Polissacarídeos sulfatados extraídos da alga dictyota mertensii protegem células de linhagem óssea do estresse oxidativo e induzem a diferenciação óssea

Abstract

Some antioxidant compounds are able to reduce the levels of reactive oxygen species (ROS) and consequently reduce the deleterious effects of ROS in osteoblasts, thus avoiding an imbalance in bone tissue homeostasis, since damage to osteoblasts converges to bone fragility. Fucans and fucoidans are polysaccharides rich in sulfated L-fucose, they are found mainly in brown seaweed, this group of polysaccharides are attributed pharmacological activities as anticoagulant, anti-tumor and antioxidant. Considering its antioxidant character and vast abundance of macroalgae on the Brazilian coast, the present study aimed evaluate the antioxidant and osteogenic potential of rich fractions of fucoidan from the brown macroalgae Dictyota mertensii – a tropical algae easily found in the Brazil’s northeast coast, through the presence of these compounds in pre-osteoblastic and mesenchymal cells. Thought techniques of precipitation in increasing volumes of acetone, 6 fractions rich in fucoidan (FRFs) were obtained from the D. mertensii algae: F03, F0.5, F0.7, F1.0, F1.5 and F2.1, confirming the presence of fucoidans by physicochemical analyzes. Except for F0.3, which only had relevant activity in the test of total antioxidant capacity, the other fractions had antioxidant activity in vitro in most of the assays evaluated. Hydroxyl radical and superoxide ions scavenging activity were not detected activity higher than 50% in none of the FRFs at the concentrations studied. Similarly, when MCT3-E1 pre-osteoblastic cells were exposed to an oxidative stress by hydrogen peroxide (H2O2) and treated with FRFs, the MTT reduction test showed that, except F0.3, all others FRFs had protective effect, with emphasis on F0.7, F1.5 and F2.1. This protective effect was also observed in the evaluation of the activity of caspase-3 and caspase-9, again showing that cells damaged by H2O2 and treated with F0.7, F1.5 and F2.1 fractions had less apoptotic activity, being protective for the cells. The activity of the antioxidant enzyme superoxide dismutase (SOD) such as alkaline phosphatase (ALP) – a marker of osteoblastic activity, are also increased in the MC3T3 cells treated with the FRFs, when compared to the control even in the presence of H2O2, this indicates a osteoblast differentiation. To evaluate the osteogenic activity of FRFs, human mesenchymal cells (Msc) were treated with the FRFs and evaluated the viability of these cells by the MTT reduction assay. As noted, the FRFs studied are not toxic to Msc cells when applied at up to 0.1 mg / mL; moreover, through cytochemistry, it was observed that on days 14 and 21 Msc treated with the fractions F0.7 and F1.5 increased the synthesis of collagen and alkaline phosphatase, the first one intensely more at the beginning and the second later. These data indicate a possible induction of these mesenchymal cells to osteoblasts, which is confirmed by scanning electron microscopy (Mev), through which it is observed mineralization points in the cells treated with these fractions, confirmed by energy-dispersive X-ray spectroscopy (EDS). The results described here indicate that F0.7 and F1.5 fractions obtained from D. mertensii algae as well as antioxidants present an osteogenic potential.