Effects of 3-methoxy-2(5H)-furanone-containing extracts from Narthecium ossifragum (L.) Huds. on renal tubular cells in vitro

Narthecium ossifragum, a perennial herb of the lily family, causes toxic renal tubular necrosis in several ruminant species. 3-methoxy-2(5H)-furanone (3M2F) has been identified as a nephrotoxin present in N. ossifragum extracts. We studied effects of three different 3M2F-containing fractions isolated from N. ossifragum and synthetic 3M2F on the porcine kidney cell line LLC-PK1. In some of the experiments, we included the glioma cell lines U251 and BT4Cn to compare the effects of the toxin on LLC-PK1 cells to the effect on these cell lines. The synthetic 3M2F was shown to be only mildly toxic, and the most purified fraction from N. ossifragum showed the highest degree of toxicity in our studies. When monolayer cultures were exposed to increasing amounts of 3M2F-containing extract, a dose-dependent increase in cell death was observed. Similarly, reduced neutral red uptake and 3H-thymidine uptake (DNA synthesis) was observed. There was increased apoptotic activity in the LLC-PK1 cells with increasing concentration of 3M2F-containing extract. Multicellular three-dimensional spheroids from LLC-PK1 cells stopped fluid transport, showed degenerative changes and collapsed totally 6 h after extract exposure. Our findings indicate junctional damage, reduced cellular endocytosis and DNA-synthesis as well as induction of apoptosis as possible mechanisms for the acute tubular necrosis observed in ruminant species.