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It is now well established that children with sickle cell disease (SCD) demonstrate cognitive deficits even in the absence of clinical stroke, but studies in children who have not experienced a stroke or other neurological event are lacking. Systemic processes that occur in SCD, like chronic inflammation and hypoxia, have been associated with hippocampal damage and episodic memory deficits in a range of clinical populations and animal models. However, studies examining episodic memory performance in children with SCD and in relation to systemic processes are largely absent. The present study addressed these gaps in young children with SCD (Mage = 7.37 years, SD = 1.51) who had not experienced a clinical stroke. Participants (N = 31) completed various memory measures as part of a larger neuropsychological protocol and participated in routine clinical blood draws. Latent class analysis (LCA) was used to empirically define SCD groups based on measures of specific visual memory processes, and results revealed two distinct visual memory groups, characterized by (1) visual memory deficits, specifically in delayed recognition abilities, and (2) intact visual memory. Follow-up analyses revealed that the two classes did not significantly differ on verbal memory performance. The relation between memory processes and both biomarkers of inflammation and adaptive functioning also were examined with variable-centered analyses. Results showed only one significant relation between C-reactive protein (CRP) and a measure of verbal delayed recognition. In sum, young children with SCD demonstrate variable episodic memory performance, with most notable deficits in visual delayed recognition. Higher levels of CRP, a biomarker of inflammation, were associated with poorer verbal delayed recognition. The results indicate that young children with SCD experience deficits in memory, even in the absence of a neurological event, and specific memory processes should be assessed in these children to guide targeted interventions. / Psychology

The genetic disorder sickle cell anemia causes hemolytic anemia and sickle pain crisis, episodes of microvascular occlusion resulting in painful ischemic tissue damage. Pain crisis is thought to occur when sickle erythrocytes adhere in the post-capillary venule, partially occluding the vessel. The resulting slowed blood flow causes more extensive cell adherence and entrapment of rigid, deoxygenated erythrocytes until the vessel is entirely occluded. It was hypothesized that the inflammatory mediators histamine and tumor necrosis factor-, factors known to cause endothelial expression of adhesive ligands, might significantly increase sickle erythrocyte adhesion, and thus be capable of initiating sickle pain crisis. It was also hypothesized that the perfusion shear stress environment of the endothelium, known to be oscillatory and reduced in sickle cell patients, was a significant mediating factor of sickle cell adhesion. An in-vitro flow chamber using cultured endothelial cells and erythrocytes from blood samples of sickle cell anemic patients was used to quantify sickle erythrocyte adherence to stimulated and unstimulated endothelial cells under shear stresses from 1.0 to 0.1 dyne/cm2. Results showed that both endothelial stimulation and reduction of the perfusion shear stress increased sickle erythrocyte adherence. In combination, the use of inflammatory stimulation with reduced shear stress resulted in further increased adhesion, but only when above the range of 0.1 V 0.2 or 0.4 dyne/cm2, depending on the inflammatory mediator. Adhesion below this level of shear is not significantly increased by endothelial stimulation. The mechanism by which histamine mediates adhesion was investigated, and found to involve the endothelial H2 and H4 receptors and expression of the P-selectin ligand. These data suggest that irregular flow, typical of sickle microvasculature, may act in conjunction with the pro-inflammatory state of sickle vasculature and the histaminergic nature of some pain treatments to initiate or propagate sickle vaso-occlusion. Findings concerning histamine, tumor necrosis factor-alpha, and shear stress effects on adherence are discussed in relation to their possible applicability to patient health, future studies are outlined to confirm the relation of in vitro data to in vivo patient condition, and proposals are made for applying these methodologies to other potential mediators of sickle erythrocyte adhesion.

Adhesion of sickle erythrocytes to vascular endothelium may initiate or propagate occlusive events in sickle cell anemia, many of which are accompanied by infection and the associated inflammatory response. Inflammatory markers are also present in sickle patients during asymptomatic periods. Inflammatory cytokines upregulate expression of endothelial adhesion molecules that promote adhesion of sickle erythrocytes. The data in this work demonstrate that after 2 hrs of stimulation with the cytokine TNF- and alpha;, E-selectin, but not VCAM-1 is upregulated on human dermal microvascular endothelial cells. After 6 hrs of TNF- and alpha; stimulation, both VCAM-1 and E-selectin expression are upregulated on MECs, and sickle erythrocytes bind to both receptors. Because strategies to control inflammation-associated adhesion in vivo may need to account for both VCAM-1 and E-selectin mediated events, control of intracellular signaling pathways leading to receptor expression is an attractive strategy for inhibiting adhesion. Cyclic AMP and nitric oxide are two intracellular signaling molecules important to cytokine-induced receptor expression. The data in this work demonstrate that TNF- and alpha; induced VCAM-1 and E-selectin expression on endothelial cells and sickle erythrocyte adhesion are abated by increasing endothelial cyclic AMP concentrations using Forskolin, IBMX, or Bt2cAMP. Conversely, when sickle erythrocytes, rather than endothelial cells, are treated with reagents that increase intracellular cAMP, adhesion to unstimulated endothelial cells is increased in some patients. Treatment of endothelial cells with reagents such as SNP and DETA-NO that increase nitric oxide significantly inhibits VCAM-1, but not E-selectin expression, induced by TNF- and alpha; stimulation and significantly inhibits sickle erythrocyte adhesion. Treatment of sickle erythrocytes directly with these reagents may also inhibit adhesion. Together these data suggest that cAMP- and nitric oxide-dependent signaling are useful therapeutic targets to inhibit cytokine-induced sickle erythrocyte adhesion to endothelium.

A Anemia Falciforme (AF) é uma doença autossômica recessiva e, dentre as hemoglobinopatias, é a mais comum das alterações hematológicas hereditárias conhecidas no homem. Sua distribuição é ampla, abrangendo todos os continentes. AF foi a primeira doença caracterizada em nível molecular. É causada por uma mutação no gene beta da globina, produzindo uma alteração estrutural na molécula. No gene da globina beta S, há a substituição de uma base nitrogenada do códon GAG para GTG, resultando na substituição do ácido glutâmico (Glu) pela valina (Val) na posição número seis da globina beta. Essa troca dos aminoácidos que resulta na HbS, altera estruturalmente a molécula e, sob determinadas condições, ocorre a polimerização, trazendo graves conseqüências ao indivíduo sintomático. As espécies reativas de oxigênio (ERO) podem causar profundas lesões em eritrócitos, diminuindo seu período de vida útil, em especial nos pacientes com AF. Acredita-se que, os eritrócitos falcizados estejam sob constante estresse oxidativo e, assim, liberem produtos de degradação da HbS, contribuindo para a progressão da doença. Dessa forma, o dano oxidativo agrava a fisiopatologia dos doentes falciformes. Utilizando técnica espectrofotométrica, foram determinadas as atividades das enzimas antioxidantes catalase (CAT), glutationa peroxidase (GPx) e superóxido dismutase (SOD) e quantificada a glutationa total (GSH) nos eritrócitos dos pacientes. Também determinou-se o dano oxidativo nas proteínas plasmáticas e no hemolisado celular pelo método do carbonil a 360 nm. Os níveis da peroxidação lipídica (MDA) e da vitamina C foram determinados por cromatografia líquida de alta performance (HPLC). Quantificou-se, por fim, os níveis de proteína C reativa ultra-sensível (CRPus) por técnica turbidimétrica. Os participantes da pesquisa (30 HbAA, 28 HbAS e 20 HbSS) foram selecionados junto ao Centro de Apoio do Portador de Anemia Falciforme (CAPAF/RS) e/ou cadastrados no Laboratório de Hematologia da Faculdade de Farmácia (UFRGS). Os doentes falciformes foram identificados por HPLC e confirmados por estudo molecular, utilizando a reação da polimerase em cadeia (PCR). Todos os indivíduos assinaram o termo de consentimento livre e esclarecido e foram submetidos a um questionário nutricional. Os dados obtidos foram expressos como médias ± desvio padrão e analisados utilizando-se o Teste ANOVA de uma via com posterior teste ad hoc. Os resultados do trabalho mostram que os indivíduos traço falciforme (HbAS) apresentam atividade significativamente elevada da CAT em relação aos indivíduos controle. Por outro lado, os doentes falciformes possuem maior atividade da GPx e SOD. O nível de GSH foi proporcionalmente maior nos HbSS seguido dos HbAS e HbAA. Observamos também, dano oxidativo em proteínas plasmáticas, mas não no hemolisado celular. Os HbSS possuem dano oxidativo em proteínas plasmáticas significativamente maior que nos demais grupos. Um aumento significativo da produção de MDA no soro dos HbSS foi observado, como um indicativo do aumento da auto-oxidação dos lipídios sob condições de estresse oxidativo. Os níveis séricos da vitamina C foram significativamente maiores nos HbSS que nos indivíduos controle. A CRPus apresentou-se significativamente elevada nos HbSS em relação aos HbAA. Esses resultados reforçam a idéia de que os pacientes com AF estão sujeitos a um estresse oxidativo crônico, o que contribui para a progressão das complicações dessa anemia hemolítica. Já, os indivíduos traço possuem elevada atividade das defesas antioxidantes capazes de reduzir o dano oxidativo em biomoléculas como proteínas e lipídios. / Sickle cell anaemia (SCA) is an autossomal recessive disease and, among the hemoglobinopaties, is the most common of the known hereditary hematologic alterations in man. Its distribution is ample, enclosing all the continents. Sickle cell anaemia (SCA) was the first disease to be characterized on the molecular level. It is basically a red blood cell (RBC) disorder in which the gene encoding the human β-globin subunit presents a mutation with the resulting replacement of β6 glutamic acid (Glu) by valine (Val). This exchange of the amino acids that results in the HbS modifies the molecule structurally, and under determined conditions, the polymeration occurs, bringing serious consequences to the symptomatic individual. The reactive oxygen species (ROS) can cause deep injuries in erythrocytes, diminishing its period of useful life, specially in patients with sickle cell anaemia. Sickled erythrocytes are under constant oxidative stress and thus liberate products of degradation of the HbS, contributing for the progression of the disease. Because of the oxidative damage aggravates the pathophysiology of sickle cell patients. Using spectrophotometrically technique, catalase (CAT), glutathione peroxidase (GPx), superoxide dismutase (SOD), were determined the activities of antioxidant enzymes and total glutathione (GSH) quantified in the erythrocytes of the patients. We also determined oxidative damage of plasma proteins and in hemolysate using carbonyl assay at 360 nm. The levels of lipid peroxidation (MDA) and vitamin C were determined by high-performance liquid chromatography (HPLC). Finally, we quantified the levels of high sensitivity Creactive protein (hsCRP) using turbidimetry technique. The participants of the research (30 HbAA, 28 HbAS and 20 HbSS) were selected from Centro de Apoio ao Portador de Anemia Falciforme (CAPAF/RS) and/or registered in cadastre in Laboratório de Hematologia da Faculdade de Farmácia (UFRGS). Sickle cell patients had been identified by HPLC and confirmed by molecular study, using the polymerase chain reaction amplification (PCR). All individuals signed the term of free and clarified assent and were submitted to a nutricional questionnaire. Data were expressed as average ±SD and analyzed using ANOVA test of one way via with the post ad hoc test. Results show that sickle cell trait subjects (HbAS) had significantly high activity of CAT when compared to healthy controls. On the other hand, sickle cell patients had greater activity of GPx and SOD. The GSH levels was proportionally higher in the followed HbSS of the HbAS and HbAA. We also observe oxidative damage in plasma proteins, but not in the cellular hemolysate. HbSS have significantly higher oxidative damage in plasma proteins than other groups. A significant increasing of the production of MDA in the serum of the HbSS was observed as an indicative of the increasing of the auto-oxidation of the lipids under oxidative estress. Serum vitamin C levels in HbSS were significantly higher than healthy control. The hsCRP was presented significantly higher in HbSS than HbAA. These results reinforce the idea that patients with SCA are subjected to chronic oxidative stress, that contributes for the progression of this hemolytic anaemia. On the other hand, sickle cell trait subjects have higher antioxidant defenses that are able to reduce oxidative damage in biological macromolecules such as proteins and lipids.

Made available in DSpace on 2014-06-11T19:32:15Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-07-01Bitstream added on 2014-06-13T18:43:20Z : No. of bitstreams: 1 shimauti_elt_dr_sjrp.pdf: 703318 bytes, checksum: bf063f504644666407e22683da45d271 (MD5) / A anemia falciforme (Hb SS), caracterizada por estados de inflamação crônica e eventos de agressão isquêmica e reperfusão pode causar danos oxidativos às macromoléculas biológicas. Os genes epistáticos e os haplótipos do gene da globina βS são possíveis moduladores de variabilidade fenotípica na Hb SS. A melatonina pode atenuar o estresse oxidativo e abrandar a morbidade. O objetivo deste foi analisar a influência de talassemia alfa (-α3.7), haplótipos do gene βS e melatonina na capacidade antioxidante total e expressão fenotípica, em indivíduos com Hb SS e Hb AS. Para avaliar o nível de melatonina sérica foram selecionadas 15 amostras de pacientes com Hb SS e 24 com Hb AA da região noroeste do estado do Paraná (NOPR). A triagem das hemoglobinas foram realizadas por métodos eletroforéticos e cromatográfico, e a caracterização genotípica da Hb S por meio de PCR- RFLP e PCR-AE. As espécies reativas de ácido tiobarbitúrico (TBARS), e a capacidade antioxidante total em equivalência ao Trolox (TEAC) foram determinadas por meio da espectrofotometria. A melatonina foi estimada por meio de HPLC. O estado redox foi analisado selecionando amostras de 68 indivíduos com Hb SS, 53 com Hb AS e 149 com Hb AA, das regiões sul e sudeste do Brasil. Para analisar a influência dos genes epistáticos sobre a TEAC, melatonina e expressão fenotípica, foram selecionadas 17 amostras com Hb SS, 30 com Hb AS, e 30 com Hb AA, da NOPR. O critério para elegebilidade foi ser não fumantes não gestantes, não etilistas e estar na fase estável para os indivíduos com HbSS. Os haplótipos do gene βS e a talassemia -α3.7 foram identificados por meio de PCR-RFLP e PCR multiplex, respectivamente. Os níveis séricos de melatonina apresentaram-se reduzidos nos indivíduos com Hb SS (P<0,001), e os TBARS e TEAC elevados quando comparados a grupo controle (P<0,001 e P<0,01 respectivamente)... / Sickle cell disease (Hb SS), characterized by chronic inflammation and ischemic damage episodes and reperfusion, comprises excessive production of free radicals which can cause oxidative damage to biologic macromolecules. The epistatic genes and the polymorphism of the βS globin gene region are possible modulators of phenotypical variability of sickle cell disease in Hb SS. Melatonin can diminish both the oxidative stress and morbidity. The objective of this study was to analyze the influence of -α-3.7 thalassemia and from βS gene haplotypes on total anti-oxidant capacity, melatonin serum levels and phenotypical expression in individuals with Hb SS and Hb AS. To evaluate melatonin serum level 15 samples with HbSS and 24 with Hb AA from the northwest region of Paraná state (NOPR). The hemoglobin screenings were performed by electrophoretic and chromatographic methods and genotypic characterization of Hb S by PCR- RFLP and PCR-AE. The concentrations of thiobarbituric acid reactive substances (TBARS), used as markers of oxidative stress and total antioxidant capacity in equivalence to Trolox (TEAC), were determined by spectrophotometry. Melatonin was estimated by HPLC. The redox state was analyzed through selecting 68 individuals with Hb SS, 53 with Hb AS and 149 with Hb AA from Brazil’s south and southeast regions. To analyze the influence of epistatic genes on TEAC, melatonin and phenotypical expression, there were selected 17 samples with Hb SS, 30 with Hb AS, and 30 with Hb AA from NOPR. The βS gene haplotypes and -α-3.7 thalassemia were identified through RFLP-PCR and multiplex PCR, respectively. The melatonin serum levels appeared significantly reduced in individuals with Hb SS (P<0.001); and the TBARS and TEAC appeared significantly increased when compared to the control group (P<0.001 and P<0.01, respectively). The correlation between TBARS and TEAC (r=0.51; P=0.04)... (Complete abstract click electronic access below)

A Anemia Falciforme (AF) é uma doença autossômica recessiva e, dentre as hemoglobinopatias, é a mais comum das alterações hematológicas hereditárias conhecidas no homem. Sua distribuição é ampla, abrangendo todos os continentes. AF foi a primeira doença caracterizada em nível molecular. É causada por uma mutação no gene beta da globina, produzindo uma alteração estrutural na molécula. No gene da globina beta S, há a substituição de uma base nitrogenada do códon GAG para GTG, resultando na substituição do ácido glutâmico (Glu) pela valina (Val) na posição número seis da globina beta. Essa troca dos aminoácidos que resulta na HbS, altera estruturalmente a molécula e, sob determinadas condições, ocorre a polimerização, trazendo graves conseqüências ao indivíduo sintomático. As espécies reativas de oxigênio (ERO) podem causar profundas lesões em eritrócitos, diminuindo seu período de vida útil, em especial nos pacientes com AF. Acredita-se que, os eritrócitos falcizados estejam sob constante estresse oxidativo e, assim, liberem produtos de degradação da HbS, contribuindo para a progressão da doença. Dessa forma, o dano oxidativo agrava a fisiopatologia dos doentes falciformes. Utilizando técnica espectrofotométrica, foram determinadas as atividades das enzimas antioxidantes catalase (CAT), glutationa peroxidase (GPx) e superóxido dismutase (SOD) e quantificada a glutationa total (GSH) nos eritrócitos dos pacientes. Também determinou-se o dano oxidativo nas proteínas plasmáticas e no hemolisado celular pelo método do carbonil a 360 nm. Os níveis da peroxidação lipídica (MDA) e da vitamina C foram determinados por cromatografia líquida de alta performance (HPLC). Quantificou-se, por fim, os níveis de proteína C reativa ultra-sensível (CRPus) por técnica turbidimétrica. Os participantes da pesquisa (30 HbAA, 28 HbAS e 20 HbSS) foram selecionados junto ao Centro de Apoio do Portador de Anemia Falciforme (CAPAF/RS) e/ou cadastrados no Laboratório de Hematologia da Faculdade de Farmácia (UFRGS). Os doentes falciformes foram identificados por HPLC e confirmados por estudo molecular, utilizando a reação da polimerase em cadeia (PCR). Todos os indivíduos assinaram o termo de consentimento livre e esclarecido e foram submetidos a um questionário nutricional. Os dados obtidos foram expressos como médias ± desvio padrão e analisados utilizando-se o Teste ANOVA de uma via com posterior teste ad hoc. Os resultados do trabalho mostram que os indivíduos traço falciforme (HbAS) apresentam atividade significativamente elevada da CAT em relação aos indivíduos controle. Por outro lado, os doentes falciformes possuem maior atividade da GPx e SOD. O nível de GSH foi proporcionalmente maior nos HbSS seguido dos HbAS e HbAA. Observamos também, dano oxidativo em proteínas plasmáticas, mas não no hemolisado celular. Os HbSS possuem dano oxidativo em proteínas plasmáticas significativamente maior que nos demais grupos. Um aumento significativo da produção de MDA no soro dos HbSS foi observado, como um indicativo do aumento da auto-oxidação dos lipídios sob condições de estresse oxidativo. Os níveis séricos da vitamina C foram significativamente maiores nos HbSS que nos indivíduos controle. A CRPus apresentou-se significativamente elevada nos HbSS em relação aos HbAA. Esses resultados reforçam a idéia de que os pacientes com AF estão sujeitos a um estresse oxidativo crônico, o que contribui para a progressão das complicações dessa anemia hemolítica. Já, os indivíduos traço possuem elevada atividade das defesas antioxidantes capazes de reduzir o dano oxidativo em biomoléculas como proteínas e lipídios. / Sickle cell anaemia (SCA) is an autossomal recessive disease and, among the hemoglobinopaties, is the most common of the known hereditary hematologic alterations in man. Its distribution is ample, enclosing all the continents. Sickle cell anaemia (SCA) was the first disease to be characterized on the molecular level. It is basically a red blood cell (RBC) disorder in which the gene encoding the human β-globin subunit presents a mutation with the resulting replacement of β6 glutamic acid (Glu) by valine (Val). This exchange of the amino acids that results in the HbS modifies the molecule structurally, and under determined conditions, the polymeration occurs, bringing serious consequences to the symptomatic individual. The reactive oxygen species (ROS) can cause deep injuries in erythrocytes, diminishing its period of useful life, specially in patients with sickle cell anaemia. Sickled erythrocytes are under constant oxidative stress and thus liberate products of degradation of the HbS, contributing for the progression of the disease. Because of the oxidative damage aggravates the pathophysiology of sickle cell patients. Using spectrophotometrically technique, catalase (CAT), glutathione peroxidase (GPx), superoxide dismutase (SOD), were determined the activities of antioxidant enzymes and total glutathione (GSH) quantified in the erythrocytes of the patients. We also determined oxidative damage of plasma proteins and in hemolysate using carbonyl assay at 360 nm. The levels of lipid peroxidation (MDA) and vitamin C were determined by high-performance liquid chromatography (HPLC). Finally, we quantified the levels of high sensitivity Creactive protein (hsCRP) using turbidimetry technique. The participants of the research (30 HbAA, 28 HbAS and 20 HbSS) were selected from Centro de Apoio ao Portador de Anemia Falciforme (CAPAF/RS) and/or registered in cadastre in Laboratório de Hematologia da Faculdade de Farmácia (UFRGS). Sickle cell patients had been identified by HPLC and confirmed by molecular study, using the polymerase chain reaction amplification (PCR). All individuals signed the term of free and clarified assent and were submitted to a nutricional questionnaire. Data were expressed as average ±SD and analyzed using ANOVA test of one way via with the post ad hoc test. Results show that sickle cell trait subjects (HbAS) had significantly high activity of CAT when compared to healthy controls. On the other hand, sickle cell patients had greater activity of GPx and SOD. The GSH levels was proportionally higher in the followed HbSS of the HbAS and HbAA. We also observe oxidative damage in plasma proteins, but not in the cellular hemolysate. HbSS have significantly higher oxidative damage in plasma proteins than other groups. A significant increasing of the production of MDA in the serum of the HbSS was observed as an indicative of the increasing of the auto-oxidation of the lipids under oxidative estress. Serum vitamin C levels in HbSS were significantly higher than healthy control. The hsCRP was presented significantly higher in HbSS than HbAA. These results reinforce the idea that patients with SCA are subjected to chronic oxidative stress, that contributes for the progression of this hemolytic anaemia. On the other hand, sickle cell trait subjects have higher antioxidant defenses that are able to reduce oxidative damage in biological macromolecules such as proteins and lipids.