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Myeloid-derived suppressor cells (MDSCs) contribute to high mortality rates during sepsis, but how sepsis induces MDSCs is unclear. Previously we reported that microRNA (miR)-21 and miR-181b reprogram MDSCs in septic mice by increasing levels of DNA binding transcription factor, nuclear factor 1 (NFI-A). Here, we provide evidence that miR-21 and miR-181b stabilize NFI-A mRNA and increase NFI-A protein levels by recruiting RNA-binding proteins HuR and Ago1 to its 3′ untranslated region (3′UTR). We also find that the NFI-A GU-rich element (GRE)-binding protein CUGBP1 counters miR-21 and miR-181b dependent NFI-A mRNA stabilization and decreases protein production by replacing 3′UTR bound Ago1 with Ago2. We confirmed the miR-21 and miR-181b dependent reprogramming pathway in MDSCs transfected with a luciferase reporter construct containing an NFI-A 3′UTR fragment with point mutations in the miRNA binding sites. These results suggest that targeting NFI-A in MDSCs during sepsis may enhance resistance to uncontrolled infection.

Sepsis-induced immunosuppression increases the risk of chronic infection and reduces survival. Myeloid-derived suppressor cells (MDSCs) expand in the bone marrow and spleen during murine polymicrobial sepsis, contributing to immunosuppression. A better understanding of molecular controls of MDSC production is needed to identify treatment targets. We previously reported that miR-21 and miR-181b couple with transcription factor NFI-A to induce MDSCs during murine sepsis. Here, we expand upon these observations by showing that conditional deletion of the Nfia gene in the myeloid lineage precludes MDSC development. NFI-A-deficient Gr1+CD11b+ myeloid cells are not immunosuppressive and differentiate normally into macrophages and dendritic cells. In contrast, ectopically expressed NFI-A prevents differentiation of these immature Gr1+CD11b+ cells, while converting them into MDSCs. In addition, NFI-A-deficient Gr1+CD11b+ cells decreased, and cells transfected with NFI-A increase expression of miR-21 and miR181b. Our results support a myeloid cell loop in which NFI-A and miR-21 and miR-181b sustain Gr1+CD11b+ MDSC-dependent immunosuppression during sepsis.

Myeloid progenitor-derived suppressor cells (MDSCs) arise from myeloid progenitors and suppress both innate and adaptive immunity. MDSCs expand during the later phases of sepsis in mice, promote immunosuppression, and reduce survival. Here, we report that the myeloid differentiation-related transcription factor nuclear factor I-A (NFI-A) controls MDSC expansion during sepsis and impacts survival. Unlike MDSCs, myeloid cells with conditional deletion of the Nfia gene normally differentiated into effector cells during sepsis, cleared infecting bacteria, and did not express immunosuppressive mediators. In contrast, ectopic expression of NFI-A in myeloid progenitors from NFI-A myeloid cell-deficient mice impeded myeloid cell maturation and promoted immune repressor function. Importantly, surviving septic mice with conditionally deficient NFI-A myeloid cells were able to respond to challenge with bacterial endotoxin by mounting an acute inflammatory response. Together, these results support the concept of NFI-A as a master molecular transcriptome switch that controls myeloid cell differentiation and maturation and that malfunction of this switch during sepsis promotes MDSC expansion that adversely impacts sepsis outcome.

La famille des facteurs de transcription NFI comprend les isoformes NFI-A, -B, -C et -X, qui peuvent agir comme activateurs ou répresseurs de la transcription génique. Récemment, nous avons démontré le rôle répresseur de NFI dans la transcription du gène encodant la sous-unité α5 de l'intégrine α5βl. Dans cette étude, nous avons démontré que la suppression de certains isoformes du facteur de transcription NFI altère l'expression du gène α5 dans une lignée particulièrement agressive de mélanome uvéal humain (T97). Nous avons démontré que la diminution de l'expression de certains isoformes permet l'augmentation significative l'expression de l'intégrine α5β1 à la surface des cellules de la lignée T97. Nous avons également identifié quelques protéines qui intéragissent in vitro avec NFI et qui pourraient ainsi modifier ses propriétés modulatrices en regard du gène de α5β1.

La régulation de l'expression des gènes est à la base de tous processus cellulaires tels que la différenciation, la migration, la réponse aux dommages, la survie et l'apoptose. La compréhension globale des mécanismes cellulaires passe donc par l'étude de la transcription. Définir les rôles, fonctions ainsi que les voies de signalisation pour chacun des facteurs de transcription d'une cellule est maintenant un incontournable dans la poursuite de la compréhension du génome humain. Les travaux de cette thèse cadrent dans cette optique en concentrant nos efforts sur la caractérisation des facteurs de transcription ± nuclear factor I ¿ (NFI). Nous démontrons, entre autre, la première preuve de liaison directe d'un répresseur au promoteur du gène p21. Cette répression exercée par les NFI est essentielle à la bonne progression du cycle cellulaire dans les cellules en prolifération, tout en permettant l'expression basale de p21. Nous avons aussi caractérisé les mécanismes de régulation des facteurs de transcription Spl, Sp3 et NFI sur le promoteur de l'intégrine α6, en présence de laminine et de la fibronectine. Nos résultats suggèrent une répression de l'expression de l'intégrine a6 par la liaison de NFI et la diminution des facteurs Spl et Sp3 en présence de laminine, pour ainsi expliquer les mécanismes de migration et prolifération cellulaire dans un modèle de cicatrisation cornéenne. Finalement, nous examinons l'étendue de la liaison des facteurs NFI dans les kératinocytes humains de peau, par une analyse globale d'immunoprécipitation de chromatine couplée aux puces à ADN îlots CpG, dans le but d'établir une meilleure compréhension des voies de signalisation dans lesquelles les facteurs de transcription NFI sont impliqués. Ces analyses permettront de mieux comprendre les mécanismes de régulation de la transcription, particulièrement en ce qui a trait à la régulation par les facteurs de transcription NFI

This dissertation elucidates several independent molecular mechanisms that function in astrocytes and glial tumor cells, and suggest that developmental and inflammatory signals may contribute to the development of brain tumors. First, we analyzed the mechanism of TIMP-1 activation in astrocytes and glioblastoma cells. TIMP-1 expression is activated by IL-1, which is the major neuroinflammatory cytokine, via simultaneous activation of IKK/NF-kB and MEK3/6/p38/ATF-2 pathways in primary human astrocytes. In contrast to astrocytes, TIMP-1 is expressed at lower levels in glioblastomas, and is not regulated by IL-1 due to either dysfunctional IKK/NF-kB or MEK3/6/p38/ATF-2 activation. Thus, we propose a novel mechanism of TIMP-1 regulation, which ensures an increased supply of the inhibitor after tissue injury to limit the ECM degradation. This mechanism does not operate in gliomas, and may in part explain the increased invasiveness of glioma cells.Inflammation has been associated with the development of several cancers, including glioblastoma multiforme. However, it has not been linked to other brain tumors. Here we show for the first time that inflammation is associated with oligodendroglioma tumors as pro-inflammatory cytokines, such as OSM, IL-6, MCP1, MIP1α, and MIP1β and inflammatory markers, such as ACT and COX-2, were expressed at higher levels in oligodendroglioma samples. In addition, cytokine-induced STAT3 signaling, but not NF-kB, is highly activated in the oligodendroglioma patients. Moreover, OSM promotes oligodendroglioma cell proliferation in vitro, and this effect is mediated through STAT3. In summary, oligodendroglioma tumors secrete and respond to inflammatory mediators, with OSM being the major cytokine that activates STAT3 to promote the growth of tumor cells, and express ACT and COX-2 as a hallmark of ongoing inflammation. Since STAT3 promotes the growth of oligodendroglioma, as well as glioblastoma cells, and also regulates gliogenesis, we studied molecular mechanisms of this process in an in vitro differentiation model. We turn our attention to the NFI family of transcription factors since they have recently emerged as novel regulators of the development of vertebral neocortex. We developed a stem cell-neural progenitor-astrocyte differentiation model, in which the generated astrocytes were characterized by proper morphology, increased glutamate uptake, and expression of early and late astrocyte markers. Moreover, we found that NFI-X and NFI-C but not NFI-A or NFI-B, control the expression of GFAP and SPARCL1, the markers of terminal differentiation of astrocytes.In summary, the three mechanisms of gene regulation we studied, provided new insights into astrocyte biology, with the important implications for understanding the basis leading to the development and progression of brain tumors.

YKL-40 is a secreted protein that is highly up-regulated in malignant glioblastoma (GBM). Its expression is correlated with the invasive nature of GBMs and poor diagnosis of patients (Nigro et al., 2005). Previous research has shown that in astrocytes and GBM cells, YKL-40 expression is regulated by two transcription factors, NFI-X3 and STAT3, which form a complex with each other (Singh et al., 2011). Here, we show that the N-terminal domain of NFI-X3 is sufficient and required for its interaction with STAT3. We also show that the DNA-binding domain of NFI-X3 is required to induce YKL-40 expression. Thus, the interaction of NFI-X3 with STAT3 may play a role in stabilizing the otherwise weak binding of NFI-X3 to the YKL-40 promoter. Collectively, the observations made in this study shed light on the mechanisms by which NFI-X3, in concert with STAT3 regulate YKL-40 expression.

Humans have affected the Swedish forests for a long time, mainly through agriculture and forestry. Since the beginning of the 20th century, a lot of changes have taken place in forest policy, which have affected the structures of our forests, such as tree age, size and species distributions. By using information from different types of historical data, we can increase our understanding of the earlier structures of forests and how to manage them in the future. This study investigated changes in annual growth and changes in age, diameter, height and tree species distribution in the forests of Östergötland from year 1927 to the period 1983-2017, using data from the Swedish National Forest Inventory. Six tree species where used, Picea spp, Pinus spp, Quercus robur, Betula spp, Alnus spp and Populus tremula. There was an increase in the proportion of Picea spp, and a decrease in the proportion of Pinus spp since 1927. The proportion of deciduous trees also increased but not as much. Interestingly, the proportion of Betula spp had fallen since 1927, but the now larger proportion of young trees indicate that the proportion of Betula spp is increasing again. Q. robur, P. tremula, Picea spp and Pinus spp also had a larger proportion of younger trees in the more recent period. There was a significantly higher proportion of tall trees during the latter period for all tree species. For growth rate in width, the results varied among tree species. For Betula spp it had decreased and for Q. robur it had increased. For the coniferous trees there was hardly any difference between the periods. Overall, the results show that Östergötland's forests have been influenced by forestry and increased ungulate populations since the first inventory 1927 and that there have been changes in the composition of the forests. / Människan har länge påverkat de svenska skogarna. Sedan 1900-talets början har en hel del förändringar skett inom skogspolitiken, vilket har påverkat bland annat skogarnas ålder- och storleksfördelning, samt trädslagsfördelning. Genom att använda information från olika typer av historiska data kan vi öka vår förståelse om hur skogar såg ut förr och vad som har påverkat dem. Denna studie undersökte skillnader i tillväxt, höjd-och diameterfördelning, samt åldersfördelning i Östergötlands skogar från år 1927 till perioden 1983–2017, med hjälp av data från riksskogstaxeringen. Sex trädslag användes; gran (Picea spp), tall (Pinus spp), ek (Quercus robur), björk (Betula spp), al (Alnus spp) och asp (Populus tremula). En kraftig ökning av andelen gran hade skett sedan 1927, medan andelen tall hade minskat. För lövträden hade det endast skett en liten ökning i andel. Intressant var också att andelen björk hade minskat sedan 1927 och att andelen björkar yngre än 50 år var större den senare perioden, vilket indikerar att björkens andel kan vara på väg att öka igen. Ek, asp, gran och tall hade också en större andel yngre träd under den senare perioden. För alla trädslag fanns det en betydligt mindre andel höga träd år 1927. För årlig tillväxt i bredd varierade resultaten trädslagen emellan. För björk hade årsringsbredden minskat sedan 1927, medan den hade ökat för ek. För barrträden var det knappt någon skillnad mellan perioderna. Resultaten visar att Östergötlands skogar har påverkats mycket av skogsbruket, samt klövviltsbete, sedan 1927 och att det har skett flera strukturella förändringar i skogarnas sammansättning.

Old and coarse trees implement vital functions for humans, animals, and nature. Swedish forests have for a long time been affected by man, which affects the trees’ age and size distribution. The proportion of old-trees is currently low and it is desirable to increase it. Using data on sample trees’ from the Swedish National Forest Inventory, I analysed old-tree distribution in different land-use types and age-distribution, as well as coarse trees in Östergötland under the years 1983-2017. Furthermore, I analysed the sample trees form the year 1927 to compare with the sample trees in period 1983-2017. The defined age of an old-tree 5% older trees in the data was used while the coarse trees were defined by “miljömålets” definition. The tree sample data involved Pinus sylvestris, Picea abies, Quercus robur, Populus tremula, and Alnus glutinosa. Their occurrence in four different land-use types were considered: (i) productive woodland, (ii) arable land and natural pasture, (iii) mountains and other wastelands, and (iv) peatland. The results shows that a higher percentage of old and coarse trees occur in (i) productive woodland during the years 1983-2017. The coarse trees had similar distribution in the different land-use types during the years 1983-2017 and over time. Comparison with 1927 showed that the number of old-trees has decreased while the coarse trees had increased. Thus, the land-use types, except (i), do not constitute a reservoir for old and coarse trees and that the coarse trees individuals overlap with the old trees. / Gamla träd och grova träd utför livsviktiga funktioner för både människa, djur och natur. Sveriges skogar har länge påverkats av människan och det har påverkat trädens ålders- och storleksfördelning. Idag är andelen gamla träd låg i Sverige och det manifesteras åtgärder för att öka andelen äldre och grövre träd i skogarna. Genom att använda riksskogstaxeringens provträdsdata kan vi veta hur gamla och grova träd är fördelade bland marktyper. Denna studie undersökte hur gamla provträd var fördelade bland marktyper och deras åldersfördelning, samt motsvarande för grova träd i Östergötlands län. Dessutom jämfördes perioderna 1983–2017 och 1927. Gamla träd utgjorde per definition 5% av de äldre träden, medan grova träd definierades utifrån miljömålet levande skogar. Provträdsdata var på trädslagen; Pinus sylvestris, Picea abies, Quercus robur, Populus tremula och Alnus glutinosa som fanns inom marktyperna; (i) produktiv skogsmark, (ii) åkermark och naturbete, (iii) berg och vissa andra impediment, och (iv) myr. Resultaten visar att en hög andel gamla liksom grova provträd fanns inom produktiv skogsmark i Östergötlands län under 1983–2017. Bland marktyperna hade de grova provträd liknande odds och en liknande grovleksfördelning för båda perioderna. Dock sedan 1927 har andelen gamla träd minskat, medan andelen grova provträd har ökat. Därmed utgör marktyperna utöver (i) inte en reservoar för gamla som grova träd samt att de grova träden kan spegla de gamla trädens förekomst.