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Journal of Peking University (Health Sciences) ›› 2022, Vol. 54 ›› Issue (3): 421-426. doi: 10.19723/j.issn.1671-167X.2022.03.005

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Inhibition connexin 43 by mimetic peptide Gap27 mediates protective effects on 6-hydroxydopamine induced Parkinson's disease mouse model

Hui-hui QUAN,Wei-xing XU,Yu-ze QI,Qing-ru LI,Hui ZHOU,Jing HUANG*()   

  1. Department of Occupational and Environmental Health, Peking University School of Public Health, Beijing 100191, China
  • Received:2020-08-31 Online:2022-06-18 Published:2022-06-14
  • Contact: Jing HUANG E-mail:jing_huang@bjmu.edu.cn
  • Supported by:
    the National Natural Science Foundation of China(21577004);the Beijing Natural Science Foundation of(7162104)

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Abstract:

Objective: To explore whether the using of mimetic peptide Gap27, a selective inhibitor of connexin 43 (Cx43), could block the death of dopamine neurons and influence the expression of Cx43 in 6-hydroxydopamine (6-OHDA)-induced Parkinson's disease mouse models. Methods: Eighteen C57BL/6 mice were randomly divided into control group, 6-OHDA group and 6-OHDA+Gap27 group, with 6 mice in each group. Bilateral substantia nigra stereotactic injection was performed. The control group was injected with ascorbate solution, 6-OHDA group was injected with 6-OHDA solution, and 6-OHDA+Gap27 group was injected with 6-OHDA and Gap27 mixed solution. Immuno-histochemical staining was used to detect the number of dopamine neurons, quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of Cx43 messenger ribonucleic acid (mRNA), immuno-fluorescence staining was used to detect the distribution of Cx43 protein, the contents of Cx43 protein and Cx43 phosphorylation at serine 368 (Cx43-ps368) in mouse midbrain were detected by Western blot. Results: After injection of 6-OHDA, numerous dopamine neurons in substantia nigra died as Cx43 content increased, Cx43-ps368 content decreased. Mixing Gap27 while injecting 6-OHDA could reduce the number of death dopamine neurons and weaken the changes of Cx43 and Cx43-ps368 content caused by 6-OHDA. The number of tyrosine hydroxylase (TH) immunoreactive positive neurons in 6-OHDA group decreased to 27.7% ± 0.02% of the control group (P < 0.01); The number of TH immunoreactive positive neurons in 6-OHDA+Gap27 group was (1.64±0.16) times higher than that in 6-OHDA group (P < 0.05); The content of total Cx43 protein in 6-OHDA group was (1.44±0.07) times higher than that in 6-OHDA+Gap27 group (P < 0.05) while (1.68±0.07) times higher than that in control group (P < 0.01). In 6-OHDA group, the content of Cx43-ps368 protein and its proportion in total Cx43 protein were significantly lower than that in 6-OHDA+Gap27 group (P < 0.05). Conclusion: In 6-OHDA mouse models, mimetic peptide Gap27 played a protective role in reducing the damage to substantia nigra dopamine neurons, which was induced by 6-OHDA. The overexpression of Cx43 protein might have neurotoxicity to dopamine neuron. Meanwhile, decreasing Cx43 protein level and keeping Cx43-ps368 protein level may be the protective mechanisms of Gap27.

Key words: Connexin 43, 6-hydroxydopamine, Peptide Gap27, Parkinson's disease

CLC Number: 

  • R392.3

Figure 1

Variation of DA neuron number in substantia nigra A, TH immunoreactive neurons of control group (DAB staining, n=3); B, TH immunoreactive neurons of 6-OHDA group (DAB staining, n=3); C, TH immunoreactive neurons of 6-OHDA+Gap27 group (DAB staining, n=3); D, relative number of TH positive neurons in substantia nigra. * P < 0.05, # P < 0.01. TH, tyrosine hydroxylase; 6-OHDA, 6-hydroxydopamine."

Figure 2

Changes of Cx43 protein content, gene expression and distribution A, the differences of Cx43 and Cx43-ps368 protein content among groups (n=3); B, relative expression of Cx43 protein, Cx43-ps368 protein, and Cx43-ps368/Cx43; C, relative expression of Cx43 mRNA (n=3); D, immunofluorescence co-staining of astrocytes (green) and Cx43 protein (red) in substantia nigra (n=3); E, relative expression of Cx43 protein fluorescence intensity. * P < 0.05, # P < 0.01. 6-OHDA, 6-hydroxydopamine; DAPI, 4′, 6-diamidino-2-phenylindole."

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