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Differential transcriptional profiling of damaged and intact adjacent dorsal root ganglia neurons in neuropathic pain.

Reinhold AK, Batti L, Bilbao D, Buness A, Rittner HL, Heppenstall PA - PLoS ONE (2015)

Bottom Line: Data for damaged neurons reveal an mRNA expression pattern consistent with established upregulated genes like galanin, which supports our approach.Moreover, novel genes were found strongly regulated such as corticotropin-releasing hormone (CRH), providing novel targets for further research.Differential fluorescent neuronal labelling and sorting allows for a clear distinction between primarily damaged neuropathic neurons and "bystanders," thereby facilitating a more detailed understanding of their respective roles in neuropathic processes in the DRG.

View Article: PubMed Central - PubMed

Affiliation: European Molecular Biology Laboratory, Monterotondo, Italy; Department of Anaesthesiology, University Hospital, Würzburg, Germany.

ABSTRACT
Neuropathic pain, caused by a lesion in the somatosensory system, is a severely impairing mostly chronic disease. While its underlying molecular mechanisms are not thoroughly understood, neuroimmune interactions as well as changes in the pain pathway such as sensitization of nociceptors have been implicated. It has been shown that not only are different cell types involved in generation and maintenance of neuropathic pain, like neurons, immune and glial cells, but, also, intact adjacent neurons are relevant to the process. Here, we describe an experimental approach to discriminate damaged from intact adjacent neurons in the same dorsal root ganglion (DRG) using differential fluorescent neuronal labelling and fluorescence-activated cell sorting (FACS). Two fluorescent tracers, Fluoroemerald (FE) and 1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine perchlorate (DiI), were used, whose properties allow us to distinguish between damaged and intact neurons. Subsequent sorting permitted transcriptional analysis of both groups. Results and qPCR validation show a strong regulation in damaged neurons versus contralateral controls as well as a moderate regulation in adjacent neurons. Data for damaged neurons reveal an mRNA expression pattern consistent with established upregulated genes like galanin, which supports our approach. Moreover, novel genes were found strongly regulated such as corticotropin-releasing hormone (CRH), providing novel targets for further research. Differential fluorescent neuronal labelling and sorting allows for a clear distinction between primarily damaged neuropathic neurons and "bystanders," thereby facilitating a more detailed understanding of their respective roles in neuropathic processes in the DRG.

No MeSH data available.


Related in: MedlinePlus

Microarray cluster analysis.Samples of damaged neurons (red) exhibit within-group similarity and considerable group difference to both adjacent (yellow) and contralateral (green) neurons. In contrast, these two groups cannot be discriminated from each other (hierarchical clustering with complete linkage for 200 probe sets with highest MAD. (Color key: row z-score -2 (dark blue) to +2 (white)).
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pone.0123342.g003: Microarray cluster analysis.Samples of damaged neurons (red) exhibit within-group similarity and considerable group difference to both adjacent (yellow) and contralateral (green) neurons. In contrast, these two groups cannot be discriminated from each other (hierarchical clustering with complete linkage for 200 probe sets with highest MAD. (Color key: row z-score -2 (dark blue) to +2 (white)).

Mentions: From sorted cell populations (ipsilateral DiI+/FE+, ipsilateral DiI+/FE- and contralateral DiI+ cells), RNA was purified and further analysed using the Affymetrix Gene Expression array. Cluster analysis of all three runs revealed a homogenous expression profile in damaged neurons Samples of adjacent, non-damaged neurons, in contrast, exhibit a broader within-group variety of gene expression (Fig 3).


Differential transcriptional profiling of damaged and intact adjacent dorsal root ganglia neurons in neuropathic pain.

Reinhold AK, Batti L, Bilbao D, Buness A, Rittner HL, Heppenstall PA - PLoS ONE (2015)

Microarray cluster analysis.Samples of damaged neurons (red) exhibit within-group similarity and considerable group difference to both adjacent (yellow) and contralateral (green) neurons. In contrast, these two groups cannot be discriminated from each other (hierarchical clustering with complete linkage for 200 probe sets with highest MAD. (Color key: row z-score -2 (dark blue) to +2 (white)).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4400143&req=5

pone.0123342.g003: Microarray cluster analysis.Samples of damaged neurons (red) exhibit within-group similarity and considerable group difference to both adjacent (yellow) and contralateral (green) neurons. In contrast, these two groups cannot be discriminated from each other (hierarchical clustering with complete linkage for 200 probe sets with highest MAD. (Color key: row z-score -2 (dark blue) to +2 (white)).
Mentions: From sorted cell populations (ipsilateral DiI+/FE+, ipsilateral DiI+/FE- and contralateral DiI+ cells), RNA was purified and further analysed using the Affymetrix Gene Expression array. Cluster analysis of all three runs revealed a homogenous expression profile in damaged neurons Samples of adjacent, non-damaged neurons, in contrast, exhibit a broader within-group variety of gene expression (Fig 3).

Bottom Line: Data for damaged neurons reveal an mRNA expression pattern consistent with established upregulated genes like galanin, which supports our approach.Moreover, novel genes were found strongly regulated such as corticotropin-releasing hormone (CRH), providing novel targets for further research.Differential fluorescent neuronal labelling and sorting allows for a clear distinction between primarily damaged neuropathic neurons and "bystanders," thereby facilitating a more detailed understanding of their respective roles in neuropathic processes in the DRG.

View Article: PubMed Central - PubMed

Affiliation: European Molecular Biology Laboratory, Monterotondo, Italy; Department of Anaesthesiology, University Hospital, Würzburg, Germany.

ABSTRACT
Neuropathic pain, caused by a lesion in the somatosensory system, is a severely impairing mostly chronic disease. While its underlying molecular mechanisms are not thoroughly understood, neuroimmune interactions as well as changes in the pain pathway such as sensitization of nociceptors have been implicated. It has been shown that not only are different cell types involved in generation and maintenance of neuropathic pain, like neurons, immune and glial cells, but, also, intact adjacent neurons are relevant to the process. Here, we describe an experimental approach to discriminate damaged from intact adjacent neurons in the same dorsal root ganglion (DRG) using differential fluorescent neuronal labelling and fluorescence-activated cell sorting (FACS). Two fluorescent tracers, Fluoroemerald (FE) and 1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine perchlorate (DiI), were used, whose properties allow us to distinguish between damaged and intact neurons. Subsequent sorting permitted transcriptional analysis of both groups. Results and qPCR validation show a strong regulation in damaged neurons versus contralateral controls as well as a moderate regulation in adjacent neurons. Data for damaged neurons reveal an mRNA expression pattern consistent with established upregulated genes like galanin, which supports our approach. Moreover, novel genes were found strongly regulated such as corticotropin-releasing hormone (CRH), providing novel targets for further research. Differential fluorescent neuronal labelling and sorting allows for a clear distinction between primarily damaged neuropathic neurons and "bystanders," thereby facilitating a more detailed understanding of their respective roles in neuropathic processes in the DRG.

No MeSH data available.


Related in: MedlinePlus