Large Cohort Screening of G6PD Deficiency and the Mutational Spectrum in the Dongguan District in Southern China

Background

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is a common enzymatic disorder of the erythrocytes that affects 400 million people worldwide. We developed a PCR-reverse dot blot (RDB) assay to screen twenty genotypes of seventeen Chinese G6PD mutations and investigate the spectrum of G6PD deficiency mutations in Dongguan District, Guangdong Province, in southern China.

Method

The PCR-RDB assay consists of multiplex PCR amplification of seven fragments in the G6PD target sequence of wild-type and mutant genomic DNA samples followed by hybridization to a test strip containing allele-specific oligonucleotide probes. A total of 16,464 individuals were analyzed by a combination of phenotypic screening and genotypic detection using the PCR-RDB assay and DNA sequence analysis.

Results

The PCR-RDB assay had a detection rate of 98.1%, which was validated by direct sequencing in a blind study with 100% concordance. The G6PD deficiency incidence rate in Dongguan District is 4.08%. Thirty-two genotypes from 469 individuals were found. The two most common variants were c.1376G>T and c.1388G>A, followed by c.95A>G, c.871G>A, c.392G>T, and c.1024 C>T. In addition, two rare mutations (c.703C>A and c.406C>T) were detected by DNA sequencing analysis. In our study, 65 cases harbored the C1311T/IVS polymorphism and 67 cases were homozygote.

Conclusion

The PCR-RDB assay we established is a reliable and effective method for screening G6PD mutations in the Chinese population. Data on the spectrum of mutations in the Dongguan District is beneficial to the clinical diagnosis and prevention of G6PD deficiency.     

Evaluation of the Diagnostic Accuracy of CareStart G6PD Deficiency Rapid Diagnostic Test (RDT) in a Malaria Endemic Area in Ghana,Africa

Background

Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most widespread enzyme defect that can result in red cell breakdown under oxidative stress when exposed to certain medicines including antimalarials. We evaluated the diagnostic accuracy of CareStart G6PD deficiency Rapid Diagnostic Test (RDT) as a point-of-care tool for screening G6PD deficiency.

Methods

A cross-sectional study was conducted among 206 randomly selected and consented participants from a group with known G6PD deficiency status between February 2013 and June 2013. A maximum of 1.6ml of capillary blood samples were used for G6PD deficiency screening using CareStart G6PD RDT and Trinity qualitative with Trinity quantitative methods as the “gold standard”. Samples were also screened for the presence of malaria parasites. Data entry and analysis were done using Microsoft Access 2010 and Stata Software version 12. Kintampo Health Research Centre Institutional Ethics Committee granted ethical approval.

Results

The sensitivity (SE) and specificity (SP) of CareStart G6PD deficiency RDT was 100% and 72.1% compared to Trinity quantitative method respectively and was 98.9% and 96.2% compared to Trinity qualitative method. Malaria infection status had no significant (P=0.199) change on the performance of the G6PD RDT test kit compared to the “gold standard”.

Conclusions

The outcome of this study suggests that the diagnostic performance of the CareStart G6PD deficiency RDT kit was high and it is acceptable at determining the G6PD deficiency status in a high malaria endemic area in Ghana. The RDT kit presents as an attractive tool for point-of-care G6PD deficiency for rapid testing in areas with high temperatures and less expertise. The CareStart G6PD deficiency RDT kit could be used to screen malaria patients before administration of the fixed dose primaquine with artemisinin-based combination therapy.     

Assessment of Point-of-Care Diagnostics for G6PD Deficiency in Malaria Endemic Rural Eastern Indonesia

Background

Patients infected by Plasmodium vivax or Plasmodium ovale suffer repeated clinical attacks without primaquine therapy against latent stages in liver. Primaquine causes seriously threatening acute hemolytic anemia in patients having inherited glucose-6-phosphate dehydrogenase (G6PD) deficiency. Access to safe primaquine therapy hinges upon the ability to confirm G6PD normal status. CareStart G6PD, a qualitative G6PD rapid diagnostic test (G6PD RDT) intended for use at point-of-care in impoverished rural settings where most malaria patients live, was evaluated.

Methodology/Principal Findings

This device and the standard qualitative fluorescent spot test (FST) were each compared against the quantitative spectrophotometric assay for G6PD activity as the diagnostic gold standard. The assessment occurred at meso-endemic Panenggo Ede in western Sumba Island in eastern Indonesia, where 610 residents provided venous blood. The G6PD RDT and FST qualitative assessments were performed in the field, whereas the quantitative assay was performed in a research laboratory at Jakarta. The median G6PD activity ≥5 U/gHb was 9.7 U/gHb and was considered 100% of normal activity. The prevalence of G6PD deficiency by quantitative assessment (<5 U/gHb) was 7.2%. Applying 30% of normal G6PD activity as the cut-off for qualitative testing, the sensitivity, specificity, positive predictive value, and negative predictive value for G6PD RDT versus FST among males were as follows: 100%, 98.7%, 89%, and 100% versus 91.7%, 92%, 55%, and 99%; P = 0.49, 0.001, 0.004, and 0.24, respectively. These values among females were: 83%, 92.7%, 17%, and 99.7% versus 100%, 92%, 18%, and 100%; P = 1.0, 0.89, 1.0 and 1.0, respectively.

Conclusions/Significance

The overall performance of G6PD RDT, especially 100% negative predictive value, demonstrates suitable safety for G6PD screening prior to administering hemolytic drugs like primaquine and many others. Relatively poor diagnostic performance among females due to mosaic G6PD phenotype is an inherent limitation of any current practical screening methodology.     

Haplotype analyses of haemoglobin C and haemoglobin S and the dynamics of the evolutionary response to malaria in Kassena-Nankana District of Ghana

Background

Haemoglobin S (HbS) and C (HbC) are variants of the HBB gene which both protect against malaria. It is not clear, however, how these two alleles have evolved in the West African countries where they co-exist at high frequencies. Here we use haplotypic signatures of selection to investigate the evolutionary history of the malaria-protective alleles HbS and HbC in the Kassena-Nankana District (KND) of Ghana.

Methodology/Principal Findings

The haplotypic structure of HbS and HbC alleles was investigated, by genotyping 56 SNPs around the HBB locus. We found that, in the KND population, both alleles reside on extended haplotypes (approximately 1.5 Mb for HbS and 650 Kb for HbC) that are significantly less diverse than those of the ancestral HbA allele. The extended haplotypes span a recombination hotspot that is known to exist in this region of the genome

Significance

Our findings show strong support for recent positive selection of both the HbS and HbC alleles and provide insights into how these two alleles have both evolved in the population of northern Ghana.     

Comparison of Three Screening Test Kits for G6PD Enzyme Deficiency: Implications for Its Use in the Radical Cure of Vivax Malaria in Remote and Resource-Poor Areas in the Philippines

Objective

We evaluated a battery of Glucose-6-Phosphate Dehydrogenase diagnostic point-of-care tests (PoC) to assess the most suitable product in terms of performance and operational characteristics for remote areas.

Methods

Samples were collected in Puerto Princesa City, Palawan, Philippines and tested for G6PD deficiency with a fluorescent spot test (FST; Procedure 203, Trinity Biotech, Ireland), the semiquantitative WST8/1-methoxy PMS (WST; Dojindo, Japan) and the Carestart G6PD Rapid Diagnostic Test (CSG; AccessBio, USA). Results were compared to spectrophotometry (Procedure 345, Trinity Biotech, Ireland). Sensitivity and specificity were calculated for each test with cut-off activities of 10%, 20%, 30% and 60% of the adjusted male median.

Results

The adjusted male median was 270.5 IU/10¹² RBC. FST and WST were tested on 621 capillary blood samples, the CSG was tested on venous and capillary blood on 302 samples. At 30% G6PD activity, sensitivity for the FST was between 87.7% (95%CI: 76.8% to 93.9%) and 96.5% (95%CI: 87.9% to 99.5%) depending on definition of intermediate results; the WST was 84.2% (95%CI: 72.1% to 92.5%); and the CSG was between 68.8% (95%CI: 41.3% to 89.0%) and 93.8% (95%CI: 69.8% to 99.8%) when the test was performed on capillary or venous blood respectively. Sensitivity of FST and CSG (tested with venous blood) were comparable (p>0.05). The analysis of venous blood samples by the CSG yielded significantly higher results than FST and CSG performed on capillary blood (p<0.05). Sensitivity of the CSG varied depending on source of blood used (p<0.05).

Conclusion

The operational characteristics of the CSG were superior to all other test formats. Performance and operational characteristics of the CSG performed on venous blood suggest the test to be a good alternative to the FST.     

Glucose-6-Phosphate Dehydrogenase Deficiency and Haemoglobin Drop after Sulphadoxine-Pyrimethamine Use for Intermittent Preventive Treatment of Malaria during Pregnancy in Ghana – A Cohort Study

Background

Sulphadoxine-Pyrimethamine (SP) is still the only recommended antimalarial for use in intermittent preventive treatment of malaria during pregnancy (IPTp) in some malaria endemic countries including Ghana. SP has the potential to cause acute haemolysis in G6PD deficient people resulting in significant haemoglobin (Hb) drop but there is limited data on post SP-IPTp Hb drop. This study determined the difference, if any in proportions of women with significant acute haemoglobin drop between G6PD normal, partial deficient and full deficient women after SP-IPTp.

Methods and Findings

Prospectively, 1518 pregnant women who received SP for IPTp as part of their normal antenatal care were enrolled. Their G6PD status were determined at enrollment followed by assessments on days 3, 7,14 and 28 to document any adverse effects and changes in post-IPTp haemoglobin (Hb) levels. The three groups were comparable at baseline except for their mean Hb (10.3 g/dL for G6PD normal, 10.8 g/dL for G6PD partial deficient and 10.8 g/dL for G6PD full defect women).The prevalence of G6PD full defect was 2.3% and 17.0% for G6PD partial defect. There was no difference in the proportions with fractional Hb drop ≥ 20% as compared to their baseline value post SP-IPTp among the 3 groups on days 3, 7, 14. The G6PD full defect group had the highest median fractional drop at day 7. There was a weak negative correlation between G6PD activity and fractional Hb drop. There was no statistical difference between the three groups in the proportions of those who started the study with Hb ≥ 8g/dl whose Hb level subsequently fell below 8g/dl post-SP IPTp. No study participant required transfusion or hospitalization for severe anaemia.

Conclusions

There was no significant difference between G6PD normal and deficient women in proportions with significant acute haemoglobin drop post SP-IPTp and lower G6PD enzyme activity was not strongly associated with significant acute drug-induced haemoglobin drop post SP-IPTp but a larger study is required to confirm consistency of findings.     

Risks of Hemolysis in Glucose-6-Phosphate Dehydrogenase Deficient Infants Exposed to Chlorproguanil-Dapsone,Mefloquine and Sulfadoxine-Pyrimethamine as Part of Intermittent Presumptive Treatment of Malaria in Infants

Background

Chlorproguanil-dapsone (CD) has been linked to hemolysis in symptomatic glucose-6-phosphate dehydrogenase deficient (G6PDd) children. Few studies have explored the effects of G6PD status on hemolysis in children treated with Intermittent Preventive Treatment in infants (IPTi) antimalarial regimens. We sought to examine the joint effects of G6PD status and IPTi antimalarial treatment on incidence of hemolysis in asymptomatic children treated with CD, sulfadoxine-pyrimethamine (SP), and mefloquine (MQ).

Methods

A secondary analysis of data from a double-blind, placebo-controlled trial of IPTi was conducted. Hemoglobin (Hb) measurements were made at IPTi doses, regular follow-up and emergency visits. G6PD genotype was determined at 9 months looking for SNPs for the A- genotype at coding position 202. Multivariable linear and logistic regression models were used to examine hemolysis among children with valid G6PD genotyping results. Hemolysis was defined as the absolute change in Hb or as any post-dose Hb <8 g/dL. These outcomes were assessed using either a single follow-up Hb on day 7 after an IPTi dose or Hb obtained 1 to 14 or 28 days after each IPTi dose.

Findings

Relative to placebo, CD reduced Hb by approximately 0.5 g/dL at day 7 and within 14 days of an IPTi dose, and by 0.2 g/dL within 28 days. Adjusted declines in the CD group were larger than in the MQ and SP groups. At day 7, homo-/hemizygous genotype was associated with higher odds of Hb <8 g/dL (adjusted odds ratio = 6.7, 95% CI 1.7 to 27.0) and greater absolute reductions in Hb (-0.6 g/dL, 95% CI -1.1 to 0.003). There was no evidence to suggest increased reductions in Hb among homo-/hemizygous children treated with CD compared to placebo, SP or MQ.

Conclusions

While treatment with CD demonstrated greater reductions in Hb at 7 and 14 days after an IPTi dose compared to both SP and MQ, there was no evidence that G6PD deficiency exacerbated the adverse effects of CD, despite evidence for higher hemolysis risk among G6PDd infants.     

Prevalence of α-thalassemia 3.7 kb deletion in the adult population of Rio Grande do Norte,Brazil

α-Thalassemia, arising from a defect in α-globin chain synthesis, is often caused by deletions involving one or both of the α-genes on the same allele. With the aim of investigating the prevalence of α-thalassemia 3.7 kb deletion in the adult population of Rio Grande do Norte, 713 unrelated individuals, between 18 and 59 years-of-age, were analyzed. Red blood cell indices were electronically determined, and A₂ and F hemoglobins evaluated by HPLC. PCR was applied to the molecular investigation of α-thalassemia 3.7 kb deletion. Eighty (11.2%) of the 713 individuals investigated presented α-thalassemia, of which 79 (11.1%) were heterozygous (-α^3.7/αα) deletions and 1 (0.1%) homozygous (-α^3.7/-α^3.7). Ethnically, heterozygous deletions were higher (24.8%) in Afro-Brazilians. Comparison of hematological parameters between individuals with normal genotype and those with heterozygous α⁺-thalassemia showed a statistically significant difference in the number of erythrocytes (p < 0.001), MCV (p < 0.001), MCH (p < 0.001) and Hb A₂ (p = 0.007). This study is one of the first dedicated to investigating α-thalassemia 3.7 kb deletion in the population of the State Rio Grande do Norte state. Results obtained demonstrate the importance of investigating this condition in order to elucidate the causes of microcytosis and hypochromia.     

Single Low Dose Primaquine (0.25mg/kg) Does Not Cause Clinically Significant Haemolysis in G6PD Deficient Subjects

Background

Primaquine is the only drug consistently effective against mature gametocytes of Plasmodium falciparum. The transmission blocking dose of primaquine previously recommended was 0.75mg/kg (adult dose 45mg) but its deployment was limited because of concerns over haemolytic effects in patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency. G6PD deficiency is an inherited X-linked enzymatic defect that affects an estimated 400 million people around the world with high frequencies (15–20%) in populations living in malarious areas. To reduce transmission in low transmission settings and facilitate elimination of P. falciparum, the World Health Organization now recommends adding a single dose of 0.25mg/kg (adult dose 15mg) to Artemisinin-based Combination Therapies (ACTs) without G6PD testing. Direct evidence of the safety of this low dose is lacking. Adverse events and haemoglobin variations after this treatment were assessed in both G6PD normal and deficient subjects in the context of targeted malaria elimination in a malaria endemic area on the North-Western Myanmar-Thailand border where prevalence of G6PD deficiency (Mahidol variant) approximates 15%.

Methods and Findings

The tolerability and safety of primaquine (single dose 0.25 mg base/kg) combined with dihydroartemisinin-piperaquine (DHA-PPQ) given three times at monthly intervals was assessed in 819 subjects. Haemoglobin concentrations were estimated over the six months preceding the ACT + primaquine rounds of mass drug administration. G6PD deficiency was assessed with a phenotypic test and genotyping was performed in male subjects with deficient phenotypes and in all females. Fractional haemoglobin changes in relation to G6PD phenotype and genotype and primaquine round were assessed using linear mixed-effects models. No adverse events related to primaquine were reported during the trial. Mean fractional haemoglobin changes after each primaquine treatment in G6PD deficient subjects (-5.0%, -4.2% and -4.7%) were greater than in G6PD normal subjects (0.3%, -0.8 and -1.7%) but were clinically insignificant. Fractional drops in haemoglobin concentration larger than 25% following single dose primaquine were observed in 1.8% of the population but were asymptomatic.

Conclusions

The single low dose (0.25mg/kg) of primaquine is clinically well tolerated and can be used safely without prior G6PD testing in populations with high prevalence of G6PD deficiency. The present evidence supports a broader use of low dose primaquine without G6PD testing for the treatment and elimination of falciparum malaria.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT01872702","term_id":"NCT01872702"}}NCT01872702     

Field Trial Evaluation of the Performances of Point-of-Care Tests for Screening G6PD Deficiency in Cambodia

Background

User-friendly, accurate, point-of-care rapid tests to detect glucose-6-phosphate dehydrogenase deficiency (G6PDd) are urgently needed at peripheral level to safely recommend primaquine for malaria elimination.

Methods

The CareStart G6PD RDT (AccessBio, New Jersey, USA), a novel rapid diagnostic test and the most commonly used test, the fluorescent spot test (FST) were assessed against the quantitatively measured G6PD enzyme activity for detecting G6PDd. Subjects were healthy males and non-pregnant females aged 18 years or older residing in six villages in Pailin Province, western Cambodia.

Findings

Of the 938 subjects recruited, 74 (7.9%) were severe and moderately severe G6PD deficient (enzyme activity <30%), mostly in male population; population median G6PD activity was 12.0 UI/g Hb. The performances of the CareStart G6PD RDT and the FST, according to different cut-off values used to define G6PDd were very similar. For the detection of severe and moderately severe G6PDd (enzyme activity <30%, <3.6 UI/g Hb) in males and females, sensitivity and negative (normal status) predictive value were 100% for both point-of-care tools. When the G6PDd cut-off value increased (from <40% to <60%), the sensitivity for both PoCs decreased: 93.3% to 71.7% (CareStart G6PD RDT, p = 10⁻⁶) and 95.5% to 73.2% (FST, p = 10⁻⁶) while the specificity for both PoCs remained similar: 97.4% to 98.3% (CareStart G6PD RDT, p = 0.23) and 98.7% to 99.6% (FST, p = 0.06). The cut-off values for classifying individuals as normal were 4.0 UI/g Hb and 4.3 UI/g Hb for the CareStart G6PD RDT and the FST, respectively.

Conclusions

The CareStart G6PD RDT reliably detected moderate and severe G6PD deficient individuals (enzyme activity <30%), suggesting that this novel point-of-care is a promising tool for tailoring appropriate primaquine treatment for malaria elimination by excluding individuals with severe G6PDd for primaquine treatment.     

Excessive Weight Gain during the First Year of Peritoneal Dialysis Is Associated with Inflammation,Diabetes Mellitus,and a Rapid Decrease in Residual Renal Function

Objectives

Significant weight gain is a potential problem in most patients starting peritoneal dialysis (PD); however, few studies have explored the clinical effects of increased body weight (BW) in these patients. We evaluated the effect of excess weight gain during the first year after PD on residual renal function (RRF).

Methods

A total of 148 incident PD patients were analyzed in a longitudinal observational study. The mean duration of follow-up was 23.8 months. RRF was measured at baseline (within 1 month of starting PD) and thereafter at 6-month intervals for 2–3 years or until loss of RRF. BW was measured at the time of RRF measurement, and excess weight gain was defined as a BW increase over the median value (3.0%).

Results

The median 1-year increase in BW was 2.3kg (IQR, 1.01–4.58) or 3.0% (IQR, 1.13–5.31). The mean slope of RRF decline was –0.068 ± 0.053 mL/min/month/1.73m², and RRF loss developed in 48 patients at a mean follow-up time of 19.4 ± 6.8 months. Patients with BW increases > 3.0% showed significantly increased RRF decline rate compared to those without excess weight gain (p<0.001), and the BW increase (%/year) correlated significantly with higher hs-CRP levels and RRF decline rate. High systolic blood pressure, diabetes, large amount of proteinuria and excess BW gain significantly influenced the RRF decline rate. Also, it increased the risk of RRF loss by 4.17-fold (95% confidence intervals, 1.87–9.28; p<0.001).

Conclusions

Excess weight gain during the first year of PD was closely linked to systemic inflammation, diabetes and rapid decline in RRF.     

Distinctive Features of NREM Parasomnia Behaviors in Parkinson’s Disease and Multiple System Atrophy

Objective

To characterize parasomnia behaviors on arousal from NREM sleep in Parkinson’s Disease (PD) and Multiple System Atrophy (MSA).

Methods

From 30 patients with PD, Dementia with Lewy Bodies/Dementia associated with PD, or MSA undergoing nocturnal video-polysomnography for presumed dream enactment behavior, we were able to select 2 PD and 2 MSA patients featuring NREM Parasomnia Behviors (NPBs). We identified episodes during which the subjects seemed to enact dreams or presumed dream-like mentation (NPB arousals) versus episodes with physiological movements (no-NPB arousals). A time-frequency analysis (Morlet Wavelet Transform) of the scalp EEG signals around each NPB and no- NPB arousal onset was performed, and the amplitudes of the spectral frequencies were compared between NPB and no-NPB arousals.

Results

19 NPBs were identified, 12 of which consisting of ‘elementary’ NPBs while 7 resembling confusional arousals. With quantitative EEG analysis, we found an amplitude reduction in the 5-6 Hz band 40 seconds before NPBs arousal as compared to no-NPB arousals at F4 and C4 derivations (p<0.01).

Conclusions

Many PD and MSA patients feature various NREM sleep-related behaviors, with clinical and electrophysiological differences and similarities with arousal parasomnias in the general population.

Significance

This study help bring to attention an overlooked phenomenon in neurodegenerative diseases.     

Body Mass Index and Risk of Parkinson’s Disease: A Dose-Response Meta-Analysis of Prospective Studies

Background

A number of epidemiologic studies examining the relationship between body mass index (BMI) and the future occurrence of Parkinson’s disease (PD) reported largely inconsistent findings. We conducted a dose-response meta-analysis of prospective studies to clarify this association.

Methods

Eligible prospective studies were identified by a search of PubMed and by checking the references of related publications. The generalized least squares trend estimation was employed to compute study-specific relative risks (RR) and 95% confidence intervals (CI) for an increase in BMI of 5 kg/m², and the random-effects model was used to compute summary RR and 95% CI.

Results

A total of 10 prospective studies were included in the final analysis. An increase in BMI of 5 kg/m² was not associated with PD risk, with a summary RR of 1.00 (95% CI = 0.89-1.12). Results of subgroup analysis found similar results except for a week positive association in studies that adjusted for alcohol consumption (RR = 1.13, 95% CI = 0.99-1.29), and a week inverse association in studies that did not (RR = 0.90, 95% CI = 0.78-1.04). In a separate meta-analysis, no significant association between overweight (25 kg/m² ≤ BMI ≤29.9 kg/m²), obesity (BMI≥30 kg/m²) or excess weight (BMI≥25 kg/m²) and PD risk was observed.

Conclusion

This meta-analysis does not support the notion that higher BMI materially increases PD risk. However, a week positive BMI-PD association that may be masked by confounders still cannot be excluded, and future prospective studies with a good control for potential confounding factors are needed.     

Blood Lead Levels in Children Aged 0–6 Years Old in Hunan Province,China from 2009–2013

Objectives

The aim of this study is to describe blood lead levels (BLLs) and the prevalence of elevated blood lead levels (EBLLs) in children aged 0–6 years old and to analyze the BLL trend in children from 2009 to 2013 in China.

Methods

A total of 124,376 children aged 0–6 years old were recruited for this study from January 1^st 2009 to December 31^st 2013. Their blood lead levels were analyzed using atomic absorption spectrometry.

Results

The median BLL was 64.3 μg/L (IQR: 49.6–81.0), and the range was 4.3–799.0 μg/L. Blood lead levels were significantly higher in boys (66.0 μg/L) than in girls (61.9 μg/L) (P<0.001). The overall prevalence of BLLs≥100 μg/L was 10.54% in children aged 0–6 years in Hunan Province. Between 2009 and 2013, the prevalence of EBLLs (≥100 μg/L) decreased from 18.31% to 4.26% in children aged 0–6 years and increased with age. The prevalence of EBLLs has dramatically decreased in two stages (2009–2010 and 2012–2013), with a slight fluctuation in 2010 and 2011.

Conclusions

Both BLLs and the prevalence of EBLLs in children aged 0–6 years old declined substantially from 2009 to 2013 in Hunan Province; however, both remain at unacceptably high levels compared to developed countries. Comprehensive strategies are required to further reduce blood lead levels in children.     

Fractalkine Signaling Regulates the Inflammatory Response in an α-Synuclein Model of Parkinson Disease

Background

Parkinson disease (PD) is a progressive neurodegenerative disorder characterized by loss of dopamine neurons in the substantia nigra pars compacta (SNpc) and widespread aggregates of the protein alpha-synuclein (α-syn). Increasing evidence points to inflammation as a chief mediator; however, the role of α-syn in triggering and sustaining inflammation remains unclear. In models of Alzheimer’s disease (AD), multiple sclerosis (MS) and neurotoxin models of PD, the chemokine CX3CL1 (fractalkine) and its receptor (CX3CR1) have important roles in modulating neuroinflammation.

Methods

To examine the role of fractalkine signaling in α-syn-induced-neuroinflammation and neurodegeneration, we used an in vivo mouse model in which human α-syn is overexpressed by an adeno associated viral vector serotype 2 (AAV2) and in vitro phagocytosis and protein internalization assays with primary microglia treated with aggregated α-syn.

Results

We observed that loss of CX3CR1 expression led to a reduced inflammatory response, with reduced IgG deposition and expression of MHCII 4 weeks post-transduction. Six months post transduction, AAV2 mediated overexpression of α-syn leads to loss of dopaminergic neurons, and this loss was not exacerbated in animals with deletion of CX3CR1. To determine the mechanism by which CX3CR1affects inflammatory responses in α-syn-induced inflammation, phagocytosis was assessed using a fluorescent microsphere assay as well as by microglial uptake of aggregated α-syn. CX3CR1-/- microglia showed reduced uptake of fluorescent beads and aggregated α-syn.

Conclusion

Our results suggest that one mechanism by which CX3CR1-/- attenuates inflammation is at the level of phagocytosis of aggregated α-syn by microglia. These data implicate fractalkine signaling as a potential therapeutic target for regulating inflammatory response in α-syn models PD.     

Inflammation,Endothelial Dysfunction and Increased Left Ventricular Mass in Chronic Kidney Disease (CKD) Patients: A Longitudinal Study

Introduction

Within this longitudinal study we investigated the association of inflammation markers C-reactive protein (CRP), interleukin-6 (IL-6) and tumor necrosis factor-α (TNFα) and endothelial dysfunction markers intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) with left ventricular mass indexed for height²·⁷¹ (LVMI) in hypertensive predialysis CKD patients.

Material and Methods

From 2004 to 2005, 182 incident consecutive adult patients from the outpatient CKD clinics of two hospitals in Greece with CKD and hypertension or using antihypertensive medication, were included. Of these, 107 patients underwent CRP (mg/l) and LVMI (g/height²·⁷¹) measurements annually for three years.

Results

In the longitudinal analyses, using linear mixed modeling, a higher IL-6 (ß = 1.9 (95%ci:0.38;3.5), inflammation score based on CRP, IL-6 and TNF-α (ß = 5.0 (95%ci:0.72; 9.4) and VCAM-1 (ß = 0.01 (95%ci:0.005;0.02) were associated with higher LVMI. These models were adjusted for age, gender and primary renal disease, and for confounders that on top changed the beta with ≥10%, i.e. diuretic use (for IL-6 and inflammation score).

Conclusion

The results suggest that in predialysis CKD patients, inflammation as well as endothelial dysfunction may play an important role towards the increase in LVMI.     

Hemoglobins S and C in Upper Volta

Summary We have studied the incidence of hemoglobinopathies in 1059 individuals in Upper Volta. We have found that this population has a high frequency of HbS and HbC and -thalassemia. The gene frequency of HbS was high (0.1 for the ^S gene) in the arid Sahel portion of Upper Volta accompanied by a lower frequency for HbC (0.05 for the ^c gene). The reverse was true in the humid Savanna region of this country (0.03 for the ^S gene and 0.14 for the ^c gene). There was no age dependency of the HbS gene frequency, but -thalassemia, detectable in HbS heterozygotes, showed a statistically significant decrease with age. No homozygote for HbS was detected after the age of 1 year, and SC and CC genotypes were found at a lower incidence than expected. The environmental and medical conditions in Upper Volta preclude the survival of SS individuals and decrease the survival of SC and CC genotypes.     

Combined Antigen-Specific Interferon-γ and Interleukin-2 Release Assay (FluoroSpot) for the Diagnosis of Mycobacterium tuberculosis Infection

Background

To evaluate interleukin (IL)-2 and interferon (IFN)-γ secreting T-cells in parallel for the differentiation of latent infection with Mycobacterium tuberculosis infection (LTBI) from active tuberculosis.

Methods

Following ex-vivo stimulation of peripheral blood mononuclear cells (PBMC) with M. tuberculosis-specific antigens early secretory antigenic target (ESAT)-6 and culture filtrate protein (CFP)-10, immune responses were assessed by enzyme-linked immunospot IFN-γ release assay (EliSpot-IGRA) and a novel dual cytokine detecting fluorescence-linked immunospot (FluoroSpot) in 18 patients with pulmonary tuberculosis, 10 persons with previously cured tuberculosis, 25 individuals with LTBI and 16 healthy controls.

Results

Correlation of IFN- γ⁺ spot-forming cells in EliSpot-IGRA and FluoroSpot were R² = 0.67 for ESAT-6 and R² = 0.73 for CFP-10. The number of IL-2^- IFN- γ⁺ producing cells was higher in patients with tuberculosis compared with past tuberculosis (CFP-10-induced p = 0.0068) or individuals with LTBI (ESAT-6-induced p = 0.0136). A cutoff value of >16 CFP-10-induced IFN-γ⁺ secreting cells/200.000 PBMC in the EliSpot-IGRA discriminated with highest sensitivity and specificity (89% and 76%, respectively). However, overlap in cytokine responses precludes distinction between the cohorts on an individual basis.

Conclusions

Combined analysis of IFN-γ and IL-2 secretion by antigen specific T-cells does not allow a reliable differentiation between different states of M. tuberculosis infection in clinical practice.     

Investigation on Abnormal Iron Metabolism and Related Inflammation in Parkinson Disease Patients with Probable RBD

Objective

To investigate potential mechanisms involving abnormal iron metabolism and related inflammation in Parkinson disease (PD) patients with probable rapid eye movement sleep behavior disorder (PRBD).

Methods

Total 210 PD patients and 31 controls were consecutively recruited. PD patients were evaluated by RBD Screening Questionnaire (RBDSQ) and classified into PRBD and probable no RBD (NPRBD) groups. Demographics information were recorded and clinical symptoms were evaluated by series of rating scales. Levels of iron and related proteins and inflammatory factors in cerebrospinal fluid (CSF) and serum were detected. Comparisons among control, NPRBD and PRBD groups and correlation analyses between RBDSQ score and levels of above factors were performed.

Results

(1)The frequency of PRBD in PD patients is 31.90%. (2)PRBD group has longer disease duration, more advanced disease stage, severer motor symptoms and more non-motor symptoms than NPRBD group. (3)In CSF, levels of iron, transferrin, NO and IL–1β in PRBD group are prominently increased. RBDSQ score is positively correlated with the levels of iron, transferrin, NO and IL–1β in PD group. Iron level is positively correlated with the levels of NO and IL–1β in PD group. (4)In serum, transferrin level is prominently decreased in PRBD group. PGE₂ level in PRBD group is drastically enhanced. RBDSQ score exhibits a positive correlation with PGE₂ level in PD group.

Conclusions

PRBD is common in PD patients. PRBD group has severer motor symptoms and more non-motor symptoms. Excessive iron in brain resulted from abnormal iron metabolism in central and peripheral systems is correlated with PRBD through neuroinflammation.