Impact of siRNA targeting of β-catenin on differentiation of rat neural stem cells and gene expression of Ngn1 and BMP4 following in vitro hypoxic-ischemic brain damage

Zhang,Xiaoying; Zhu,Cuicui; Luo,Qiong; Dong,Jv; Liu,Lv; Li,Min; Zhu,Hongtao; Ma,Xiangping; Wang,Jun

doi:10.3892/mmr.2016.5667

Impact of siRNA targeting of β-catenin on differentiation of rat neural stem cells and gene expression of Ngn1 and BMP4 following in vitro hypoxic-ischemic brain damage

Authors:
- Xiaoying Zhang
- Cuicui Zhu
- Qiong Luo
- Jv Dong
- Lv Liu
- Min Li
- Hongtao Zhu
- Xiangping Ma
- Jun Wang
View Affiliations

Affiliations: Department of Pediatrics, The First Affiliated Hospital of Xinjiang Medical University, Ürümqi, Xinjiang 830054, P.R. China
Published online on: August 24, 2016 https://doi.org/10.3892/mmr.2016.5667
Pages: 3595-3601
Copyright: © Zhang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

The aim of the present study was to investigate the possible damage-repair mechanisms of neural stem cells (NSCs) following hypoxic-ischemic brain damage (HIBD). NSCs obtained from Sprague Dawley rats were treated with tissue homogenate from normal or HIBD tissue, and β‑catenin expression was silenced using siRNA. The differentiation of NSCs was observed by immunofluorescence, and semiquantitative reverse transcription‑polymerase chain reaction and western blot analysis were applied to detect the mRNA and protein expression levels of Ngn1 and BMP4 in the NSCs. Compared with control NSCs, culture with brain tissue homogenate significantly increased the differentiation of NSCs into neurons and oligodendrocytes (P<0.05), whereas differentiation into astrocytes was significantly reduced (P<0.05). Compared with negative control‑transfected cells, knockdown of β‑catenin expression significantly decreased the differentiation of NSCs into neurons and oligodendrocytes (P<0.01), whereas the percentage of NSCs differentiated into astrocytes was significantly increased (P<0.01). Compared with control NSCs, the mRNA and protein expression levels of Ngn1 were significantly increased (P<0.01) and BMP4 levels were significantly reduced (P<0.01) by exposure of the cells to brain tissue homogenate. Compared with the negative control plasmid‑transfected NSCs, the levels of Ngn1 mRNA and protein were significantly reduced by β‑catenin siRNA (P<0.01), whereas BMP4 levels were significantly increased (P<0.01). In summary, the damaged brain tissues in HIBD may promote NSCs to differentiate into neurons for self‑repair processes. β‑Catenin, BMP4 and Ngn1 may be important for the coordination of NSC proliferation and differentiation following HIBD.

View Figures

View References

1	Nanavati T, Seemaladinne N, Regier M, Yossuck P and Pergami P: Can we predict functional outcome in neonates with hypoxic ischemic encephalopathy by the combination of neuroimaging and electroencephalography? Pediatr Neonatol. 56:307–316. 2015. View Article : Google Scholar : PubMed/NCBI
2	Buonocore G, Perrone S, Longini M, Paffetti P, Vezzosi P, Gatti MG and Bracci R: Non protein bound iron as early predictive marker of neonatal brain damage. Brain. 126:1224–1230. 2013. View Article : Google Scholar
3	Cai Q, Xue XD and Fu JH: Research status and progress of neonatal hypoxic ischemic encephalopathy. Zhong Guo Shi Yong Er Ke Za Zhi. 24:968–971. 2009.In Chinese.
4	Vanucci RC and Perlman JM: Interventions for perinatal hypoxic-ischemic encephalopathy. Pediatrics. 100:1004–1014. 1997. View Article : Google Scholar
5	Sun D: Endogenous neurogenic cell response in the mature mammalian brain following traumatic injury. Exp Neurol. 275:405–410. 2016. View Article : Google Scholar
6	Sun D: The potential of endogenous neurogenesis for brain repair and regeneration following traumatic brain injury. Neural Regen Res. 9:688–692. 2014. View Article : Google Scholar : PubMed/NCBI
7	Edelmann K, Glashauser L, Sprungala S, Hesl B, Fritschle M, Ninkovic J, Godinho L and Chapouton P: Increased radial glia quiescence, decreased reactivation upon injury and unaltered neuroblast behavior underlie decreased neurogenesis in the aging zebrafish telencephalon. J Comp Neurol. 521:3099–3115. 2013. View Article : Google Scholar : PubMed/NCBI
8	Oliva CA and Inestrosa NC: A novel function for Wnt signaling modulating neuronal firing activity and the temporal structure of spontaneous oscillation in the entorhinal-hippocampal circuit. Exp Neurol. 269:43–55. 2015. View Article : Google Scholar : PubMed/NCBI
9	Oliva CA, Vargas JY and Inestrosa NC: Wnts in adult brain: From synaptic plasticity to cognitive deficiencies. Front Cell Neurosci. 7:2242013. View Article : Google Scholar : PubMed/NCBI
10	Inestrosa NC and Varela-Nallar L: Wnt signaling in the nervous system and in Alzheimer's disease. J Mol Cell Biol. 6:64–74. 2014. View Article : Google Scholar : PubMed/NCBI
11	Liu W, Zhou H, Liu L, Zhao C, Deng Y, Chen L, Wu L, Mandrycky N, McNabb CT, Peng Y, et al: Disruption of neurogenesis and cortical development in transgenic mice misex-pressing Olig2, a gene in the Down syndrome critical region. Neurobiol Dis. 77:106–116. 2015. View Article : Google Scholar : PubMed/NCBI
12	Liu F, Xuan A, Chen Y, Zhang J, Xu L, Yan Q and Long D: Combined effect of nerve growth factor and brain derived neurotrophic factor on neuronal differentiation of neural stem cells and the potential molecular mechanisms. Mol Med Rep. 10:173917–173945. 2014.
13	Lei ZN, Liu F, Zhang LM, Huang YL and Sun FY: Bcl-2 increases stroke-induced striatal neurogenesis in adult brains by inhibiting BMP-4 function via activation of β-catenin signaling. Neurochem Int. 61:34–42. 2012. View Article : Google Scholar : PubMed/NCBI
14	Chuang CY, Lin KI, Hsiao M, Stone L, Chen HF, Huang YH, Lin SP, Ho HN and Kuo HC: Meiotic competent human germ cell-like cells derived from human embryonic stem cells induced by BMP4/WNT3A signaling and OCT4/EpCAM (epithelial cell adhesion molecule) selection. J Biol Chem. 287:14389–14401. 2012. View Article : Google Scholar : PubMed/NCBI
15	Zhang XY, Yang YJ, Xu PR, Zheng XR, Wang QH, Chen CF and Yao Y: The role of β-catenin signaling pathway on proliferation of rats neural stem cells after hyperbaric oxygen therapy in vitro. Cell Mol Neurobiol. 31:101–109. 2011. View Article : Google Scholar
16	Rice JE III, Vannucci RC and Brierley JB: The influence of immaturity on hypoxic-ischemic brain damage in the rat. Ann Neurol. 9:131–141. 1981. View Article : Google Scholar : PubMed/NCBI
17	Chen CF, Yang YJ, Wang QH, Yao Y and Li M: Effect of hyperbaric oxygen administered at different pressures and diffrernt exposure time on differentiation of neural stem cells in vitro. Zhongguo Dang Dai Er Ke Za Zhi. 12:368–372. 2010.In Chinese. PubMed/NCBI
18	Shyam R, Ren Y, Lee J, Braunstein KE, Mao HQ and Wong PC: Intraventricular delivery of siRNA nanoparticles to the central nervous system. Mol Ther Nucleic Acids. 4:e2422015. View Article : Google Scholar : PubMed/NCBI
19	Li TS, Yawata T and Honke K: Efficient siRNA delivery and tumor accumulation mediated by ionically cross-linked folic acid-poly(ethylene glycol)-chitosan oligosaccharide lactate nanoparticles: For the potential targeted ovarian cancer gene therapy. Eur J Pharm Sci. 52:48–61. 2014. View Article : Google Scholar
20	Zhang XY, Yang YJ, Chen CF, Yao Y and Wang QH: Construction and screening of eukaryotic expression plasmids containing shRNA targeting β-catenin gene. J Med Mol Biol. 7:136–142. 2010.
21	Gigek CO, Chen ES, Ota VK, Maussion G, Peng H, Vaillancourt K, Diallo AB, Lopez JP, Crapper L, Vasuta C, et al: A molecular model for neurodevelopmental disorders. Transl Psychiatry. 5:e5652015. View Article : Google Scholar : PubMed/NCBI
22	Chen ES, Gigek CO, Rosenfeld JA, Diallo AB, Maussion G, Chen GG, Vaillancourt K, Lopez JP, Crapper L, Poujol R, et al: Molecular convergence of neurodevelopmental disorders. Am J Hum Genet. 95:490–508. 2014. View Article : Google Scholar : PubMed/NCBI
23	Rak K, Völker J, Jürgens L, Völker C, Frenz S, Scherzad A, Schendzielorz P, Jablonka S, Mlynski R, Radeloff A and Hagen R: Cochlear nucleus whole mount explants promote the differentiation of neuronal stem cells from the cochlear nucleus in co-culture experiments. Brain Res. 1616:58–70. 2015. View Article : Google Scholar : PubMed/NCBI
24	Schafer ST, Han J, Pena M, von Bohlen Und Halbach O, Peters J and Gage FH: The Wnt adaptor protein ATP6AP2 regulates multiple stages of adult hippocampal neurogenesis. J Neurosci. 35:4983–4998. 2015. View Article : Google Scholar : PubMed/NCBI
25	Wang L, Liu Y, Li S, Long ZY and Wu YM: Wnt signaling pathway participates in valproic acid-induced neuronal differentiation of neural stem cells. Int J Clin Exp Pathol. 8:578–585. 2015.PubMed/NCBI
26	Ma YX, Wu ZQ, Feng YJ, Xiao ZC, Qin XL and Ma QH: G protein coupled receptor 50 promotes self-renewal and neuronal differentiation of embryonic neural progenitor cells through regulation of notch and wnt/β-catenin signalings. Biochem Biophys Res Commun. 458:836–842. 2015. View Article : Google Scholar : PubMed/NCBI
27	Yuan L and Hassan BA: Neurogenins in brain development and disease: An overview. Arch Biochem Biophys. 558:10–13. 2014. View Article : Google Scholar : PubMed/NCBI
28	Imayoshi I and Kageyama R: bHLH factors in self-renewal, multipotency, and fate choice of neural progenitor cells. Neuron. 82:9–23. 2014. View Article : Google Scholar : PubMed/NCBI
29	Zhang Z, Shi Y, Zhao S, Li J, Li C and Mao B: Xenopus Nkx6.3 is a neural plate border specifier required for neural crest development. PLoS One. 9:e1151652014. View Article : Google Scholar : PubMed/NCBI
30	An SM, Ding Q, Zhang J, Xie J and Li L: Targeting stem cell signaling pathways for drug discovery: Advances in the Notch and Wnt pathways. Sci China Life Sci. 57:575–580. 2014. View Article : Google Scholar : PubMed/NCBI