The Protective Effects of Silk Sericin Against Retinal Oxidative Stress: In Vitro and In Vivo Assays with a Fluorometric Nitroxide Molecular Probe

Sericin is a major polypeptidic constituent of the silk in the cocoons produced by the <i>Bombyx mori</i> silkworm. Certain fractions isolated from sericin exhibited antioxidant properties in a variety of reported experimental settings. In a previous study, we found that only the non-protein fraction, extracted from crude sericin, displayed antioxidative activity in cultures of murine retinal photoreceptor cells (661W), a cell line that is highly sensitive to oxidative stress associated with diseases of the retina. In the same assay, the protein fraction (purified sericin) did not show any such activity. To check these findings, in the present study, two additional different assays were employed: an <i>in vitro</i> assay based on the dose-dependent mitigating effects exerted by each sericin fraction on the activity of antimycin A in cultures of 661W cells and an <i>in vivo</i> assay based on an animal (rat) model of retinal ischemia/reperfusion injury. In both assays, nitroxide was appended as a fluorometric molecular probe, and fluorescent intensity was monitored by either flow cytometry (<i>in vitro</i>) or the Micron IV retinal imaging system (<i>in vivo</i>). The <i>in vitro</i> assay indicated unequivocally antioxidative capacity for the non-protein fraction and a lack of it for the purified sericin. The <i>in vivo</i> assay indicated that each fraction was able to act as an antioxidant. We hypothesized that the ability of purified sericin to display antioxidative activity <i>in vivo</i>, but not <i>in vitro</i>, was the result of the metabolic degradation of sericin, a process that delivered serine, an amino acid with known antioxidant properties. However, this hypothesis needs experimental confirmation.