Search

Search Results

1 results found. Results per page: [ 20 ][ 40 ][ 60 ][ 80 ][ 100 ][ 200 ][ 300 ]

Sort by: [ Publication Date ][ Score ]

Year of publication: [ 2025 ][ 2024 ][ 2023 ][ 2022 ][ 2021 ][ 2020 ][ 2019 ][ 2018 ][ 2017 ][ 2016 ][ 2015 ][ 2014 ][ 2013 ][ 2012 ][ 2011 ][ 2010 ][ 2009 ][ Any ]

Direction: [ Desc ][ Asc ]

• Research Paper Volume 12, Issue 11 pp 11004-11024

  Upregulation of Mlxipl induced by cJun in the spinal dorsal horn after peripheral nerve injury counteracts mechanical allodynia by inhibiting neuroinflammation

  Relevance score: 16.163801

  Hongrui Zhan, Yaping Wang, Shi Yu, Guiyuan Cai, Yanyan Zeng, Junqin Ma, Wei Liu, Wen Wu

  Keywords: mechanical allodynia, neuroinflammation, microglia, Mlxipl, cJun

  Published in Aging on June 9, 2020

  Show abstract
  Hide abstract

  Mlxipl regulates glucose metabolism, lipogenesis and tumorigenesis and has a wide-ranging impact on human health and disease. However, the role of Mlxipl in neuropathic pain remains unknown. In this study, we found that Mlxipl was increased in the ipsilateral L4–L6 spinal dorsal horn after Spared Nerve Injury surgery. Knockdown of Mlxipl in the ipsilateral L4–L6 spinal dorsal horn by intraspinal microinjection aggravated Spared Nerve Injury-induced mechanical allodynia and inflammation in the spinal dorsal horn, on the contrary, overexpression of Mlxipl inhibited mechanical allodynia and inflammation. Subsequently, the rat Mlxipl promoter was analyzed using bioinformatics methods to predict the upstream transcription factor cJun. Luciferase assays and ChIP-qPCR confirmed that cJun bound to the promoter of Mlxipl and enhanced its expression. Finally, we demonstrated that Mlxipl inhibited the inflammatory responses of lipopolysaccharide-induced microglia and that Mlxipl was regulated by the transcription factor cJun. These findings suggested that cJun-induced Mlxipl upregulation in the spinal dorsal horn after peripheral nerve injury provided a protective mechanism for the development and progression of neuropathic pain by inhibiting microglial-derived neuroinflammation. Targeting Mlxipl in the spinal dorsal horn might represent an effective strategy for the treatment of neuropathic pain.

  

  Mlxipl was upregulated in ipsilateral SDH of SNI-induced mechanical allodynia rats. (A and B) The Von Frey test were carried out before and after surgery (Sham or SNI). Mechanical allodynia was induced by SNI surgery in the ipsilateral paw. N = 5. ***P < 0.001 vs. SNI BL Ipsi; ^$$$P < 0.001, ^$$P < 0.01 vs. SNI Contra; ^###P < 0.001 vs. Sham Ipsi. QPCR (C) and western blot (D) western blot (E) were performed before and after surgery. Mlxipl in the ipsilateral SDH was upregulated after SNI surgery. Quantification of the western blot (F). N = 5. ***P < 0.001, **P < 0.01 vs. SNI BL Ipsi; ^$$$P < 0.001, ^$$P < 0.01 vs. SNI BL Contra. (G) and immunofluorescence (I) were performed at day 7 after Sham or SNI surgery. Mlxipl was significantly upregulated in the ipsilateral SDH. Quantification of the western blot (H) and immunofluorescence (J). N = 5. ^$$$P < 0.001, ^$$P < 0.01 vs. SNI Contra; ^###P < 0.001, ^##P < 0.01 vs. Sham Ipsi. (K) Double immunofluorescence staining showed that Mlxipl was co-localized with Iba1 (white arrows) and NeuN (yellow arrows) but not GFAP. BL, baseline (before surgery); ipsi, ipsilateral; Contra, contralateral; SNI, spare nerve injury; SDH, spinal dorsal horn.

  
  
  

  

  Knockdown of Mlxipl in the SDH promoted mechanical allodynia and neuroinflammation. (A) A schematic that illustrates the timing of the main experimental procedures. (B) The injection location was confirmed by intraspinal microinjection of trypan blue. (C–G) Mlxipl in the ipsilateral SDH was knockdown by intraspinal microinjection of shMLXIPL. SNI surgery was performed on day 28 after intraspinal microinjection. QPCR (C), western blot (D) and immunofluorescence (F) were performed at day 7 after SNI surgery. Quantification of western blot (E) and immunofluorescence (G). N = 5. ***P < 0.001, **P < 0.01 vs. SNI Ipsi; ^###P < 0.001, ^##P < 0.01 vs. SNI vehicle Ipsi; ^$$$P < 0.001, ^$$P < 0.01 vs. SNI shNC Ipsi. (H and I) Knockdown of Mlxipl promoted mechanical allodynia in the ipsilateral paw. The Von Frey test was performed before and after SNI surgery with or without pre-microinjection. N = 5. **P < 0.01, *P < 0.05 vs. SNI Ipsi. ^##P < 0.01, ^#P < 0.05 vs. SNI vehicle Ipsi; ^$$P < 0.01, ^$P < 0.05 vs. SNI shNC Ipsi. (J–L) Knockdown of Mlxipl promoted neuroinflammation in the ipsilateral SDH. Proinflammatory cytokines were detected using qPCR at day 7 after SNI surgery. N = 5. ***P < 0.001 vs. SNI Ipsi; ^###P < 0.001 vs. SNI vehicle Ipsi; ^$$$P < 0.001 vs. SNI shNC Ipsi. BFMI, before microinjection; BL, baseline (before surgery); ipsi, ipsilateral; Contra, contralateral; SNI, spare nerve injury; SDH, spinal dorsal horn.

  
  
  

  

  Overexpression of Mlxipl in the SDH inhibited mechanical allodynia and neuroinflammation. (A–E) Mlxipl in the ipsilateral SDH was overexpressed by intraspinal microinjection of OeMLXIPL. SNI surgery was performed on day 28 after intraspinal microinjection. QPCR (A), western blot (B) and immunofluorescence (D) were performed at day 7 after SNI surgery. Quantification of western blot (C) and immunofluorescence (E). N = 5. ***P < 0.001 vs. SNI Ipsi; ^###P < 0.001 vs. SNI vehicle Ipsi; ^$$$P < 0.001 vs. SNI shNC Ipsi. (F and G) Overexpression of Mlxipl inhibited mechanical allodynia in the ipsilateral paw. The Von Frey test was performed before and after SNI surgery with or without pre-microinjection. N = 5. **P < 0.01, *P < 0.05 vs. SNI Ipsi 0D; ^##P < 0.01, ^#P < 0.05 vs. SNI vehicle Ipsi 0D; ^$$P < 0.01, ^$P < 0.05 vs. SNI OeNC Ipsi 0D. (H–J) Overexpression of Mlxipl inhibited neuroinflammation in the ipsilateral SDH. Proinflammatory cytokines were detected using qPCR at day 7 after SNI surgery. N = 5. ***P < 0.001 vs. SNI Ipsi; ^###P < 0.001 vs. SNI vehicle Ipsi; ^$$$P < 0.001 vs. SNI shNC Ipsi. BFMI, before microinjection; BL, baseline (before surgery); ipsi, ipsilateral; Contra, contralateral; SNI, spare nerve injury; SDH, spinal dorsal horn.

  
  
  

  

  cJun directly regulated Mlxipl expression at the transcriptional level. (A) Schematic diagrams showed the Mlxipl promoter and its fragments containing predicting binding sites. The diagrams show the Mlxipl promoter and its fragments, which were used to predict potential binding sites. (B) Luciferase reporter assays of full-length Mlxipl reporter. PcDNA3.1-cJun or pcDNA3.1-NC was co-transfected with full-length Mlxipl reporter in microglia. PcDNA3.1-cJun promoted luciferase activity of full-length Mlxipl reporter luciferase activity. The data were relative to the pcDNA3.1-NC group. N = 3. ***P < 0.001 vs. pcDNA3.1-NC. (C) Luciferase reporter assays of fragments of Mlxipl reporter. PcDNA3.1-cJun or pcDNA3.1-NC was co-transfected with fragmented Mlxipl promoters (pGL3-Mlxipl1, pGL3-Mlxipl2 and pGL3-Mlxipl3), respectively. PcDNA3.1-cJun promoted luciferase activity of pGL3-Mlxipl3 reporter luciferase activity. N = 3. **P < 0.01 vs. pcDNA3.1-NC. (D) Luciferase reporter assays of mutation of pGL3-Mlxipl3 reporter. PcDNA3.1-cJun or pcDNA3.1-NC was co-transfected with pGL3-Mlxipl3 reporter with or without mutation. N = 3. **P < 0.01 vs. pcDNA3.1-cJun+pGL3-Mlxipl3. (E and F) ChIP assays with anti-cJun antibody were performed followed by qPCR or agarose gel electrophoresis. The DNA product of ChIP was detected by fragmented Mlxipl promoter primers. PcDAN3.1-cJun transfection enriched more Mlxipl3 fragments. IgG was used as a negative control; RNA pol II was used as a positive control system; ChIP DNA mixture was used as Input, and the sample loading was 10% of other groups. Quantification of agarose gel electrophoresis (G). N= 3. *P < 0.05, **P < 0.01 vs. pcDNA3.1-NC. TSS, transcription starting site.

  
  
  

  

  cJun-induced Mlxipl upregulation protectively inhibited inflammation in the SDH and improved mechanical allodynia. (A) The schematic illustrates the timing of the main experimental process. (B and D–F) QPCR (B) and western blot (D) were performed before and after surgery. CJun or p-cJun in the ipsilateral SDH was upregulated after SNI surgery. Quantification of the western blot (E-F). N = 5. ***P < 0.001, **P < 0.01, *P < 0.05 vs. SNI BL Ipsi; ^$$$P < 0.001, ^$$P < 0.01, ^$P < 0.05 vs. SNI BL Contra. (C and G–J) QPCR(C), western blot (G) and immunofluorescence (I) were performed at day 7 after Sham or SNI surgery. CJun or p-cJun was significantly upregulated in the ipsilateral SDH. cJun was co-localized with Mlxipl (white arrows). Quantification of the western blot (H) and immunofluorescence (J). N = 5. ^$$$P < 0.001, ^$$P < 0.01 vs. SNI Contra; ^###P < 0.001, ^##P < 0.01 vs. Sham Ipsi. (K–O) Intrathecal injections of cJun small interfering RNA (sicJun) or lentivirus encoding cJun (LvcJun) were performed at day 7 after SNI surgery. The injections were carried out once a day for 7 consecutive days and then cJun expression was detected by qPCR (K) and western blot (L). Knockdown of cJun inhibited the expression of Mlxipl and p-cJun. Overexpression of cJun promoted the expression of Mlxipl and p-cJun (L-O). Quantification of the western blots (M–O). N = 5. ^$$$P < 0.001, ^$$P < 0.01 vs. vehicle; ^###P < 0.001, ^##P < 0.01 vs. sicJun+OeMlxipl; ***P < 0.001, ^**P < 0.01 vs. LvcJun+shMlxipl. (P–S) The effect of cJun on neuroinflammation (P–R) and mechanical allodynia (S), with or without the pre-microinjection of OeMlxipl or shMlxipl. N = 5. ^$$$P < 0.001, ^$$P < 0.01 vs. vehicle; ^###P < 0.001, ^##P < 0.01 vs. sicJun+OeMlxipl; ***P < 0.001, ^**P < 0.01 vs. LvcJun+shMlxipl. BL, baseline (before surgery); ipsi, ipsilateral; Contra, contralateral; SNI, spare nerve injury; SDH, spinal dorsal horn.

  
  
  

  

  Mlxipl inhibited the cJun-mediated inflammatory response in microglia. (A–F) The Mlxipl expression was detected by QPCR (A–B), western blot (E) and immunofluorescence (E). The primary microglia were co-transfected with siMlxipl or LvMlxipl for 48 hr, and then treated with LPS (1 μg/ml) for 24 hr. Quantification of the western blot (D) and immunofluorescence (F). Data relative to vehicle. N = 3. ^$$$P < 0.001, ^$$P < 0.01, ^$P < 0.05 vs. LPS; ^###P < 0.001, ^##P < 0.01 vs. LPS-siNC; ***P < 0.001, **P < 0.01, *P < 0.05 vs. LPS-LvNC. (G–I) ELISA were performed to detect the expression of proinflammation cytokines. Mlxipl inhibited inflammation response in LPS-induced microglia. N = 3. ^$$$P < 0.001, ^$$P < 0.01, ^$P < 0.05 vs. LPS; ^###P < 0.001, ^##P < 0.01 vs. LPS-siNC; ***P < 0.001, **P < 0.01, *P < 0.05 vs. LPS-LvNC. (J–O) The cJun expression was detected by QPCR (J–K), western blot (L) and immunofluorescence (N). The primary microglia were co-transfected with sicJun or LvcJun for 48 hr, and then treated with LPS (1 μg/ml) for 24 hr. Knockdown of cJun inhibited the expression of Mlxipl and p-cJun. Overexpression of cJun promoted the expression of Mlxipl and p-cJun. Quantification of the western blot (M) and immunofluorescence (O). Data relative to vehicle. N = 3. ^$$$P < 0.001, ^$$P < 0.01, ^$P < 0.05 vs. LPS; ^###P < 0.001, ^##P < 0.01 vs. LPS-siNC; ***P < 0.001, **P < 0.01, *P < 0.05 vs. LPS-LvNC. (P–R) ELISA were performed to detect the expression of proinflammation cytokines. CJun promoted inflammation response in LPS-induced microglia. N = 3. ^$$$P < 0.001, ^$$P < 0.01, ^$P < 0.05 vs. LPS; ^###P < 0.001, ^##P < 0.01 vs. LPS-siNC; ***P < 0.001, **P < 0.01, *P < 0.05 vs. LPS-LvNC.

  
  
  

  

  Upregulation of Mlxipl induced by cJun in the spinal dorsal horn after peripheral nerve injury counteracts mechanical allodynia by inhibiting neuroinflammation. CJun expression in the SDH was upregulated after peripheral nerve injury, and this increased Mlxipl transcription. Upregulation of Mlxipl counteracts mechanical allodynia by inhibiting neuroinflammation derived from microglia in the SDH.