A Metabolomic Signature of Ischemic Stroke Showing Acute Oxidative and Energetic Stress.

Metabolomics is a powerful data-driven tool for in-depth biological phenotyping that could help identify the specific metabolic profile of cryptogenic strokes, for which no precise cause has been identified. We performed a targeted quantitative metabolomics study in West African patients who had recently suffered an ischemic stroke, which was either cryptogenic (n = 40) or had a clearly identified cause (n = 39), compared to a healthy control group (n = 40). Four hundred fifty-six metabolites were accurately measured. Multivariate analyses failed to reveal any metabolic profile discriminating between cryptogenic ischemic strokes and those with an identified cause but did show superimposable metabolic profiles in both groups, which were clearly distinct from those of healthy controls. The blood concentrations of 234 metabolites were significantly affected in stroke patients compared to controls after the Benjamini-Hochberg correction. Increased methionine sulfoxide and homocysteine concentrations, as well as an overall increase in saturation of fatty acids, were indicative of acute oxidative stress. This signature also showed alterations in energetic metabolism, cell membrane integrity, monocarbon metabolism, and neurotransmission, with reduced concentrations of several metabolites known to be neuroprotective. Overall, our results show that cryptogenic strokes are not pathophysiologically distinct from ischemic strokes of established origin, and that stroke leads to intense metabolic remodeling with marked oxidative and energetic stresses.

Biomarkers of sickle cell nephropathy in Senegal.

Sickle cell anemia (SCA) is caused by a single point variation in the ß-globin gene (HBB): c.20A> T (p.Glu7Val), in homozygous state. SCA is characterized by sickling of red blood cells in small blood vessels which leads to a range of multiorgan complications, including kidney dysfunction. This case-control study aims at identifying sickle cell nephropathy biomarkers in a group of patients living with SCA from Senegal. A total of 163 patients living with SCA and 177 ethnic matched controls were investigated. Biological phenotyping included evaluation of glycemia, glucosuria, albuminuria, proteinuria, tubular proteinuria, serum creatinine, urine creatinine, urine specific gravity and glomerular filtration rate. Descriptive statistics of biomarkers were performed using the χ2 -test, with the significance level set at p<0.05. Patients living with SCA had a median age of 20 years (range 4 to 57) with a female sex frequency of 53.21%. The median age of the control participants was 29 years (range: 4-77) with a female sex frequency of 66.09%. The following proportions of abnormal biological indices were observed in SCA patients versus (vs.) controls, as follows: hyposthenuria: 35.3%vs.5.2% (p<0.001); glomerular hyperfiltration: 47.66%vs.19.75% (p<0.001), renal insufficiency: 5.47%vs.3.82% (p = 0.182); microalbuminuria: 42.38%vs.5.78% (p<0.001); proteinuria: 39.33%vs.4.62% (p<0.001); tubular proteinuria: 40.97%vs.4.73% (p<0.001) and microglucosuria: 22.5%vs.5.1% (p<0.001). This study shows a relatively high proportion of SCA nephropathy among patients living with SCA in Senegal. Microglucosuria, proteinuria, tubular proteinuria, microalbuminuria, hyposthenuria and glomerular hyperfiltration are the most prevalent biomarkers of nephropathy in this group of Senegalese patients with SCA.

Effects of Senegal haplotype (Xmn1-rs7412844), alpha-thalassemia (3.7kb HBA1/HBA2 deletion), NPRL3-rs11248850 and BCL11A-rs4671393 variants on sickle cell nephropathy.

OBJECTIVE: Sickle cell anemia (SCA) can cause substantial kidney dysfunction resulting in sickle cell nephropathy, which may be affected by the presence of modifier genes. This study evaluates the effects of some modifier genes on sickle cell nephropathy. METHODS: Patients living with SCA were recruited. Alpha-thalassemia (3.7kb HBA1/HBA2 deletion) was genotyped using gap PCR multiplex. Senegal haplotype (Xmn1-rs7412844), BCL11A-rs4671393 and NPRL3-rs11248850 were genotyped using Mass Array. The effects of variants on kidney dysfunction were then evaluated using multivariate analysis. RESULTS: The number of patients living with SCA included in this study was 162 with a median age of 20 years [minimum-maximum: 4-57] and a female frequency of 53.21%. Senegal haplotype, BCL11A-rs4671393 variant were protective factors against albuminuria stage A2 with an odds ratio (OR) of 0.22 (95% CI 0.05-0.90) and 0.27 (95% CI 0.08-0.96) respectively. The combination NPRL3-rs11248850 variant - 3.7kb HBA1/HBA2 deletion was a protective factor against albuminuria stage A2 (OR = 0.087, 95% Cl 0.01-0.78) but it was a risk factor for glomerular hyperfiltration (OR = 17.69, 95% CI 1.85-169.31). CONCLUSIONS: All four variants displayed a protective effect against albuminuria stage A2. The combination alpha-thalassemia - NPRL3-rs11248850 variant is a risk factor for glomerular hyperfiltration.

Influence of Oxidative Stress Biomarkers and Genetic Polymorphisms on the Clinical Severity of Hydroxyurea-Free Senegalese Children with Sickle Cell Anemia.

Oxidative stress would play a role in the pathophysiology of sickle cell anemia (SCA). We tested the impact of common SCA genetic modifiers (alpha-thalassemia, G6PD deficiency, HbF quantitative trait loci; QTL) and pro/antioxidant genes polymorphisms (SOD2 rs4880, XO rs207454, MPO rs233322) on oxidative stress biomarkers (AOPP, MDA, MPO, XO, MnSOD, CAT, GPx) and clinical severity in 301 Senegalese SCA hydroxyurea-free children at steady-state (median age 9.1 years, sex ratio H/F = 1.3). Plasma oxidative stress biomarkers were compared with those of a control group (AA). CAT activity, AOPP, and MDA levels were higher in SCA than in AA individuals while XO, GPX, and MnSOD activities were lower. The presence of alpha-thalassemia decreased MDA level and MPO activity but no effect of the HbF QTL or G6PD deficiency was observed. SCA children who experienced their first hospitalized complication before 3 years old had higher MnSOD and CAT activities than the other children while those with no hospitalized VOC in the previous 2 years presented higher GPX activity. Age of the first hospitalized complication and AOPP levels were affected by the MPO rs2333227 SNP. Our results suggest that alpha-thalassemia modulates oxidative stress in SCA, presumably because of a reduction in the MPO activity.

Combined and differential effects of alpha-thalassemia and HbF-quantitative trait loci in Senegalese hydroxyurea-free children with sickle cell anemia.

BACKGROUND: Our objective was to investigate the combined and differential effects of alpha-thalassemia -3.7 kb deletion and HbF-promoting quantitative trait loci (HbF-QTL) in Senegalese hydroxyurea (HU)-free children and young adults with sickle cell anemia (SCA). PROCEDURE: Steady-state biological parameters and vaso-occlusive crises (VOC) requiring emergency admission were recorded over a 2-year period in 301 children with SCA. The age of the first hospitalized VOC was also recorded. These data were correlated with the alpha-globin and HbF-QTL genotypes. For the latter, three different genetic loci were studied (XmnI, rs7482144; BCL11A, rs1427407; and the HBS1L-MYB region, rs28384513) and a composite score was calculated, ranging from zero (none of these three polymorphisms) to six (all three polymorphisms at the homozygous state). RESULTS: A positive clinical impact of the HbF-QTL score on VOC rate, HbF, leucocytes, and C-reactive protein levels was observed only for patients without alpha-thalassemia deletion. Conversely, combination of homozygous -3.7 kb deletion with three to six HbF-QTL was associated with a higher VOC rate. The age of the first hospitalized VOC was delayed for patients with one or two alpha-thalassemia deletions and at least two HbF-QTL. CONCLUSION: Alpha-thalassemia -3.7 kb deletion and HbF-QTL are modulating factors of SCA clinical severity that interact with each other. They should be studied and interpreted together and not separately, at least in HU-free children.

Genetic Background of the Sickle Cell Disease Pediatric Population of Dakar, Senegal, and Characterization of a Novel Frameshift ß-Thalassemia Mutation [HBB: c.265_266del; p.Leu89Glufs*2].

Sickle cell disease is a genetic disorder with a large variability in the pattern and severity of clinical manifestations. Different genetic modulators have been identified but very few epidemiologic data are available on these modifier genes in Senegal. This study aimed to determine their prevalence in a Senegalese sickle cell disease pediatric population. The following genetic parameters were genotyped in 295 sickle cell disease children of the Dakar pediatric hospital: sickle cell disease genotype [ßS/ßS (HBB: c.20A>T), ßS/ßC (HBB: c.19G>A), ßS/ß0-thalassemia (ß0-thal)], XmnI polymorphism, the five most common α-thalassemia (α-thal) deletions and the A(-) and Betica glucose-6-phosphate-dehydrogenase (G6PD) deficient variants. Despite very few ßS/ßC and ßS/ß0-thal children (1.0% each), a novel frameshift ß0-thal mutation was characterized: HBB: c.265_266del; p.Leu89Glufs*2. The -α3.7 (rightward) deletion was the only α-thal deletion identified in this cohort (12.0% allelic frequency). Most of ßS/ßS patients (61.9%) were homozygous for the XmnI polymorphism and assumed to carry a Senegal/Senegal ßS haplotype. The remaining haplotypes were predominantly of the Benin type. While the Betica G6PD variant was quite frequent (13.0%), a low frequency of the A(-) variant was detected (1.0-2.0%). The systematic genotyping of the -α3.7 deletion and of the G6PD Betica variant in sickle cell disease patients from Senegal could be useful to identify patients at risk for several complications, such as cerebral vasculopathy, where it has been demonstrated that a normal α-globin genotype and G6PD deficiency are predisposing factors. These patients should be eligible for a transcranial Doppler examination that is not routinely offered in Senegal.

Analysis of topoisomerase mutations in fluoroquinolone-resistant and -susceptible Campylobacter jejuni strains isolated in Senegal.

In this study, topoisomerase mutations in ciprofloxacin-resistant and -susceptible Campylobacter jejuni were analysed by DNA sequencing. In certain ciprofloxacin-resistant C. jejuni, the mechanism of resistance was complex. The Thr86-Ala substitution in the GyrA protein appears to play a role in increasing the minimum inhibitory concentration of nalidixic acid only. In addition, isolates with this amino acid change and those resistant to quinolones but lacking a mutation in the GyrA quinolone resistance-determining region could be derived from two different clones. Based on gyrA and gyrB polymorphisms, C. jejuni isolates from the Dakar region of Senegal appeared to be less diverse than those from other countries. Moreover, C. jejuni isolates in Senegal appeared to differ from European isolates by lack of a silent mutation at codon 120 of the gyrA gene.

Campylobacter coli isolates derived from chickens in Senegal: Diversity, genetic exchange with Campylobacter jejuni and quinolone resistance.

We used the multilocus sequence typing (MLST) method to study the genetic diversity of Campylobacter coli isolated from chickens in Senegal, and to check the presence of genetic exchange with Campylobacter jejuni. In addition, we assessed the resistance of the isolates to ciprofloxacin and nalidixic acid, and their gyrA sequences. MLST revealed a low level of diversity and the absence of lineages among C. coli isolates. In addition, an exchange of alleles with C. jejuni was found. Twenty percent of the ciprofloxacin-resistant isolates lacked mutations within the quinolone resistance-determining region (QRDR) of GyrA. There was no link between quinolone resistance and sequence type (ST).

Genetic diversity and quinolone resistance in Campylobacter jejuni isolates from poultry in Senegal.

We used the multilocus sequence typing (MLST) method to evaluate the genetic diversity of 46 Campylobacter jejuni isolates from chickens and to determine the link between quinolone resistance and sequence type (ST). There were a total of 16 ST genotypes, and the majority of them belonged to seven clonal complexes previously identified by using isolates from human disease. The ST-353 complex was the most common complex, whereas the ST-21, ST-42, ST-52, and ST-257 complexes were less well represented. The resistance phenotype varied for each ST, and the Thr-86-Ile substitution in the GyrA protein was the predominant mechanism of resistance to quinolone. Nine of the 14 isolates having the Thr-86-Ile substitution belonged to the ST-353 complex. MLST showed that the emergence of quinolone resistance is not related to the diffusion of a unique clone and that there is no link between ST genotype and quinolone resistance. Based on silent mutations, different variants of the gyrA gene were shown to exist for the same ST. These data provide useful information for understanding the epidemiology of C. jejuni in Senegal.

Fluoroquinolone susceptibility of Campylobacter strains, Senegal.

To assess fluoroquinolone susceptibility of Campylobacter strains in Senegal, skin samples were collected from 250 chicken carcasses from January 2001 to October 2002. Among 205 isolated Campylobacter strains, 59% and 41% were identified as Campylobacter jejuni and C. coli, respectively; the overall ciprofloxacin-resistance rate was 34%.