Journal de la maladie d'Alzheimer et du parkinsonisme

Abstrait

Hydroxynonenal Causes Lysosomal and Autophagic Failure in the Monkey POMC Neurons

Tetsumori Yamashima Levy, Piyakarn Boontem, Hidenori Kido, Masahiro Yanagi, Takuya Seike, Daisuke Yamamiya, Shihui Li, Tatsuya Yamashita, Mitsuru Kikuchi, Eishiro Mizukoshi

Nowadays, overweight people have increased worldwide, giving rise to global ‘obesity epidemic’. Appetite and energy balance are regulated by the arcuate nucleus of hypothalamus. Pro-opiomelanocortin (POMC) neurons play an important role for the appetite suppression by sensing leptin, insulin, free fatty acids, etc. in the blood. High-fat diet is known to be associated with POMC neurodegeneration and related lifestyle-related diseases by generating reactive oxygen species. However, the mechanism by which excessive fatty acids induce POMC neurodegeneration remains unknown. We recently reported that lipid-peroxidation product ‘hydroxynonenal’ causes lysosomal membrane disintegrity via calpain-mediated cleavage of the oxidized (carbonylated) Hsp70.1. After the consecutive injections of synthetic hydroxynonenal in monkeys to make the serum concentration relevant to human 60’s, here we studied its adverse effect upon POMC neurons. By light microscopy, many POMC neurons showed dissolution of the cytoplasm and nuclear chromatin, and were positive for Fluoro-Jade C staining. Immunoreactivities of GPR40 (free fatty acid receptor), μ-calpain (Ca 2+ -dependent papain-like protease), Hsp70.1 (chaperone protein and lysosomal stabilizer), and p62 (chaperone for autophagic removal), showed an increased immunoreactivity within the degenerating POMC neurons. Albeit the decrease of POMC neurons in the arcuate nucleus tissue, Western blotting could confirm slight upregulation of these proteins. Furthermore, permeabilization of the lysosomal limiting membrane was suggested by the enlarged immunoreactive area double-positive for Lamp-2 and cathepsin B. Electron-microscopic analysis showed a remarkable decrease of lysosomes and a concomitant increase of autophagosomes. Lysosomes measuring 300~500 nm were round or oval with the distinct limiting membrane, whereas autophagosomes measuring 350~800 nm had an irregular configuration, being devoid of the limiting membrane. Lysosomes prior to fusing with autophagosomes showed permeabilization of the limiting membrane. Although up-regulation of the calpain-mediated Hsp70.1 cleavage was hardly demonstrated on immunoblots because of the POMC neuronal loss, Hsp70.1 disorder presumably contributed to the lysosomal membrane disintegrity and autophagy deficiency.