Molecular background of three lethal fetal syndromes

This study identified the molecular defects underlying three lethal fetal syndromes. Lethal Congenital Contracture Syndrome 1 (LCCS1, MIM 253310) and Lethal Arthrogryposis with Anterior Horn Cell Disease (LAAHD, MIM 611890) are fetal motor neuron diseases. They affect the nerve cells that control voluntary muscle movement, and eventually result in severe atrophy of spinal cord motor neurons and fetal immobility. Both LCCS1 and LAAHD are caused by mutations in the GLE1 gene, which encodes for a multifunctional protein involved in posttranscriptional mRNA processing. LCCS2 and LCCS3, two syndromes that are clinically similar to LCCS1, are caused by defective proteins involved in the synthesis of inositol hexakisphosphate (IP6), an essential cofactor of GLE1. This suggests a common mechanism behind these fetal motor neuron diseases, and along with accumulating evidence from genetic studies of more late-onset motor neuron diseases such as Spinal muscular atrophy (SMA) and Amyotrophic lateral sclerosis (ALS), implicates mRNA processing as a common mechanism in motor neuron disease pathogenesis. We also studied gle1-/- zebrafish in order to investigate whether they would be a good model for studying the pathogenesis of LCCS1 and LAAHD. Mutant zebrafish exhibit cell death in their central nervous system at two days post fertilization, and the distribution of mRNA within the cells of mutant zebrafish differs from controls, encouraging further studies. The third lethal fetal syndrome is described in this study for the first time. Cocoon syndrome (MIM 613630) was discovered in a Finnish family with two affected individuals. Its hallmarks are the encasement of the limbs under the skin, and severe craniofacial abnormalities, including the lack of skull bones. We showed that Cocoon syndrome is caused by a mutation in the gene encoding the conserved helix-loop-helix ubiquitous kinase CHUK, also known as IκB kinase α (IKKα). The mutation results in the complete lack of CHUK protein expression. CHUK is a subunit of the IκB kinase enzyme that inhibits NF-κB transcription factors, but in addition, it has an essential, independent role in controlling keratinocyte differentiation, as well as informing morphogenetic events such as limb and skeletal patterning. CHUK also acts as a tumor suppressor, and is frequently inactivated in cancer. This study has brought significant new information about the molecular background of these three lethal fetal syndromes, as well as provided knowledge about the prerequisites of normal human development.

On formation, growth and concentrations of air ions

Aerosol particles have effect on climate, visibility, air quality and human health. However, the strength of which aerosol particles affect our everyday life is not well described or entirely understood. Therefore, investigations of different processes and phenomena including e.g. primary particle sources, initial steps of secondary particle formation and growth, significance of charged particles in particle formation, as well as redistribution mechanisms in the atmosphere are required. In this work sources, sinks and concentrations of air ions (charged molecules, cluster and particles) were investigated directly by measuring air molecule ionising components (i.e. radon activity concentrations and external radiation dose rates) and charged particle size distributions, as well as based on literature review. The obtained results gave comprehensive and valuable picture of the spatial and temporal variation of the air ion sources, sinks and concentrations to use as input parameters in local and global scale climate models. Newly developed air ion spectrometers (Airel Ltd.) offered a possibility to investigate atmospheric (charged) particle formation and growth at sub-3 nm sizes. Therefore, new visual classification schemes for charged particle formation events were developed, and a newly developed particle growth rate method was tested with over one year dataset. These data analysis methods have been widely utilised by other researchers since introducing them. This thesis resulted interesting characteristics of atmospheric particle formation and growth: e.g. particle growth may sometimes be suppressed before detection limit (~ 3 nm) of traditional aerosol instruments, particle formation may take place during daytime as well as in the evening, growth rates of sub-3 nm particles were quite constant throughout the year while growth rates of larger particles (3-20 nm in diameter) were higher during summer compared to winter. These observations were thought to be a consequence of availability of condensing vapours. The observations of this thesis offered new understanding of the particle formation in the atmosphere. However, the role of ions in particle formation, which is not well understood with current knowledge, requires further research in future.

Arrested Drunk Drivers : Trends, social background, recidivism and mortality

The aim of this study was to examine the trends, incidence and recidivism of drunken driving during a 20-year period (1988 - 2007) using the data on all suspected drunken driving in this period. Furthermore, the association between social background and drunken driving, and the mortality of drunk drivers were studied by using administrative register data provided by Statistics Finland. The study was completely register-based. In 1989 - 1991, every year 30,000 drivers were suspected of drunken driving, but the number fell to less than 20,000 by 1994, during the economic recession. The changes in the arrest incidence of the youngest age groups were especially pronounced, most of all in the age group of 18 - 19-year olds. Even though the incidence among youth decreased dramatically, their incidence rate was still twice that of the general population aged 15 - 84 years. Drunken driving was associated with a poor social background among youth and working-aged men and women. For example, a low level of education, unemployment, divorce, and parental factors in youth were associated with a higher risk of being arrested for drunken driving. While a low income was related to more drunken driving among working-aged people, the effect among young persons was the opposite. Every third drunk driver got rearrested during a 15-year period, whereas the estimated rearrest rate was 44%. Findings of drugs only or in combination with alcohol increased the risk of rearrest. The highest rearrest rates were seen among drivers who were under the influence of amphetamines or cannabis. Also male gender, young age, high blood alcohol concentration, and arrest during weekdays and in the daytime predicted rearrest. When compared to the general population, arrested drunk drivers had significant excess mortality. The greatest relative differences were seen in alcohol-related causes of death (including alcohol diseases and alcohol poisoning), accidents, suicides and violence. Also mortality due to other than alcohol-related diseases was elevated among drunk drivers. Drunken driving was associated with multiple factors linked to traffic safety, health and social problems. Social marginalization may expose a person to harmful use of alcohol and drunken driving, and the associations are seen already among the youth. Recidivism is common among drunk drivers, and driving under the influence of illicit and/or medicinal drugs is likely to indicate worse substance abuse problems, judging from the high rearrest rates. High alcohol-related mortality in this population shows that drunken driving is clearly an indicator of alcohol abuse. More effective measures of preventing alcohol-related harms are needed, than merely preventing convicted drunk drivers from driving again.

Hostility: A prospective study of the genetic and environmental background and associations with cardiovascular risk

Hostility is a multidimensional construct having wide effects on society. In its different forms, hostility is related to a large array of social and health problems, such as criminality, substance abuse, depression, and cardiovascular risks. Identifying and tackling early-life factors that contribute to hostility may have public health significance. Although the variance in hostility is estimated to be 18-50 percent heritable, there are significant gaps in knowledge regarding the molecular genetics of hostility. It is known that a cold and unsupportive home atmosphere in childhood predicts a child s later hostility. However, the long-term effects of care-giving quality on hostility in adulthood and the role of genes in this association are unclear. The present dissertation is part of the ongoing population-based prospective Young Finns study, which commenced in 1980 with 3596 3-18-year-old boys and girls who were followed for 27 years. The specific aims of the dissertation were first to study the antecedents of hostility by looking at 1) the genetic background, 2) the early environmental predictors, and 3) the gene environment interplay behind hostility. As a second aim, the thesis endeavored to examine 4) the association between hostility and cardiovascular risks, and 5) the moderating effect of demographic factors, such as gender and socioeconomic status, on this association. The study found potential gene polymorphisms from chromosomes 7, 14, 17, and 22 suggestively associated with hostility. Of early environmental influences, breastfeeding and early care-giving were found to predict hostility in adulthood. In addition, a serotonin receptor 2A polymorphism rs6313 moderated the effect of early care-giving on later hostile attitudes. Furthermore, hostility was shown to predict cardiovascular risks, such as metabolic syndrome and inflammation. Finally, parental socioeconomic status was found to moderate the association between anger and early atherosclerosis. The new genetic and early environmental antecedents of hostility identified in this research may help in understanding the development of hostility and its health risks, and in planning appropriate prevention. The significance of early influences on this development is stressed. Although the markers studied are individual- and family-related factors, these may be influenced at the societal level by giving accurate information to all individuals concerned and by improving the societal circumstances.

Development of atmospheric pressure ionization ion mobility spectrometry and ion mobility spectrometry mass spectrometry

This study is focused on the development and evaluation of ion mobility instrumentation with various atmospheric pressure ionization techniques and includes the following work. First, a high-resolution drift tube ion mobility spectrometer (IMS), coupled with a commercial triple quadrupole mass spectrometer (MS), was developed. This drift tube IMS is compatible with the front-end of commercial Sciex mass spectrometers (e.g., Sciex API-300, 365, and 3000) and also allows easy (only minor modifications are needed) installation between the original atmospheric pressure ion source and the triple quadrupole mass spectrometer. Performance haracteristics (e.g.,resolving power, detection limit, transmission efficiency of ions) of this IMS-MS instrument were evaluated. Development of the IMS-MS instrument also led to a study where a proposal was made that tetraalkylammonium ions can be used as chemical standards for ESI-IMS. Second, the same drift tube design was also used to build a standalone ion mobility spectrometer equipped with a Faraday plate detector. For this highresolution (resolving power about 100 shown) IMS device, a multi-ion source platform was built, which allows the use of a range of atmospheric pressure ionization methods, such as: corona discharge chemical ionization (CD-APCI), atmospheric pressure photoionization (APPI), and radioactive atmospheric pressure chemical ionization (R-APCI). The multi-ion source platform provides easy switching between ionization methods and both positive and negative ionization modes can be used. Third, a simple desorpion/ionization on silicon (DIOS) ion source set-up for use with the developed IMS and IMS-MS instruments was built and its operation demonstrated. Fourth, a prototype of a commercial aspiration-type ion mobility spectrometer was mounted in front of a commercial triple quadrupole mass spectrometer. The set-up, which is simple, easy to install, and requires no major modifications to the MS, provides the possibility of gathering fundamental information about aspiration mobility spectrometry.

Dissection of the genetic background of childhood onset progressive myoclonic epilepsies

The progressive myoclonic epilepsies (PMEs) are a clinically and etiologically heterogeneous group of symptomatic epilepsies characterized by myoclonus, tonic-clonic seizures, psychomotor regression and ataxia. Different disorders have been classified as PMEs. Of these, the group of neuronal ceroid lipofuscinoses (NCLs) comprise an entity that has onset in childhood, being the most common cause of neurodegeneration in children. The primary aim of this thesis was to dissect the molecular genetic background of patients with childhood onset PME by studying candidate genes and attempting to identify novel PME-associated genes. Another specific aim was to study the primary protein properties of the most recently identified member of the NCL-causing proteins, MFSD8. To dissect the genetic background of a cohort of Turkish patients with childhood onset PME, a screen of the NCL-associated genes PPT1, TPP1, CLN3, CLN5, CLN6, MFSD8, CLN8 and CTSD was performed. Altogether 49 novel mutations were identified, which together with 56 mutations found by collaborators raised the total number of known NCL mutations to 364. Fourteen of the novel mutations affect the recently identified MFSD8 gene, which had originally been identified in a subset of mainly Turkish patients as the underlying cause of CLN7 disease. To investigate the distribution of MFSD8 defects, a total of 211 patients of different ethnic origins were evaluated for mutations in the gene. Altogether 45 patients from nine different countries were provided with a CLN7 molecular diagnosis, denoting the wide geographical occurrence of MFSD8 defects. The mutations are private with only one having been established by a founder-effect in the Roma population from the former Czechoslovakia. All mutations identified except one are associated with the typical clinical picture of variant late-infantile NCL. To address the trafficking properties of MFSD8, lysosomal targeting of the protein was confirmed in both neuronal and non-neuronal cells. The major determinant for this lysosomal sorting was identified to be an N-terminal dileucine based signal (9-EQEPLL-14), recognized by heterotetrameric AP-1 adaptor proteins, suggesting that MFSD8 takes the direct trafficking pathway en route to the lysosomes. Expression studies revealed the neurons as the primary cell-type and the hippocampus and cerebellar granular cell layer as the predominant regions in which MFSD8 is expressed. To identify novel genes associated with childhood onset PME, a single nucleotide polymorphism (SNP) genomewide scan was performed in three small families and 18 sporadic patients followed by homozygosity mapping to determine the candidate loci. One of the families and a sporadic patient were positive for mutations in PLA2G6, a gene that had previously been shown to cause infantile neuroaxonal dystrophy. Application of next-generation sequencing of candidate regions in the remaining two families led to identification of a homozygous missense mutation in USP19 for the first and TXNDC6 for the second family. Analysis of the 18 sporadic cases mapped the best candidate interval in a 1.5 Mb region on chromosome 7q21. Screening of the positional candidate KCTD7 revealed six mutations in seven unrelated families. All patients with mutations in KCTD7 were reported to have early onset PME, rapid disease progression leading to dementia and no pathologic hallmarks. The identification of KCTD7 mutations in nine patients and the clinical delineation of their phenotype establish KCTD7 as a gene for early onset PME. The findings presented in this thesis denote MFSD8 and KCTD7 as genes commonly associated with childhood onset symptomatic epilepsy. The disease-associated role of TXNDC6 awaits verification through identification of additional mutations in patients with similar phenotypes. Completion of the genetic spectrum underlying childhood onset PMEs and understanding of the gene products functions will comprise important steps towards understanding the underlying pathogenetic mechanisms, and will possibly shed light on the general processes of neurodegeneration and nervous system regulation, facilitating the diagnosis, classification and possibly treatment of the affected cases.

Oral malodour – background and diagnostics

Bad breath or oral malodour can be related to gingival diseases, trimethylaminuria, various inflammation diseases of upper respiratory tract, foreign bodies in nasal cavity etc. Bad breath is usually, in 85 % to 95 % of cases, inflicted by gram negative anaerobic bacteria in tongue coating. These bacteria have a tendency of producing foul-smelling sulphur containing gases called volatile sulphur compounds or VSC. Main cause of bad breath is parodontitis or postnasal drip into posterior part of the tongue. Detecting bad breath is most efficiently done by organoleptic method. By skilled analyser the reason for oral malodour can be determined with great accuracy. For scientific study the most effective method is gas chromatography (GC) with flame photometric detector (FPD). With it almost every component of exhaled air can be detected both quantitative and qualitative. Effective chairside methods include portable sulphur monitors and saliva tests.