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A role for WISP2 in colorectal cancer cell invasion and motility
Kathryn A Frewer1, Andrew J Sanders, Sioned Owen
1Metastasis and Angiogenesis Research Group, Institute of Cancer and Genetics, Cardiff University School of Medicine, Heath Park, Cardiff CF14 4XN, UK.
Background:
WNT inducible secreted protein 2 (WISP2) has been linked with a variety of human cancer types and may contribute to cancer metastasis. The current study investigated the importance of WISP2 in colorectal cancer cells, examining the impact of targeting WISP2 on Caco-2 cell invasion and motility together with potential mechanisms of action.
Materials And Methods:
WISP2 expression was targeted in Caco-2 cells using a ribozyme transgene system and successful knockdown was verified using reverse transcription-polymerase chain reaction (RT-PCR). The impact of WISP2 knockout (Caco-2(WISP2 KO)) on cell growth, adhesion, motility and invasion was examined using a number of in vitro functional assays. In vitro invasion assays were repeated in the presence of wingless-type MMTV integration site family (WNT) inhibitors (FH535 and IWP-2) to investigate the role of the WNT-signalling pathway in the regulation of cell invasion by WISP2. Quantitative-PCR was conducted to measure matrix metalloproteinase (MMP) expression in control [wild-type (Caco-2(WT)) and cells containing the empty pEF6 plasmid (Caco-2(pEF6))] and Caco-2(WISP2 KO) cells.
Results:
WISP2 knockout resulted in a significant increase in Caco-2 cell invasion and motility (p<0.05 in comparison to wild-type and plasmid control Caco-2 cells). WISP2 knockout had no significant effect on Caco-2 cell growth rate in 3- and 5-day incubation and no significant impact on Caco-2 cell-matrix adhesion rates (p>0.05). Expression analysis of a number of MMPs indicated an insignificant up-regulation of MMP2, MMP9 (p>0.05) but significant up-regulation of MMP7 (p=0.025) in Caco-2(WISP2 KO) cells compared to controls. Inhibition of WNT signalling using FH535 and IWP-2 brought about a significant or borderline significant decrease in Caco-2(WISP2 KO) cell invasion (FH535 p=0.065) and (IWP-2 p=0.002) and negated the pro-invasive effect of targeting WISP2 in Caco-2 cells.
Conclusion:
WISP2 knockout significantly increased Caco-2 cell invasion and motility. Up-regulation of MMP2, -7 and -9 may indicate that WISP2 regulates invasion and motility through MMPs. Regulation of invasion by WISP2 may involve the WNT signalling pathway.
Insights
Targeting WNT inducible secreted protein 2 (WISP2) in colorectal cancer cells significantly increased invasion and motility. WISP2 may regulate cancer cell invasion through matrix metalloproteinases and the WNT signaling pathway.
Area of Science:
- Oncology
- Molecular Biology
- Cell Biology
Background:
- WNT inducible secreted protein 2 (WISP2) is implicated in various cancers and metastasis.
- The role of WISP2 in colorectal cancer (CRC) progression requires further investigation.
Purpose of the Study:
- To investigate the functional role of WISP2 in Caco-2 colorectal cancer cells.
- To determine the impact of WISP2 targeting on cell invasion and motility.
- To explore the underlying mechanisms involving WNT signaling and matrix metalloproteinases (MMPs).
Main Methods:
- WISP2 expression was knocked down in Caco-2 cells using a ribozyme system.
- In vitro assays assessed cell growth, adhesion, motility, and invasion.
- WNT signaling inhibitors (FH535, IWP-2) were used to probe pathway involvement.
- Quantitative PCR measured MMP expression (MMP2, MMP7, MMP9).
Main Results:
- WISP2 knockout significantly increased Caco-2 cell invasion and motility (p<0.05).
- Cell growth and adhesion rates were not significantly affected by WISP2 knockout.
- MMP7 expression was significantly upregulated (p=0.025) in WISP2 knockout cells.
- WNT pathway inhibition reduced invasion in WISP2 knockout cells.
Conclusions:
- WISP2 knockout enhances Caco-2 cell invasion and motility.
- WISP2 may regulate cancer cell invasion via MMPs, particularly MMP7.
- The WNT signaling pathway is involved in WISP2-mediated regulation of invasion.
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