Steatohepatitis-inducing drugs trigger cytokeratin cross-links in hepatocytes. Possible contribution to Mallory-Denk body formation

Mallory-Denk bodies (MDB) are hepatocyte inclusions containing cytokeratin 8 (CK8) which can develop, along with other steatohepatitis lesions, in patients treated with amiodarone, perhexiline maleate or 4,4′-diethylaminoethoxyhexestrol. These drugs accumulate lipids, whose subsequent peroxidation liberates reactive by-products, like malondialdehyde (MDA). The formation of MDB has been previously reproduced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine or griseofulvin administration which cross-link CK8 by tissue transglutaminase, thus forming an entangled network, from which MDB progressively arise. The present study depicts the mechanisms initiating MDB formation by steatohepatitis-inducing drugs. Short incubation of hepatocytes with amiodarone (50 μM), 4,4′-diethylaminoethoxyhexestrol (50 μM) or perhexiline maleate (25 μM) increased the pool of CK8 monomers and increased cell calcium to activate Ca++-dependent transglutaminases which cross-linked the CK8 monomers into CK8-containing oligomers. The present study also provides the first evidence that MDA might directly participate in MDB formation, as this reactive agent cross-linked purified CK8 or albumin in vitro, disrupted the cytokeratin network of isolated hepatocytes, and bridged CK8 molecules. In conclusion, steatohepatitis-inducing drugs increase cell calcium and activate tissue transglutaminase, which cross-links CK8 to form a molecular scaffold, from which MDB might secondarily arise. Malondialdehyde also cross-links CK8, albeit through a different mechanism, and might also contribute to MDB formation.