Vascular diseases such as hypertension are marked by changes in calcium (Ca2+) and extracellular signal regulated kinase (ERK) signaling in the arterial wall. The overall goal of this project was to better understand pathways leading to altered gene regulation in cerebral arteries. Two models were tested to determine if, 1) Ca2+/cAMP response element binding protein (CREB) is regulated in intact cerebral arteries by multiple sources of Ca2+, and 2) hypertensive disease causes changes in genes regulated by ERK and CREB. Ca2+-mediated phosphorylation of CREB (P-CREB) was measured by immunofluorescence in both cultured vascular smooth muscle cells (VSMCs) and in intact cerebral arteries. The level of P-CREB was increased by both Ca2+ influx through voltage-dependent calcium channels (VDCCs) and store-operated Ca2+ entry (SOCE) in VSMCs. A similar increase in P-CREB was observed following stimulation of VDCCs and SOCE in intact cerebral arteries. However, unlike the results obtained from VSMCs phosphorylation of CREB following Ca2+ store depletion using thapsigargin, was partially dependent on Ca2+ entry through VDCCs, suggesting that communication between Ca2+ entry pathways in intact arteries may be lost during cell culture. The second model was tested using immunocytochemistry and RNA analysis to measure differences in cerebral artery signal transduction and gene expression caused by chronic hypertension in the Dahl salt sensitive genetic hypertensive rat model. Arteries from hypertensive animals exhibited increased phosphorylation of ERK and expression of Ki-67, a marker of proliferation, when compared to controls. In addition, microarray analysis of arterial RNA revealed overexpression of the matricellular ERK-regulated genes osteopontin (OPN), and plasminogen activator inhibitor 1 (PAI-1), and the activator protein transcription factor (AP-1) member junB in cerebral arteries, with validation using RT qPCR. To elucidate a role for CREB, ERK and JunB in the transcriptional regulation of OPN and PAI-1, VSMCs were treated with angiotensin II (Ang II), a vasoconstrictor linked to hypertension, and confirmed activator of OPN and PAI-1 transcription. Ang II induced an ERK-dependent transient increase in junB mRNA and protein prior to OPN, and PAI-1 induction. Gene silencing experiments indicated that OPN and PAI-1 are reciprocally regulated by junB and CREB, respectively, and that CREB is a negative regulator of OPN. Data from cell culture confirms that the Ang II response in VSMCs is transient, in contrast to the hypertensive in vivo model, suggesting that the CREB and ERK response induces long term changes. Together, these data have revealed mechanisms for regulation of gene expression that are linked to proliferation and remodeling in the arterial wall. Future experiments will explore an in vivo role for Ang II and SOCE in the mediation of ERK- and CREBregulated gene expression. This research has the potential to help in defining therapeutic strategies to prevent arterial remodeling caused by arterial pathologies such as hypertension.

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Die Einschleppung gebietsfremder Parasiten durch den Menschen ist einer der Hauptgründe für das Auftreten neuer Krankheiten in the letzten Jahrzehnten. Neue Wirte sind oftmals anfälliger für diese invasiven Parasiten als die ursprünglichen Wirte. In schwerwiegenden Fällen können invasive Parasiten zu Massensterben und zum Aussterben ihrer neuen Wirte führen. Der ursprüngliche Wirt des Aalparasiten Anguillicola crassus ist der Japanische Aal. In den frühen 1980er Jahren wurde der Parasit in die Population des Europäischen Aals eingeschleppt. Er ist einer der Faktoren, die für den Populationsrückgang des Europäischen Aals verantwortlich sind. Die molekularen Prozesse, die zur stärkeren Anfälligkeit des Europäischen Aals im Vergleich zum Japanischen Aal führen, sind noch nicht zureichend bekannt. Die Analyse transkriptomweiter differenzieller Genexpression von Immungewebe ergab, dass im Europäischen Aal sowohl Immun- als auch Nichtimmungene differenziell exprimiert waren. Dies war im Japanischen Aal nicht der Fall und deutet darauf hin, dass der Europäische Aal eine ineffiziente und kostspielige Immunantwort auslöst. Die Resultate ensprechen jenen die schon in anderen Wirbeltierwirt-invasiven Parasiten-Systemen beobachtet wurden. Ausserdem stützen diese Resultate die Hypothese, dass neuen Wirten eine wirksame Immunantwort fehlt und sie deuten darauf hin, dass Nichtimmunprozesse wesentlich zur höheren Anfälligkeit von neuen Wirten beitragen. Als Reaktion of die negativen Fitnesseffekte können neue Wirte Abwehrmechanismen entwickeln. Im Europäischen Aal entspricht das der Einkapselung von A. crassus. Einkapselung führte zu eine niedrigere Abundanz adulter A. crassus. Dies deutet darauf hin, dass das Einkapseln sich positiv auf die Gesundheit infizierter Aale auswirkt. Jedoch war die Abundanz zweier nativer Parasiten höher in Aalen, die A. crassus einkapselten. Eine verbesserte Abwehr des eingeschleppten Parasiten könnte daher die Abwehr nativer Parasiten beeinträchtigen. / The human-mediated translocation of non-native parasites into foreign regions is one of the primary factors for the emergence of new diseases in recent decades. Novel hosts are often more susceptible to these invasive parasites than the native host. In severe cases, invasive parasites can lead to population declines and extinctions of their novel hosts. The eel parasite Anguillicola crassus is native to the Japanese eel. In the early 1980s it was introduced into the European eel population and is now considered to be one factor contributing to the population decline of its novel host. The underlying molecular processes determining higher susceptibility in the European eel compared to the Japanese eel are not well understood. Using whole-transcriptome differential gene expression analysis of immune organs, I found that genes involved in both immune and non-immune processes were differentially expressed in the European eel but not the Japanese eel, suggestive of an ineffective and costly immune response in the former. These results are in line with those observed between susceptible and resistant hosts in other vertebrate host-invasive parasite systems. Furthermore, the results support the hypothesis that novel hosts lack an effective immune response. The results also suggest that alteration of non-immune processes contributes substantially higher susceptibilities of novel hosts. In response to the negative fitness effects of invasive parasites, novel hosts can evolve coping mechanisms. The European eel has the capacity to encapsulate and kill A. crassus. Using natural infections, I found a lower abundance of adult A. crassus, the most costly parasitic stage in those eels encapsulating the parasite, suggesting that encapsulation can potentially improve health of infected eels. At the same time, the abundance of two native parasites was higher in those eels encapsulating A. crassus. Thus, coping with A. crassus may come at the expense of coping with native parasites.