Background
Histological analysis of brain tissue samples provide valuable information about the pathological processes leading to common neurodegenerative disorders such as Alzheimer’s or Parkinson’s diseases. In particular, high resolution and specific analysis of intra-neuronal lesions is crucial to understand the pathogenesis and progression of these diseases. In this context, the development of novel imaging approaches is a current challenge in neuroscience.
Methods
To this end, we used a recent super-resolutive imaging technique called STochastic Optical Reconstruction Microscopy (STORM) to analyze human brain sections. We combined STORM cell imaging protocols with neuropathological techniques and imaged cryopreserved brain samples from control subjects and patients with neurodegenerative diseases.
Results
This approach allowed us to perform 2D-, 3D- and two-color-STORM in central nervous system tissue sections, and to characterize with a nanometer-scale precision the architecture of physiological and pathological structures in neocortex, white matter and brainstem samples. STORM proved to be particularly effective to visualize the organization of dense protein inclusions, and allowed us to image with unprecedented details Aβ, Tau, α-synuclein and TDP-43 pathological aggregates within the central nervous system of patients with neurodegenerative disorders.
Conclusions
STORM imaging of human brain samples opens further gates to a more comprehensive understanding of the underlying mechanisms responsible for common neurological diseases. The convenience of this technique should open a straightforward extension of its application for super-resolution imaging of the human brain, with promising avenues to current challenges in neuroscience.
The centrosome, as the main microtubule organizing center, plays key roles in cell polarity, genome stability and ciliogenesis. The recent identification of ribosomes, RNA-binding proteins and transcripts at the centrosome suggests local protein synthesis. In this context, we hypothesized that TDP-43, a highly conserved RNA binding protein involved in the pathophysiology of amyotrophic lateral sclerosis and frontotemporal lobar degeneration, could be enriched at this organelle. Using dedicated high-magnification sub-diffraction microscopy on human cells, we discovered a novel localization of TDP-43 at the centrosome during all phases of the cell cycle. These results were confirmed on purified centrosomes by western blot and immunofluorescence microscopy. In addition, the colocalization of TDP-43 and pericentrin suggested a pericentriolar enrichment of the protein, leading us to hypothesize that TDP-43 might interact with local mRNAs and proteins. Supporting this hypothesis, we found 4 conserved centrosomal mRNAs and 16 centrosomal proteins identified as direct TDP-43 interactors. More strikingly, all the 16 proteins are implicated in the pathophysiology of TDP-43 proteinopathies, suggesting that TDP-43 dysfunction in this organelle contributes to neurodegeneration. This first description of TDP-43 centrosomal enrichment paves the way for a more comprehensive understanding of TDP-43 physiology and pathology.
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