Purpose: Schizophrenia is a progressive degenerative illness that has been causally linked to mitochondrial dysfunction, oxidative stress and a pro-inflammatory state. Social isolation rearing (SIR) in rats models the neurodevelopmental aspects of schizophrenia. The antioxidant and glutamate modulator, N-acetyl cysteine (NAC), has demonstrated therapeutic potential in schizophrenia as adjunctive treatment, although this has not been tested in the SIR model. The purpose of this study was to assess whether SIR induces changes in mitochondrial function (adenosine triphosphate (ATP)), pro- vs. anti-inflammatory cytokine balance, tryptophan metabolism, a disturbance in cortico-striatal monoamines and related metabolites, and associated alterations in behaviors akin to schizophrenia, viz. social interaction, object recognition memory and prepulse inhibition (PPI). Moreover, I evaluated whether these bio-behavioral alterations could be reversed with sub-chronic clozapine, or NAC, and whether NAC may bolster the response to clozapine treatment. Methods: The objectives of the study were pursued through separately conducted studies. Male Sprague-Dawley (SD) rats (10 rats/group) were used in this study (Ethics number: NWU-0035-08-S5). Rats were randomly allocated to either social rearing or SIR for 8 weeks receiving either no treatment, vehicle, NAC (150 mg/kg/day), clozapine (5 mg/kg/day) or a combination of clozapine + NAC (CLZ + NAC) during the last 11 or 14 days of social rearing or SIR. After the 8 weeks, rats were tested for social interactive behaviors, object recognition memory and prepulse inhibition (PPI). Peripheral tryptophan metabolites (determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS)) and pro- and anti-inflammatory cytokines (IL-4, IL-6, TNF-α, IFN-γ) (enzyme-linked immunosorbent assay (ELISA)) were determined. Cortico-striatal ATP (bioluminescence assay) and monoamines (high performance liquid chromatography (HPLC)) were also determined. Results: SIR-induced significant deficits in social interactive behaviours, object recognition memory and PPI, associated with increased peripheral kynurenine, quinolinic acid (QA), and pro-inflammatory cytokines, as well as a decrease in kynurenic acid (KYNA), neuroprotective ratio and anti-inflammatory cytokines. I also observed an increase in striatal, but reduced frontal cortical ATP, dopamine, serotonin as well as their metabolites and noradrenaline’s metabolite, with noradrenaline increased in both brain regions in SIR rats. A separate dose-response study of NAC (50, 150, 250 mg/kg/day) found 150 mg/kg to be the most appropriate dose for the NAC and CLZ + NAC studies. Clozapine, NAC as well as CLZ + NAC reversed all these changes, with NAC being less effective than CLZ alone. CLZ + NAC was found to be more effective than clozapine alone in reversing certain bio-behavioral alterations induced by SIR. In addition NAC alone dose dependently reversed most of the SIR induced alterations. Conclusion: SIR induces behavioral alterations, a pro-inflammatory state, mitochondrial dysfunction and cortico-striatal monoamine alterations, closely resembling evidence in schizophrenia. Importantly, all these bio-behavioral alterations were reversed with clozapine, NAC and CLZ + NAC treatment. However, CLZ + NAC was more effective than clozapine alone in reversing some bio-behavioral alterations, supporting the therapeutic application of NAC as adjunctive treatment in schizophrenia. In addition, NAC dose dependently reversed SIR-induced cortico-striatal serotonin, noradrenaline and metabolites, emphasizing NAC’s potential use in other anxiety and stress- related disorders. / Thesis (PhD (Pharmacology))--North-West University, Potchefstroom Campus, 2013

Purpose: Schizophrenia is a progressive degenerative illness that has been causally linked to mitochondrial dysfunction, oxidative stress and a pro-inflammatory state. Social isolation rearing (SIR) in rats models the neurodevelopmental aspects of schizophrenia. The antioxidant and glutamate modulator, N-acetyl cysteine (NAC), has demonstrated therapeutic potential in schizophrenia as adjunctive treatment, although this has not been tested in the SIR model. The purpose of this study was to assess whether SIR induces changes in mitochondrial function (adenosine triphosphate (ATP)), pro- vs. anti-inflammatory cytokine balance, tryptophan metabolism, a disturbance in cortico-striatal monoamines and related metabolites, and associated alterations in behaviors akin to schizophrenia, viz. social interaction, object recognition memory and prepulse inhibition (PPI). Moreover, I evaluated whether these bio-behavioral alterations could be reversed with sub-chronic clozapine, or NAC, and whether NAC may bolster the response to clozapine treatment. Methods: The objectives of the study were pursued through separately conducted studies. Male Sprague-Dawley (SD) rats (10 rats/group) were used in this study (Ethics number: NWU-0035-08-S5). Rats were randomly allocated to either social rearing or SIR for 8 weeks receiving either no treatment, vehicle, NAC (150 mg/kg/day), clozapine (5 mg/kg/day) or a combination of clozapine + NAC (CLZ + NAC) during the last 11 or 14 days of social rearing or SIR. After the 8 weeks, rats were tested for social interactive behaviors, object recognition memory and prepulse inhibition (PPI). Peripheral tryptophan metabolites (determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS)) and pro- and anti-inflammatory cytokines (IL-4, IL-6, TNF-α, IFN-γ) (enzyme-linked immunosorbent assay (ELISA)) were determined. Cortico-striatal ATP (bioluminescence assay) and monoamines (high performance liquid chromatography (HPLC)) were also determined. Results: SIR-induced significant deficits in social interactive behaviours, object recognition memory and PPI, associated with increased peripheral kynurenine, quinolinic acid (QA), and pro-inflammatory cytokines, as well as a decrease in kynurenic acid (KYNA), neuroprotective ratio and anti-inflammatory cytokines. I also observed an increase in striatal, but reduced frontal cortical ATP, dopamine, serotonin as well as their metabolites and noradrenaline’s metabolite, with noradrenaline increased in both brain regions in SIR rats. A separate dose-response study of NAC (50, 150, 250 mg/kg/day) found 150 mg/kg to be the most appropriate dose for the NAC and CLZ + NAC studies. Clozapine, NAC as well as CLZ + NAC reversed all these changes, with NAC being less effective than CLZ alone. CLZ + NAC was found to be more effective than clozapine alone in reversing certain bio-behavioral alterations induced by SIR. In addition NAC alone dose dependently reversed most of the SIR induced alterations. Conclusion: SIR induces behavioral alterations, a pro-inflammatory state, mitochondrial dysfunction and cortico-striatal monoamine alterations, closely resembling evidence in schizophrenia. Importantly, all these bio-behavioral alterations were reversed with clozapine, NAC and CLZ + NAC treatment. However, CLZ + NAC was more effective than clozapine alone in reversing some bio-behavioral alterations, supporting the therapeutic application of NAC as adjunctive treatment in schizophrenia. In addition, NAC dose dependently reversed SIR-induced cortico-striatal serotonin, noradrenaline and metabolites, emphasizing NAC’s potential use in other anxiety and stress- related disorders. / Thesis (PhD (Pharmacology))--North-West University, Potchefstroom Campus, 2013

L’hyperglycémie associée au diabète de type 2 induit des complications vasculaires menant à des désordres cérébrovasculaires, comme l’accident vasculaire cérébral. En effet, l’hyperglycémie altère l’intégrité de la barrière hémato-encéphalique, et les cellules endothéliales cérébrales qui la composent sont particulièrement affectées. Le stress oxydant et l’état pro-inflammatoire engendrés par l’hyperglycémie jouent un rôle causal. Dans ce contexte, un intérêt croissant est accordé aux polyphénols d’origine végétale qui pourraient exercer une action antioxydante et anti-inflammatoire protectrice. L’objectif du travail de thèse était d’évaluer l’impact de l’hyperglycémie sur des marqueurs redox, inflammatoires et vasoactifs des cellules endothéliales cérébrales. L’action protectrice de l’insuline en tant qu’hormone hypoglycémiante clé a été explorée. De plus, nous avons étudié le rôle protecteur de polyphénols antioxydants extraits de la plante médicinale Antirhea borbonica de La Réunion. Pour ce faire, un modèle de cellules endothéliales cérébrales murines ainsi qu’un modèle animal d’ischémie cérébrale en condition d’hyperglycémie ont été utilisés. Nos résultats ont montré que l’hyperglycémie a induit un stress oxydant et une réponse pro-inflammatoire contribuant à une altération de la fonction endothéliale. Plusieurs cibles moléculaires ont été identifiées dont les protéines redox Nox4, Cu/ZnSOD, MnSOD, catalase et HO-1 ainsi que les facteurs vasoactifs ET-1, eNOS et NO. L’implication des médiateurs de signalisation Nrf2, AMPK, PI3K, JNK, ERK, p38 MAPK, NFκB et de la cytokine pro-inflammatoire IL-6 a été mise en évidence. L’insuline et les polyphénols ont exercé des effets antioxydants et anti- inflammatoires protégeant la fonction endothéliale. En situation d’ischémie cérébrale, l’hyperglycémie a exacerbé la dérégulation de l’état redox et pro-inflammatoire cérébral, associée à une altération de la barrière hémato-encéphalique. De plus, l’hyperglycémie a aggravé le déficit neurologique, le volume de l’infarctus cérébral et la transformation hémorragique. Les polyphénols ont exercé un rôle protecteur. L’acide caféique et son métabolite circulant, l’acide férulique, détectés au niveau cérébral, pourraient rendre compte de cette action protectrice. Des travaux complémentaires ont montré que les polyphénols protègent également contre l’altération de la fonction de cellules endothéliales aortiques humaines et la perte de vasorelaxation d’anneaux aortiques isolés de souris exposés à une hyperglycémie associée aux lipopolysaccharides de la bactérie Escherichia coli, qui sont des endotoxines liées de manière causale au contexte diabétique. En conclusion, ce travail de thèse a mis en évidence l’effet délétère de l’hyperglycémie sur la fonction endothéliale et le rôle protecteur de polyphénols antioxydants. L’utilisation des modèles expérimentaux développés permettra d’approfondir l’exploration des voies moléculaires impliquées et d’identifier de possibles cibles thérapeutiques innovantes. / Type 2 diabetes promotes vascular complications, leading to cerebrovascular disorders such as stroke. Indeed, hyperglycemia alters the blood-brain barrier integrity by deregulating the cerebral endothelial cell function. Oxidative stress and pro-inflammatory response may play a causal role. Thus, the biological effect of plant polyphenols known to exert antioxidant and anti-inflammatory capacities is of high interest. We evaluated the impact of hyperglycemia on the production of redox, inflammatory and vasoactive markers of cerebral endothelial cells, and the protective effect of polyphenols from the medicinal plant Antirhea borbonica from Reunion island. The murine bEnd.3 cerebral endothelial cells and an ischemia-reperfusion mouse model exposed to hyperglycemia were used. Our results demonstrated that hyperglycemia induced an oxidative stress and a pro-inflammatory state, leading to cerebral endothelial dysfunction through the activation of specific signaling molecules. Importantly, polyphenols extracted from Antirhea borbonica counteracted hyperglycemia deleterious effects and protected cerebral endothelial cells. Moreover, hyperglycemia exacerbated oxidative stress and pro-inflammatory state promoting cerebrovascular damages and loss of endothelial barrier integrity in ischemia-reperfusion mice model. Polyphenols exerted antioxidant and anti-inflammatory activities, attenuating cerebrovascular damages. These findings suggest that polyphenols extracted from Antirhea borbonica exerted protective effects on cerebral endothelial cells and an ischemia-reperfusion mouse model against deleterious effects of hyperglycemia.