The effect of CPT-1 inhibition on myocardial function and resistance to ischemia/reperfusion injury in a rodent model of the metabolic syndrome

Maarman, Gerald Jerome (Stellenbosch : University of Stellenbosch, 2010-12)

Thesis

ENGLISH ABSTRACT: Background: Obesity is associated with dyslipidemia, insulin resistance and glucose intolerance and together these components characterise the metabolic syndrome (Dandona et al. 2005). In the state of obesity, there are high levels of circulating free fatty acids and increased rates of fatty oxidation which inhibit glucose oxidation. This: (i) reduce the heart‘s contractile ability, (ii) exacerbates ischemic/reperfusion injury and (iii) decreases cardiac mechanical function during reperfusion (Kantor et al. 2000; Liu et al. 2002; Taegtmeyer, 2000). Aim: The aim of our study was to investigate the effect of inhibiting fatty acid oxidation, with oxfenicine (4-Hydroxy-L-phenylglycine), on (i) cardiac mechanical function, (ii) mitochondrial respiration, (iii) myocardial tolerance to ischemia/reperfusion injury, (iv) CPT-I expression, MCAD expression, IRS-1 activation, total GLUT- 4 expression and (v) the RISK pathway (ERK42/44 and PKB/Akt). Methods: Male Wistar rats were fed a control rat chow diet or a high calorie diet (HCD) for 16 weeks. The HCD caused diet induced obesity (DIO). The animals were randomly divided into 4 groups [Control, DIO, Control + oxfen and DIO + oxfen]. The drug was administered for the last 8 weeks of feeding (200mg/kg/day). Animals were sacrificed and the hearts were perfused on the Langendorff perfusion system. After being subjected to regional ischemia and two hours of reperfusion, infarct size was determined. A separate series of animals were fed and/or treated and hearts were collected after 25 minutes global ischemia followed by 30 min reperfusion for determination of GLUT- 4, CPT-1, IRS -1, MCAD, ERK (42/44) and PKB/Akt expression/phosphorylation using Western blot analysis. A third series of hearts were excised and used for the isolation of mitochondria. Results: In the DIO rats, chronic oxfenicine treatment improved cardiac mechanical function by improving mitochondrial respiration. Oxfenicine inhibited CPT-1 expression but had no effect on MCAD or GLUT- 4 expression. Oxfenicine decreased IRS-1 iv expression, but not IRS-1 activation. Oxfenicine also improved myocardial tolerance to ischemia/reperfusion without activation of the RISK pathway (ERK & PKB). In the control rats, chronic oxfenicine treatment worsened cardiac mechanical function by adversely affecting mitochondrial respiration. Oxfenicine also worsened myocardial tolerance to ischemia/reperfusion in the control rats without changes in the RISK pathway (ERK & PKB). Oxfenicine had no effect on CPT-1, MCAD or GLUT- 4 expression. Oxfenicine increased IRS-1 expression, but not IRS-1 activity. Conclusion: Chronic oxfenicine treatment improved cardiac mechanical function and myocardial resistance to ischemia/reperfusion injury in obese animals, but worsened it in control animals. The improved cardiac mechanical function and tolerance to ischemia/reperfusion injury may be due to improvement in mitochondrial respiration.

AFRIKAANSE OPSOMMING: Agtergrond: Vetsug word geassosieer met dislipidemie, insulien weerstandigheid en glukose intoleransie, wat saam die metaboliese sindroom karakteriseer (Dandona et al. 2005). Met vetsug is daar ‗n hoë sirkulasie van vetsure, sowel as verhoogde vertsuur oksidasie wat gevolglik glukose oksidasie onderdruk. Dit: (i) verlaag die hart se vermoë om saam te trek, (ii) vererger isgemiese/herperfusie skade en (iv) verlaag kardiale effektiwiteit gedurende herperfusie (Kantor et al. 2000; Liu et al. 2002; Taegtmeyer, 2000). Doel: Die doel van die studie was om die effekte van vetsuur onderdrukking m.b.v. oksfenisien (4-Hidroksie-L-fenielglisien) op (i) meganiese hart funksie, (ii) mitokondriale respirasie, (iii) miokardiale toleransie teen isgemiese/herperfusie skade, (iv) CPT-I uitdrukking, MCAD uitdrukking, IRS-1 aktiwiteit, totale GLUT-4 uitdrukking en (v) die RISK pad (ERK42/44 en PKB/Akt) te ondersoek. Metodes: Manlike Wistar rotte was gevoer met ‗n kontrole rot dieet of ‗n hoë kalorie dieet (HKD) vir 16 weke. Die HKD lei tot dieet-geïnduseerde vetsug (DGV). Die diere was lukraak verdeel in 4 groepe [kontrole, DGV, kontrole + oksfen en DGV + oksfen]. Die behandeling met die middel was toegedien vir die laaste 8 weke van die voeding protokol (200mg/kg/dag). Die diere was geslag en die harte was geperfuseer op die Langendorff perfusie sisteem. Na blootstelling aan streeks- of globale isgemie en 2 ure herperfusie was infark groottes bepaal. ‗n Aparte reeks diere was gevoer en/of behandel en die harte was versamel na 25 minute globale isgemie gevolg deur 30 minute herperfusie vir die bepaling van GLUT-4, CPT 1, IRS -1, MCAD, ERK (42/44) en PKB/Akt uitdrukking/aktivering d.m.v. Western blot analise. ‗n Derde reeks diere was gebruik vir die isolasie van mitokondria. Resultate: In die DGV diere, het kroniese oksfenisien behandeling meganiese hart funksie verbeter d.m.v. die verbetering van mitokondriale respirasie. Oksfenisien het CPT-1 uitdrukking verlaag terwyl GLUT- 4 en MCAD uitdrukking nie geaffekteer was vi nie. Oksfenisien het IRS-1 uitdrukking verlaag, maar nie IRS-1 aktiwiteit nie. Oksfenisien het ook miokardiale weerstand teen isgemiese/herperfusie verbeter met sonder aktivering van die RISK pad (ERK & PKB). In die kontrole diere, het kroniese oksfenisien behandeling die meganiese hart funksie versleg d.m.v. negatiewe effekte op mitokondriale respirasie. Oksfenisien het die miokardiale weerstand teen isgemiese/herperfusie van die kontrole rotte versleg sonder veranderinge in die RISK pad (ERK & PKB). Oksfenisien het geen effek gehad op CPT-1, MCAD en GLUT-4 uitdrukking nie. Oksfenisien het IRS-1 uitdrukking verhoog, maar nie IRS-1 aktiwiteit nie. Samevatting: Kroniese oksfenisien behandeling het die meganiese hart funksie en miokardiale weerstand teen isgemiese/herperfusie skade in die vet diere verbeter, maar versleg in die kontrole diere. Hierdie verbetering van meganiese hart funksie en weerstand teen isgemiese/herperfusie skade kon dalk wees a.g.v. ‗n verbetering in mitokondriale respirasie.

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