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Heliyon
2020 Jan 16;61:e03199. doi: 10.1016/j.heliyon.2020.e03199.
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Down-regulation of TGF-β, VEGF, and bFGF in vascular endothelial cells of chicken induced by a brittle star (Ophiocoma erinaceus) extract.
Kachooei SA
,
Rahmani R
,
Zareh N
,
Donyadideh F
,
Kachooei SA
,
Nabiuni M
,
Yazdansetad S
.
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Great attention has been focused on the discovery of anti-angiogenic natural and synthetic compounds to be finally used as or at least a part of the treatment of tumors. The marine ecosystems provide diversity in natural chemicals with the potential of being exploited as medicines in the treatment of diseases. Several studies have investigated Ophiuroids as a source of anti-tumor and anti-metastatic organisms. Here, we described the inhibitory effects of an ethanolic crude extract of brittle star (Ophiocoma erinaceus) on angiogenesis and the expression level of TGF-β, VEGF, and bFGF in chicken chorioallantoic membrane (CAM) as an experimental model. To do this 45 embryonated eggs were randomly divided into six groups including the control group, sham, three experimental groups and positive. The number and the length of vessels were calculated using ImageJ® software. The relative mRNA levels of the genes in different groups were evaluated by qRT-PCR method. Our study was suggestive of an anti-angiogenesis effect of brittle star ethanolic crude extract in a CAM model. The extract also showed a pharmacological effect of down-regulation of mRNA related to VEGF, TGF-β, and bFGF genes on chicken vascular endothelial cells. It was also showed that the observed inhibitory effect is with a dose-dependent manner in which the highest inhibitory effect belonged to the highest used dose. We indicated the anti-angiogenesis properties of the Persian Gulf brittle star. Further studies are needed in other aspects of the brittle star extract in the treatment of angiogenesis, hyperplasia, and cancers.
Figure 1. Photomicrographs of CAM in the control and treatment groups. The effect of the extract on angiogenesis is showed in these figures. (A) positive control (100 μg/ml sutent®), (B) negative control (C) sham exposure, (D) first treatment (25 μg/ml), (E) second treatment (50 μg/ml), (F) third treatment (100 μg/ml). As indicated, a dose dependent manner was seen in treatment groups by extracts. The picture magnification is 5.5.
Figure 2. Mean of the number of blood vessels in experimented (treated) groups (n = 3, Mean ± SD, ***p < 0.001). The decreasing effect of the extract on the number of blood vessels was shown quantitatively. As seen, the extract affected the number of blood vessels in a dose dependent manner.
Figure 3. Mean of the length of blood vessels in experimented (treated) groups (n = 3, Mean ± SD, ***p < 0.001). The effect of the extract on the length of blood vessels was shown quantitatively. As seen, the extract affected the length of blood vessels in a dose dependent manner.
Figure 4. The relative expression of VEGF, bFGF, and TGF-β genes in exprimented (treated) groups (n = 3, Mean ± SD, ***P<0.001, **P<0.05). The figure shows the results of real time PCR assay using bar chart. As it can be seen, the extract of brittle star reduces the expression of VEGF, bFGF, and TGF-β genes in treatment and positive control groups so that the highest reduction in mRNA was related to positive control group and the treatment groups also showed a dose dependent decrease in expression of these genes. This reduction in mRNA levels related to TGF-β gene in the first and second treatment groups were not significant (p>0.05) and was significant for third treatment and positive control groups with a level significance of p<0.001. The reduction of the mRNA levels related to bFGF gene was significant for first treated group with a level significance of p<0.05 and for second and third groups with a level significance of p<0.001. The reduction level related to VEGF gene was significant for first and second treated groups with a level significance of p<0.05 and for third group and positive control with a level significance of p<0.001.
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