A novel proanthocyanidin IH636 grape seed extract increases in vivo Bcl-XLexpression and prevents acetaminophen-induced programmed and unprogrammed cell death in mouse liver

Several molecular events in the apoptotic or necrotic death of hepatocytes induced by acetaminophen (AAP) now appear to be well defined. Recent studie s also indicate that select expression of bcl-XL is possibly modified durin g AAP-induced liver injury. The purpose of this study was several-fold: (i) to examine the hepatoprotective ability of short-term (3-day) and long-ter m (7-day) exposures of a grape seed pro-anthocyanidin extract (GSPE) on AAP -induced liver injury and animal lethality; (ii) to monitor effects of GSPE on one of the prime targets of AAP, i.e., hepatocellular genomic DNA and a ssociated apoptotic and necrotic death; and (iii) to unravel changes in the pattern of expression of an antiapoptotic gene, bcl-XL in the liver. In or der to investigate these events, male ICR mice (30-40 g) were administered nontoxic doses of GSPE (3 or 7 days, 100 mg/kg, po), followed by hepatotoxi c doses of AAP (400 and 500 mg/kg, ip), and sacrificed 24 h later. Serum wa s analyzed for alanine aminotransferase activity (ALT) and the liver for hi stopathological diagnosis of apoptosis/necrosis. The ability of AAP to prom ote apoptotic DNA fragmentation and its counteraction by GSPE in the liver was also evaluated quantitatively (by a sedimentation assay) and qualitativ ely (by agarose gel electrophoresis). Portions of livers were also subjecte d to Western blot analysis (21,000g fraction of liver homogenates) to exami ne the pattern of expression of cell death inhibitory gene bcl-XL. Results indicate that 7-day GSPE preexposure induced dramatic protection and marked ly decreased liver injury and animal lethality culminated by AAP, when comp ared to a short-term 3-day exposure. Abrogation of toxicity was also mirror ed in DNA fragmentation. Histopathological evaluation of liver sections sho wed remarkable counteraction of AAP-toxicity by this novel GSPE and substan tial inhibition of both apoptotic and necrotic liver cell death. Agarose ge l electrophoresis revealed that 7-day GSPE preexposure prior to AAP adminis tration completely blocked Ca2+/Mg2+-Ca2+/Ng(2+)-dependent-endonuclea media ted ladder-like fragmentation of genomic DNA and significantly altered the bcl-XL expression. The most dramatic changes observed in this; study were: (i) substantial increase in the expression of bcP-XL in the liver by 7-day GSPE exposure alone; (ii) significant modification bcl-XL expression by AAP alone; and (iii) dramatic inhibition of AAP-induced modification of bcl-XL (phosphorylation?) expression by GSPE. In summary, these observations demo nstrate that GSPE preexposure may significantly attenuate AAP-induced hepat ic DNA damage, apoptotic and necrotic cell death of liver cells, and, most remarkably, antagonize the influence of AAP-induced changes in bcP-XL expre ssion in vivo. (C) 1999 Academic Press.