Elevated Cerebral Spinal Fluid Cytokine Levels in Boys with Cerebral Adrenoleukodystrophy Correlates with MRI Severity

Background: X-linked adrenoleukodystrophy (ALD) is a metabolic, peroxisomal disease that results from a mutation in the ABCD1 gene. The most severe course of ALD progression is the cerebral inflammatory and demyelinating form of the disease, cALD. To date there is very little information on the cytokine mediators in the cerebral spinal fluid (CSF) of these boys. Methodology/Principal Findings: Measurement of 23 different cytokines was performed on CSF and serum of boys with cerebral ALD and patients without ALD. Significant elevations in CSF IL-8 (29.362.2 vs 12.861.1 pg/ml, p = 0.0001), IL-1ra (166630 vs 8.666.5 pg/ml, p = 0.005), MCP-1 (610647 vs 328634 pg/ml, p = 0.002), and MIP-1b (14.261.3 vs 2.061.4 pg/ml, p,0.0001) were found in boys with cALD versus the control group. The only serum cytokine showing an elevation in the ALD group was SDF-1 (21246155 vs 11756125 pg/ml, p = 0.0001). The CSF cytokines of IL-8 and MCP-1b correlated with the Loes MRI severity score (p = 0.04 and p = 0.008 respectively), as well as the serum SDF-1 level (p = 0.002). Finally, CSF total protein was also significantly elevated in boys with cALD and correlated with both IL-8, MCP-1b (p = 0.0001 for both), as well as Loes MRI severity score (p = 0.0007). Conclusions/Significance: IL-8, IL-1ra, MCP-1, MIP-1b and CSF total protein were significantly elevated in patients with cALD; IL-8, MCP-1b, and CSF total protein levels correlated with disease severity determined by MRI. This is the largest repor

Elevated levels of CSF total protein correlate to IL-8, MCP-1, and Loes MRI severity score.

<p>CSF total protein levels were taken from the patients' medical record. Boxes show min and max, and the bar is at the mean. Regression analysis was performed using the previously determined CSF cytokine levels for IL-8 and MCP-1 as well as each patient's pretransplant Loes score using Prism software.</p

CSF inflammatory cytokines are elevated in boys with cerebral ALD.

<p>CSF cytokines were evaluated using a Lumunix system and SDF-1 by ELISA. Boxes show min and max, and the bar is at the mean. A Student's unpaired t-test calculated p-values shown.</p

List of inflammatory factors evaluated in CSF and serum samples.

<p>List of inflammatory factors evaluated in CSF and serum samples.</p

Correlation of CSF inflammatory cytokine levels and Loes MRI severity score.

<p>Regression analysis was performed using the previously determined CSF cytokine levels and each patient's pretransplant Loes score using Prism software.</p

Correlation of serum SDF-1 levels and Loes MRI severity score.

<p>Regression analysis was performed using serum SDF-1 levels and each patient's pretransplant Loes score using Prism software.</p

Correlation of CSF MCP-1 levels and chitotriosidase activity.

<p>Regression analysis was performed using CSF MCP-1 levels and CSF chitotriosidase activity as described by Orchard et al <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0032218#pone.0032218-Orchard1" target="_blank">[10]</a>. Statistical analysis was performed using Prism software.</p

Markers of oxidative stress in umbilical cord blood from G6PD deficient African newborns

<div><p>Glucose-6-phosphate dehydrogenase (G6PD) deficiency is an X-linked disorder that affects as many as 400 million people worldwide, making it the most common enzymatic defect. Subjects with G6PD deficiency are more likely to develop neonatal hyperbilirubinemia potentially leading to kernicterus and are at increased risk for acute hemolytic anemia when exposed to pro-oxidant compounds such as anti-malarial drugs. We collected umbilical cord blood from 300 males born in Uganda to assess for novel markers of systemic oxidative stress. We determined that 10.7% of the samples collected were G6PD A- deficient (G202A/A376G) and when these were compared with unaffected controls, there was significantly higher 8-hydroxy-2’-deoxyguanosine (8-OHdG) concentration, elevated ferritin, increased leukocyte count and higher small molecule antioxidant capacity. These data suggest increased baseline oxidative stress and an elevated antioxidant response in umbilical cord blood of patients with G6PD deficiency.</p></div

Complete blood count indices in G6PD deficient UCB from male neonates.

<p>(A-E) Total white blood cell (WBC), neutrophil, lymphocyte, platelet counts and hemoglobin were in G6PD deficient samples (n = 32) compared to normal controls (n = 240). Columns indicate the mean values and standard errors. P-values in two-way comparisons were derived from a Student’s t-test. Platelet values are from 23 G6PD deficient and 136 normal controls due to equipment malfunction. See also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0172980#pone.0172980.s001" target="_blank">S1 Table</a>.</p

Markers of oxidative stress/inflammation in G6PD deficient UCB from male neonates.

<p>(A) Levels of plasma 8-OHdG, n = 32 G6PD deficient, n = 35 Normal samples. (B) Pearson’s correlation analysis of plasma 8-OHdG levels and absolute lymphocyte counts. (C) Comparison of G6PD deficient UCB lymphocyte counts between high and low 8-OHdG levels determine by the median 8-OHdG value (31.9 nM). (D) Plasma ferritin concentration in UCB. (E) Plasma small molecule antioxidant capacity expressed in equivalents of Trolox. (F) Plasma protein carbonylation as nmol/mg. (G) Plasma catalase activity in UCB. See also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0172980#pone.0172980.s001" target="_blank">S1 Table</a>. Columns indicate the mean values and standard deviation. P-values in two-way comparisons were derived from a Student’s t-test.</p