Zytokinabhängige Expression von EGF und VEGF und ihrer Rezeptoren EGFR und VEGFR-1 im Tumormikromilieu des kolorektalen Karzinoms

Cytokine-dependent gene expression of EGF, VEGF and their receptors EGFR and VEGFR-1 in the microenvironment of colorectal carcinoma

by Florentine Sattler
Doctoral thesis
Date of Examination:2014-07-08
Date of issue:2014-06-23
Advisor:Prof. Dr. Sabine Mihm
Referee:PD Dr. Jochen Gaedcke
Referee:Prof. Dr. Martin Oppermann
Persistent Address: http://dx.doi.org/10.53846/goediss-4568

 

 

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Abstract

English

Several diseases associated with chronic inflammation - for example gastro-esophageal reflux, Helicobacter pylori infection and chronic (gastro)enteritis in the context of inflammatory bowel disease (IBD) - promote tumor formation and growth. Secretion of cytokines such as TNF-α, IL-1β and IFN-γ, and of growth factors such as epidermal growth factor (EGF) and vascular endothelial growth factor (VEGF) by tumor cells or surrounding inflammatory cells causes development of a tumor growth-promoting microenvironment. This work was to investigate gene expression of the growth factors EGF and VEGF, as well as their receptors EGFR and VEGFR-1 in immune cells and cells of colorectal carcinoma in the setting of chronic inflammation. Furthermore, this work aims to investigate whether application of specific antibodies against TNF-α, EGF and VEGF can be of therapeutic use in colorectal carcinoma. Immune cells were isolated from the blood of male, healthy subjects and were incubated with phytohemagglutinin (PHA) or lipopolysaccharide (LPS), anti-VEGF antibody, anti-EGF antibody and anti-TNF-α antibody. Colon cancer cells (DLD-1 and HT-29) were incubated with the cytokines TNF-α, IL-1β or IFN-γ, and RNA was isolated and transcribed into cDNA. Gene expression of EGF, VEGF and their receptors EGFR and VEGFR-1 was measured by real-time PCR. PBMCs express EGF at low level; expression of EGFR was not detected . PBMCs do express VEGF, and its expression increased after stimulation of the cells with LPS. Treatment of cells with infliximab resulted in inhibition of both LPS-induced and basal VEGF expression. Incubation of PBMCS with TNF-α or PHA had no significant effect on the expression of VEGF. VEGFR-1 was expressed by PBMCs and gene expression increased after incubation with LPS. LPS-induced increase in gene expression was diminished by addition of infliximab. Addition of anti-VEGF antibody caused further increase of VEGFR-1 gene expression. Incubation of cells with PHA and TNF-α resulted in an increase of VEGFR-1 gene expression, addition of infliximab inhibited this effect. Colon carcinoma cells DLD-1 and HT-29 express EGF and EGFR as well as VEGF. DLD-1 cells also express VEGFR-1 at low level. Gene expression of EGF/R and VEGF/R-1 in both cell lines was not significantly altered by treatment with TNF-α, IL-1β or IFN-γ - each applied individually - during the observation period. Combination of IL-1-β and IFN-γ had a strong stimulatory effect on VEGF gene expression in both cell lines, and on gene expression of EGFR in DLD-1 cells. Combination of TNF-α with IFN-γ caused an increase of VEGF gene expression in both cell lines, and of VEGFR-1 gene expression in DLD-1 cells . Anti-inflammatory therapy with infliximab thus counteracts expression of VEGF in certain leukocyte subpopulations and may inhibit VEGF mediated tumor growth. In the context of cancer therapy, inhibition of VEGF appears to be effective rather via inhibition of tumor angiogenesis, whereas anti-inflammatory therapy by inhibition of VEGFR-1 might be a new target for antibody therapy.
Keywords: EGF; EGFR; VEGF; VEGFR; Colorectal carcinoma
Schlagwörter: EGF; EGFR; VEGF; VEGFR; kolorektales Karzinom
 
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