Evolution of Bordetella pertussis post vaccination

Evolution of Bordetella pertussis post vaccination Whooping cough or pertussis is caused by the gram-negative bacterium Bordetella pertussis. It is a highly contiguous disease in the human respiratory tract. Characteristic of pertussis is a paroxysmal cough with whooping sound during gasps of breath after coughing episodes. It is potentially fatal to unvaccinated infants. The best approach to fight pertussis is to vaccinate. Vaccinations against pertussis have been available from the 1940s. Traditionally vaccines were whole-cell pertussis (wP) preparations as part of the combined diphtheria-tetanus-pertussis (DTP) vaccines. More recently acellular pertussis (aP) vaccines have replaced the wP vaccines in many countries. The aP vaccines are less reactogenic and can also be administered to school children and adults. There are several publications reporting variation in the i>B. pertussis virulence factors that are also aP vaccine antigens. This has occurred in the genes coding for pertussis toxin and pertactin about 15 to 30 years after the introduction of pertussis vaccines to immunisation programs. Resurgence of pertussis has also been reported in many countries with high vaccination coverage. In this study the evolution of B. pertussis was investigated in Finland, the United Kingdom, Poland, Serbia, China, Senegal and Kenya. These represent countries with a long history of high vaccination coverage with stable vaccines or changes in the vaccine formulation; countries which established high vaccination coverage late; and countries where vaccinations against pertussis were started late. With bacterial cytotoxicity and cytokine measurements, comparative genomic hybridisation, pulsed-field gel electrophoresis (PFGE), genotyping and serotyping it was found that changes in the vaccine composition can postpone the emergence of antigenic variants. It seems that the change in PFGE profiles and the loss of genetic material in the genome of B. pertussis are similar in most countries and the vaccine-induced immunity is selecting non-vaccine type strains. However, the differences in the formulation of the vaccines, the vaccination programs and in the coverage of pertussis vaccination have affected the speed and timing of these changes.

Quantitative refinement of reaction-based biomodels

In the field of molecular biology, scientists adopted for decades a reductionist perspective in their inquiries, being predominantly concerned with the intricate mechanistic details of subcellular regulatory systems. However, integrative thinking was still applied at a smaller scale in molecular biology to understand the underlying processes of cellular behaviour for at least half a century. It was not until the genomic revolution at the end of the previous century that we required model building to account for systemic properties of cellular activity. Our system-level understanding of cellular function is to this day hindered by drastic limitations in our capability of predicting cellular behaviour to reflect system dynamics and system structures. To this end, systems biology aims for a system-level understanding of functional intraand inter-cellular activity. Modern biology brings about a high volume of data, whose comprehension we cannot even aim for in the absence of computational support. Computational modelling, hence, bridges modern biology to computer science, enabling a number of assets, which prove to be invaluable in the analysis of complex biological systems, such as: a rigorous characterization of the system structure, simulation techniques, perturbations analysis, etc. Computational biomodels augmented in size considerably in the past years, major contributions being made towards the simulation and analysis of large-scale models, starting with signalling pathways and culminating with whole-cell models, tissue-level models, organ models and full-scale patient models. The simulation and analysis of models of such complexity very often requires, in fact, the integration of various sub-models, entwined at different levels of resolution and whose organization spans over several levels of hierarchy. This thesis revolves around the concept of quantitative model refinement in relation to the process of model building in computational systems biology. The thesis proposes a sound computational framework for the stepwise augmentation of a biomodel. One starts with an abstract, high-level representation of a biological phenomenon, which is materialised into an initial model that is validated against a set of existing data. Consequently, the model is refined to include more details regarding its species and/or reactions. The framework is employed in the development of two models, one for the heat shock response in eukaryotes and the second for the ErbB signalling pathway. The thesis spans over several formalisms used in computational systems biology, inherently quantitative: reaction-network models, rule-based models and Petri net models, as well as a recent formalism intrinsically qualitative: reaction systems. The choice of modelling formalism is, however, determined by the nature of the question the modeler aims to answer. Quantitative model refinement turns out to be not only essential in the model development cycle, but also beneficial for the compilation of large-scale models, whose development requires the integration of several sub-models across various levels of resolution and underlying formal representations.

Autologous Stem Cell Transplantation In Multiple Myeloma

Background. Multiple myeloma (MM) is the second most common hematologic malignancy after lymphomas In Finland: the annual incidence of MM is approximately 200. For three decades the median survival remained at 3 to 4 years from diagnosis until high-dose melphalan treatment supported by autologous stem cell transplantation (ASCT) became the standard of care for newly diagnosed MM since the mid 1990’s and the median survival increased to 5 – 6 years. This study focuses on three important aspects of ASCT, namely 1) stem cell mobilization, 2) single vs. double ASCT as initial treatment, and 3) the role of minimal residual disease (MRD) for longterm outcome. Aim. The aim of this series of studies was to evaluate the outcomes of MM patients and the ASCT procedure at the Turku University Central Hospital, Finland. First, we tried to identify which factors predict unsuccessful mobilization of autologous stem cells. Second, we compared the use of short-acting granulocyte-colony stimulating factor (GCSF) with long-acting G-CSF as mobilization agents. Third, one and two successive ASCTs were compared in 100 patients with MM. Fourth, for patients in complete response (CR) after stem cell transplantation (SCT), patient-specific probes for quantitative allele-specific oligonucleotide polymerase-chain reaction (qASO-PCR) measurements were designed to evaluate MRD and its importance for long-term outcome. Results. The quantity of previous chemotherapy and previous interferon use were significant pre-mobilization factors that predicted mobilization failure, together with some factors related to mobilization therapy itself, such as duration and degree of cytopenias and occurrence of sepsis. Short-acting and long-acting G-CSF combined with chemotherapy were comparable as stem cells mobilizers. The progression free (PFS) and overall survival (OS) tended to be longer after double ASCT than after single ASCT with a median follow-up time of 4 years, but this difference disappeared as the follow-up time increased. qASO-PCR was a good and sensitive divider of the CR patients into two prognostic groups: MRD low/negative (≤ 0.01%) and MRD high (>0.01%) groups with a significant difference in PFS and suggestively also in OS. Conclusions. When the factors prediciting a poor outcome of stem cell mobilization prevail, it is possible to identify those patients who need specific efforts to maximize the mobilization efficacy. Long-acting pegfilgrastim is a practical and effective alternative to short-acting filgrastim for mobilization therapy. There is no need to perform double ASCT on all eligible patients. MRD assessment with qASO-PCR is a sensitive method for evaluation of the depth of the CR response and can be used to predict long-term outcome after ACST.

Molecular markers for progression of squamous cell carcinoma of the skin

Incidence of nonmelanoma skin cancer (NMSC) is increasing. Ultraviolet (UV) –light is a major risk factor for the development of cutaneous SCC. Cutaneous SCCs that develop to chronic ulcers are known to progress and metastasize more easily than UV-induced SCCs. Matrix metalloproteinases (MMPs) are a group of proteolytic enzymes which are suggested to have a role in cancer growth and invasion. The molecular background for progression of cutaneous SCC was examined by immunohistochemistry (IHC) using tissue samples of recessive dystrophic epidermolysis bullosa (RDEB) –associated SCC, sporadic UV-induced SCC, and SCC precursors. IHC studies using tissue microarray (TMA) technique revealed overexpression of MMP-7 and MMP-13 in SCC tumor cells. MMP-7 expression was enhanced especially in the SCC tumor cells of the RDEB –associated SCCs. Studies with SCC cell lines showed that tumor cell derived MMP-7 activated heparin binding epidermal growth factor –like growth factor (HB-EGF) which enhanced the growth of SCC tumor cells. Further, it was shown that type VII collagen (COL7) is expressed in sporadic SCC tumor cells. Interestingly, it was shown that SCC –associated MMP-13 is capable of cleaving COL7 in vitro. COL7 cleavage may have a role in the progression of cutaneous SCC. Studies on serine proteinase inhibitor gene family using SCC tumor cell gene array, quantitative real-time PCR, SCC cell lines, normal human epidermal keratinocytes and IHC of TMA samples showed that serine proteinase inhibitor clade A, member 1 (serpinA1, alpha-1-antitrypsin) is expressed and produced by human SCC tumor cells but not by normal keratinocytes. Moreover, serpinA1 expression was shown to correlate with the progression of cutaneous SCC using transformed HaCaT-cell lines and mouse chemically induced skin SCC model. SerpinA1 may serve as a novel biomarker for the progression of cutaneous SCC. This study elucidated putative mechanisms of the progression of cutaneous SCC and revealed novel biomarker candidates for the progression of SCC of the skin.

Battery Cell Modeling for Battery Management System

Usage of batteries as energy storage is emerging in automotive and mobile working machine applications in future. When battery systems become larger, battery management becomes an essential part of the application concerning fault situations of the battery and safety of the user. A properly designed battery management system extends one charge cycle of battery pack and the whole life time of the battery pack. In this thesis main objectives and principles of BMS are studied and first order Thevenin’s model of the lithium-titanate battery cell is built based on laboratory measurements. The battery cell model is then verified by comparing the battery cell model and the actual battery cell and its suitability for use in BMS is studied.

Tuning cell motility : roles of nestin and vimentin in cancer cell invasion and motility

The cytoskeleton is a key feature of both prokaryotic and eukaryotic cells. Itis comprised of three protein families, one of which is the intermediate filaments (IFs). Of these, the IFs are the largest and most diverse. The IFs are expressed throughout life, and are involved in the regulation of cell differentiation, homeostasis, ageing and pathogenesis. The IFs not only provide structural integrity to the cell, they are also involved in a range of cellular functions from organelle trafficking and cell migration to signalling transduction. The IFs are highly dynamic proteins, able to respond and adapt their network rapidly in response to intra- and extra- cellular cues. Consequently they interact with a whole host of cellular signalling proteins, regulating function, and activity, and cellular localisation. While the function of some of the better-known IFs such as the keratins is well studied, the understanding of the function of two IFs, nestin and vimentin, is poor. Nestin is well known as a marker of differentiation and is expressed in some cancers. In cancer, nestin is primarily described as is a promoter of cell motility, however, how it fulfils this role remains undefined. Vimentin too is expressed in cancer, and is known to promote cell motility and is used as a marker for epithelial to mesenchymal transition (EMT). It is only in the last decade that studies have addressed the role that vimentin plays in cell motility and EMT. This work provides novel insight into how the IFs, nestin and vimentin regulate cell motility and invasion. In particular we show that nestin regulates the cellular localisation and organisation of two key facilitators of cell migration, focal adhesion kinase and integrins. We identify nestin as a regulator of extracellular matrix degradation and integrin-mediated cell invasion. Two further studies address the specific regulation of vimentin by phosphorylation. A detailed characterisation study identified key phosphorylation sites on vimentin, which are critical for proper organisation of the vimentin network. Furthermore, we show that the bioactive sphingolipids are vimentin network regulators. Specifically, the sphingolipids induced RhoA kinasedependent (ROCK) phosphorylation at vimentin S71, which lead to filament reorganisation and inhibition of cell migration. Together these studies shed new light into the regulation of nestin and vimentin during cell motility.

Gene Regulation in The Human Immune System. Gene Expression Signatures of Th2 Cell Differentiation, Type 1 Diabetes, and Intrauterine Immune Adaptation

The human immune system is constantly interacting with the surrounding stimuli and microorganisms. However, when directed against self or harmless antigens, these vital defense mechanisms can cause great damage. In addition, the understanding the underlying mechanism of several human diseases caused by aberrant immune cell functions, for instance type 1 diabetes and allergies, remains far from being complete. In this Ph.D. study these questions were addressed using genome-wide transcriptomic analyses. Asthma and allergies are characterized by a hyperactive response of the T helper 2 (Th2) immune cells. In this study, the target genes of the STAT6 transcription factor in naïve human T cells were identified with RNAi for the first time. STAT6 was shown to act as a central activator of the genes expression upon IL-4 signaling, with both direct and indirect effects on Th2 cell transcriptome. The core transcription factor network induced by IL-4 was identified from a kinetic analysis of the transcriptome. Type 1 diabetes is an autoimmune disease influenced by both the genetic susceptibility of an individual and the disease-triggering environmental factors. To improve understanding of the autoimmune processes driving pathogenesis in the prediabetic phase in humans, a unique series of prospective whole-blood RNA samples collected from HLA-susceptible children in the Finnish Type 1 Diabetes Prediction and Prevention (DIPP) study was studied. Changes in different timewindows of the pathogenesis process were identified, and especially the type 1 interferon response was activated early and throughout the preclinical T1D. The hygiene hypothesis states that allergic diseases, and lately also autoimmune diseases, could be prevented by infections and other microbial contacts acquired in early childhood, or even prenatally. To study the effects of the standard of hygiene on the development of neonatal immune system, cord blood samples from children born in Finland (high standard of living), Estonia (rapid economic growth) and Russian Karelia (low standard of living) were compared. Children born in Russian Karelia deviated from Finnish and Estonian children in many aspects of the neonatal immune system, which was developmentally more mature in Karelia, resembling that of older infants. The results of this thesis offer significant new information on the regulatory networks associated with immune-mediated diseases in human. The results will facilitate understanding and further research on the role of the identified target genes and mechanisms driving the allergic inflammation and type 1 diabetes, hopefully leading to a new era of drug development.