Gene Expression Differences in Peripheral Blood of Parkinson's Disease Patients With Distinct Progression Profiles

Abstract

The prognosis of neurodegenerative disorders is clinically challenging due to the inexistence of established biomarkers for predicting disease progression. Here, we performed an exploratory cross-sectional, case-control study aimed at determining whether gene expression differences in peripheral blood may be used as a signature of Parkinson's disease (PD) progression, thereby shedding light into potential molecular mechanisms underlying disease development. We compared transcriptional profiles in the blood from 34 PD patients who developed postural instability within ten years with those of 33 patients who did not develop postural instability within this time frame. Our study identified >200 differentially expressed genes between the two groups. The expression of several of the genes identified was previously found deregulated in animal models of PD and in PD patients. Relevant genes were selected for validation by real-time PCR in a subset of patients. The genes validated were linked to nucleic acid metabolism, mitochondria, immune response and intracellular-transport. Interestingly, we also found deregulation of these genes in a dopaminergic cell model of PD, a simple paradigm that can now be used to further dissect the role of these molecular players on dopaminergic cell loss. Altogether, our study provides preliminary evidence that expression changes in specific groups of genes and pathways, detected in peripheral blood samples, may be correlated with differential PD progression. Our exploratory study suggests that peripheral gene expression profiling may prove valuable for assisting in prediction of PD prognosis, and identifies novel culprits possibly involved in dopaminergic cell death. Given the exploratory nature of our study, further investigations using independent, well-characterized cohorts will be essential in order to validate our candidates as predictors of PD prognosis and to definitively confirm the value of gene expression analysis in aiding patient stratification and therapeutic intervention.

Fig 1. Hierarchical clustering of genes detected as differentially expressed in rapid versus slow progression PD patients.

Classification based on expression signals of the bootstrapping t-test-detected genes resulted in only five mis-classified slow and 11 mis-classified rapid progression patients. The color bar denotes z-score adjusted expression values, green used for lower expression and purple for higher expression levels. Eucledian distance and average linkage methods were used.

Fig 2. Functional network of the differentially expressed genes altered in PD patients having slow versus rapid disease progression.

GO network analysis on the GO biological process terms that were found as highly enriched in blood cells from slow versus rapid progression patients. The node circle size represents the number of altered genes in the category, and the categories colours correspond to the functional group.

Fig 3. Selected genes investigated by qPCR in PD patients-derived samples.

-ΔCts plotted for 10 genes chosen for qPCR validation in cDNA obtained from 16 patients with either slow or rapid progression of the disease. Data is expressed as means ± SD of triplicates. T-test was used for statistical analysis with significance level of p<0.05. *p<0.05; **p<0.01; ***p<0.0005.

Fig 4. Selected genes investigated in a LUHMES/MPP⁺ cell model of PD.

(A) LUHMES cells treated with solvent (DMSO) or 2.5μM MPP⁺ stained for Tyrosine Hydroxylase (TH), TUJ1 and nucleus or (B) observed in bright field, showed robust loss of neurite integrity upon MPP⁺ treatment. (C) Cell viability analysis assessed by ToxiLigh assay revealed a significant increase of adenylate kinase content in the supernatants of MPP⁺ treated cells. (D) -Δcts plotted for seven genes chosen for validation using D8 differentiated LUHMES cells exposed to 2.5μm MPP⁺. (E) Correlation between fold-change expression values obtained in LUHMES/MPP⁺ model and PD patients. Data is expressed as means ± SD of triplicate samples. T-test was used for statistical analysis with significance level of p<0.05. *p<0.05; **p<0.01. Scale bar 100 μm.