Everolimus does not prevent Lafora body formation in murine Lafora disease

(A) Representative brain cortical (A.a) and skeletal muscle (A.b) sections stained with periodic acid–Schiff–diastase for detection of Lafora bodies (arrows). (B) Western blots on skeletal muscle extracts with antibodies against glycogen synthase (GS), phosphorylated GS (pGS), and glycogen phosphorylase (GP), and autophagy markers p62 and LC3. Glyceraldehyde 3-phosphate dehydrogenase (Ga) is the loading control for each blot. (C) Skeletal muscle glycogen content in millimoles per gram of tissue. (D) GS activity ratio, i.e., GS activity with no added G6P (G6P is the potent allosteric activator of GS) divided by GS activity with 7.2 mM G6P (the latter resulting in maximal activation of GS). This ratio conveys how much of total potential existing GS activity in the tissue is actually active, i.e., the enzyme's activation state (ratio). (E) Components of the above ratio (i.e., separate activities with no G6P or with 7.2 mM G6P) in nanomoles per minute per milligram of protein. (F) GP (a glycogen-degrading enzyme) activity ratio (±3 mM of the GP allosteric activator adenosine monophosphate). n = 6 for all experiments (e-Methods).