Rolle von Exosomen-lokalisierter RNAs in der Interaktion zwischen Tumorzellen und Makrophagen im Pankreaskarzinom

Tumorassoziierte Makrophagen machen einen erheblichen Teil des tumorassoziierten Stromas von Pankreaskarzinomen aus. Verschiedene Studien haben bereits gezeigt, dass Makrophagen in vielen humanen Tumoren, auch in Pankreastumoren, einen überwiegend tumorfördernden M2-funktionalen Phänotyp besitzen, w...

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Bibliographic Details
Main Author: Landmann, Emelie Maximiliane
Contributors: Buchholz, Malte (Prof. Dr.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2019
Online Access:PDF Full Text
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Tumor associated macrophages represent a considerable part of the tumor-associated stroma in pancreatic cancer. Various studies have shown that many of the macrophages in human tumors, including PDAC, show a tumor promoting M2 functional phenotype, which is associated with poor prognosis. miRNAs and other non-coding RNAs play important regulatory roles in the development and progression of malignant tumors, including the functional interaction of tumor cells and cells of the surrounding inflammatory stroma. Currently, it is assumed that tumor cells use, among others, extracellular vesicles for cell-to-cell exchange of biomolecules. Via these vesicles proteins, lipids or nucleic acids, especially miRNAs, are exchanged and contribute to altering cellular processes of target cells and thereby have effects on the tumor microenvironment. After establishing a protocol for primary human macrophage and macrophage cell line polarisation, real-time PCR analysis of THP1/BlaER1 cells after functional co-culture experiments with pancreatic tumor cell lines showed that M1-Markers reduce rapidly during co-culture experiments, while the M2 polarization especially in co-culture with LON 556 tumor cells is retained to a higher degree. The treatment of THP1 macrophages with conditioned tumor cell medium of S2-007 and PaTu 8988t cells promotes the M2 polarization much stronger. To check if these findings were caused by tumor cell secreted extra cellular vesicles, a purification protocol for exosomes was established. Next, THP1/ BlaER1 macrophages were treated with exosomes which were screted by pancreatic tumor cells. Real-time PCR analysis of THP1/ BlaER1 cells showed here that in THP1 cells exosomes from cancer cells caused a markedly increased expression of the M2-Marker CCl13, while in BlaER1 cells the cytokines CCL13, CCL17 and CCL18 were increased. In both cell lines, tumor-derived exosomes exhibit no effects on the M1-Markers CXCL9, CXCL10 and CCL2. The sequencing of secreted exosomes from S2-007 cells and THP1 macrophages after co-culture enabled the identification of some promising candidate molecules, like IGFII mRNA and miRNAs 6724, 4497 and 5787. The exact function of these candidates concerning the differentiation of tumor associated macrophages in pancreatic tumors have to be examined in more detail using knock down methods for IGFII or miRNA inhibition. To summarize, this study showed that tumor cell derived exosomes promote the polarisation of M2 macrophages. The identified candidates in tumor derived exosomes, IGFII and miRNAs 6724, 4497 and 5787, could play an important role in differentiation of tumor associated macrophages and thereby promote tumor progression in exocrine pancreatic tumors.