Intranuclear inclusions in hepatocellular carcinoma contain autophagy-associated proteins and correlate with prolonged survival

GND
1113307943
ORCID
0000-0002-0320-4653
Affiliation
Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Schwertheim, Suzan;
GND
1171437064
Affiliation
Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Westerwick, Daniela;
GND
136830617
Affiliation
Institute of Anatomy, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Jastrow, Holger;
Affiliation
Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Theurer, Sarah;
Affiliation
Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Schaefer, Christoph M.;
GND
1014026865
LSF
60868
Affiliation
Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Kälsch, Julia;
Affiliation
Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Möllmann, Dorothe;
Affiliation
Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Schlattjan, Martin;
GND
124865798
LSF
60121
Affiliation
Department of Gastroenterology and Hepatology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Wedemeyer, Heiner;
GND
1140538640
LSF
14799
Affiliation
Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Schmid, Kurt Werner;
GND
1208859102
Affiliation
Institute of Pathology, University Hospital of Essen, University of Duisburg-Essen, Essen, Germany.
Baba, Hideo A.

For decades, intranuclear inclusions in many normal and neoplastic cells have been considered to be mere invaginations of cytoplasm into the nucleus without any notable function or influence on disease. We investigated such inclusions in 75 specimens of hepatocellular carcinoma (HCC). In this context we demonstrate that these inclusions are true inclusions, completely closed and delimited by the nuclear membrane, containing degenerate cell organelles and lysosomal proteins. Moreover, their occurrence was positively associated with patient survival but not with tumour grade or stage. In a standardised area a mean of 124 inclusions per specimen was present in the tumorous liver tissue in contrast to 5 inclusions in the non-tumorous adjacent section and 89% of all scrutinised HCC showed at least one membrane-bound nuclear inclusion. Ultrastructural characterisation by transmission electron microscopy revealed degenerative materials such as residues of lysosomes, endoplasmic reticulum and Golgi apparatus within the inclusions. Due to the fact that the content of the inclusions appears to be more condensed than cytoplasm and contains fewer intact cell organelles, we assume that they are not mere invaginations of cytoplasm. Three dimensional (3D) reconstruction of isolated and immunofluorescence stained nuclei showed that the inclusions are completely located within the nucleus without any connection to the cytoplasm. The limiting membrane of the inclusions contained lamin B suggesting nuclear membrane origin. The content of the inclusions stained for the autophagy-associated proteins p62, ubiquitin, LC3B, cathepsin B and cathepsin D. Triple immunofluorescence staining followed by 3D reconstruction revealed co-localisation of p62, ubiquitin and LC3B in the same inclusion. Our observations uncover that these inclusions are real inclusions completely surrounded by the nucleus. We propose that the presence of autophagy-associated proteins and proteases within the inclusions contribute to beneficial survival.

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