Limits...
Mitochondrial Translocation of High Mobility Group Box 1 Facilitates LIM Kinase 2-Mediated Programmed Necrotic Neuronal Death.

Hyun HW, Ko AR, Kang TC - Front Cell Neurosci (2016)

Bottom Line: LIMK2 knockdown effectively attenuated SE-induced neuronal death and HMGB1 import into mitochondria accompanied by inhibiting nuclear HMGB1 release and abnormal mitochondrial elongation.However, LMB did not prevent mitochondrial elongation induced by SE, but inhibited the HMGB1 import into mitochondria.The efficacy of LMB was less effective to attenuate SE-induced neuronal death than that of LIMK2 siRNA.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy and Neurobiology, Institute of Epilepsy Research, College of Medicine, Hallym University Chuncheon, Kangwon-Do, South Korea.

ABSTRACT
High mobility group box 1 (HMGB1) acts a signaling molecule regulating a wide range of inflammatory responses in extracellular space. HMGB1 also stabilizes nucleosomal structure and facilitates gene transcription. Under pathophysiological conditions, nuclear HMGB1 is immediately transported to the cytoplasm through chromosome region maintenance 1 (CRM1). Recently, we have reported that up-regulation of LIM kinase 2 (LIMK2) expression induces HMGB1 export from neuronal nuclei during status epilepticus (SE)-induced programmed neuronal necrosis in the rat hippocampus. Thus, we investigated whether HMGB1 involves LIMK2-mediated programmed neuronal necrosis, but such role is not reported. In the present study, SE was induced by pilocarpine in rats that were intracerebroventricularly infused with saline, control siRNA, LIMK2 siRNA or leptomycin B (LMB, a CRM1 inhibitor) prior to SE induction. Thereafter, we performed Fluoro-Jade B staining, western blots and immunohistochemical studies. LIMK2 knockdown effectively attenuated SE-induced neuronal death and HMGB1 import into mitochondria accompanied by inhibiting nuclear HMGB1 release and abnormal mitochondrial elongation. LMB alleviated SE-induced neuronal death and nuclear HMGB1 release. However, LMB did not prevent mitochondrial elongation induced by SE, but inhibited the HMGB1 import into mitochondria. The efficacy of LMB was less effective to attenuate SE-induced neuronal death than that of LIMK2 siRNA. These findings indicate that nuclear HMGB1 release and the subsequent mitochondrial import may facilitate and deteriorate programmed necrotic neuronal deaths. The present data suggest that the nuclear HMGB1 release via CRM1 may be a potential therapeutic target for the programmed necrotic neuronal death induced by SE.

No MeSH data available.


Related in: MedlinePlus

Effect of LIM Kinase 2 (LIMK2) knockdown on status epilepticus (SE)-induced alterations in LIMK2 protein expression at 3 days after SE. (A) Western blot image of LIMK2 protein expression in the hippocampus. As compared to control siRNA, LIMK2 siRNA infusion effectively inhibits up-regulation of LIMK2 expression following SE. (B) Quantitative values (mean ± SEM) of LIMK2 expression level in the hippocampus, based on western blot (n = 7, respectively). *p < 0.05 vs. non-SE; #p < 0.05 vs. control siRNA. (C) Representative photographs of LIMK2 and NeuN in CA1 neurons. Following SE, LIMK2 expression is elevated in CA1 neurons. LIMK2 siRNA attenuates SE-induced LIMK2 induction in CA1 neurons. Scale bar = 25 μm. (D) Quantitative values (mean ± SEM) of LIMK2 and NeuN expression intensity in CA1 neurons, based on immunofluorescent images (n = 7, respectively). *p < 0.05 vs. non-SE; #p < 0.05 vs. control siRNA.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4829584&req=5

Figure 1: Effect of LIM Kinase 2 (LIMK2) knockdown on status epilepticus (SE)-induced alterations in LIMK2 protein expression at 3 days after SE. (A) Western blot image of LIMK2 protein expression in the hippocampus. As compared to control siRNA, LIMK2 siRNA infusion effectively inhibits up-regulation of LIMK2 expression following SE. (B) Quantitative values (mean ± SEM) of LIMK2 expression level in the hippocampus, based on western blot (n = 7, respectively). *p < 0.05 vs. non-SE; #p < 0.05 vs. control siRNA. (C) Representative photographs of LIMK2 and NeuN in CA1 neurons. Following SE, LIMK2 expression is elevated in CA1 neurons. LIMK2 siRNA attenuates SE-induced LIMK2 induction in CA1 neurons. Scale bar = 25 μm. (D) Quantitative values (mean ± SEM) of LIMK2 and NeuN expression intensity in CA1 neurons, based on immunofluorescent images (n = 7, respectively). *p < 0.05 vs. non-SE; #p < 0.05 vs. control siRNA.

Mentions: Consistent with our previous report (Kim et al., 2014), LIMK2 expression in the hippocampus was increased to 1.99-fold of non-SE level 3 days after SE (p < 0.05 vs. non-SE animals, Figures 1A,B). LIMK2 siRNA effectively inhibited SE-mediated LIMK2 induction in the hippocampus (p < 0.05 vs. control siRNA, Figures 1A,B). Immunohistochemical study revealed that LIMK2 expression was increased to 2.37-fold of non-SE level in the CA1 pyramidal cells 3 days after SE (p < 0.05 vs. non-SE animals, Figures 1C,D), while intensity of NeuN expression was reduced 0.35-fold of non-SE level in this region (p < 0.05 vs. non-SE animals, Figures 1C,D). LIMK2 siRNA infusion attenuated up-regulation of LIMK2 induction and decrease in NeuN expression induced by SE (p < 0.05 vs. control siRNA, Figures 1C,D). LMB infusion did not affect LIMK2 expression in CA1 pyramidal cells 3 days after SE (data not shown). These findings indicate that up-regulation of LIMK2 expression may play an important role in the SE-induced CA1 neuronal death, and nuclear HMGB1 release may not be involved in up-regulation of LIMK2 expression induced by SE.


Mitochondrial Translocation of High Mobility Group Box 1 Facilitates LIM Kinase 2-Mediated Programmed Necrotic Neuronal Death.

Hyun HW, Ko AR, Kang TC - Front Cell Neurosci (2016)

Effect of LIM Kinase 2 (LIMK2) knockdown on status epilepticus (SE)-induced alterations in LIMK2 protein expression at 3 days after SE. (A) Western blot image of LIMK2 protein expression in the hippocampus. As compared to control siRNA, LIMK2 siRNA infusion effectively inhibits up-regulation of LIMK2 expression following SE. (B) Quantitative values (mean ± SEM) of LIMK2 expression level in the hippocampus, based on western blot (n = 7, respectively). *p < 0.05 vs. non-SE; #p < 0.05 vs. control siRNA. (C) Representative photographs of LIMK2 and NeuN in CA1 neurons. Following SE, LIMK2 expression is elevated in CA1 neurons. LIMK2 siRNA attenuates SE-induced LIMK2 induction in CA1 neurons. Scale bar = 25 μm. (D) Quantitative values (mean ± SEM) of LIMK2 and NeuN expression intensity in CA1 neurons, based on immunofluorescent images (n = 7, respectively). *p < 0.05 vs. non-SE; #p < 0.05 vs. control siRNA.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4829584&req=5

Figure 1: Effect of LIM Kinase 2 (LIMK2) knockdown on status epilepticus (SE)-induced alterations in LIMK2 protein expression at 3 days after SE. (A) Western blot image of LIMK2 protein expression in the hippocampus. As compared to control siRNA, LIMK2 siRNA infusion effectively inhibits up-regulation of LIMK2 expression following SE. (B) Quantitative values (mean ± SEM) of LIMK2 expression level in the hippocampus, based on western blot (n = 7, respectively). *p < 0.05 vs. non-SE; #p < 0.05 vs. control siRNA. (C) Representative photographs of LIMK2 and NeuN in CA1 neurons. Following SE, LIMK2 expression is elevated in CA1 neurons. LIMK2 siRNA attenuates SE-induced LIMK2 induction in CA1 neurons. Scale bar = 25 μm. (D) Quantitative values (mean ± SEM) of LIMK2 and NeuN expression intensity in CA1 neurons, based on immunofluorescent images (n = 7, respectively). *p < 0.05 vs. non-SE; #p < 0.05 vs. control siRNA.
Mentions: Consistent with our previous report (Kim et al., 2014), LIMK2 expression in the hippocampus was increased to 1.99-fold of non-SE level 3 days after SE (p < 0.05 vs. non-SE animals, Figures 1A,B). LIMK2 siRNA effectively inhibited SE-mediated LIMK2 induction in the hippocampus (p < 0.05 vs. control siRNA, Figures 1A,B). Immunohistochemical study revealed that LIMK2 expression was increased to 2.37-fold of non-SE level in the CA1 pyramidal cells 3 days after SE (p < 0.05 vs. non-SE animals, Figures 1C,D), while intensity of NeuN expression was reduced 0.35-fold of non-SE level in this region (p < 0.05 vs. non-SE animals, Figures 1C,D). LIMK2 siRNA infusion attenuated up-regulation of LIMK2 induction and decrease in NeuN expression induced by SE (p < 0.05 vs. control siRNA, Figures 1C,D). LMB infusion did not affect LIMK2 expression in CA1 pyramidal cells 3 days after SE (data not shown). These findings indicate that up-regulation of LIMK2 expression may play an important role in the SE-induced CA1 neuronal death, and nuclear HMGB1 release may not be involved in up-regulation of LIMK2 expression induced by SE.

Bottom Line: LIMK2 knockdown effectively attenuated SE-induced neuronal death and HMGB1 import into mitochondria accompanied by inhibiting nuclear HMGB1 release and abnormal mitochondrial elongation.However, LMB did not prevent mitochondrial elongation induced by SE, but inhibited the HMGB1 import into mitochondria.The efficacy of LMB was less effective to attenuate SE-induced neuronal death than that of LIMK2 siRNA.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy and Neurobiology, Institute of Epilepsy Research, College of Medicine, Hallym University Chuncheon, Kangwon-Do, South Korea.

ABSTRACT
High mobility group box 1 (HMGB1) acts a signaling molecule regulating a wide range of inflammatory responses in extracellular space. HMGB1 also stabilizes nucleosomal structure and facilitates gene transcription. Under pathophysiological conditions, nuclear HMGB1 is immediately transported to the cytoplasm through chromosome region maintenance 1 (CRM1). Recently, we have reported that up-regulation of LIM kinase 2 (LIMK2) expression induces HMGB1 export from neuronal nuclei during status epilepticus (SE)-induced programmed neuronal necrosis in the rat hippocampus. Thus, we investigated whether HMGB1 involves LIMK2-mediated programmed neuronal necrosis, but such role is not reported. In the present study, SE was induced by pilocarpine in rats that were intracerebroventricularly infused with saline, control siRNA, LIMK2 siRNA or leptomycin B (LMB, a CRM1 inhibitor) prior to SE induction. Thereafter, we performed Fluoro-Jade B staining, western blots and immunohistochemical studies. LIMK2 knockdown effectively attenuated SE-induced neuronal death and HMGB1 import into mitochondria accompanied by inhibiting nuclear HMGB1 release and abnormal mitochondrial elongation. LMB alleviated SE-induced neuronal death and nuclear HMGB1 release. However, LMB did not prevent mitochondrial elongation induced by SE, but inhibited the HMGB1 import into mitochondria. The efficacy of LMB was less effective to attenuate SE-induced neuronal death than that of LIMK2 siRNA. These findings indicate that nuclear HMGB1 release and the subsequent mitochondrial import may facilitate and deteriorate programmed necrotic neuronal deaths. The present data suggest that the nuclear HMGB1 release via CRM1 may be a potential therapeutic target for the programmed necrotic neuronal death induced by SE.

No MeSH data available.


Related in: MedlinePlus