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1.	Pettersson, Ulf (författare) Människoraser, finns dom? 2009 Ingår i: Rasen och vetenskapen. - Uppsala : Centrum för multietnisk forskning / Programmet för studier kring Förintelsen och folkmord. - 9789197743426 ; , s. 13-22 Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract Föredrag från föreläsningsserien Rasen och vetenskapen vid Uppsala universitet våren 2008. Artikeln av professorn i medicinsk genetik Ulf Pettersson (Uppsala) behandlar rasbegreppets ställning i människans utvecklingshistoria inom genetiken.
2.	Bin Kaderi, Mohamed Arifin, 1978- (författare) Assessment of Novel Molecular Prognostic Markers in Chronic Lymphocytic Leukemia 2010 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract The clinical course of chronic lymphocytic leukemia (CLL) is highly heterogeneous, which has prompted the search for biomarkers that can predict prognosis in this disease. The IGHV gene mutation status and certain genomic aberrations have been identified as reliable prognostic markers of clinical outcome for this disorder. However, the search for more feasible prognostic markers in CLL is still being pursued. Recently, certain single nucleotide polymorphisms (SNPs) in the GNAS1, BCL2 and MDM2 genes and the RNA expression levels of the LPL, ZAP70, TCL1, CLLU1 and MCL1 genes were suggested as novel prognostic markers in CLL. In papers I-III, we performed genotyping analyses of the GNAS1 T393C, BCL2 -938C>A and MDM2 SNP309 polymorphisms in 268-418 CLL patients and related the genotypes with clinical data. Association studies between the polymorphisms and established prognostic markers (i.e. IGHV mutation status, genomic aberrations, CD38 expression) were also performed. Our studies did not find any significant relationship between these SNPs with either clinical outcome or other known prognostic markers in CLL. In paper IV, we measured the RNA expression levels of LPL, ZAP70, TCL1, CLLU1 and MCL1 in 252 CLL cases and correlated these levels with clinical outcome. Here, we verified that high expression of all these RNA-based markers, except MCL1, were associated with an unfavourable prognosis. We also confirmed a close relationship between IGHV mutation status and the RNA-based markers, especially for LPL and CLLU1 expression. Among the RNA-based markers, multivariate analysis revealed LPL expression as the strongest independent prognostic marker for overall survival and time to treatment. Furthermore, the RNA-based markers could add further prognostic information to established markers in subgroups of patients, with LPL expression status giving the most significant results. In summary, data from papers I-III could not verify the GNAS1 T393C, BCL2 -938C>A and MDM2 SNP309 polymorphisms as prognostic markers in CLL. Future SNP markers must hence be confirmed in large, independent cohorts before being proposed as prognostic marker in CLL. In paper IV, we conclude that LPL expression appears to be the strongest among the RNA-based markers for CLL prognostication. Further efforts to standardize LPL quantification are required before it can be applied in the clinical laboratory to predict clinical outcome in this disease.
3.	Delgado Vega, Angélica María, 1982- (författare) Dissecting the Genetic Basis of Systemic Lupus Erythematosus : The Pursuit of Functional Variants 2013 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Systemic lupus erythematosus (SLE) is a chronic and systemic autoimmune disease that primarily affects women during the childbearing years. SLE is characterized by the production of autoantibodies against nucleic acids and their interacting proteins. The exact molecular mechanisms leading to the breakdown of self-tolerance remain to a large extent unknown, but it is well established that they are influenced by both non-genetic (i.e. environmental and hormonal) and genetic factors. SLE is a complex, polygenic disease. Several susceptibility variants have been identified in SLE. However, the functional role in disease pathogenesis for the majority of them remains largely unknown.This thesis includes case-control association studies where the role of the genes TNFSF4 (Paper I), STAT4 (Paper II), CD226 (Paper III), and BLK (Papers IV and V) in the susceptibility of developing SLE was investigated. The primary focus was on the identification of the functional variants underlying the association. For each of these genes, fine mapping was performed using single nucleotide polymorphisms (SNPs), the linkage disequilibrium (LD) was characterized, and the association was narrowed down to specific haplotypes by means of several different statistical genetic strategies. Candidate variants were prioritized for further functional analysis on the basis of their potential effect on the gene function, their association, and/or biological plausibility. In Paper I, the association of TNFSF4 with SLE was validated and attributed to a risk haplotype tagged by SNPs rs1234317-T and rs12039904-T. Paper II provides evidence supporting the presence of at least two independent genetic effects within the STAT4 gene represented by rs3821236-A and rs7574865-A, which correlated with increased levels of gene expression. In Paper III, a functional allele in CD226 (rs727088-C) was identified, which was responsible for decreased levels in both mRNA and protein expression. In Paper IV, two independent genetic effects in the BLK gene were demonstrated. The first one comprised multiple regulatory variants in high LD that were enriched for NFκB and IRF4 binding sites and correlated with low BLK mRNA levels. The second was a low-frequency missense substitution (Ala71Thr) that decreased the BLK protein half-life. In Paper V, a genetic epistatic interaction between BANK1 rs10516487 (GG) and BLK rs2736340 (TT+TC) was demonstrated. Additional molecular analyses established that these molecules interact physically. These studies have contributed to the dissection of the genetic architecture of SLE. They highlight the allelic heterogeneity of the disease and provide functional links to the associated variants, which has significantly aided in the understanding of SLE disease pathogenesis.
4.	Olsson, Malin, 1972- (författare) Familial amyloidosis with polyneuropathy : studies of genetic factors modifying the phenotype of the disease 2010 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Background. Familial Amyloidosis with Polyneuropathy (FAP) is an autosomal dominantly inherited systemic amyloid disease. The disease is caused by mutations in the transthyretin (TTR) gene, where close to 100 different amyloidogenic mutations have been identified. FAP is found worldwide, but endemic areas with a high frequency of patients are found in Portugal, Japan and northern Sweden. Cases from these endemic areas all share the same TTR c.148G>A, p.V50M ("V30M") mutation, but the phenotype of the disease varies between the areas, and also within the endemic areas. The mean onset of the disease is two decades earlier in Portugal and Japan compared to Sweden, but late as well as early age at onset cases occur within all the populations. Interestingly, the different populations all display a maternal anticipation, where an earlier onset is observed for those individuals who inherit the trait from their mother. Since substantial variation in the phenotype is observed for different populations, epigenetic/genetic and/or environmental factors must exert a significant impact on the penetrance of the disease. Amyloid formation is caused by conformational changes of proteins, which facilitates their assembly into fibrils, amyloid. Oxidative stress can mediate conformational changes of proteins and since the mitochondria regulate oxidative processes within the cell, mitochondrial function may affect amyloid formation. The mitochondrial DNA is a non-nuclear DNA, which is entirely maternally inherited, and therefore could be related to the observed maternal anticipation of the disease. In addition, differences within the surrounding regions of the TTR gene may have an impact on the transcription of the gene and thereby on the expression of the different alleles. Material and methods. DNA from early and late onset V30M cases and from non-carriers (the latter utilised as controls) from Swedish, French, Japanese and Portuguese populations were analysed. In addition, DNA from healthy Swedish V30M carriers was analysed. Conventional analytical methods were employed, such as PCR, sequencing and genotyping. Conventional statistical methods used were t-test, Chi-squared test and maximum likelihood. Results. The study of V30M carrier frequency in two counties (Lycksele and Skellefteå) within the Swedish endemic area revealed a carrier frequency of 2.14% and 2.54%, respectively. The mitochondrial haplogroup analysis showed that in populations with generally late onset (French and Swedish), the haplogroup distribution of late onset cases resembled that of the controls derived from the same area, whereas haplogroup distribution for early onset patients was significantly different. The most pronounced difference was for the rare haplogroup K, of which early onset cases had a higher frequency than the controls. Analysis of the Portuguese population, with predominantly early onset, showed that haplogroup distribution for early onset cases were similar to the Portuguese control group, which had a different distribution than the Swedish control group. By analysis of pedigrees from Swedish and Portuguese patients it could be shown that mitochondrial genetic variation entirely could explain maternal anticipation in the Portuguese patients, whereas for Swedish patients, an additional parent of origin effect is present. Our analysis of the TTR gene disclosed a polymorphism (rs62093482) in the 3'UTR region of the Swedish patients. This polymorphism was found in all V30M carriers, irrespective of symptoms. In addition, homozygous TTR V30M carriers were homozygous also for the polymorphism. Since Swedish patients share a common founder this polymorphism thus is localised on the V30M allele. This polymorphism was found in only 4% of the Swedish controls. French controls showed the same frequency, but none of the French V30M patients displayed the polymorphism. In the Japanese population the polymorphism was not present at all. Interestingly, this polymorphism generates a potential binding site for microRNA and thereby possibly could down-regulate the expression of the mutated TTR allele. Conclusions. The carrier frequency in the endemic area is remarkably high, above 2% in the Lycksele and Skellefteå areas. The prevailing haplogroup distributions in the different endemic areas are consistent between the general population and the patient group with the predominant phenotype of that area. Mitochondrial genetic differences may explain maternal anticipation in Portuguese patients, and have an influence in Swedish patients. A polymorphism in the 3'UTR regulatory region of the mutated TTR allele is found in all Swedish patients. This polymorphism may down-regulate TTR V30M expression and thereby contribute to the late onset of the disease noted in the Swedish population.
5.	Norberg, Maria, 1976- (författare) In Vitro Drug Sensitivity and Apoptosis in Chronic Lymphocytic Leukemia 2010 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Chronic lymphocytic leukemia (CLL) is a heterogeneous malignancy displaying varying clinical outcome, where molecular markers today can divide patients into prognostic subgroups. Despite the introduction of new agents for treatment, remissions are usually not sustained in CLL and resistance towards treatment can partly be explained by aberrant apoptosis. The aim of this thesis was to find new drugs for CLL patients resistant to conventional therapy and to analyze genes involved in apoptosis within different prognostic subgroups. In paper I-II, the in vitro activity of substances was investigated using the fluorometric microculture cytotoxicity assay (FMCA). When evaluating rapamycin (paper I), an inhibitor of mTOR, in 97 tumor samples from different entities, CLL was found to be one of the most sensitive tumor types. Combination experiments on patient CLL cells indicated that rapamycin acted synergistically with the CLL drugs vincristine and chlorambucil. An investigation of 20 anti-cancer agents in cells from 40 CLL patients (paper II) revealed that prednisolone and rolipram displayed high activity in poor-prognostic patients, in particular IGHV unmutated CLL. Furthermore, when used in combination these agents were found to produce a synergistic effect. In paper III, the anti-apoptotic BCL2 family member BFL1 was evaluated in 37 CLL cases. Levels of BFL1 were higher in fludarabine-resistant patients compared to fludarabine-sensitive patients. In addition, the high expression of BFL1 inversely correlated to fludarabine-induced apoptosis in CLL cells. A single nucleotide polymorphism in the anti-apoptotic BCL2 gene (-938C>A) has been suggested as a novel poor-prognostic marker in CLL. In paper IV, we investigated this BCL2 polymorphism in 268 CLL patients and correlated genotypes to clinical data. However, no association could be confirmed between this polymorphism and clinical outcome or established prognostic markers. In conclusion, this thesis has shown that rapamycin is a potential drug for treatment in CLL. Furthermore, prednisolone and rolipram were identified as interesting candidates for treatment of poor-prognostic patients. Finally, the anti-apoptotic protein BFL1 may contribute to chemoresistance and hence represents a potential therapeutic target in CLL, whereas from our data, the BCL2 -938C>A polymorphism does not appear to have any prognostic significance.
6.	Nord, Helena, 1980- (författare) Application of Genomic and Expression Arrays for Identification of new Cancer Genes 2010 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Copy number variation (CNV) comprises a recently discovered kind of variation involving deletion and duplication of DNA segments of variable size, ranging from a few hundred basepairs to several million. By altering gene dosage levels or disrupting proximal or distant regulatory elements CNVs create human diversity. They represent also an important factor in human evolution and play a role in many disorders including cancer. Array-based comparative genomic hybridization as well as expression arrays are powerful and suitable methods for determination of copy number variations or gene expression changes in the human genome. In paper I we established a 32K clone-based genomic array, covering 99% of the current assembly of the human genome with high resolution and applied it in the profiling of 71 healthy individuals from three ethnic groups. Novel and previously reported CNVs, involving ~3.5% of the genome, were identified. Interestingly, 87% of the detected CNV regions overlapped with known genes indicating that they probably have phenotypic consequences. In papers II through IV we applied this platform to different tumor types, namely two collections of brain tumors, glioblastoma (paper II) and medulloblastoma (paper III), and a set of bladder carcinoma (paper IV) to identify chromosomal alterations at the level of DNA copy number that could be related to tumor initiation/progression. Tumors of the central nervous system represent a heterogeneous group of both benign and malignant neoplasms that affect both children and adults. Glioblastoma and medulloblastoma are two malignant forms. Glioblastoma often affects adults while the embryonal tumor medulloblastoma is the most common malignant brain tumor among children. The detailed profiling of 78 glioblastomas, allowed us to identify a complex pattern of aberrations including frequent and high copy number amplicons (detected in 79% of samples) as well as a number of homozygously deleted loci. These regions encompassed not only previously reported oncogenes and tumor suppressor genes but also numerous novel genes. In paper III, a subset of 26 medulloblastomas was analyzed using the same genomic array. We observed that alterations involving chromosome 17, especially isochromosome 17q, were the most common genomic aberrations in this tumor type, but copy number alterations involving other chromosomes: 1, 7 and 8 were also frequent. Focal amplifications, on chromosome 1 and 3, not previously described, were also detected. These loci may encompass novel genes involved in medulloblastoma development. In paper IV we examined for the presence of DNA copy number alterations and their effect on gene expression in a subset of 21 well-characterized Ta bladder carcinomas, selected for the presence or absence of recurrences. We identified a number of novel genes as well as a significant association between amplifications and high-grade and recurrent tumors which might be clinically useful. The results derived from these studies increase our understanding of the genetic alterations leading to the development of these tumor forms and point out candidate genes that may be used in future as targets for new diagnostic and therapeutic strategies.
7.	Afrakhte, Mozhgan (författare) Growth control mechanisms in normal and neoplastic mammalian cells 1998 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract The main theme of the studies presented in this thesis is, the growth control mechanisms whose loss in normal cells predispose to or cause cancer. The balance between growth inhibitory and stimulatory mechanisms is crucial for the development and maintenance of a normal animal.PDGF, a growth factor for cells of mesenchymal origin, is implicated in normal developmental processes as well as neoplasia. The alternative splicing of exon 6 in PDGF-A gene transcripts gives rise to two different proteins with different compartmentalization properties. The PDGF-A chain homodimers, PDGF-AAL, encoded PDGF A-splice variant remain associated with the cell membrane. Studies of a human fibrosarcoma cell line, U-2197, revealed a high expression level of the cell associated PDGF-AAL which upon release increased autophosphorylation of the endogenous PDGF receptors, suggesting an autocrine loop. PDGF-A gene and PDGFR-α gene found to be co-amplified in the U-2197, indicating an optimised system for growth in these cells, i.e. amplified growth factor receptor as well as a local autocrine supply of the mitogen.Members of TGFβ superfamily are potent regulators of the growth and differentiation of a wide range of cell types. Intracellular mediators of TGF-β signalling, SMADs, transduce signals from serine/threonine kinase receptors to the nucleus where they affect transcription of target genes. A new class of SMAD proteins has been identified whose members, the inhibitory SMADS, antagonise TGF-β signals by interfering with agonistic SMADs activity. Smad6 and Smad7 are two closely related TGF-β antagonists identified in mammalian cells. Overexpression of Smad7 inhibited the cellular response to TGF-β whereas expression of an anti-sense Smad7 construct showed an enhancing effect on this response. The inhibitory SMADs may act in a negative feedback loop, as their expression is induced by the same ligands whose action they antagonise.Density dependent growth inhibition is a growth control mechanism often lost in transformed and malignant cells. Cells in dense culture are refractory to the mitogen stimulation although, the mitogenic signals were shown to be processed to some extent. The expression of immediate-early genes in dense culture stimulated with mitogen was induced. The activity of cyclin dependent kinases (CDKs), the pivotal kinases in G1/S transition, showed to be density dependent and decreased by increasing cell density. pRb, a tumour suppressor and growth regulatory protein, remained unphosphorylated in mitogen treated dense culture. The cessation of CDKs kinase activity in dense cultures was shown to be accompanied with increasing expression of inhibitory proteins of these kinases, CKIs. The impaired expression of a positive regulator of CDKs, Cdc25A phosphatase, was another feature of dense cultures.
8.	Antson, Dan-Oscar (författare) Genotyping RNA and DNA using padlock probes 2001 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Novel techniques are needed to investigate the genetic variation revealed in the first draft of the human genome sequence. Padlock probes are recently developed reagents, suitable for detecting single-nucleotide variations of DNA and RNA in situ or in solution. The probes are oligonucleotides of about 70-140 nucleotides that can be circularized by ligation in the presence of a correct target sequence. Standard chemical synthesis of padlock probes is difficult due to the requirement for intact 5' and 3' ends of these long oligonucleotides. A novel PCR-based method is presented in this thesis, whereby longer, densely labeled padlock probes can be made as compared to conventional chemical synthesis. PCR-generated padlock probes produced a stronger signal and a more resolved staining pattern, compared to chemically synthesized probes in fluorescence in situ analysis of an alpha-satellite sequence variant present in human chromosomes 13 and 21. Padlock probes used for in situ analysis of metaphase chromosomes had an optimal length of 140 nucleotides. They were used to identify individual chromosomes 7 and 15, and to follow the transmission of chromosome homologues for two consecutive generations. The specificity of the padlock probes to detect single copy genes in genomic DNA samples was demonstrated by detecting a single-nucleotide mutation in the ATP7B gene. It has not previously been known if T4 DNA ligase can be used for RNA sequence analysis. In this thesis, it is demonstrated that T4 DNA ligase can be used for distinguishing single-nucleotide RNA sequence variants. Reaction conditions were defined where most mismatches could be discriminated by a factor of 80 and all mismatches by a factor of at least 20. Under these conditions padlock probes could detect and distinguish RNA sequence variants with ligation efficiency almost as high as on the corresponding DNA sequence. A detailed study of the parameters influencing RNA-templated DNA ligation revealed that DNA ligation on RNA templates proceeds at a much slower rate compared to the same reaction on DNA, and that a molar excess of enzyme is required. Furthermore, the ligation reaction is inhibited by high concentrations of the cofactor ATP and NaCl. The work presented in this thesis demonstrates that PCR-generated padlock probes can detect and distinguish single-nucleotide variation in both RNA and DNA.
9.	Aspegren, Anders (författare) Nuclear Organization of Gene Expression in Adenovirus Infected Cells 2001 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Adenovirus infected cells provide a good model system for studying nuclear organization during RNA production and transport. This thesis is focused on the dynamic organization of splicing factors during the late phase of Adenovirus infection in HeLa cells, the nuclear localization of viral RNA, and the pathway used for viral RNA transport to the cytoplasm.Splicing factors are relocalized from interchromatin granule clusters to sites of transcription in Adenovirus infected cells at intermediate times of infection. Later, splicing factors and viral RNA accumulate posttranscriptionally in interchromatin granule clusters. The release of the splicing factors from transcription sites was energy dependent or preceded by energy requiring mechanisms. Our data indicated that phosphorylation events inhibited by staurosporine, and 3' cleavage of the transcript are two possible mechanisms involved prior to the release of the RNP complex from transcription sites.A viral protein derived from orf6 of early region 4, 34K, is important for the nuclear stability and transport of late viral mRNA derived from the major late transcription unit. A viral mutant lacking this region is defective for posttranscriptional accumulation of viral mRNA in interchromatin granule clusters, and for the accumulation of viral RNA in the cytoplasm. These results suggest that posttranscriptional accumulation of viral RNA in interchromatin granule clusters may contribute to the maturation of the RNP complex or sorting of RNAs and proteins, to prepare the final RNP complex for transport to the cytoplasm.A previous model suggested that adenoviral late mRNA is transported to the cytoplasm by utilizing the CRM-1 pathway. This pathway can be blocked by the drug leptomycin B. The data presented in paper IV suggests that this model might not be applicable, since leptomycin B did not inhibit adenoviral late gene expression.
10.	Ata, Ahmad Khaled (författare) Expression of TGF- isoforms, their receptors and related SMAD proteins in brain pathology : Immunohistochemical studies focusing on infarcts, abscesses and malignant gliomas 1999 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract This thesis focuses on the immunohistochemical expression of transforming growth factor beta(TGFβ) isoforms, their receptors and TGF-β-related SMAD proteins in brain pathology, chiefly in-farcts. One key question was whether the expressions of these compounds are altered within glial cells, endothelial cells of microvessels and other cell types in the vicinity of infarcts. Studies on human and animal brain infarcts were made. Immunoreactivities to TGF-β isoforms -β1, -β2 and -β3, and TGF-βreceptor (TβR) type I were seen in astrocytes, macrophages, neurons, endothelial and vascular smooth muscle cells of human brain infarcts. Similar observations were made in an experimental model of rat brain infarct at day 1 and 3 following occlusion of the middle cerebral artery (MCA). Increased expression of Smad2, -3, -4, -6 and -7 was seen already at 6 h after MCA occlusion in neurons, microvascular endothelial cells, astroglial cells and inflammatory cells. Later on, immunopositive macrophages were present in the infarcts. The changes persisted even at day 7 after MCA occlusion.Several alterations thus occur during the evolution of brain infarcts with regard to the immunohistochemical expression of TGF-β, its receptors and related SMAD proteins. Such changes are, however, not unique to brain infarcts. Thus, patterns of high expression for TGF-β- isoforms -β1, -β2 -β3, and TβR-I in cases of brain abscess (human), and of Smad2, -3, -4, -6 and -7 in tumor cells and neoplastic blood vessels of malignant gliomas (human) were also observed.In addition, immunohistochemical expression of vascular endothelial growth factor (VEGF) andits receptors was investigated since this growth factor is involved in angiogenesis and edemaformation, two cardinal features of brain infarcts. Increased immunoreactivities, seen particularly in the edges of infarcts, were observed already at day 1 after MCA occlusion.In conclusion, the various TGF-β isoforms, receptors and related SMAD proteins, together with other factors, seem to be involved in the very complicated and important changes taking place in the vicinity of brain infarcts.
11.	Balciuniene, Jorune (författare) Genetic studies of two inherited human phenotypes : Hearing loss and monoamine oxidase activity 2001 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract This thesis focuses on the identification of genetic factors underlying two inherited human phenotypes: hearing loss and monoamine oxidase activity. Non-syndromic hearing loss segregating in a Swedish family was tested for linkage to 13 previously reported candidate loci for hearing disabilities. Linkage was found to two loci: DFNA12 (llq22-q24) and DFNA2 (lp32). A detailed analysis of the phenotypes and haplotypes shared by the affected individuals supported the hypothesis of digenic inheritance of hearing disability in the Swedish family. Mutation screening of α-tectorin, a gene residing within the DFNA12 region revealed a mutation of a conserved amino acid (Cys to Ser), that segregated with the disease. The identification of the mutation added support to the involvement of α-tectorin in hearing disabilities. In contrast, no mutations were identified in two candidate genes at the DFNA2 locus, that were reported to cause hearing loss in other families. It is possible that the DFNA2 locus contains a third, not yet identified, hearing loss gene. Monoamine oxidase A (MAOA) and B (MAOB) catalyze the degradation of certain neurotransmitters in the central nervous system and are associated with specific behavioral and neuropsychiatric human traits. Activity levels of both monoamine oxidases (MAO) are highly variable among humans and are determined by unknown genetic factors. This study investigated the relationship of different MAO alleles with MAO mRNA levels and enzyme activity in human brain. Several novel DNA polymorphisms were identified in a group of Swedish individuals. Haplotypes containing several closely located MAOA polymorphisms were assessed in Asian, African, and Caucasian populations. The haplotype distribution and diversity pattern found among the three populations supported the occurrence of a bottleneck during the dispersion of modem humans from Africa. Allelic association studies conducted on postmortem human brain samples, revealed the association between a SNP in the MAOB intron 13, and different levels of both MAO enzyme activities. This suggested that this SNP is in linkage disequilibrium with at least one novel functional DNA polymorphism that controls MAO enzyme activities in human brain. The identification of functional polymorphisms regulating the activity of these enzymes will help to elucidate the involvement of MAO in human behavior and neuropsychiatric conditions.
12.	Beskow, Anna, 1972- (författare) Genetic Risk Factors for Cervical Carcinoma in situ 2003 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Oncogenic human papillomaviruses (HPVs) are implicated in 99.7 % of cervical cancer cases but require the co-operation of other factors. To investigate potential genetic risk factors we have typed the HLA class II DRB1 and DQB1 loci in 478 women diagnosed with cervical carcinoma in situ and in 608 age-matched controls. Quantitative measurements of HPV 16, HPV 18/45 and HPV 31 were obtained. The DRB11501 and DQB10602 alleles were found to increase the risk of HPV 16 infection. Carriers of DRB11501 and DQB10602 were also shown to have an increased risk of a higher viral load compared to non-carriers. The DRB11301 and DQB10603 alleles were found to protect from HPV 18/45 and 31 infections as well as resulting in a lower viral load in carriers compared to non-carriers. Women with a high HPV 16, 18/45 or 31 viral load were more prone to long-term infections and women with a low HPV 16 viral load were more prone to short-term infections. Carriers of DRB11501 and DQB10602 alleles were also shown to have an increased risk of long-term infections compared to short-term infections. We also tested if an HPV susceptibility locus found for epidermodysplasia verruciformis (EV) was also linked to HPV susceptibility in cervical cancer. We did not find any linkage to this locus in a set of 77 families, each with at least three cases diagnosed with cervical carcinoma in situ. Other potential risk factors tested were HPV 16 E6 variants together with a p53 codon 72 polymorphism and HLA class II alleles. We found an association between the E6 L83V variant and the HLA DR4-DQ3 haplotype, as well as an increased frequency of Arg homozygosity of p53 in women infected with the L83V variant. These results show that alleles at HLA class II loci represents risk factors for persistent HPV infection and thereby also contribute to the risk of development of cervical carcinoma in situ.
13.	Birve, Anna, 1968- (författare) Suppressor of zeste 12, a Polycomb group gene in Drosophila melanogaster; one piece in the epigenetic puzzle 2003 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract In multicellular organisms all cells in one individual have an identical genotype, and yet their bodies consist of many and very different tissues and thus many different cell types. Somehow there must be a difference in how genes are interpreted. So, there must be signals that tell the genes when and where to be active and inactive, respectively. In some instances a specific an expression pattern (active or inactive) is epigenetic; it is established and maintained throughout multiple rounds of cell divisions. In the developing Drosophila embryo, the proper expression pattern of e.g. the homeotic genes Abd-B and Ubx is to be kept active in the posterior part and silenced in the anterior. Properly silenced homeotic genes are crucial for the correct segmentation pattern of the fly and the Polycomb group (Pc-G) proteins are vital for maintaining this type of stable repression.As part of this thesis, Suppressor of zeste 12 (Su(z)12) is characterized as a Drosophila Pc-G gene. Mutations in the gene cause widespread misexpression of several homeotic genes in embryos and larvae. Results show that the silencing of the homeotic genes Abd-B and Ubx, probably is mediated via physical binding of SU(Z)12 to Polycomb Response Elements in the BX-C. Su(z)12 mutations are strong suppressors of position-effect-variegation and the SU(Z)12 protein binds weakly to the heterochromatic centromeric region. These results indicate that SU(Z)12 has a function in heterochromatin-mediated repression, which is an unusual feature for a Pc-G protein. The structure of the Su(z)12 gene was determined and the deduced protein contains a C2-H2 zinc finger domain, several nuclear localization signals, and a region, the VEFS box, with high homology to mammalian and plant homologues. Su(z)12 was originally isolated in a screen for modifiers of the zeste-white interaction and I present results that suggests that this effect is mediated through an interaction between Su(z)12 and zeste. I also show that Su(z)12 interact genetically with other Pc-G mutants and that the SU(Z)12 protein binds more than 100 euchromatic bands on polytene chromosomes. I also present results showing that SU(Z)12 is a subunit of two different E(Z)/ESC embryonic silencing complexes, one 1MDa and one 600 kDa complex, where the larger complex also contains PCL and RPD3.In conclusion, results presented in this thesis show that the recently identified Pc-G gene, Su(z)12, is of vital importance for correct maintenance of silencing of the developmentally important homeotic genes.
14.	Cahill, Nicola, 1983- (författare) Molecular Genetic and DNA Methylation Profiling of Chronic Lymphocytic Leukaemia : A Focus on Divergent Prognostic Subgroups and Subsets 2012 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Advancements in prognostication have improved the subdivision of chronic lymphocytic leukaemia (CLL) into diverse prognostic subgroups. In CLL, IGHV unmutated and IGHV3-21 genes are associated with a poor-prognosis, conversely, IGHV mutated genes with a favourable outcome. The finding of multiple CLL subsets expressing ‘stereotyped’ B-cell receptors (BCRs) has suggested a role for antigen(s) in leukemogenesis. Patients belonging to certain stereotyped subsets share clinical and biological characteristics, yet limited knowledge exists regarding the genetic and epigenetic events that may influence their clinical behaviour. This thesis aimed to, further investigate Swedish IGHV3-21-utilising patients, screen for genetic and DNA-methylation events in CLL subgroups/subsets and study DNA methylation over time and within different CLL compartments. In paper I, IGHV gene sequencing of 337 CLL patients from a Swedish population-based cohort revealed a lower (6.5%) IGHV3-21 frequency relative to previous Swedish hospital-based studies (10.1-12.7%). Interestingly, this frequency remained higher compared to other Western CLL (2.6-4.1%) hospital-based cohorts. Furthermore, we confirmed the poor-outcome for IGHV3-21 patients to be independent of mutational and stereotypy status. In paper II, genomic events in stereotyped IGHV3-21-subset #2, IGHV4-34-subset #4 and subset #16 and their non-stereotyped counterparts were investigated via SNP arrays (n=101). Subset #2 and non-subset #2 carried a higher frequency of events compared to subset #4. A high frequency of del(11q) was evident in IGHV3-21 patients particularly subset #2 cases, which may partially explain their poor-prognosis. In contrast, the lower prevalence of aberrations and absence of poor-prognostic alterations may reflect the inherent low-proliferative disease seen in subset #4 cases. In papers III and IV, differential methylation profiles in IGHV mutated and IGHV unmutated patients were identified using DNA-methylation microarrays. CLL prognostic genes (CLLU1, LPL), tumor-suppressor genes (TSGs) (ABI3, WISP3) and genes belonging to TGF-ß and NF-kB/TNFR1 pathways were differentially methylated between the subgroups. Additionally, the re-expression of methylated TSGs by use of methyl and deacetyl inhibitors was demonstrated. Interestingly, analysis of patient-paired diagnostic/follow-up samples and patient-matched lymph node (LN) and peripheral blood (PB) cases revealed global DNA methylation to be relatively stable over time and remarkably similar within the different compartments. Altogether, this thesis provides insight into the aberrant genomic and DNA methylation events in divergent CLL subgroups. Moreover this thesis helps distinguish the extent to which DNA methylation changes with respect to time and microenvironment in CLL.
15.	Campino, Susana, 1975- (författare) Genetic analysis of murine malaria 2003 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Malaria, an infectious disease caused by Plasmodium parasites, is one of the major world-scale health problems. Despite the efforts aimed at finding an effective way to control the disease, the success has been thwarted by the emergence of parasite drug resistance and mosquito resistance to insecticides. This thesis focuses on the genetic analysis of resistance to murine malaria induced by the lethal Plasmodium berghei ANKA using a wild-derived-inbred strain (WDIS). The aim of this thesis was to exploit the genetic diversity represented among WDIS for identifying loci contributing to resistance/susceptibility to murine malaria. The work included a genome-wide polymorphism survey using microsatellite markers performed on 10 WDIS. Comparisons of these strains to laboratory inbred strains confirmed a higher rate of polymorphism among the WDIS. We conclude that these WDIS represent repositories of unique naturally occurring genetic variability that may prove to be invaluable for the study of complex phenotypes. Next, we used the WDIS to search for novel phenotypes related to malaria pathogenesis. Whereas most laboratory strains were susceptible to experimental cerebral malaria (ECM) after infection with P. berghei ANKA, several WDIS were found to be resistant. To study the genetic inheritance of resistant/susceptibility to P. berghei ANKA infection we analysed backcross and F2 cohorts derived from crossing the WLA wild-derived strain with a laboratory mouse strain (C57BL/6). A novel phenotype represented by the cure of infection, clearance of parasitaemia and establishment of immunological memory was observed in the F2 progeny. The backcross progeny was used to genetically map one locus on chromosome 1 (Berr1) and one locus on chromosome 11 (Berr2) that mediate control of resistance to ECM induced by P. berghei ANKA. Genetic mapping using the F2 progeny showed that a locus on chromosome 1 (Berr1) and a locus on chromosome 9 (Berr3) were contributing to control survival time after infection with lethal Plasmodium. Finally, we identified, a locus on chromosome 4 (Berr4) that appears to control time of death due to hyperparasitaemia. This thesis underlines the value of using WDIS to reveal genetic factors involved in the aetiology of disease phenotypes. The characterisation of the genetic factors represented by the malaria resistance loci identified here are expected to provide a better understanding of the malaria pathology.
16.	Carlsson, Martin (författare) Phylogeography of the Adder, Vipera berus 2003 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract The phylogeography of a wide ranging temperate species, the adder, Vipera berus, was investigated using several genetic tools, with special emphasis on the post-glacial colonisation pattern of Fennoscandia. The area was colonised from two directions by adder populations representing different glacial refugia. The two populations meet in three places and the main contact zone is situated in Northern Finland. The two other contact zones are the result of dispersal across the Baltic Sea to the Umeå archepelago and South-Western Finland. Asymmetrically distributed nuclear genetic variation compared to mitochondrial DNA in the northern contact zone suggests a skewed gene flow from the east to the west across the zone. This pattern might reflect differences in dispersal among sexes and lineages, or may be accounted for by a selective advantage for nuclear variation of eastern origin among Fennoscandian adders.The phylogeographic pattern for adders across the entire species range was addressed by sequencing part of the mitochondrial genome and scoring microsatellite markers. The adder can be divided into three major genetic groups. One group is confined to the Balkan peninsula harbouring the distribution range of V. b. bosniensis. A second, well differentiated group is restricted to the Southern Alps. These two areas have probably served as refugia for adders during a number of ice ages for the adders. The third group is distributed across the remainder of the species’ range, from extreme Western Europe to Pacific Russia and can be further divided into one ancestral group inhabiting the Carpathians refugial area, and three more recent groups inhabiting areas west, north and east of the Alps. The adder provides an example of a species where the Mediterranean areas are housing endemic populations, rather than the sources for post-glacial continental colonisation. Continent-wide colonisation has instead occurred from up to three cryptic northern refugia.
17.	Cederquist, Kristina, 1971- (författare) Genetic and epidemiological studies of hereditary colorectal cancer 2005 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Lynch syndrome (Hereditary Nonpolyposis Colorectal Cancer, HNPCC) is the most common hereditary syndrome predisposing to colorectal cancer, accounting for 1-3% of all colorectal cancer. This multi-organ cancer predisposition syndrome is caused by mutations in the mismatch repair (MMR) genes, especially MLH1 and MSH2, and to lesser extents MSH6 and PMS2, which lead to widespread genetic instability and thus microsatellite instability (MSI). Hereditary cancer often manifests in two or more tumours in a single individual; 35-40% of Lynch syndrome patients have synchronous or metachronous tumours of the two major Lynch syndrome-related cancers: colorectal and endometrial. The main purposes of the work underlying this thesis were to identify persons at risk of Lynch syndrome or other types of hereditary colorectal cancer, to estimate the cancer risks associated with these predispositions and to identify the underlying genetic causes. A population-based cohort of 78 persons with double primary colorectal or colorectal and endometrial cancer was identified. Cancer risks in their 649 first-degree relatives were estimated in relation to tumour MSI status (positive or negative) and age at diagnosis (before or after 50 years of age) in the probands. The overall standardised incidence ratio was 1.69 (95% CI; 1.39-2.03). The highest risks for Lynch syndrome-associated cancers: (colorectal, endometrial, ovarian and gastric) were found in families with young MSI-positive probands, likely representing Lynch syndrome families. Importantly, no overall risk was found in families with old probands, irrespective of MSI status. Blood samples were available from 24 MSI-positive patients for mutation screening of MLH1, MSH2 and MSH6. Sequence variants or rearrangements predicted to affect protein function were found in 16 patients. Six novel variants were found: two large rearrangements, two truncating and two missense mutations. The missense mutations were found to segregate in the families. Studies of allele frequencies, MSI and loss of immunostaning in tumours from family members further supports the hypothesis that these missense changes play a role in Lynch syndrome, as do the non-conservative nature and evolutionary conservation of the amino acid exchanges. Five families had mutations in MLH1, five in MSH2, and six in MSH6. The unexpectedly large impact of MSH6 was in genealogical studies shown to be due to a founder effect. Cumulative risk studies showed that the MSH6 families, despite their late age of onset, have a high lifetime risk for all Lynch syndrome-related cancers, significantly higher in women (89% by age 80 years) than in men (69%). The gender differences are in part due to high endometrial (70%) and ovarian cancer risk (33%) in addition to the high colorectal cancer risk (60%). These findings are of great importance for counselling and surveillance of families with MSH6 mutations. Finally, in a large family with MSI-negative hereditary colorectal cancer for which the MMR genes and APC had been excluded as possible causes, a genome-wide linkage analysis was performed, resulting in a suggested linkage to chromosome 7. Conclusions: Relatives of probands with MSI-positive, double primary colorectal and endometrial cancer diagnosed before the age of 50 years have significantly increased risks of Lynch syndrome-related cancers. MSH6 mutations, which have unusually high impact in this study population due to a founder effect, confer high cumulative risks of cancer despite the generally late age of onset.
18.	Einarsdottir, Elisabet, 1974- (författare) Mapping genetic diseases in northern Sweden 2005 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract The population of northern Sweden has previously been shown to be well suited for the mapping of monogenic diseases. In this thesis we have tested the hypothesis that this population could also be used for efficient identification of risk genes for common diseases. In Paper I we have hypothesised that despite the admixture of Swedish, Finnish and Sami, the northern Swedish population consists of sub-populations geographically restricted by the main river valleys running through the region. This geographic isolation, in combination with founder effects and genetic drift, could represent a unique resource for genetic studies. On the other hand, it also underlines the importance of accounting for this e.g. in genetic association studies. To test this hypothesis, we studied the patterns of marriage within and between river valley regions and compared allelic frequencies of genetic markers between these regions. The tendency to find a spouse and live in the river valley where one was born is strong, and allelic frequencies of genetic markers vary significantly between adjacent regions. These data support our hypothesis that the river valleys are home to distinct sub-populations and that this is likely to affect mapping of genetic diseases in these populations. In Paper II, we tested the applicability of the population in mapping HSAN V, a monogenic disease. This disease was identified in only three consanguineous individuals suffering from a severe loss of deep pain perception and an impaired perception of heat. A genome-wide scan combined with sequencing of candidate genes resulted in the identification of a causative point mutation in the nerve growth factor beta (NGFB) gene. In Paper III, a large family with multiple members affected by familial forms of type 1 diabetes mellitus (T1DM) and autoimmune thyroiditis (AITD) was studied. This syndrome was mapped to the IDDM12 region on 2q33, giving positive lodscores when conditioning on HLA haplotype. The linkage to HLA and to the IDDM12 region thus confirmed previous reports of linkage and/or association of T1DM and AITD to these loci and provided evidence that the same genetic factors may be mediating these diseases. This also supported the feasibility of mapping complex diseases in northern Sweden by the use of familial forms of these diseases. In Paper IV, we applied the same approach to study type 2 diabetes mellitus (T2DM). A non-parametric genome-wide scan was carried out on a family material from northern Sweden, and linkage was found to the calpain-10 locus, a previously described T2DM-susceptibility gene on 2q37. Together, these findings demonstrate that selecting for familial forms of even complex diseases, and choosing families from the same geographical region can efficiently reduce the genetic heterogeneity of the disease and facilitate the identification of risk genes for the disease.
19.	Emahazion, Tesfai (författare) Genomic strategies towards the dissection of human complex disease 2001 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Complex diseases are caused by the activities of many genes acting in concert with environmental factors. A major challenge facing human genetic research is to unravel the genetic factors behind complex disorders. Oxidative damage by free oxygen radicals is believed to contribute to many complex disorders, particularly degenerative disorders such as Alzheimer's disease. A principle source of oxygen free radicals in a human cell is the mitochondrial Oxidative Phosphorylation pathway. This consists of five protein complexes whose subunits are encoded both by the mitochondrial and the nuclear genomes. The least genetically defined complex is Complex I, which comprises 41 genes, of which 34 are encoded by the nuclear genome and the remaining seven by the mitochondrial genome. By exploiting publicly available EST sequences and by radiation hybrid mapping, I successfully characterized and mapped 20 nuclear complex I genes to precise chromosomal locations. These genes can now be tested in positional candidate scenarios. Turning my attention to encephalomyopathies, I analyzed eight patients from Italy suffering severe myopathic disease. All eight individuals had confirmed Oxidative Phosphorylation Complex I biochemical deficiency but did not carry known mitochondrial DNA mutations. We mutation scanned over 20 Complex I cDNAs and found one rare alteration causing the substitution of an evolutionary conserved glycine to arginine at amino acid position 32 in the NDUFA1 gene. The pathogenic role of this mutation is unclear. Polymorphism is an important investigation tool in genetic research, without which disease genes could not be identified. Single nucleotide polymorphisms (SNPs) are one base differences within a population. They have high prevalence in the human genome, are easy to score, and are suitable for automated genotyping techniques. I therefore worked to identify 167 SNPs in 88 candidate genes for neurodegenerative disorders. These SNPs would provide a valuable resource for subsequent association analysis. Alzheimer's disease (AD) is a complex disease where many risk genes have been reported to be associated with the disease, but only one (APOE) has been independently replicated. I performed large scale (60 SNPs) association analysis to test for association with early onset form of AD. Despite observing 12 positive association signals in a set of 121 patients and matched controls, the signals were not robust enough to survive an independent confirmation study in a set of 117 patients and 176 controls.
20.	Fedorov, Vadim B. (författare) Phylogeography and mitochondrial DNA diversity in arctic lemmings 1998 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract The aim of this thesis is to evaluate the effects of Quaternary environmental fluctuations on the present patterns of mitochondrial DNA variation in the wood lemming, Myopus schisticolor, and the two genera of Arctic lemmings: collared lemmings, Dicrostonyx, and true lemmings, Lemmus.The phylogeographic pattern and the limited mtDNA diversity in the Scandinavian populations of the wood lemming reflect recent colonization by a limited number of founders during the Holocene boreal forest expansion.The mtDNA phylogeny in the two genera of Arctic lemmings gives no support for the existence of a single Beringian refungium since the mid Pleistocene. Isolation by intermittent inundation of the Bering Strait during the interglacials was most probably important for speciation in both genera, Dicrostonyx and Lemmus.Comparative phylogeography of the two genera of Arctic lemmings, Lemmus and Dicrostonyx gives evidence for the vicariant separation by the glacial barriers in the Eurasian Arctic. There is genetic support for the importance of the Asian Beringia as a refugial area for the tundra specialist, D. torquatus, during warming periods in the interglacials.There is no evidence for the direct effect of the last glaciation on the present level of mtDNA diversity and population structure in Arctic lemmings. Intrapopulation and intraregion mtDNA diversity estimates in each genus reflect the demographic events which were likely a result of environmental changes in the Holocene. Comparisons of mtDNA diversity estimates across the two genera of Arctic lemmings suggest that historically and presently codistributed Lemmus and Dicrostonyx responded differently to environmental changes resulting from the Holocene warming events in the Eurasian Arctic.The mtDNA phylogeny supports the importance of vicariant events generated by the glacial-interglacial periods for allopatric speciation in chromosomally variable collared lemmings, Dicrostonyx, in the North America Arctic. In the Eurasian Arctic, the congruence between the phylogeographic structure in mtDNA variation and geographic distribution of chromosome races indicates that the fragmentation by the glacial barriers during the late Pleistocene and bottleneck events due to warming events in the Holocene were important for the origin of intraspecific chromosome races in the collared lemming, D. torquatus
21.	Hagberg, Anette (författare) Expression profiling using manifold supports 2001 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Analyses of mRNA provides a condensed view of gene structure and quantitative analysis can reveal the induction of physiological or pathological gene expression programs. This thesis describes a new method for mRNA isolation, followed by sensitive real time detection via polymerase chain reaction (PCR), in order to quantitate transcripts of interest. Chimeric genes that result from chromosomal translocations can be used as disease-specific markers for the malignant clone to detect minimal residual diseases. It is important to detect an expanding clone as early as possible to increase the chance of a successful treatment. Accordingly, RT-PCR (reverse transcription PCR) of such chimeric transcripts has gained interest as a means to monitor patients due to its sensitivity. Expression of BCR-ABL in bone marrow or blood can be used as a measure of minimal residual disease (MRD) in patients with chronic myeloid leukemia. The newly described method for mRNA isolation was used to analyse the tumor burden in patient samples via real time detection using PCR. The proposed method constitues a promising, reproducible, and sensitive means to quantify BCR-ABL mRNA and it is suitable to monitor MRD in leukemic patients. Recombinant human erythropoietin (r-HuEpo) has an important role in the treatment of anemic patients. β-globin mRNA was monitored in order to elucidate if it could serve as a new marker for monitoring the response to r-HuEpo. Because of the high cost of EPO treatment, an early indicator of whether a patient responds to the therapy would be of great value. The response pattern for mRNA was compared to the reticulocyte count, levels of hemoglobin, transferrin receptor and ferritin in healty individuals receiving r-HuEpo or in controls. Following treatment, β-globin mRNA showed a more distinct increase compared to all other laboratory measurements and is therefore promising as a marker for the response to EPO treatment. The fourth project was undertaken to investigate fluctuations of mRNA expression levels for cytokines important for the rejection of xenotransplants. Porcine islet xenotransplantation could potentially solve the problem of the limited supply of suitable human donors for transplanation of islets, in order to offer a curative treatment of insulin dependent diabetes mellitus (IDDM). The rejection process was studied in the pig→rat model. Earlier studies reported a Th2 associated response. However, both morphological pattern and mRNA expression profiling supported the view that rejection is primarily due to a Th1 response.
22.	Ingman, Max, 1970- (författare) Mitochondria and Human Evolution 2003 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Mitochondrial DNA (mtDNA) has been a potent tool in studies of the evolution of modern humans, human migrations and the dynamics of human populations over time. The popularity of this cytoplasmic genome has largely been due to its clonal inheritance (in Man) allowing the tracing of a direct genetic line. In addition, a comparatively high rate of nucleotide substitution facilitates phylogenetic resolution among relatively closely related individuals of the same species.In this thesis, a statistically supported phylogeny based on complete mitochondrial genome sequences is presented which, for the first time, unambiguously places the root of modern human mitochondrial lineages in Africa in the last 200 thousand years. This conclusion provides strong support for the “recent African origin” hypothesis. Also, the complete genome data underline the problematic nature of traditional approaches to analyses of mitochondrial phylogenies.The dispersal of anatomically modern humans from the African continent is examined through single nucleotide polymorphism (SNP) and sequence data. These data imply an expansion from Africa about 57 thousand years ago and a subsequent population dispersal into Asia. The dispersal coincides with a major population division that may be the result of multiple migratory routes to East Asia.Also investigated is the question of a common origin for the indigenous peoples of Australia and New Guinea. Previous studies have been equivocal on this question with some presenting evidence for a common genetic origin and other proposing separate histories. Our data reveals an ancient genetic link between Australian Aborigines and the peoples of the New Guinea highlands.
23.	Jiao, Xiang (författare) Somatic Mutations in Breast Cancer Genomes : Discovery and Validation of Breast Cancer Genes 2012 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Breast cancer is the most common cancer in women worldwide. However, the genetic alterations that lead to breast cancer are not fully understood. This thesis aims to identify novel genes of potential mechanistic, diagnostic or therapeutic interest in breast cancers by mutational analysis and whole-genome sequencing.In paper I, sequencing of 36 previously identified candidate genes in 96 breast tumors with patient-matched normal DNA determined the somatic mutation prevalence of these candidate genes and identified additional mutations in Notch, NF-κB, PI3K, and Hedgehog pathways as well as in processes mediating DNA methylation, RNA processing and calcium signaling.In paper II, comparison of massively parallel mate-pair sequencing results of a human genome before and after phi29-mediated multiple displacement amplification (MDA) revealed that MDA introduces structural alteration artifacts, with an emphasis on false positive inversions, and impairs the sensitivity to detect true inversions. Therefore, MDA has limited value in sample preparation for whole-genome sequencing for structural alteration detection.In paper III, massively parallel paired-end sequencing identified gene rearrangements in 15 hormone receptor negative breast cancers. Forty validated rearrangements were predicted to directly affect 30 genes, involved in epigenetic regulation, cell mitosis, signalling transduction and glycolytic flux. RNA interference-based assays revealed the potential roles in cell growth of some affected genes, among which DDX10 was implicated to be involved in apoptosis.In paper IV, a method for statistical evaluation of putative translocations detected by massively parallel paired-end sequencing was proposed. In an application of this method to analyse translocations detected by cancer genome deep paired-end sequencing, 76 putative translocations were classified into four categories, with the majority likely to be caused by mismapping due to repetitive regions.Taken together, this thesis provides insights into genes and pathways mutated in sporadic breast cancer genomes, which broaden our understanding of the genetic basis of breast cancer and may ultimately facilitate the diagnosis and treatment of this disease.
24.	Josefsson, Agnetha (författare) Genetic and environmental risk factors of cervical carcinoma in Situ 2000 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Invasive cervical cancer is the third most common cancer form among women. In 1990 there were approximately 370,000 new cases diagnosed in the world. The risk of morbidity and mortality of cervical cancer among women is a problem, particularly in the developing countries.For the purpose of investigating various aspects of the natural history of cervical carcinoma, we have evaluated a PCR based fluorescent 5´exonuclease assay for typing and quantitating of human papillomavirus (HPV). Using this new technique, we have succeeded in detecting and quantitating HPV DNA from archival cervical smears and tumour biopsy specimens.To study the relationship between HPV 16 infection and risk of cancer in situ, including the effect of recurrent and persistent infection, we examined archival cervical smears collected from women over a time period of 26 years, spanning the time from infection to development of cancer in situ. Archival smears from 484 women with carcinoma in situ and the corresponding 619 individually matched controls were studied for presence for HPV 16. Women with multiple infected smears had an increased risk of cervical cancer development. For instance, women with three HPV infected smears had a dramatically elevated risk (OR=47.3 (95% CI 14.3-156.7)) of developing cervical cancer in situ.Further, we examined the relationship between amount of HPV 16 DNA and risk of developing cervical carcinoma in situ. A positive correlation was found between amount of HPV 16 DNA and risk of cervical cancer. The OR showed an approximately 70-fold (OR=68.8 95% CI 15.8-299.6) higher risk for women in the quintile with the highest amount of HPV 16 DNA in relation to women testing negative. This relationship was seen also when including only the first, cytologically normal, smear from each woman, taken on average 7.8 years before diagnosis. For young women (20-24 years old) in the quintile with the highest amount of HPV DNA the positive predictive value (PPV) was found to be 48%. Thus, HPV DNA amount appears to be a strong predictor of cancer risk.Finally, we evaluated a hypothesis concerning the importance of homozygosity for arginine (p53Arg) at codon 72 in the p53 gene for the risk of developing cervical cancer. We were unable to find any association between the genotype at codon 72 and risk of cervical cancer in our patient cohort, despite the large sample size.
25.	Karsten, Stanislav L. (författare) Molecular investigation of mucopolysaccharidosis type II (Hunter syndrome) in man 2000 Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract Mucopolysaccharidosis type II or Hunter syndrome is a rare lysosomal storage disorder caused by a deficiency in the enzyme iduronate-2-sulfatase (IDS). The disorder is inherited in an X-linked recessive fashion and has a broad spectrum of clinical phenotypes ranging from severe to mild. Different mutations in the IDS locus affecting enzyme function or stability explain a wide spectrum of clinical phenotypes.The aim of the work described in this thesis is to identify and analyse the mutations in the human iduronate-2-sulfatase (IDS) gene resulting in the development of Hunter syndrome. The DNA samples from 63 unrelated patients were analysed by Southern blot analysis, PCR, RT-PCR, SSCP and DNA sequencing. Among 55 mutant alleles, there were 33 missense/nonsense mutations, 6 mutations affecting splicing, and 12 major structural alterations resulting in various rearrangements. Of the 45 different mutations 26 were novel and unique. The study of large group of patients originated from Russia anddiagnosed by the same clinician allowed us to investigate the correlation between genotype and phenotype in the Hunter syndrome.The structural rearrangements and deletions of the entire IDS gene are observed in about 20% of the patients, suggesting the presence of recombinational "hotspots" in the locus. We have isolated and characterised the recombinational junctions from 9 unrelated MPSII patients with structural rearrangements in the IDS gene. It was shown that the IDS gene is frequently involved in the homologous recombination with the IDS-2 pseudogene resulting in inversions. Moreover, the presence of homologous regions in the IDS locuspossibly promotes nonhomologous recombination resulting in various intragenic deletions and complex rearrangements of the IDS gene. The models explaining recombinational events are proposed.The data reported in this thesis contributes to our understanding of the nature of mutation events and provides valuable insights into the mechanisms of DNA recombination. The knowledge of the IDS gene defects permits direct detection of carriers of that mutation, allows to make the prognostic predictions about clinical severity and gives us information about the functionally important parts of the iduronate-2-sulfatase.

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