Multi‐centre reproducibility of diffusion MRI parameters for clinical sequences in the brain

The purpose of this work was to assess the reproducibility of diffusion imaging, and in particular the apparent diffusion coefficient (ADC), intra‐voxel incoherent motion (IVIM) parameters and diffusion tensor imaging (DTI) parameters, across multiple centres using clinically available protocols with limited harmonization between sequences. An ice–water phantom and nine healthy volunteers were scanned across fives centres on eight scanners (four Siemens 1.5T, four Philips 3T). The mean ADC, IVIM parameters (diffusion coefficient D and perfusion fraction f) and DTI parameters (mean diffusivity MD and fractional anisotropy FA), were measured in grey matter, white matter and specific brain sub‐regions. A mixed effect model was used to measure the intra‐ and inter‐scanner coefficient of variation (CV) for each of the five parameters. ADC, D, MD and FA had a good intra‐ and inter‐scanner reproducibility in both grey and white matter, with a CV ranging between 1% and 7.4%; mean 2.6%. Other brain regions also showed high levels of reproducibility except for small structures such as the choroid plexus. The IVIM parameter f had a higher intra‐scanner CV of 8.4% and inter‐scanner CV of 24.8%. No major difference in the inter‐scanner CV for ADC, D, MD and FA was observed when analysing the 1.5T and 3T scanners separately. ADC, D, MD and FA all showed good intra‐scanner reproducibility, with the inter‐scanner reproducibility being comparable or faring slightly worse, suggesting that using data from multiple scanners does not have an adverse effect compared with using data from the same scanner. The IVIM parameter f had a poorer inter‐scanner CV when scanners of different field strengths were combined, and the parameter was also affected by the scan acquisition resolution. This study shows that the majority of diffusion MRI derived parameters are robust across 1.5T and 3T scanners and suitable for use in multi‐centre clinical studies and trials. © 2015 The Authors NMR in Biomedicine Published by John Wiley & Sons Ltd.     

用正糖钳夹试验评估基础胰岛素类似物作用特点的研究

健康人通过体内胰岛素的分泌调节，可以保持正常的血糖水平。各种外源性胰岛素制剂在代谢过程上总是尽可能地模拟内源性胰岛素的分泌动力学。理想的外源性基础胰岛素制剂可以模拟健康人的基础胰岛素分泌，以致使用者可以恢复两餐之间和夜间正常生理情况下的血浆胰岛素水平。     

Systemic hematologic effects of PEG-rHuMGDF-induced megakaryocyte hyperplasia in mice

PEG-rHuMGDF injected daily in normal mice causes a rapid dose-dependent increase in megakaryocytes and platelets. At the same time that platelet numbers are increased, the mean platelet volume (MPV) and platelet distribution width (PDW) can be either decreased, normal, or increased depending on the dose and time after administration. Thus, PEG-rHuMGDF at a low dose causes decreases in MPV and PDW, MGDF at an intermediate dose causes an initial increase followed by a decrease in MPV and PDW, and PEG-rHuMGDF at higher doses causes an increase in MPV and PDW followed by a gradual normalization of these platelet indices. In addition to the expected thrombocytosis after 7 to 10 days of daily injection of high doses of PEG-rHuMGDF, a transient decrease in peripheral red blood cell numbers and hemoglobin is noted accompanied in the bone marrow by megakaryocytic hyperplasia, myeloid hyperplasia, erythroid and lymphoid hypoplasia, and deposition of a fine network of reticulin fibers. Splenomegaly, an increase in splenic megakaryocytes, and extramedullary hematopoiesis accompany the hematologic changes in the peripheral blood and marrow to complete a spectrum of pathologic features similar to those reported in patients with myelofibrosis and megakaryocyte hyperplasia. However, all the PEG-rHuMGDF-initiated hematopathology including the increase in marrow reticulin is completely and rapidly reversible upon the cessation of administration of PEG-rHuMGDF. Thus, transient hyperplastic proliferation of megakaryocytes does not cause irreversible tissue injury. Furthermore, PEG-rHuMGDF completely ameliorates carboplatin-induced thrombocytopenia at a low-dose that does not cause the hematopathology associated with myelofibrosis.     

Clinical anatomy of vertebrae in scoliosis: global analysis in four different diseases by multiplanar reconstructive computed tomography

Background contextFew accurate analyses of clinically useful vertebral anatomy have been conducted, and most have focused on thoracic idiopathic scoliosis.PurposeTo evaluate the different anatomic characteristics in scoliosis by disease type and level.Study designObservational cohort study.Patient sampleForty-eight patients with scoliosis were included in this study.Outcome measuresSubjects underwent computed tomography (CT) of the whole spine.MethodsForty-eight patients with scoliosis were included in this study: 15 adolescent idiopathic, 11 cerebral palsy (CP), 10 muscular dystrophy (MD), and 12 congenital (CG) scoliosis patients with similar demographics. Subjects underwent CT of the whole spine, preoperatively. Eight anatomic parameters were measured in multiplanar reconstructive CT images, and statistical analysis was performed to investigate differences.ResultsIn general, values in the anatomic parameters were similar for the four diseases. Each parameter showed the unique change pattern according to the spinal level regardless of curvature shape, direction, or magnitude. In particular, chord length (CL) in MD and CG scoliosis was lower than in adolescent idiopathic scoliosis (AIS) and CP, and pedicle rib unit length was lower in CG scoliosis than in the other diseases (p<.05). Comparisons of convex and concave anatomies in AIS showed that inner pedicle width (PWI) and outer pedicle width (PWO) were wider for convex side, CL, pedicle width, and transverse pedicle angle were greater for concave side (p<.05), and differences were more significant at apices. However, in CP, PWI and PWO were similar between convex and concaves sides (p>.05). Although PWI and PWO were wider for convex sides and CL and pedicle length were greater for concave sides in MD (p<.05), differences were less significant at apices. Particularly, CG scoliosis showed severely deformed anatomy, with differences of seven parameters at apical vertebrae (p<.05).ConclusionClinical anatomies of vertebrae in scoliosis were found to differ significantly at different levels and in terms of convexity and disease type.     

Urea unfolding of peptide helices as a model for interpreting protein unfolding.

To provide a model system for understanding how the unfolding of protein alpha-helices by urea contributes to protein denaturation, urea unfolding was measured for a homologous series of helical peptides with the repeating sequence Ala-Glu-Ala-Ala-Lys-Ala and chain lengths varying from 14 to 50 residues. The dependence of the helix propagation parameter of the Zimm-Bragg model for helix-coil transition theory (s) on urea molarity ([urea]) was determined at 0 degree C with data for the entire set of peptides, and a linear dependence of In s on [urea] was found. The results were fitted by the binding-site model and by the solvent-exchange model for the interaction of urea with the peptides. Each of these thermodynamic models is able to describe the data quite well and we are not able to discern any difference between the ability of each model to fit the data. Thus a linear relation, ln s = ln s0 - (m/RT).[urea], fits the data for alpha-helix unfolding, just as others have found for protein unfolding. When the m value determined here for alpha-helix unfolding is multiplied by the number of helical residues in partly helical protein molecules, the resulting values agree within a factor of 2 with observed m values for these proteins. This result indicates that the interaction between urea and peptide groups accounts for a major part of the denaturing action of urea on proteins, as predicted earlier by some model studies with small molecules.     

An experimentally determined protein folding energy landscape

Energy landscapes have been used to conceptually describe and model protein folding but have been difficult to measure experimentally, in large part because of the myriad of partly folded protein conformations that cannot be isolated and thermodynamically characterized. Here we experimentally determine a detailed energy landscape for protein folding. We generated a series of overlapping constructs containing subsets of the seven ankyrin repeats of the Drosophila Notch receptor, a protein domain whose linear arrangement of modular structural units can be fragmented without disrupting structure. To a good approximation, stabilities of each construct can be described as a sum of energy terms associated with each repeat. The magnitude of each energy term indicates that each repeat is intrinsically unstable but is strongly stabilized by interactions with its nearest neighbors. These linear energy terms define an equilibrium free energy landscape, which shows an early free energy barrier and suggests preferred low-energy routes for folding.     

Medial patellofemoral ligament reconstruction in patellar instability

Background:

Medial patellofemoral ligament (MPFL) is one of the major static medial stabilising structures of the patella. MPFL is most often damaged in patients with patellar instability. Reconstruction of MPFL is becoming a common surgical procedure in treating patellar instability. We hypothesised that MPFL reconstruction was adequate to treat patients with patellar instability if the tibial tubercle and the centre of the trochlear groove (TT-TG) value was less than 20 mm and without a dysplastic trochlea.

Materials and Methods:

30 patients matching our inclusion criteria and operated between April 2009 and May 2011 were included in the study. MPFL reconstruction was performed using gracilis tendon fixed with endobutton on the patellar side and bio absorbable interference screw or staple on the femoral side. Patients were followed up with subjective criteria, Kujala score and Lysholm score.

Results:

The mean duration of followup was 25 months (range 14-38 months). The mean preoperative Kujala score was 47.5 and Lysholm score was 44.7. The mean postoperative Kujala score was 87 and Lysholm score was 88.06. None of the patients had redislocation.

Conclusion:

MPFL reconstruction using gracilis tendon gives excellent results in patients with patellar instability with no redislocations. Some patients may have persistence of apprehension.     

From the Cover: Recursive genomewide recombination and sequencing reveals a key refinement step in the evolution of a metabolic innovation in Escherichia coli

Evolutionary innovations often arise from complex genetic and ecological interactions, which can make it challenging to understand retrospectively how a novel trait arose. In a long-term experiment, Escherichia coli gained the ability to use abundant citrate (Cit⁺) in the growth medium after ∼31,500 generations of evolution. Exploiting this previously untapped resource was highly beneficial: later Cit⁺ variants achieve a much higher population density in this environment. All Cit⁺ individuals share a mutation that activates aerobic expression of the citT citrate transporter, but this mutation confers only an extremely weak Cit⁺ phenotype on its own. To determine which of the other >70 mutations in early Cit⁺ clones were needed to take full advantage of citrate, we developed a recursive genomewide recombination and sequencing method (REGRES) and performed genetic backcrosses to purge mutations not required for Cit⁺ from an evolved strain. We discovered a mutation that increased expression of the dctA C₄-dicarboxylate transporter greatly enhanced the Cit⁺ phenotype after it evolved. Surprisingly, strains containing just the citT and dctA mutations fully use citrate, indicating that earlier mutations thought to have potentiated the initial evolution of Cit⁺ are not required for expression of the refined version of this trait. Instead, this metabolic innovation may be contingent on a genetic background, and possibly ecological context, that enabled citT mutants to persist among competitors long enough to obtain dctA or equivalent mutations that conferred an overwhelming advantage. More generally, refinement of an emergent trait from a rudimentary form may be crucial to its evolutionary success.Key innovations in the history of life are often caused by the acquisition of a qualitatively new trait that is “an evolutionary novelty which allows the exploitation of new resources or habitats and thus triggers an adaptive radiation” (1). Such innovations are typically rare and difficult to predict because they result from complex nonadditive (i.e., epistatic) genetic interactions or ecological interactions, within or between species, that develop only over the course of long evolutionary trajectories (2). Evolution of a new trait can be conceptually divided into three steps: potentiation, actualization, and refinement (3). First, one or more potentiating events may be necessary to generate a genetic background or environmental conditions that make a new trait accessible to evolution. Genetic potentiation, for example, may involve a period of nonadaptive genetic drift wherein a phenotype stays constant or the accumulation of mutations that are immediately advantageous for reasons unrelated to the new trait (4, 5). Then, a keystone actualizing mutation or environmental shift may lead to expression of the new trait, possibly by coopting latent changes in a cellular network or physical structure for a new use (6–8). Finally, there may be many subsequent opportunities for further refinement mutations that improve an emergent trait so that a newly colonized niche can be fully exploited (1).The appearance of citrate utilization in a >25-y long-term evolution experiment (LTEE) with Escherichia coli provides an opportunity to study a deep historical record leading to a key metabolic innovation (3, 9, 10). E. coli cannot ordinarily grow on citrate as a sole carbon source under aerobic conditions (11, 12), a phenotype that has been used to define it as a species (13). Just 1 of 12 replicate LTEE populations gained the ability to aerobically use citrate (Cit⁺), and this rare innovation happened only after ∼31,500 generations of growth in glucose-limited media, despite the presence of an excess of citrate as an untapped carbon source all along (10). The actualizing event for the Cit⁺ trait is known: tandem amplifications of a chromosomal region that place a copy of an aerobically active promoter upstream of a citrate transporter (citT) are present in all Cit⁺ isolates from this population (3).However, the Cit⁺ trait was surprisingly weak when it first appeared. The earliest individuals with the citT mutation exhibit little or no growth on citrate as a sole carbon source and appear to have derived only a small benefit from this mutation under the conditions of the LTEE (3). In fact, a majority of the population remained Cit^– for at least 1,500 generations (225 d) after the citT mutation evolved, and these initial Cit⁺ individuals were only detected retrospectively in historical samples of the population by using a sensitive indicator agar test for citrate utilization and allowing days to weeks for a positive result (10). Adding a high-copy plasmid with a module containing the new promoter configuration and citT gene to the ancestral strain of the LTEE leads to a phenotype similar to that of the early Cit⁺ clones, indicating that this mutation is, at least qualitatively, sufficient on its own for this rudimentary version of the new trait (3).Shortly after ∼33,000 generations, this LTEE population experienced a massive increase in the final cell density it reached at the end of each daily growth cycle (10). This population expansion was due to the evolution of new Cit⁺ variants that fully use the abundant citrate in the media after glucose depletion. We call this strong phenotype Cit⁺⁺, to differentiate it from the weak Cit⁺ phenotype of earlier isolates with just the citT mutation. Cit⁺⁺ cells contain one or more additional refinement mutations that make robust growth on citrate as a sole carbon source possible. Strains with the Cit⁺⁺ trait can be readily distinguished from Cit⁺ strains by their ability to form colonies within 48 h on minimal agar containing citrate as the only carbon source.Experiments that replayed the evolution of a Cit⁺⁺ phenotype many times from Cit^– isolates taken at different generational time points from the focal LTEE population, found evidence that one or more as-yet-unknown potentiating mutations accumulated in the lineage leading to Cit⁺ that made later strains more likely to access this metabolic innovation (10). Determining what evolved alleles are required for efficient citrate utilization would shed light on the evolutionary pathway that led to the appearance and refinement of this trait. However, identifying these mutations is confounded by the presence of >70 evolved alleles in the earliest strains that have the Cit⁺⁺ phenotype, most of which likely improved growth on glucose by mechanisms that were not relevant for the emergence of efficient citrate utilization.Finding the minimal set of mutations required for Cit⁺⁺, as well as many other complex evolved phenotypes, is currently a daunting task in asexual microbes. Methods such as genome shuffling (14), multiplex automated genome engineering (MAGE) (15), and array-based discovery of adaptive mutations (ADAM) (16) can be used to create and test of libraries consisting of many genetic variants to dissect complex traits. However, no current technique is suitable in a bacterium like E. coli for concurrently (i) capturing the structure of epistatic networks involving multiple alleles spread across the entire chromosome, (ii) introducing point mutations without associated genetic markers, (iii) broad compatibility with different strain backgrounds, and (iv) reconstructing complex chromosomal rearrangements, mobile element insertions, or large genomic duplications, which are common features of these evolved strains (17, 18). To overcome these limitations, we developed recursive genomewide recombination and sequencing (REGRES), a method that uses conjugative chromosomal transfer, phenotypic selection, and whole-genome sequencing to identify the alleles required for an evolved trait.Qualitative traits, including evolutionary innovations, may be the result of all-or-none epistatic interactions, where no subset of mutations is sufficient for expression of a new phenotype (5). This absolute genetic interdependence will lead to a sudden and discrete change in a trait when an actualizing or refinement mutation occurs, as observed with the two steps in the evolution of citrate utilization in the LTEE. In accordance with this model, we expected REGRES with selection for the Cit⁺⁺ phenotype to preserve the citT amplification and some combination of unknown potentiation and refinement mutations. We found that a single refinement mutation affecting the dctA gene was the only evolved allele other than citT conserved across REGRES genomes. These two mutations were sufficient for qualitatively reproducing full utilization of citrate, suggesting a physiological mechanism for refinement that involved activation of a second transporter that makes citrate import by the CitT transporter independent of the production of a cosubstrate by central metabolism. Finally, we discuss an alternative to the all-or-none epistasis hypothesis for how mutations could potentiate the evolution of the strong Cit⁺⁺ trait without being strictly required for its expression.     

Successful Aortic Valve Replacement Surgery in a Patient with Severe Haemophilia a with Low Titre Inhibitor

Acute leukemia, secondary myelodysplasia and paroxysmal nocturnal hemoglobinuria evolving from severe aplastic anemia (AA) following immunosuppressive therapy are well recognized. However, severe AA occurring after complete remission of acute promyelocytic leukemia (APL) has been documented only once in 2009. We report a case of 30-year-old male diagnosed with APL who achieved complete cytogenetic remission with all-trans retinoic acid based induction regimen and developed severe AA few months later during maintenance therapy.     

The effect of pulsed ultrasound on the survival of Drosophila

There are reports in the literature that the temporal peak intensity of pulsed ultrasound may be a better predictor of biological damage than is average intensity. The effects of temporal peak intensity on the survival of Drosophila melanogaster pupae exposed to pulsed ultrasound were investigated. Pulse average intensity and pulse repetition frequency were varied, while temporal average intensity and exposure duration were held constant. Thus, the total energy delivered was held constant. Survival was found to be directly dependent on pulse average intensity. We observed decreased survival at spatial average pulse average intensities as low as 0.5 W/cm2.