Shikonin regulates HeLa cell death via caspase-3 activation and blockage of DNA synthesis

Abstract

Shikonin, isolated from the plant Lithospermum erythrorhizon Sieb. Et Zucc, inhibited tumor cell growth and induced cell death in various tumor cells, with 50% growth inhibition of human cervical cancer cells, HeLa, at {\rm 18.9 \pm 1.1 \hspace{0.167em} \mgreek{m} mol \hspace{0.167em} L^{-1}.} Treated with 40 μmol L^-1 shikonin, HeLa cells underwent marked apoptotic morphological changes such as a round shape, membrane blebbing and apoptotic bodies derived from the fragmented nuclei. Another hallmark of apoptosis, DNA fragmentation, was observed by gel electrophoresis. Shikonin (10 μmol L^-1) significantly blocked the transition from G1 to S phase in the HeLa cell cycle. Pan-caspase inhibitor (Z-VAD-FMK), caspase-3 inhibitor (Z-DEVD-FMK) or caspase-8 inhibitor (Z-IETD-FMK) effectively inhibited shikonin-induced cell death, while caspase-1 inhibitor (Ac-YVAD-CMK) and caspase-9 inhibitor (Z-LEHD-FMK) failed to affect cell death. Caspase-3 activity significantly increased within 12 h after shikonin treatment. Reduced expression of inhibitor of caspase-activated deoxyribonuclease (ICAD) after exposure to shikonin for 12 h suggests the resultant activation of caspase-activated deoxyribonuclease (CAD), leading to apoptosis.

Keywords:

• Shikonin
• Apoptosis
• HeLa cells
• Caspases