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To provide precise age-specific risk estimates of cancers other than female breast and ovarian cancers associated with pathogenic variants (PVs) inWe used data from 3,184In addition to female breast and ovarian cancers

Reconstitution of T cell repertoire after allogeneic stem cell transplantation is a long and often incomplete process. As a result, reactivation of Epstein-Barr virus (EBV) is a frequent complication that may be treated by adoptive transfer of donor-derived EBV-specific T cells. We generated donor-derived EBV-specific T cells by peptide stimulation and adoptively transferred them to a patient with angioimmunoblastic T-cell lymphoma (AITL), who had developed persisting high titers of EBV concomitant to relapse after transplantation. T cell receptor beta (TCRβ) deep sequencing showed that the T cell repertoire of the patient early after transplantation (day 60) was strongly reduced and only very low numbers of EBV-specific T cells were detectable. Manufacturing andin vitroexpansion of donor-derived EBV-specific T cells resulted in enrichment of EBV epitope-specific, HLA-restricted T cells. Monitoring after adoptive transfer revealed that the dominant TCR sequences from peptide-stimulated T cells persisted long-term and established an EBV-specific TCR clonotype repertoire in the host, with many of the EBV-specific TCRs present in the donor. This reconstituted repertoire was associated with immunological control of EBV and with lack of further AITL relapse.Author summaryA characteristic feature of all herpesviruses is their persistence in the host’s body after primary infection. Hence, the host’s immune system is confronted with the problem to control these viruses life-long. Well-known representative of the herpesvirus group are the classic Herpes-Simplex Virus (HSV-1) and Varicella Zoster Virus (VZV, causing chicken pox); a less known representative is Epstein Barr Virus (EBV, causing mononucleosis). When the immune system is severely compromised, for example after stem cell transplantation from a foreign (allogeneic) donor, these viruses can reappear, as they are already in the host’s body. Especially EBV cause life-threatening complications after stem cell transplantation and only reinforcement of the host’s immune system can reestablish viral control. Here we show thatex vivomanufactured EBV-specific T cells can reestablish long-term control of EBV and that these cells persist in the host’s body over months. These results give us a better understanding of viral immune reconstitution post-transplant and of clinically-relevant T cell populations against EBV.

Zusammenfassung Hintergrund Die Therapie mit CPAP (continuous positive airway pressure therapy) ist die Referenztherapie beim mittel- und hochgradigen obstruktiven Schlafapnoesyndrom (OSAS). Die Therapietreue bei diesem Verfahren ist jedoch nicht zufriedenstellend. Ziel der Untersuchung war die Analyse der Einflussfaktoren auf die Therapietreue sowie die Evaluation, ob eine strukturierte Nachsorge die Therapietreue verbessert. Material und Methoden In dieser Studie wurden alle Patienten (n = 237) mit der Diagnose eines milden bis schweren OSAS und einer neu angepassten CPAP-Therapie im Zeitraum zwischen 2011 und 2013 untersucht. Die Nachsorgetermine fanden alle 1593,7 ± 77,4 CPAP-Betriebsstunden statt. Ergebnisse Unter Therapie nahm der AHI-Wert (Apnoe-Hypopnoe-Index) signifikant ab (5,6 ± 8,5/h; p < 0,001). 79 % erfüllten die Kriterien einer CPAP-Therapietreue (mindestens 4 Betriebsstunden an 7 Tagen pro Woche). Es gab keine signifikanten Einflüsse von demografischen, klinischen oder pharmakologischen Charakteristika der Patienten auf die Therapietreue. Die häufigsten Probleme unter CPAP-Therapie waren Trockenheit der Nasen- und Rachenschleimhäute (43,7 %) und Druckstellen (22,4 %). Die CPAP-Therapietreue korrelierte nicht mit den Werten der Epworth Sleepiness Scale. Die Zufriedenheit mit der Nachsorge korrelierte signifikant mit der Therapietreue (r = 0,185; p = 0,032), während der Therapiedruck keinen signifikanten Einfluss hierauf zeigte (r = −0,072; p = 0,383). Schlussfolgerungen Eine strukturierte, individuell angepasste Nachsorge ist wichtig für die Therapietreue. In Zukunft sollte im Rahmen der Nachsorge neben der Kontrolle des Therapieerfolges mehr Aufmerksamkeit auf Aspekte der Problemlösung, Rückkopplung, Schulung und Motivation gerichtet werden.

Zusammenfassung Ärzte in Ausbildung haben häufig bei Berufseintritt aufgrund mangelnder Trainingsangebote im Rahmen ihres Medizinstudiums kaum Erfahrung in der Anwendung und Beurteilung der Sonografie. Daher sollte ein multimediales Programm für das schrittweise Erlernen und das Training der Schilddrüsensonografie entwickelt werden. Es wurde eine Software zur Gestaltung eines multimedialen Lernprogrammes für ein tragbares Ultraschallsystem genutzt. Die Software erlaubt die Darstellung von Texten und Bildmaterial parallel zur Untersuchung, um eigene Befunde mit Beispielen aus einer Datenbank vergleichen zu können. Das fertiggestellte Tutorial wurde durch 8 Studierende evaluiert und mit einem Tutor-geführten Training verglichen. Es wurde eine strukturierte, stufenweise Anleitung zur Schilddrüsensonografie mit einer Vielzahl von Beispiel-Sonografien in verschiedenen Schnittbildebenen gestaltet, die der Nutzer parallel am Probanden mit dem Ultraschallgerät nachstellt. Der Informationsgehalt des Tutorials und die Nachvollziehbarkeit der Untersuchungsschritte wurden positiv eingeschätzt. Die Übersichtlichkeit, die Eignung für Anwender mit fehlender Erfahrung und somit auch der Lernerfolg wurden hingegen unterschiedlich bewertet. Das Tutorial zum Erlernen und zum Training der Schilddrüsensonografie ist ein Instrument zum Selbstlernen und zur Verbesserung der praktischen Ausbildung. In der nächsten Version wird die Untersuchungsanleitung noch detaillierter aufgebaut sein.

Obstructive sleep apnea represents the most common form of sleep-disordered breathing and has a high prevalence in patients with cardiovascular disease. Disturbed sleep is a potential risk factor for cardiovascular disorders such as arterial hypertension, cardiac ischemia, sudden cardiac death, and stroke. In this study we investigated polysomnographic records and analyzed the ECG, plethysmogram, respiration and SpO2 time series during wakefulness (WK), NREM, and REM sleep stages applying variability and coupling analyses methods. We enrolled 55 patients with obstructive sleep apnea syndrome (OSAS) and 29 healthy control subjects (CON: 45.9±14.9 years, 21 male) in this study. OSAS patients were subdivided into a low- and high-risk group (LR: 50.8±14.1 years, n=29, 21 male; HR: 57.2±13.4 years, n=26, 19 male) based on the Apnea–Hypopnea Index (AHI) (CON: 0-5 AHI, LR: 5-15 AHI, HR: >15 AHI). We could demonstrate the presence of an altered autonomic function in OSAS patients, differing from healthy controls. This altered autonomic function was mainly based on heart rate-, respiratory-, SpO2- and plethysmogram variability and their couplings. The discriminant analysis showed that the optimal set consisting of two autonomic indices revealed a high classification power (ACC=86.7%, AUC=90.3%, SENS=89.5% and SPEC=84.6%) when comparing low-risk and high-risk OSAS patients during WK. These results were slightly improved when analyzing REM sleep stages. Based on these results it seems to be possible to perform optimal risk stratification for OSAS patients based on autonomic indices. Based on our findings it is possible to differentiate between high-risk OSAS patient and low-risk OSAS patient at an early stage and in a promising manner allowing to set up therapy strategies for those patients in an early stage.

Cell alterations during isolation and preparation for flow cytometry cell sorting by antibodies, temperature, homogenization, buffer composition and mitogens are well known. In contrast, little is known about cell alteration caused by the instrument or the sorting process itself. We systematically evaluated cellular responses to different sorter-induced physical forces. In summary, flow cytometry cell-sorting induced forces can affect cellular signaling cascades, especially the MAPK p38. Functional assays, related to the p38 MAPK pathway, of human primary T cells after flow cytometry sorting did lead to minor physiological modulation but no functional impairments. © 2020 The Authors. Cytometry Part A published by Wiley Periodicals, Inc. on behalf of International Society for Advancement of Cytometry.

The multifactorial likelihood analysis method has demonstrated utility for quantitative assessment of variant pathogenicity for multiple cancer syndrome genes. Independent data types currently incorporated in the model for assessing BRCA1 and BRCA2 variants include clinically calibrated prior probability of pathogenicity based on variant location and bioinformatic prediction of variant effect, co-segregation, family cancer history profile, co-occurrence with a pathogenic variant in the same gene, breast tumor pathology, and case-control information. Research and clinical data for multifactorial likelihood analysis were collated for 1,395 BRCA1/2 predominantly intronic and missense variants, enabling classification based on posterior probability of pathogenicity for 734 variants: 447 variants were classified as (likely) benign, and 94 as (likely) pathogenic; and 248 classifications were new or considerably altered relative to ClinVar submissions. Classifications were compared with information not yet included in the likelihood model, and evidence strengths aligned to those recommended for ACMG/AMP classification codes. Altered mRNA splicing or function relative to known nonpathogenic variant controls were moderately to strongly predictive of variant pathogenicity. Variant absence in population datasets provided supporting evidence for variant pathogenicity. These findings have direct relevance for BRCA1 and BRCA2 variant evaluation, and justify the need for gene-specific calibration of evidence types used for variant classification., Ohio State University Comprehensive Cancer Center Barretos Cancer Hospital. Grant Number: FINEP ‐ CT‐INFRA (02/2010) Breast Cancer Foundation of New Zealand Canadian Institutes of Health Research. Grant Number: PSR‐SIIRI‐701 Cancer Research UK. Grant Numbers: C8197/A16565, C5047/A8384, C1281/A12014, C12292/A11174, C1287/A10710, C1287/A10118, C1287/A16563, C5047/A10692, C5047/A15007 Department of Defence, USA. Grant Number: W81XWH‐10‐1‐0341 Helsinki University Hospital Research fund Scientific Foundation Asociación Española Contra el Cáncer Leiden University Medical Centre. Grant Number: Grant 30.925 Generalitat de Catalunya. Grant Numbers: PERIS_MedPerCan, URDCat, 2017SGR1282, 2017SGR496 Royal Society of New Zealand Cancer Council Victoria Netherlands Organization for Scientific Research (NWO). Grant Number: Grant 017.008.022 Breast Cancer Research Foundation Cancer Foundation of Western Australia EU H2020. Grant Number: 634935 Fundación Mutua Madrileña Seventh Framework Programme. Grant Numbers: 634935, 223175, 633784 Cancer Council South Australia Government of Galicia. Grant Number: Consolidation and structuring program: IN607B Cancer Council Tasmania Italian Association of Cancer Research. Grant Number: 15547 Queensland Cancer Fund AstraZeneca National Institute of Health (USA). Grant Numbers: 1U19 CA148065‐01, CA128978, CA192393, 1U19 CA148537, P50 CA1162091, CA116167, 1U19 CA148112 Newcastle University Dutch Cancer Society KWF. Grant Numbers: KWF/Pink Ribbon‐11704, UL2012‐5649 National Institute for Health Research. Grant Number: Manchester Biomedical Research centre (IS‐BRC‐1215 National Council of Technological and Scientific Development (CNPq) Instituto de Salud Carlos III. Grant Numbers: FIS PI15/00355, FIS PI13/01711, CIBERONC, FIS PI16/01218, PI16/00563 French National Institute of Cancer National Breast Cancer Foundation National Health and Medical Research Council. Grant Numbers: ID1061778, ID1104808 Carlos III National Health Centro de Investigación Biomédica en Red de Enferemdades Raras. Grant Number: ACCI 2016: ER17P1AC7112/2018 Cancer Council NSW Deutsche Krebshilfe. Grant Numbers: (#110837, #70111850 Fondazione Pisa. Grant Number: Grant “Clinical characterization of BRCA 1/2 Mis

Summary Multinucleated giant cells (MGCs) form by fusion of macrophages and are presumed to contribute to the removal of debris from tissues. In a systematic in vitro analysis, we show that IL-4-induced MGCs phagocytosed large and complement-opsonized materials more effectively than their unfused M2 macrophage precursors. MGC expression of complement receptor 4 (CR4) was increased, but it functioned primarily as an adhesion integrin. In contrast, although expression of CR3 was not increased, it became functionally activated during fusion and was located on the extensive membrane ruffles created by excess plasma membrane arising from macrophage fusion. The combination of increased membrane area and activated CR3 specifically equips MGCs to engulf large complement-coated targets. Moreover, we demonstrate these features in vivo in the recently described complement-dependent therapeutic elimination of systemic amyloid deposits by MGCs. MGCs are evidently more than the sum of their macrophage parts., Graphical Abstract, Highlights • MGCs are specialized for phagocytosis of large and complement-opsonized particles • MGCs show extensive membrane ruffles containing pre-activated complement receptor 3 • Membrane ruffles provide excess membrane for ingestion of large materials • MGCs eliminate systemic amyloid deposits after immunotherapeutic targeting, Macrophage-derived multinucleated giant cells (MGCs) form in diverse chronic inflammatory diseases, but their functional role remains unclear. Milde et al. show that MGCs are specialized for complement-mediated phagocytosis and destruction of large targets and demonstrate their key role in the therapeutic elimination of the pathogenic amyloid deposits in systemic amyloidosis.

Continuous positive airway pressure (CPAP) therapy is the reference therapy for moderate and severe obstructive sleep apnea syndrome (OSAS). However, CPAP adherence is dissatisfying. We analyzed influencing factors on CPAP adherence and we evaluated if a structured follow-up enhances its adherence.In this study all patients (n = 237) with a newly adjusted CPAP therapy due to the diagnosis of mild to severe OSAS between 2011 and 2013 were investigated. Follow-up took place every 1593.7 ± 77.4 CPAP operation hours.AHI (Apnea-hypopnea index) decreased significantly during therapy (5.6 ± 8.5/h; p0.001). 79 % fulfilled the criteria of CPAP adherence (at least 4 operation hours at 7 days a week). There were no significant influences on CPAP adherence seen in patients' demographic, clinical and pharmacological characteristics. The most common problems of patients receiving a CPAP therapy were dryness of the mucous membranes (43.7 %) and pressure marks (22.4 %). Considering the changes in Epworth Sleepiness Scale there were no significant relations to CPAP adherence. Satisfaction with our follow-up significantly correlated with CPAP adherence (r = 0.185; p = 0.032), whereas therapy pressure did not have any significant influence on CPAP adherence (r = -0.072; p = 0.383).A structured and individually adapted follow-up is important. In future, apart from control of therapy success, greater attention in the follow-up of CPAP therapy should be given to aspects of problem solution, feedback, education, and motivation.Die Therapie mit CPAP (continuous positive airway pressure therapy) ist die Referenztherapie beim mittel- und hochgradigen obstruktiven Schlafapnoesyndrom (OSAS). Die Therapietreue bei diesem Verfahren ist jedoch nicht zufriedenstellend. Ziel der Untersuchung war die Analyse der Einflussfaktoren auf die Therapietreue sowie die Evaluation, ob eine strukturierte Nachsorge die Therapietreue verbessert.In dieser Studie wurden alle Patienten (n = 237) mit der Diagnose eines milden bis schweren OSAS und einer neu angepassten CPAP-Therapie im Zeitraum zwischen 2011 und 2013 untersucht. Die Nachsorgetermine fanden alle 1593,7 ± 77,4 CPAP-Betriebsstunden statt.Unter Therapie nahm der AHI-Wert (Apnoe-Hypopnoe-Index) signifikant ab (5,6 ± 8,5/h; p0,001). 79 % erfüllten die Kriterien einer CPAP-Therapietreue (mindestens 4 Betriebsstunden an 7 Tagen pro Woche). Es gab keine signifikanten Einflüsse von demografischen, klinischen oder pharmakologischen Charakteristika der Patienten auf die Therapietreue. Die häufigsten Probleme unter CPAP-Therapie waren Trockenheit der Nasen- und Rachenschleimhäute (43,7 %) und Druckstellen (22,4 %). Die CPAP-Therapietreue korrelierte nicht mit den Werten der Epworth Sleepiness Scale. Die Zufriedenheit mit der Nachsorge korrelierte signifikant mit der Therapietreue (r = 0,185; p = 0,032), während der Therapiedruck keinen signifikanten Einfluss hierauf zeigte (r = −0,072; p = 0,383).Eine strukturierte, individuell angepasste Nachsorge ist wichtig für die Therapietreue. In Zukunft sollte im Rahmen der Nachsorge neben der Kontrolle des Therapieerfolges mehr Aufmerksamkeit auf Aspekte der Problemlösung, Rückkopplung, Schulung und Motivation gerichtet werden.

Objective Refractory coeliac disease (RCD) is a potentially hazardous complication of coeliac disease (CD). In contrast to RCD type I, RCD type II is a precursor entity of enteropathy-associated T-cell lymphoma (EATL), which is associated with clonally expanding T-cells that are also found in the sequentially developing EATL. Using high-throughput sequencing (HTS), we aimed to establish the small-intestinal T-cell repertoire (TCR) in CD and RCD to unravel the role of distinct T-cell clonotypes in RCD pathogenesis. Design DNA extracted from duodenal mucosa specimens of controls (n=9), active coeliacs (n=10), coeliacs on a gluten-free diet (n=9), RCD type I (n= 8), RCD type II (n= 8) and unclassified Marsh I cases (n= 3) collected from 2002 to 2013 was examined by TCR beta-complementarity- determining regions 3 (CDR3) multiplex PCR followed by HTS of the amplicons. Results On average, 106 sequence reads per sample were generated consisting of up to 900 individual TCR beta rearrangements. In RCD type II, the most frequent clonotypes (ie, sequence reads with identical CDR3) represent in average 42.6% of all TCR beta rearrangements, which was significantly higher than in controls (6.8%; p

Null mutations in genes involved in V(D)J recombination cause a block in B- and T-cell development, clinically presenting as severe combined immunodeficiency (SCID). Hypomorphic mutations in the non-homologous end-joining gene DCLRE1C (encoding ARTEMIS) have been described to cause atypical SCID, Omenn syndrome, Hyper IgM syndrome and inflammatory bowel disease-all with severely impaired T-cell immunity. By whole-exome sequencing, we investigated the molecular defect in a consanguineous family with three children clinically diagnosed with antibody deficiency. We identified perfectly segregating homozygous variants in DCLRE1C in three index patients with recurrent respiratory tract infections, very low B-cell numbers and serum IgA levels. In patients, decreased colony survival after irradiation, impaired proliferative response and reduced counts of naïve T cells were observed in addition to a restricted T-cell receptor repertoire, increased palindromic nucleotides in the complementarity determining regions 3 and long stretches of microhomology at switch junctions. Defective V(D)J recombination was complemented by wild-type ARTEMIS protein in vitro. Subsequently, homozygous or compound heterozygous DCLRE1C mutations were identified in nine patients from the same geographic region. We demonstrate that DCLRE1C mutations can cause a phenotype presenting as only antibody deficiency. This novel association broadens the clinical spectrum associated with ARTEMIS mutations. Clinicians should consider the possibility that an immunodeficiency with a clinically mild initial presentation could be a combined immunodeficiency, so as to provide appropriate care for affected patients.

Physicians in education often have poor experience in practice and assessment of ultrasonography on entering their profession, due to a deficiency of training offers during their study of medicine. Hence, a multimedia device for stepwise learning and training ultrasonography of the thyroid was developed. A software for a portable ultrasonography system was used to design a multimedia device for ultrasonography of the thyroid. It allows the user to illustrate texts and pictorial material simultaneously with ultrasound examination in order to compare own findings with examples from a database. The device was evaluated by 8 medical students and compared to a tutor-guided training. A structured, stepwise manual for ultrasonography of the thyroid with a large content of examples in different sectional images was designed for simultaneous reconstruction with the ultrasonography device. The informative content of the device and the replicability of the examination procedure were evaluated positively. Assessment respecting clarity, eligibility for users without experience and learning success was varying. The tutorial to learn and train ultrasonography of the thyroid is an instrument for self-learning and improving practical education in ultrasonography in medical education. In the next version, the manual for the examination will be structured in greater detail.Ärzte in Ausbildung haben häufig bei Berufseintritt aufgrund mangelnder Trainingsangebote im Rahmen ihres Medizinstudiums kaum Erfahrung in der Anwendung und Beurteilung der Sonografie. Daher sollte ein multimediales Programm für das schrittweise Erlernen und das Training der Schilddrüsensonografie entwickelt werden. Es wurde eine Software zur Gestaltung eines multimedialen Lernprogrammes für ein tragbares Ultraschallsystem genutzt. Die Software erlaubt die Darstellung von Texten und Bildmaterial parallel zur Untersuchung, um eigene Befunde mit Beispielen aus einer Datenbank vergleichen zu können. Das fertiggestellte Tutorial wurde durch 8 Studierende evaluiert und mit einem Tutor-geführten Training verglichen. Es wurde eine strukturierte, stufenweise Anleitung zur Schilddrüsensonografie mit einer Vielzahl von Beispiel-Sonografien in verschiedenen Schnittbildebenen gestaltet, die der Nutzer parallel am Probanden mit dem Ultraschallgerät nachstellt. Der Informationsgehalt des Tutorials und die Nachvollziehbarkeit der Untersuchungsschritte wurden positiv eingeschätzt. Die Übersichtlichkeit, die Eignung für Anwender mit fehlender Erfahrung und somit auch der Lernerfolg wurden hingegen unterschiedlich bewertet. Das Tutorial zum Erlernen und zum Training der Schilddrüsensonografie ist ein Instrument zum Selbstlernen und zur Verbesserung der praktischen Ausbildung. In der nächsten Version wird die Untersuchungsanleitung noch detaillierter aufgebaut sein.

Accumulation of CD3(+) T-cell receptor (TCR)αβ(+)CD4(-)CD8(-) double-negative T cells (DNT) is a hallmark of autoimmune lymphoproliferative syndrome (ALPS). DNT origin and differentiation pathways remain controversial. Here we show that human ALPS DNT have features of terminally differentiated effector memory T cells reexpressing CD45RA(+) (TEMRA), but are CD27(+)CD28(+)KLRG1(-) and do not express the transcription factor T-bet. This unique phenotype was also detected among CD4(+) or CD8(+) ALPS TEMRA cells. T-cell receptor β deep sequencing revealed a significant fraction of shared CDR3 sequences between ALPS DNT and both CD4(+) and CD8(+)TEMRA cells. Moreover, in ALPS patients with a germ line FAS mutation and somatic loss of heterozygosity, in whom biallelic mutant cells can be tracked by absent Fas expression, Fas-negative T cells accumulated not only among DNT, but also among CD4(+) and CD8(+)TEMRA cells. These data indicate that in human Fas deficiency DNT cannot only derive from CD8(+), but also from CD4(+) T cells. Furthermore, defective Fas signaling leads to aberrant transcriptional programs and differentiation of subsets of CD4(+) and CD8(+) T cells. Accumulation of these cells before their double-negative state appears to be an important early event in the pathogenesis of lymphoproliferation in ALPS patients.

Adoptive immunotherapy against viral infections is a promising treatment option for patients after hematopoietic stem cell transplantation. However, the generation of virus-specific T cells is either cost-intensive or time-consuming. We developed the first GMP-compliant protocol to generate donor-derived adenovirus (HAdV), cytomegalovirus, and Epstein-Barr virus-specific T-cell lines (TCLs) within 12 days by the use of overlapping polypeptides derived from different viruses in combination with IL-15. Two patients after undergoing haploidentical hematopoietic stem cell transplantation with HAdV viremia displaying rising viral loads despite treatment with cidofovir received 1×10 donor-derived short-term expanded HAdV-specific TCLs per kg body weight. In both patients, HAdV-specific T cells could be detected by IFN-γ-ELISpot 30 and 22 days postinfusion, and resulted in complete clearance or >1.5 log reduction of viral load within 15 and 18 days, respectively. This protocol facilitates rapid and cost-effective generation of virus-specific TCLs, which appear to provide an effective treatment option.

The degree and type of T cell infiltration influence rectal cancer prognosis regardless of classical tumor staging. We asked whether clonal expansion and tumor infiltration are restricted to selected-phenotype T cells; which clones are accessible in peripheral blood; and what the spatial distribution of their target antigens is. From five rectal cancer patients, we isolated paired tumor-infiltrating T cells (TILs) and T cells from unaffected rectum mucosa (T(UM)) using 13-parameter FACS single cell index sorting. TCRαβ sequences, cytokine, and transcription factor expression were determined with single cell sequencing. TILs and T(UM) occupied distinct phenotype compartments and clonal expansion predominantly occurred within CD8(+) T cells. Expanded TIL clones identified by paired TCRαβ sequencing and exclusively detectable in the tumor showed characteristic PD-1 and TIM-3 expression. TCRβ repertoire sequencing identified 49 out of 149 expanded TIL clones circulating in peripheral blood and 41 (84%) of these were PD-1(-) TIM-3(-). To determine whether clonal expansion of predominantly tumor-infiltrating T cell clones was driven by antigens uniquely presented in tumor tissue, selected TCRs were reconstructed and incubated with cells isolated from corresponding tumor or unaffected mucosa. The majority of clones exclusively detected in the tumor recognized antigen at both sites. In summary, rectal cancer is infiltrated with expanded distinct-phenotype T cell clones that either i) predominantly infiltrate the tumor, ii) predominantly infiltrate the unaffected mucosa, or iii) overlap between tumor, unaffected mucosa, and peripheral blood. However, the target antigens of predominantly tumor-infiltrating TIL clones do not appear to be restricted to tumor tissue.

Electroconvulsive therapy (ECT) is an established treatment option for major depressive disorder when other treatments have failed. However, the underlying mechanisms responsible for these therapeutical effects are insufficiently understood to date. Furthermore, treatment outcome is difficult to predict. Recent research suggested an important role of autonomic modulation for successful treatment. We aimed to examine putative associations between autonomic modulation and response to ECT treatment and hypothesized a role for vagal modulation prior to therapy. Twenty-four patients with MDD who received ECT were assessed by means of heart rate and blood pressure variability analysis as well as baroreflex sensitivity measurements before, during and after a course of ECT. Autonomic parameters from the complete study population revealed that ECT did not significantly alter basic autonomic modulation after six sessions. Analyses showed a significant association of the reduction of HAMD scores during therapy when compared with baseline autonomic function as reflected in SDNN(RR) (p

The aim was to determine the patients’ characteristics, comorbidity, and inpatient treatment features of very old otorhinolaryngology patients (80+ years) compared to old patients (75–79 years). A single-center cohort study in a tertiary and university care center was performed with 144 old and 143 very old patients who were hospitalized in 2012. Predictors for differences between old and very old patients were analyzed univariately and multivariately using regression models. Ear (30 %) and nose/paranasal sinus (23 %) diseases were the most frequent reasons for hospitalization. Baseline and disease characteristics were not different between the two groups of elderly patients. Duration of hospitalization was no longer in very old patients (p = 0.827). Mobility (p = 0.017), dietary intake (p = 0.017), and having hearing aid (p

ObjectiveWe propose that the fetal heart is highly resilient to hypoxic stress. Our objective was to elucidate the human fetal gene expression profile in response to simulated ischemia and reperfusion to identify molecular targets that account for the innate cardioprotection exhibited by the fetal phenotype.MethodsPrimary cultures of human fetal cardiac myocytes (gestational age, 15–20 weeks) were exposed to simulated ischemia and reperfusion in vitro by using a simulated ischemic buffer under anoxic conditions. Total RNA from treated and baseline cells were isolated, reverse transcribed, and labeled with Cy3 or Cy5 and hybridized to a human cDNA microarray for expression analysis. This analysis revealed a highly significant (false discovery rate

The global myocardial stress response during cardiac surgery has not been systematically studied, nor is it known whether the response of the neonatal myocardium is intrinsically different from that of older children. To determine the age-related molecular basis of this response, we conducted microarray-based differential gene expression profiling on right ventricular tissue samples acquired in patients of varying ages with right ventricular outflow tract obstruction.We studied gene expression profiles in 24 patients during operations for lesions involving right ventricular outflow tract obstruction age stratified into group I (7 patients, aged 5 to 66 days; mean, 30 days) and group II (17 patients, aged 4 months to 12.5 years; mean, 2.8 years). Myocardial samples were taken from the right ventricular outflow tract after aortic occlusion and archived in liquid nitrogen. RNA isolation, fluorescence labeling of complementary DNA, hybridization to spotted arrays containing 19,008 characterized or unknown human complementary DNAs, and quantitative fluorescence scanning of gene-expression intensity were performed at the University of Toronto Health Network Microarray Centre. Data were analyzed with the Significance Analysis for Microarrays program. Minimum Information About Microarray Experiments-compliant, log2-normalized data sets were compared to ascertain potential statistical differences in gene expression between patient groups.There were no hospital deaths or major postoperative morbid events. We identified 50 transcripts differentially expressed in the neonatal group (the predicted false discovery rate was0.8 transcripts). The neonatal pattern of gene expression (group I) was dominated by genes with literature-validated cardioprotective, antihypertrophic, and antiproliferative properties, including increases in atrial natriuretic peptide, protein phosphatase 2A, small GTPase rap1, and protein inhibitor of activated STAT protein, PIASy. Several transcripts have not been previously reported in heart.Neonatal myocardium has a unique pattern of gene expression, which may result from developmental (age-related) differences or reflect a more severe disease phenotype independent of age effects per se. The neonatal transcript profile seems to reflect a stress-induced protective program composed of genes with functions diametrically opposed to those expected to be related to the pathogenesis of critical right ventricular outflow tract obstruction, thus revealing a novel and compensatory antidisease transcriptional response in the neonatal heart.

This study was undertaken to evaluate the antiadhesive properties of a polymeric agent in infants undergoing staged surgical correction of congenital heart abnormalities.Sixteen infants having staged surgical repair were treated with a polymeric matrix at the completion of the initial surgery. There were 5 untreated controls. The tenacity and extent of adhesions at five separate regions of the heart were evaluated at the follow-up surgery.For all sites combined, there was a threefold difference in median tenacity scores in favor of the experimental treatment (1.0 vs 3.0, p0.01). Significant differences were achieved separately at the right ventricle and the anterior surface of the great vessels (p = 0.02 for both comparisons). Analysis of adhesion scores reflecting the extent of adhesions similarly favored the experimental treatment for all sites (80 vs 270, p0.01), with significant differences persisting at the right atrium (p0.01) and the anterior surface of the great vessels (p = 0.04). There were no treatment-related adverse events.Use of this polymeric agent at the completion of open cardiac surgery may prevent the occurrence or reduce the severity of pericardial adhesions.

Objectives To measure autonomic nervous system function after idiopathic sudden sensorineural hearing loss (ISSHL). Study design Diagnostic prospective cohort single-center study. Setting Tertiary referral university hospital. Patients Twenty-three adult patients with ISSHL and 10 normal-hearing control patients without ISSHL (CON) matched with respect to age, sex, hypertension, and medication. Main outcome measures Bivariate analysis of autonomic regulation (ISSHL versus CON) using 30-minute heart rate (HR) and systolic and diastolic blood pressure (BP) time series at baseline, based on cardiovascular coupling, HR and BP regulatory patterns, high-resolution coupling analysis based on joint symbolic dynamics (High-Resolution Joint Symbolic Dynamics). Intervention No intervention. Results Multivariate discrimination between ISSHL and CON achieved values of area under the receiver operator characteristic curve = 95.5, sensitivity = 90.9%, and specificity = 88.9%. Independent from medication and hypertension increased the complexity of nonlinear HR regulation and reduced cardiovascular coupling of ISSHL patients and independent from hypertension altered nonlinear systolic and diastolic BP regulation. Coupling patterns are characterized by a less pronounced strong and fast decrease of systolic BP when HR increases and rapidly changes in ISSHL patients. Disturbed BP modulation and complexity by impaired baroreflex activities resulting in short-term BP fluctuations, altered peripheral resistance, and reduced cochlear blood flow. Increased values for the pulse wave velocity in the aorta and carotid-femoral were early indicators that the elasticity of the arteries might be restricted in ISSHL patients. Conclusion ISSHL patients show an altered autonomic regulation. At least a subgroup of ISSHL patients seems to exist where a vascular impairment might play an important role in the pathogenesis of this disease.

Background: Reactivation of CMV and EBV negatively impacts on outcome after allogeneic stem cell transplantation (aSCT). Specific antiviral therapy is only available for CMV. With the exception of ganciclovir all drugs are being used off-label. 40-50% of patients reactivate CMV following aSCT. For the 20-30% of patients reactivating EBV, only rituximab is available to control EBV. Rituximab leads to long term B-cell depletion requiring frequent administration of immunoglobulins. To cover the unmet medical need of CMV and EBV control after aSCT, we investigate a somatic cell therapy approach by means of CMV- and EBV-specific peptide-stimulated T cells. We have set up a prospective randomized controlled phase I/IIa multi-center clinical trial to evaluate the preventive and preemptive adoptive transfer of this ATMP in patients after aSCT (EudraCT number 2012-004240-30). The multi-center trial is currently recruiting. Methods: For manufacturing of the cell product two peptide pools (CMV and EBV) each covering 17 well-defined HLA class I and class II epitopes for stimulation of donor derived PBMC are used. PBMC collected by leukapheresis of mobilized or non-mobilized donors can be used as starting material. To avoid a second leukapheresis of the donor, CMV- and EBV-specific T cells are preferentially expanded from a small fraction of the stem cell graft. A strong expansion of virus-specific T cells could be observed in the cell products as determined by HLA class I multimer analysis. Reconstitution and cell counts of leukocytes after aSCT are monitored for both treatment and control group. To obtain further insights in the expansion of transferred T cells, the TCR beta (TCRb) repertoire of the T-cell product before and after adoptive transfer in the patient is monitored by high throughput sequencing. Specificities of TCRb sequences can be assigned by determining the repertoire of HLA/peptide-multimer-sorted CD8+ T cells. New virus-specific TCRb sequences can be identified thereby. Furthermore, TCR sequences within the T-cell product can be tracked in the patient. Study design: After recruitment patients are randomized in intervention or control group. Patients of the intervention group receive three applications of virus specific T cells (5x10e4/kg bodyweight) starting the first adoptive transfer 30 days after allogeneic stem cell transplantation. Cells are transferred as preventive, preemptive, or also as therapeutic treatment. Patients are monitored for occurrence of GvHD, for viral load, as well as for immune reconstitution, especially of virus-specific T cells. Results: So far, 10 patients have been randomized. The reconstitution of virus-specific T cells of treated patients looks encouraging after transfer so far. The immunomonitoring of six included patients is completed. New CMV- and EBV-specific TCRb sequences could be identified and tracked. Conclusion: Our first observations show promising results regarding feasibility and efficacy of our approach under clinical trial conditions. Disclosures Hennig: HS Diagnomics GmbH: Employment, Equity Ownership.

Objective: Establishment of Fontan circulation in complex univentricular hearts often requires several surgical procedures. We developed a procedure which maintains the advantages of a staged approach, however, during the initial surgery additional preparatory measures are performed to allow subsequent non-surgical Fontan completion. Methods: The operation is a lateral baffle Fontan procedure. The baffle bears multiple perforations to allow the inferior vena cava blood to drain into the systemic atrium. Total cavopulmonary connection is performed as usual and the cardiac end of the superior vena cava is subtotally banded. Formally the operation establishes a bi-directional Glenn physiology. During subsequent catheter intervention the banding of the superior vena cava is dilated and the holes in the baffle are closed with appropriate devices. Results: From April 1994 to December 1995, 18 children having at least two risk factors for Fontan operation received the above described operation. Ages ranged from 3 months to 15 years. Ten patients had one or more previous operations. Bypass time ranged from 86 to 128 min and cross clamp time from 14 to 79 min. O 2 saturation after discontinuation of cardiopulmonary bypass was 76% (70-81%). The postoperative recovery of all patients was rapid with early extubation (mean 6 h) and discharge to the ward the morning of the first postoperative day. One patient died. No fluid retention as pericardial, pleural or abdominal fluid effusions occurred. At discharge O 2 saturation was 77% (75-82%). In thirteen children successful conversion to total cavopulmonary connection with interventional debanding of the superior vena cava and closure of the fenestrations was performed. After a hospital stay of only a couple of days the children were discharged with normal O 2 saturation after Fontan completion. Conclusions: We suggest that this modification of the staged Fontan procedure reduces the need for surgical interventions by applying balloon angioplasty and occluder technology to this unique subset of patients.

The causes of idiopathic sudden sensorineural hearing loss (SSNHL) are still unclear while SSNHL seems to be a multicausal disease. To date limited information about autonomic regulation and, especially, cardiovascular coupling (CVC) are available for those patients. The objective of this study was to characterize short-term (30min) CVC in 23 SSNHL patients in comparison to 23 healthy age and gender matched normal hearing control subjects (CON). Further on, the results from CVC should be compared with those from standard heart rate variability (HRV) and blood pressure variability (BPV) analyses. The results showed that HRV is not affected by the disease whereas BPV analysis revealed significant differences between both groups (p

Previous studies have proposed varying causes for idiopathic sudden sensorineural hearing loss (SSNHL), including vascular occlusion, ruptured inner ear membrane, acoustic tumours and circulatory disturbances in the inner ear. The objective of this study was to characterise the autonomic regulation in 19 SSNHL patients in comparison to 19 healthy age-gender matched normal-hearing control subjects (CON) in order to improve the diagnostics of vascular caused hearing loss in SSNHL patients. A high-resolution short-term electrocardiogram (ECG) and the continuous noninvasive blood pressure signal were simultaneously recorded under resting conditions (30min). Linear and nonlinear indices of heart rate- and blood pressure variability (HRV, BPV) were calculated to characterise autonomic regulation. The results showed that HRV analysis did not produce significantly different results between SSNHL and CON, whereas linear and nonlinear BPV indices showed significant differences between both groups (p