Karyolysis is the complete dissolution of nuclear components of a dying cell. However, the generation mechanism has not been clarified. We studied a necrotic DNA fragmentation factor DNase γ (also known as DNase1L3) and previously found that karyolysis was inhibited in DNase γ deficient (DNase γ^−/−) mice. To confirm this, we transiently expressed DNase γ in the liver of DNase γ^−/− mice and caused hepatocyte necrosis by acetaminophen overdose. As expected, karyolysis was induced in the necrotic hepatocytes. We also found that the depletion of Kupffer cells from wild type mice reduced the expression and activity of DNase γ in the liver. Thus, we concluded that DNase γ produced from Kupffer cells caused karyolysis of necrotic hepatocytes.