Chlorella vulgaris modulates hydrogen peroxide-induced DNA damage and telomere shortening of human fibroblasts derived from different aged individuals

Abstract

The objective of this study was to investigate the modulatory effect of Chlorella vulgaris on cultured fibroblast cells derived from young and old aged individuals focusing on DNA damage, telomere length and telomerase activity. Dose-response test of the algal extract on cells in both age groups revealed that optimum viability was observed at a concentration of 50 microg/ml. Results obtained showed that Chlorella vulgaris exhibited protective effects against H(2)O(2)-induced oxidative stress as shown by the reduction in damaged DNA caused by H(2)O(2) treatment (p<0.05) in Chlorella vulgaris pre- and post-treated groups (p<0.05). Pre-treatment of Chlorella vulgaris resulted in a significant decrease in DNA damage suggesting a bioprotective effect against free radical attacks. A decline in DNA damage was observed in post-treated cells which proves Chlorella vulgaris to present bioremediative properties. In cells induced with oxidative stress, telomere length decreased significantly coupled with a concomitant decline of telomerase activity (p<0.05). However, these reductions were prevented with prior and post treatment of Chlorella vulgaris. Therefore, we concluded that Chlorella vulgaris exhibited bioprotective effects especially in cells obtained from young donor but were more bioremediative for cells obtained from old donor as indicated by DNA damage, telomere shortening and reduction in telomerase activity.

Keywords: Chlorella vulgaris; DNA damage; ageing; telomerase; telomere.

Figure 1

Effect of CVE hot water extract on fibroblasts derived from young and old aged human as assessed by MTS assay. Percent MTS reduction corresponds to the viable cell number. Fibroblasts were incubated with increasing concentrations of CVE for 24 hours at 37oC. Incubation with CVE caused a significant increase in the number of cell viable. The percentage of the viable cell was highest at CVE concentration of 50 ug/ml for fibroblasts from both young and old donors. The number of cell viable decreased with high concentration of CVE incubation. ^a,b Denotes p<0.05 compared to control, ^a*,b* p<0.05 compared to previous concentration for fibroblasts derived from both young and old donors respectively. Data is presented as means ± SD, n = 3.

Figure 2

Comets analysis by fluorescence microscopy with visual inspection of tail length of nuclei. The cell nuclei were classified into five categories: (0) undamaged (nuclei without comet tail); (1) low damaged (nuclei with comet tails up to two fold longer than nucleus diameter); (2) damaged (nuclei with comet tail two to three fold longer than nucleus diameter), (3) highly damaged (nuclei with comet tails three fold longer than nucleus diameter), and (4) severely damaged, respectively.

Figure 3a

Protective effects of CVE against H₂O₂-induced DNA damage in fibroblasts cultures. Fibroblasts derived from young aged human were treated with optimum concentration of CVE for 24 hours at 37oC before or after H₂O₂ exposure (IC₅₀) for 2 hours. DNA damage was assessed using Comet assay. Protective effect of CVE against H₂O₂-induced DNA damage was seen in fibroblast cells derived from young aged human. ^a Denotes p<0.05 compared to control, ^b p<0.05 compared to H₂O₂. Data is presented as mean ± SD, n=2.

Figure 3b

Protective effects of CVE against H₂O₂-induced DNA damage in fibroblasts cultures. Fibroblasts derived from old aged human were treated with optimum concentration of CVE for 24 hours at 37oC before or after H₂O₂ exposure (IC₅₀) for 2 hours. DNA damage was assessed using Comet assay. Protective effect of CVE against H₂O₂-induced DNA damage was seen in fibroblast cells derived from old aged human. ^a Denotes p<0.05 compared to control, ^b p<0.05 compared to H₂O₂. Data is presented as mean ± SD, n=2.

Figure 4a

Effect of CVE on telomere length (TRF length) of human skin fibroblast cell line derived from young aged human. Cells were treated with optimum dose of CVE prior or after H₂O₂ exposure. Protective effects of CVE against H₂O₂-induced telomere shortening was observed in fibroblast cells derived from young aged human. ^a Denotes p<0.05 compared to control, ^b p<0.05 compared to CVE, ^c p<0.05 compared to H₂O₂. Data is presented as means ± SD, n = 3.

Figure 4b

Effect of CVE on telomere length (TRF length) of human skin fibroblast cell line derived from old aged human. Cells were treated with optimum dose of CVE prior or after H₂O₂ exposure. Protective effects of CVE against H₂O₂-induced telomere shortening was observed in fibroblast cells derived from old aged human. ^a Denotes p<0.05 compared to control, ^b p<0.05 compared to CVE, ^c p<0.05 compared to H₂O₂. Data is presented as means ± SD, n = 3

Figure 5a

Effect of CVE on telomerase activity (Total Product Generated, TPG) of human skin fibroblast cell line derived from young aged human. Cells were treated with optimum dose of CVE prior or after H₂O₂ exposure. Pretreatment with optimum dose of CVE protected against H₂O₂-induced decreased telomerase activity in fibroblast cells derived from young aged human. ^a Denotes p<0.05 compared to control, ^b p<0.05 compared to CVE, ^c p<0.05 compared to H₂O₂. Data is presented as means ± SD, n = 3.

Figure 5b

Effect of CVE on telomerase activity (Total Product Generated, TPG) of human skin fibroblast cell line derived from old aged human. Cells were treated with optimum dose of CVE prior or after H₂O₂ exposure. Pretreatment with optimum dose of CVE protected against H₂O₂-induced decreased telomerase activity in fibroblast cells derived from old aged human. ^a Denotes p<0.05 compared to control, ^b p<0.05 compared to CVE, ^c p<0.05 compared to H₂O₂. Data is presented as means ± SD, n = 3.