Revealing protein profiles in Parkinson's disease by mass spectrometric analysis of living brain tissue during deep brain stimulation

Introduction

In an ageing population, the increase in neurodegenerative disorders such as Parkinson's disease (PD) poses a major challenge. A deeper understanding of the pathophysiology and identification of biomarkers is therefore of utmost interest to develop new diagnostic and therapeutic interventions. PD manifests through a spectrum of motor and non-motor symptoms, rooted in the selective loss of dopaminergic neurons in the substantia nigra. Current diagnostic approaches rely on post-mortem examinations of brain samples, underscoring the urgency for novel methodologies. Single-use deep brain stimulation (DBS) electrodes provide a promising tool for acquiring living brain tissue, enabling proteomic analysis to elucidate disease mechanisms and identify biomarkers, advancing our understanding of PD's progression.

Methods

Immediately after DBS surgery, electrodes were collected and associated proteins were added to lysis buffer. Protein concentration was determined and 20 µg of the samples were digested using the SP3 method. Peptides were cleaned and desalted using C18 stop and go extraction tips. Liquid-chromatography tandem mass-spectrometry (LC-MS/MS) was performed with an ion mobility TOF instrument using a 44min gradient and measured in DIA-PASEF approach. Spectra were analysed using DIA-NN and bioinformatic data analysis was performed using Perseus and InstantClue to obtain disease-specific protein profiles from different disease entities.

Preliminary data

We collected 38 samples from 12 patients undergoing DBS surgery to explore recovering neuronal cells from surgical. The patient cohort exhibited various neurological conditions, including Alzheimer"s Disease (AD), dystonia, essential tremor, Tourette Syndrome, and PD. Protein concentrations ranged from 25 µg to 390 µg per test electrode and LC-MS/MS identified up to 1182 proteins per sample, with a total of 3276 proteins from living brain tissue. The most abundant proteins were neurofilament and myelin-related proteins (PLP1, CA1, MBP), indicating the presence of neurons and glial cells. Principal component analysis (PCA) didn't show patient group segregation, possibly due to sample size limitations. However, a-synuclein (SNCA) showed a significant enrichment in PD whereas the amyloid precursor protein (APP) or tau protein (MAPT) showed and increased expression in AD patients.

The preliminary findings suggest feasibility in acquiring sufficient material from DBS electrodes for analysis and reveal PD and AD-related proteins. In future experiments we will expand the patient cohort with non-neurodegenerative conditions like Tourette Syndrome and pain as a control. This approach enables comprehensive examination of protein changes in specific and intact brain regions of living patients, overcoming the challenges of post-mortem tissue use, such as protein degeneration. This method provides a valuable tool for studying the molecular mechanisms underlying neurodegenerative diseases.