodied individuals, lending credence for its use in neurophysiological assessment of motor control. Additionally, the BMCA as an assessment tool to measure neurorecovery in an individual with chronic SCI after intense ES interventions was demonstrated.

Improved Gait symmetry with spinal cord transcutaneous stimulation in individuals with spinal cord injury.

A previous study by our group showed preliminary results showcasing the usage of optimal and individualized spinal cord transcutaneous stimulation (scTS) parameters during overground gait training to facilitate more repeatable gait kinematics profiles for one participant with an incomplete spinal cord injury (SCI). The goal of this study was to use the combined targeted scTS and training to improve spatial and temporal parameters to achieve a symmetrical gait profile after completing activity-based training (ABT) (stepping and stand training) and exoskeleton training with and without scTS. Our data indicates that stimulation with optimal and individualized parameters can lead to more effective, stable and symmetric gait patterns in participants with varied levels of SCI.Clinical Relevance- This analysis will enable us to better understand the combined effect of exercise based training and tonic scTS on improving the quality and symmetry of gait pattern in participants with incomplete SCI.

Differential Corticospinal Excitability and Cortical Functional Connectivity Modulation by Spinal Cord Transcutaneous Stimulation-based Motor Training versus Motor Training alone in Able-bodied and SCI participants: A Multiple Case Study.

Spinal cord transcutaneous stimulation (scTS) has shown its potential for boosting motor, sensory, and autonomic function recovery after a spinal cord injury. Despite the demonstrated benefits, little is known about the exact neuromodulatory mechanisms triggered by scTS and the cortex involvement in the beneficial effects observed. Here, we examine the effects of scTS-based motor training and motor training alone on sensorimotor cortical functional connectivity and corticospinal excitability in able-bodied and SCI participants.Clinical Relevance- The results show preliminary evidence of differential cortical involvement and modulation by scTS-based motor training in uninjured and spinal-cord injured individuals. A better understanding of scTS mechanisms of action could help optimize the intervention design and potentiate its benefits.

GaAs-based nanoneedle light emitting diode and avalanche photodiode monolithically integrated on a silicon substrate.

Monolithic integration of III-V compound semiconductor devices with silicon CMOS integrated circuits has been hindered by large lattice mismatches and incompatible processing due to high III-V epitaxy temperatures. We report the first GaAs-based avalanche photodiodes (APDs) and light emitting diodes, directly grown on silicon at a very low, CMOS-compatible temperature and fabricated using conventional microfabrication techniques. The APDs exhibit an extraordinarily large multiplication factor at low voltage resulting from the unique needle shape and growth mode.

Exploring the Galectin Network by Light and Fluorescence Microscopy.

Dynamic changes of a cell's glycophenotype are increasingly interpreted as shifts in the capacity to interact with tissue (endogenous) lectins. The status of glycan branching or chain length (e.g., core 1 vs core 2 mucin-type O-glycans and polyLacNAc additions) as well as of sialylation/sulfation has been delineated to convey signals. They are "read" by galectins, for example regulating lattice formation on the membrane and cell growth. Owing to the discovery of the possibility that these effectors act in networks physiologically resulting in functional antagonism or cooperation, their detection and distribution profiling need to be expanded from an individual (single) protein to the-at best-entire family. How to work with non-cross-reactive antibodies and with the labeled tissue-derived proteins (used as probes) is exemplarily documented for chicken and human galectins including typical activity and specificity controls. This description intends to inspire the systematic (network) study of members of a lectin family and also the application of tissue proteins beyond a single lectin category in lectin histochemistry.

Evaluating the Impact of Chemical Complexity and Horizontal Resolution on Tropospheric Ozone Over the Conterminous US With a Global Variable Resolution Chemistry Model.

A new configuration of the Community Earth System Model (CESM)/Community Atmosphere Model with full chemistry (CAM-chem) supporting the capability of horizontal mesh refinement through the use of the spectral element (SE) dynamical core is developed and called CESM/CAM-chem-SE. Horizontal mesh refinement in CESM/CAM-chem-SE is unique and novel in that pollutants such as ozone are accurately represented at human exposure relevant scales while also directly including global feedbacks. CESM/CAM-chem-SE with mesh refinement down to ∼14 km over the conterminous US (CONUS) is the beginning of the Multi-Scale Infrastructure for Chemistry and Aerosols (MUSICAv0). Here, MUSICAv0 is evaluated and used to better understand how horizontal resolution and chemical complexity impact ozone and ozone precursors over CONUS as compared to measurements from five aircraft campaigns, which occurred in 2013. This field campaign analysis demonstrates the importance of using finer horizontal resolution to accurately simulate ozone precursors such as nitrogen oxides and carbon monoxide. In general, the impact of using more complex chemistry on ozone and other oxidation products is more pronounced when using finer horizontal resolution where a larger number of chemical regimes are resolved. Large model biases for ozone near the surface remain in the Southeast US as compared to the aircraft observations even with updated chemistry and finer horizontal resolution. This suggests a need for adding the capability of replacing sections of global emission inventories with regional inventories, increasing the vertical resolution in the planetary boundary layer, and reducing model biases in meteorological variables such as temperature and clouds.

Tracking plant physiological properties from multi-angular tower-based remote sensing.

Imaging spectroscopy is a powerful technique for monitoring the biochemical constituents of vegetation and is critical for understanding the fluxes of carbon and water between the land surface and the atmosphere. However, spectral observations are subject to the sun-observer geometry and canopy structure which impose confounding effects on spectral estimates of leaf pigments. For instance, the sun-observer geometry influences the spectral brightness measured by the sensor. Likewise, when considering pigment distribution at the stand level scale, the pigment content observed from single view angles may not necessarily be representative of stand-level conditions as some constituents vary as a function of the degree of leaf illumination and are therefore not isotropic. As an alternative to mono-angle observations, multi-angular remote sensing can describe the anisotropy of surface reflectance and yield accurate information on canopy structure. These observations can also be used to describe the bi-directional reflectance distribution which then allows the modeling of reflectance independently of the observation geometry. In this paper, we demonstrate a method for estimating pigment contents of chlorophyll and carotenoids continuously over a year from tower-based, multi-angular spectro-radiometer observations. Estimates of chlorophyll and carotenoid content were derived at two flux-tower sites in western Canada. Pigment contents derived from inversion of a CR model (PROSAIL) compared well to those estimated using a semi-analytical approach (r(2) = 0.90 and r(2) = 0.69, P < 0.05 for both sites, respectively). Analysis of the seasonal dynamics indicated that net ecosystem productivity was strongly related to total canopy chlorophyll content at the deciduous site (r(2) = 0.70, P < 0.001), but not at the coniferous site. Similarly, spectral estimates of photosynthetic light-use efficiency showed strong seasonal patterns in the deciduous stand, but not in conifers. We conclude that multi-angular, spectral observations can play a key role in explaining seasonal dynamics of fluxes of carbon and water and provide a valuable addition to flux-tower-based networks.

Theoretical analysis of subwavelength high contrast grating reflectors.

A simple analytic analysis of the ultra-high reflectivity feature of subwavelength dielectric gratings is developed. The phenomenon of ultra high reflectivity is explained to be a destructive interference effect between the two grating modes. Based on this phenomenon, a design algorithm for broadband grating mirrors is suggested.