Longitudinal confocal microscopy imaging of solid tumor destruction following adoptive T cell transfer

A fluorescence-based, high-resolution imaging approach was used to visualize longitudinally the cellular events unfolding during T cell-mediated tumor destruction. The dynamic interplay of T cells, cancer cells, cancer antigen loss variants, and stromal cells—all color-coded in vivo—was analyzed in established, solid tumors that had developed behind windows implanted on the backs of mice. Events could be followed repeatedly within precisely the same tumor region—before, during and after adoptive T cell therapy—thereby enabling for the first time a longitudinal in vivo evaluation of protracted events, an analysis not possible with terminal imaging of surgically exposed tumors. T cell infiltration, stromal interactions, and vessel destruction, as well as the functional consequences thereof, including the elimination of cancer cells and cancer cell variants were studied. Minimal perivascular T cell infiltrates initiated vascular destruction inside the tumor mass eventually leading to macroscopic central tumor necrosis. Prolonged engagement of T cells with tumor antigen-crosspresenting stromal cells correlated with high IFNγ cytokine release and bystander elimination of antigen-negative cancer cells. The high-resolution, longitudinal, in vivo imaging approach described here will help to further a better mechanistic understanding of tumor eradication by T cells and other anti-cancer therapies

The Two-Component Signal Transduction System CopRS of Corynebacterium glutamicum Is Required for Adaptation to Copper-Excess Stress

Copper is an essential cofactor for many enzymes but at high concentrations it is toxic for the cell. Copper ion concentrations ≥50 µM inhibited growth of Corynebacterium glutamicum. The transcriptional response to 20 µM Cu2+ was studied using DNA microarrays and revealed 20 genes that showed a ≥ 3-fold increased mRNA level, including cg3281-cg3289. Several genes in this genomic region code for proteins presumably involved in the adaption to copper-induced stress, e. g. a multicopper oxidase (CopO) and a copper-transport ATPase (CopB). In addition, this region includes the copRS genes (previously named cgtRS9) which encode a two-component signal transduction system composed of the histidine kinase CopS and the response regulator CopR. Deletion of the copRS genes increased the sensitivity of C. glutamicum towards copper ions, but not to other heavy metal ions. Using comparative transcriptome analysis of the ΔcopRS mutant and the wild type in combination with electrophoretic mobility shift assays and reporter gene studies the CopR regulon and the DNA-binding motif of CopR were identified. Evidence was obtained that CopR binds only to the intergenic region between cg3285 (copR) and cg3286 in the genome of C. glutamicum and activates expression of the divergently oriented gene clusters cg3285-cg3281 and cg3286-cg3289. Altogether, our data suggest that CopRS is the key regulatory system in C. glutamicum for the extracytoplasmic sensing of elevated copper ion concentrations and for induction of a set of genes capable of diminishing copper stress

The role of open abdomen in non-trauma patient : WSES Consensus Paper

The open abdomen (OA) is defined as intentional decision to leave the fascial edges of the abdomen un-approximated after laparotomy (laparostomy). The abdominal contents are potentially exposed and therefore must be protected with a temporary coverage, which is referred to as temporal abdominal closure (TAC). OA use remains widely debated with many specific details deserving detailed assessment and clarification. To date, in patients with intra-abdominal emergencies, the OA has not been formally endorsed for routine utilization; although, utilization is seemingly increasing. Therefore, the World Society of Emergency Surgery (WSES), Abdominal Compartment Society (WSACS) and the Donegal Research Academy united a worldwide group of experts in an international consensus conference to review and thereafter propose the basis for evidence-directed utilization of OA management in non-trauma emergency surgery and critically ill patients. In addition to utilization recommendations, questions with insufficient evidence urgently requiring future study were identified.Peer reviewe

Research and Design of a Routing Protocol in Large-Scale Wireless Sensor Networks

无线传感器网络，作为全球未来十大技术之一，集成了传感器技术、嵌入式计算技术、分布式信息处理和自组织网技术，可实时感知、采集、处理、传输网络分布区域内的各种信息数据，在军事国防、生物医疗、环境监测、抢险救灾、防恐反恐、危险区域远程控制等领域具有十分广阔的应用前景。 本文研究分析了无线传感器网络的已有路由协议，并针对大规模的无线传感器网络设计了一种树状路由协议，它根据节点地址信息来形成路由，从而简化了复杂繁冗的路由表查找和维护，节省了不必要的开销，提高了路由效率，实现了快速有效的数据传输。 为支持此路由协议本文提出了一种自适应动态地址分配算——ADAR（AdaptiveDynamicAddre...As one of the ten high technologies in the future, wireless sensor network, which is the integration of micro-sensors, embedded computing, modern network and Ad Hoc technologies, can apperceive, collect, process and transmit various information data within the region. It can be used in military defense, biomedical, environmental monitoring, disaster relief, counter-terrorism, remote control of haz...学位：工学硕士院系专业：信息科学与技术学院通信工程系_通信与信息系统学号：2332007115216

Broadband Multi-wavelength Properties of M87 during the 2017 Event Horizon Telescope Campaign

Abstract: In 2017, the Event Horizon Telescope (EHT) Collaboration succeeded in capturing the first direct image of the center of the M87 galaxy. The asymmetric ring morphology and size are consistent with theoretical expectations for a weakly accreting supermassive black hole of mass ∼6.5 × 109 M ⊙. The EHTC also partnered with several international facilities in space and on the ground, to arrange an extensive, quasi-simultaneous multi-wavelength campaign. This Letter presents the results and analysis of this campaign, as well as the multi-wavelength data as a legacy data repository. We captured M87 in a historically low state, and the core flux dominates over HST-1 at high energies, making it possible to combine core flux constraints with the more spatially precise very long baseline interferometry data. We present the most complete simultaneous multi-wavelength spectrum of the active nucleus to date, and discuss the complexity and caveats of combining data from different spatial scales into one broadband spectrum. We apply two heuristic, isotropic leptonic single-zone models to provide insight into the basic source properties, but conclude that a structured jet is necessary to explain M87’s spectrum. We can exclude that the simultaneous γ-ray emission is produced via inverse Compton emission in the same region producing the EHT mm-band emission, and further conclude that the γ-rays can only be produced in the inner jets (inward of HST-1) if there are strongly particle-dominated regions. Direct synchrotron emission from accelerated protons and secondaries cannot yet be excluded

Optimierung von retroviralem Gentransfer in T-Lymphocyten für die Neuausrichtung von T-Zellspezifität

TITLE AND CONTENTS SUMMARY 1 ZUSAMMENFASSUNG 2 GENERAL INTRODUCTION 3 HIGH-LEVEL GENE EXPRESSION IN T CELLS 30 ENGINEERING RCC-REACTIVE CTL 45 DISCUSSION 58 APPENDIX 78 ABBREVIATIONS 80 REFERENCES 83 ACKNOWLEDGEMENTS 100 PUBLICATIONS 101 EIDESSTATTLICHE ERKLÄRUNG 102Adoptive T cell therapy is a promising approach for the treatment of cancer. Donor lymphocyte infusions in patients with cytomegalovirus-mediated disease, Epstein-Barr virus-positive B cell lymphomas, and chronic myelogenous leukaemia have proven the efficacy of transferred T cells to kill infected or malignant cells. The main hindrance of using this approach for the cure of other tumour entities is the often unsuccessful isolation and amplification of tumour-reactive T cells from patients. A potential method to circumvent this problem is the transfer of anti-tumour specificity to peripheral blood lymphocytes by retroviral T cell receptor (TCR) gene transfer. In this thesis I have compared several retroviral vectors regarding their gene transfer potential. For this purpose the green fluorescent protein (GFP) was cloned into vectors containing different cis-regulatory elements. Promoter and enhancer elements of two different long terminal repeats (LTR) and a cellular gene were compared. The LTR originating from the myeloproliferative sarcoma virus (MPSV) was shown to be superior to the Moloney murine leukaemia virus (MoMLV) LTR and a truncated MoMLV LTR/CD2 enhancer combination. Furthermore, the impact of a modified leader sequence and the Woodchuck hepatitis virus post-transcriptional regulatory element (PRE) was analysed. These elements showed an enhancing effect on transgene expression and retroviral titre, respectively. Taken together, the MP71 retrovirus containing the MPSV LTR, the modified leader, and the PRE demonstrated strong and durable transgene expression in murine and human T lymphocytes. High transduction rates were obtained when using retroviruses pseudotyped with an ecotropic MoMLV or the amphotropic 10A1 MLV envelope for murine or human cells, respectively. The same results were obtained after exchanging GFP with TCR α and β -chain genes, linked by an internal ribosomal entry site. The redirected primary human T lymphocytes showed the same specificity as the tumour infiltrating lymphocyte (TIL)-26 from which the TCR originated. Furthermore, the intensity of anti- tumour reactivity, measured as cytokine release and tumour cell lysis, was comparable for both the TCR-26-grafted T cells and the TIL-26. The TCR expression, analysed for a period of 100 days, remained stable. This approach enables us to generate functional renal cell carcinoma (RCC)-specific T lymphocytes by TCR gene transfer. These data provide the rationale for adoptive T cell therapy for RCC.Der adoptive T-Zelltransfer ist eine viel versprechende Form der Tumortherapie. Die Infusion von Donor T-Lymphozyten für die Behandlung von Virusinfektionen (Cytomegalovirus und Epstein-Barr-Virus) und einigen wenigen Krebsarten hat sich als sehr effektiv erwiesen. Für die meisten Tumorarten ist jedoch die Isolierung und in vitro Vermehrung von Tumor-spezifischen T-Zellen nicht möglich. Eine Abhilfe für dieses Problem kann der Transfer von anti- Tumorspezifität schaffen, wie zum Beispiel der retrovirale Transfer von T-Zell Rezeptor (TCR)-Genen in periphere Blut-Lymphozyten (PBL). In dieser Arbeit habe ich verschiedene retrovirale Vektoren bezüglich ihrer Gentransfereffizienz in T-Lymphozyten analysiert. Dazu wurde das grün fluoreszierende Protein (GFP) in Vektoren mit verschiedenen cis- regulatorischen Elementen kloniert. Das Promotor/"Enhancer" Paar des "long terminal repeats" (LTR) des myeloproliferativen Sarkom-Virus (MPSV) war dabei dem Paar des LTR des Moloney-Maus-Leukämie-Virus (MoMLV) und einem Paar aus dem MoMLV Promotor und dem "Enhancer" des CD2-Gens überlegen. Der Einsatz einer modifizierten "leader"-Sequenz und des posttranskriptionellen regulatorischen Elements (PRE) des Waldmurmeltier-Hepatitis-B-Virus führte zu einer Erhöhung der Transgenexpression bzw. des Virustiters. Die Expressionsuntersuchungen ergaben, dass die Kombination aus der LTR des MPSV, der modifizierten "leader"-Sequenz und des PRE geeignet ist, um Gene in Maus- und humanen T-Zellen in hohem Maße und für längere Zeit zu exprimieren. Des weiteren wurde gezeigt, dass sich das ecotrope MoMLV, bzw. das amphotrope 10A1 -MLV-Hüllprotein, gut eignen, um Maus-, bzw. humane T-Zellen, zu transduzieren. Der Austausch des GFP durch TCR α und β -Ketten-Gene, verbunden durch eine interne Ribosomen-Eintrittsstelle, ergab ähnliche Expressionsergebnisse. Der transferierte TCR-26 wurde aus dem Nierenzellkarzinom-spezifischen Tumor-infiltrierenden Lymphozyten Klon-26 gewonnen. Die TCR-26-modifizierten T-Zellen zeigten dieselbe Spezifität wie der Ursprungsklon. Auch war die anti-Tumorreaktivität, gemessen als Zytokinfreisetzung und Zelllyse, der beiden Zellpopulationen vergleichbar. Die über einen Zeitraum von 100 Tagen analysierte TCR-Expression war stabil. Diese Ergebnisse zeigen, dass der Transfer von anti-Nierenzellkarzinomspezifität auf primäre T-Zellen möglich ist, und somit ein adoptiver T-Zelltransfer als Nierenzellkarzinomtherapie denkbar