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No description available.

Objetivos. Avaliar o papel do transplante de células hematopoiéticas (TCH) e da interleucina 6 (IL – 6) sobre vários parâmetros de estresse oxidativo em pacientes com Adrenoleucodistrofia ligada ao cromossomo X (X-ALD). Métodos. A concentração de malondialdeído (MDA), o conteúdo de carbolinas e sulfidrilas e a concentração de ácido hexacosanóico (C26:0) foram quantificados no plasma de pacientes X-ALD antes e após serem submetidos ao TCH. E, a concentração de MDA, a formação de carbonilas e a concentração de IL-6 foram quantificados em plasma e o conteúdo de glutationa reduzida (GSH) foi quantificado em eritrócitos de pacientes X-ALD com fenótipos cerebral infantil (cALD) ou assintomáticos no momento diagnóstico. Resultados. Observamos um aumento significativo na concentração de MDA em plasma de pacientes X-ALD antes e após o TCH em comparação ao grupo controle e uma redução significativa nesses valores após o transplante em comparação aos anteriores ao procedimento. Verificamos uma redução significativa no conteúdo de sulfidrilas no plasma de pacientes X-ALD antes do TCH em comparação ao grupo controle e um aumento significativo desses níveis após o TCH. Não observamos diferenças significativas no conteúdo de carbonilas no plasma de X-ALD antes e após o TCH, em comparação aos controles, apesar de observarmos uma redução significativa nesta determinação nos pacientes após o transplante em relação a antes do TCH. Os pacientes X-ALD apresentam níveis plasmáticos de C26:0 significativamente aumentados antes do TCH em comparação aos controles e, após o TCH, as concentrações de C26:0 foram reduzidas. Observamos uma correlação negativa significativa entre a medida do conteúdo de sulfidrilas e os níveis plasmáticos de C26:0 de indivíduos X-ALD antes do TCH. Também evidenciamos elevados níveis de MDA e da formação de carbonilas no plasma de pacientes cALD e assintomáticos em comparação ao grupo controle. Ainda, observamos redução significativa do conteúdo de GSH nos dois grupos testados comparados aos controles. A quantificação de IL-6 foi significativamente maior nos pacientes cALD, o que não foi observado nos pacientes assintomáticos, apesar destes mostrarem uma tendência de aumento da concentração de IL-6. Conclusões. Os resultados obtidos a partir do plasma de pacientes X-ALD antes e após o TCH demonstram que esta terapia, quando bem indicada e bem sucedida, tem alta efetividade em reduzir a concentração plasmática de C26:0 e é eficaz em reduzir a peroxidação lipídica e o dano oxidativo às proteínas nos pacientes X-ALD. Ainda, é possível relacionar o acúmulo de C26:0 e o dano oxidativo na patogênese da X-ALD. Nossos dados permitem sugerir que a lipoperoxidação e o dano oxidativo às proteínas possam de alguma forma estar envolvidos na fisiopatologia da X-ALD. Além disso, podemos presumir que, nos pacientes X-ALD assintomáticos estudados, o dano oxidativo e os aspectos inflamatórios desempenham papéis importantes na evolução e nas futuras manifestações do fenótipo neuronal. Também podemos supor que a administração de antioxidantes deve ser considerada como uma terapia adjuvante potencial para os pacientes assintomáticos e sintomáticos afetados pela X-ALD, inclusive para aqueles submetidos ao TCH. / Objective. We aimed to evaluate the role of hematopoietic stem cell transplantation (HSCT) and interleukin 6 (IL – 6) on various parameters of oxidative stress in X-linked adrenoleukodystrophy (X-ALD) patients. Methods. Malondialdehyde (MDA), sulfhydryl, carbonyl and hexacosanoic acid (C26:0) levels were measured in plasma from X-ALD patients before and after HSCT. And, MDA, carbonyl and IL-6 levels were measured in plasma and reduced glutathione (GSH) content was measured in erythrocytes from X-ALD patients with different phenotype (asymptomatic and childhood cerebral (CCER patients) at diagnosis moment. Results. We observed increased levels of MDA in plasma from X-ALD before and after HSCT compared to control group, but there was a significant reduction in MDA values after transplantation compared to levels found before the procedure. We verified a significant decrease in sulfhydryl content in plasma of X-ALD patients before HSCT compared with the control group and we also verified a significant increase in the levels of sulfhydryl content after HSCT. No significant differences were observed in carbonyl content in plasma of X-ALD before and after HSCT, compared to controls. However, we observed a significant reduction of plasma carbonyl content from X-ALD patients after HSCT compared to before HSCT. X-ALD patients presented a significant increase of C26:0 plasma level before HSCT when compared to controls and an important reduction of C26:0 plasma concentration in X-ALD patients after HSCT when compared to before HSCT C26:0 levels. We observed an inverse significant correlation between sulfhydryl content and plasma C26:0 levels of X-ALD individuals before HSCT. We also evidenced high levels of MDA and carbonyl formation in plasma from CCER and asymptomatic patients compared to controls. Still, we observed a significant decrease of GSH content in both groups tested compared to controls. The quantification of IL-6 is significantly higher in CCER patients, which is not observed in asymptomatic patients, despite these patients show a tendency of increased concentration of IL-6. Conclusions. The results obtained from plasma of X-ALD patients before and after HSCT demonstrate that this therapy, when well indicated and successful, has high effectiveness in reducing C26:0 plasma and is effective in reducing lipid peroxidation and oxidative damage to proteins in X-ALD patients. Still, it is possible to relate the accumulation of C26:0 and oxidative damage in the pathogenesis of X-ALD. Our data also suggest that lipid peroxidation and protein damage may somehow be involved in the pathophysiology of X-ALD. Moreover, we can assume that in our asymptomatic X-ALD patients, oxidative damage and inflammatory issues seem to play an important role in the evolution and future manifestations of neuronal phenotype. We can also assume that the administration of antioxidants should be considered as a potential adjuvant therapy for asymptomatic and symptomatic patients affected by X-ALD, including those that are submitted to HSCT.

Objetivos. Avaliar o papel do transplante de células hematopoiéticas (TCH) e da interleucina 6 (IL – 6) sobre vários parâmetros de estresse oxidativo em pacientes com Adrenoleucodistrofia ligada ao cromossomo X (X-ALD). Métodos. A concentração de malondialdeído (MDA), o conteúdo de carbolinas e sulfidrilas e a concentração de ácido hexacosanóico (C26:0) foram quantificados no plasma de pacientes X-ALD antes e após serem submetidos ao TCH. E, a concentração de MDA, a formação de carbonilas e a concentração de IL-6 foram quantificados em plasma e o conteúdo de glutationa reduzida (GSH) foi quantificado em eritrócitos de pacientes X-ALD com fenótipos cerebral infantil (cALD) ou assintomáticos no momento diagnóstico. Resultados. Observamos um aumento significativo na concentração de MDA em plasma de pacientes X-ALD antes e após o TCH em comparação ao grupo controle e uma redução significativa nesses valores após o transplante em comparação aos anteriores ao procedimento. Verificamos uma redução significativa no conteúdo de sulfidrilas no plasma de pacientes X-ALD antes do TCH em comparação ao grupo controle e um aumento significativo desses níveis após o TCH. Não observamos diferenças significativas no conteúdo de carbonilas no plasma de X-ALD antes e após o TCH, em comparação aos controles, apesar de observarmos uma redução significativa nesta determinação nos pacientes após o transplante em relação a antes do TCH. Os pacientes X-ALD apresentam níveis plasmáticos de C26:0 significativamente aumentados antes do TCH em comparação aos controles e, após o TCH, as concentrações de C26:0 foram reduzidas. Observamos uma correlação negativa significativa entre a medida do conteúdo de sulfidrilas e os níveis plasmáticos de C26:0 de indivíduos X-ALD antes do TCH. Também evidenciamos elevados níveis de MDA e da formação de carbonilas no plasma de pacientes cALD e assintomáticos em comparação ao grupo controle. Ainda, observamos redução significativa do conteúdo de GSH nos dois grupos testados comparados aos controles. A quantificação de IL-6 foi significativamente maior nos pacientes cALD, o que não foi observado nos pacientes assintomáticos, apesar destes mostrarem uma tendência de aumento da concentração de IL-6. Conclusões. Os resultados obtidos a partir do plasma de pacientes X-ALD antes e após o TCH demonstram que esta terapia, quando bem indicada e bem sucedida, tem alta efetividade em reduzir a concentração plasmática de C26:0 e é eficaz em reduzir a peroxidação lipídica e o dano oxidativo às proteínas nos pacientes X-ALD. Ainda, é possível relacionar o acúmulo de C26:0 e o dano oxidativo na patogênese da X-ALD. Nossos dados permitem sugerir que a lipoperoxidação e o dano oxidativo às proteínas possam de alguma forma estar envolvidos na fisiopatologia da X-ALD. Além disso, podemos presumir que, nos pacientes X-ALD assintomáticos estudados, o dano oxidativo e os aspectos inflamatórios desempenham papéis importantes na evolução e nas futuras manifestações do fenótipo neuronal. Também podemos supor que a administração de antioxidantes deve ser considerada como uma terapia adjuvante potencial para os pacientes assintomáticos e sintomáticos afetados pela X-ALD, inclusive para aqueles submetidos ao TCH. / Objective. We aimed to evaluate the role of hematopoietic stem cell transplantation (HSCT) and interleukin 6 (IL – 6) on various parameters of oxidative stress in X-linked adrenoleukodystrophy (X-ALD) patients. Methods. Malondialdehyde (MDA), sulfhydryl, carbonyl and hexacosanoic acid (C26:0) levels were measured in plasma from X-ALD patients before and after HSCT. And, MDA, carbonyl and IL-6 levels were measured in plasma and reduced glutathione (GSH) content was measured in erythrocytes from X-ALD patients with different phenotype (asymptomatic and childhood cerebral (CCER patients) at diagnosis moment. Results. We observed increased levels of MDA in plasma from X-ALD before and after HSCT compared to control group, but there was a significant reduction in MDA values after transplantation compared to levels found before the procedure. We verified a significant decrease in sulfhydryl content in plasma of X-ALD patients before HSCT compared with the control group and we also verified a significant increase in the levels of sulfhydryl content after HSCT. No significant differences were observed in carbonyl content in plasma of X-ALD before and after HSCT, compared to controls. However, we observed a significant reduction of plasma carbonyl content from X-ALD patients after HSCT compared to before HSCT. X-ALD patients presented a significant increase of C26:0 plasma level before HSCT when compared to controls and an important reduction of C26:0 plasma concentration in X-ALD patients after HSCT when compared to before HSCT C26:0 levels. We observed an inverse significant correlation between sulfhydryl content and plasma C26:0 levels of X-ALD individuals before HSCT. We also evidenced high levels of MDA and carbonyl formation in plasma from CCER and asymptomatic patients compared to controls. Still, we observed a significant decrease of GSH content in both groups tested compared to controls. The quantification of IL-6 is significantly higher in CCER patients, which is not observed in asymptomatic patients, despite these patients show a tendency of increased concentration of IL-6. Conclusions. The results obtained from plasma of X-ALD patients before and after HSCT demonstrate that this therapy, when well indicated and successful, has high effectiveness in reducing C26:0 plasma and is effective in reducing lipid peroxidation and oxidative damage to proteins in X-ALD patients. Still, it is possible to relate the accumulation of C26:0 and oxidative damage in the pathogenesis of X-ALD. Our data also suggest that lipid peroxidation and protein damage may somehow be involved in the pathophysiology of X-ALD. Moreover, we can assume that in our asymptomatic X-ALD patients, oxidative damage and inflammatory issues seem to play an important role in the evolution and future manifestations of neuronal phenotype. We can also assume that the administration of antioxidants should be considered as a potential adjuvant therapy for asymptomatic and symptomatic patients affected by X-ALD, including those that are submitted to HSCT.

Objetivos. Avaliar o papel do transplante de células hematopoiéticas (TCH) e da interleucina 6 (IL – 6) sobre vários parâmetros de estresse oxidativo em pacientes com Adrenoleucodistrofia ligada ao cromossomo X (X-ALD). Métodos. A concentração de malondialdeído (MDA), o conteúdo de carbolinas e sulfidrilas e a concentração de ácido hexacosanóico (C26:0) foram quantificados no plasma de pacientes X-ALD antes e após serem submetidos ao TCH. E, a concentração de MDA, a formação de carbonilas e a concentração de IL-6 foram quantificados em plasma e o conteúdo de glutationa reduzida (GSH) foi quantificado em eritrócitos de pacientes X-ALD com fenótipos cerebral infantil (cALD) ou assintomáticos no momento diagnóstico. Resultados. Observamos um aumento significativo na concentração de MDA em plasma de pacientes X-ALD antes e após o TCH em comparação ao grupo controle e uma redução significativa nesses valores após o transplante em comparação aos anteriores ao procedimento. Verificamos uma redução significativa no conteúdo de sulfidrilas no plasma de pacientes X-ALD antes do TCH em comparação ao grupo controle e um aumento significativo desses níveis após o TCH. Não observamos diferenças significativas no conteúdo de carbonilas no plasma de X-ALD antes e após o TCH, em comparação aos controles, apesar de observarmos uma redução significativa nesta determinação nos pacientes após o transplante em relação a antes do TCH. Os pacientes X-ALD apresentam níveis plasmáticos de C26:0 significativamente aumentados antes do TCH em comparação aos controles e, após o TCH, as concentrações de C26:0 foram reduzidas. Observamos uma correlação negativa significativa entre a medida do conteúdo de sulfidrilas e os níveis plasmáticos de C26:0 de indivíduos X-ALD antes do TCH. Também evidenciamos elevados níveis de MDA e da formação de carbonilas no plasma de pacientes cALD e assintomáticos em comparação ao grupo controle. Ainda, observamos redução significativa do conteúdo de GSH nos dois grupos testados comparados aos controles. A quantificação de IL-6 foi significativamente maior nos pacientes cALD, o que não foi observado nos pacientes assintomáticos, apesar destes mostrarem uma tendência de aumento da concentração de IL-6. Conclusões. Os resultados obtidos a partir do plasma de pacientes X-ALD antes e após o TCH demonstram que esta terapia, quando bem indicada e bem sucedida, tem alta efetividade em reduzir a concentração plasmática de C26:0 e é eficaz em reduzir a peroxidação lipídica e o dano oxidativo às proteínas nos pacientes X-ALD. Ainda, é possível relacionar o acúmulo de C26:0 e o dano oxidativo na patogênese da X-ALD. Nossos dados permitem sugerir que a lipoperoxidação e o dano oxidativo às proteínas possam de alguma forma estar envolvidos na fisiopatologia da X-ALD. Além disso, podemos presumir que, nos pacientes X-ALD assintomáticos estudados, o dano oxidativo e os aspectos inflamatórios desempenham papéis importantes na evolução e nas futuras manifestações do fenótipo neuronal. Também podemos supor que a administração de antioxidantes deve ser considerada como uma terapia adjuvante potencial para os pacientes assintomáticos e sintomáticos afetados pela X-ALD, inclusive para aqueles submetidos ao TCH. / Objective. We aimed to evaluate the role of hematopoietic stem cell transplantation (HSCT) and interleukin 6 (IL – 6) on various parameters of oxidative stress in X-linked adrenoleukodystrophy (X-ALD) patients. Methods. Malondialdehyde (MDA), sulfhydryl, carbonyl and hexacosanoic acid (C26:0) levels were measured in plasma from X-ALD patients before and after HSCT. And, MDA, carbonyl and IL-6 levels were measured in plasma and reduced glutathione (GSH) content was measured in erythrocytes from X-ALD patients with different phenotype (asymptomatic and childhood cerebral (CCER patients) at diagnosis moment. Results. We observed increased levels of MDA in plasma from X-ALD before and after HSCT compared to control group, but there was a significant reduction in MDA values after transplantation compared to levels found before the procedure. We verified a significant decrease in sulfhydryl content in plasma of X-ALD patients before HSCT compared with the control group and we also verified a significant increase in the levels of sulfhydryl content after HSCT. No significant differences were observed in carbonyl content in plasma of X-ALD before and after HSCT, compared to controls. However, we observed a significant reduction of plasma carbonyl content from X-ALD patients after HSCT compared to before HSCT. X-ALD patients presented a significant increase of C26:0 plasma level before HSCT when compared to controls and an important reduction of C26:0 plasma concentration in X-ALD patients after HSCT when compared to before HSCT C26:0 levels. We observed an inverse significant correlation between sulfhydryl content and plasma C26:0 levels of X-ALD individuals before HSCT. We also evidenced high levels of MDA and carbonyl formation in plasma from CCER and asymptomatic patients compared to controls. Still, we observed a significant decrease of GSH content in both groups tested compared to controls. The quantification of IL-6 is significantly higher in CCER patients, which is not observed in asymptomatic patients, despite these patients show a tendency of increased concentration of IL-6. Conclusions. The results obtained from plasma of X-ALD patients before and after HSCT demonstrate that this therapy, when well indicated and successful, has high effectiveness in reducing C26:0 plasma and is effective in reducing lipid peroxidation and oxidative damage to proteins in X-ALD patients. Still, it is possible to relate the accumulation of C26:0 and oxidative damage in the pathogenesis of X-ALD. Our data also suggest that lipid peroxidation and protein damage may somehow be involved in the pathophysiology of X-ALD. Moreover, we can assume that in our asymptomatic X-ALD patients, oxidative damage and inflammatory issues seem to play an important role in the evolution and future manifestations of neuronal phenotype. We can also assume that the administration of antioxidants should be considered as a potential adjuvant therapy for asymptomatic and symptomatic patients affected by X-ALD, including those that are submitted to HSCT.

X–linked adrenoleukodystrophy (X–ALD) is the most common peroxisomal enzyme deficiency disorder, characterized by inborn mutations in the ABCD1 gene, an ATP–binding cassette (ABC) half–transporter. The ABCD1 gene encodes the adrenoleukodystrophy protein (ALDP), the transporter for the very–long–chain fatty acids (VLCFA; C > 22:0) from the cytosol into the peroxisomes to enter the peroxisomal B–oxidation pathway. The diagnostic disease marker is the elevated levels of VLCFAs which accumulate in different tissues and body fluids, leading to inflammatory demyelination, neuro–deterioration and adrenocortical insufficiency. At present, there is no satisfactory therapy for X–ALD available. However, another peroxisomal ABC half–transporter, ALDRP can compensate for the functional loss of ALDP and is encoded by the ABCD2 gene. This prompted a new approach to treatment strategies. Phenylbutyric acid (PBA) over–expresses the ABCD2 gene, leading to an increased expression of ALDRP and PBA decreases VLCFA levels by increasing peroxisomal B–oxidation. This study had a dual aim: to determine the antioxidant capacity of PBA and to verify known and identify new metabolites of PBA. In vitro, HeLa cells were cultivated and treated with 0.5 mM, 1 mM, 2 mM and 5 mM PBA for 48 hours. The ROS, lipid peroxidation, apoptosis and cell viability were determined using fluorescein–based flow cytometry. Images were taken to visualize the peroxisome proliferation. In vivo, a vervet monkey was given a single dose of 130 mg/kg PBA. Blood was collected before treatment and 15 minutes, 30 minutes, 1, 2 and 3 hours after treatment. ROS, apoptosis and lipid peroxidation were determined by fluorescein–based flow cytometry. Urine was collected before treatment and 15 minutes, 30 minutes, 1, 2, 3, 7 and 24 hours after PBA treatment. A standardised method, employing gas chromatography–mass spectrometry (GC/MS), was used to analyse the organic acids in the urine and fatty acids in the blood. In vitro results showed decreased levels of ROS and lipid peroxidation with increased concentrations of PBA. PBA showed a protective effect towards the HeLa cells with reduced apoptosis and a high number of viable cells. In vivo levels of ROS en lipid peroxidation decreased over time of treatment with PBA. The fluorescence microscope images confirmed an increased number of peroxisomes after PBA treatment. The short term effect of PBA showed an initial, but small decrease in the levels of the fatty acids, suggesting induction over a longer period rather than activation of peroxisomal B–oxidation. New metabolites of phenylbutyrate were identified in the urine of a vervet monkey. These new metabolites originated from monooxygenase, N–phenylacetyl–glutamine synthases and B–oxidation byproducts. Recently discovered metabolites in humans and rats were also verified and confirmed in the vervet monkey. We therefore propose that treatment with PBA, on account of its beneficial effects of restoring VLCFA levels and reducing oxidative stress, could be considered a novel approach for the treatment of X–ALD. / Thesis (M.Sc. (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2011.

X–linked adrenoleukodystrophy (X–ALD) is the most common peroxisomal enzyme deficiency disorder, characterized by inborn mutations in the ABCD1 gene, an ATP–binding cassette (ABC) half–transporter. The ABCD1 gene encodes the adrenoleukodystrophy protein (ALDP), the transporter for the very–long–chain fatty acids (VLCFA; C > 22:0) from the cytosol into the peroxisomes to enter the peroxisomal B–oxidation pathway. The diagnostic disease marker is the elevated levels of VLCFAs which accumulate in different tissues and body fluids, leading to inflammatory demyelination, neuro–deterioration and adrenocortical insufficiency. At present, there is no satisfactory therapy for X–ALD available. However, another peroxisomal ABC half–transporter, ALDRP can compensate for the functional loss of ALDP and is encoded by the ABCD2 gene. This prompted a new approach to treatment strategies. Phenylbutyric acid (PBA) over–expresses the ABCD2 gene, leading to an increased expression of ALDRP and PBA decreases VLCFA levels by increasing peroxisomal B–oxidation. This study had a dual aim: to determine the antioxidant capacity of PBA and to verify known and identify new metabolites of PBA. In vitro, HeLa cells were cultivated and treated with 0.5 mM, 1 mM, 2 mM and 5 mM PBA for 48 hours. The ROS, lipid peroxidation, apoptosis and cell viability were determined using fluorescein–based flow cytometry. Images were taken to visualize the peroxisome proliferation. In vivo, a vervet monkey was given a single dose of 130 mg/kg PBA. Blood was collected before treatment and 15 minutes, 30 minutes, 1, 2 and 3 hours after treatment. ROS, apoptosis and lipid peroxidation were determined by fluorescein–based flow cytometry. Urine was collected before treatment and 15 minutes, 30 minutes, 1, 2, 3, 7 and 24 hours after PBA treatment. A standardised method, employing gas chromatography–mass spectrometry (GC/MS), was used to analyse the organic acids in the urine and fatty acids in the blood. In vitro results showed decreased levels of ROS and lipid peroxidation with increased concentrations of PBA. PBA showed a protective effect towards the HeLa cells with reduced apoptosis and a high number of viable cells. In vivo levels of ROS en lipid peroxidation decreased over time of treatment with PBA. The fluorescence microscope images confirmed an increased number of peroxisomes after PBA treatment. The short term effect of PBA showed an initial, but small decrease in the levels of the fatty acids, suggesting induction over a longer period rather than activation of peroxisomal B–oxidation. New metabolites of phenylbutyrate were identified in the urine of a vervet monkey. These new metabolites originated from monooxygenase, N–phenylacetyl–glutamine synthases and B–oxidation byproducts. Recently discovered metabolites in humans and rats were also verified and confirmed in the vervet monkey. We therefore propose that treatment with PBA, on account of its beneficial effects of restoring VLCFA levels and reducing oxidative stress, could be considered a novel approach for the treatment of X–ALD. / Thesis (M.Sc. (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2011.

No description available.