A Modified Enzymatic Method for Measurement of Glycogen Content in Glycogen Storage Disease Type IV
- 360 Downloads
Deficiency of glycogen branching enzyme in glycogen storage disease type IV (GSD IV) results in accumulation of less-branched and poorly soluble polysaccharides (polyglucosan bodies) in multiple tissues. Standard enzymatic method, when used to quantify glycogen content in GSD IV tissues, causes significant loss of the polysaccharides during preparation of tissue lysates. We report a modified method including an extra boiling step to dissolve the insoluble glycogen, ultimately preserving the glycogen content in tissue homogenates from GSD IV mice. Muscle tissues from wild-type, GSD II and GSD IV mice and GSD III dogs were homogenized in cold water, and homogenate of each tissue was divided into two parts. One part was immediately clarified by centrifugation at 4°C (STD-prep); the other part was boiled for 5 min then centrifuged (Boil-prep) at room temperature. When glycogen was quantified enzymatically in tissue lysates, no significant differences were found between the STD-prep and the Boil-prep for wild-type, GSD II and GSD III muscles. In contrast, glycogen content for GSD IV muscle in the STD-prep was only 11% of that in the Boil-prep, similar to wild-type values. Similar results were observed in other tissues of GSD IV mice and fibroblast cells from a GSD IV patient. This study provides important information for improving disease diagnosis, monitoring disease progression, and evaluating treatment outcomes in both clinical and preclinical clinical settings for GSD IV. This report should be used as an updated protocol in clinical diagnostic laboratories.
KeywordsGlycogen quantitation method Glycogen storage disease type IV Lafora disease Polyglucosan body
We thank Dr. Craigen and Dr. Akman of Baylor College of Medicine for sharing their new mouse model of GSD IV (Gbe1 ys/ys mice). We would also like to thank Cecelia Mizelle for helping edit the manuscript. This work was supported by the Alice and Y.T. Chen Research Center for Genetics and Genomics at Duke University.
- Chen Y-T, Kishnani PS, Koeberl DD (2009) Glycogen storage diseases. In: Valle D, Beaudet A, Vogelstein B, Kinzler K, Antonarakis S, Ballabio A (eds) Scriver’s online metabolic & molecular bases of inherited disease. McGraw-Hill, New YorkGoogle Scholar
- Raben N, Nagaraju K, Lee E, Kessler P, Byrne B, Lee L, LaMarca M, King C, Ward J, Sauer B et al (1998) Targeted disruption of the acid alpha-glucosidase gene in mice causes an illness with critical features of both infantile and adult human glycogen storage disease type II. J Biol Chem 273:19086–19092CrossRefPubMedGoogle Scholar
- Van Hove JL, Yang HW, Wu JY, Brady RO, Chen YT (1996) High-level production of recombinant human lysosomal acid alpha-glucosidase in Chinese hamster ovary cells which targets to heart muscle and corrects glycogen accumulation in fibroblasts from patients with Pompe disease. Proc Natl Acad Sci U S A 93:65–70CrossRefPubMedPubMedCentralGoogle Scholar