BackgroundDeep brain stimulation (DBS) of the subthalamic nucleus (STN) or the internal segment of the globus pallidus (GPi) has been established as a highly effective symptomatic therapy for Parkinson’s disease (PD). An intriguing biological aspect related to the DBS procedure is that a temporary contact establishes between surgical instruments and the surrounding brain tissue. In this exploratory study, we took advantage of this unique context to harvest brain material adhering to the stylet routinely used during surgery, and to examine the biological value of these samples, here referred to as “brain tissue imprints” (BTIs).ResultsNineteen BTIs from 12 STN- or GPi-electrode implanted patients were obtained in vivo during DBS surgery, without any modification of the surgical procedure. Immunofluorescence analyses confirmed that our approach allowed the harvesting of many neural cells including neurons harboring distinct neurotransmitter markers. Shotgun proteomic and transcriptomic analyses provided for the first time molecular information from DBS-associated brain samples, and confirmed the compatibility of this new type of sample with poly-omic approaches. The method appears to be safe and results consistent.ConclusionsWe here propose BTIs as original and highly valuable brain samples, and DBS-related brain imprinting as a new conceptual approach to biological research in living patients with PD.Electronic supplementary materialThe online version of this article (doi:10.1186/s13024-016-0077-4) contains supplementary material, which is available to authorized users.
Dopaminergic neurons (DA) of the substantia nigra pars compacta (SNpc) selectively and progressively degenerate in Parkinson’s disease (PD). Until now, molecular analyses of DA in PD have been limited to genomic or transcriptomic approaches, whereas, to the best of our knowledge, no proteomic or combined multiomic study examining the protein profile of these neurons is currently available. In this exploratory study, we used laser capture microdissection to extract regions from DA in 10 human SNpc obtained at autopsy in PD patients and control subjects. Extracted RNA and proteins were identified by RNA sequencing and nanoliquid chromatography–mass spectrometry, respectively, and the differential expression between PD and control group was assessed. Qualitative analyses confirmed that the microdissection protocol preserves the integrity of our samples and offers access to specific molecular pathways. This multiomic analysis highlighted differential expression of 52 genes and 33 proteins, including molecules of interest already known to be dysregulated in PD, such as LRP2, PNMT, CXCR4, MAOA and CBLN1 genes, or the Aldehyde dehydrogenase 1 protein. On the other hand, despite the same samples were used for both analyses, correlation between RNA and protein expression was low, as exemplified by the CST3 gene encoding for the cystatin C protein. This is the first exploratory study analyzing both gene and protein expression of laser-dissected neuronal parts from SNpc in PD. Data are available via ProteomeXchange with identifier PXD024748 and via GEO with identifier GSE 169755.
Besides genetic abnormalities in MPN patients, several studies have reported alterations in protein expression that could contribute towards the clinical phenotype. However, little is known about protein modifications in Ph MPN erythrocytes. In this context, we used a quantitative mass spectrometry proteomics approach to study the MPN erythrocyte proteome. LC-MS/MS (LTQ Orbitrap) analysis led to the identification of 51 and 86 overexpressed proteins in Polycythemia Vera and Essential Thrombocythemia respectively, compared with controls. Functional comparison using pathway analysis software showed that the Rho GTPase family signaling pathways were deregulated in MPN patients. In particular, IQGAP1 was significantly overexpressed in MPNs compared with controls. Additionally, Western-blot analysis not only confirmed IQGAP1 overexpression, but also showed that IQGAP1 levels depended on the patient's genotype. Moreover, we found that in JAK2V617F patients IQGAP1 could bind RhoA, Rac1 and Cdc42 and consequently recruit activated GTP-Rac1 and the cytoskeleton motility protein PAK1. In CALR(+) patients, IQGAP1 was not overexpressed but immunoprecipitated with RhoGDI. In JAK2V617F transduced Ba/F3 cells we confirmed JAK2 inhibitor-sensitive overexpression of IQGAP1/PAK1. Altogether, our data demonstrated alterations of IQGAP1/Rho GTPase signaling in MPN erythrocytes dependent on JAK2/CALR status, reinforcing the hypothesis that modifications in erythrocyte signaling pathways participate in Ph MPN pathogenesis.
Nanoparticle (NP)–protein interactions in complex samples have not yet been clearly understood. Nevertheless, several studies demonstrated that NP’s physicochemical features significantly impact on the protein corona composition. Taking advantage of the NP potential to harvest different subsets of proteins, we assessed for the first time the capacity of three kinds of superparamagnetic NPs to highlight the erythrocyte minor proteome. Using both qualitative and quantitative proteomics approaches, nano-liquid chromatography–tandem mass spectrometry allowed the identification of 893 different proteins, confirming the reproducible capacity of NPs to increase the number of identified proteins, through a reduction of the sample concentration range and the capture of specific proteins on the three different surfaces. These NP-specific protein signatures revealed significant differences in their isoelectric point and molecular weight. Moreover, this NP strategy offered a deeper access to the erythrocyte proteome highlighting several signaling pathways implicated in important erythrocyte functions. The automated potentiality, the reproducibility, and the low-consuming sample demonstrate the strong compatibility of our strategy for large-scale clinical studies and may become a standardized sample preparation in future erythrocyte-associated proteomics studies.
Background Dopaminergic (DA) neurons of the substantia nigra pars compacta (SNpc) selectively and progressively degenerate in Parkinson’s disease (PD). Until now, molecular analyses of DA neurons in PD have been limited to genomic and transcriptomic approaches, whereas, to the best of our knowledge, no proteomic or combined polyomic study examining the protein profile of these neurons, is currently available. Methods In this exploratory study, we used laser microdissection to extract DA neurons from 10 human SNpc samples obtained at autopsy in PD patients and control subjects. Extracted RNA and proteins were identified by RNA sequencing and nano-LC-MS/MS, respectively, and the differential expression between the PD and control group was assessed. Results Qualitative analyses confirmed that the microdissection protocol preserves the integrity of our samples and offers access to specific molecular pathways. This polyomic analysis highlighted differential expression of 52 genes and 33 proteins, including molecules of interest already known to be dysregulated in PD, such as LRP2, PNMT, CXCR4, MAOA and CBLN1 genes, or the Aldehyde dehydrogenase 1 protein. On the other hand, despite the same samples were used for both analyses, correlation between RNA and protein expression was low, as exemplified by the CST3 gene encoding for the cystatin C protein. Conclusion This is the first exploratory study analyzing both gene and protein expression of LMD-dissected DA neurons from SNpc in PD. Although correlation between RNA and protein expressions was limited, this polyomic study provides an extensive and integrated overview of molecular changes identified in the PD SNpc and may offer novel insights into specific pathological processes at work in PD degeneration.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.