Effect of Native and Degraded CGN on THP-1 Proliferation and Cell Cycle
Preliminary observations by enumeration of THP-1 cells exposed to different concentrations of native and dCGN (10 and 40 kDa) during 2, 5 and 7 days, showed a decline in cell number (data not shown). This suggested that dCGN might cause an alteration in the cell cycle. Cell cycle analysis using flow cytometry showed an accumulation of THP-1 cells in G0/G1 phase, which was associated with a decrease number of cells in the S phase (Fig. 3A). The percentage of cells in the G0/G1 phase was 45.2% for control cells, 62.6% for C10 dCGN (at 2 mg/ml), and 64.2% for C40 dCGN-treated cells (at 2 mg/ml) (Fig. 3B). The effect of dCGN on cell cycle was dose-dependent (Fig. 3B). Neither native nor dCGN had an effect on the number of cells in the G2/M phase (Fig. 3). This effect is not due to cytotoxicity of dCGN even at the highest concentration (i.e. 2 mg/ml) since cell viability was not affected (data not shown).
10.1371/journal.pone.0008666.g003 Figure 3  Degraded CGN induced THP1 cell cycle arrest in G1 phase.
THP-1 cells in exponential growth phase were incubated in the presence or absence of carrageenan for 24 h before being stained with propidium iodide. Cell DNA content was then analyzed by flow cytometry. A: Histograms of cells treated with medium only (control), 10 kDa dCGN (C10), or 40 kDa dCGN (C40). B: Percentage of cells in each phase of the cell cycle when treated with medium only (control), different concentrations of 10 kDa dCGN (C10), of 40 kDa dCGN (C40), or of native CGN (Native).

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