Dataset Information

Cancer associated fibroblast-derived WNT2 increases tumor angiogenesis in colon cancer

ABSTRACT: WNT2 is important for placenta vascularization and acts as a pro-angiogenic factor for liver and other endothelial cells (ECs). WNT2 induction has been shown in many carcinomas and is associated with tumor progression. In colorectal cancer (CRC) WNT2 is selectively elevated in cancer associated fibroblasts (CAFs), leading to increased invasion and metastasis. However, if there is a role for WNT2 in colon cancer angiogenesis has not been addressed so far. Here, we demonstrate that WNT2 enhances EC migration and invasion, while it induces ß catenin dependent signaling in only a small subset of HUVECs. We show that siRNA-mediated knockdown of WNT2 in CAFs reduced the growth of vessel-like structures significantly in a co-culture assay, while the overexpression of WNT2 in skin fibroblasts otherwise being devoid of WNT2 led to increased angiogenesis in vitro. In a xenograft model, overexpression of WNT2 in HCT116 led to enhanced tumor volume and vessel density. Moreover, WNT2 expression correlates with vessel markers in human CRC. Secretome profiling of CAFs revealed that proteins related to angiogenesis and extracellular matrix (ECM) remodeling are regulated by WNT2. Thus, stroma-derived WNT2 positively affects angiogenesis in CRC by shifting the balance towards pro-angiogenic signals.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture, Stromal Cell, Fibroblast

SUBMITTER: Christopher Gerner

LAB HEAD: Christopher Gerner

PROVIDER: PXD014019 | Pride | 2020-05-26

REPOSITORIES: Pride

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Publications

Cancer-associated fibroblast-derived WNT2 increases tumor angiogenesis in colon cancer.

Unterleuthner Daniela D Neuhold Patrick P Schwarz Katharina K Janker Lukas L Neuditschko Benjamin B Nivarthi Harini H Crncec Ilija I Kramer Nina N Unger Christine C Hengstschläger Markus M Eferl Robert R Moriggl Richard R Sommergruber Wolfgang W Gerner Christopher C Dolznig Helmut H

Angiogenesis 20191030 2

WNT2 acts as a pro-angiogenic factor in placental vascularization and increases angiogenesis in liver sinusoidal endothelial cells (ECs) and other ECs. Increased WNT2 expression is detectable in many carcinomas and participates in tumor progression. In human colorectal cancer (CRC), WNT2 is selectively elevated in cancer-associated fibroblasts (CAFs), leading to increased invasion and metastasis. However, if there is a role for WNT2 in colon cancer, angiogenesis was not addressed so far. We demo ...[more]

PMID: 31667643

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Project description:Carcinoma-associated fibroblasts (CAFs) that express ?-smooth-muscle-actin (?SMA+) contribute to cancer progression, but their precise origin and role in tumorigenesis is not established. Using mouse models of inflammation-induced gastric cancer, we show that at least 20% of CAFs originate from bone marrow and derive from mesenchymal stem cells (MSCs). Surprisingly, we find that ?SMA+ myofibroblasts (MF) are niche cells normally present in bone marrow and increase markedly in the bone marrow and blood during progression to dysplasia. MSC-derived CAFs that are recruited to the dysplastic stomach express IL-6, Wnt5? and BMP4 and show DNA hypomethylation. Bone marrow (BM)-derived CAFs strongly promote tumor growth in organotypic and xenograft models. In addition, CAFs are generated from MSCs and are recruited to distant tumor sites in a TGF-?- and SDF-1?-dependent manner. Carcinogenesis therefore involves the expansion and relocation of normal bone marrow niche cells to the tumor site where they create a new niche to sustain cancer progression. Since resident (non-BM-derived) CAFs could not be cultured and directly compared to BM-derived CAFs, we additionally isolated total RFP(+) gastric CAFs from aSMA-RFP mice with Helicobacter felis-induced dysplasia, and compared them to GFP(+) BM-derived gastric CAFs from mice with H. felis-induced dysplasia mice that had been transplanted with UBC-EGFP bone marrow. The RFP+ CAFs (HF CAF) represent total CAFs (of which only 20% were BM-derived), while the latter represented only BM-derived CAFs (BM CAF). We compared their gene expression using the Illumina array (MouseWG-6v2) directly after FACS sorting. Interestingly, the GFP+ BM-derived CAFs expressed higher levels of inflammatory genes (IL-6, IL-1?, IL-33) and a number of tumor and stem cell associated factors (CCL5, SPP1, Notch3, MMP9, CD47, CXCR4, PARP10,) compared to the total (RFP+) population of gastric CAFs. Comparison of bone marrow-derived GFP-labeled gastric CAFs versus all gastric CAFs.

Project description:The cyclin D1 oncogene encodes the regulatory subunit of a holoenzyme that phosphorylates and inactivates the Rb protein and promotes progression through G1 to S phase of the cell cycle. Several prostate cancer cell lines and a subset of primary prostate cancer samples have increased cyclin D1 protein expression. However, the relationship between cyclin D1 expression and prostate tumor progression has yet to be clearly characterized. This study examined the effects of manipulating cyclin D1 expression in either human prostatic epithelial or stromal cells using a tissue recombination model. The data showed that overexpression of cyclin D1 in the initiated BPH-1 cell line increased cell proliferation rate, but did not elicit tumorigenicity in vivo. However, overexpression of cyclin D1 in Normal Prostate Fibroblasts (NPF) that were subsequently recombined with BPH-1 did induce malignant transformation of the epithelial cells. The present study also showed that recombination of BPH-1 + cyclin D1 overexpressing fibroblasts (NPF cyclin D1) resulted in permanent malignant transformation of epithelial cells (BPH-1 NPF-cyclin D1 cells) similar to that seen with Carcinoma Associated Fibroblasts (CAFs). Microarray analysis showed that the expression profiles between CAFs and NPF cyclin D1 cells were highly concordant including cyclin D1 upregulation. These data indicated that the tumor-promoting activity of cyclin D1 may be tissue-specific. Keywords: cyclin D1; stromal-epithelial interactions; prostate cancer; cDNA microarray NPF cyclin D1 cells were generated from NPFs, which were isolated from two different patient samples; CAFs were isolated from two different patient samples as well. RNA was isolated from NPFs, CAFs, and NPF cyclin D1 cells using total RNA isolation kit (Qiagen). Custom spotted cDNA microarrays were constructed as previously described (True, L., Coleman, I., Hawley, S., Huang, C. Y., Gifford, D., Coleman, R., Beer, T. M., Gelmann, E., Datta, M., Mostaghel, E., Knudsen, B., Lange, P., Vessella, R., Lin, D., Hood, L., and Nelson, P. S. A molecular correlate to the Gleason grading system for prostate adenocarcinoma. Proc Natl Acad Sci U S A, 103: 10991-10996, 2006) using a non-redundant set of 6,700 prostate-derived cDNA clones identified from the Prostate Expression Data Base (PEDB), a public sequence repository of expressed sequence tag data derived from human prostate cDNA libraries. Total RNA was amplified through one round of linear amplification using the MessageAmp aRNA kit (Ambion, Austin, TX). Sample quality and quantification was assessed by agarose gel electrophoresis and absorbance at A260. Cy3 and Cy5 labeled cDNA probes were made from 4 M-BM-5g of amplified RNA. Two NPF cyclin D1 and two CAF samples (labeled with Cy3) were hybridized head-to-head with an NPF control sample labeled with Cy5. Probes were hybridized competitively to microarrays under a coverslip for 16 h at 63M-BM-0C. Fluorescent array images were collected for both Cy3 and Cy5 by using a GenePix 4000B fluorescent scanner, and image intensity data were gridded and extracted using GenePix Pro 4.1 software. Differences in gene expression between NPF cyclin D1/NPF and CAF/NPF groups were determined using a two-sample t-test with Significance Analysis of Microarrays (SAM) software (http://www-stat.stanford.edu/_tibs/SAM/) with a False Discovery Rate (FDR) of M-bM-^IM-$ 10% considered significant (Tusher, PNAS, 2001). Similarities in gene expression between NPF cyclin D1/NPF and CAF/NPF groups were determined using a one-sample t-test in SAM with an FDR of M-bM-^IM-$ 0.1% considered significant. These results were reduced to unique genes by eliminating all but the highest scoring clones for each gene. A Pearson correlation coefficient was calculated in Excel to assess the strength of the linear relationship between NPF cyclin D1/NPF and CAF/NPF average log2 ratios.

Dataset Information

Cancer associated fibroblast-derived WNT2 increases tumor angiogenesis in colon cancer

Publications

Cancer-associated fibroblast-derived WNT2 increases tumor angiogenesis in colon cancer.

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