Stereological measurement of porto-central gradients in gene expression in mouse liver

Gieling, Roben G.; Markman, Marry; Hagoort, Jaco; Lamers, Wouter H.

doi:10.1002/hep.20068

Cited by 20 publications

(14 citation statements)

References 32 publications

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“…Inmunostaining experiments show a PEPCK-C gradient across the porto-central axis of the liver acinus (21). Silencing efficiency was higher in the perivenous portion of the acinus (Fig.…”

Section: Discussionmentioning

confidence: 91%

Pck1 Gene Silencing in the Liver Improves Glycemia Control, Insulin Sensitivity, and Dyslipidemia in db/db Mice

Méndez-Lucas²,

et al. 2008

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OBJECTIVE-Cytosolic phosphoenolpyruvate carboxykinase (PEPCK-C; encoded by Pck1) catalyzes the first committed step in gluconeogenesis. Extensive evidence demonstrates a direct correlation between PEPCK-C activity and glycemia control. Therefore, we aimed to evaluate the metabolic impact and their underlying mechanisms of knocking down hepatic PEPCK-C in a type 2 diabetic model. RESEARCH DESIGN AND METHODS-PEPCK-C gene targeting was achieved using adenovirus-transduced RNAi. The study assessed several clinical symptoms of diabetes and insulin signaling in peripheral tissues, in addition to changes in gene expression, protein, and metabolites in the liver. Liver bioenergetics was also evaluated.RESULTS-Treatment resulted in reduced PEPCK-C mRNA and protein. After treatment, improved glycemia and insulinemia, lower triglyceride, and higher total and HDL cholesterol were measured. Unsterified fatty acid accumulation was observed in the liver, in the absence of de novo lipogenesis. Despite hepatic lipidosis, treatment resulted in improved insulin signaling in the liver, muscle, and adipose tissue. O 2 consumption measurements in isolated hepatocytes demonstrated unaltered mitochondrial function and a consequent increased cellular energy charge. Key regulatory factors (FOXO1, hepatocyte nuclear factor-4␣, and peroxisome proliferator-activated receptor-␥ coactivator [PGC]-1␣) and enzymes (G6Pase) implicated in gluconeogenesis were downregulated after treatment. Finally, the levels of Sirt1, a redox-state sensor that modulates gluconeogenesis through PGC-1␣, were diminished. CONCLUSIONS-Our observations indicate that silencing PEPCK-C has direct impact on glycemia control and energy metabolism and provides new insights into the potential significance of the enzyme as a therapeutic target for the treatment of diabetes.

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“…Inmunostaining experiments show a PEPCK-C gradient across the porto-central axis of the liver acinus (21). Silencing efficiency was higher in the perivenous portion of the acinus (Fig.…”

Section: Discussionmentioning

confidence: 91%

Pck1 Gene Silencing in the Liver Improves Glycemia Control, Insulin Sensitivity, and Dyslipidemia in db/db Mice

Méndez-Lucas²,

et al. 2008

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“…The two-layer-thick cuff of GS-positive hepatocytes around the central veins represents only 5% of all hepatocytes in mouse liver. (26) Pericentral hepatocytes are, therefore, 20-fold more efficient per cell at removing ammonia than periportal hepatocytes. Accordingly, the downstream, pericentral hepatocytes are well equipped to remove residual ammonia from the hepatic circulation (4,27) because they express the ammonia transporters RhBG (28) and APQ9, (29) the glutamate transporter SLC1A2, (30,31) the glutamate-producing enzyme ornithine aminotransferase, (32) the glutamine-producing enzyme GS, (33,34) and the glutamine transporter SLC38A3.…”

Section: Gs Activity Provides For Systemic High-affinity Ammonia Detomentioning

confidence: 99%

Pivotal role of glutamine synthetase in ammonia detoxification

et al. 2017

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Glutamine synthetase (GS) catalyzes condensation of ammonia with glutamate to glutamine. Glutamine serves, with alanine, as a major nontoxic interorgan ammonia carrier. Elimination of hepatic GS expression in mice causes only mild hyperammonemia and hypoglutaminemia but a pronounced decrease in the whole-body muscle-to-fat ratio with increased myostatin expression in muscle. Using GS-knockout/liver and control mice and stepwise increments of enterally infused ammonia, we show that 35% of this ammonia is detoxified by hepatic GS and 35% by urea-cycle enzymes, while 30% is not cleared by the liver, independent of portal ammonia concentrations £ 2 mmol/L. Using both genetic (GSknockout/liver and GS-knockout/muscle) and pharmacological (methionine sulfoximine and dexamethasone) approaches to modulate GS activity, we further show that detoxification of stepwise increments of intravenously (jugular vein) infused ammonia is almost totally dependent on GS activity. Maximal ammonia-detoxifying capacity through either the enteral or the intravenous route is 160 lmol/hour in control mice. Using stable isotopes, we show that disposal of glutaminebound ammonia to urea (through mitochondrial glutaminase and carbamoylphosphate synthetase) depends on the rate of glutamine synthesis and increases from 7% in methionine sulfoximine-treated mice to 500% in dexamethasone-treated mice (control mice, 100%), without difference in total urea synthesis. Conclusions: Hepatic GS contributes to both enteral and systemic ammonia detoxification. Glutamine synthesis in the periphery (including that in pericentral hepatocytes) and glutamine catabolism in (periportal) hepatocytes represents the high-affinity ammonia-detoxifying system of the body. The dependence of glutamine-bound ammonia disposal to urea on the rate of glutamine synthesis suggests that enhancing peripheral glutamine synthesis is a promising strategy to treat hyperammonemia. Because total urea synthesis does not depend on glutamine synthesis, we hypothesize that glutamate dehydrogenase complements mitochondrial ammonia production. (HEPATOLOGY 2017;65:281-293). G lutamine synthetase (GS) catalyzes condensation of ammonia with glutamate to glutamine. (Note: NH 3 is protonated for 98% to NH 4 1 at physiological pH. We use the term "ammonia" to refer to the sum of NH 3 and NH 4 1 unless specified as either "NH 3 " or "NH 41 .") Glutamine serves, with alanine, as a major nontoxic interorgan ammonia shuttle in the body (1) and as an aminomoiety donor for the synthesis of nucleotides, amino acids, amino-sugars, and oxidized nicotinamide adenine dinucleotide. Its intracellular turnover rate exceeds that of all other amino acids. GS deficiency causes only moderate hyperammonemia, but affected humans and mice suffer from encephalopathy and die neonatally. (2,3) GS is predominantly expressed in the nervous system, kidney, and liver, that is, in established glutamine-consuming organs, (4)(5)(6) whereas skeletal muscle, with a much lower expression but large mass, is considered the main net ...

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“…For measurement of the staining intensity, optical density (OD) images were acquired as recently described (Ruijter et al 2004). In short, a Photometrics cooled CCD camera (Tucson, AZ) with a 12-bit dynamic range (1317 3 1035 pixels) attached to an Axiophot microscope, a stabilized power supply, a 580 nm bandpass filter, and an infrared-blocking filter was used.…”

Section: Image Acquisition and Image Processingmentioning

confidence: 99%

Atrial and Ventricular Myosin Heavy-chain Expression in the Developing Chicken Heart: Strengths and Limitations of Non-radioactive In Situ Hybridization

Somi

Klein

Houweling

et al. 2006

J Histochem Cytochem.

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Myosin heavy-chain (MHC) isoforms are major structural components of the contractile apparatus of the heart muscle. Their spatio-temporal patterns of expression have been used as a tool to dissect cardiac development and differentiation. Although extensively investigated, controversy still exists concerning the expression patterns of atrial (AMHC), ventricular (VMHC), and cardiac myosin heavy-chain (CMHC) during development in the heart. In this study, we describe that probe length, probe concentration, and staining time in the non-radioactive in situ hybridization procedure seriously influence the observed pattern of MHC expression and the subsequent interpretation, explaining the divergent opinions in the field. Using a variety of external and internal controls for the in situ hybridization procedure, we demonstrate that both AMHC and VMHC are expressed throughout the entire heart tube during early development. During subsequent development, VMHC becomes restricted to the ventricles, whereas AMHC remains expressed in the atria, and, at substantially lower levels, is detected in the ventricles. These results are discussed in the context of methodological constraints of demonstrating patterns of gene expression. This manuscript contains online supplemental material at http://www.jhc.org . Please visit this article online to view these materials.

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Stereological measurement of porto-central gradients in gene expression in mouse liver

Cited by 20 publications

References 32 publications

Pck1 Gene Silencing in the Liver Improves Glycemia Control, Insulin Sensitivity, and Dyslipidemia in db/db Mice

Pck1 Gene Silencing in the Liver Improves Glycemia Control, Insulin Sensitivity, and Dyslipidemia in db/db Mice

Pivotal role of glutamine synthetase in ammonia detoxification

Atrial and Ventricular Myosin Heavy-chain Expression in the Developing Chicken Heart: Strengths and Limitations of Non-radioactive In Situ Hybridization

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