Functional lung imaging with synchrotron radiation: Methods and preclinical applications

Many lung disease processes are characterized by structural and functional heterogeneity that is not directly appreciable with traditional physiological measurements. Experimental methods and lung function modeling to study regional lung function are crucial for better understanding of disease mechanisms and for targeting treatment. Synchrotron radiation offers useful properties to this end: coherence, utilized in phase-contrast imaging, and high flux and a wide energy spectrum which allow the selection of very narrow energy bands of radiation, thus allowing imaging at very specific energies. K-edge subtraction imaging (KES) has thus been developed at synchrotrons for both human and small animal imaging. The unique properties of synchrotron radiation extend X-ray computed tomography (CT) capabilities to quantitatively assess lung morphology, and also to map regional lung ventilation, perfusion, inflammation and biomechanical properties, with microscopic spatial resolution. Four-dimensional imaging, allows the investigation of the dynamics of regional lung functional parameters simultaneously with structural deformation of the lung as a function of time. This review summarizes synchrotron radiation imaging methods and overviews examples of its application in the study of disease mechanisms in preclinical animal models, as well as the potential for clinical translation both through the knowledge gained using these techniques and transfer of imaging technology to laboratory X-ray sources.     

Orchid seed sensitivity to nitrate reflects habitat preferences and soil nitrate content

• Orchids are distributed around the world, however, the factors shaping their specific distribution and habitat preferences are largely unknown. Moreover, many orchids are at risk of becoming threatened as landscapes change, sometimes declining without apparent reason. One important factor affecting plant distribution is nutrient levels in the environment. Nitrates can inhibit not only orchid growth and persistence, but also seed germination.
• We used in vitro axenic cultures to exactly determine the germination sensitivity of seven orchid species to nitrates and correlated this with soil properties of the natural sites and with the species’ habitat preferences.
• We found high variation in response to nitrate between species. Orchids from oligotrophic habitats were highly sensitive, while orchids from more eutrophic habitats were almost insensitive. Sensitivity to nitrate was also associated with soil parameters that indicated a higher nitrification rate.
• Our results indicate that nitrate can affect orchid distribution via direct inhibition of seed germination. Nitrate levels in soils are increasing rapidly due to intensification of agricultural processes and concurrent soil pollution, and we propose this increase could cause a decline in some orchid species.

     

Single pot sequential crystallization-distillation as a new purification procedure

A new purification procedure exploiting the simultaneous presence of a solid, liquid, and gas phase in a low surface area system is proposed and discussed. The assumptions of vanishingly low diffusion coefficients in the solid phase and that of the presence of a single “effective impurity” allow to plan the sequence of operations starting from the knowledge of just the melting and boiling points of the substance to be purified and of those of the “effective impurity”. Examples and results are presented.     

A linkage group on pig chromosome 4 comprising the loci for blood group L, GBA, ATP1B1 and three microsatellites

Restriction fragment length polymorphisms (RFLPs) were described for the porcine loci for β-glucosidase (GBA) and the β-polypeptide 1 of the Na⁺, K⁺-transporting ATPase (ATP1B1). Linkage analyses using a three-generation pedigree provided evidence for the assignment of ATP1B1, GBA and two microsatellite loci (S0001 and S0067) to a previously described linkage group comprising the loci for blood group L (EAL) and an anonymous microsatellite (S0097). The linear order of the six markers was determined with confidence by multipoint analyses and the length of the linkage group was estimated at 88 CM. This linkage group was assigned to pig chromosome 4 on the basis of a previous physical localization of the ATP1B1 gene. In situ hybridization data for S0001 presented in this study were consistent with a localization on chromosome 4 and suggested a regional localization to 4pl2-pl3. The present study reveals conflicting data concerning the genetic localization of the K88 loci controlling the expression of the receptors for the E. coli pilus antigens. One group has reported data suggesting a loose linkage between K88 and EAL, now mapped to chromosome 4, whereas two other groups have found linkage between K88 and the transferrin locus (TF), mapped to chromosome 13 by in situ hybridization.     

In vitro reactivation of sarin-inhibited brain acetylcholinesterase from different species by various oximes

In vitro as well as in vivo evaluation of the reactivating efficacy of various oximes against nerve agent-inhibited acetylcholinesterase has been usually done with the help of animal experiments. Nevertheless, previously published data indicate that the reactivation potency of oximes may be different in human and animal species, which may hamper the extrapolation of animal data to human data. Therefore, to better evaluate the efficacy of various oximes (pralidoxime, obidoxime, HI-6, K033) to reactivate brain acetylcholinesterase inhibited by sarin by in vitro methods, human, rat and pig brain acetylcholinesterase were used to calculate kinetic parameters for the reactivation. Our results show differences among the species, depending on the type of oxime, and indicate that data from animal experiments needs to be carefully evaluated before extrapolation to humans.     

Prenatal diagnosis of Pallister-Killian syndrome and literature review

Pallister-Killian syndrome (PKS) is a rare sporadic genetic disorder usually caused by mosaicism of an extra isochromosome of 12p (i(12p)). This retrospective study analysed the prenatal ultrasound manifestations and molecular and cytogenetic results of five PKS foetuses. Samples of amniotic fluid and/or cord blood, skin biopsy and placenta were collected. Conventional karyotyping and single nucleotide polymorphism array (SNP array) were performed on all the amniotic fluid or cord blood samples. Copy number variants sequencing (CNV-seq) and fluorescence in situ hybridization (FISH) were also used for the validation for one foetus. All the five foetuses were from pregnancies with advanced parental age. Two foetuses involved structural abnormalities and one foetus had only soft markers, all of which included increased nuchal translucency. The rest two foetuses had normal ultrasounds in the second trimester, which has rarely been reported before. The karyotype revealed typical i(12p) in four cases and a small supernumerary marker chromosome consisting of 12p and 20p in the remaining one case. The proportion of cells with i(12p) ranged from 0 to 100% in cultural cells, while SNP array results suggested 2−4 copies of 12p. For one foetus, metaphase FISH showed normal results, but the interphase FISH suggested cell lines with two, three and four copies of 12p in the amniotic fluid. Advanced parental age may be an important risk factor for PKS, and there were no typical ultrasound manifestations related to PKS. A combination of karyotype analysis and molecular diagnosis is an effective method for the diagnosis of PKS.     

Comparison of Southern Appalachian high-elevation outcrop plant communities with their Northern Appalachian counterparts

Abstract. Southern Appalachian high-elevation outcrops harbour six regionally rare Northern Appalachian taxa usually considered relicts of a Pleistocene alpine flora. For five of the six taxa, minimum elevation in the south was 367–1113 m higher than in the north. While habitats compared between the two regions share only 9% of their total flora, individual plots had up to 70% of their species occurring in the opposite region. The northern affinity of southern outcrops increased with elevation, slope steepness, soil Cu, B and SO₄ and decreased with potential solar radiation and soil Na. As a result, communities above 1600 m on felsic bedrock, and above 1350 m on mafic bedrock, were most northern in composition. Northern affinity of southern outcrops also increased with latitude, which may partly result from closer geographic proximity to past communities that provided progenitors for the current northern flora. Northern treeless habitats increased in southern affinity with increased slope steepness, perennial seepage, vegetation height, shade, soil pH, Al, Mn, Na and decreased elevation and organic matter. As a result, northern outcrop communities below treeline were most similar to those on southern outcrops. This suggests that southern outcrop vegetation may be more similar to Pleistocene outcrop vegetation than to Pleistocene alpine vegetation. Partial constrained ordination showed that while compositional differences between the Northern and Southern Appalachian habitats were largely explained by environmental differences, there was a significant component of residual variation explained by north or south position that was unrelated to environment. These residual compositional differences may result from historical influences on community structure involving stochastic extinction and colonization processes.     

COMPARISON OF VERTICAL AERIAL PHOTOGRAPHIC AND GROUND CENSUSES OF STELLER SEA LIONS AT AÑO NUEVO ISLAND, JULY 1990-1993

Counts of Steller sea lion ( Eumetopias jubatus ) pups and non-pups (adults and juveniles) from aerial photographs of rookeries at Año Nuevo Island between 1990 and 1993 were significantly higher than those made on the ground. Based on regression of natural logs of photographic counts versus year, the number of pups declined at a rate of −0.099yr while non-pup numbers declined at −0.315/yr. Examination of ground count data for the same period revealed a significant decline in non-pups (−0.139/yr), but no trend was detected in the ground counts of pups. The regression coefficients from photographic and ground counts of non-pups did not differ significantly. Power analyses using the program TRENDS indicated that detectable rates of change in abundance from four annual surveys were much lower for counts of pups than counts of non-pups where sampling precision was based on fits to linear models.     

High frequency residues: A new set of signals for detectability studies of an X-ray imaging system

A new set of signals for studying detectability of an X-ray imaging system is presented. The results obtained with these signals are intended to complement the NEQ results.The signals are generated from line spread profiles by progressively removing their lower frequency components and the resulting high frequency residues (HFRs) form the set of signals to be used in detectability studies. Detectability indexes for these HFRs are obtained using a non-prewhitening (NPW) observer and a series of edge images are used to obtain the HFRs, the covariance matrices required by the NPW model and the MTF and NPS used in NEQ calculations. The template used in the model is obtained by simulating the processes of blurring and sampling of the edge images. Comparison between detectability indexes for the HFRs and NEQ are carried out for different acquisition techniques using different beam qualities and doses.The relative sensitivity shown by detectability indexes using HFRs is higher than that of NEQ, especially at lower doses. Also, the different observers produce different results at high doses: while the ideal Bayesian observer used by NEQ distinguishes between beam qualities, the NPW used with the HFRs produces no differences between them.Delta functions used in HFR are the opposite of complex exponential functions in terms of their support in the spatial and frequency domains. Since NEQ can be interpreted as detectability of these complex exponential functions, detectability of HFRs is presented as a natural complement to NEQ in the performance assessment of an imaging system.     

Quantifying Single Microvessel Permeability in Isolated Blood-perfused Rat Lung Preparation

The isolated blood-perfused lung preparation is widely used to visualize and define signaling in single microvessels. By coupling this preparation with real time imaging, it becomes feasible to determine permeability changes in individual pulmonary microvessels. Herein we describe steps to isolate rat lungs and perfuse them with autologous blood. Then, we outline steps to infuse fluorophores or agents via a microcatheter into a small lung region. Using these procedures described, we determined permeability increases in rat lung microvessels in response to infusions of bacterial lipopolysaccharide. The data revealed that lipopolysaccharide increased fluid leak across both venular and capillary microvessel segments. Thus, this method makes it possible to compare permeability responses among vascular segments and thus, define any heterogeneity in the response. While commonly used methods to define lung permeability require postprocessing of lung tissue samples, the use of real time imaging obviates this requirement as evident from the present method. Thus, the isolated lung preparation combined with real time imaging offers several advantages over traditional methods to determine lung microvascular permeability, yet is a straightforward method to develop and implement.