Sequential MRI reveals individual level deformities in the growing scoliotic spine that are clinically masked by the Cobb angle

Introduction Clinically, the Cobb angle method measures the overall scoliotic curve in the coronal plane but does not measure individual vertebra and disc wedging. The contributions of the vertebrae and discs in the growing scoliotic spine were measured to investigate coronal plane deformity progression with growth. Methods A 0.49mm isotropic 3D MRI technique was developed to investigate the level-by-level changes that occur in the growing spine of a group of Adolescent Idiopathic Scoliosis (AIS) patients, who received two to four sequential scans (spaced 3-12 months apart). The coronal plane wedge angles of each vertebra and disc in the major curve were measured to capture any changes that occurred during their adolescent growth phase. Results Seventeen patients had at least two scans. Mean patient age was 12.9 years (SD 1.5 years). Sixteen were classified as right-sided major thoracic Lenke Type 1 (one left sided). Mean standing Cobb angle at initial presentation was 31° (SD 12°). Six received two scans, nine three scans and two four scans, with 65% showing a Cobb angle progression of 5° or more between scans. Overall, there was no clear pattern of deformity progression of individual vertebrae and discs, nor between patients who progressed and those who didn’t. There were measurable changes in the wedging of the vertebrae and discs in all patients. In sequential scans, change in direction of wedging was also seen. In several patients there was reverse wedging in the discs that counteracted increased wedging of the vertebrae such that no change in overall Cobb angle was seen. Conclusion Sequential MRI data showed complex patterns of deformity progression. Changes to the wedging of individual vertebrae and discs may occur in patients who have no increase in Cobb angle measure; the Cobb method alone may be insufficient to capture the complex mechanisms of deformity progression.

Sequential MRI of growing scoliotic spines reveals individual level deformities that are clinically masked by the cobb angle

INTRODUCTION. Clinically, the Cobb angle method measures the overall scoliotic curve in the coronal plane but does not measure individual vertebra and disc wedging. The contributions of the vertebrae and discs in the growing scoliotic spine were measured to investigate coronal plane deformity progression with growth. METHODS. A 0.49mm isotropic 3D MRI technique was developed to investigate the level-by-level changes that occur in the growing spine of a group of Adolescent Idiopathic Scoliosis (AIS) patients, who received two to four sequential scans (spaced 3-12 months apart). The coronal plane wedge angles of each vertebra and disc in the major curve were measured to capture any changes that occurred during their adolescent growth phase. RESULTS. Seventeen patients had at least two scans. Mean patient age was 12.9 years (SD 1.5 years). Sixteen were classified as right-sided major thoracic Lenke Type 1 (one left sided). Mean standing Cobb angle at initial presentation was 31° (SD 12°). Six received two scans, nine three scans and two four scans, with 65% showing a Cobb angle progression of 5° or more between scans. Overall, there was no clear pattern of deformity progression of individual vertebrae and discs, nor between patients who progressed and those who didn’t. There were measurable changes in the wedging of the vertebrae and discs in all patients. In sequential scans, change in direction of wedging was also seen. In several patients there was reverse wedging in the discs that counteracted increased wedging of the vertebrae such that no change in overall Cobb angle was seen. CONCLUSION. Sequential MRI data showed complex patterns of deformity progression. Changes to the wedging of individual vertebrae and discs may occur in patients who have no increase in Cobb angle measure; the Cobb method alone may be insufficient to capture the complex mechanisms of deformity progression.

Supine to standing cobb angle change in idiopathic scoliosis: The effect of endplate preselection

INTRODUCTION Standing radiographs are the ‘gold standard’ for clinical assessment of adolescent idiopathic scoliosis (AIS), with the Cobb Angle used to measure the severity and progression of the scoliotic curve. Supine imaging modalities can provide valuable 3D information on scoliotic anatomy, however, due to changes in gravitational loading direction, the geometry of the spine alters between the supine and standing position which in turn affects the Cobb Angle measurement. Previous studies have consistently reported a 7-10° [1-3] Cobb Angle increase from supine to standing, however, none have reported the effect of endplate pre-selection and which (if any) curve parameters affect the supine to standing Cobb Angle difference. CONCLUSION There is a statistically significant relationship between supine to standing Cobb Angle change and fulcrum flexibility. Therefore, this difference can be considered a measure of spinal flexibility. Pre-selecting vertebral endplates causes only minor changes.

Sequential MRI reveals individual level deformities in the growing scoliotic spine that are not seen clinically by the cobb angle

Clinically, the Cobb angle method measures the overall scoliotic curve in the coronal plane but does not measure individual vertebra and disc wedging. The contributions of the vertebrae and discs in the growing scoliotic spine were measured to investigate coronal plane deformity progression with growth. Sequential MRI data in this project showed complex patterns of deformity progression. Changes to the wedging of individual vertebrae and discs may occur in patients who have no increase in Cobb angle measure; the Cobb method alone may be insufficient to capture the complex mechanisms of deformity progression.

Two monoclonal antibodies to precisely the same epitope of type ii collagen select non-crossreactive phage clones by phage display: Implications for autoimmunity and molecular mimicry

Two monoclonal antibodies (mAb) CB268 and CII-C1 to type II collagen (CII) react with precisely the same conformational epitope constituted by the residues ARGLT on the three chains of the CII triple helix. The antibodies share structural similarity, with most differences in the complementarity determining region 3 of the heavy chain (HCDR3). The fine reactivity of these mAbs was investigated by screening two nonameric phage-displayed random peptide libraries. For each mAb, there were phage clones (phagotopes) that reacted strongly by ELISA only with the selecting mAb, and inhibited binding to CII only for that mAb, not the alternate mAb. Nonetheless, a synthetic peptide RRLPFGSQM corresponding to an insert from a highly reactive CII-C1-selected phagotope, which was unreactive (and non-inhibitory) with CB268, inhibited the reactivity of CB268 with CII. Most phage-displayed peptides contained a motif in the first part of the molecule that consisted of two basic residues adjacent to at least one hydrophobic residue (e.g. RRL or LRR), but the second portion of the peptides differed for the two mAbs. We predict that conserved CDR sequences interact with the basic-basic-hydrophobic motif, whereas non-conserved amino acids in the binding sites (especially HCDR3) interact with unique peptide sequences and limit cross-reactivity. The observation that two mAbs can react identically with a single epitope on one antigen (CII), but show no cross-reactivity when tested against a second (phagotope) indicates that microorganisms could exhibit mimics capable of initiating autoimmunity without this being evident from conventional assays.

Phagotopes derived by antibody screening of phage-displayed random peptide libraries vary in immunoreactivity: Studies using an exemplary monoclonal antibody, CII-C1, to type II collagen

Antibody screening of phage-displayed random peptide libraries to identify mimotopes of conformational epitopes is promising. However, because interpretations can be difficult, an exemplary system has been used in the present study to investigate whether variation in the peptide sequences of selected phagotopes corresponded with variation in immunoreactivity. The phagotopes, derived using a well-characterized monoclonal antibody, CII-C1, to a known conformational epitope on type II collagen, C1, were tested by direct and inhibition ELISA for reactivity with CII-C1. A multiple sequence alignment algorithm, PILEUP, was used to sort the peptides expressed by the phagotopes into clusters. A model was prepared of the C1 epitope on type II collagen. The 12 selected phagotopes reacted with CII-C1 by both direct ELISA (titres from < 100-11 200) and inhibition ELISA (20-100% inhibition); the reactivity varied according to the peptide sequence and assay format. The differences in reactivity between the phagotopes were mostly in accord with the alignment, by PILEUP, of the peptide sequences. The finding that the phagotopes functionally mimicked the C1 epitope on collagen was validated in that amino acids RRL at the amino terminal of many of the peptides were topographically demonstrable on the model of the C1 epitope. Notably, one phagotope that expressed the widely divergent peptide C-IAPKRHNSA-C also mimicked the C1 epitope, as judged by reactivity in each of the assays used: these included cross-inhibition of CII-C1 reactivity with each of the other phagotopes and inhibition by a synthetic peptide corresponding to that expressed by the most frequently selected phagotope, RRLPFGSQM. Thus, it has been demonstrated that multiple phage-displayed peptides can mimic the same epitope and that observed immunoreactivity of selected phagotopes with the selecting mAb can depend on the primary sequence of the expressed peptide and also on the assay format used.

Rheumatoid arthritis sera react with a phage-displayed peptide selected by a monoclonal antibody to type II collagen that has homology to EBNA-1

Antibodies to type II collagen, and to Epstein Barr virus nuclear antigen-1 (EBNA-1) have been associated with rheumatoid arthritis (RA). In studies involving probing of phage-displayed random peptide libraries with an antibody to type II collagen, CII-C1, we observed that among 17 phagotopes selected 5 expressed peptides with homology with the sequence of EBNA-1. The residues in common were RLPFG. Hence we tested sera from 50 patients with RA, of whom 26 had antibodies to native type II collagen, and 43 healthy controls, for reactivity by ELISA with a phagotope selected 4 times, which expressed the peptide RRLPFGSQM. Eight RA sera (16%) but no normal sera reacted with the phagotope (p = 0.025). This reactivity could not be correlated with reactivity of RA sera with EBNA-1 by semi-quantitative western blot, with which reactivity occurred in 78% of RA patients and 81% of controls. Evidence for molecular mimicry was not found insofar as the phagotope did not inhibit reactivity of RA sera with EBNA-1 and CII-C1 was not reactive with EBNA-1. We conclude that the reactivity of the RA sera with the phagotope is most likely due to the phagotope being a mimic of an epitope of type II collagen for a proportion of RA sera.

Mimotopes identified by phage display for the monoclonal antibody CII- C1 to type II collagen

The characterization of B cell epitopes has been advanced by the use of random peptide libraries displayed within the coat protein of bacteriophage. This technique was applied to the monoclonal antibody (mAb) C1 to type II collagen (CII-C1). CII-C1 is known to react with a conformational epitope on type II collagen that includes residues 359-363. Three rounds of selection were used to screen two random nonameric phage libraries and 18 phagotopes were isolated. CII-C1 reacted by ELISA with 17 of the 18 phagotopes: one phagotope contained a stop codon. Of the eight most reactive phage, seven inhibited the reactivity by ELISA of CII-C1 with type II collagen. Of the 18 phage isolated, 11 encoded the motif F-G-x-Q with the sequence F-G-S-Q in 6, 2 encoded F-G-Q, and one the reverse motif Q-x-y-F. Most phagotopes that inhibited the reactivity of CII-C1 encoded two particular motifs consisting of two basic amino acid residues and a hydrophobic residue in the first part of the insert and the F-G-x-Q or F-G-Q motif ill the second part; phagotopes which contained only one basic residue in the first part of the sequence were less reactive. These motifs are not represented in the linear sequence of type II collagen and thus represent mimotopes of the epitope for CII-C1 on type II collagen. There were five phagotopes with peptide inserts containing the sequence RLPFG occurring in the Epstein-Barr virus nuclear antigen, EBNA- 1. This is of interest because EBV has been implicated in the initiation of rheumatoid arthritis (RA) by reason of increased reactivity to EBNA-1 in RA sera. In conclusion, the phage display technique disclosed mimotopes for a conformational epitope of type II collagen, and revealed an interesting homology with a sequence of the EBNA-1 antigen from Epstein Barr virus.

New evidence of the reproductive organs of Glossopteris based on permineralized fossils from Queensland, Australia. II: pollen-bearing organ Ediea gen. nov

Ediea homevalensis H. Nishida, Kudo, Pigg & Rigby gen. et sp. nov. is proposed for permineralized pollen-bearing structures from the Late Permian Homevale Station locality of the Bowen Basin, Queensland, Australia. The taxon represents unisexual fertile shoots bearing helically arranged leaves on a central axis. The more apical leaves are fertile microsporophylls bearing a pair of multi-branched stalks on their adaxial surfaces that each supports a cluster of terminally borne pollen sacs. Proximal to the fertile leaves there are several rows of sterile scale-like leaves. The pollen sacs (microsporangia) have thickened and dark, striate walls that are typical of the Arberiella type found in most pollen organs presumed to be of glossopterid affinity. An examination of pollen organs at several developmental stages, including those containing in situ pollen of the Protohaploxypinus type, provides the basis for a detailed analysis of these types of structures, which bear similarities to both compression/impression Eretmonia-type glossopterid microsporangiate organs and permineralized Eretmonia macloughlinii from Antarctica. These fossils demonstrate that at least some Late Permian pollen organs were simple microsporophyll-bearing shoot systems and not borne directly on Glossopteris leaves.

A rapid and label-free dual detection of Hg (II) and cysteine with the use of fluorescence switching of graphene quantum dots

A simple and rapid method of analysis for mercury ions (Hg2+) and cysteine (Cys) was developed with the use of graphene quantum dots (GQDs) as a fluorescent probe. In the presence of GQDs, Hg2+ cations are absorbed on their negatively charged surface by means of electrostatic interactions. Thus, the fluorescence (FL) of the GQDs would be significantly quenched as a result of the FL charge transfer, e.g. 92% quenching at 450 nm occurs for a 5 μmol L−1 Hg2+ solution. However, when Cys was added, a significant FL enhancement was observed (510% at 450 nm for a 8.0 μmol L−1 Cys solution), and Hg2+ combined with Cys rather than with the GQDs in an aqueous solution. This occurred because a strong metalsingle bondthiol bond formed, displacing the weak electrostatic interactions, and this resulted in an FL enhancement of the GQDs. The limits of detection (LOD) for Hg2+ and Cys were 0.439 nmol L−1 and 4.5 nmol L−1, respectively. Also, this method was used successfully to analyze Hg2+ and Cys in spiked water samples.

Missing in space: An evaluation of imputation methods for missing data in spatial analysis of risk factors for type II diabetes

Background Spatial analysis is increasingly important for identifying modifiable geographic risk factors for disease. However, spatial health data from surveys are often incomplete, ranging from missing data for only a few variables, to missing data for many variables. For spatial analyses of health outcomes, selection of an appropriate imputation method is critical in order to produce the most accurate inferences. Methods We present a cross-validation approach to select between three imputation methods for health survey data with correlated lifestyle covariates, using as a case study, type II diabetes mellitus (DM II) risk across 71 Queensland Local Government Areas (LGAs). We compare the accuracy of mean imputation to imputation using multivariate normal and conditional autoregressive prior distributions. Results Choice of imputation method depends upon the application and is not necessarily the most complex method. Mean imputation was selected as the most accurate method in this application. Conclusions Selecting an appropriate imputation method for health survey data, after accounting for spatial correlation and correlation between covariates, allows more complete analysis of geographic risk factors for disease with more confidence in the results to inform public policy decision-making.

Common variants in the HLA-DRB1-HLA-DQA1 HLA class II region are associated with susceptibility to visceral leishmaniasis

To identify susceptibility loci for visceral leishmaniasis, we undertook genome-wide association studies in two populations: 989 cases and 1,089 controls from India and 357 cases in 308 Brazilian families (1,970 individuals). The HLA-DRB1-HLA-DQA1 locus was the only region to show strong evidence of association in both populations. Replication at this region was undertaken in a second Indian population comprising 941 cases and 990 controls, and combined analysis across the three cohorts for rs9271858 at this locus showed P combined = 2.76 × 10 -17 and odds ratio (OR) = 1.41, 95% confidence interval (CI) = 1.30-1.52. A conditional analysis provided evidence for multiple associations within the HLA-DRB1-HLA-DQA1 region, and a model in which risk differed between three groups of haplotypes better explained the signal and was significant in the Indian discovery and replication cohorts. In conclusion, the HLA-DRB1-HLA-DQA1 HLA class II region contributes to visceral leishmaniasis susceptibility in India and Brazil, suggesting shared genetic risk factors for visceral leishmaniasis that cross the epidemiological divides of geography and parasite species. © 2013 Nature America, Inc. All rights reserved.

Genome-wide association study identifies susceptibility loci for open angle glaucoma at TMCO1 and CDKN2B-AS1

We report a genome-wide association study for open-angle glaucoma (OAG) blindness using a discovery cohort of 590 individuals with severe visual field loss (cases) and 3,956 controls. We identified associated loci at TMCO1 (rs4656461[G] odds ratio (OR) = 1.68, P = 6.1 × 10-10) and CDKN2B-AS1 (rs4977756[A] OR = 1.50, P = 4.7 × 10-9). We replicated these associations in an independent cohort of cases with advanced OAG (rs4656461 P = 0.010; rs4977756 P = 0.042) and two additional cohorts of less severe OAG (rs4656461 combined discovery and replication P = 6.00 × 10-14, OR = 1.51, 95% CI 1.35-1.68; rs4977756 combined P = 1.35 × 10-14, OR = 1.39, 95% CI 1.28-1.51). We show retinal expression of genes at both loci in human ocular tissues. We also show that CDKN2A and CDKN2B are upregulated in the retina of a rat model of glaucoma. © 2011 Nature America, Inc. All rights reserved.

Comparison of the Cosmed K4 b2 and the Deltatrac II™ metabolic cart in measuring resting energy expenditure in adults

Background and Aims: The objective of the study was to compare data obtained from the Cosmed K4 b2 and the Deltatrac II™ metabolic cart for the purpose of determining the validity of the Cosmed K4 b2 in measuring resting energy expenditure. Methods: Nine adult subjects (four male, five female) were measured. Resting energy expenditure was measured in consecutive sessions using the Cosmed K4 b2, the Deltatrac II™ metabolic cart separately and the Cosmed K4 b2 and Deltatrac II™ metabolic cart simultaneously, performed in random order. Resting energy expenditure (REE) data from both devices were then compared with values obtained from predictive equations. Results: Bland and Altman analysis revealed a mean bias for the four variables, REE, respiratory quotient (RQ), VCO2, VO2 between data obtained from Cosmed K4 b2 and Deltatrac II™ metabolic cart of 268 ± 702 kcal/day, -0.0±0.2, 26.4±118.2 and 51.6±126.5 ml/min, respectively. Corresponding limits of agreement for the same four variables were all large. Also, Bland and Altman analysis revealed a larger mean bias between predicted REE and measured REE using Cosmed K4 b2 data (-194±603 kcal/day) than using Deltatrac™ metabolic cart data (73±197 kcal/day). Conclusions: Variability between the two devices was very high and a degree of measurement error was detected. Data from the Cosmed K4 b2 provided variable results on comparison with predicted values, thus, would seem an invalid device for measuring adults. © 2002 Elsevier Science Ltd. All rights reserved.

The ascidian natural product eusynstyelamide B is a novel topoisomerase II poison that induces DNA damage and growth arrest in prostate and breast cancer cells

As part of an anti-cancer natural product drug discovery program, we recently identified eusynstyelamide B (EB), which displayed cytotoxicity against MDA-MB-231 breast cancer cells (IC50 = 5 μM) and induced apoptosis. Here, we investigated the mechanism of action of EB in cancer cell lines of the prostate (LNCaP) and breast (MDA-MB-231). EB inhibited cell growth (IC50 = 5 μM) and induced a G2 cell cycle arrest, as shown by a significant increase in the G2/M cell population in the absence of elevated levels of the mitotic marker phospho-histone H3. In contrast to MDA-MB-231 cells, EB did not induce cell death in LNCaP cells when treated for up to 10 days. Transcript profiling and Ingenuity Pathway Analysis suggested that EB activated DNA damage pathways in LNCaP cells. Consistent with this, CHK2 phosphorylation was increased, p21CIP1/WAF1 was up-regulated and CDC2 expression strongly reduced by EB. Importantly, EB caused DNA double-strand breaks, yet did not directly interact with DNA. Analysis of topoisomerase II-mediated decatenation discovered that EB is a novel topoisomerase II poison.