Trine Borup Andersen

Measuring glomerular filtration rate in children; can cystatin C replace established methods? A review

Our aim was to evaluate published methods that use serum cystatin C (s-CysC) for measuring glomerular filtration rate (GFR) in children and to discuss advantages and limitations of s-CysC and of established GFR methods. A comprehensive literature review of clinical studies in children evaluating s-CysC or CysC-based formulas and plasma creatinine or creatinine-based formulas against an exogenous reference method using receiver operating characteristics (ROC) curves or Bland–Altman plots is presented. The comparison of s-CysC with plasma creatinine indicated that s-CysC was superior to plasma creatinine in five of 13 studies; four studies showed no difference, and, in four studies, no statistical comparison was made. Comparison of s-CysC and the Schwartz formula showed that s-CysC was superior to the Schwartz formula in two of seven studies; two studies demonstrated no difference, and, in one study, the Schwartz formula was superior to s-CysC. In two studies no statistical comparison was made. The CysC-based prediction equations all had high accuracy but low agreement when compared with a reference GFR, in the range of 30–40% at best. S-CysC is most likely superior to plasma creatinine and at least equal to creatinine-based formulas. CysC-based prediction equations are at least as good as creatinine-based formulas but cannot replace exogenous methods.

GFR Prediction From Cystatin C and Creatinine in Children: Effect of Including Body Cell Mass

BACKGROUND Aiming to develop a more accurate cystatin C-based model for estimation of glomerular filtration rate (GFR) in children, we hypothesized that inclusion of body cell mass (BCM) would increase the accuracy of the GFR estimate in comparison to a well-established GFR reference method. STUDY DESIGN Diagnostic test accuracy study. SETTINGS & PARTICIPANTS 119 children (mean age, 8.8; range, 2.3-14.9 years) referred for GFR measurement by chromium 51 ethylenediaminetetraacetic acid ((51)Cr-EDTA) clearance (mean GFR, 98; range, 13.7-147.4 mL/min/1.73 m(2)). INDEX TEST GFR estimations by the 2 prediction models resulting from theoretical considerations corroborated by forward stepwise variable selection: GFR (mL/min) = 0.542 × (BCM/SCysC)(0.40) × (height × BSA/SCr)(0.65) and GFR (mL/min) = 0.426 × (weight/SCysC)(0.39) × (height × BSA/SCr)(0.64), where SCysC is serum cystatin C level, BSA is body surface area, and SCr is serum creatinine level. The accuracy and precision of these models were compared with 7 previously published prediction models using random subsampling cross-validation. Local constants and coefficients were calculated for all models. Root mean square error, R(2), and percentage of predictions within ±10% and ±30% of the reference GFR were calculated for all models. Based on 1,000 runs of the cross-validation procedure, median values and 2.5th and 97.5th quantiles of the validation parameters were calculated. REFERENCE TEST GFR measurement by (51)Cr-EDTA clearance. RESULTS The BCM model predicted 98% within ±30% of reference GFR and 66% within ±10%, which was higher than for any other model. The weight model predicted 97.5% within ±30% of reference GFR and 62% within ±10%. The BCM model had the highest R(2) and the smallest root mean square error. LIMITATIONS Included only 9 children with GFR <60 mL/min/1.73 m(2). Lack of independent validation cohort. CONCLUSIONS The novel BCM model predicts GFR with higher accuracy than previously published models. The weight model is almost as accurate as the BCM model and allows for GFR estimation without knowledge of BCM. However, endogenous methods are still not sufficiently accurate to replace exogenous markers when GFR must be determined with high accuracy.

Comparison of within- and between-subject variation of serum cystatin C and serum creatinine in children aged 2–13 years

Abstract Background. Previously, data on both the within-subject (SDI) and the between-subject (SDG) variation of cystatin C in children has not been reported. Thus, this study aimed to determine this biological variation including analytical variation (SDA) of both cystatin C and creatinine to characterize the two analytes as renal function markers in children. Methods. On two consecutive days blood samples for duplicate analysis of cystatin C (nephelometric, Dade Behring) and creatinine (enzymatic, Roche) were obtained from 30 children (11 females and 19 males, mean age 8.3 range 2–13 years) referred for GFR measurements by 51Cr-EDTA clearance. For determination of the between-subject variation only children with normal GFR (n=21) were included. Data were adjusted for the well known age-related increase in creatinine. Results. The results are given as coefficients of variation. The within-subject variations were identical for both analytes (6.4%). The between-subject variation was 11.1% for cystatin C and 28.4% for creatinine, though decreasing to 20.1% after adjusting for age. The analytical variation was 1.7% and 2.5% for cystatin C and creatinine, respectively. The index of individuality (IOI = SDI/SDG) was 0.65 for cystatin C and 0.25 for creatinine, though increasing to 0.36 after age-adjustment. Conclusion. The within-subject variation was identical and low for cystatin C and creatinine suggesting that the two are equally suitable for serial monitoring of renal function in children. Based on the low IOI neither analyte, however, seems suitable as a screening marker of renal function in a healthy population of children using population-based reference intervals.

Precision and within- and between-day variation of bioimpedance parameters in children aged 2–14 years

BACKGROUND & AIMS Bioimpedance spectroscopy (BIS) offers the possibility to perform rapid estimates of fluid distribution and body composition. Few studies, however, have addressed the precision and biological variation in a pediatric population. Our objectives were to evaluate precision, variation within- and between-days for the BIS-determined parameters total body fluid, extra-cellular fluid, intra-cellular fluid, body cell mass, fat-free mass, extra-cellular resistance, intra-cellular resistance and percentage body fat using a Xitron 4200. METHODS All 133 children (81 boys, 52 girls; 2.4-14.9 years) had one series measured on day one (precision population). Forty-four children had a second series on day one (within-day sub-population). Thirty-two children had a series measured on the next day (between-day sub-population). Each measurement series consisted of three repeated measurements. A linear mixed model was used for statistical analysis. RESULTS The precision was 0.3-0.8% in children ≥6 years and 0.5-2.4% in children <6 years with a statistically significant difference between the two age-groups (p<0.001). Within-day variation was 1.1-2.8% and between-day variation 2.4-5.7%. Total variation and reference change values are reported. CONCLUSION The Xitron 4200 has a very good but age-dependent precision. The median value of three repeated measurements is recommended in order to avoid incorrect measurements.

Bioimpedance Spectroscopy in Healthy Children

Prediction equations compromise the usefulness of bioimpedance spectroscopy (BIS) in children. This preliminary study explored the possibility of establishing reference curves in children, based on BIS-modeled data and/or resistance index (RI) values. In addition, the precision of repeated measurements was calculated. A total of 105 children (2-14 years old) were enrolled. A BIS device was used and a wrist-ankle electrode configuration was applied. Three repeated measurements were performed in each child. Strongest correlations were seen between RI values and weight for both sexes range, r = 0.94 to 0.97, P < .001. The precision (coefficient of variation in percent) of repeated measurements showed to be low (range, 0.4% ± 0.3% to 1.1% ± 3.1%) We established sex-specific reference curves of RI values, and the precision of repeated measurements showed to be excellent.

Effect of Recent Contrast-Enhanced CT and Patient Age on Image Quality of Thyroid Scintigraphy

Purpose of the Report When thyroid scintigraphy (TS) is performed after contrast-enhanced CT (CE-CT), tracer uptake of 99mTcO4 in the thyroid gland can be inhibited by free iodide. Currently, it is recommended to postpone TS until 4 to 8 weeks after CE-CT, but few data exist to support this recommendation. The purpose was to investigate the effect of CE-CT and other variables for the diagnostic quality of TS. Patients and Methods This retrospective study included 196 patients subjected to TS less than 3 months after a CE-CT (median, 66 days). Patients with elevated thyroid-stimulating hormone (>4.5 mIU/L) or suspected thyroiditis were excluded. Logistic regression was used to calculate the probability of a TS of diagnostic quality with the variables days since CE-CT, age, thyroid-stimulating hormone, and kidney function (eGFR). Results Days since CT and age were highly significant (P < 0.001) predictors for diagnostic TS. The probability of diagnostic quality TS after CE-CT increased with time and reached approximately 70% to 80% 6 to 8 weeks after CE-CT. Analysis of age-specific populations showed age to be a strong independent factor. Conclusions Our findings are in consensus with the currently recommended interval of 6 to 8 weeks between CE-CT and TS. However, our results indicate that patient age should be taken into account, and we suggest the following delay from CE-CT to TS: 4 weeks for patients aged younger than 50 years, 6 weeks for patients aged 50 to 60 years, and 8 weeks for patients aged older than 60 years.

Influence of a Recent Contrast-Enhanced CT on Image Quality of Thyroid Scintigraphy: A retrospective Study of 197 Patients

Aim: Inflammatory bowel disease (IBD) is defined as a chronic relapsing idiopathic inflammation of the gastrointestinal tract. The two main clinical forms of this disease family are Crohn’s Disease (CD) and Ulcerative Colitis (UC). IBD affects an estimated 3.6 million individuals in Europe and North America. To date it is thought that IBD is the result of continual activation of the mucosal immune system. In order to better understand this disease family an in-house developed animal model was implemented and characterized with [18F]FDG (used to illustrate the increased glucose consumption associated with inflammatory processes) and also with TSPO 18 kDa radioligand [18F]DPA-714, an established radiotracer for the study of inflammation within the central nervous system. Materials and Methods: Colonic inflammation was induced in male Wistar rats weighing between 200-250 g by rectal administration of trinitrobenzenesulfonic acid (TNBS) at 4cm from the anal orifice. Control animals were administered, 0.9% aq. sodium chloride analogously. A Siemens Inveon PET/CT tomograph, dedicated to small animals, was used to acquire [18F]FDG images on day 7 post TNBS administration and [18F]DPA714 images the following day. Rats were then sacrificed by an i.v. injection of pentobarbital, and then the lower intestine was extracted and analyzed by immunohistochemistry to determine macrophage infiltration and the presence of TSPO. Results: PET image analysis clearly shows an important accumulation of both radiotracers within the intestinal walls of treated animals in comparison to control animals. Mean levels of [18F]FDG uptake in treated and control animals were 1.20 ± 0.56 %ID/cc and 0.43 ± 0.18 %ID/cc, respectively. Comparable results were found when using [18F]DPA-714, with mean level of uptake in treated and control animals of 1.21 ± 0.62 %ID/cc and 0.46 ± 0.23 %ID/cc, respectively. Immunohistochemistry analysis revealed a higher presence of macrophages in TNBS treated animals. Expression of TSPO was largely increased in the treated animals, when compared to the controls animals, and mainly localized in macrophages cells. Conclusion: Preliminary results seem to indicate that [18F]DPA-714 is an adapted tracer for the study of inflammation of IBD in our animal model. Beyond this, data demonstrating that [18F]DPA-714 could be used to characterize and quantify the level of inflammation during the disease evolution, within the TNBS treated animals, will also be presented. OP366 Surface displayed SNAP-tag as a novel tool for study of Grampositive bacterial infections. B. Mills, V. Steele, J. C. A. Luckett, R. O. Awais, P. Duncanson, V. Griffiths, A. Cockayne, M. Xu, I. Correa, A. C. Perkins, P. Williams, P. Hill; School of Molecular Medical Sciences, University of Nottingham, Nottingham, UNITED KINGDOM, Radiological and Imaging Sciences, University of Nottingham, Nottingham, UNITED KINGDOM, School of Biological and Chemical Sciences, Queen Mary University of London, London, UNITED KINGDOM, New England Biolabs, Inc, Ipswich, ME, UNITED STATES, School of Biosciences, University of Nottingham, Nottingham, UNITED KINGDOM. Introduction: The design of specific probes for in vivo molecular imaging of microbial infections remains one of the greatest challenges to overcome before useful, functional data can be obtained. An increasingly attractive approach for probe design is to express a ligand-binding protein within a cell, which may then covalently bind specific synthetic ligands with attached imaging moieties. One such labelling system is the commercially available SNAP-tag. SNAP-tag specifically and covalently binds O2-benzylguanine (BG) compounds, which may have fluorophores or other functional elements attached at the 4’ position of their benzyl ring. We have designed a BG ligand labelled with Tc, suitable for SPECT imaging. We propose to utilise this technology for the imaging of Staphylococcal infection in vivo with the view to investigate bacterial pathogenicity and to visualise the effect potential antimicrobials may have on bacterial load. Methods: The SNAP-tag gene was codon optimised for expression in the Gram positive bacterium Staphylococcus aureus and fused with an N-terminal spa secretion leader sequence and a Cterminal spa cell-wall anchoring domain. The N-terminal fusion directs the expressed SNAP-tag towards the cell exterior where the C-terminal domain is recognised by the cell-wall sorting enzyme sortase A, covalently anchoring SNAPtag in such a way that the ligand binding domain decorates the cell surface. A novel 99m Tc-HYNIC–NH-BG ligand for SPECT imaging was prepared by coupling BG to HYNIC and radiolabelling with NaTcO4 in the presence of tricine as co-ligand. Radiochemical yields >99% were obtained. nanoSPECT-CT imaging will be used to assess the functional data produced by using SNAP-tag expressing S. aureus cells in in vivo infection models. Results: We have demonstrated that SNAP-tag was expressed and exported to the cell wall where it was covalently anchored. Deletion of the sortase A enzyme prevented attachment of the SNAP-tag to the cell wall, as determined by Western blot. Once situated within the cell wall, SNAP-tag was functional and able to specifically bind cell-impermeable fluorescent BG ligands and our synthesised precursor HYNIC-NH2-BG ligand, as determined by confocal microscopy and fluorometry assay. Pilot in vivo studies for fluorescence optical imaging and nanoSPECT-CT imaging with the novel 99m Tc-HYNIC-NH-BG ligand are currently under development to visualise S. aureus infections in mouse models. Conclusions: This approach should allow a higher sensitivity to be achieved when investigating bacterial infections in real time compared to current molecular imaging techniques, thus allowing bacterial virulence and the potential effects of new antimicrobials to be assessed. OP367 Dual imaging of lipopolysaccharides (LPS) by SPECT-CT and Confocal Microscopy. M. Moreau, V. Duheron, B. Collin, W. Sali, C. Bernhard, C. Goze, T. Gautier, J. Pais de Barros, V. Deckert, F. Brunotte, L. Lagrost, F. Denat; ICMUB UMR CNRS 6302, Dijon, FRANCE, INSERM UMR866, Dijon, FRANCE, Centre Georges-François Leclerc, Dijon, FRANCE, Centre Hospitalier Universitaire, Dijon, FRANCE. Introduction: Lipopolysaccharides (LPS) or endotoxins are found inserted in the outer membrane of Gram-negative bacterias. Their appearance in blood stream triggers a massive secretion of pro-inflammatory cytokines in mammals. A controlled response allows the neutralization and elimination of LPS, whereas an excessive inflammatory response leads to severe circulatory and respiratory defects. It is the endotoxemic shock or septic shock that can leads to death. Many approaches are used to study LPS, including labeling with radiochemicals (3H, 125I, 99mTc or 51Cr) or with fluorophores (FITC, Alexa488, Bodipy). Bimodality is attracting more and more interest in the field of molecular imaging since the combination of two different techniques may provide complementary information, thus improving the accuracy of diagnosis. Combining nuclear modalities (PET or SPECT) with optical imaging is of particular interest, and the similar sensitivities of the two techniques allows to fuse the signaling moieties into a unique molecule, called monomolecular multimodality imaging agent (MOMIA), ensuring a same biodistribution of the two probes. Method: A recently described bimodal probe, namely DOTA-Bodipy-NCS, has been covalently attached to LPS. The integrity of the LPS after labeling procedure was checked by SDS-PAGE electrophoresis and βhydroxymyristate titration (BHM). Pro-inflammatory activity of LPS was assessed by quantification of cytokines released by differentiated THP-1 cells. This bioconjugate was then radiometallated for SPECT-CT biodistribution imaging. Results: DOTABodipy-LPS was metallated with 111In to yield a high specific activity (600 MBq.mg1), with a radiochemical purity >98 % after purification. Biodistribution of the radiolabeled compound was then evaluated in vivo in WT mice by SPECT-CT imaging. Radiolabeled LPS is rapidly eliminated from the bloodstream and accumulates in spleen and liver. Liver slices were then analyzed by confocal microscopy, and specific fluorescent signals in the cytoplasm of hepatocytes were detected, confirming the accumulation of 111In-DOTA-Bodipy-LPS in the liver. Conclusion: These results demonstrate the efficiency of the conjugation process of our bimodal probe. It made it possible to perform both non-invasive SPECT and ex vivo fluorescence imaging of LPS biodistribution, underlining its liver uptake for further detoxification. The 111In-DOTA-Bodipy-LPS probe arises here as a relevant tool to identify key components of LPS detoxification in vivo paving the way to therapeutic issues in the field of sepsis. Acknowlegement: Support was provided by the CNRS, the University of Burgundy, the Conseil Régional de Bourgogne. O P _ M o nd ay S178 Eur J Nucl Med Mol Imaging (2013) 40 (Suppl 2):S89–S567Affibody molecules constitute a class of small (7 kDa) scaffold proteins that can be engineered to have excellent tumor targeting properties. High reabsorption in kidneys complicates development of ...