Induction of apoptosis in thymocytes by Hippophae rhamnoides: implications in radioprotection

Hippophae rhamnoides (RH-3), which has been recently reported to elicit dose-dependent pro- and antioxidant properties in vitro, induced apoptosis in murine thymocytes. In a concentration-dependent manner, RH-3 induced apoptosis in thymocytes in ex vivo conditions. The maximum effect was observed with 100 microg/mL of RH-3. Beyond this dose, the induction of apoptosis was inhibited, as seen on the ladder formation. However, apoptotic body formation, another indicator of apoptosis, was not manifested when various doses of RH-3 (20-200 microg/mL) were administered. RH-3 (>100 microg/mL) compacted chromatin in the form of densely stained masses, and subsequent treatment with proteinase-K loosened them and developed a halo around each mass. RH-3 treatment of cells that had already undergone apoptosis induced chromatin compaction, which made the ladder invisible. During in vivo experiments in mice, the radioprotective dose of RH-3 (30 mg/kg b.w.) induced significant DNA fragmentation in thymocytes studied spectrofluorimetrically. RH-3 treatment before irradiation in vivo enhanced radiation-induced apoptosis. These results were confirmed by hypodiploid population studied flow-cytometrically and also by ladder formation. RH-3 treatment was prooxidative in nature because it depleted thiols and enhanced lipids peroxidation after 8 hours of treatment. The paradox between the prooxidant and the antioxidant effects of RH-3 in the context of its overall radioprotective efficacy has been explained.