Het onderzoek beschreven in dit proefschrift is gericht op de identificatie en karakterisering van regulatoren en/of effectoren van TGF-β-signalering en TGF-β-geïnduceerde EMT in pancreas-, long-... Show moreHet onderzoek beschreven in dit proefschrift is gericht op de identificatie en karakterisering van regulatoren en/of effectoren van TGF-β-signalering en TGF-β-geïnduceerde EMT in pancreas-, long- en borstkankertypes. Deze nieuw geïdentificeerde componenten en mechanismen kunnen worden onderzocht voor de ontwikkeling van geneesmiddelen voor kankertherapie. Show less
In dit proefschrift begin ik met een algemene inleiding in Hoofdstuk 1 om kort de relevantie van EC-gedrag in vasculaire morfogenese en in angiogenese te presenteren. Bovendien bespreek ik hoe de... Show moreIn dit proefschrift begin ik met een algemene inleiding in Hoofdstuk 1 om kort de relevantie van EC-gedrag in vasculaire morfogenese en in angiogenese te presenteren. Bovendien bespreek ik hoe de EC-functie ingewikkeld wordt gereguleerd door positieve en negatieve factoren, en hoe hun functie kan worden gemanipuleerd voor therapeutische winst bij kanker en hart- en vaatziekten. In Hoofdstuk 2 bespreken we in detail de rol van de TGF-β-signaleringsroute in EndMT en bespreken we de bijdrage van dit proces aan de ontwikkeling van ziekten, evenals de mogelijke toepassingen ervan in weefselmanipulatie. In Hoofdstuk 3 onthullen we gedetailleerde werkprotocollen om TGF-β-geïnduceerde EndMT te onderzoeken en hoe de betrokkenheid van EndMT-effectoren te beoordelen met behulp van CRISPR/Cas9-genediting. In Hoofdstuk 4 hebben we de functie van EndMT transcriptiefactoren onderzocht en hun werkingsmechanisme opgehelderd. We ontdekten dat de EndMT-transcriptiefactoren (TF's) SNAIL en SLUG cruciaal zijn voor EndMT in endotheelcellen van muizen en dat de ID-eiwitten hun functie in EndMT compenseren. In Hoofdstuk 5 geven we een technisch overzicht van embryonale zebravis-xenotransplantaattesten om TGF-β-familiesignalering in de progressie van borstkanker bij de mens te onderzoeken, waaronder intravasatie/extravasatie van tumorcellen en tumorangiogenese. In Hoofdstuk 6 identificeren en onderzoeken we twee nieuwe BMP type I receptor macrocyclische kinaseremmers met therapeutisch potentieel om angiogenese in normale en tumorvatvorming bij zebravissen te normaliseren. In Hoofdstuk 7 vat ik alle studies in het proefschrift samen en geef ik enkele toekomstperspectieven met betrekking tot onze resultaten. Show less
Gerrits, T.; Zandbergen, M.; Wolterbeek, R.; Bruijn, J.A.; Baelde, H.J.; Scharpfenecker, M. 2020
Diabetic nephropathy (DN) is a complication of diabetes mellitus that can lead to proteinuria and a progressive decline in renal function. Endoglin, a co-receptor of TGF-beta, is known primarily... Show moreDiabetic nephropathy (DN) is a complication of diabetes mellitus that can lead to proteinuria and a progressive decline in renal function. Endoglin, a co-receptor of TGF-beta, is known primarily for regulating endothelial cell function; however, endoglin is also associated with hepatic, cardiac, and intestinal fibrosis. This study investigates whether endoglin contributes to the development of interstitial fibrosis in DN. Kidney autopsy material from 80 diabetic patients was stained for endoglin and Sirius Red and scored semi-quantitatively. Interstitial endoglin expression was increased in samples with DN and was correlated with Sirius Red staining (p < 0.001). Endoglin expression was also correlated with reduced eGFR (p = 0.001), increased creatinine (p < 0.01), increased systolic blood pressure (p < 0.05), hypertension (p < 0.05), and higher IFTA scores (p < 0.001). Biopsy samples from DN patients were also co-immunostained for endoglin together with CD31, CD68, vimentin, or alpha-SMA Endoglin co-localized with both the endothelial marker CD31 and the myofibroblast marker alpha-SMA. Finally, we used shRNA to knockdown endoglin expression in a human kidney fibroblast cell line. We found that TGF-beta 1 stimulation upregulated SERPINE1, CTGF, and ACTA2 mRNA and alpha-SMA protein, and that these effects were significantly reduced in fibroblasts after endoglin knockdown. Taken together, these data suggest that endoglin plays a role in the pathogenesis of interstitial fibrosis in DN. Show less
Several cellular processes and pathways were altered both by fluid shear stress and Pkd1 gene disruption in renal epithelial cells. Many of these signaling pathways are implicated in ADPKD as... Show moreSeveral cellular processes and pathways were altered both by fluid shear stress and Pkd1 gene disruption in renal epithelial cells. Many of these signaling pathways are implicated in ADPKD as well. However, more than 20 years after the discovery of PKD1 and PKD2 as genetic cause of ADPKD, the exact cellular function of the polycystins still remains unclear. Our data indicate that polycystin-1 is not a direct mechano-sensor, but it restrains shear stress induced gene expression via an unknown mechanism. Additional research is required to identify the cellular function of polycystins and the mechanism of mechanotransduction. This is needed to refine the mechanism of cyst formation in ADPKD and other ciliopathies, which could identify potential targets for therapy. Nevertheless, we showed that inhibition of activin signaling is a promising therapy to slow cyst progression in Pkd1del mice. Although other treatment strategies have been tested successfully to reduce PKD progression in pre-clinical studies, the efficacy in human patients is sometimes minimal or absent. Therefore, it has been suggested to target multiple signaling pathways affected in ADPKD. These combined therapies should reestablish the balance in cellular signaling of renal epithelial cells and maintain cellular homeostasis within physiological boundaries. Show less
In this thesis, we identify and/or analyse a couple of regulators of the TGF-β and Wnt signal transduction pathways that play important roles in breast cancer. Our aim was to understand how these... Show moreIn this thesis, we identify and/or analyse a couple of regulators of the TGF-β and Wnt signal transduction pathways that play important roles in breast cancer. Our aim was to understand how these regulators affect the different steps of these pathways, including receptor activation, post-translational modification of other transducing molecules, target gene transcription, as well as crosstalk with other cell signaling pathways. Insight in these mechanisms is expected to help to design strategies for therapeutic intervention in TGF-β and Wnt mediated breast cancer progression. Show less
