The Expression of Active CD11b Monocytes in Blood and Disease Progression in Amyotrophic Lateral Sclerosis

Abstract

Monocytes expressing the inflammation suppressing active CD11b, a beta2 integrin, may regulate neuroinflammation and modify clinical outcomes in amyotrophic lateral sclerosis (ALS). In this single site, retrospective study, peripheral blood mononuclear cells from 38 individuals living with ALS and 20 non-neurological controls (NNC) were investigated using flow cytometry to study active CD11b integrin classical (CM), intermediate (IM) and non-classical (NCM) monocytes during ALS progression. Seventeen ALS participants were sampled at the baseline (V1) and at two additional time points (V2 and V3) for longitudinal analysis. Active CD11b+ CM frequencies increased steeply between the baseline and V3 (ANOVA repeated measurement, p < 0.001), and the V2/V1 ratio negatively correlated with the disease progression rate, similar to higher frequencies of active CD11b+ NCM at the baseline (R = −0.6567; p = 0.0031 and R = 0.3862; p = 0.0168, respectively). CD11b NCM, clinical covariates and neurofilament light-chain plasma concentration at the baseline predicted shorter survival in a multivariable and univariate analysis (CD11b NCM—HR: 1.05, CI: 1.01−1.11, p = 0.013. Log rank: above median: 43 months and below median: 21.22 months; p = 0.0022). Blood samples with the highest frequencies of active CD11b+ IM and NCM contained the lowest concentrations of soluble CD11b. Our preliminary data suggest that the levels of active CD11b+ monocytes and NCM in the blood predict different clinical outcomes in ALS.

Keywords: CD11b; amyotrophic lateral sclerosis; beta2 integrin; monocytes.

Figure 1

The expression of non-classical and classical monocytes and rate of ALS progression. (A) Higher log-transformed frequencies of CD11b+ NCM are positively correlated with ΔFRS. (B) The log-transformed ratio of active CD11b+ CM between V2 and baseline is negatively correlated with ΔFRS.

Figure 2

Blood monocyte frequencies and survival. Kaplan–Meier survival analysis shows a shorter survival for ALS patients with higher frequencies (above median) of NCM (p = 0.0118) (A) and with higher frequency of CD11b+ NCM, (p = 0.0023) (B). Higher V2/V1 ratios of active CD11b+ CM frequencies predict longer survival for ALS patients (p = 0.0022) (C). Red lines indicate ALS subgroups with higher analyte levels (above median), and black lines ALS subgroups with lower level (below median). Log rank chi-square and p values as well as survival in months calculated for each subset of ALS patients are reported for each Kaplan–Meier figure. p-value was obtained from log rank test chi-square. Survival is calculated from V1.

Figure 3

Changes in monocyte subset frequencies between longitudinal time points. Analysis of stability of expression of monocyte subsets between V2 and baseline (V1) in individuals with ALS and in slow progressing ALS (A-S). Active CD11b+ CM (red dots) are the only monocyte subset to increase over time, showing a significant elevation between baseline and V2 in ALS (A), in A-S (B) but not in A-F (C). In contrast, CD11b+ CM appear stable over time (green dot, A–C). Box plot representation of ANOVA analysis for repeated measures (mixed model with missing values at random) of the changes from V1 to V3 of active CD11b+ CM. In line with the results of the stability analysis, active CD11b+ CM (F) present an up-regulation in the later time points compared to baseline, both for ALS (p = 0.0011) and for A-S (p < 0.0001) (D). Spaghetti plot representing the change over time of active CD11b+ CM between V1 and V3 with a difference in the pattern of expression of A-F compared to A-S (E).

Figure 4

Pairwise correlation analysis between soluble CD11b plasma protein (sCD11b) concentrations and active CD11b monocytes frequencies in blood. In ALS samples only, sCD11b plasma concentrations are inversely correlated with the frequencies of active CD11b+IM (R = −0.44, p = 0.0038) and active CD11b+ CM (R = −0.41, p = 0.0043) (A). In all samples (ALS + NNC), sCD11b plasma concentrations show an inverse correlation with frequencies of active CD11b+IM (R = −0.41, p = 0.0025) and active CD11b+ CM (R = −0.40, p = 0.0037) (B). Analysis performed without 4 outliers with a sCD11b concentration >400 ng/mL (Supplementary Figure S4 shows a similar level of significance with all samples included).