A modified PAS stain combined with immunofluorescence for quantitative analyses of glycogen in muscle sections

Schaart, Gert; Hesselink, Reinout P.; Keizer, H. A.; Kranenburg, Guido; Drost, Maarten R.; Hesselink, Matthijs K. C.

doi:10.1007/s00418-004-0690-0

Cited by 67 publications

(55 citation statements)

References 23 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The method for the combined PAS and immunofluorescence staining was adapted from Ref. 29. The samples with both PAS and immunofluorescence staining were viewed on a Zeiss Axio Observer Z1 microscope, and images were processed with Zeiss Axiovision 4.7.…”

Section: Methodsmentioning

confidence: 99%

Starch Binding Domain-containing Protein 1/Genethonin 1 Is a Novel Participant in Glycogen Metabolism

Jiang

Heller

Tagliabracci

et al. 2010

Journal of Biological Chemistry

127

118

View full text Add to dashboard Cite

Stbd1 is a protein of previously unknown function that is most prevalent in liver and muscle, the major sites for storage of the energy reserve glycogen. The protein is predicted to contain a hydrophobic N terminus and a C-terminal CBM20 glycan binding domain. Here, we show that Stbd1 binds to glycogen in vitro and that endogenous Stbd1 locates to perinuclear compartments in cultured mouse FL83B or Rat1 cells. When overexpressed in COSM9 cells, Stbd1 concentrated at enlarged perinuclear structures, co-localized with glycogen, the late endosomal/lysosomal marker LAMP1 and the autophagy protein GABARAPL1. Mutant Stbd1 lacking the N-terminal hydrophobic segment had a diffuse distribution throughout the cell. Point mutations in the CBM20 domain did not change the perinuclear localization of Stbd1, but glycogen was no longer concentrated in this compartment. Stable overexpression of glycogen synthase in Rat1WT4 cells resulted in accumulation of glycogen as massive perinuclear deposits, where a large fraction of the detectable Stbd1 co-localized. Starvation of Rat1WT4 cells for glucose resulted in dissipation of the massive glycogen stores into numerous and much smaller glycogen deposits that retained Stbd1. In vitro, in cells, and in animal models, Stbd1 consistently tracked with glycogen. We conclude that Stbd1 is involved in glycogen metabolism by binding to glycogen and anchoring it to membranes, thereby affecting its cellular localization and its intracellular trafficking to lysosomes.

show abstract

Section: Methodsmentioning

confidence: 99%

Starch Binding Domain-containing Protein 1/Genethonin 1 Is a Novel Participant in Glycogen Metabolism

Jiang

Heller

Tagliabracci

et al. 2010

Journal of Biological Chemistry

127

118

View full text Add to dashboard Cite

show abstract

“…For each muscle biopsy a total of 58±7 and 49±4 muscle fibres were analysed for lipid content for diabetes patients and control subjects respectively. To permit quantification of intramyocellular glycogen we used the modified PAS stain [35]. For each muscle biopsy, 148±15 and 157±13 muscle fibres were analysed for glycogen content in diabetes and control subjects respectively.…”

Section: Methodsmentioning

confidence: 99%

Substrate source utilisation in long-term diagnosed type 2 diabetes patients at rest, and during exercise and subsequent recovery

et al. 2006

View full text Add to dashboard Cite

Aims/hypothesis Disturbances in substrate source metabolism and, more particularly, in fatty acid metabolism, play an important role in the aetiology and progression of type 2 diabetes. However, data on substrate source utilisation in type 2 diabetes are inconclusive. Methods [U-13 C]palmitate and [6,6-2 H 2 ]glucose tracers were used to assess plasma NEFA and glucose oxidation rates and to estimate the use of muscle-and/or lipoproteinderived triacylglycerol and muscle glycogen. Subjects were ten male patients who had a long-term (7±1 years) diagnosis of type 2 diabetes and were overweight, and ten matched healthy, male control subjects. Muscle biopsy samples were collected before and after exercise to assess muscle fibre type-specific intramyocellular lipid and glycogen content.

show abstract

“…where F is the infusion rate (μmol kg [29] or immunolabelled oil red O [21] to measure fiber-type-specific glycogen or intramyocellular lipid content, respectively. To determine the muscle-fiber typing, (type I vs type II), we performed a myosin adenosine 5′-triphosphatase stain [22].…”

Section: Calculationsmentioning

confidence: 99%

Carbohydrate Supplementation During Prolonged Cycling Exercise Spares Muscle Glycogen But Does Not Affect Intramyocellular Lipid Use.

Stellingwerff

Boon

Gijsen

et al. 2007

Medicine &Amp; Science in Sports &Amp; Exercise

View full text Add to dashboard Cite

Using contemporary stable-isotope methodology and fluorescence microscopy, we assessed the impact of carbohydrate supplementation on whole-body and fibertype-specific intramyocellular triacylglycerol (IMTG) and glycogen use during prolonged endurance exercise. Ten endurance-trained male subjects were studied twice during 3 h of cycling at 63±4% of maximal O 2 uptake with either glucose ingestion (CHO trial; 0.7 g CHO kg −1 h −1 ) or without (CON placebo trial; water only). Continuous infusions with [U-13 C] palmitate and [6,6-2 H 2 ] glucose were applied to quantify plasma free fatty acids (FFA) and glucose oxidation rates and to estimate intramyocellular lipid and glycogen use. Before and after exercise, muscle biopsy samples were taken to quantify fiber-type-specific IMTG and glycogen content. Plasma glucose rate of appearance (R a ) and carbohydrate oxidation rates were substantially greater in the CHO vs CON trial. Carbohydrate supplementation resulted in a lower muscle glycogen use during the first hour of exercise in the CHO vs CON trial, resulting in a 38±19 and 57±22% decreased utilization in type I and II muscle-fiber glycogen content, respectively. In the CHO trial, both plasma FFA R a and subsequent plasma FFA concentrations were lower, resulting in a 34±12% reduction in plasma FFA oxidation rates during exercise (P<0.05). Carbohydrate intake did not augment IMTG utilization, as fluorescence microscopy revealed a 76 ± 21 and 78 ± 22% reduction in type I muscle-fiber lipid content in the CHO and CON trial, respectively. We conclude that carbohydrate supplementation during prolonged cycling exercise does not modulate IMTG use but spares muscle glycogen use during the initial stages of exercise in endurance-trained men.

show abstract

A modified PAS stain combined with immunofluorescence for quantitative analyses of glycogen in muscle sections

Cited by 67 publications

References 23 publications

Starch Binding Domain-containing Protein 1/Genethonin 1 Is a Novel Participant in Glycogen Metabolism

Starch Binding Domain-containing Protein 1/Genethonin 1 Is a Novel Participant in Glycogen Metabolism

Substrate source utilisation in long-term diagnosed type 2 diabetes patients at rest, and during exercise and subsequent recovery

Carbohydrate Supplementation During Prolonged Cycling Exercise Spares Muscle Glycogen But Does Not Affect Intramyocellular Lipid Use.

Contact Info

Product

Resources

About