28,585 research outputs found

    Estabelecimento de fibroblastos derivados da pele para o estudo da fosforilação de proteínas na Distrofia Miotónica tipo 1

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    Muscular dystrophies are a molecularly, genetically, and clinically heterogeneous group of disorders characterized mainly by progressive muscle weakness and degeneration. Within this group, the most common muscular dystrophy in adults is Myotonic Dystrophy type 1 (DM1), an inherited autosomal dominant disorder caused by an expansion of the (CTG)n trinucleotide repeats on the 3‚Äô untranslated region of the DMPK gene. Patients with DM1 not only present muscular symptoms such as myotonia and muscle wasting, but also extramuscular symptoms such as cataracts, cardiac conduction abnormalities and insulin resistance. In DM1, the increased length of triplet expansions leads to the accumulation of (CUG)n mRNA, that forms hairpin-like structures in the nucleus, leading to a toxic ‚Äúgain-of-function‚ÄĚ that deregulates RNA binding proteins such as MBNL1 and CUGBP1. This consequently affects alternative splicing of different mRNAs, which damages the normal function of different signalling pathways regulated through phosphorylation, an important regulatory mechanism. To understand the different impaired phosphorylation signalling pathways affected in DM1 we firstly conducted a systematic review about protein phosphorylation in DM1. The results provided a compilation of the altered protein phosphorylation events, namely the signalling pathways which regulate key cellular events. Some main findings are the underactivation of signalling pathways, such as AKT/mTOR and AMPK upon insulin signalling and starvation conditions, respectively. Also, myoblast differentiation was impaired since, during differentiation, there was an increase of activity of signalling pathways that stimulate cell proliferation (e.g. MEK/ERK, PKR/PERK) and a decrease of important proteins for muscle development, such as DMPK. In order to be able to study the mechanisms underlying the impaired signalling pathways described in the systematic review, it is necessary to establish DM1 cell models. For that reason, fibroblasts have been widely used for the study of this disorder due to its versatility and easy manipulation. We then successfully established skin-derived human fibroblast cell lines from patients with DM1 through a skin punch biopsy explant. These cell lines were subsequently characterized through indirect immunocytochemistry using a fibroblast-specific marker TE-7. The intracellular levels and localization of DMPK were also evaluated. It was possible to detect differences, although not statistically significant, of a reduced expression of DMPK in DM1-derived fibroblasts from late and juvenile onset with controls. To conclude, these fibroblasts can be an important cell model for the study of phosphorylation pathways and other mechanisms, such as nuclear envelope alterations and being an important research tool for DM1. As future perspectives, these cell models can be used to study protein phosphatases, such as PP1 and PP2, since there is not enough evidence of how these are altered in DM1 and could highly contribute to unravel new molecular mechanisms.As distrofias musculares s√£o um grupo de patologias cl√≠nica e geneticamente heterog√©neas, caracterizadas por fraqueza e degenera√ß√£o muscular progressivas. Dentro deste grupo, a distrofia muscular mais comum em adultos √© a distrofia miot√≥nica tipo 1 (DM1), uma doen√ßa heredit√°ria autoss√≥mica dominante causada por uma expans√£o das repeti√ß√Ķes de tripletos (CTG)n na regi√£o 3' n√£o traduzida do gene DMPK. Os pacientes com DM1 apresentam n√£o s√≥ sintomas musculares, como miotonia e perda de massa muscular, mas tamb√©m extramusculares, como cataratas, problemas na condu√ß√£o card√≠aca e resist√™ncia √† insulina. Na DM1, o aumento das expans√Ķes CTG levam ao ac√ļmulo de mRNA (CUG)n, que forma estruturas em hairpin no n√ļcleo, levando a um "ganho de fun√ß√£o" t√≥xico que desregula prote√≠nas de liga√ß√£o ao RNA, como a MBNL1 e CUGBP1. Isso, consequentemente, afeta o splicing alternativo de diferentes mRNAs, o que prejudica a fun√ß√£o normal de diferentes vias de sinaliza√ß√£o reguladas por fosforila√ß√£o, um importante mecanismo regulat√≥rio. Para perceber as diferentes vias de sinaliza√ß√£o de fosforila√ß√£o afetadas na DM1, executamos uma revis√£o sistem√°tica sobre a fosforila√ß√£o de prote√≠nas em DM1. Os resultados forneceram uma compila√ß√£o das vias de sinaliza√ß√£o alteradas e que regulam eventos celulares chave. Alguns dos principais resultados s√£o a reduzida ativa√ß√£o das vias da AKT/mTOR e da AMPK quando estimuladas com insulina ou com condi√ß√Ķes de priva√ß√£o de nutrientes, respetivamente. Adicionalmente, a miog√©nese estava tamb√©m alterada devido ao aumento de vias estimuladoras de prolifera√ß√£o celular (e.g. MEK/ERK, PKR/PERK) e uma diminui√ß√£o de prote√≠nas importantes para o desenvolvimento muscular, como a DMPK. Para poder estudar os mecanismos subjacentes √†s vias de sinaliza√ß√£o prejudicadas descritas na revis√£o sistem√°tica, √© necess√°rio estabelecer modelos de c√©lulas DM1. Por esse motivo, os fibroblastos t√™m sido amplamente utilizados para o estudo dessa doen√ßa devido √† sua versatilidade e f√°cil manipula√ß√£o. Em seguida, estabelecemos com sucesso linhas de c√©lulas de fibroblastos humanos derivadas da pele de pacientes com DM1 atrav√©s de um explante de bi√≥psia cut√Ęnea. Essas linhas celulares foram posteriormente caracterizadas por imunocitoqu√≠mica indireta usando um marcador espec√≠fico para fibroblastos TE-7. Os n√≠veis intracelulares e a localiza√ß√£o de DMPK tamb√©m foram avaliados. Foi poss√≠vel detetar diferen√ßas, embora n√£o estatisticamente significativas, de uma express√£o reduzida de DMPK em fibroblastos derivados de DM1 com fen√≥tipos de in√≠cio tardio e juvenil comparando com controlos. Concluindo, esses fibroblastos podem ser um importante modelo celular para o estudo das vias de fosforila√ß√£o e outros mecanismos, como altera√ß√Ķes do envelope nuclear, sendo uma importante ferramenta de pesquisa para DM1. Como perspetivas futuras, estas linhas celulares podem ser usadas para estudar as fosfatases, como a PP1 e PP2, uma vez que n√£o h√° evid√™ncias suficientes de como estas se alteram no DM1 e podem contribuir fortemente para desvendar novos mecanismos moleculares.Mestrado em Biomedicina Molecula

    Low Protein Programming Causes Increased Mitochondrial Fusion and Decreased Oxygen Consumption in the Hepatocytes of Female Rats

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    The liver is one of the major organs involved in the regulation of glucose and lipid homeostasis. The effectiveness of metabolic activity in hepatocytes is determined by the quality and quantity of its mitochondria. Mitochondrial function is complex, and they act via various dynamic networks, which rapidly adapt to changes in the cellular milieu. Our present study aims to investigate the effects of low protein programming on the structure and function of mitochondria in the hepatocytes of adult females. Pregnant rats were fed with a control or isocaloric low-protein diet from gestational day 4 until delivery. A normal laboratory chow was given to all dams after delivery and to pups after weaning. The rats were euthanized at 4 months of age and the livers were collected from female offspring for investigating the mitochondrial structure, mtDNA copy number, mRNA, and proteins expression of genes associated with mitochondrial function. Primary hepatocytes were isolated and used for the analysis of the mitochondrial bioenergetics profiles. The mitochondrial ultrastructure showed that the in utero low-protein diet exposure led to increased mitochondrial fusion. Accordingly, there was an increase in the mRNA and protein levels of the mitochondrial fusion gene Opa1 and mitochondrial biogenesis genes Pgc1a and Essra, but Fis1, a fission gene, was downregulated. Low protein programming also impaired the mitochondrial function of the hepatocytes with a decrease in basal respiration ATP-linked respiration and proton leak. In summary, the present study suggests that the hepatic mitochondrial dysfunction induced by an in utero low protein diet might be a potential mechanism linking glucose intolerance and insulin resistance in adult offspring

    Nitrite and insulin lower the oxygen cost of ATP synthesis in skeletal muscle cells by pleiotropic stimulation of glycolysis

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    Dietary nitrate lowers the oxygen cost of submaximal exercise, but precise mechanistic insight into how this occurs is lacking. Research suggests that dietary nitrate may render oxidative ATP synthesis more efficient, but evidence is inconclusive at present. This thesis aimed to establish how nitrite (a reduced form of nitrate) affects the bioenergetics of cultured skeletal muscle cells. Comparison between the acute effects of nitrite and insulin, a hormonal regulator of muscle function that increases mitochondrial efficiency, was explored to assess possible mechanistic overlap. Calculation of real-time intracellular ATP synthesis rates from simultaneous oxygen consumption and medium acidification measurements revealed the effects of sodium nitrite and insulin on intact rat (L6) myoblasts and myotubes. These extracellular flux data were also used to determine how mitochondrial and glycolytic ATP supply is used to fuel ATP-demanding processes. The data presented in this thesis revealed that both nitrite and insulin acutely stimulate glycolytic ATP synthesis. This stimulation occurs without significant mitochondrial ATP supply changes, thus increasing the glycolytic index of myocytes. Consequently, nitrite and insulin lower the oxygen cost of cellular ATP supply. Notably, insulin lowers oxygen consumption linked to mitochondrial proton leak, thus increasing mitochondrial efficiency. Nitrite does not improve coupling efficiency in myoblasts or myotubes. Further investigations revealed that stimulation of glycolytic ATP supply is not secondary to increased glucose availability. In myotubes, glycolytic stimulation persists in the presence of a mitochondrial uncoupler, suggesting that glycolysis is increased directly. In myoblasts, stimulation is annulled by uncoupler, suggesting that glycolysis increases indirectly, via increased ATP consumption. The molecular targets of nitrite and insulin remain unclear, but the data exclude stimulation of protein synthesis. Together, the data demonstrate that nitrite and insulin lower the oxygen cost of ATP synthesis in skeletal muscle cells by pleiotropic stimulation of glycolysis. The data inform the ongoing debate regarding the mechanism by which dietary nitrate lowers the oxygen cost of exercise, suggesting a push toward a more glycolytic phenotype. Such mechanistic insight is crucial for achieving the full translational potential of dietary nitrate

    In vitro investigation of the effect of disulfiram on hypoxia induced NFőļB, epithelial to mesenchymal transition and cancer stem cells in glioblastoma cell lines

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    A thesis submitted in partial fulfilment of the requirements of the University of Wolverhampton for the degree of Doctor of Philosophy.Glioblastoma multiforme (GBM) is one of the most aggressive and lethal cancers with a poor prognosis. Advances in the treatment of GBM are limited due to several resistance mechanisms and limited drug delivery into the central nervous system (CNS) compartment by the blood-brain barrier (BBB) and by actions of the normal brain to counteract tumour-targeting medications. Hypoxia is common in malignant brain tumours such as GBM and plays a significant role in tumour pathobiology. It is widely accepted that hypoxia is a major driver of GBM malignancy. Although it has been confirmed that hypoxia induces GBM stem-like-cells (GSCs), which are highly invasive and resistant to all chemotherapeutic agents, the detailed molecular pathways linking hypoxia, GSC traits and chemoresistance remain obscure. Evidence shows that hypoxia induces cancer stem cell phenotypes via epithelial-to-mesenchymal transition (EMT), promoting therapeutic resistance in most cancers, including GBM. This study demonstrated that spheroid cultured GBM cells consist of a large population of hypoxic cells with CSC and EMT characteristics. GSCs are chemo-resistant and displayed increased levels of HIFs and NFőļB activity. Similarly, the hypoxia cultured GBM cells manifested GSC traits, chemoresistance and invasiveness. These results suggest that hypoxia is responsible for GBM stemness, chemoresistance and invasiveness. GBM cells transfected with nuclear factor kappa B-p65 (NFőļB-p65) subunit exhibited CSC and EMT markers indicating the essential role of NFőļB in maintaining GSC phenotypes. The study also highlighted the significance of NFőļB in driving chemoresistance, invasiveness, and the potential role of NFőļB as the central regulator of hypoxia-induced stemness in GBM cells. GSC population has the ability of self-renewal, cancer initiation and development of secondary heterogeneous cancer. The very poor prognosis of GBM could largely be attributed to the existence of GSCs, which promote tumour propagation, maintenance, radio- and chemoresistance and local infiltration. In this study, we used Disulfiram (DS), a drug used for more than 65 years in alcoholism clinics, in combination with copper (Cu) to target the NFőļB pathway, reverse chemoresistance and block invasion in GSCs. The obtained results showed that DS/Cu is highly cytotoxic to GBM cells and completely eradicated the resistant CSC population at low dose levels in vitro. DS/Cu inhibited the migration and invasion of hypoxia-induced CSC and EMT like GBM cells at low nanomolar concentrations. DS is an FDA approved drug with low toxicity to normal tissues and can pass through the BBB. Further research may lead to the quick translation of DS into cancer clinics and provide new therapeutic options to improve treatment outcomes in GBM patients

    Metabolic and nutritional triggers associated with increased risk of liver complications in SARS-CoV-2

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    Obesity, diabetes, cardiovascular and respiratory diseases, cancer and smoking are risk factors for negative outcomes in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which can quickly induce severe respiratory failure in 5% of cases. Coronavirus disease-associated liver injury may occur during progression of SARS-CoV-2 in patients with or without pre-existing liver disease, and damage to the liver parenchyma can be caused by infection of hepatocytes. Cirrhosis patients may be particularly vulnerable to SARS-CoV-2 if suffering with cirrhosis-associated immune dysfunction. Furthermore, pharmacotherapies including macrolide or quinolone antibiotics and steroids can also induce liver damage. In this review we addressed nutritional status and nutritional interventions in severe SARS-CoV-2 liver patients. As guidelines for SARS-CoV-2 in intensive care (IC) specifically are not yet available, strategies for management of sepsis and SARS are suggested in SARS-CoV-2. Early enteral nutrition (EN) should be started soon after IC admission, preferably employing iso-osmolar polymeric formula with initial protein content at 0.8 g/kg per day progressively increasing up to 1.3 g/kg per day and enriched with fish oil at 0.1 g/kg per day to 0.2 g/kg per day. Monitoring is necessary to identify signs of intolerance, hemodynamic instability and metabolic disorders, and transition to parenteral nutrition should not be delayed when energy and protein targets cannot be met via EN. Nutrients including vitamins A, C, D, E, B6, B12, folic acid, zinc, selenium and ŌČ-3 fatty acids have in isolation or in combination shown beneficial effects upon immune function and inflammation modulation. Cautious and monitored supplementation up to upper limits may be beneficial in management strategies for SARS-CoV-2 liver patients

    RNA pull-down-confocal nanoscanning (RP-CONA), a novel method for studying RNA/protein interactions in cell extracts that detected potential drugs for Parkinson’s disease targeting RNA/HuR complexes

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    MicroRNAs (miRNAs, miRs) are a class of small non-coding RNAs that regulate gene expression through specific base-pair targeting. The functional mature miRNAs usually undergo a two-step cleavage from primary miRNAs (pri-miRs), then precursor miRNAs (pre-miRs). The biogenesis of miRNAs is tightly controlled by different RNA-binding proteins (RBPs). The dysregulation of miRNAs is closely related to a plethora of diseases. Targeting miRNA biogenesis is becoming a promising therapeutic strategy. HuR and MSI2 are both RBPs. MiR-7 is post-transcriptionally inhibited by the HuR/MSI2 complex, through a direct interaction between HuR and the conserved terminal loop (CTL) of pri-miR-7-1. Small molecules dissociating pri-miR-7/HuR interaction may induce miR-7 production. Importantly, the miR-7 levels are negatively correlated with Parkinson‚Äôs disease (PD). PD is a common, incurable neurodegenerative disease causing serious motor deficits. A hallmark of PD is the presence of Lewy bodies in the human brain, which are inclusion bodies mainly composed of an aberrantly aggregated protein named őĪ-synuclein (őĪ-syn). Decreasing őĪ-syn levels or preventing őĪ-syn aggregation are under investigation as PD treatments. Notably, őĪ-syn is negatively regulated by several miRNAs, including miR-7, miR-153, miR-133b and others. One hypothesis is that elevating these miRNA levels can inhibit őĪ-syn expression and ameliorate PD pathologies. In this project, we identified miR-7 as the most effective őĪ-syn inhibitor, among the miRNAs that are downregulated in PD, and with őĪ-syn targeting potentials. We also observed potential post-transcriptional inhibition on miR-153 biogenesis in neuroblastoma, which may help to uncover novel therapeutic targets towards PD. To identify miR-7 inducers that benefit PD treatment by repressing őĪ-syn expression, we developed a novel technique RNA Pull-down Confocal Nanoscaning (RP-CONA) to monitor the binding events between pri-miR-7 and HuR. By attaching FITC-pri-miR-7-1-CTL-biotin to streptavidin-coated agarose beads and incubating them in human cultured cell lysates containing overexpressed mCherry-HuR, the bound RNA and protein can be visualised as quantifiable fluorescent rings in corresponding channels in a confocal high-content image system. A pri-miR-7/HuR inhibitor can decrease the relative mCherry/FITC intensity ratio in RP-CONA. With this technique, we performed several small-scale screenings and identified that a bioflavonoid, quercetin can largely dissociate the pri-miR-7/HuR interaction. Further studies proved that quercetin was an effective miR-7 inducer as well as őĪ-syn inhibitor in HeLa cells. To understand the mechanism of quercetin mediated őĪ-syn inhibition, we tested the effects of quercetin treatment with miR-7-1 and HuR knockout HeLa cells. We found that HuR was essential in this pathway, while miR-7 hardly contributed to the őĪ-syn inhibition. HuR can directly bind an AU-rich element (ARE) at the 3‚Äô untranslated region (3‚Äô-UTR) of őĪ-syn mRNA and promote translation. We believe quercetin mainly disrupts the ARE/HuR interaction and disables the HuR-induced őĪ-syn expression. In conclusion, we developed and optimised RP-CONA, an on-bead, lysate-based technique detecting RNA/protein interactions, as well as identifying RNA/protein modulators. With RP-CONA, we found quercetin inducing miR-7 biogenesis, and inhibiting őĪ-syn expression. With these beneficial effects, quercetin has great potential to be applied in the clinic of PD treatment. Finally, RP-CONA can be used in many other RNA/protein interactions studies

    Proof of Concept of Therapeutic Gene Modulation of MBNL1/2 in Myotonic Dystrophy

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    La distrofia miot√≥nica tipo 1 es una enfermedad gen√©tica rara multisist√©mica que afecta a 1 de cada 3000-8000 personas. La causa molecular de la enfermedad proviene de repeticiones t√≥xicas ‚ÄúCTG‚ÄĚ en el gen DMPK (DM Protein Kinase). Tras la transcripci√≥n, estas repeticiones forman una estructura de horquilla que se une con alta afinidad a la familia de prote√≠nas MBNL (Muscleblind-like) que agota su funci√≥n de regulaci√≥n de la poliadenilaci√≥n y el splicing alternativo postranscripcional en numerosos transcritos. La p√©rdida de funci√≥n de MBNL provoca una cascada de efectos posteriores, que eventualmente conducen a s√≠ntomas cl√≠nicos que incluyen mioton√≠a, debilidad y atrofia muscular, cataratas, disfunci√≥n card√≠aca y trastorno cognitivo. La restauraci√≥n de la funci√≥n de la prote√≠na MBNL es clave para aliviar los s√≠ntomas debilitantes de esta enfermedad. Se han utilizado oligonucle√≥tidos antisentido (AON) para apuntar a las repeticiones de DMPK y liberar MBNL del secuestro, lo que da como resultado resultados terap√©uticos prometedores en modelos celulares y animales de la enfermedad. Otro factor que interviene en la p√©rdida de funci√≥n de las prote√≠nas MBNL son los miRNAs que regulan su traducci√≥n. Aqu√≠ se muestra el uso de AON dirigidos a la actividad de miR-23b y miR-218, que se ha demostrado previamente que regulan directamente MBNL1 y MBNL2. Estos antimiRs recibieron modificaciones FANA para aumentar su entrega en las c√©lulas y reducir la toxicidad. Tambi√©n se probaron los AON, denominados blockmiRs, que se unen de manera complementaria a los sitios de uni√≥n confirmados de miR-23b y miR-218 en los 3'-UTR de las transcripciones de MBNL1 y MBNL2. De esta manera, los miRNAs no pueden unirse y regular la traducci√≥n de MBNL, lo que aumenta la cantidad de prote√≠na MBNL producida en una c√©lula deficiente. Aqu√≠ se propone el uso de AON de nuevo dise√Īo dirigidos a la actividad de miR-23b y miR-218 para regular MBNL1 y MBNL2 a trav√©s de (1) exploraci√≥n del bloqueo de miRNA a trav√©s de FANA-antimiR AON in vitro, (2) exploraci√≥n del bloqueo del sitio de uni√≥n de miRNA a trav√©s de la estrategia blockmiR in vitro e in vivo con el uso de modificaciones qu√≠micas de LNA, y (3) mejora de la qu√≠mica de la estrategia blockmiR mediante el uso de tecnolog√≠a de p√©ptidos de penetraci√≥n celular in vitro e in vivo.Myotonic Dystrophy Type 1 is a multi-systemic rare genetic disease affecting 1 in 3000-8000 people. The molecular cause of the disease stems from toxic ‚ÄúCTG‚ÄĚ repetitions in the DMPK (DM Protein Kinase) gene. Upon transcription, these repetitions form a hairpin structure that binds with high affinity to the MBNL (Muscleblind-like) family of proteins depleting their function of post-transcriptional alternative splicing and polyadenylation regulation on numerous transcripts. MBNL loss-of-function causes a cascade of downstream effects, which eventually lead to clinical symptoms including myotonia, muscle weakness and atrophy, cataracts, cardiac dysfunction, and cognitive disorder. The restoration of MBNL protein function is key to relieving the debilitating symptoms of this disease. Antisense oligonucleotides (AONs) have been used to target the DMPK repeats and release MBNL from sequestration resulting in promising therapeutic results in cellular and animal models of the disease. Another factor playing a role in the loss-of-function of MBNL proteins are the miRNAs that regulate their translation. Here is shown the use of AONs targeting miR-23b and miR-218 activity, which have been previously shown to directly regulate MBNL1 and MBNL2. These antimiRs were given FANA modifications to increase their delivery in cells and lower toxicity. Also tested are AONs, termed blockmiRs, that complementary bind to the confirmed binding sites of miR-23b and miR-218 in the 3‚Äô-UTRs of MBNL1 and MBNL2 transcripts. In this way, the miRNAs are unable to bind and regulate the translation of MBNL thereby augmenting the amount of MBNL protein made in an otherwise deficient cell. Proposed here is the use of newly designed AONs targeting miR-23b and miR-218 activity in order to regulate MBNL1 and MBNL2 through (1) exploration of miRNA blocking through FANA-antimiR AONs in vitro, (2) exploration of miRNA binding site blocking through blockmiR strategy in vitro and in vivo with the use of LNA chemical modifications, and (3) improvement of the chemistry of the blockmiR strategy through the use of cell penetrating peptide technology in vitro and in vivo

    Innate immunity and metabolism in the bovine ovarian follicle

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    Postpartum uterine disease in dairy cows is associated with reduced fertility. One of the first and most prevalent bacteria associated with uterine disease is Escherichia coli. The bacterial endotoxin, lipopolysaccharide (LPS), accumulates in the ovarian follicular fluid of animals with uterine disease. The granulosa cells of the ovarian follicle respond to LPS by secreting pro-inflammatory cytokines, such as interleukin (IL)-1a, IL-1b and IL-8, and oocyte health is perturbed. Dairy cows also experience metabolic energy stress in the postpartum period, which is associated with an increased risk of developing uterine disease and ovarian dysfunction. This thesis explored the crosstalk between innate immunity and metabolic energy stress in bovine granulosa cells and cumulus-oocyte complex. Firstly, we found that glycolysis, AMP-activated protein kinase and the mechanistic target of rapamycin, regulate the innate immune responses to LPS in granulosa cells isolated from bovine ovarian follicles. Activation of AMP-activated protein kinase decreased the LPS-induced secretion of IL-1a, IL-1b, and IL8, and was associated with shortened duration of ERK1/2 and JNK phosphorylation. Next, we found that decreasing the availability of cholesterol or inhibiting cholesterol biosynthesis using short-interfering RNA impaired the LPS-induced secretion of IL-1a and IL-1b by granulosa cells. Furthermore, metabolic energy stress or inhibiting cholesterol biosynthesis in the bovine cumulus-oocyte complex modulated the innate immune responses to LPS, and perturbed meiotic progression during in vitro maturation. Finally, we explored an in vivo model of uterine disease in heifers, using RNAseq to investigate alterations to the transcriptome of the reproductive tract. We found that uterine disease altered the transcriptome of the endometrium, oviduct, granulosa cells and oocyte, several months after bacterial infusion; these changes were most evident in the granulosa cells and oocyte of the ovarian follicle. The findings from this thesis imply that there is crosstalk between innate immunity and metabolism in the bovine ovarian follicle

    MicroRNAs as Future Treatment Tools and Diagnostic Biomarkers in Alzheimer’s Disease

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    Alzheimer‚Äôs disease (AD) is a neurodegenerative disorder and is considered to be the most common form of dementia. This disorder is characterized by the formation of amyloid ő≤ (Aő≤) plaques, neurofibrillary tangles, and alterations in synaptic function, all of which cause memory loss and behavioral disturbances. Despite the high prevalence of AD, effective therapeutic and diagnostic tools remain unavailable. MicroRNAs (miRNAs, miRs) are regulatory non-coding RNAs that target mRNAs. MiRNAs are involved in the regulation of the expressions of APP and BACE1, Aő≤ clearance, and the formation of neuro-fibrillary tangles. Furthermore, there are evidences that show alteration in the expression of several miRs in AD. MicroRNA is emerging as a biomarker because they have high specificity and, efficiency, and can be detected in biological fluids such as cerebrospinal fluid, tear, urine, blood. Moreover, miRNAs may be acquired and measured easily by utilizing real-time PCR, next-generation sequencing, or microarray. These techniques are cost-effective in comparison with imaging techniques such as magnetic resonance imaging, positron emission tomography. These features make miRNAs viable therapeutic as well as diagnostic tools in the treatment of AD. This review covers the regulatory function of miRNAs in AD, as well as their prospective applications as diagnostic biomarkers
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