Generation and transmission of mtDNA mutations and their effect on aging

Mutations in mtDNA are known to cause neuromuscular diseases and have been associated with several common age-associated diseases as well as being a contributor to the aging process. Despite their importance there are still important aspects of mtDNA inheritance and mtDNA mutations that remain unknown and need to be addressed. The aim of this thesis is to examine how mtDNA mutations behave in in vivo systems with regard to inheritance and distribution. A further aim is to investigate the phenotypic consequences of mtDNA mutations, specifically in the aging process. For this purpose we used mouse models and different approaches for DNA sequence analysis. Our group previously demonstrated that increased somatic mtDNA mutagenesis can cause premature aging in the mouse, thus providing the first experimental data for the involvement of mtDNA mutations in aging. This work was based on analysis of the mtDNA mutator mouse, which is a knock- in mouse model with increased mtDNA mutation load due to a proofreading deficiency of mitochondrial DNA polymerase The work presented in this thesis demonstrates that a significant proportion of mtDNA mutations are passed on via the maternal germline and expand clonally in the offspring to contribute significantly to the observed phenotypes. By performing a variety of mouse crosses to generate mice with different combinations of maternally inherited and somatic mtDNA mutations we found that even low levels of inherited mtDNA mutations can lead to premature aging phenotypes. These phenotypes could be rescued by reintroduction of wild-type mtDNA. Our findings show that clonal expansion of maternally transmitted mtDNA mutations can be an important factor in the aging process. This finding is very intriguing in light of recent findings that inherited heteroplasmy is common in humans. Furthermore, a combination of inherited and somatic mtDNA mutations can disturb development to cause stochastic brain malformations. There is a lack of understanding how pathogenic mtDNA mutations are inherited and studies in the area have been hampered by the lack of appropriate animal models. By backcrossing mtDNA mutator females we were able to isolate a mouse line carrying a single base pair deletion in the mitochondrial tRNAMet gene. Although this mutation was under selection in living offspring, we failed to observe any clear selection in germ cells and primary oocytes. The selection is instead occurring after fertilization as the embryo develops. Finally, we investigated the clonality of mtDNA inheritance. Mutations in mtDNA are the most commonly used marker in population genetics studies, and much of this popularity is based on the assumed lack of recombination in mtDNA. This assumption has now been questioned in a number of studies that report frequent recombination of mtDNA. In this thesis, I developed a novel direct cloning protocol to trap single mtDNA molecules, allowing for DNA sequence analysis without prior amplification by PCR. Applying this technique, there was no evidence for germline recombination of mtDNA in mice that has been heteroplasmic for >50 generations

The importance of fat and alcohol for progression and prognosis in chronic liver disease

Chronic liver disease is an increasing cause of global morbidity and mortality. The popular belief is that liver disease is caused mainly by alcoholic liver disease or viral hepatitis. However, the most common cause of chronic liver disease today is non-alcoholic fatty liver disease (NAFLD), which is associated with obesity and insulin resistance rather than alcohol. NAFLD is considered to become the most common cause for need of liver transplantation in the coming years. Today, the most common cause of liver transplantation in Sweden is primary sclerosing cholangitis (PSC) - a rare but very serious disease of the bile ducts that become inflamed and obliterated, and is associated with a high risk of development of cholangiocarcinoma. The role of concurrent use of alcohol in NAFLD and PSC is controversial. Part of this thesis explores the effect of alcohol on the degree of liver damage in these two diseases. We found that a low consumption of alcohol, around one unit per day, is not associated with a higher stage of fibrosis in the liver in PSC and should be safe in these patients. For NAFLD, we found that a low to moderate consumption of alcohol was associated with a lower risk for a higher fibrosis stage, up to thirteen units of alcohol per week. However, patients who had biochemical evidence of high alcohol consumption had a higher risk of more severe liver damage. This is well in line with other studies and indicates a J-formed risk profile for alcohol consumption in NAFLD. In another part of the thesis we studied the long-term risk of having fat accumulation in the liver and if overweight per se can predict development of severe liver disease. We found that the strongest histological marker for disease-specific mortality in NAFLD after a follow-up of in mean 26 years was the stage of fibrosis, and found no excess mortality in patients with signs of inflammation in the liver after adjustment for the stage of fibrosis. The risk of being overweight was studied in close to 45.000 men in their late adolescence who were conscribed to military service in 1969-1970 after adjustment of potential confounders, such as alcohol consumption. Body mass index (BMI) was found to be an independent predictor of development of severe liver disease after a mean follow-up of 39 years. Taken together, this thesis indicates that a low to moderate consumption of alcohol is safe in PSC and possibly protective in NAFLD. Furthermore, we found that the strongest predictor of disease-specific mortality in NAFLD is the stage of fibrosis, which can have implications for the design of endpoints in future clinical studies. Also, the finding that overweight per se is a predictor for development of severe liver disease is important for public health decisionmaking

Summer life - memories, dreams and materiality

Kerstin Gunnemark (ed.) 2016. Sommarliv. Minnen, drömmar och materialitet. Göteborg: Makadam. 335 pp. Ill. ISBN: 978-91-7061-218-3

The Exemption Status of the Bona Fide Pledgee of Unregistered Securities Under the Securities Act of 1933

The number of connections of photovoltaic (PV) to distribution network is increasing. Very few PV connection guidelines that distribution system operators (DSOs) can refer to have been found. This paper deals with network planning guidelines for distribution networks with PV. The paper aims to identify planning rules that are relatively easy to implement.QC 20140625</p