Thesis (PhD (Biomedical Sciences. Medical Physiology))--University of Stellenbosch, 2010. / ENGLISH ABSTRACT: Background: There has been a dramatic increase in the use of dietary creatine
supplementation among sports men and women, and by clinicians as a therapeutic
agent in muscular and neurological diseases. The effects of creatine have been studied
extensively in skeletal muscle, but knowledge of its myocardial effects is limited.
Objectives: To investigate the effects of dietary creatine supplementation with and
without exercise on 1) basal cardiac function, 2) susceptibility to ischaemia/reperfusion
injury and 3) myocardial protein expression and phosphorylation and 4) mitochondrial
oxidative function.
Methods: Male Wistar rats were randomly divided into control or creatine supplemented
groups. Half of each group was exercise trained by swimming for a period of 8 weeks, 5
days per week. At the end of the 8 weeks the open field test was performed and blood
corticosterone levels were measured by RIA to determine whether the swim training
protocol had any effects on stress levels of the rats. Afterwards hearts were excised and
either freeze-clamped for biochemical and molecular analysis or perfused on the
isolated heart perfusion system to assess function and tolerance to ischaemia and
reperfusion. Five series of experiments were performed: (i) Mechanical function was
documented before and after 20 minutes global ischaemia using the work heart model,
(ii) A H2O filled balloon connected to a pressure transducer was inserted into the left
ventricle to measure LVDP and ischaemic contracture in the Langendorff model, (iii)
The left coronary artery was ligated for 35 minutes and infarct size determined after 30
minutes of reperfusion by conventional TTC staining methods. (iv) Mitochondrial
oxidative capacity was quantified. (v) High pressure liquid chromatography (HPLC) and
Western Blot analysis were performed on blood and heart tissue for determination of
high energy phosphates and protein expression and phosphorylation.
Results: Neither the behavioural studies nor the corticosterone levels showed any
evidence of stress in the groups investigated. Hearts from creatine supplemented
sedentary (33.5 ± 4.5%), creatine supplemented exercised rats (18.22 ± 6.2%) as well
as control exercised rats (26.1 ± 5.9%) had poorer aortic output recoveries than the
sedentary control group (55.9 ± 4.35% p < 0.01) and there was also greater ischaemic
contracture in the creatine supplemented exercised group compared to the sedentary
control group (10.4 ± 4.23 mmHg vs 31.63 ± 4.74 mmHg). There were no differences in
either infarct size or in mitochondrial oxygen consumption between the groups. HPLC
analysis revealed elevated phosphocreatine content (44.51 ±14.65 vs 8.19 ±4.93
nmol/gram wet weight, p < 0.05) as well as elevated ATP levels (781.1 ±58.82 vs 482.1
±75.86 nmol/gram wet weight, p<0.05) in blood from creatine supplemented vs control
sedentary rats. These high energy phosphate elevations were not evident in heart
tissue and creatine tranporter expression was not altered by creatine supplementation.
GLUT4 and phosphorylated AMPK and PKB/Akt were all significantly higher in the
creatine supplemented exercised hearts compared to the control sedentary hearts.
Conclusion: This study suggests that creatine supplementation has no effects on basal
cardiac function but reduces myocardial tolerance to ischaemia in hearts from exercise
trained animals by increasing the ischaemic contracture and decreasing reperfusion
aortic output. Exercise training alone also significantly decreased aortic output recovery.
However, the exact mechanisms for these adverse myocardial effects are unknown and
need further investigation. / AFRIKAANSE OPSOMMING: Agtergrond: Die gebruik van kreatien as dieetaanvulling het in die afgelope aantal jaar
dramaties toegeneem onder sportlui, sowel as mediese praktisyns wat dit as ‘n
terapeutiese middel vir die behandeling van spier- en neurologiese siektes aanwend.
Die effekte van kreatien op skeletspier is reeds deeglik ondersoek, maar inligting
aangaande die miokardiale effekte van die preperaat is beperk.
Doelwitte: Om die effekte van kreatien dieetaanvulling met of sonder oefening ten
opsigte van die volgende aspekte te ondersoek: 1) basislyn miokardiale funksie, 2)
vatbaarheid vir iskemie/herperfusie besering, 3) proteïenuitdrukking en -fosforilering in
die miokardium en 4) mitochondriale oksidatiewe funksie.
Metodes: Manlike Wistar rotte is ewekansig in kontrole of kreatien aanvullings groepe
verdeel. Helfte van elke groep is aan oefening in die vorm van swemsessies, vir ‘n
periode van 8 weke, 5 dae per week blootgestel. Gedrags- en biochemiese toetse is
aangewend om die moontlike effek van die swemprotokol op die rotte se stres vlakke te
bepaal. In hierdie verband is die oop area toets gebruik, asook bloed kortikosteroon
vlakke gemeet deur radioaktiewe immuunessais. Harte is daarna uit die rotte
gedissekteer en gevriesklamp vir biochemiese en molekulêre analise, of geperfuseer op
die geïsoleerde werkhart perfusiesisteem om sodoende funksie en weerstand teen
iskemie en herperfusie beskadeging te bepaal. Vyf eksperimentele reekse is uitgevoer:
(i) Meganiese funksie is noteer voor en na 20 minute globale isgemie in die werkhart
model; (ii) ‘n Water gevulde plastiek ballon, gekoppel aan ‘n druk omsetter, is in die
linker ventrikel geplaas om sodoende linker ventrikulêre ontwikkelde druk (LVDP),
asook iskemiese kontraktuur te meet; (iii) Linker koronêre arterie afbinding is vir ‘n
periode van 35 minute toegepas en die infarktgrootte bepaal na 30 minute herperfusie
deur gebruik te maak van standaard kleuringsmetodes; (iv) Mitochondriale oksidatiewe
kapasiteit is gemeet; (v) Hoë druk vloeistof chromatografie (HPLC) en Western Blot
analises is uitgevoer op bloed en hartweefsel vir die bepaling van hoë energie fosfate
(HEFe), sowel as proteïenuitdrukking en -fosforilering.
Resultate: Beide gedragsstudies en kortikosteroonvlakke het geen teken van stres in
die betrokke groepe getoon nie. Die groep blootgestel aan kreatienaanvulling en
oefening se harte het na iskemie funksioneel swakker herstel as harte van die
onaktiewe kontrole groep (18.22±6.2% vs 55.9±4.35%; p<0.01), asook ‘n groter
ikgemiese kontraktuur in vergelyking met die onaktiewe kontrole groep ontwikkel
(31.63±4.74 mmHg vs 10.4±4.23 mmHg). Daar was geen verskille in infarktgrootte of
mitochondriale suurstofverbruik tussen die verskillende groepe waargeneem nie. HPLC
analise het verhoogde fosfokreatien (44.51±14.65 vs 8.19±4.93 nmol/gram nat gewig,
p<0.05) en adenosientrifosfaat (ATP) bloedvlakke (781.1±58.82 vs 482.1±75.86
nmol/gram nat gewig, p<0.05) in kreatien aanvullings vergelyk met die kontrole groepe
getoon. Daar was egter geen meetbare veranderings in HEF vlakke in hartweefsel nie.
Gepaardgaande hiermee het kreatienaanvulling geen effek gehad op die uitdrukking va
die kreatien transporter nie. In vergelyking met onaktiewe kontrole harte was GLUT4, en
fosforileerde AMPK en PKB/ Akt beduidend hoër in harte van geoefende rotte met
kreatienaangevulling.
Gevolgtrekking: Hierdie data dui daarop dat kreatienaanvulling geen effek op basislyn
miokardiale funksie het nie. Kreatienaanvulling het egter die miokardium se weerstand
teen iskemiese skade verlaag in harte van rotte blootgestel aan oefening: iskemiese
kontraktuur is verhoog en aorta-uitset tydens herperfusie is verlaag. Die presiese
meganismes hierby betrokke is egter onbekend en vereis dus verdere studie. / Division of Medical Physiology (University of Stellenbosch), The National Research
Foundation and the Harry Crossley Fund for financial support.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/5250 |
| Date | 12 1900 |
| Creators | Webster, Ingrid |
| Contributors | Du Toit, E. F., Huisamen, B., University of Stellenbosch. Faculty of Health Sciences. Dept. of Biomedical Sciences. Medical Physiology. |
| Publisher | Stellenbosch : University of Stellenbosch |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | xxvi, 239 p. : ill. (some col.) |
| Rights | University of Stellenbosch |
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