https://orcid.org/0000-0003-2820-1982
Figures & data
Figure 1. The role of amygdala nerve activity in rats with neuropathic pain-like behaviors is related to NPY expression. A-B, Heatmap (a) and volcano map (b) of differentially expressed genes between samples of control (n = 3) and CCI (n = 12) rats in the GSE30691 dataset. (c) The logFC values of differentially expressed genes between samples of control and CCI rats in the GSE30691 dataset. FC, fold change. (d) The MWT of CCI rats with/without BLA kindling at different time points. * p < 0.05 vs. untreated CCI rats. (e) The MWT of CCI rats at different time points. * p < 0.05 vs. untreated CCI rats. (f)-(g) Statistics (f) and NPY immunohistochemistry-positive staining (g) of sciatic nerve tissue of the CCI rats and the sham-operated rats on the 1st, 7th, 14th, and 21st day after modeling (The arrow points to NPY positive cells). * p < 0.05 vs. sham-operated rats n = 5.
Figure 2. High expression of NPY occurred in the amygdala of rats with neuropathic pain-like behaviors. (a) The mRNA level of NPY in amygdala of the CCI rats and the sham-operated rats determined by RT-qPCR. (b) NPY expression in amygdala of the CCI rats and the sham-operated rats measured by immunofluorescence. (c) NPY protein level in amygdala of the CCI rats and the sham-operated rats determined by Western blot assay (n = 5).
Figure 3. NPY2R-mediated activation of MAPK signaling pathway is involved in neuropathic pain. (a) The mRNA levels of NPY1R and NPY2R in the CCI rats and the sham-operated rats at different time points determined by RT-qPCR. (b) The protein levels of NPY1R and NPY2R in the sham-operated rats and the CCI rats and the sham-operated rats at different time points determined by Western blot assay. (c) The expression of NPY1R and NPY2R in the amygdala of CCI rats and the sham-operated rats determined by immunofluorescence staining. (d) The binding of NPY and NPY2R in the CCI rats and the sham-operated rats as observed by co-IP 21 days after operation. (e) The protein levels of ERK, p38, and JNK in the CCI rats and the sham-operated rats at different time points determined by Western blot assay. (f) The MWT and TWL (0 represents before injection) and the expression changes of MAPK signaling pathway-related proteins after injection of NPY2R agonist and NPY2R antisense ODN in the CCI rats. (g) The changes of MWT and TWL in CCI rats injected with MAPK signaling pathway antagonists (ERK antagonist, JNK antagonist, and p38 MAPK antagonist) at different time points (n = 5).
Figure 4. NPY2R-activated MAPK signaling pathway is associated with microglia apoptosis. (a) The mRNA and protein levels of NPY2R in MN9D, C6, and BV-2 cells determined by RT-qPCR (left) and Western blot assay (right). (b) The knockdown effect of three shRNAs targeting NPY2R in BV-2 cells determined by RT-qPCR (left) and Western blot assay (right). (c) The expression effect of three NPY2R overexpression plasmids in BV-2 cells determined by RT-qPCR (left) and Western blot assay (right). (d) The viability of BV-2 cells in response to NPY2R overexpression or knockdown as examined by CCK-8 assay. (e) The viability of BV-2 cells in response to NPY2R overexpression or knockdown as examined by EdU assay. (f) The effect of NPY2R overexpression on the expression of MAPK signaling pathway related factors (ERK, p38, and JNK) in response to NPY2R overexpression or knockdown as determined by Western blot assay. (g) The apoptosis of BV-2 cells in response to NPY2R overexpression or knockdown as as examined by flow cytometry. Cell experiments were repeated three times independently.
