Luparello C et al. (2022), Cytotoxic capability and the associated proteom...

ECB-ART-50275

EXCLI J 2022 Apr 25;21:722-743. doi: 10.17179/excli2022-4825.

Show Gene links Show Anatomy links

Cytotoxic capability and the associated proteomic profile of cell-free coelomic fluid extracts from the edible sea cucumber Holothuria tubulosa on HepG2 liver cancer cells.

Luparello C , Branni R , Abruscato G , Lazzara V , Drahos L , Arizza V , Mauro M , Di Stefano V , Vazzana M .

???displayArticle.abstract???
Hepatocellular carcinoma (HCC) is an aggressive cancer histotype and one of the most common types of cancer worldwide. The identification of compounds that might intervene to restrain neoplastic cell growth appears imperative due to its elevated overall mortality. The marine environment represents a reservoir rich in bioactive compounds in terms of primary and secondary metabolites produced by aquatic animals, mainly invertebrates. In the present study, we determined whether the water-soluble cell-free extract of the coelomic fluid (CFE) of the edible sea cucumber Holothuria tubulosa could play an anti-HCC role in vitro by analyzing the viability and locomotory behavior, cell cycle distribution, apoptosis and autophagy modulation, mitochondrial function and cell redox state of HepG2 HCC cells. We showed that CFE causes an early block in the cell cycle at the G2/M phase, which is coupled to oxidative stress promotion, autophagosome depletion and mitochondrial dysfunction ultimately leading to apoptotic death. We also performed a proteomic analysis of CFE identifying a number of proteins that are seemingly responsible for anti-cancer effects. In conclusion, H. tubulosa's CFE merits further investigation to develop novel promising anti-HCC prevention and/or treatment agents and also beneficial supplements for formulation of functional foods and food packaging material.

???displayArticle.pubmedLink??? 35721581
???displayArticle.pmcLink??? PMC9203982
???displayArticle.link??? EXCLI J

???attribute.lit??? ???displayArticles.show???

References [+] :

Auguste, Conservation of Cell Communication Systems in Invertebrate Host-Defence Mechanisms: Possible Role in Immunity and Disease. 2020, Pubmed

	Table 1. Cytotoxic activity-associated protein profile of the CFE from H. tubulosa
	Figure 1. A Holothuria tubulosa sea cucumber specimen
	Figure 2. Dose-response effect of CFE from H. tubulosa at 2.5, 5, 10 and 15 µg/ml concentration on the viability of HepG2 cells after either 24 (brown circles) or 48 h (green circles) of exposure. Error bars correspond to the standard error of the mean (s.e.m.) of three independent measurements. P values comparing viability ratios to controls were < 0.05 for every measurement.
	Figure 3. DNA profiles of control (blue line) and CFE-treated (red line) HepG2 cells after 24 and 48 h of exposure to the IC5024 and IC5048 of the preparation, respectively. Cell cycle distribution is reported in the annexed tables for both cell samples (mean ± s.e.m. of three independent experiments).
	Figure 4. Flow cytometric assays for apoptosis in HepG2 cells cultured in control conditions or exposed to CFE IC5024 for 24 h. The plots show the results of representative experiments and the percentages, indicated as the mean ± s.e.m. of three independent experiments, refer to viable annexin-V-/PI- cells (bottom left quadrant), early apoptotic annexin-V+/PI- cells (bottom right quadrant), late apoptotic annexin-V+/PI+ cells (top right quadrant) and necrotic annexin-V-/PI+ cells (top left quadrant).
	Figure 5. Flow cytometric assays for MMP in HepG2 cells cultured for 24 or 48 h in control conditions, in the presence of 1 µM valinomycin, and of either CFE IC5024 or CFE IC5048. The plots show the results of representative experiments and the percentages in the bottom quadrants of each frame, indicated as the mean ± s.e.m. of three independent experiments, are referred to dim red-emitting cells that underwent MMP dissipation.
	Figure 6. Bar graphs showing the ROS-associated MFI in ROS+ cell subpopulations (A) and the ROS+ cell/ROS- cell ratio (B) in parallel preparations of control and IC5024 and IC5048 CFE-treated HepG2 cells for 24 and 48 h (mean ± s.e.m. of three independent experiments). * p < 0.001 (A) and = 0.029 (B) determined with independent t-tests.
	Figure 7. Flow cytometric assays for AVOs in HepG2 cells cultured in control conditions for 24 (A) and 48 h (C) or exposed to CFE IC5024 for 24 h (B) and CFE IC5048 for 48 h (D). The plots show the results of representative experiments and the percentages, indicated as the mean ± s.e.m. of three independent experiments, refer to AVO+ cells (right quadrants) and AVO- cells (left quadrants).
	Figure 8. Representative phase-contrast micrographs acquired during wound-healing experiments at different time intervals under control conditions, and in the presence of CFE at IC5024 and 5 µg/mL concentration. The assay was performed in triplicate. Microscopic magnification = 20X