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Energy determinants GAPDH and NDPK act as genetic modifiers for hepatocyte inclusion formation.

Snider NT, Weerasinghe SV, Singla A, Leonard JM, Hanada S, Andrews PC, Lok AS, Omary MB - J. Cell Biol. (2011)

Bottom Line: Prominent histological features of some chronic human liver diseases are hepatocyte ballooning and Mallory-Denk bodies.GAPDH knockdown depleted bioenergetic and antioxidant enzymes and elevated hepatocyte ROS, whereas GAPDH overexpression decreased hepatocyte ROS.We propose that GAPDH and NDPK are genetic modifiers of murine DDC-induced liver injury and potentially human liver disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, MI 48109, USA. nsnider@umich.edu

ABSTRACT
Genetic factors impact liver injury susceptibility and disease progression. Prominent histological features of some chronic human liver diseases are hepatocyte ballooning and Mallory-Denk bodies. In mice, these features are induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) in a strain-dependent manner, with the C57BL and C3H strains showing high and low susceptibility, respectively. To identify modifiers of DDC-induced liver injury, we compared C57BL and C3H mice using proteomic, biochemical, and cell biological tools. DDC elevated reactive oxygen species (ROS) and oxidative stress enzymes preferentially in C57BL livers and isolated hepatocytes. C57BL livers and hepatocytes also manifested significant down-regulation, aggregation, and nuclear translocation of glyceraldehyde 3-phosphate dehydrogenase (GAPDH). GAPDH knockdown depleted bioenergetic and antioxidant enzymes and elevated hepatocyte ROS, whereas GAPDH overexpression decreased hepatocyte ROS. On the other hand, C3H livers had higher expression and activity of the energy-generating nucleoside-diphosphate kinase (NDPK), and knockdown of hepatocyte NDPK augmented DDC-induced ROS formation. Consistent with these findings, cirrhotic, but not normal, human livers contained GAPDH aggregates and NDPK complexes. We propose that GAPDH and NDPK are genetic modifiers of murine DDC-induced liver injury and potentially human liver disease.

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Proteomic comparison of C3H and C57BL mouse livers before and after DDC treatment by 2D DIGE analysis and classification of differentially expressed proteins into function-based groups. (A) Liver homogenates were prepared from untreated mice (Control) or mice fed a DDC-containing diet for 3 mo. The liver proteins from the C3H and C57BL mice were labeled with the fluorescent dyes Cy3 (green) and Cy5 (red), respectively. The circled spots indicate differentially expressed proteins that were selected for identification by MS. (B) From an initial analysis of 80 selected gel spots, proteins that were identified as having significant expression differences (ratio greater than two) between the two mouse strains under basal conditions or after DDC treatment are shown. Note that PRDX6 is represented twice, reflecting the acidic and basic forms of the protein. (C) The proteins shown in B were classified into three groups based on their known functional properties.
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fig1: Proteomic comparison of C3H and C57BL mouse livers before and after DDC treatment by 2D DIGE analysis and classification of differentially expressed proteins into function-based groups. (A) Liver homogenates were prepared from untreated mice (Control) or mice fed a DDC-containing diet for 3 mo. The liver proteins from the C3H and C57BL mice were labeled with the fluorescent dyes Cy3 (green) and Cy5 (red), respectively. The circled spots indicate differentially expressed proteins that were selected for identification by MS. (B) From an initial analysis of 80 selected gel spots, proteins that were identified as having significant expression differences (ratio greater than two) between the two mouse strains under basal conditions or after DDC treatment are shown. Note that PRDX6 is represented twice, reflecting the acidic and basic forms of the protein. (C) The proteins shown in B were classified into three groups based on their known functional properties.

Mentions: A 2D DIGE analysis was conducted to compare the liver proteome of C3H (MDB resistant) and C57BL (MDB susceptible) mice. This approach yielded a large number of differentially expressed proteins in untreated and DDC-treated mice from the two strains (Fig. 1 A). The circled green- and red-colored spots (Fig. 1 A) were subsequently subjected to tandem mass spectrometry (MS/MS) for protein identification and determination of the relative expression ratios between the two strains. Of 80 targets whose identity was characterized by MS/MS, those that had the highest expression difference either before of after DDC treatment were selected for further characterization (Fig. 1 B). Function-based classification of the identified proteins resulted in three major functional categories: (1) protein processing, (2) energy metabolism, and (3) oxidative stress, with some overlap between the latter two (Fig. 1 C).


Energy determinants GAPDH and NDPK act as genetic modifiers for hepatocyte inclusion formation.

Snider NT, Weerasinghe SV, Singla A, Leonard JM, Hanada S, Andrews PC, Lok AS, Omary MB - J. Cell Biol. (2011)

Proteomic comparison of C3H and C57BL mouse livers before and after DDC treatment by 2D DIGE analysis and classification of differentially expressed proteins into function-based groups. (A) Liver homogenates were prepared from untreated mice (Control) or mice fed a DDC-containing diet for 3 mo. The liver proteins from the C3H and C57BL mice were labeled with the fluorescent dyes Cy3 (green) and Cy5 (red), respectively. The circled spots indicate differentially expressed proteins that were selected for identification by MS. (B) From an initial analysis of 80 selected gel spots, proteins that were identified as having significant expression differences (ratio greater than two) between the two mouse strains under basal conditions or after DDC treatment are shown. Note that PRDX6 is represented twice, reflecting the acidic and basic forms of the protein. (C) The proteins shown in B were classified into three groups based on their known functional properties.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3198167&req=5

fig1: Proteomic comparison of C3H and C57BL mouse livers before and after DDC treatment by 2D DIGE analysis and classification of differentially expressed proteins into function-based groups. (A) Liver homogenates were prepared from untreated mice (Control) or mice fed a DDC-containing diet for 3 mo. The liver proteins from the C3H and C57BL mice were labeled with the fluorescent dyes Cy3 (green) and Cy5 (red), respectively. The circled spots indicate differentially expressed proteins that were selected for identification by MS. (B) From an initial analysis of 80 selected gel spots, proteins that were identified as having significant expression differences (ratio greater than two) between the two mouse strains under basal conditions or after DDC treatment are shown. Note that PRDX6 is represented twice, reflecting the acidic and basic forms of the protein. (C) The proteins shown in B were classified into three groups based on their known functional properties.
Mentions: A 2D DIGE analysis was conducted to compare the liver proteome of C3H (MDB resistant) and C57BL (MDB susceptible) mice. This approach yielded a large number of differentially expressed proteins in untreated and DDC-treated mice from the two strains (Fig. 1 A). The circled green- and red-colored spots (Fig. 1 A) were subsequently subjected to tandem mass spectrometry (MS/MS) for protein identification and determination of the relative expression ratios between the two strains. Of 80 targets whose identity was characterized by MS/MS, those that had the highest expression difference either before of after DDC treatment were selected for further characterization (Fig. 1 B). Function-based classification of the identified proteins resulted in three major functional categories: (1) protein processing, (2) energy metabolism, and (3) oxidative stress, with some overlap between the latter two (Fig. 1 C).

Bottom Line: Prominent histological features of some chronic human liver diseases are hepatocyte ballooning and Mallory-Denk bodies.GAPDH knockdown depleted bioenergetic and antioxidant enzymes and elevated hepatocyte ROS, whereas GAPDH overexpression decreased hepatocyte ROS.We propose that GAPDH and NDPK are genetic modifiers of murine DDC-induced liver injury and potentially human liver disease.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Molecular and Integrative Physiology, University of Michigan, Ann Arbor, MI 48109, USA. nsnider@umich.edu

ABSTRACT
Genetic factors impact liver injury susceptibility and disease progression. Prominent histological features of some chronic human liver diseases are hepatocyte ballooning and Mallory-Denk bodies. In mice, these features are induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) in a strain-dependent manner, with the C57BL and C3H strains showing high and low susceptibility, respectively. To identify modifiers of DDC-induced liver injury, we compared C57BL and C3H mice using proteomic, biochemical, and cell biological tools. DDC elevated reactive oxygen species (ROS) and oxidative stress enzymes preferentially in C57BL livers and isolated hepatocytes. C57BL livers and hepatocytes also manifested significant down-regulation, aggregation, and nuclear translocation of glyceraldehyde 3-phosphate dehydrogenase (GAPDH). GAPDH knockdown depleted bioenergetic and antioxidant enzymes and elevated hepatocyte ROS, whereas GAPDH overexpression decreased hepatocyte ROS. On the other hand, C3H livers had higher expression and activity of the energy-generating nucleoside-diphosphate kinase (NDPK), and knockdown of hepatocyte NDPK augmented DDC-induced ROS formation. Consistent with these findings, cirrhotic, but not normal, human livers contained GAPDH aggregates and NDPK complexes. We propose that GAPDH and NDPK are genetic modifiers of murine DDC-induced liver injury and potentially human liver disease.

Show MeSH
Related in: MedlinePlus