Activity of microRNAs and transcription factors in Gene Regulatory Networks

In biological research, diverse high-throughput techniques enable the investigation of whole systems at the molecular level. The development of new methods and algorithms is necessary to analyze and interpret measurements of gene and protein expression and of interactions between genes and proteins. One of the challenges is the integrated analysis of gene expression and the associated regulation mechanisms. The two most important types of regulators, transcription factors (TFs) and microRNAs (miRNAs), often cooperate in complex networks at the transcriptional and post-transcriptional level and, thus, enable a combinatorial and highly complex regulation of cellular processes. For instance, TFs activate and inhibit the expression of other genes including other TFs whereas miRNAs can post-transcriptionally induce the degradation of transcribed RNA and impair the translation of mRNA into proteins. The identification of gene regulatory networks (GRNs) is mandatory in order to understand the underlying control mechanisms. The expression of regulators is itself regulated, i.e. activating or inhibiting regulators in varying conditions and perturbations. Thus, measurements of gene expression following targeted perturbations (knockouts or overexpressions) of these regulators are of particular importance. The prediction of the activity states of the regulators and the prediction of the target genes are first important steps towards the construction of GRNs. This thesis deals with these first bioinformatics steps to construct GRNs. Targets of TFs and miRNAs are determined as comprehensively and accurately as possible. The activity state of regulators is predicted for specific high-throughput data and specific contexts using appropriate statistical approaches. Moreover, (parts of) GRNs are inferred, which lead to explanations of given measurements. The thesis describes new approaches for these tasks together with accompanying evaluations and validations. This immediately defines the three main goals of the current thesis: 1. The development of a comprehensive database of regulator-target relation. Regulators and targets are retrieved from public repositories, extracted from the literature via text mining and collected into the miRSel database. In addition, relations can be predicted using various published methods. In order to determine the activity states of regulators (see 2.) and to infer GRNs (3.) comprehensive and accurate regulator-target relations are required. It could be shown that text mining enables the reliable extraction of miRNA, gene, and protein names as well as their relations from scientific free texts. Overall, the miRSel contains about three times more relations for the model organisms human, mouse, and rat as compared to state-of-the-art databases (e.g. TarBase, one of the currently most used resources for miRNA-target relations). 2. The prediction of activity states of regulators based on improved target sets. In order to investigate mechanisms of gene regulation, the experimental contexts have to be determined in which the respective regulators become active. A regulator is predicted as active based on appropriate statistical tests applied to the expression values of its set of target genes. For this task various gene set enrichment (GSE) methods have been proposed. Unfortunately, before an actual experiment it is unknown which genes are affected. The missing standard-of-truth so far has prevented the systematic assessment and evaluation of GSE tests. In contrast, the trigger of gene expression changes is of course known for experiments where a particular regulator has been directly perturbed (i.e. by knockout, transfection, or overexpression). Based on such datasets, we have systematically evaluated 12 current GSE tests. In our analysis ANOVA and the Wilcoxon test performed best. 3. The prediction of regulation cascades. Using gene expression measurements and given regulator-target relations (e.g. from the miRSel database) GRNs are derived. GSE tests are applied to determine TFs and miRNAs that change their activity as cellular response to an overexpressed miRNA. Gene regulatory networks can constructed iteratively. Our models show how miRNAs trigger gene expression changes: either directly or indirectly via cascades of miRNA-TF, miRNA-kinase-TF as well as TF-TF relations. In this thesis we focus on measurements which have been obtained after overexpression of miRNAs. Surprisingly, a number of cancer relevant miRNAs influence a common core of TFs which are involved in processes such as proliferation and apoptosis

miRSel: automated extraction of associations between microRNAs and genes from the biomedical literature.

Background MicroRNAs have been discovered as important regulators of gene expression. To identify the target genes of microRNAs, several databases and prediction algorithms have been developed. Only few experimentally confirmed microRNA targets are available in databases. Many of the microRNA targets stored in databases were derived from large-scale experiments that are considered not very reliable. We propose to use text mining of publication abstracts for extracting microRNA-gene associations including microRNA-target relations to complement current repositories. Results The microRNA-gene association database miRSel combines text-mining results with existing databases and computational predictions. Text mining enables the reliable extraction of microRNA, gene and protein occurrences as well as their relationships from texts. Thereby, we increased the number of human, mouse and rat miRNA-gene associations by at least three-fold as compared to e.g. TarBase, a resource for miRNA-gene associations. Conclusions Our database miRSel offers the currently largest collection of literature derived miRNA-gene associations. Comprehensive collections of miRNA-gene associations are important for the development of miRNA target prediction tools and the analysis of regulatory networks. miRSel is updated daily and can be queried using a web-based interface via microRNA identifiers, gene and protein names, PubMed queries as well as gene ontology (GO) terms. miRSel is freely available online at http://services.bio.ifi.lmu.de/mirse

The Use of Targeted Next Generation Sequencing to Explore Candidate Regulators of TGF-β1’s Impact on Kidney Cells

Aims/Hypothesis: Transforming growth factor-beta (TGF-β1) plays an important regulatory role in the progression of chronic kidney failure. Further, damage to kidney glomerular mesangial cells is central to the progression of diabetic nephropathy. The aim of this study was to explore the genetic associations between mRNA, microRNA, and epigenetics in mesangial cells in response to TGF-β1.Methods: The regulatory effects of TGF-β1 on mesangial cells were investigated at different molecular levels by treating mesangial cells with TGF-β1 for 3 days followed by genome-wide miRNA, RNA, DNA methylation, and H3K27me3 expression profiling using next generation sequencing (NGS).Results: Our results provide the first comprehensive, computationally integrated report of RNA-Seq, miRNA-Seq, and epigenomic analyses across all genetic variations, confirming the occurrence of DNA methylation and H3K27me3 in response to TGF-β1. Our findings show that the expression of KLF7 and Gja4 are involved in TGF-β1 regulated DNA methylation. Our data also provide evidence of the association between epigenetic changes and the expression of genes closely related to TGF-β1 regulation.Conclusion: This study has advanced our current knowledge of mechanisms that contribute to the expression of TGF-β1-regulated genes involved in the pathogenesis of kidney disease. The molecular underpinnings of TGF-β1 stimulation of kidney cells was determined, thereby providing a robust platform for further target exploration

Comparative analysis of AC and DC distribution system with respect to harmonic distortion considering daily load profile

Electrical energy is the most efficient and the cleanest form of energy at the moment that is being transmitted and distributed amongst end-users. From its earlier days, the AC system was preferred as an economical solution for transmission and distribution. However, the development in the power electronics technology and the evolution of highly efficient power electronic converters have established the resurgence of DC power system. Furthermore, the trend is shifting towards DC loads as various energy efficient appliances, such as DC inverter air conditioners, operate on DC nowadays. This further advocates the shift towards the DC power system. This research works is an effort to perform the comparative analysis of AC Distribution System (ACDS) and DC Distribution System (DCDS), with regards to power quality and harmonic distortion in particular. The comparison is performed considering load profile and load variation on daily basis. Simulations are performed in MATLAB. It has been concluded at the end that ACDS is better than DCDS in terms of power quality as total harmonic distortion of the DCDS under the same loading and same load variation during the whole day was significantly higher than that of ACDS

Fungal Sinusitis with Intracranial Extension in Immuno-competent Patients: Surgical Planning According to Relation to Eloquent Neurovascular Structures

AbstractObjectiveTo evaluate the results of combined neurosurgical and ENT surgical management for patients with fungal sinusitis with intracranial extension.MethodsWe managed 10 cases with fungal sinusitis with intracranial extension. This included 7 females and 3 males. Four patients presented with unilateral proptosis, 2 with deterioration of level of consciousness due to meningitis, 2 with chronic headache, one with epilepsy and one patient presented with trigeminal pain. All cases had long history of chronic headache and nasal obstruction. CT was done in all cases, MR in 8 patients, CT angiography and conventional cerebral angiography in one patient. Surgical intervention was decided according to the relation of the fungal granuloma to eloquent intracranial structures. Endonasal approach alone was used when the granuloma was not related to the optic nerve, internal carotid artery or cavernous sinus (n=3). Combined subfrontal and endonasal approach was used when the granuloma was closely related to one or more of these structures (n=5). Transcranial approach alone was done for 2 patients with isolated sphenoid fungal sinusitis that was associated with a mycotic internal carotid artery aneurysm in one patient and with a temporal lobe abscess in the other. In addition, antifungal treatment was used for 8-12 weeks. Patients were followed up clinically and radiologically for 6-36 month period.ResultsNo morbidity related to the operative procedures was recorded in the study group. One patient died two month post-operatively due to fungal meningitis. In survivors (n=9): headache and nasal obstruction improved, proptosis was corrected, epilepsy and trigeminal pain were controlled by medication. Follow-up CT showed eradication of the fungal granuloma in all survivors. Histopathological results showed mucormycosis (n=2), aspirgillosis (n=4), and no fungus (4 patients).ConclusionTeam work by ENT and neurosurgical staff and early diagnosis are mandatory in the management of fungal sinusitis with intracranial extension in immune-competent patients. Surgical planning according to the relation of fungal granuloma to eloquent neurovascular structures is the corner stone for save removal of granuloma

In vivo analgesic, antipyretic and anti-inflammatory activities of ethanol extract of Pericampylus glaucus in experimental animals

Pericampylus glaucus is a common Malaysian plant used traditionally in the treatments of joint pain, abdominal pain and headache. Hence, the present research was aimed to evaluate ethanolic extract of Pericampylus glaucus for analgesic, antipyretic and anti-inflammatory activities in experimental animals. The central and peripheral analgesic activity was determined by acetic acid induced writhing and hot plate method by examining the number of writhing and paw licking or jumping time. Meanwhile, the antipyretic activity was determined by Brewer’s yeast- test that induced pyrexia and carrageenan- that induced hind paw inflammation was used for anti-inflammatory activities. The ethanolic extract of Pericampylus glaucus at doses (300 and 600 mg/kg b.wt) and Ibuprofen (100 mg/kg (b.wt) was used as a reference drug in the whole experiment. Intraperitoneal administration of Pericampylus glaucus produced significant (p<0.01) inhibition in writhing response in acetic acid induced writhing test and dose-dependent (p<0.001) prolonged paw licking in hot plate test as compared to control (normal saline treated group). Similarly, significant (p<0.001) attenuation in lowering rectal temperature was noted in animal groups that were treated with ethanolic extract of Pericampylus glaucus at different doses. The attenuation was, almost the same as produced by ibuprofen treated group. Furthermore, Pericampylus glaucus extract also produced significant reduction in hind paw edema (p<0.001), 4 h after administration of carrageenan and inhibition was 60.19% and 42.17% as compared to control (normal saline treated group). The findings of this study indicated that Pericampylus glaucus possess significant analgesic, antipyretic and anti-inflammatory activities and could possibly be used in the management of fever, pain and inflammation

Anti-urolithiatic effect of Pericampylus glaucus against ethylene glycol induced urolithiasis in male sprague dawley rats

Pericampylus glaucus is a common plant used traditionally for the remedies of urinary and kidney stones. Hence, present research was aimed to investigate the antiurolithiatic effect of ethanol extract of P. glaucus against ethylene glycol induced urolithiasis in male Sprague Dawley rats. The nephrolithiasis was induced by ethylene glycol (0.75% v/v) in drinking water over a period of 30 days. The animals were divided into normal, lithiatic and extract treated group that received ethanolic extract of P. glaucus at dose rate of 200 and 400 mg/kg (b.wt) by mouth. The calcium, phosphate, urine pH, oxalate, magnesium in urine and histopathalogical examination of the kidney were observed. A significant (p<0.01) increased in the levels of phosphate, calcium, oxalate, and an imbalance in kidney with decreased magnesium excretion in urine were observed in ethylene glycol induced urolithiasis as compared to normal control. Treatment with ethanolic extract of P. glaucus at dose of 200 and 400 mg/kg (b.wt) decreased significantly (p<0.01) the hyperoxaluria, calcium, phosphate, oxalate with significant (p<0.05) increased retention of urinary magnesium and improvement in the renal function as matched to lithiatic group. The histopathological examination showed the deposition of a large number of calcium oxalate crystal in lithiatic group while smaller depositions in 200 mg/kg (b.wt) extract treated group. The results confirmed the antiurolithiatic effect of P. glaucus in male Sprague Dawley rats and validate its traditional uses against urinary stone

Post-traumatic stress disorder and association with low birth weight in displaced population following conflict in Malakand division, Pakistan: a case control study.

BACKGROUND:The northern part of the province of Khyber Pakhtunkhwa in Pakistan experienced armed conflict since September 2007 till the autumn of 2011. Conflict involved widespread insurgency activity and military intervention including in 2009 internally displacing the 2.5 million people of the valley of Swat to live in camps, with relatives, or in rented accommodation across the region for approximately 4 months. It was during this period the current study was conducted to determine whether Post-Traumatic Stress Disorder in pregnant women was independently associated with Low Birth Weight (LBW) in an area affected by conflict and militancy. METHODS:A case control study was conducted in tertiary care hospitals of district Peshawar, Khyber Pakhtunkhwa. Two hundred twenty-five cases (neonates with birth weight  2.5 kg) were enrolled within 24 h of delivery. Post-Traumatic Stress Disorder was assessed through the MINI Neuropsychiatric Interview 5.0, a validated questionnaire along with the birth weight of the newborn. Maternal anthropometry, anemia and other sociodemographic details were also obtained during data collection. Data was analyzed using statistical package (STATA version 14). Logistic regression analysis of the association between LBW and all variables collected with a p-value of < 0.25 on uni-variate analysis were entered. RESULTS:A total of 450 newborn and mother pairs participated in the study with 225 cases and 225 controls. On univariate analysis factors significantly associated with LBW include: less than 5 years of paternal schooling and PTSD. On logistic regression, PTSD was independently associated with low birth weight in the presence of other factors like maternal/paternal schooling, gravida, history of preterm, BMI of the mother and maternal anemia. CONCLUSION:PTSD was found to be independently associated with LBW. In light of the current findings and other similar literature, intervention programs should be considered for pregnant women exposed to traumatic events