Impact of the transcription factor Nrf2 on the metabolic state of colorectal cells during inflammation associated carcinogenesis. The transcription factor Nrf2 plays a key role in oxidative stress response and mediates profound cytoprotection. Contrary to the previous view, it was recently found that deregulated Nrf2 activity contributes to carcinogenesis. Deregulated Nrf2 activity can result from persistent oxidative stress, which is often seen in the permanent presence of e.g. macrophages during the course of a chronic inflammation such as inflammatory bowel disease (IBD) or in the context of colorectal cancer (CRC). Under such conditions, epithelial cells adapt to oxidative stress through the enhanced and durable activation of Nrf2. One consequence is an increased resistance to apoptosis, manifesting in chemoresistance, but several other growth advantages are conferred by an enhanced Nrf2 activation, too. Recent data indicates that Nrf2 is involved in metabolic reprogramming, as well. In fact, the altered condition not only covers the high energy demand, but also ensures masses of glucose as intermediate metabolite for biomass production along with cell proliferation and tumor growth. Recently, it was shown that oxidative stress modulates the expression of monocarboxylate transporters (MCT) - these are involved in the cellular balance of lactate -, thus pointing to a possible contribution of Nrf2 in cancer metabolism. This would particularly apply to inflammation associated cancers (IAC) initiated and promoted by persistent oxidative stress and Nrf2 dependent adaptation processes.
In order to explore the impact of Nrf2 on the metabolic state of colorectal cancer cells and non-malignant colonic enterocytes, the work plan will address the following questions:
Does Nrf2 affect metabolic gene expression?
How does this influence the cellular phenotype?
Does the Nrf2 controlled metabolic reprogramming relate to exposition to inflammatory cells like M1-macrophages?
Is the Nrf2 dependent metabolic signature present in human disease like IBD and CRC?
To gain insight in Nrf2 dependent cell responses, enterocytes will be analyzed after achieving Nrf2 activation by treatment with anti-oxidants and established Nrf2 inducers.
To address the impact of Nrf2 on metabolic reprogramming, expression analysis of Nrf2 dependent miRs will be conducted by qPCR with samples from Nrf2 induced/inhibited non-malignant and malignant colonic epithelial cells.
To explore the impact of the inflammatory microenvironment, premalignant and malignant colonic enterocytes will be studied in indirect co-culture with M1- and M2-polarized macrophages. After knocking down Nrf2 in the enterocytes, both cell compartments will be analyzed for Nrf2-dependent cell responses, performing western blot and qPCR.
To further extend these findings towards a signature relating to metabolic reprogramming and enhanced Nrf2 activation, formalin fixed and paraffin embedded colon tissue samples from patients with IBD or colon cancer will be analyzed by immunohistochemistry.