Citation for published version (APA): Oosterveer, M. H. (2009). Control of metabolic flux by nutrient sensors Groningen: s.n.
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1 University of Groningen Control of metabolic flux by nutrient sensors Oosterveer, Maaike IMPORTANT NOTE: You are advised to consult the publisher's version (publisher's PDF) if you wish to cite from it. Please check the document version below. Document Version Publisher's PDF, also known as Version of record Publication date: 2009 Link to publication in University of Groningen/UMCG research database Citation for published version (APA): Oosterveer, M. H. (2009). Control of metabolic flux by nutrient sensors Groningen: s.n. Copyright Other than for strictly personal use, it is not permitted to download or to forward/distribute the text or part of it without the consent of the author(s) and/or copyright holder(s), unless the work is under an open content license (like Creative Commons). Take-down policy If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim. Downloaded from the University of Groningen/UMCG research database (Pure): For technical reasons the number of authors shown on this cover page is limited to 10 maximum. Download date:
2 Supplemental Figure 1 Schematic model of hepatic carbohydrate metabolism [U - 13 C]-glucose [1-2 H]-galactose 4 2 glucose glucose-6-phosphate glycogen blood sample glucose 3 5 UDP-glucose [2-13 C]-glycerol 1 paracetamol triose phosphate urine sample paracetamol glucuronic acid Major metabolic pathways and enzymatic reactions are depicted, sharing G6P as a central metabolite. The pathways included are: (1) Gluconeogenic flux toward G6P, (2) Glycogen phosphorylase flux, (3) Glucose-6-phosphatase flux, (4) Glucokinase flux and (5) Glycogen synthase flux. Mice received an infusion containing [U- 13 C]glucose, [2-13 C]glycerol, [1-2 H]galactose and paracetamol for six hours. MIDA was applied on blood glucose and urinary paracetamol glucuronide samples. Supplemental Material 123
3
4 Supplemental Table 1 Primer and probe sequences used for qpcr Gene Sense Antisense Probe Accession number Acc1 CCA TCC AAA CAG AGG GAA CAT C CTA CAT GAG TCA TGC CAT AGT GGT T Acc2 CCC AGG AGG CTG CAT TGA AGA CAT GCT GGG CCT CAT AGT A ACG CTA AAC AGA ATG TCC TTT GCC TCC AAC CAC AAG TGA TCC TGA ATC TCA CGC GC NM_ NM_ Acs GGA GCT TCG CAG TGG CAT C CCC AGG CTC GAC TGT ATC TTG T Aox GCC ACG GAA CTC ATC TTC GA CCA GGC CAC CAC TTA ATG GA Elovl5 TGG CTG TTC TTC CAG ATT GGA CCC TTT CTT GTT GTA AGT CTG AAT GTA Elovl6 ACA CGT AGC GAC TCC GAA GAT AGC GCA GAA AAC AGG AAA GAC T Fads1 CCT TCG CGG ACA TTG TTT ACT C TAT GGA GGT CTG CTG CTG CTA T Fads2 CCC TGA TCG ACA TTG TGA GTT C GAC GGC AGC TTC ATT TAT GGA Fgf-21 CCG CAG TCC AGA AAG TCT CC TGA CAC CCA GGA TTT GAA TGA C G6pdh GCA ACA GAT ACA AGA ATG TGA AGC T AGG CTT CCC TGA GTT CAT CAC T CAG AAA CAA CAG CCT GTG GGA TAA ACT CAT CTT CCA CTG CCA CAT ATG ACC CCA AGA CCC CAT GAT TTC CCT GAT TGC TCT CTT CAC AAA C TTT CCT GCA TCC ATT GGA TGG CTT C CTC TGG TTG GAC GCT TAC CTT CAC CA CCA GCC ACA GCT CCC CAG ACT TCT CCT GGC TTC AAG GCT TTG AGC TCC A CCT ATG AAC GCC TCA TCC TGG ATG TCT T ACC GCT CCA TTG TGA CAG CTG ACA NM_ NM_ NM_ NM_ NM_ NM_ NM_ NM_ Gyk GGG TTG GTG TGT GGA GTC TTG GAT TTC GCT TTC TTC AGC ATT GA NM_ Hmgcs1 CGA TGG TGT AGA TGC TGG AAA G CAT CAG TTT CTG AAC CAC AGT CGA TCC GTG CAG AAG CCC ATC C NM_ Lcad TAC GGC ACA AAA GAA CAG ATC G CAG GCT CTG TCA TGG CTA TGG CAC TTG CCC GCC GTC ATC TGG NM_ Me1 AGG CAG CGT CTT CCA AAT ATG TCG ATA CTT GTT CAG GAG ACG AA TGG CAA AAT CTT CAA ACT GAA TAA GGC AAT TC CTT TAT GCT GCT CAG ACA GGC AGC CAC NM_ Pdgh GGA CAT CCG TAA GGC CCT CTA T ATT GAG GGT CCA GCC AAA CTC NM_ Pgc-1β GAG ACA CAG ATG AAG ATC CAA GCT CTT GCC AAG AGA GTC GCT TTG T CCA GGT GCC TCA TGC TGG CCT NM_ Sglt1 GTT GGA GTC TAC GCA ACA GCA A GGG CTT CTG TGT CTA TTT CAA TTG T TCC TCC TCT CCT GCA TCC AGG TCG NM_ Taldo1 CAG AAG TTG ATG CAA GGC TTT C CCA GCT TCT TTG TAA AGC TCG A CTC GGG CCA CCA TGG CAT CC NM_ Tkt GCA TCC TGT CCC GAA ACA AG CAA TAG ACT CGG TAG CTG GCT TT CCC TGG CCC AGG GAG CCA GT NM_ Supplemental Material 125
5 Supplemental Table 2 Parameters and equations used to calculate hepatic glucose metabolism Parameter Equation Primary isotopic parameters 1. d(glc) (glc)blood/ (glc)infusate 2. d(udpglc) M 1 (pglcua)urine/m 1 (gal)infusate 3. c(glc) (pglcua)urine/ (glc)blood 4. c(udpglc) M 1 (glc)blood/m 1 (pglcua)urine 5. f(glc) M 2 (glc)blood/m 2 (FBP:MIDA)glc 6. f(udpglc) M 2 (pglcua)urine/m 2 (FBP:MIDA)pGlcUA Whole body glucose metabolism 7. Ra(glc;whole body) Inf(glc;total)/d(glc) 8. Ra(UDPglc;whole body) Inf(gal;total)/d(UDPglc) 9. MCR(glc) Ra(glc;whole body)/glc conc 10. Ra(glc;endo) Ra(glc;whole body) - Inf(gal;total) 11. Ra(UDPglc;endo) Ra(UDPglc;whole body) - Inf(gal;total) 12. Rr(glc) {c(glc)/[1-c(glc)]}/ra(glc;endo) 13. Rr(UDPglc) {c(udpglc)/[1-c(udpglc)]}/ra(udpglc;endo) 14. Total Ra(glc;endo) Ra(glc;endo) + Rr(glc) 15. Total Ra(UDPglc;endo) Ra(UDPglc;endo) + Rr(UDPglc) 16. UDPglc(glc) c(udpglc) x [Ra(glc;endo) + Inf(glc;total)] 17. glc(udpglc) c(glc) x [Ra(UDPglc;endo) + Inf(gal;total)] 18. GNG(glc) f(glc) x [Ra(glc;whole body) + Rr(glc)] 19. GNG(UDPglc) f(udpglc) x [Ra(UDPglc;whole body) + Rr(UDPglc)] 20. GNG(glc;indirect) [f(udpglc) x UDPglc(glc)] + [f(glc] x Rr(glc)] 21. GNG(UDPglc;indirect) [f(glc) x glc(udpglc)] + [f(udpglc) x Rr(UDPglc)] 22. GNG(glc;direct) GNG(glc) - GNG(glc;indirect) 23. GNG(UDPglc;direct) GNG(UDPglc) - GNG(UDPglc;indirect) 24. GLY(glc) Ra(glc;endo) - GNG(glc;direct) - [f(udpglc) x UDPglc(glc)] 25. GLY(UDPglc) Ra(UDPglc;endo) - GNG(UDPglc;direct) - glc(udpglc) Individual fluxes comprising hepatic G6P metabolism 26. GNG(G6P) GNG(glc;direct) + GNG(UDPglc;direct) 27. GK glc(udpglc) + Rr(glc) 28. G6Pase GNG(glc) + GLY(glc) 29. GS GNG(UDPglc) + GLY(UDPglc) 30. GP GLY(UDPglc) + GLY(glc) + {[ 1- c(glc)] x Rr(UDPglc)} 126 Supplemental Material
6 d(glc) (glc)infusate fractional contribution of infused glucose to blood glucose mole percent enrichments (MPE) of [U- 13 C]glucose in the infusate glc conc Ra(glc;endo) blood glucose concentration in mm rate of endogenous blood glucose appearance, not corrected for recycling of tracer (glc)blood MPE of [U- 13 C]glucose in blood Ra(UDPglc;endo) rate of endogenous UDP glucose appearance, not corrected for recycling of tracer d(udpglc) fractional contribution of infused galactose Rr(glc) rate of recycling of glucose tracer to UDPglucose M 1 (gal)infusate MPE of [1-2 H]galactose in the infusate Rr(UDPglc) rate of recycling of UDPglc tracer M 1 (pglcua)urine MPE of [1-2 H]-UDPglucose measured in urinary Par-GlcUA totalra(glc;endo) total endogenous glucose production, including recycling of tracer c(glc) (pglcua)urine c(udpglc) fractional contribution of blood glucose to UDP-glucose formation MPE of [U- 13 C]- UDPglucose measured in urinary Par-GlcUA fractional contribution of UDPglucose to blood glucose formation totalra(udpglc) UDPglc(glc) glc(udpglc) total endogenous UDPglucose production, including recycling of tracer rate of UDPglucose conversion into blood glucose rate of blood glucose conversion into UDPglucose M 1 (glc)blood MPE of [1-2 H]- glucose in blood GNG(glc) rate of gluconeogenesis into blood glucose (pglcua)urine f(glc) MPE of [U- 13 C]-UDPglucose measured in urinary Par-GlcUA fractional contribution of newly synthesized glucose to blood glucose GNG(UDPglc) GNG(glc;indirect) rate of gluconeogenesis into UDPglucose rate of gluconeogenesis into blood glucose indirectly via glycogen M 2 (glc)blood MPE of [ 13 C 2 ]-glucose in blood GNG(UDPglc;indirect) rate of gluconeogenesis into UDPglucose indirectly via blood glucose M 2 (FBP:MIDA)glc theoretical MPE of [ 13 C 2 ]-Fructose 1,6 biphosphate, calculated by MIDA using 13 C-enrichment data of glucose f(udpglc) fractional contribution of newly synthesized glucose to UDPglc pool M 2 (pglcua)urine MPE of [ 13 C 2 ]-UDPglc, sampled as urinary Par-GlcUA M 2 (FBP:MIDA)pGlcUA theoretical MPE of [ 13 C 2 ]-Fructose 1,6 biphosphate, calculated by MIDA using 13 C enrichment data of urinary Par-GlcUA Ra(glc;whole body) Inf(glc) Ra(UDPglc; whole body) Inf(gal) whole body rate of appearance of glucose into the blood glucose pool rate of infusion of [U- 13 C]glucose in μmol/kg/min whole body rate of appearance of UDPglc rate of infusion of [1-2 H]galactose in μmol/kg/min GNG(glc;direct) GNG(UDPglc;direct) GLY(glc) GLY (UDPglc) GNG(G6P) GK G6Pase GS rate of gluconeogenesis directly into blood glucose rate of gluconeogenesis directly into UDPglucose rate of glycogenolysis contributing to blood glucose formation rate of glycogenolysis contributing to UDPglucose formation total flux of G6P de novo synthesis, corrected for the exchange between blood glucose and UDPglucose pools glucokinase flux glucose-6-phosphatase flux glycogen synthase flux MCR(glc) metabolic clearance rate of blood glucose GP glycogen phosphorylase flux Supplemental Material 127
7 Supplemental Table 3 Formulas used to calculate the concentration vs. time curves and the kinetic parameters in a first order absorption process in an one-compartment model Parameter Equation Oral [U- 13 C]-glucose blood concentration C t = x [glc] [U- 13 C]-glucose in blood over time C t = C(0) el x e -k(el) x t C(0) ab x e -k(ab) x t Lag time t lag = (ln(c(0) ab ) ln(c(0) el ))/(k ab k el ) [U- 13 C]-glucose blood concentration at tlag C lag = C(0) el x e -k(el) x t(lag) = C(0) ab -k(ab) x t(lag) x e Curve of absorption and elimination C t = C lag x (e -k(el) x (t-t(lag)) - e -k(ab) x (t-t(lag)) ) Time were curve reaches its maximum t max = 1/(k ab k el ) x ln(k ab /k el ) [U- 13 C]-glucose blood concentration at tmax C max = C lag x (e -k(el) x t(max) e -k(ab) x t(max) ) Half life t ½ = ln(2)/k el Mean residence time MRT = 1/k ab + 1/k el Bioavailability F = 1 ((C(0) ab x k el )/(k ab x C(0) el )) Volume of distribution V D = F x D L / C lag C(0) ab initial concentration by extrapolation of the absorption period C(0) el concentration by extrapolation of the elimination period C lag concentration at lag time calculated from elimination or absorption curve C max concentration at t max D L oral dose administrated F fractional contribution to the sampled compartment [glc] total blood glucose concentration k ab absorption rate constant k el elimination rate constant mole percent enrichment of blood glucose MRT mean residence time in sampled compartment t(½) el half-life t lag time between administration and appearance in sampled compartment t max time of maximal concentration V D appearant volume of distribution 128 Supplemental Material
8 Supplemental Table 4 Fatty acid composition of the experimental diets chow high-fat high-fat/fish oil C14: C16: C16: C18: C18: C18: C18: C Values are given in g/kg. Supplemental Material 129
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