AB0722 LOSS OF GLYCOSAMINOGLYCANS OF LUMBAR INTERVERTEBRAL DISCS IN PATIENTS WITH ANKYLOSING SPONDYLITIS

Background:To evaluate the glycosaminoglycan (GAG) content of lumbar intervertebral discs (IVD) in patients with ankylosing spondylitis (AS) using GAG chemical exchange saturation transfer (gagCEST).Objectives:Does local GAG content in non-degenerative IVDs measured by gagCEST MRI differs between AS patients and HC?Methods:195 lumbar IVD of 15 patients with AS (mean age 50 ±10 years) and 25 healthy control patients (HC) were prospectively examined with 3 T magnetic resonance imaging (MRI). MRI protocol contained morphological T2 weighted (T2w) images to grade IVD according to the Pfirrmann classification and biochemical imaging with gagCEST to calculate a region of interest (ROI) of the nucleus pulposus (NP) and annulus fibrosus (AF). Prior to statistical testing of gagCEST effects in patients and HC, IVD were classified according to Pfirrmann.Results:Significantly lower gagCEST values of NP and AF were found in non-degenerative IVD (Pfirrmann 1 and 2) of AS patients compared to HC (NP: 1.88 % ±1.21% vs. 3.38 % ±1.71%; p<0.01; confidence interval (CI): 0.89%/2.11%. AF: 1.11 % ± 1.07 % vs. 1.96 %± 1.23 %; p<0.01; CI 0.39%/1.3%).Conclusion:GagCEST analysis of morphologically non-degenerative IVDs in T2w images showed significantly lower GAG values in patients with AS in the NP and AF compared to HC. Our results potentially allow for the detection of GAG loss prior to morphological degeneration.Figure 1.Comparison of morphological T2 weighted (T2w) images to grade IVD according to the Pfirrmann classification and biochemical imaging with gagCEST between HC (A and C) and AS patients (B and D) showing significant lower GAG levels in AS patients.Disclosure of Interests:Philipp Sewerin Grant/research support from: AbbVie Deutschland GmbH & Co. KGBristol-Myers Squibb Celgene GmbHLilly Deutschland GmbHNovartis Pharma GmbH Pfizer Deutschland GmbHRheumazentrum Rhein-Ruhr, Consultant of: AMGEN GmbH AbbVie Deutschland GmbH & Co. KG Biogen GmbHBristol-Myers Squibb Celgene GmbH Chugai Pharma arketing Ltd. / Chugai Europe GmbHHexal Pharma Janssen-CilagGmbH Johnson & Johnson Deutschland GmbHLilly Deutschland GmbH / Lilly Europe / Lilly Global Novartis Pharma GmbH Pfizer Deutschland GmbH Roche Pharma Rheumazentrum Rhein-Ruhr Sanofi-Genzyme Deutschland GmbH Swedish Orphan Biovitrum GmbH UCB Pharma GmbH, Speakers bureau: AMGEN GmbH AbbVie Deutschland GmbH & Co. KG Biogen GmbHBristol-Myers Squibb Celgene GmbH Chugai Pharma arketing Ltd. / Chugai Europe GmbHHexal Pharma Janssen-CilagGmbH Johnson & Johnson Deutschland GmbHLilly Deutschland GmbH / Lilly Europe / Lilly Global Novartis Pharma GmbH Pfizer Deutschland GmbH Roche Pharma Rheumazentrum Rhein-Ruhr Sanofi-Genzyme Deutschland GmbH Swedish Orphan Biovitrum GmbH UCB Pharma GmbH, Daniel Abrar: None declared, Miriam Frenken: None declared, Xenofon Baraliakos Grant/research support from: Grant/research support from: AbbVie, BMS, Celgene, Chugai, Merck, Novartis, Pfizer, UCB and Werfen, Consultant of: AbbVie, BMS, Celgene, Chugai, Merck, Novartis, Pfizer, UCB and Werfen, Speakers bureau: AbbVie, BMS, Celgene, Chugai, Merck, Novartis, Pfizer, UCB and Werfen, Matthias Schneider Grant/research support from: GSK, UCB, Abbvie, Consultant of: Abbvie, Alexion, Astra Zeneca, BMS, Boehringer Ingelheim, Gilead, Lilly, Sanofi, UCB, Speakers bureau: Abbvie, Astra Zeneca, BMS, Chugai, GSK, Lilly, Pfizer, Sanofi, Benedikt Ostendorf: None declared, Christoph Schleich: None declared

Modulation of Lipoteichoic Acids and Exopolysaccharides Prevents Streptococcus mutans Biofilm Accumulation

Dental caries is a diet–biofilm-dependent disease. Streptococcus mutans contributes to cariogenic biofilms by producing an extracellular matrix rich in exopolysaccharides and acids. The study aimed to determine the effect of topical treatments with compound 1771 (modulates lipoteichoic acid (LTA) metabolism) and myricetin (affects the synthesis of exopolysaccharides) on S. mutans biofilms. In vitro S. mutans UA159 biofilms were grown on saliva-coated hydroxyapatite discs, alternating 0.1% sucrose and 0.5% sucrose plus 1% starch. Twice-daily topical treatments were performed with both agents alone and combined with and without fluoride: compound 1771 (2.6 µg/mL), myricetin (500 µg/mL), 1771 + myricetin, fluoride (250 ppm), 1771 + fluoride, myricetin + fluoride, 1771 + myricetin + fluoride, and vehicle. Biofilms were evaluated via microbiological, biochemical, imaging, and gene expression methods. Compound 1771 alone yielded less viable counts, biomass, exopolysaccharides, and extracellular LTA. Moreover, the combination 1771 + myricetin + fluoride decreased three logs of bacterium counts, 60% biomass, >74% exopolysaccharides, and 20% LTA. The effect of treatments on extracellular DNA was not pronounced. The combination strategy affected the size of microcolonies and exopolysaccharides distribution and inhibited the expression of genes linked to insoluble exopolysaccharides synthesis. Therefore, compound 1771 prevented the accumulation of S. mutans biofilm; however, the effect was more pronounced when it was associated with fluoride and myricetin.

Novel strategies of Raman imaging for monitoring intracellular retinoid metabolism in cancer cells

We developed a label-free Raman method for whole-cell biochemical imaging to detect molecular processes that occur in normal and cancer brain cells due to retinol transport in human cancers at the level of isolated organelles. Our approach allows the creation of biochemical maps of lipid droplets, mitochondria and nuclei in single cells. The maps were capable of discriminating triglycerides (TAG) from retinyl ester (RE) in lipid droplets (LD), providing an excellent tool to monitor intracellular retinoid metabolism. We detected spectral changes that arose in proteins and lipids due to retinoid metabolism in human cell lines of normal astrocytes and high-grade cancer cells of glioblastoma as well as in human medulloblastoma and glioblastoma tissue. Raman imaging is an effective tool for monitoring of retinoids and retinol binding proteins involved in carcinogenesis, as monitored by the unique spectral signatures of vibrations. We found two functionally distinct lipid droplets: TAG-LD, for energy storage, and RE-LD, for regulating the level of apo-CRBP1 in cytosol. Raman polarization measurements revealed the occurrence of conformational changes affecting discrete regions of proteins associated with retinol binding. Aberrant expression of retinoids and retinol binding proteins in human tumours were localized in lipid droplets, mitochondria and nuclei according Raman imaging.

Differential Diagnosis

Diseases that should be considered in the differential diagnosis of Hashimoto's thyroiditis (HT) include subacute thyroiditis and Reidle's thyroiditis, Graves' disease, euthyroid sick syndrome, goiter, hypopituitarism, lithium-induced goiter, simple (non-toxic) goiter, thyroid lymphoma, toxic nodular goiter, and types I and II polyglandular autoimmune syndrome. The characteristic diagnostic clinical, biochemical, imaging (sonographic), and histological/cytological features of HT will help make a differential diagnosis. This chapter explores the differential diagnosis of Hashimoto's disease.

Photon-statistics in sensitized emission FRET and FLIM: a comparative theoretical analysis

FRET imaging is an essential analytical method in biomedical research. The limited photon-budget experimentally available, however, imposes compromises between spatiotemporal and biochemical resolutions, photodamage and phototoxicity. The study of photon-statistics in biochemical imaging is thus important in guiding the efficient design of instrumentation and assays. Here, we show a comparative analysis of photon-statistics in FRET imaging demonstrating how the precision of FRET imaging varies vastly with imaging parameters. Therefore, we provide analytical and numerical tools for assay optimization. FLIM is a very robust technique with excellent photon-efficiencies but also intensity-based FRET imaging can reach very high precision by utilizing also information within acceptor fluorescence.

Transthyretin Amyloidosis with Gastrointestinal Manifestation: a Case Report

Transthyretin amyloidosis (ATTR) is a rare, progressive, life-threatening, hereditary disorder caused by mutations in the transthyretin gene. Due to the phenotypic heterogeneity, ATTR is difficult to recognize and it is often diagnosed very late. In ATTR gastrointestinal (GI) disorders play an important role in the patients’ morbidity and mortality. In some cases, GI symptoms are present even before the onset of the peripheral polyneuropathy. However, the complaints are various and it is really difficult to differentiate them from other GI disorders. We present a 61-year old male referred for diarrhea, unintentional weight loss and early satiety. He had hypotension after longstanding hypertension, numbness and tingling in the feet. We considered a broad differential diagnosis spectrum of chronic diarrhea syndrome and performed numerous laboratory, biochemical, imaging, endoscopic, histological and genetic tests. Transthyretin amyloidosis with a Glu89Gln mutation was diagnosed. Transthyretin amyloidosis is frequently misdiagnosed, representing a diagnostic challenge in GI practice. The presence of certain clinical combinations could help gastroenterologists to include ATTR in their diagnostic work-up.

Pancreatic kaposiform hemangioendothelioma complicated by Kasabach–Merritt phenomenon: A rare entity

Primary pancreatic tumours are a rare and unusual entity in children. In this article, we present the case of an 8-month-old girl who presented with obstructive jaundice. The differential diagnosis based on imaging studies was that of a pancreatic vascular neoplasm; however, with the laboratory evidence of Kasabach–Merritt phenomenon (KMP), this prompted the diagnosis of pancreatic kaposiform hemangioendothelioma. A core biopsy of the pancreatic mass was taken at laparotomy and confirmed this diagnosis. The pancreas is an exceedingly rare site of occurrence for this tumour, with only nine cases being published to date. The clinical, biochemical, imaging and pathological findings are discussed to highlight a rare and potentially life-threatening vascular tumour.

THE ROLE OF CECOSTOMY IN THE SURGICAL PATHOLOGY OF THE COLON

In spite of a correct surgical technique, colon interventions can lead to complications of the fistula, sometimes real abdominal dramas, worsening the prognosis of these patients and hardening the surgical technique. We present the evolution of a patient initially treated endoscopically for a sigmoid polyp. The histopathological examination of the biopsy confirmed its malignant character, with invasion of the pedicle, for which reason the endoscopic polypectomy is considered insufficient from an oncological point of view and a colonic resection is advisable. In the postoperative evolution of the patient there appear complications through the occurrence of an anastomotic fistula and peritonitis, whose presence leads to the necessity of performing a terminal colostomy which permits the cessation of the inflammatory syndrome. The further reinstatement of transit is done by means of an “a minima” cecostomy, thus reducing the clinical impact in case of other complications of the fistula. The following elements were analyzed: the patient’s history, associated diseases, fistula-related risk factors, biochemical, imaging and clinical samples, elements related to the surgical technique and the entire postoperative evolution of the patient. This paper stands as an argument in supporting the advantages of “a minima” cesostomy in case of colon surgery, proving how a technical artifice which is minimally invasive for the patient and easy to achieve may become very useful in solving or even preventing some severe complications arising on the background of a simple pathology.

Multiplexed biochemical imaging reveals caspase activation patterns underlying single cell fate

The biochemical activities underlying cell-fate decisions vary profoundly even in genetically identical cells. But such non-genetic heterogeneity remains refractory to current imaging methods, because their capacity to monitor multiple biochemical activities in single living cells over time remains limited1. Here, we deploy a family of newly designed GFP-like sensors (NyxBits) with fast photon-counting electronics and bespoke analytics (NyxSense) in multiplexed biochemical imaging, to define a network determining the fate of single cells exposed to the DNA-damaging drug cisplatin. By simultaneously imaging a tri-nodal network comprising the cell-death proteases Caspase-2, -3 and -92, we reveal unrecognized single-cell heterogeneities in the dynamics and amplitude of caspase activation that signify survival versus cell death via necrosis or apoptosis. Non-genetic heterogeneity in the pattern of caspase activation recapitulates traits of therapy resistance previously ascribed solely to genetic causes3,4. Chemical inhibitors that alter these patterns can modulate in a predictable manner the phenotypic landscape of the cellular response to cisplatin. Thus, multiplexed biochemical imaging reveals cellular populations and biochemical states, invisible to other methods, underlying therapeutic responses to an anticancer drug. Our work develops widely applicable tools to monitor the dynamic activation of biochemical networks at single-cell resolution. It highlights the necessity to resolve patterns of network activation in single cells, rather than the average state of individual nodes, to define, and potentially control, mechanisms underlying cellular decisions in health and disease.