Inflammation and Complement Activation in Intracranial Artery Aneurysms

Intracranial artery aneurysms (IAs) are estimated to be present in 2.3% of the population. A rupture of an IA causes subarachnoid hemorrhage, with up to 50% mortality. The annual low rupture risk of an IA indicates that most IAs never rupture. The current treatment options are invasive and somewhat risky. Thus rupture-prone IAs should be identified and this requires a better understanding of the IA wall pathobiology. Inflammatory cell infiltrations have been found to precede IA rupture, indicating the role of inflammation in IA wall degeneration and rupture. The complement system is a key mediator of inflammation and house-hold processing of injured tissue. This study aimed at identifying the role of complement activation in IA wall degeneration and the complement activators involved and determining how the complement system is regulated in the IA wall. In immunostainings, the end-product of complement activation, the terminal complement complex (TCC), was located mainly in the outer part of the IA wall, in areas that had also sustained loss of cells. In electron microscopy, the area of maximum TCC accumulation contained cellular debris and evidence of both apoptotic and necrotic cell death. Complement activation correlated with IA wall degeneration and rupture, de-endothelialization, and T-cell and CD163-positive macrophage infiltration. The complement system was found to become activated in all IAs by the classical pathway, with recruitment of alternative pathway amplification. Of the potential activators immunoglobulins G and M and oxidatively modified lipids were found in large areas. Lipid accumulation was observed to clearly colocalize with TCC and C-reactive protein. In the luminal parts of the IA wall, complement activation was limited by cellular expression of protectin (CD59) and extracellular matrix-bound inhibitors, C4b binding protein and factor H whereas the outer part of the wall lacked cells expressing protectin as well as matrix-bound factor H. In single nucleotide polymorphism-analysis, age-related macular degeneration-associated factor H Y402H polymorphism did not associate with the presence of IAs or their rupture The data suggest that complement activation and TCC formation are involved in IA wall degeneration and rupture. Complement seems to become activated by more than one specific activator. The association of complement with de-endothelialization and expression of several complement activators indicate a possible role of endothelial dysfunction and/or impaired clearance mechanisms. Impaired complement regulation seems to be associated with increased complement activation in IA walls. These results stress the role of chronic inflammation in IA wall pathobiology and the regulatory role of complement within this process. Imaging inflammation would possibly enhance the diagnostics of rupture-prone IAs, and targeting IA treatment to prevent chronic inflammation might improve IA treatment in the future.

Markers of systemic inflammation in diagnostics and in prediction of outcome of community-acquired infection

Sepsis is associated with a systemic inflammatory response. It is characterised by an early proinflammatory response and followed by a state of immunosuppression. In order to improve the outcome of patients with infection and sepsis, novel therapies that influence the systemic inflammatory response are being developed and utilised. Thus, an accurate and early diagnosis of infection and evaluation of immune state are crucial. In this thesis, various markers of systemic inflammation were studied with respect to enhancing the diagnostics of infection and of predicting outcome in patients with suspected community-acquired infection. A total of 1092 acutely ill patients admitted to a university hospital medical emergency department were evaluated, and 531 patients with a suspicion of community-acquired infection were included for the analysis. Markers of systemic inflammation were determined from a blood sample obtained simultaneously with a blood culture sample on admission to hospital. Levels of phagocyte CD11b/CD18 and CD14 expression were measured by whole blood flow cytometry. Concentrations of soluble CD14, interleukin (IL)-8, and soluble IL-2 receptor α (sIL-2Rα) were determined by ELISA, those of sIL-2R, IL-6, and IL-8 by a chemiluminescent immunoassay, that of procalcitonin by immunoluminometric assay, and that of C-reactive protein by immunoturbidimetric assay. Clinical data were collected retrospectively from the medical records. No marker of systemic inflammation, neither CRP, PCT, IL-6, IL-8, nor sIL-2R predicted bacteraemia better than did the clinical signs of infection, i.e., the presence of infectious focus or fever or both. IL-6 and PCT had the highest positive likelihood ratios to identify patients with hidden community-acquired infection. However, the use of a single marker failed to detect all patients with infection. A combination of markers including a fast-responding reactant (CD11b expression), a later-peaking reactant (CRP), and a reactant originating from inflamed tissues (IL-8) detected all patients with infection. The majority of patients (86.5%) with possible but not verified infection showed levels exceeding at least one cut-off limit of combination, supporting the view that infection was the cause of their acute illness. The 28-day mortality of patients with community-acquired infection was low (3.4%). On admission to hospital, the low expression of cell-associated lipopolysaccharide receptor CD14 (mCD14) was predictive for 28-day mortality. In the patients with severe forms of community-acquired infection, namely pneumonia and sepsis, high levels of soluble CD14 alone did not predict mortality, but a high sCD14 level measured simultaneously with a low mCD14 raised the possibility of poor prognosis. In conclusion, to further enhance the diagnostics of hidden community-acquired infection, a combination of inflammatory markers is useful; 28-day mortality is associated with low levels of mCD14 expression at an early phase of the disease.

Genetics of Cardiovascular Disease: A Candidate Gene Study of USF1

Cardiovascular diseases (CVD) are major contributors to morbidity and mortality worldwide. Several interacting environmental, biochemical, and genetic risk factors can increase disease susceptibility. While some of the genes involved in the etiology of CVD are known, many are yet to be discovered. During the last few decades, scientists have searched for these genes with genome-wide linkage and association methods, and with more targeted candidate gene studies. This thesis investigates variation within the upstream transcription factor 1 (USF1) gene locus in relation to CVD risk factors, atherosclerosis, and incidence and prevalence of CVD. This candidate gene was first identified in Finnish families ascertained for familial combined hyperlipidemia, a common dyslipidemia predisposing to coronary heart disease. The gene is a ubiquitously expressed transcription factor regulating expression of several genes from lipid and glucose metabolism, inflammation, and endothelial function. First, we examined association between USF1 variants and several CVD risk factors, such as lipid phenotypes, body composition measures, and metabolic syndrome, in two prospective population cohorts. Our data suggested that USF1 contributes to these CVD risk factors at the population level. Notably, the associations with quantitative measurements were mostly detected among study subjects with CVD or metabolic syndrome, suggesting complex interactions between USF1 effects and the pathophysiological state of an individual. Second, we investigated how variation at the USF1 locus contributes to atherosclerotic lesions of the coronary arteries and abdominal aorta. For this, we used two study samples of middle-aged men with detailed measurements of atherosclerosis obtained in autopsy. USF1 variation significantly associated with areas of several types of lesions, especially with calcification of the arteries. Next, we tested what effect the USF1 risk variants have on sudden cardiac death and incidence of CVD. The atherosclerosis-associated risk variant increased the risk of sudden cardiac death of the same study subjects. Furthermore, USF1 alleles associated with incidence of CVD in the Finnish population follow-up cohorts. These associations were especially prominent among women, suggesting a sex specific effect, which has also been detected in subsequent studies. Finally, as some of the low-yield DNA samples of the Finnish follow-up study cohort needed to be whole-genome amplified (WGA) prior to genotyping, we evaluated whether the produced WGA genotypes were of good quality. Although the samples giving genotype discrepancies could not be detected before genotyping with standard laboratory quality control methods, our results suggested that enhanced quality control at the time of the genotyping could identify such samples. In addition, combining two WGA reactions into one pooled DNA sample for genotyping markedly reduced the number of discrepancies and samples showing them. In conclusion, USF1 seems to have a role in the etiology of CVD. Additional studies are warranted to identify functional variants and to study interactions between USF1 and other genetic or environmental factors. This USF1 study, and other studies with low DNA yield of some samples, can benefit from whole genome amplification of the low-yield samples prior to genotyping. Careful quality control procedures are, however, needed in WGA genotyping.

GDP-L-fucose: synthesis and role in inflammation

GDP-L-fucose: synthesis and role in inflammation The migration of leukocytes from intravascular locations to extravascular sites is essential to the immune responses. The initial attachment of leukocytes to the endothelium and the rolling step of the leukocyte extravasation cascade are mediated by selectins, a family of cell adhesion molecules on cell surfaces. Selectins are able to recognize glycoproteins and glycolipids containing the tetrasaccharide sialyl Lewis x (sLex, Neu5Acα2-3Galβ1-4(Fucα1-3)GlcNAc). Several glycosyltransferases are involved in the biosynthesis of sLex, fucosyltransferase VII (Fuc-TVII) being the last enzyme to modify the sLex structure. Fuc-TVII transfers L-fucose from GDP-L-fucose to sialylated N-acetyllactosamine. GDP-L-fucose is synthesized in the cytosol via two different metabolic pathways. The major, constitutively active de novo pathway involves conversion of GDP-α-D-mannose to GDP-β-L-fucose. In the alternative salvage pathway, L-fucokinase synthesizes from free fucose L-fucose-1-phosphate, which is further converted to GDP-L-fucose by GDP-L-fucose pyrophosphorylase. GDP-L-fucose is translocated from the cytosol to Golgi for fucosylation via the GDP-fucose transporter. This thesis involved the study of the synthesis of GDP-L-fucose via the salvage pathway: cloning and expression of murine L-fucokinase and GDP-L-fucose pyrophosphorylase. The gene expression levels of these enzymes were found to be relatively high in various tissues; the mRNA levels were highest in brain, ovary and testis. This study also describes molecular cloning of rat fucosyltransferase VII (FUT7) and its expression as a functional enzyme. Gene expression levels of GDP-L-fucose synthesizing enzymes, GDP-fucose transporter and FUT7 were determined in inflamed tissues as well as cancer cells. Our results revealed a clear upregulation of the enzymes involved in the synthesis of GDP-L-fucose via de novo pathway, GDP-fucose transporter and FUT7 in inflamed tissues and in cancer cells. On the contrary, the GDP-L-fucose salvage pathway was found to be irrelevant in inflammation and in tumorigenesis. Furthermore, our results indicated the transcriptional coregulation of Golgi transporters involved in the synthesis of sulfo sLex, i.e. CMP-sialic acid, GDP-fucose and 3 phosphoadenosine 5 -phosphosulfate transporters, in inflammation.

Systemic Inflammation in Preterm Infants

Even though mortality among preterm infants has decreased, their risk for chronic complications such as bronchopulmonary dysplasia (BPD) and neurological disability remains significant. One common risk factor for these is exposure to inflammation. The fetus may be exposed prenatally during maternal chorioamnionitis. Pre-eclampsia is also associated with low-grade maternal inflammation. Postnatally, local and systemic inflammation is present during respiratory distress syndrome (RDS). Furthermore, septic infections in the preterm infant are an important source of inflammatory stimuli and can lead to death in only a few hours. The diagnosis of septic infection is difficult, since reliable diagnostic markers are unavailable. This thesis evaluates peri- and postnatal systemic inflammation in preterm infants in septic infections, in RDS treated with mechanical ventilation and surfactant treatment, and in preterm infants prenatally exposed to chorioamnionitis and pre-eclampsia. Surface expressions of the activation markers CD11b, CD54, and CD62L, determined by flow cytometry on circulating phagocytes and T lymphocytes, serve as indicators of systemic inflammation. The main findings: I) In preterm infants with developing late-onset sepsis and fulminant necrotizing enterocolitis, a significant increase in CD11b expression on circulating phagocytes is already present on the day of onset of clinical symptoms. II) In preterm infants with RDS, circulating phagocytes become activated within hours after start of mechanical ventilation. In preterm infants treated for RDS with nasal continuous positive airway pressure, no such activation occurs. III) In preterm infants, RDS is associated during the first days of life with fewer circulating helper and cytotoxic T lymphocytes, of which the greater proportions are activated. Even greater proportions of circulating T cells are activated in infants subsequently developing BPD. IV) In preterm infants born after maternal pre-eclampsia, RDS-associated phagocyte CD11b up-regulation is greater than in preterm infants not exposed to pre-eclampsia during the first week of life. These findings suggest that I) an increase in CD11b expression on circulating phagocytes can identify preterm infants with late-onset sepsis as early as at sampling for blood culture and may thus aid in the diagnosis. II) In preterm infants with RDS, initiation of mechanical ventilation, but not the use of nasal continuous positive airway pressure, promotes a systemic inflammatory reaction; exogenous surfactant does not seem to promote inflammation. III) In addition to activation of circulating cells of the innate immunity in preterm infants with RDS, the circulating cells of the adaptive immunity are activated. The activation of adaptive immunity may link acute inflammation and development of chronic inflammation-associated problems such as BPD. IV) Maternal pre-eclampsia may prime neonatal immunity to react more strongly to postnatal stimuli. In conclusion, the preterm infant is exposed to numerous potentially injurious events such as intrauterine inflammation, respiratory distress syndrome (RDS), and systemic infections, all evoking systemic inflammation. Due to ongoing development of the lung and the brain, this may, in addition to acute injury, lead to aberrant lung and brain development and to clinical syndromes of BPD and neurologic sequelae.

On the environmental genetics of depressive symptoms : Focus on two serotonergic genes

Depression is a complex psychiatric disorder influenced by several genes, environmental factors, and their interplay. Serotonin receptor 2A (HTR2A) and tryptophan hydroxylase 1 (TPH1) genes have been implicated in vulnerability to depression and other psychiatric disorders, but the results have been inconsistent. The present study examined whether these two genes moderated the influence of different depressogenic environmental factors on subthreshold depressive symptoms (assessed on a modified version of Beck s Depression Inventory, BDI) and depression-related temperament, i.e., harm avoidance (assessed on the Temperament and Character Inventory, TCI). The environmental factors included measures of childhood and adolescence exposure, i.e., maternal nurturance and parental socioeconomic status, and adulthood social circumstances, i.e., perceived social support and urban/rural residence. The participants were two randomly selected subsamples (n = 1246, n = 341) from the longitudinal population-based Cardiovascular Risk in Young Finns study (n = 3596). Childhood environmental factors were assessed when the participants were 3 to 18 years of age, and three years after the baseline. Adulthood environmental factors and outcome measures were assessed 17 and 21 years later when the participants were 21 to 39 years of age. The T102C polymorphism of the HTR2A gene moderated the association between childhood maternal nurturance and adulthood depressive symptoms, such that exposure to high maternal nurturance predicted low depressive symptoms among individuals carrying the T/T or T/C genotypes, but not among those carrying the C/C genotype. Likewise, high parental SES predicted low adulthood harm avoidance in individuals carrying the T/T or T/C genotype, but not in C/C-genotype carriers. Individuals carrying the T/T or T/C genotype were also sensitive to urban/rural residence, such that they had lower depressive symptoms in urban than in rural areas, whereas those carrying the C/C genotype were not sensitive to urban/rural residence difference. HTR2A did not moderate the influence of social support. TheA779C/A218C haplotype of the TPH1 gene was not involved in the association between childhood environment and adulthood outcomes. However, individuals carrying A alleles of the TPH1 haplotype were more vulnerable to the lack of adulthood social support in terms of high depressive symptoms than their counterparts carrying no A alleles. Furthermore, individuals living in remote rural areas and carrying the A/A haplotype had higher depressive symptoms than those carrying other genotypes of the TPH1. The findings suggest that the HTR2A and TPH1 genes may be involved in the development of depression by influencing individual s sensitivity to depressogenic environmental influences.

Inflammation and tissue destruction in arthritides and periodontal disease

The principal aim of this study was to examine diseases characterized by inflammatory injury, especially human arthritides and periodontitis, with specific interest to final effector enzymes of tissue destruction and address the possible future tools to prevent permanent tissue loss. We used biochemical and immunological methods applied to synovial tissue samples, samples of synovial fluid, and samples of peripheral blood. In Study IV, we used established clinical inflammatory injury indicator probing pocket depth and used it to derive a new clinical measure of systemic burden, periodontal inflammatory burden index. In study I, we showed a difference in the effector enzymes of peripheral blood leukocytes and leukocytes from inflamed synovial fluid of rheumatoid arthritis and reactive arthritis patients. The effector enzyme activities were higher in synovial fluid than in peripheral blood. In study II, we showed the presence of collagenase-3 in rheumatoid synovial tissue samples, relative resistance of the enzyme to inhibition in vitro and developed an electrophoretic method for detection of collagenase-3 in presence of collagenase-1. In study III, we carried out an open label study of doxycycline treatment of 12 RA patients. During the treatment period, we observed an improvement in several of the biochemical and psychosocial variables used to assess the status of the patients. In study IV, we showed a clearly lower level of periodontal inflammatory injury in chronic periodontitis patients referred for periodontal treatment. In this cross-sectional pilot study, we showed lower levels of inflammatory injury in periodontitis patients using statin than in those not receiving statin treatment. The difference was of same magnitude in patients using simvastatin or atorvastatin. The weighted index of inflammatory burden, PIBI, which emphasizes the burden imposed by the deepest pathological pockets on the system showed values consistent with a wider scale to ease future studies on the inflammatory burden associated with periodontitis.

Inflammation-induced atherogenesis, liver alterations, and cardiovascular outcome

Cardiovascular diseases, which presently are considered inflammatory diseases, affect millions of people worldwide. Chronic infections may contribute to the systemic inflammation suggested to increase the risk for cardiovascular diseases. Such chronic infections are periodontitis and Chlamydia pneumoniae infection. They are highly prevalent as approximately 10% of adult population and 30% of people over 50 years old are affected by severe periodontitis and 70-80% of elderly people are seropositive for C. pneumoniae. Our general aim was to investigate the role of infection and inflammation in atherosclerosis both in animal and human studies. We aimed to determine how the two pathogens alter the atherosclerosis-associated parameters, and how they affect the liver inflammation and lipid composition. Furthermore, we evaluated the association between matrix metalloproteinase-8 (MMP-8), a proteinase playing a major role in inflammation, and the future cardiovascular diseases (CVD) events in a population-based cohort. For the animal experiments, we used atherosclerosis-susceptible apolipoprotein E deficient (apoE-/-) mice. They were kept in germ free conditions and fed with a normal chow diet. The bacteria were administered either intravenously (A. actinomycetemcomitans) or intranasally (C. pneumoniae). Several factors were determined from serum as well as from aortic and hepatic tissues. We also determined how cholesterol efflux, a major event in the removal of excess cholesterol from the tissues, and endothelial function were affected by these pathogens. In the human study, serum MMP-8 and its tissue inhibitor (TIMP-1) concentrations were measured and their associations during the follow-up time of 10 years with CVD events were determined. An infection with A. actinomycetemcomitans increased concentrations of inflammatory mediators, MMP production, and cholesterol deposit in macrophages, decreased lipoprotein particle size, and induced liver inflammation. C. pneumoniae infection also elicited an inflammatory response and endothelial dysfunction, as well as induced liver inflammation, microvesicular appearance and altered fatty acid profile. In the population-based cohort, men with increased serum MMP-8 concentration together with subclinical atherosclerosis (carotid artery intima media thickness > 1mm) had a three-fold increased risk for CVD death during the follow-up. The results show that infections with A. actinomycetemcomitans and C. pneumoniae induce proatherogenic changes, as well as affect the liver. These data therefore support the concept that common infections have systemic effects and could be considered as cardiovascular risk factors. Furthermore, our data indicate that, as an independent predictor of fatal CVD event, serum MMP-8 could have a clinical significance in diagnosing cardiovascular diseases.

Living on the edge : Population genetics of Finno-Ugric-speaking humans in North Eurasia

The major aim of this thesis was to examine the origins and distribution of uniparental and autosomal genetic variation among the Finno-Ugric-speaking human populations living in Boreal and Arctic regions of North Eurasia. In more detail, I aimed to disentangle the underlying molecular and population genetic factors which have produced the patterns of uniparental and autosomal genetic diversity in these populations. Among Finno-Ugrics the genetic amalgamation and clinal distribution of West and East Eurasian gene pools were observed within uniparental markers. This admixture indicates that North Eurasia was colonized through Central Asia/ South Siberia by human groups already carrying both West and East Eurasian lineages. The complex combination of founder effects, gene flow and genetic drift underlying the genetic diversity of the Finno-Ugric- speaking populations were emphasized by low haplotype diversity within and among uniparental and biparental markers. A high prevalence of lactase persistence allele among the North Eurasian Finno- Ugric agriculturalist populations was also shown indicating a local adaptation to subsistence change with lactose rich diet. Moreover, the haplotype background of lactase persistence allele among the Finno- Ugric-speakers strongly suggested that the lactase persistence T-13910 mutation was introduced independently more than once to the North Eurasian gene pool. A significant difference in genetic diversity, haplotype structure and LD distribution within the cytochrome P450 CYP2C and CYP2D regions revealed the unique gene pool of the Finno-Ugric Saami created mainly by population genetic processes compared to other Europeans and sub-Saharan Mandenka population. From all studied populations the Saami showed also significantly the highest allele frequency of a CYP2C19 gene mutation causing variable drug reactions. The diversity patterns observed within CYP2C and CYP2D regions emphasize the strong effect of demographic history shaping genetic diversity and LD especially among such small and constant size populations as the Finno-Ugric-speaking Saami. Moreover, the increased LD in Saami due to genetic drift and/or admixture was shown to offer an advantage for further attempts to identify alleles associated to common complex pharmacogenetic traits.

Molecular Background of Common Dyslipidemias

The leading cause of death in the Western world continues to be coronary heart disease (CHD). At the root of the disease process is dyslipidemia an aberration in the relevant amounts of circulating blood lipids. Cholesterol builds up in the arterial wall and following rupture of these plaques, myocardial infarction or stroke can occur. Heart disease runs in families and a number of hereditary forms are known. The leading cause of adult dyslipidemia presently however is overweight and obesity. This thesis work presents an investigation of the molecular genetics of common, hereditary dyslipidemia and the tightly related condition of obesity. Familial combined hyperlipidemia (FCHL) is the most common hereditary dyslipidemia in man with an estimated population prevalence of 1-6%. This complex disease is characterized by elevated levels of serum total cholesterol, triglycerides or both and is observed in about 20% of individuals with premature CHD. Our group identified the disease to be associated with genetic variation in the USF1 transcription factor gene. USF1 has a key role in regulating other genes that control lipid and glucose metabolism as well as the inflammatory response all central processes in the progression of atherosclerosis and CHD. The first two works of this thesis aimed at understanding how these USF1 variants result in increased disease risk. Among the many, non-coding single-nucleotide polymorphisms (SNPs) that associated with the disease, one was found to have a functional effect. The risk-enhancing allele of this SNP seems to eradicate the ability of the important hormone insulin to induce the expression of USF1 in peripheral tissues. The resultant changes in the expression of numerous USF1 target genes over time probably enhance and accelerate the atherogenic processes. Dyslipidemias often represent an outcome of obesity and in the final work of this thesis we wanted to address the metabolic pathways related to acquired obesity. It is recognized that active processes in adipose tissue play an important role in the development of dyslipidemia, insulin resistance and other pathological conditions associated with obesity. To minimize the confounding effects of genetic differences present in most human studies, we investigated a rare collection of identical twins that differed significantly in the amount of body fat. In the obese, but otherwise healthy young adults, several notable changes were observed. In addition to chronic inflammation, the adipose tissue of the obese co-twins was characterized by a marked (47%) decrease in amount of mitochondrial DNA (mtDNA) a change associated with mitochondrial dysfunction. The catabolism of branched chain amino acids (BCAAs) was identified as the most down-regulated process in the obese co-twins. A concordant increase in the serum level of these insulin secretagogues was identified. This hyperaminoacidemia may provide the feed-back signal from insulin resistant adipose tissue to the pancreas to ensure an appropriately augmented secretory response. The down regulation of BCAA catabolism correlated closely with liver fat accumulation and insulin. The single most up-regulated gene (5.9 fold) in the obese co-twins was osteopontin (SPP1) a cytokine involved in macrophage recruitment to adipose tissue. SPP1 is here implicated as an important player in the development of insulin resistance. These studies of exceptional study samples provide better understanding of the underlying pathology in common dyslipidemias and other obesity associated diseases important for future improvement of intervention strategies and treatments to combat atherosclerosis and coronary heart disease.

Molecular Genetics of Lactase Deficiencies

Congenital lactase deficiency (CLD) (MIM 223000) is a rare autosomal recessive gastrointestinal disorder characterized by watery diarrhea in infants fed with breast milk or other lactose-containing formulas. The CLD locus was previously assigned by linkage and linkage disequilibrium analyses on 2q21 in 19 Finnish families. In this study, the molecular background of this disorder is reported. The CLD locus was refined in 32 CLD patients in 24 families by using microsatellite and single nucleotide polymorphism (SNP) haplotypes. Mutation analyses were performed by direct sequencing. We identified 5 distinct mutations in the lactase (LCT) gene, encoding the enzyme that hydrolyzes lactose in the intestinal lumen. These findings facilitate genetic testing of CLD in clinical practice and enable genetic counseling. The present data also provide the basis for detailed characterization of the molecular pathogenesis of this disorder. Adult-type hypolactasia (MIM 223100) (lactase non-persistence, lactose intolerance) is an autosomal recessive gastrointestinal condition that is a result of a decline in the activity of lactase in the intestinal lumen after weaning. Adult-type hypolactasia is considered to be a normal phenomenon among mammals and symptoms are remarkably milder than experienced in CLD. Recently, a variant C/T-13910 was shown to associate with the adult-type hypolactasia trait, locating 13.9 kb upstream of the LCT gene. In this study, the functional significance of the C/T-13910 variant was determined by studying the LCT mRNA levels in intestinal biopsy samples in children and adults with different genotypes. RT-PCR followed by solid-phase minisequencing was applied to determine the relative expression levels of the LCT alleles using an informative SNP located in exon 1. In children, the C-13910 allele was observed to be downregulated after five years of age in parallel with lactase enzyme activity. The expression of the LCT mRNA in the intestinal mucosa in individuals with the T-13910 A-22018 alleles was 11.5 times higher than that found in individuals with the C-13910, G-22018 alleles. These findings suggest that the C/T-13910 associated with adult-type hypolactasia is associated with the transcriptional regulation of the LCT gene. The presence of the T-13910 A-22018 allele also showed significant elevation lactase activity. Galactose, the hydrolysing product of the milk sugar lactose, has been hypothesized to be poisonous to ovarian epithelial cells. Hence, consumption of dairy products and lactase persistence has been proposed to be a risk factor for ovarian carcinoma. To investigate whether lactase persistence is related to the risk of ovarian carcinoma the C/T-13910 genotype was determined in a cohort of 782 women with ovarian carcinoma 1331 individuals serving as controls. Lactase persistence did not associate significantly with the risk for ovarian carcinoma in the Finnish, in the Polish or in the Swedish populations. The findings do not support the hypothesis that lactase persistence increases the risk for ovarian carcinoma.