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Analysis of Apoptosis in Zebrafish Embryos by Whole-mount Immunofluorescence to Detect Activated Caspase 3
Published on: December 20, 2013
Zebrafish embryo proteins induce apoptosis in human colon cancer cells (Caco2)
Alessandra Cucina1, Pier-Mario Biava, Fabrizio D'Anselmi
1Department of Surgery, Pietro Valdoni, Università La Sapienza, Roma, Rome.
Abstract:
Previous studies have shown that proteins extracted from Zebrafish embryo share some cytostatic characteristics in cancer cells. Our study was conducted to ascertain the biological properties of this protein network. Cancer cell growth and apoptosis were studied in Caco2 cells treated with embryonic extracts. Cell proliferation was significantly inhibited in a dose-dependent manner. Cell-cycle analysis in treated cells revealed a marked accumulation in the G(2)/M phase preceding induction of apoptosis. Embryo proteins induced a significant reduction in FLIP levels, and increased caspase-3 and caspase-8 activity as well as the apoptotic rate. Increased phosphorylated pRb values were obtained in treated Caco2 cells: the modified balance in pRb phosphorylation was associated with an increase in E2F1 values and c-Myc over-expression. Our data support previous reports of an apoptotic enhancing effect displayed by embryo extracts, mainly through the pRb/E2F1 apoptotic pathway, which thus suggests that Zebrafish embryo proteins have complex anti-cancer properties.
Insights
Zebrafish embryo proteins inhibit cancer cell growth and induce apoptosis by affecting cell cycle progression and key regulatory proteins. These findings highlight Zebrafish embryo extracts as a potential source of novel anti-cancer compounds.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Previous research indicates Zebrafish embryo extracts possess cytostatic properties against cancer cells.
- Understanding the specific biological mechanisms of these extracts is crucial for therapeutic development.
Purpose of the Study:
- To investigate the biological activities of Zebrafish embryo protein networks in cancer cells.
- To elucidate the anti-cancer mechanisms, including effects on cell proliferation, cell cycle, and apoptosis.
Main Methods:
- Caco2 cancer cells were treated with Zebrafish embryonic extracts.
- Cell proliferation, cell cycle distribution, and apoptosis assays were performed.
- Western blotting and enzymatic activity assays were used to analyze protein levels and activity (FLIP, caspases, pRb, E2F1, c-Myc).
Main Results:
- Zebrafish embryonic extracts significantly inhibited Caco2 cell proliferation in a dose-dependent manner.
- Treatment led to G(2)/M cell cycle arrest and subsequent apoptosis induction.
- Extracts reduced FLIP levels, increased caspase-3 and caspase-8 activity, and elevated phosphorylated pRb, E2F1, and c-Myc levels.
Conclusions:
- Zebrafish embryo proteins exhibit significant anti-cancer properties by promoting apoptosis.
- The pRb/E2F1 pathway is implicated in the apoptotic effect of Zebrafish embryo extracts.
- These findings suggest Zebrafish embryo proteins have complex anti-cancer potential.
