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Expression of DNA methyltransferases in adult dorsal root ganglia is cell-type specific and up regulated in a rodent model of neuropathic pain.

Pollema-Mays SL, Centeno MV, Apkarian AV, Martina M - Front Cell Neurosci (2014)

Bottom Line: Real-time RT PCR analysis revealed robust and time-dependent changes in DNMT transcript expression in ipsilateral DRGs from spared nerve injury (SNI) but not sham rats.Interestingly, DNMT3b transcript showed a robust upregulation that appeared already 1 week after surgery and persisted at 4 weeks (our endpoint); in contrast, DNMT1 and DNMT3a transcripts showed only moderate upregulation that was transient and did not appear until the second week.We suggest that DNMT regulation in adult DRGs may be a contributor to the pain phenotype and merits further study.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Northwestern University Feinberg School of Medicine Chicago, IL, USA.

ABSTRACT
Neuropathic pain is associated with hyperexcitability and intrinsic firing of dorsal root ganglia (DRG) neurons. These phenotypical changes can be long lasting, potentially spanning the entire life of animal models, and depend on altered expression of numerous proteins, including many ion channels. Yet, how DRGs maintain long-term changes in protein expression in neuropathic conditions remains unclear. DNA methylation is a well-known mechanism of epigenetic control of gene expression and is achieved by the action of three enzymes: DNA methyltransferase (DNMT) 1, 3a, and 3b, which have been studied primarily during development. We first performed immunohistochemical analysis to assess whether these enzymes are expressed in adult rat DRGs (L4-5) and found that DNMT1 is expressed in both glia and neurons, DNMT3a is preferentially expressed in glia and DNMT3b is preferentially expressed in neurons. A rat model of neuropathic pain was then used to determine whether nerve injury may induce epigenetic changes in DRGs at multiple time points after pain onset. Real-time RT PCR analysis revealed robust and time-dependent changes in DNMT transcript expression in ipsilateral DRGs from spared nerve injury (SNI) but not sham rats. Interestingly, DNMT3b transcript showed a robust upregulation that appeared already 1 week after surgery and persisted at 4 weeks (our endpoint); in contrast, DNMT1 and DNMT3a transcripts showed only moderate upregulation that was transient and did not appear until the second week. We suggest that DNMT regulation in adult DRGs may be a contributor to the pain phenotype and merits further study.

No MeSH data available.


Related in: MedlinePlus

DNA methyltransferase expression in DRG is cell-type specific. 20-micron L4 and L5 DRG sections from control rats aged 5–7 weeks were immuno-labeled with DNMT antibodies as well as incubated with the nuclear labeling dye 4’,6-diamidino-2-phenylindole (DAPI). (A–C) Show robust and ubiquitous labeling of DNMT1 in nuclei of DRG neurons. Notably, there is little expression of DNMT1 detected in the satellite cells surrounding DRG neurons. In addition, widespread overlap of DAPI and DNMT1 was detected in axons, demonstrating ubiquitous expression in Schwann cells (D–F). (G–I) In contrast to DNMT1, DNMT3a was not detected in neuronal nuclei but rather in satellite cells surrounding DRG neurons. (J–L) DNMT3a also showed widespread overlap with DAPI in nuclei of Schwann cells. (M–O) DNMT3b was detected in nuclei of all DRG neurons. (P–R) Notably, DNMT3b staining did not reveal overlap with DAPI in DRG axons, suggesting DNMT3b is not expressed in glia. Scale bar: 20 μM.
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Figure 1: DNA methyltransferase expression in DRG is cell-type specific. 20-micron L4 and L5 DRG sections from control rats aged 5–7 weeks were immuno-labeled with DNMT antibodies as well as incubated with the nuclear labeling dye 4’,6-diamidino-2-phenylindole (DAPI). (A–C) Show robust and ubiquitous labeling of DNMT1 in nuclei of DRG neurons. Notably, there is little expression of DNMT1 detected in the satellite cells surrounding DRG neurons. In addition, widespread overlap of DAPI and DNMT1 was detected in axons, demonstrating ubiquitous expression in Schwann cells (D–F). (G–I) In contrast to DNMT1, DNMT3a was not detected in neuronal nuclei but rather in satellite cells surrounding DRG neurons. (J–L) DNMT3a also showed widespread overlap with DAPI in nuclei of Schwann cells. (M–O) DNMT3b was detected in nuclei of all DRG neurons. (P–R) Notably, DNMT3b staining did not reveal overlap with DAPI in DRG axons, suggesting DNMT3b is not expressed in glia. Scale bar: 20 μM.

Mentions: Immunochemical analysis was performed on 20 μm-thick slices obtained from L4 and L5 DRGs of 5–7 week old naïve rats to study the expression of the DNMT1, DNMT3a, and DNMT3b in L4 and L5 DRGs of adult rats. Immunostaining of DNMT1 demonstrated that expression is robust and is detected ubiquitously in the nuclei of DRG neurons as well as in Schwann cells (Figure 1). Indeed labeling with the nuclear dye 4′,6-diamidino-2-phenylindole (DAPI) revealed almost perfect overlap with DNMT1 expression in both DRG neuronal nuclei (Figures 1A–C, see arrowheads for examples) and the nuclei of Schwann cells surrounding DRG axons (Figures 1D–F). Interestingly, DNMT1 expression was not detected in the satellite cells that surround and support DRG neuronal cell bodies (Figures 1A–C).


Expression of DNA methyltransferases in adult dorsal root ganglia is cell-type specific and up regulated in a rodent model of neuropathic pain.

Pollema-Mays SL, Centeno MV, Apkarian AV, Martina M - Front Cell Neurosci (2014)

DNA methyltransferase expression in DRG is cell-type specific. 20-micron L4 and L5 DRG sections from control rats aged 5–7 weeks were immuno-labeled with DNMT antibodies as well as incubated with the nuclear labeling dye 4’,6-diamidino-2-phenylindole (DAPI). (A–C) Show robust and ubiquitous labeling of DNMT1 in nuclei of DRG neurons. Notably, there is little expression of DNMT1 detected in the satellite cells surrounding DRG neurons. In addition, widespread overlap of DAPI and DNMT1 was detected in axons, demonstrating ubiquitous expression in Schwann cells (D–F). (G–I) In contrast to DNMT1, DNMT3a was not detected in neuronal nuclei but rather in satellite cells surrounding DRG neurons. (J–L) DNMT3a also showed widespread overlap with DAPI in nuclei of Schwann cells. (M–O) DNMT3b was detected in nuclei of all DRG neurons. (P–R) Notably, DNMT3b staining did not reveal overlap with DAPI in DRG axons, suggesting DNMT3b is not expressed in glia. Scale bar: 20 μM.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 1: DNA methyltransferase expression in DRG is cell-type specific. 20-micron L4 and L5 DRG sections from control rats aged 5–7 weeks were immuno-labeled with DNMT antibodies as well as incubated with the nuclear labeling dye 4’,6-diamidino-2-phenylindole (DAPI). (A–C) Show robust and ubiquitous labeling of DNMT1 in nuclei of DRG neurons. Notably, there is little expression of DNMT1 detected in the satellite cells surrounding DRG neurons. In addition, widespread overlap of DAPI and DNMT1 was detected in axons, demonstrating ubiquitous expression in Schwann cells (D–F). (G–I) In contrast to DNMT1, DNMT3a was not detected in neuronal nuclei but rather in satellite cells surrounding DRG neurons. (J–L) DNMT3a also showed widespread overlap with DAPI in nuclei of Schwann cells. (M–O) DNMT3b was detected in nuclei of all DRG neurons. (P–R) Notably, DNMT3b staining did not reveal overlap with DAPI in DRG axons, suggesting DNMT3b is not expressed in glia. Scale bar: 20 μM.
Mentions: Immunochemical analysis was performed on 20 μm-thick slices obtained from L4 and L5 DRGs of 5–7 week old naïve rats to study the expression of the DNMT1, DNMT3a, and DNMT3b in L4 and L5 DRGs of adult rats. Immunostaining of DNMT1 demonstrated that expression is robust and is detected ubiquitously in the nuclei of DRG neurons as well as in Schwann cells (Figure 1). Indeed labeling with the nuclear dye 4′,6-diamidino-2-phenylindole (DAPI) revealed almost perfect overlap with DNMT1 expression in both DRG neuronal nuclei (Figures 1A–C, see arrowheads for examples) and the nuclei of Schwann cells surrounding DRG axons (Figures 1D–F). Interestingly, DNMT1 expression was not detected in the satellite cells that surround and support DRG neuronal cell bodies (Figures 1A–C).

Bottom Line: Real-time RT PCR analysis revealed robust and time-dependent changes in DNMT transcript expression in ipsilateral DRGs from spared nerve injury (SNI) but not sham rats.Interestingly, DNMT3b transcript showed a robust upregulation that appeared already 1 week after surgery and persisted at 4 weeks (our endpoint); in contrast, DNMT1 and DNMT3a transcripts showed only moderate upregulation that was transient and did not appear until the second week.We suggest that DNMT regulation in adult DRGs may be a contributor to the pain phenotype and merits further study.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, Northwestern University Feinberg School of Medicine Chicago, IL, USA.

ABSTRACT
Neuropathic pain is associated with hyperexcitability and intrinsic firing of dorsal root ganglia (DRG) neurons. These phenotypical changes can be long lasting, potentially spanning the entire life of animal models, and depend on altered expression of numerous proteins, including many ion channels. Yet, how DRGs maintain long-term changes in protein expression in neuropathic conditions remains unclear. DNA methylation is a well-known mechanism of epigenetic control of gene expression and is achieved by the action of three enzymes: DNA methyltransferase (DNMT) 1, 3a, and 3b, which have been studied primarily during development. We first performed immunohistochemical analysis to assess whether these enzymes are expressed in adult rat DRGs (L4-5) and found that DNMT1 is expressed in both glia and neurons, DNMT3a is preferentially expressed in glia and DNMT3b is preferentially expressed in neurons. A rat model of neuropathic pain was then used to determine whether nerve injury may induce epigenetic changes in DRGs at multiple time points after pain onset. Real-time RT PCR analysis revealed robust and time-dependent changes in DNMT transcript expression in ipsilateral DRGs from spared nerve injury (SNI) but not sham rats. Interestingly, DNMT3b transcript showed a robust upregulation that appeared already 1 week after surgery and persisted at 4 weeks (our endpoint); in contrast, DNMT1 and DNMT3a transcripts showed only moderate upregulation that was transient and did not appear until the second week. We suggest that DNMT regulation in adult DRGs may be a contributor to the pain phenotype and merits further study.

No MeSH data available.


Related in: MedlinePlus