Peroxynitrite-induced apoptosis in epithelial (T84) and macrophage (RAW 264.7) cell lines: effect of legume-derived polyphenols (phytolens)

Nitric Oxide. 1997;1(6):476-83. doi: 10.1006/niox.1997.0160.

Abstract

Peroxynitrite (ONOO-) has been proposed as a mediator of gut inflammation and as an inducer of cell death by apoptosis. Phytolens (PHY), a water-soluble extract of polyphenolic antioxidants from nonsoy legumes (Biotics Research Corp, patent pending), was evaluated as a cytoprotective agent in human colonic (T84) and murine macrophage (RAW 264.7) cell lines. In the antioxidant testing, PHY showed a significant free radical scavenging ability against 1,1-diphenyl-2-picrylhydrazyl radical (DPPH) and superoxide (O2.) radicals with an IC50 of 4.44 and 5.87 microg/ml against DPPH and O2., respectively. Apoptosis (DNA fragmentation) was measured by an ELISA technique. Cells were exposed to oxidative stress by treating them with peroxynitrite (100-300 microM) for 4 h in the presence and absence of PHY. Peroxynitrite elicited a dose-dependent increase in DNA fragmentation in both cell lines compared to the control group receiving decomposed ONOO-. PHY (10, 30, or 50 microg/ml) significantly attenuated the degree of apoptosis in T84 cells induced by ONOO- (P < 0.05). PHY (10-100 microg/ml) did not directly affect T84 cell viability or induce apoptosis after 4 h or overnight exposure. RAW 264.7 cells exposed to PHY alone (>30 microg/ml) for 4 h displayed decreased cell viability (P < 0.05) and increased apoptosis (P < 0.05). Phytolens may have beneficial effects on inflammation by attenuating peroxynitrite-induced apoptosis. The sparing of epithelial cells while compromising the viability of macrophages suggests that PHY may be beneficial in autoimmune disorders.

MeSH terms

  • Animals
  • Antioxidants / pharmacology
  • Apoptosis / drug effects*
  • Bepridil / analogs & derivatives
  • Bepridil / metabolism
  • Biphenyl Compounds
  • Cell Division / drug effects
  • Cell Line
  • Cell Survival / drug effects
  • Colonic Neoplasms / metabolism
  • DNA Fragmentation / drug effects
  • Fabaceae / chemistry
  • Flavonoids*
  • Free Radical Scavengers / metabolism
  • Humans
  • Inflammation / drug therapy
  • Macrophages / metabolism
  • Nitrates / pharmacology*
  • Phenols / pharmacology*
  • Picrates*
  • Plants, Medicinal
  • Polymers / pharmacology*
  • Polyphenols
  • Rats
  • Superoxides / metabolism

Substances

  • Antioxidants
  • Biphenyl Compounds
  • Flavonoids
  • Free Radical Scavengers
  • Nitrates
  • Phenols
  • Picrates
  • Polymers
  • Polyphenols
  • Superoxides
  • peroxynitric acid
  • Bepridil
  • 1,1-diphenyl-2-picrylhydrazyl