Activated macrophages express upregulated RIP3 signaling. (a)
RAW264.7 macrophages were exposed to varying concentrations of LPS
(0 µg/ml, 1 µg/ml, 3 µg/ml, 6 µg/ml and 10 µg/ml) for 24 hours.
Expression levels of RIP1, RIP3, and MLKL were examined by Western
blot. RIP1, RIP3, and MLKL levels were dose-dependently upregulated
following LPS stimulation. (b) RAW264.7 macrophages were incubated
with LPS (3 µg/ml) and/or Nec-1 (100 µM) as described in images.
Western blotting analysis showed that RIP1, RIP3, and MLKL levels
were downregulated in macrophages after co-treatment with Nec-1
compared to LPS alone treatment. *p < 0.05,
**p < 0.005,
***p < 0.0005. LPS: lipopolysaccharide; MLKL:
mixed lineage kinase domain-like; Nec-1: necrostatin-1; RIP:
receptor-interacting protein kinase.