3-D data handling and registration of multiple modality medical images

The many different clinical imaging modalities used in diagnosis and therapy deliver two different types of information: morphological and functional. Clinical interpretation can be assisted and enhanced by combining such information (e.g. superimposition or fusion). The handling of such data needs to be performed in 3-D. Various methods for registration developed by other authors are reviewed and compared. Many of these are based on registering external reference markers, and are cumbersome and present significant problems to both patients and operators. Internal markers have also been used, but these may be very difficult to identify. Alternatively, methods based on the external surface of an object have been developed which eliminate some of the problems associated with the other methods. Thus the methods which have been extended, developed, and described here, are based primarily on the fitting of surfaces, as determined from images obtained from the different modalities to be registered. Annex problems to that of the surface fitting are those of surface detection and display. Some segmentation and surface reconstruction algorithms have been developed to identify the surface to be registered. Surface and volume rendering algorithms have also been implemented to facilitate the display of clinical results. An iterative surface fitting algorithm has been developed based on the minimization of a least squares distance (LSD) function, using the Powell method and alternative minimization algorithms. These algorithms and the qualities of fit so obtained were intercompared. Some modifications were developed to enhance the speed of convergence, to improve the accuracy, and to enhance the display of results during the process of fitting. A common problem with all such methods was found to be the choice of the starting point (the initial transformation parameters) and the avoidance of local minima which often require manual operator intervention. The algorithm was modified to apply a global minimization by using a cumulative distance error in a sequentially terminated process in order to speed up the time of evaluating of each search location. An extension of the algorithm into multi-resolution (scale) space was also implemented. An initial global search is performed at coarse resolution for the 3-D surfaces of both modalities where an appropriate threshold is defined to reject likely mismatch transformations by testing of only a limited subset of surface points. This process is used to define the set of points in the transformation space to be used for the next level of resolution, again with appropriately chosen threshold levels, and continued down to the finest resolution level. All these processes were evaluated using sets of well defined image models. The assessment of this algorithm for 3-D surface registration of data from (3-D) MRI with MRI, MRI with PET, MRI with SPECT, and MRI with CT data is presented, and clinical examples are illustrated and assessed. In the current work, the data from multi-modality imaging of two different types phantom (e.g. Hoffman brain phantom, Jaszczak phantom), thirty routinely imaged patients and volunteer subjects, and ten patients with setting external markers on their head were used to assess and verify 3-D registration. The accuracy of the sequential multi-resolution method obtained by the distance values of 4-10 selected reference points on each data set gave an accuracy of 1.44±0.42 mm for MR-MR, 1.82±0.65 for MR-CT, 2.38±0.88 for MR-PET, and 3.17±1.12 for MR-SPECT registration. The cost of this process was determined to be of the order of 200 seconds (on a Micro-VAX II), although this is highly dependent on some adjustable parameters of the process (e.g. threshold and the size of the geometrical transformation space) by which the accuracy is aimed

Bioinformatics analysis of intron retention events associated with the minor spliceosome

In the Eukaryotes, DNA sequences in genes are often interrupted by non-coding sequences called introns. These sequences are removed from the transcripts via a process known as splicing either while the genes are being transcribed (co-transcriptionally) or after transcription (posttranscriptionally). In higher eukaryotes two separate pre-mRNA splicing machineries have been described: the U12-dependent spliceosome which is responsible for splicing of approximately 700-800 unique introns (known as the U12-type introns), and the U2-dependent spliceosome responsible for splicing all other introns (known as the U2-type introns). The two intron types show divergent sequence elements in their 5' splice site and branch point sequences. In addition, earlier reports have indicated that U12-type introns are spliced with a slower rate comparing to the U2-type introns, suggesting that the splicing of U12-type introns is rate-limiting to the expression of the U12-type intron containing genes. This slower splicing is manifested as unspliced or retained U12-type introns in the otherwise fully processed mRNA products. In this work I developed a novel computational tool called the intron-exon retention estimator (IntEREst) which allows accurate detection, quantification and differential analysis of the intron retention levels from RNAseq data. Additional features of IntEREst include a tool for identification of U12-type introns, and a number of tools to compare the retention levels of userdefined subclasses of introns across several samples. An already published RNAseq dataset (available under accession GSE63816 in NCBI Gene expression Omnibus database) from patients and control subjects of myelodysplastic syndrome (MDS) was used to assess the functionality by benchmarking IntEREst. This dataset included RNAseq data from MDS patients featuring mutations in the ZRSRS2 gene that functions in the recognition of U12-type introns, and from control subjects that were either healthy or MDS patients without ZRSR2 mutations. Additionally, I used a Maize dataset consisting of samples with mutated and wild-type RGH3 gene, which is an ortholog of human ZRSR2. My results indicate that IntEREst is a reliable tool for analyzing intron retention events from RNAseq data producing comparable or better results than the other similar methods. I used IntEREst to globally compare the retention of the U12-type introns to that of U2-type introns. I found that U12-type introns show on average a 2-fold higher retention levels compared to that of U2-type introns both in human and plant cells. This result recapitulates the findings from earlier studies using a small set of selected genes and generalizes the increased intron retention of U12-type introns to a genome-wide scale. Furthermore, the results of this work provide evidence that transcripts containing unspliced U12-type introns are degraded in the nucleus by the nuclear exosome. Together, these results support the hypothesis that U12-type introns are globally spliced less efficiently than the U2-type introns and can thus regulate the rate of mature mRNA formation with the genes containing U12-type introns. Additionally, intron retention analysis of human/plant cells containing mutations in the U12-dependent spliceosome showed that such defects lead to a further increase in the levels of unspliced U12-type introns. In conclusion, this thesis extends current knowledge concerning the significance of the correct splicing of U12-type introns and the consequences of their abnormal splicing. Furthermore, it describes a combination of available tools together with a novel software tool (i.e. IntEREst) that can be used to measure and compare the efficiency and accuracy of RNA splicing across multiple samples. We show that these tools can reveal valuable information about the molecular mechanisms involved in various conditions, e.g. diseases caused by defective spliceosome.Aitotumallisilla geenien DNA-sekvenssit sisältävät usein introneiksi kutsuttuja ei-koodaavia jaksoja. Nämä sekvenssit poistetaan transkripteista silmukoinniksi kutsutun prosessin kautta, joka voi tapahtua joko transkription aikana (kotranskriptionaalisesti) tai transkription jälkeen (posttranskriptionaalisesti). Korkeammilla aitotumallisilla on kuvattu kaksi erillistä silmukointikoneistoa: U12-tyypin spliseosomi, joka vastaa noin 700–800:n intronin silmukoinnista (U12-tyypin intronit), sekä U2-tyypin spliseosomi, joka vastaa kaikkien muiden intronien silmukoinnista (U2-tyypin intronit). Näissä kahdessa intronityypissä on erilaiset 5’-silmukointikohdan ja intronin haarautumiskohdan sekvenssielementit. Tämän lisäksi aiemmat tutkimukset ovat viitanneet siihen, että U12-tyypin intronit silmukoidaan hitaammin U2-tyypin introneihin verrattuna, minkä seurauksena U12-tyypin intronien silmukointi saattaa rajoittaa näitä introneja sisältävien geenien ekspressiota. Hidas silmukointi ilmenee silmukoimattomina tai retentoituina U12-tyypin introneina muuten täysin prosessoiduissa lähetti-RNA:issa. Tässä työssä olen kehittänyt uuden ohjelmistotyökalun nimeltä Intron-Exon Retention Estimator (IntEREst), joka mahdollistaa intronien retention havaitsemisen, kvantifikaation sekä retentiotasojen vertailun RNAsekvensointidataa käyttäen. IntEREstin muihin ominaisuuksiin kuuluvat työkalu U12-tyypin intronien tunnistamiseen sekä useita eri työkaluja käyttäjän määrittelemien intronien alaluokkien retentiotasojen vertailuun useiden näytteiden välillä. IntEREstin vertailuanalyysiin hyödynnettiin jo julkaistua RNAsekvensointidataa myelodysplastista oireyhtymää (MDS) sairastavailta potilailta ja verrokkiyksilöiltä (saatavilla tunnuksella GSE63816 NCBI Gene Expression Omnibus -tietokannasta). Tämä datasetti sisälsi RNA-sekvensointidataa MDS-potilailta, joilla on mutaatioita U12-tyypin intronien tunnistuksessa toimivassa ZRSR2-geenissä, sekä verrokkiyksilöiltä jotka olivat joko terveitä yksilöitä tai MDS-potilaita, joilla ei ole ZRSR2-mutaatioita. Tämän lisäksi käytin maissista peräisin olevaa datasettiä, joka koostui maissin ZRSR2ortologin, RGH3-geenin, suhteen villityyppisistä tai mutanteista näytteistä. Tulokseni viittaavat siihen, että IntEREst on luotettava työkalu intronien retention analyysiin RNA-sekvensointidatasta, joka tuottaa muiden menetelmien kanssa vertailukelpoisia tai niitä parempia tuloksia. Hyödynsin IntEREstiä U12-tyypin ja U2-tyypin intronien retention vertailuun globaalilla tasolla. Osoitin, että U12-tyypin inronien retentiotaso on keskimäärin kaksi kertaa korkeampi U2-tyypin introneihin verrattuna sekä ihmis- että kasvisoluissa. Tämä tulos toistaa aiemmissa tutkimuksissa muutamilla valikoiduilla geeneillä tehdyt löydökset sekä yleistää U12-tyypin intronien korkeamman retention genominlaajuiselle tasolle. Tämän lisäksi työn tulokset osoittavat, että silmukoimattomia U12-tyypin introneja sisältävät transkriptit hajotetaan tumassa tumaeksosomin toimesta. Yhdessä nämä tulokset tukevat hypoteesia, jonka mukaan U12-tyypin intronit silmukoidaan heikommalla tehokkuudella kuin U2-tyypin intronit ja ne voivat näin ollen säädellä U12-tyypin introneja sisältävistä geeneistä tuotettavan lähetti-mRNA:n muodostumisnopeutta. Lisäksi U12-tyypin spliseosomin mutaatioita sisältävien ihmis- ja kasvisolujen introniretentioanalyysi osoitti, että viat silmukoinnissa kohottavat entisestään silmukoimattomien U12tyypin intronien tasoja. Tämä väitöskirja laajentaa nykyistä tietämystä U12-tyypin intronien oikean silmukoinnin merkityksestä ja virheellisen silmukoinnin seurauksista. Lisäksi työssä kuvataan olemassa olevia työkaluja sekä uusi ohjelmistotyökalu (IntEREst), joita voidaan käyttää RNA:n silmukoinnin tehokkuuden ja tarkkuuden mittaamiseen ja vertailuun useiden näytteiden välillä. Osoitamme, että nämä työkalut voivat tuottaa arvokasta tietoa molekyylitason mekanismeista eri olosuhteissa, muun muassa viallisesti toimivasta silmukointikoneistosta johtuvissa sairauksissa

Involved brain areas in processing of Persian classical music: an fMRI study

AbstractThe purpose of this study is to investigate the neurological process of the rhythm in Persian classical music by using fMRI. The test consists of two groups of no rhythmic and rhythmic pieces that has examined on 12 right-handed musicians. The result showed that no rhythmic Persian pieces activated right middle frontal gyrus, right middle temporal gyrus, left planum temporal and right superior temporal gyrus, and rhythmic pieces activated left frontal pole, left inferior frontal gyrus and left suramarginal. These results are based on the laterality and hierarchical models

IntEREst: Intron-exon retention estimator

Background: In-depth study of the intron retention levels of transcripts provide insights on the mechanisms regulating pre-mRNA splicing efficiency. Additionally, detailed analysis of retained introns can link these introns to post-transcriptional regulation or identify aberrant splicing events in human diseases. Results: We present IntEREst, Intron-Exon Retention Estimator, an R package that supports rigorous analysis of non-annotated intron retention events (in addition to the ones annotated by RefSeq or similar databases), and support intra-sample in addition to inter-sample comparisons. It accepts binary sequence alignment/map (.bam) files as input and determines genome-wide estimates of intron retention or exon-exon junction levels. Moreover, it includes functions for comparing subsets of user-defined introns (e.g. U12-type vs U2-type) and its plotting functions allow visualization of the distribution of the retention levels of the introns. Statistical methods are adapted from the DESeq2, edgeR and DEXSeq R packages to extract the significantly more or less retained introns. Analyses can be performed either sequentially (on single core) or in parallel (on multiple cores). We used IntEREst to investigate the U12- and U2-type intron retention in human and plant RNAseq dataset with defects in the U12-dependent spliceosome due to mutations in the ZRSR2 component of this spliceosome. Additionally, we compared the retained introns discovered by IntEREst with that of other methods and studies. Conclusion: IntEREst is an R package for Intron retention and exon-exon junction levels analysis of RNA-seq data. Both the human and plant analyses show that the U12-type introns are retained at higher level compared to the U2-type introns already in the control samples, but the retention is exacerbated in patient or plant samples carrying a mutated ZRSR2 gene. Intron retention events caused by ZRSR2 mutation that we discovered using IntEREst (DESeq2 based function) show considerable overlap with the retained introns discovered by other methods (e.g. IRFinder and edgeR based function of IntEREst). Our results indicate that increase in both the number of biological replicates and the depth of sequencing library promote the discovery of retained introns, but the effect of library size gradually decreases with more than 35 million reads mapped to the introns.Peer reviewe

IntEREst: Intron-exon retention estimator

Background: In-depth study of the intron retention levels of transcripts provide insights on the mechanisms regulating pre-mRNA splicing efficiency. Additionally, detailed analysis of retained introns can link these introns to post-transcriptional regulation or identify aberrant splicing events in human diseases. Results: We present IntEREst, Intron-Exon Retention Estimator, an R package that supports rigorous analysis of non-annotated intron retention events (in addition to the ones annotated by RefSeq or similar databases), and support intra-sample in addition to inter-sample comparisons. It accepts binary sequence alignment/map (.bam) files as input and determines genome-wide estimates of intron retention or exon-exon junction levels. Moreover, it includes functions for comparing subsets of user-defined introns (e.g. U12-type vs U2-type) and its plotting functions allow visualization of the distribution of the retention levels of the introns. Statistical methods are adapted from the DESeq2, edgeR and DEXSeq R packages to extract the significantly more or less retained introns. Analyses can be performed either sequentially (on single core) or in parallel (on multiple cores). We used IntEREst to investigate the U12- and U2-type intron retention in human and plant RNAseq dataset with defects in the U12-dependent spliceosome due to mutations in the ZRSR2 component of this spliceosome. Additionally, we compared the retained introns discovered by IntEREst with that of other methods and studies. Conclusion: IntEREst is an R package for Intron retention and exon-exon junction levels analysis of RNA-seq data. Both the human and plant analyses show that the U12-type introns are retained at higher level compared to the U2-type introns already in the control samples, but the retention is exacerbated in patient or plant samples carrying a mutated ZRSR2 gene. Intron retention events caused by ZRSR2 mutation that we discovered using IntEREst (DESeq2 based function) show considerable overlap with the retained introns discovered by other methods (e.g. IRFinder and edgeR based function of IntEREst). Our results indicate that increase in both the number of biological replicates and the depth of sequencing library promote the discovery of retained introns, but the effect of library size gradually decreases with more than 35 million reads mapped to the introns.Peer reviewe

Biclustering Methods: Biological Relevance and Application in Gene Expression Analysis

Peer reviewe

Multiple Isoforms of ANRIL in Melanoma Cells: Structural Complexity Suggests Variations in Processing

The long non-coding RNA ANRIL, antisense to the CDKN2B locus, is transcribed from a gene that encompasses multiple disease-associated polymorphisms. Despite the identification of multiple isoforms of ANRIL, expression of certain transcripts has been found to be tissue-specific and the characterisation of ANRIL transcripts remains incomplete. Several functions have been associated with ANRIL. In our judgement, studies on ANRIL functionality are premature pending a more complete appreciation of the profusion of isoforms. We found differential expression of ANRIL exons, which indicates that multiple isoforms exist in melanoma cells. In addition to linear isoforms, we identified circular forms of ANRIL (circANRIL). Further characterisation of circANRIL in two patient-derived metastatic melanoma cell lines (NZM7 and NZM37) revealed the existence of a rich assortment of circular isoforms. Moreover, in the two melanoma cell lines investigated, the complements of circANRIL isoforms were almost completely different. Novel exons were also discovered. We also found the family of linear ANRIL was enriched in the nucleus, whilst the circular isoforms were enriched in the cytoplasm and they differed markedly in stability. With respect to the variable processing of circANRIL species, bioinformatic analysis indicated that intronic Arthrobacter luteus (Alu) restriction endonuclease inverted repeats and exon skipping were not involved in selection of back-spliced exon junctions. Based on our findings, we hypothesise that “ANRIL” has wholly distinct dual sets of functions in melanoma. This reveals the dynamic nature of the locus and constitutes a basis for investigating the functions of ANRIL in melanoma.Peer reviewe

A Different Olfactory Perception in Anosmic Patients: Evidence from Functional MRI

Olfactory system is a vital sensory system in mammals, giving them the ability to connect with their environment. Anosmia, or the complete loss of olfaction ability, which could be caused by injuries, is an interesting topic for inspectors with the aim of diagnosing patients. Sniffing test is currently utilized to examine if an individual is suffering from anosmia; however, functional Magnetic Resonance Imaging (fMRI) provides unique information about the structure and function of the different areas of the human brain, and therefore this noninvasive method could be used as a tool to locate the olfactory-related regions of the brain. In this study, by recruiting 31 healthy and anosmic individuals, we investigated the neural BOLD responses in the olfactory cortices following two odor stimuli, rose and eucalyptus, by using a 3T MR scanner. Comparing the two groups, we observed a network of brain areas being more active in the normal individuals when smelling the odors. In addition, a number of brain areas also showed an activation decline during the odor stimuli, which is hypothesized as a resource allocation deactivation. This study illustrated alterations in the brain activity between the normal individuals and anosmic patients when smelling odors, and could potentially help for a better anosmia diagnosis in the future

Longitudinal Effects of Bumetanide on Neuro-Cognitive Functioning in Drug-Resistant Epilepsy

Antiepileptic drugs (AEDs) have repeatedly shown inconsistent and almost contradictory effects on the neurocognitive system, from substantial impairments in processing speed to the noticeable improvement in working memory and executive functioning. Previous studies have provided a novel insight into the cognitive improvement by bumetanide as a potential antiepileptic drug. Through the current investigation, we evaluated the longitudinal effects of bumetanide, an NKCC1 co-transporter antagonist, on the brain microstructural organization as a probable underlying component for cognitive performance. Microstructure assessment was completed using SPM for the whole brain assay and Freesurfer/TRACULA for the automatic probabilistic tractography analysis. Primary cognitive operations including selective attention and processing speed, working memory capacity and spatial memory were evaluated in 12 patients with a confirmed diagnosis of refractory epilepsy. Participants treated with bumetanide (2 mg/ day) in two divided doses as an adjuvant therapy to their regular AEDs for 6 months, which followed by the re-assessment of their cognitive functions and microstructural organizations. Seizure frequency reduced in eight patients which accompanied by white matter reconstruction; fractional anisotropy (FA) increased in the cingulum-cingulate gyrus (CCG), anterior thalamic radiation (ATR), and temporal part of the superior longitudinal fasciculus (SLFt) in correlation with the clinical response. The voxel-based analysis in responder patients revealed increased FA in the left hippocampus, right cerebellum, and right medial temporal lobe, while mean diffusivity (MD) values reduced in the right occipital lobe and cerebellum. Microstructural changes in SLFt and ATR accompanied by a reduction in the error rate in the spatial memory test. These primary results have provided preliminary evidence for the effect of bumetanide on cognitive functioning through microstructural changes in patients with drug-resistant epilepsy