An Overview of the Isochronets Architecture for High Speed Networks

This paper overviews a novel switching architecture for high-speed networks: Isochronets. Isochronets time-divide network bandwidth among routing trees. Traffic moves down a routing tree to the root during its time band. Network functions such as routing and flow control are entirely governed by band timers and require no processing of frame headers bits. Frame motions need not be delayed for switch processing, allowing Isochronets to scale over a large spectrum of transmission speeds and support all-optical implementations. The network functions as a media-access layer that can support multiple framing protocols simultaneously, handled by higher layers at the periphery. Internetworking is reduced to a simple media-layer bridging. Isochronets provide flexible quality of service control and multicasting through allocation of bands to routing trees. They can be tuned to span a spectrum of performance behaviors outperforming both circuit or packet switching

Analytic modelling and resource dimensioning of optical burst switched networks

The realisation of optical network architectures may hold the key to delivering the enormous bandwidth demands of next generation Internet applications and services. Optical Burst Switching (OBS) is a potentially cost-effective switching technique that can satisfy these demands by offering a high bit rate transport service that is bandwidth-efficient under dynamic Internet traffic loads. Although various aspects of OBS performance have been extensively investigated, there remains a need to systematically assess the cost/performance trade-offs involved in dimensioning OBS switch resources in a network. This goal is essential in enabling the future deployment of OBS but poses a significant challenge due to the complexity of obtaining tractable mathematical models applicable to OBS network optimisation. The overall aim of this thesis lies within this challenge. This thesis firstly develops a novel analytic performance model of an OBS node where burst contention is resolved by combined use of Tuneable Wavelength Converters (TWCs) and Fibre Delay Lines (FDLs) connected in an efficient share-per-node configuration. The model uses a two-moment traffic representation that gives a good trade-off between accuracy and complexity, and is suitable for extension to use in network modelling. The OBS node model is then used to derive an approximate analytic model of an OBS network of switches equipped with TWCs and FDLs, again maintaining a two-moment traffic model for each end-to-end traffic path in the network. This allows evaluation of link/route loss rates under different offered traffic characteristics, whereas most OBS network models assume only a single-moment traffic representation. In the last part of this thesis, resource dimensioning of OBS networks is performed by solving single and multi-objective optimisation problems based on the analytic network model. The optimisation objectives relate to equipment cost minimisation and throughput maximisation under end-to-end loss rate constraints. Due to non-convexity of the network performance constraint equations, a search heuristic approach has been taken using a constraint-handling genetic algorithm

A Slotted Ring Test Bed for the Study of ATM Network Congestion Management

This thesis addresses issues raised by the proposed Broadband Integrated Services Digital Network which will provide a flexible combination of integrated services traffic through its cell-based Asynchronbus Transport Mode (ATM). The introduction of a cell-based, connection-oriented, transport mode brings with it new technical challenges for network management. The routing of cells, their service at switching centres, and problems of cell congestion not encountered in the existing network, are some of the key issues. The thesis describes the development of a hardware slotted ring testbed for the investigation of congestion management in an ATM network. The testbed is designed to incorporate a modified form of the ORWELL protocol to control media access. The media access protocol is analysed to give a model for maximum throughput and reset interval under various traffic distributions. The results from the models are compared with measurements carried out on the testbed, where cell arrival statistics are also varied. It is shown that the maximum throughput of the testbed is dependent on both traffic distribution and cell arrival statistics. The testbed is used for investigations in a heterogeneous traffic environment where two classes of traffic with different cell arrival statistics and quality of service requirements are defined. The effect of prioritisation, media access protocol, traffic intensity, and traffic source statistics were investigated by determining an Admissible Load Region (ALR) for a network station. Conclusions drawn from this work suggest that there are many problems associated with the reliable definition of an ALR because of the number of variable parameters which could shift the ALR boundary. A suggested direction for further work is to explore bandwidth reservation and the concept of equivalent capacity of a connection, and how this can be linked to source control parameters

Wavelength reconfigurability for next generation optical access networks

Next generation optical access networks should not only increase the capacity but also be able to redistribute the capacity on the fly in order to manage larger variations in traffic patterns. Wavelength reconfigurability is the instrument to enable such capability of network-wide bandwidth redistribution since it allows dynamic sharing of both wavelengths and timeslots in WDM-TDM optical access networks. However, reconfigurability typically requires tunable lasers and tunable filters at the user side, resulting in cost-prohibitive optical network units (ONU). In this dissertation, I propose a novel concept named cyclic-linked flexibility to address the cost-prohibitive problem. By using the cyclic-linked flexibility, the ONU needs to switch only within a subset of two pre-planned wavelengths, however, the cyclic-linked structure of wavelengths allows free bandwidth to be shifted to any wavelength by a rearrangement process. Rearrangement algorithm are developed to demonstrate that the cyclic-linked flexibility performs close to the fully flexible network in terms of blocking probability, packet delay, and packet loss. Furthermore, the evaluation shows that the rearrangement process has a minimum impact to in-service ONUs. To realize the cyclic-linked flexibility, a family of four physical architectures is proposed. PRO-Access architecture is suitable for new deployments and disruptive upgrades in which the network reach is not longer than 20 km. WCL-Access architecture is suitable for metro-access merger with the reach up to 100 km. PSB-Access architecture is suitable to implement directly on power-splitter-based PON deployments, which allows coexistence with current technologies. The cyclically-linked protection architecture can be used with current and future PON standards when network protection is required

Dynamics and Heterogeneity of Gene Expression and Epigenetic Regulation at the Single-Cell Level

The ability of cells to establish and remember their gene expression states is a cornerstone of multicellular life. This thesis explores how gene expression states are regulated dynamically, and how these regulations differ in individual cells even under the same conditions. These properties underlie cellular state decisions and often determine the balance between different cell types in a multicellular system, but are typically inaccessible to conventional techniques that rely on static snapshots and population averaging. We address these issues in two separate contexts, one natural and one synthetic, using time-lapse imaging and other single-cell techniques. In the first context, we use embryonic stem cells (ES), which were shown to exist in a mixed population of at least two cellular states with distinct differentiation propensities, as a model to study natural dynamics of cellular states. These cells display rare, stochastic, and spontaneous transitions between the two states, as well as more frequent fluctuations in gene expression levels within each state. Our system enables us to further investigate how these dynamics are modulated under a cell signaling environment that enhances pluripotency, and the role DNA methylation plays in maintaining these states. In the second context, we investigate how chromatin regulators (CRs), part of a complex system that enables cells to modulate gene expression and epigenetic memory, operate dynamically in individual cells. We build a synthetic platform to measure the isolated effect of recruitment and de-recruitment of four individual CRs. In contrast to conventional transcription factor control, all CRs tested regulate gene expression in all-or-none events, controlling the probability of stochastic transitions between fully active and silent states rather than the strength of gene expression. The qualitative and quantitative responses of a cell population are determined by the set of event rates associated with each CR, as well as the duration of CR recruitment. These results provide a framework for understanding and engineering chromatin-based cellular states and their dynamics. </p

Functional Impact of Preleukemic Mutations on the Human Hematopoietic Stem and Progenitor Cell Compartment

Dissecting functional consequences of gene mutations contributes to understanding disease development and identifying potential treatment strategies. Mutations in epigenetic regulators (especially in DNMT3A, TET2, and ASXL1, termed DTA mutations) have been identified in acute myeloid leukemia patients, as well as individuals without history of hematologic malignancies. Mutations in these genes are hypothesized to confer a clone a competitive advantage, leading to preferential expansion of this specific clone. DTA mutations are proposed to be preleukemic events that are acquired before leukemia onset and are thought to elevate the risk for acquisition of additional mutations that in turn drive malignant transformation. Clones harboring these mutations are able to survive chemotherapy, potentially increasing the risk for disease recurrence. In this thesis, I investigated the impact of DTA mutations in the human hematopoietic stem and progenitor cell compartment, as well as the role of preleukemic events in patients with AML t(8;21). Using CRISPR/Cas9, site-specific mutations were introduced in CD34+ progenitor cells freshly isolated from umbilical cord blood samples. By delivering Cas9 nuclease together with an in vitro transcribed sgRNA as ribonucleoprotein, I was able to achieve editing efficiencies ranging from 40% for the sgRNA targeting exon 13 of ASXL1 to over 90% for the sgRNA targeting exon 6 of TET2. A known hotspot mutation in DNMT3A R882H was introduced with an efficiency of up to 50% in the cell bulk. After successful genetic modification, cells were subjected to in vitro assays to assess phenotype, self-renewal properties, differentiation capacities, and long-term culture-initiating potential. Clonal composition and expansion were assessed via deep sequencing and subsequent analysis of introduced specific insertions and deletions. To investigate the role and importance of preleukemic mutations in AML leukemogenesis, a comprehensive genetic in-depth characterization of AML t(8;21) was performed. To this aim, DNA from the time of diagnosis was analyzed via next generation sequencing with a targeted gene panel including exons and coding regions of 66 genes recurrently mutated in hematologic malignancies. From 56 patients, samples from clinical remission were analyzed for the previously identified mutations by targeted re-sequencing of amplicons spanning the target region. In this thesis, I was able to demonstrate that mutations in DNMT3A, TET2, and ASXL1 influence the behavior of hematopoietic stem and progenitor cells (HSPCs) in a gene-specific fashion. Mutations in ASXL1 had only little effect on HSPCs with no observed changes in expression of differentiation markers after culture for three weeks and no enhanced proliferation in long-term culture. The only hint that ASXL1 mutations have an impact on the fitness of HSPCs was increased colony forming ability after long-term culture, meaning that although progenitor cells did not proliferate during the long-term culture period, they were able to sustain and generate colonies later. DNMT3A mutations led to a longer retention of CD34 marker expression in differentiation culture experiments, but not enhanced serial replating capacity. TET2-mutated cells, in contrast, showed delayed myeloid marker expression in short-term culture and increased self-renewal of committed progenitor cells. Both, DNMT3A and TET2 mutations led to clonal expansion of distinct cell clones during long-term culture with a competitive advantage over wild type cells. The clonal composition of TET2- and DNMT3A-mutated samples differed, with TET2mut samples being more diverse. Collectively my data indicate that DNMT3A mutations preferentially influence primitive HSC clones, whereas TET2 mutations enhance the fitness of committed progenitor cell clones. ASXL1 mutations might need additional events to drive clonal expansion. Analysis of patient samples revealed that mutations in DNMT3A and TET2 are early events in AML t(8;21) and potentially acquired before the onset of the disease. ASXL1 mutations are neither early nor late events in these patients but are potentially cooperating events in leukemia transformation. In this thesis, I was able to demonstrate that DTA mutations enhance the fitness of distinct human hematopoietic stem or progenitor cell clones, in terms of self-renewal and proliferative capacity, leading to clonal expansion. The results obtained here highlight the importance of monitoring specific preleukemic events to prevent disease recurrence as well as development of severe hematologic conditions.Um die Entwicklung von Krankheiten aufzudecken und etwaige zielgerichtete Behandlungsstrategien zu entwickeln, ist es essenziell den funktionellen Einfluss bestimmter Genmutationen zu untersuchen. Mutationen in epigenetischen Regulatoren wie DNMT3A, TET2 und ASXL1 (auch DTA-Mutationen genannt) wurden sowohl in Patienten mit akuter myeloischer Leukämie als auch in Patienten ohne hämatologische Erkrankung gefunden. Diese Mutationen führen zu einem Vorteil bestimmter Zellklone, die daraufhin expandieren. DTA-Mutationen werden auch als prä-leukämische Mutationen bezeichnet, die vor dem Ausbruch der Leukämie bereits in Stamm- und Vorläuferzellen vorkommen und ein erhöhtes Risiko für das Auftreten weiterer Mutationen bedingen. Zusätzliche Mutationen in einem prä-leukämischen Klon können wiederum zur malignen Transformation des Zellklons führen. Des Weiteren können Zellen mit prä-leukämischen Mutationen resistent gegen Chemotherapie sein und somit das Risiko eines Rezidivs erhöhen. In dieser Arbeit habe ich den Einfluss von Mutationen in DNMT3A, TET2 und ASXL1 auf humane hämatopoetische Stamm- und Vorläuferzellen untersucht, sowie die Rolle von prä-leukämischen Mutationen in Patienten mit AML t(8;21) erforscht. Mit Hilfe von CRISPR/Cas9 wurden spezifische Mutationen in CD34-positiven Vorläuferzellen eingeführt, die frisch aus Nabelschnurblut isoliert wurden. Die Cas9-Nuklease wurde zusammen mit in vitro-transkribierter sgRNA in Form von Ribonukleoproteinen in die Zellen gebracht. In Exon 13 von ASXL1 wurden Insertionen und Deletionen in mindestens 40% der Zellen generiert und in Exon 6 des Gens TET2 konnten in mehr als 90% der Zellen Insertionen und Deletionen entdeckt werden. Die spezifische Mutation DNMT3A R882H wurde in bis zu 50% der transfizierten Zellen eingeführt. Die erfolgreich mutierten Zellen wurden anschließend in in vitro-Experimenten untersucht in Hinsicht auf Veränderungen im Phänotyp, der Fähigkeit zur Selbsterneuerung, Differenzierungsfähigkeit und der Fähigkeit sich in Langzeitkulturen zu vermehren. Die klonale Zusammensetzung, sowie die Expansion bestimmter Zellklone wurden mit Hilfe von Deep Sequencing und anschließender Analyse der generierten Insertionen und Deletionen untersucht. Patienten mit AML t(8;21) wurden auf prä-leukämische Mutationen untersucht, indem DNA vom Diagnosezeitpunkt via Next-Generation Sequenzierung analysiert wurde. Kodierende Regionen von 66 Genen, die wiederholt in malignen hämatologischen Erkrankungen auftreten, wurden in die Analyse eingeschlossen. Von 56 Patienten wurden DNA-Proben vom Zeitpunkt der klinischen Remission erneut auf die bereits identifizierten Mutationen untersucht. In dieser Arbeit konnte ich zeigen, dass Mutationen in DNMT3A, TET2 und ASXL1 unterschiedliche Effekte auf hämatopoetische Stamm- und Vorläuferzellen haben. Mutationen in ASXL1 hatten den mildesten Effekt. Weder Unterschiede in der Expression von Differenzierungsmarkern noch erhöhte Replatierungsfähigkeit in Vergleich zu Wildtypzellen wurden beobachtet. Des Weiteren zeigten die Zellen kein vermehrtes Wachstum in der Langzeitkultur. Sie waren jedoch in der Lage Kolonien nach der Langzeitkultur zu bilden, was bedeuten könnte, dass diese Zellen trotz des unveränderten Proliferationsverhaltens einen Vorteil gegenüber Wildtypzellen haben und somit in der Lage sind die Langzeitkultur zu überdauern. Zellen mit DNMT3A Mutationen exprimierten den Vorläuferzellmarker CD34 länger als Kontrollzellen. Mutationen in TET2 führten zu einer verspäteten Expression von myeloischen Differenzierungsmarkern und einem erhöhten Selbsterneuerungspotential von Vorläuferzellen. Sowohl in Zellen mit DNMT3A- als auch mit TET2-Mutationen konnte ich die Expansion bestimmter mutierter Zellklone nachweisen. Die Proben mit eingeführten Mutationen in TET2 hatten jedoch eine größere Anzahl verschiedener Insertionen und Deletionen als Proben mit DNMT3A-Veränderungen. Die Daten dieser Arbeit zeigen, dass Mutationen in DNMT3A dazu führen, dass primitive Zellklone einen evolutionären Vorteil erhalten, wohingegen reifere Vorläuferzellen von Mutationen in TET2 profitieren. Mutationen in ASXL1 führen möglicherwiese nur in Anwesenheit weiterer genetischer oder zellulärer Veränderungen zu klonaler Expansion. In Patienten mit AML t(8;21) treten Mutationen in DNMT3A und TET2 wahrscheinlich früher auf als andere Mutationen und kommen möglicherweise in Vorläuferzellen vor, bevor sich die eigentliche Erkrankung entwickelt. Mutationen in ASXL1 scheinen weder frühe noch späte Ereignisse zu sein, sondern treten irgendwann dazwischen auf. Sie kooperieren möglicherweise mit anderen Mutationen, um Zellen zu transformieren. In dieser Arbeit konnte ich zeigen, dass DTA-Mutationen die Fitness von bestimmten Stamm- und Vorläuferzellen im menschlichen Stammzellkompartiment erhöhen und so zu einer Expansion der mutierten Zellklone führen. Die Ergebnisse dieser Arbeit bestätigen, dass die Untersuchung und Beobachtung prä-leukämischer Klone dazu beitragen könnte, ein Rezidiv oder sogar die Entstehung schwerer hämatologische Erkrankungen zu verhindern