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γ-Aminobutyric acid (GABA) is the main inhibitory neurotransmitter in the central nervous system. It acts upon two classes of GABA receptors: ionotropic and metabotropic. The GABAB receptor is a metabotropic G protein-coupled receptor (GPCR) that associates with the Gαi/o family of G-proteins causing inhibitory actions via the inhibition of adenylyl cyclase. Baclofen, a GABAB receptor agonist, has recently been found to be a potential treatment of alcohol addiction by suppressing the drug craving stage of the addiction cycle. However, the adverse side effects of baclofen including sedation and hypothermia, as well as its narrow therapeutic index, have limited its clinical use in the treatment. Positive allosteric modulators may be the answer to providing more selective and personalised treatments. These modulators function by enhancing or attenuating the response activated by the agonists, while having little inherent intrinsic activity. Therefore, they are less likely to cause adverse effects while also reducing the dosage of the agonistic drugs, essentially diminishing the adverse effects of the agonist. Mu-opioid receptor antagonists have been available for the treatment of alcohol and drug addiction for over 20 years. Preclinical studies have found that a continuous infusion of naltrexone has led to the increased consumption of ethanol after the drug has been metabolised out of the system. Furthermore, nalmefene, a specific and potent opioid antagonist has been developed and shown to have therapeutic advantages over naltrexone. Our study wanted to compare sex-specific brain-wide distribution of G-protein coupling after activation of GABAB receptors by baclofen and the positive allosteric modulator RacBHFF in ten RccHan®:WIST albino male and female rats, and compare species differences between the rats and five wildtype C57BL/6J male mice. Furthermore, we wanted to determine whether G-protein coupling of GABAB receptors is altered in sixty female AA (Alko, Alcohol) alcohol-preferring rats with and without various modes of treatments with naltrexone and nalmefene (7 daily injections or 7-day continuous infusion using osmotic minipumps at equipotent low and moderate doses). We did this using [35S]GTPγS autoradiography as means to measure the efficacy and potency of different concentrations of RacBHFF combined with 30 µM baclofen. Our results showed that RacBHFF does have positive allosteric activity on baclofen-stimulated GABAB receptor-mediated G-protein activation and the extent of it was regionally dependent. RacBHFF had greatest efficacy in the cerebellum, cortex and thalamus in all the samples studied. Furthermore, 30 µM baclofen alone and 10 µM RacBHFF alone are not enough to produce a statistically significant enhancement in GABAB activity in all brain regions, but RacBHFF combined with baclofen does. High concentrations of RacBHFF (100 and 300 µM) led to the complete inactivation of the receptors with G-protein coupling levels falling below non-specific binding levels, thus indicating tolerance. This was noticed in both of the sexes, as well as both of the species, indicating high system stability. Furthermore, there were no statistically significant differences between the species, and only minor regional statistically significant differences between the sexes. Comparing the regional baclofen-stimulated GABAB receptor-mediated G-protein activation between the two rat models showed that there are slight statistically significant differences, most of which are present in the nucleus accumbens, a part of the mesolimbic dopamine reward pathway. In addition, continuous 7-day infusions of a moderate dose (4 mg/kg) of nalmefene produced the greatest statistically significant enhancement of receptor stimulation by RacBHFF and baclofen when compared to the saline-treated samples. Seven daily injections of the same dose of nalmefene also produced statistically significant RacBHFF and baclofen-stimulated GABAB receptor activation in the amygdala and medial habenular nucleus. Therefore, a moderate dose of nalmefene (either as a continuous infusion or repeated injections) seems to produce the greatest overall enhancement of RacBHFF-induced stimulation of GABAB receptors. In conclusion, our results reveal that depending on the concentration of RacBHFF and the location of the receptors, RacBHFF differs in its efficacy to GABAB receptors in both wildtype and treated animal models. Furthermore, our results provide the first preclinical evidence that in the presence of opioid receptor tolerance, the GABAB receptor remains rather unaltered although the signal transductions of opioid and GABAB receptors are identical. This suggests that a combinational therapy of the GABAB receptor PAMs with a mu-opioid antagonist may potentially enhance the anti-craving, anti-alcohol-consuming habits of alcohol-addicted individuals.

Gastropares är en komplikation vid diabetes och definieras som en fördröjning av magsäckens tömningshastighet. Allmänna symptom är uppsvälldhet, snabb mättnadskänsla, illamående och uppkastning. Gastropares är en av de mest allvarliga komplikationerna vid diabetes och eftersom det tidigare gjorts förhållandevis lite forskning om komplikationen finns det ett intresse av vidare studier i ämnet. Målet med avhandlingen är att analysera sjukdomsbilden vid gastropares hos patienter med typ 1-diabetes. Utöver diagnosen gastropares undersöks också förekomsten av gastrointestinala symptom hos studiepopulationen. Studien utförs på Folkhälsans forskningscenter, som har tillgång till ett omfattande patientmaterial på patienter med typ 1-diabetes via FinnDiane-studien. Analysen gav en prevalens för gastropares på 7,2% och en prevalens för besvärliga gastrointestinala symptom på 7%. Det framkom att patienter med gastropares generellt har betydligt mera gastrointestinala symptom, lider av mera diabetiska komplikationer och att patienterna kan kopplas till en del övriga faktorer som hög ålder, lång diabetesduration, högt blodtryck och lågt BMI. Ytterligare diskuteras problematiken kring diagnostisering av gastropares, bland annat av den orsaken att komplikationen saknar en egen diagnoskod i det vårdanmälningssystem som används för tillfället. Till följd av svår diagnostisering och avsaknad av diagnoskod finns en risk att symptom i mag- och tarmkanalen inte får tillräckligt med uppmärksamhet.

Suomalainen perinnöllinen amyloidoosi on kromosomissa 9 vallitsevasti periytyvä systeeminen amyloidoosi, joka tunnetaan myös nimillä Meretojan ja Kymenlaakson tauti. Taudin kuvasi vuonna 1969 Jouko Meretoja, joka arveli potilaiden yhteisen esi-isän olevan lähtöisin 1300- luvulta Kymenlaaksosta. Taudin aiheuttaa pistemutaatio c.640G > A p.D187N gelsoliini-geenissä. Hiljattain osoitettiin SNP-siru genotyypitykseen perustuen, että suomalaisilla potilailla on todennäköisesti yhteinen esi-isä. Tutkimuksen tavoitteena on arvioida, kuinka monen sukupolven päässä sijaitsee suomalaisten gelsoliiniamyloidoosipotilaiden viimeisin yhteinen esi-isä. Toisena tavoitteena on arvioida uuden perustajahaplotyypin säilymiseen perustuvan iänmääritysmenetelmän toimivuutta. Verifioiduksi kontrollimenetelmäksi tämän rinnalle valittiin laajasti iänmääritykseen käytetty alleeliassosiaation hajoamiseen perustuva menetelmä geenin ympärillä sijaitsevia polymorfisia toistojaksoja hyödyntäen. Polymorfisiin toistojaksoihin perustuva menetelmä arvioi etäisyyden yhteiseen esi-isään olevan noin 31 sukupolvea eli noin 775 vuotta. Tämä ei ollut linjassa perustajahaplotyypin säilymiseen nojaavan menetelmän kanssa, joka arvioi etäisyyden yhteiseen esi-isään olevan noin 79 sukupolven päässä.

Objectives Autism generally refers to lingual, communicational and behavioral continuous traits, which are determined by genetics as well as environment. The purpose of this study was to examine to which extent the genetic autism risk explains the variation in autistic traits in two-year-old children in the normal population. Additionally the impact of the genetic autism risk was examined in conditions, where the mother of the child suffered from depressive symptoms during pregnancy and/or had alexithymia (trouble identifying feelings). Methods The sample was collected from the Child-Sleep cohort study (n=942). The child’s genetic autism risk was calculated by comparing the child’s genome with the genome of people with an autism diagnosis. All of the other information was collected with questionnaires. The child’s autistic traits were assessed according to the autism scale in BITSEA. The mother’s depressive symptoms during pregnancy were assessed according to the Ces-d (short) questionnaire and her trouble identifying feelings according to the TAS-20-questionnaire. The connections between the variables were examined with Pearson’s correlation coefficient. The autistic traits were examined with linear regression analysis. Results and conclusions The genetic autism risk explained 2% of the variation in the autistic traits of two-year-old children in the normal population. This result is in line with previous studies. Gender (autistic traits were more prevalent in males), maternal adolescence and the degree of maternal alexithymia were the best predictors of an autistic phenotype. Generally maternal maturity is considered a risk factor for the child’s more autistic phenotype, whereas our result possibly points out that the risk is rather created by paternal maturity instead. In addition of identifying the greatest risk factors, interactions between the genetic risk of autism and the psychological factors of the mother (depressive symptoms during pregnancy and trouble identifying emotions) were examined. However, these interactions were not statistically significant in this study. The impact of genes is already a verified fact – the next goal is to identify environments, where the genes interact, creating a more autistic phenotype.

Objectives. Maternal diabetes during the pregnancy increases the risk of pregnancy complications, but the effects of maternal diabetes on offspring cognition are less understood. Earlier studies have mainly associated the adverse effects of maternal diabetes with slight deficits in general cognitive and verbal functions in young children. Despite the earlier studies, it is unclear, does maternal diabetes per se affect cognitive development in children and adult offspring. The offspring with several developmental risks seem to be more prone to the adverse effects of maternal diabetes than offspring without the other concomitant risks. The aim of this study was to examine is maternal diabetes associated to lowered offspring general cognitive function in childhood and midlife, when the other concomitant perinatal risks occurred or either did not occur. A hypothesis was that maternal diabetes is associated to lowered general cognitive function only in children who had the other concomitant perinatal risks. Another aim was to explore is there a time related change in the risk groups. Methods. This study is a part of a prospective birth-cohort study originating in Helsinki region that follows 1971 to 1974 born risk group offspring. Out of 22,359 consecutive deliveries at the Institute of Midwifery during that time, 93 offspring had mother's diabetes obtained during the pregnancy or before it. Of the offspring with maternal diabetes, 59 had maternal diabetes as the only risk, and 34 had at least another predefined concomitant risk. General cognitive function in the subjects and controls was assessed by Wechsler Intelligence Scales at 9 and 40 years as a part of the wider neuropsychological examination. Differences between the groups were examined by group and pairwise comparisons. Longitudinal changes in general cognitive function in each group were estimated by fitting the linear multilevel models. Results and conclusions. Findings of the present study were controversial to the hypothesis. Both risk groups, with and without other concomitant risks, had lower general and verbal function in childhood than controls. At midlife, no effect of maternal diabetes was found. The results indicated that general cognitive function and acquired verbal information improved at least in the risk group with the other concomitant risks. Otherwise performance remained relatively same.

The extraembryonic placenta is composed of trophoblast cells consisting of the proliferative cytotrophoblasts (CTB) and its differentiated subtypes syncytiotrophoblast (SCT) and extravillous trophoblast (EVT). A normal trophoblast development is important as disruptions can lead to pregnancy complications such as pre-eclampsia. Therefore, it is crucial to investigate the underlying causes behind these abnormalities to discover treatments for patients suffering from pregnancy related disorders. Previously placental research was conducted largely on animal models and despite shared conservative pathways with humans, there are differences that exist. Only recently have researchers managed to successfully isolate and culture primary trophoblast stem cells (TSC)s by creating a TSC medium. Due to limited access to placental cells, pluripotent stem cells (PSC)s can be differentiated to TSCs by using the TSC medium. Naïve and primed states are described to be PSCs in different developmental stages, the former representing the pre-implantation state and the latter the post-implantation state. There lacks a consensus on whether both PSC states can be used to generate TSCs that correspond to primary trophoblasts. It has been argued that naïve cells possess more potential to differentiate into TSCs compared to the primed ones. The primed cells have been induced with the bone morphogenic protein (BMP) 4 to generate TSCs. This method is controversial as some suggest the induction resulting in other than TSCs, such as amniotic cells. Therefore, the aim of this thesis was to investigate whether both PSC states could be used to generate TSCs and its subtypes, if at all. Further, the effect of BMP4 was examined in the prime- derived differentiation protocol. The generated cells were then characterized and analyzed using imaging, immunocytochemistry (ICC) and quantitative reverse transcription PCR (RT-qPCR). The thesis found that although TSCs and its subtypes could be successfully generated from both PSC states, differences were observed. In addition to morphological differences, the most significant finding was the expression of the HLA-G gene, an EVT-specific marker, in the prime-derived TSCs (TSC(BMP4)). HLA-G was also significantly more expressed in the prime-derived EVTs (EVT(p)) compared to the naïve-derived EVTs (EVT(n)). Further, MMP2 which is also an EVT specific marker, was significantly more expressed in the EVT(n) compared to the EVT(p). As a result, the research question regarding the validity of the TSCs using both methods and the effect of BMP4 remains open. Further studies are required including single-cell RNA sequencing to obtain a better and broader view of the trophoblast profile and functional assays for subtype differentiation. Additionally, the role of BMP4 should be investigated in more depth.

Objectives. Motivational contexts exert a profound influence on behavior biasing actions in sometimes detrimental ways. In Pavlovian bias, reward-predicting conditioned cues elicit approach behavior while aversively associated cues elicit withdrawal, with capacity to impact instrumental goal-driven behavior. Similar bias has been suggested to be produced by instrumental learning. Motivational biases have been linked to dopaminergic system but the precise role of dopamine in their modulation is unclear. The present study investigated genetically driven variation in Pavlovian and instrumental learning biases by comparing task performance in subjects carrying different variants of two dopaminergic SNPs, COMT Val108/158Met and DRD2/ANKK1Taq1A. Associations with BMI, diet, age and gender were studied. All subjects were expected to show motivational bias while no direct hypotheses were made concerning genotypic or lifestyle-mediated effects due to exploratory nature of the study. Methods. 160 subjects completed a probabilistic Go/NoGo learning task in an experimental within-subject design. Generalized mixed-model logistic regressions were used to predict differences by genotype in Go responding with and without covariants. Differences by genotype in computationally modelled latent bias estimates were studied with linear regression. Results and Conclusions. Confirming expectations, an overall effect of motivational bias and a general bias towards active responding were found. Relative to Val/Met and A1+, carriers of COMT Val/Val and Taq1A A1- variants showed superior learning of correct Go responses, indicating enhanced instrumental bias. BMI was inversely associated with learning rate while diet, age and gender did not explain variance. Results partly contradict previous findings and highlight the mixed nature of research regarding associations between dopaminergic SNPs and motivational biases.

Studies have shown that there is a familial risk in developing lymphomas. This thesis aimed to find candidate predisposing mutations for inherited non-Hodgkin lymphoma susceptibility in a Finnish pedigree. Exome sequences were available from two individuals in the pedigree, and common mutations were sought from exomes using bioinformatical tools. The validity and uniqueness of these common mutations were verified, and variation segregation was assessed by sequencing variation areas from all lymphoma patients in the pedigree. After filtering, 13 common variations were found, but due to variation's presence also in healthy control samples or the lack of segregation in the kindred, no candidate variation explaining non-Hodgkin lymphoma clustering in the pedigree was found. However, this study proved a group of mutations that probably do not need further studies as candidate variations for inheritable non-Hodgkin lymphoma. This study also provided rough comparison data for two different DNA sequence analysis programs.

Introduction: Parkinson’s disease is the second most common neurodegenerative disease in the Western countries with a prevalence of about 0.3% in the population. Approximately 5 to 10% of patients are estimated to have a hereditary form of the disease. In recent years, 23 gene loci have been found, in which mutations cause hereditary Parkinson’s disease. In Finland, however, only a few disease linked gene variants have been found so far. Aims and methods: To find out if there are gene variants previously found in Parkinson’s disease patients in the Finnish population, we searched variants found in literature search in a novel genetic database, SISu, which contains genetic data of over 10 000 Finns. In addition, to confirm population findings and search for new gene variants, we analyzed 47 patient cohort with a designed gene panel and also another cohort, containing 147 patients, by minisequencing one variant found in the population data. Results: We found 16 variants in five different genes in the population data. Three of them were considered pathogenic and four likely benign after our analysis. In addition, we found nine potentially disease linked variants in eight different patients. Four of the variants were novel. Discussion: Finns seem to carry only few previously described gene variants in genes linked to Parkinson’s disease. It is likely that Finns carry their own unique variants, some of which we also found in our study. Our analysis brings valuable information about the still scarce knowledge of the genetics of this disease in the Finnish population. In addition, we were able to evaluate the disease risk of many variants further by studying their occurrence in Finns. The study of novel gene variants may bring valuable new information about the pathogenic processes related to the disease; especially the location of a novel variant in PARK2 gene found in our study turned out to be crucial for one of the previously suggested disease mechanisms.

Plötslig hjärtdöd är den vanligaste dödsorsaken i industriländerna. Kammarflimmer är en av de viktigaste orsakerna till plöstslig hjärtdöd. Hos 5-10 % av patienterna finner man ingen orsak till kammarflimret. Hos dessa patienter inleds en noggrann klinisk utredning där även gentester för genetiska rytmstörningssjukdomar spelar en mycket viktig roll. Då utredningen inte ger en förklaring till kammarflimret ställs diagnosen idiopatiskt kammarflimmer. Målet med vår studie var att med hjälp av exomsekvensering undersöka den genetiska bakgrunden till idiopatiskt kammarflimmer. I studien inkluderades 36 patienter hos vilka en förklaring till kammarflimret inte upptäcktes vid rutinmässiga kliniska undersökningar. Efter noggrann evaluering av varianterna i ljuset av släktanamnes och klinisk information om patienterna identifierades tre sannolikt patogena varianter och två varianter med okänd betydelse (VUS). Studien visar att vid exomsekvensering av patienter med idiopatiskt kammarflimmer kan avslöja en underliggande genetisk rytmstörningssjukdom hos en liten del av patienterna men hos majoriteten förblir orsaken till kammarflimret oklar.