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Entrapment neuropathy results in different microRNA expression patterns from denervation injury in rats.

Rau CS, Jeng JC, Jeng SF, Lu TH, Chen YC, Liliang PC, Wu CJ, Lin CJ, Hsieh CH - BMC Musculoskelet Disord (2010)

Bottom Line: In the DRGs, 6 miRNAs in the entrapment group (miR-9, miR-320, miR-324-3p, miR-672, miR-466b, and miR-144) and 3 miRNAs in the decompression group (miR-9, miR-320, and miR-324-3p) were significantly downregulated.Regarding the muscle-specific miRNAs, real-time RT-PCR analysis revealed an approximately 50% decrease in miR-1 and miR-133a expression levels at 3 and 6 months after entrapment, whereas miR-1 and miR-133a levels were unchanged and were decreased after decompression at 1 and 3 months.In contrast, there were no statistical differences in the expression of miR-206 during nerve entrapment and after decompression.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neurosurgery, Chang Gung Memorial Hospital - Kaohsiung Medical Center, Chang Gung University College of Medicine, Taiwan.

ABSTRACT

Background: To compare the microRNA (miRNA) expression profiles in neurons and innervated muscles after sciatic nerve entrapment using a non-constrictive silastic tube, subsequent surgical decompression, and denervation injury.

Methods: The experimental L4-L6 spinal segments, dorsal root ganglia (DRGs), and soleus muscles from each experimental group (sham control, denervation, entrapment, and decompression) were analyzed using an Agilent rat miRNA array to detect dysregulated miRNAs. In addition, muscle-specific miRNAs (miR-1, -133a, and -206) and selectively upregulated miRNAs were subsequently quantified using real-time reverse transcription-polymerase chain reaction (real-time RT-PCR).

Results: In the soleus muscles, 37 of the 47 miRNAs (13.4% of the 350 unique miRNAs tested) that were significantly downregulated after 6 months of entrapment neuropathy were also among the 40 miRNAs (11.4% of the 350 unique miRNAs tested) that were downregulated after 3 months of decompression. No miRNA was upregulated in both groups. In contrast, only 3 miRNAs were upregulated and 3 miRNAs were downregulated in the denervated muscle after 6 months. In the DRGs, 6 miRNAs in the entrapment group (miR-9, miR-320, miR-324-3p, miR-672, miR-466b, and miR-144) and 3 miRNAs in the decompression group (miR-9, miR-320, and miR-324-3p) were significantly downregulated. No miRNA was upregulated in both groups. We detected 1 downregulated miRNA (miR-144) and 1 upregulated miRNA (miR-21) after sciatic nerve denervation. We were able to separate the muscle or DRG samples into denervation or entrapment neuropathy by performing unsupervised hierarchal clustering analysis. Regarding the muscle-specific miRNAs, real-time RT-PCR analysis revealed an approximately 50% decrease in miR-1 and miR-133a expression levels at 3 and 6 months after entrapment, whereas miR-1 and miR-133a levels were unchanged and were decreased after decompression at 1 and 3 months. In contrast, there were no statistical differences in the expression of miR-206 during nerve entrapment and after decompression. The expression of muscle-specific miRNAs in entrapment neuropathy is different from our previous observations in sciatic nerve denervation injury.

Conclusions: This study revealed the different involvement of miRNAs in neurons and innervated muscles after entrapment neuropathy and denervation injury, and implied that epigenetic regulation is different in these two conditions.

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Related in: MedlinePlus

In situ hybridization analysis. In comparison with the negative control (Figure 2A), in situ hybridization analysis revealed that the positive control probe rno-U6 was abundantly and diffusely expressed in the perinuclear region of the neurons in the denervated dorsal root ganglia (DRGs) (Figure 2B). Intense signals for miR-21 in the perinuclear region of the neurons were also observed in the tissue sections at 1 month after denervation injury by using digoxigenin-labeled miR-21 probes (Figure 2C). Real time RT-PCR revealed that the expression of miR-21 in the DRGs was increased by ~6 fold; it was detected 1 week after denervation and lasted for up to 6 months (Figure 2D). Bars represent means ± standard error of 6 independent experiments; *, P < 0.05 vs. sham control.
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Figure 2: In situ hybridization analysis. In comparison with the negative control (Figure 2A), in situ hybridization analysis revealed that the positive control probe rno-U6 was abundantly and diffusely expressed in the perinuclear region of the neurons in the denervated dorsal root ganglia (DRGs) (Figure 2B). Intense signals for miR-21 in the perinuclear region of the neurons were also observed in the tissue sections at 1 month after denervation injury by using digoxigenin-labeled miR-21 probes (Figure 2C). Real time RT-PCR revealed that the expression of miR-21 in the DRGs was increased by ~6 fold; it was detected 1 week after denervation and lasted for up to 6 months (Figure 2D). Bars represent means ± standard error of 6 independent experiments; *, P < 0.05 vs. sham control.

Mentions: In the DRGs, 6 miRNAs (miR-9, miR-320, miR-324-3p, miR-672, miR-466b, and miR-144) were significantly downregulated in the entrapment group and 3 miRNAs (miR-9, miR-320, and miR-324-3p) were significantly downregulated in the decompression group. Although there also appeared to be a decrease of miR-672, miR-466b, and miR-144 in the decompression group, it was not statistically significant in all 3 specimens. No miRNAs were upregulated in the DRGs of the entrapment and decompression groups. On the other hand, after sciatic nerve denervation, we observed 1 downregulated miRNA (miR-144) and 1 upregulated miRNA (miR-21) in the DRGs (Figure 1). To localize the expression of miR-21 in the DRGs, in situ hybridization of miR-21 was performed to differentiate whether the upregulation occurred in the neurons or the interstitial connective tissue. As shown in Figure 2, in comparison with the negative control (Figure 2A), the positive control probe rno-U6 was abundantly and diffusely expressed in the perinuclear region of the DRG neurons after 6 months of denervation (Figure 2B). In addition, using DIG-labeled miR-21 probes, intense signals for miR-21 were also observed in the perinuclear region of the neurons (Figure 2C). Real time RT-PCR revealed that the expression of miR-21 in the DRGs was detected after 1 week of denervation with an ~6-fold increase that lasted for up to 6 months (Figure 2D). We were able to separate the samples into denervation, entrapment, and decompression by performing unsupervised hierarchal clustering analysis.


Entrapment neuropathy results in different microRNA expression patterns from denervation injury in rats.

Rau CS, Jeng JC, Jeng SF, Lu TH, Chen YC, Liliang PC, Wu CJ, Lin CJ, Hsieh CH - BMC Musculoskelet Disord (2010)

In situ hybridization analysis. In comparison with the negative control (Figure 2A), in situ hybridization analysis revealed that the positive control probe rno-U6 was abundantly and diffusely expressed in the perinuclear region of the neurons in the denervated dorsal root ganglia (DRGs) (Figure 2B). Intense signals for miR-21 in the perinuclear region of the neurons were also observed in the tissue sections at 1 month after denervation injury by using digoxigenin-labeled miR-21 probes (Figure 2C). Real time RT-PCR revealed that the expression of miR-21 in the DRGs was increased by ~6 fold; it was detected 1 week after denervation and lasted for up to 6 months (Figure 2D). Bars represent means ± standard error of 6 independent experiments; *, P < 0.05 vs. sham control.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2927509&req=5

Figure 2: In situ hybridization analysis. In comparison with the negative control (Figure 2A), in situ hybridization analysis revealed that the positive control probe rno-U6 was abundantly and diffusely expressed in the perinuclear region of the neurons in the denervated dorsal root ganglia (DRGs) (Figure 2B). Intense signals for miR-21 in the perinuclear region of the neurons were also observed in the tissue sections at 1 month after denervation injury by using digoxigenin-labeled miR-21 probes (Figure 2C). Real time RT-PCR revealed that the expression of miR-21 in the DRGs was increased by ~6 fold; it was detected 1 week after denervation and lasted for up to 6 months (Figure 2D). Bars represent means ± standard error of 6 independent experiments; *, P < 0.05 vs. sham control.
Mentions: In the DRGs, 6 miRNAs (miR-9, miR-320, miR-324-3p, miR-672, miR-466b, and miR-144) were significantly downregulated in the entrapment group and 3 miRNAs (miR-9, miR-320, and miR-324-3p) were significantly downregulated in the decompression group. Although there also appeared to be a decrease of miR-672, miR-466b, and miR-144 in the decompression group, it was not statistically significant in all 3 specimens. No miRNAs were upregulated in the DRGs of the entrapment and decompression groups. On the other hand, after sciatic nerve denervation, we observed 1 downregulated miRNA (miR-144) and 1 upregulated miRNA (miR-21) in the DRGs (Figure 1). To localize the expression of miR-21 in the DRGs, in situ hybridization of miR-21 was performed to differentiate whether the upregulation occurred in the neurons or the interstitial connective tissue. As shown in Figure 2, in comparison with the negative control (Figure 2A), the positive control probe rno-U6 was abundantly and diffusely expressed in the perinuclear region of the DRG neurons after 6 months of denervation (Figure 2B). In addition, using DIG-labeled miR-21 probes, intense signals for miR-21 were also observed in the perinuclear region of the neurons (Figure 2C). Real time RT-PCR revealed that the expression of miR-21 in the DRGs was detected after 1 week of denervation with an ~6-fold increase that lasted for up to 6 months (Figure 2D). We were able to separate the samples into denervation, entrapment, and decompression by performing unsupervised hierarchal clustering analysis.

Bottom Line: In the DRGs, 6 miRNAs in the entrapment group (miR-9, miR-320, miR-324-3p, miR-672, miR-466b, and miR-144) and 3 miRNAs in the decompression group (miR-9, miR-320, and miR-324-3p) were significantly downregulated.Regarding the muscle-specific miRNAs, real-time RT-PCR analysis revealed an approximately 50% decrease in miR-1 and miR-133a expression levels at 3 and 6 months after entrapment, whereas miR-1 and miR-133a levels were unchanged and were decreased after decompression at 1 and 3 months.In contrast, there were no statistical differences in the expression of miR-206 during nerve entrapment and after decompression.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Neurosurgery, Chang Gung Memorial Hospital - Kaohsiung Medical Center, Chang Gung University College of Medicine, Taiwan.

ABSTRACT

Background: To compare the microRNA (miRNA) expression profiles in neurons and innervated muscles after sciatic nerve entrapment using a non-constrictive silastic tube, subsequent surgical decompression, and denervation injury.

Methods: The experimental L4-L6 spinal segments, dorsal root ganglia (DRGs), and soleus muscles from each experimental group (sham control, denervation, entrapment, and decompression) were analyzed using an Agilent rat miRNA array to detect dysregulated miRNAs. In addition, muscle-specific miRNAs (miR-1, -133a, and -206) and selectively upregulated miRNAs were subsequently quantified using real-time reverse transcription-polymerase chain reaction (real-time RT-PCR).

Results: In the soleus muscles, 37 of the 47 miRNAs (13.4% of the 350 unique miRNAs tested) that were significantly downregulated after 6 months of entrapment neuropathy were also among the 40 miRNAs (11.4% of the 350 unique miRNAs tested) that were downregulated after 3 months of decompression. No miRNA was upregulated in both groups. In contrast, only 3 miRNAs were upregulated and 3 miRNAs were downregulated in the denervated muscle after 6 months. In the DRGs, 6 miRNAs in the entrapment group (miR-9, miR-320, miR-324-3p, miR-672, miR-466b, and miR-144) and 3 miRNAs in the decompression group (miR-9, miR-320, and miR-324-3p) were significantly downregulated. No miRNA was upregulated in both groups. We detected 1 downregulated miRNA (miR-144) and 1 upregulated miRNA (miR-21) after sciatic nerve denervation. We were able to separate the muscle or DRG samples into denervation or entrapment neuropathy by performing unsupervised hierarchal clustering analysis. Regarding the muscle-specific miRNAs, real-time RT-PCR analysis revealed an approximately 50% decrease in miR-1 and miR-133a expression levels at 3 and 6 months after entrapment, whereas miR-1 and miR-133a levels were unchanged and were decreased after decompression at 1 and 3 months. In contrast, there were no statistical differences in the expression of miR-206 during nerve entrapment and after decompression. The expression of muscle-specific miRNAs in entrapment neuropathy is different from our previous observations in sciatic nerve denervation injury.

Conclusions: This study revealed the different involvement of miRNAs in neurons and innervated muscles after entrapment neuropathy and denervation injury, and implied that epigenetic regulation is different in these two conditions.

Show MeSH
Related in: MedlinePlus