적절한 농도의 활성산소종(ROS)은 병원체에 대한 세포의 방어, 신호 전달, 세포 성장 및 유전자 발현을 포함한 다양한 정상 세포 기능을 매개한다. 최근의 연구는 ROS와 ROS를 생성하는 NADPH 산화 효소(Nox)가 성인 쥐 뇌의 뇌실 하 영역(SVZ)에 있는 신경 줄기세포의 자가 복제와 신경 세포 분화에 중요하다는 것을 보여 주었다. 본 연구에서 세포 내 ROS가 갓 태어난 쥐의 뇌에서 적출되어 배양된 SVZ 신경 줄기세포에서 검출된 것으로 나타났다. Nox 유사 유전자들 중 Nox4가 배양된 세포에서 주로 발현되었고, Nox1과 Nox2는 거의 발현되지 않았다. 또한, Nox4 유전자는 신경 세포 분화 동안 최대 10배까지 발현이 크게 증가하였다. Immunocytochemistry결과 Nox4 단백질은 신경 세포 특이적인 tubulin인 Tuj1-양성 신경 세포에서 주로 발견되었다. 이와 맥을 같이 하여, 내인성 ROS는 분화 후 축삭돌기를 가지고 있으며 신경 세포로 보이는 세포에서만 검출되었다. 또한, ROS를 제거하는N-acetyl cysteine에 의해 세포 산화 환원 상태가 교란되었을 때, 신경 세포로의 분화가 크게 감소하였다. 마지막으로, shRNA를 이용하 여 Nox4를 knockdown한 세포에서 신경 세포로의 분화가 감소하였다. 이러한 연구 결과는 Nox4가 갓 태어난 쥐의 SVZ 신경 줄기 세포의 주요한 ROS 생성 효소이고, Nox4에 의한 ROS생성이 신경 세포 분화에 중요하다는 것을 암시한다.
적절한 농도의 활성산소종(ROS)은 병원체에 대한 세포의 방어, 신호 전달, 세포 성장 및 유전자 발현을 포함한 다양한 정상 세포 기능을 매개한다. 최근의 연구는 ROS와 ROS를 생성하는 NADPH 산화 효소(Nox)가 성인 쥐 뇌의 뇌실 하 영역(SVZ)에 있는 신경 줄기세포의 자가 복제와 신경 세포 분화에 중요하다는 것을 보여 주었다. 본 연구에서 세포 내 ROS가 갓 태어난 쥐의 뇌에서 적출되어 배양된 SVZ 신경 줄기세포에서 검출된 것으로 나타났다. Nox 유사 유전자들 중 Nox4가 배양된 세포에서 주로 발현되었고, Nox1과 Nox2는 거의 발현되지 않았다. 또한, Nox4 유전자는 신경 세포 분화 동안 최대 10배까지 발현이 크게 증가하였다. Immunocytochemistry결과 Nox4 단백질은 신경 세포 특이적인 tubulin인 Tuj1-양성 신경 세포에서 주로 발견되었다. 이와 맥을 같이 하여, 내인성 ROS는 분화 후 축삭돌기를 가지고 있으며 신경 세포로 보이는 세포에서만 검출되었다. 또한, ROS를 제거하는N-acetyl cysteine에 의해 세포 산화 환원 상태가 교란되었을 때, 신경 세포로의 분화가 크게 감소하였다. 마지막으로, shRNA를 이용하 여 Nox4를 knockdown한 세포에서 신경 세포로의 분화가 감소하였다. 이러한 연구 결과는 Nox4가 갓 태어난 쥐의 SVZ 신경 줄기 세포의 주요한 ROS 생성 효소이고, Nox4에 의한 ROS생성이 신경 세포 분화에 중요하다는 것을 암시한다.
Reactive oxygen species (ROS), at appropriate concentrations, mediate various normal cellular functions, including defense against pathogens, signal transduction, cellular growth, and gene expression. A recent study demonstrated that ROS and ROS-generating NADPH oxidase (Nox) are important in self-r...
Reactive oxygen species (ROS), at appropriate concentrations, mediate various normal cellular functions, including defense against pathogens, signal transduction, cellular growth, and gene expression. A recent study demonstrated that ROS and ROS-generating NADPH oxidase (Nox) are important in self-renewal and neuronal differentiation of subventricular zone (SVZ) neural stem cells in adult mouse brains. In this study, we found that endogenous ROS were detected in SVZ neural stem cells cultured from postnatal mouse brains. Nox4 was predominantly expressed in cultured cells, while the levels of the Nox1 and Nox2 transcripts were very low. In addition, the Nox4 gene was highly upregulated (by up to 10-fold) during neuronal differentiation. Immunocytochemical analysis detected the Nox4 protein mainly in neurons positive for the neuronal specific tubulin Tuj1. After differentiation, endogenous ROS were detected exclusively in neuron-like cells with processes. In addition, perturbation of the cellular redox state with N-acetyl cysteine, a ROS scavenger, during neuronal differentiation greatly inhibited neurogenesis. Lastly, knockdown of Nox4 using short hairpin RNA decreased neurogenesis. These findings suggest that Nox4 may be a major ROS-generating enzyme in postnatal SVZ neural stem cells, and Nox4-mediated ROS generation may be important in their neuronal differentiation.
Reactive oxygen species (ROS), at appropriate concentrations, mediate various normal cellular functions, including defense against pathogens, signal transduction, cellular growth, and gene expression. A recent study demonstrated that ROS and ROS-generating NADPH oxidase (Nox) are important in self-renewal and neuronal differentiation of subventricular zone (SVZ) neural stem cells in adult mouse brains. In this study, we found that endogenous ROS were detected in SVZ neural stem cells cultured from postnatal mouse brains. Nox4 was predominantly expressed in cultured cells, while the levels of the Nox1 and Nox2 transcripts were very low. In addition, the Nox4 gene was highly upregulated (by up to 10-fold) during neuronal differentiation. Immunocytochemical analysis detected the Nox4 protein mainly in neurons positive for the neuronal specific tubulin Tuj1. After differentiation, endogenous ROS were detected exclusively in neuron-like cells with processes. In addition, perturbation of the cellular redox state with N-acetyl cysteine, a ROS scavenger, during neuronal differentiation greatly inhibited neurogenesis. Lastly, knockdown of Nox4 using short hairpin RNA decreased neurogenesis. These findings suggest that Nox4 may be a major ROS-generating enzyme in postnatal SVZ neural stem cells, and Nox4-mediated ROS generation may be important in their neuronal differentiation.
* AI 자동 식별 결과로 적합하지 않은 문장이 있을 수 있으니, 이용에 유의하시기 바랍니다.
문제 정의
In this work, we showed that the level of Nox4 transcript was strongly increased during neurogenesis of SVZ NSCs. In addition, we detected ROS in differentiated neurons and a ROS scavenger inhibited neurogenesis.
가설 설정
Scale bar=20 μm. B. Transduced NSCs as in A were imaged and GFP-positive cells were counted from 4 wells in each condition. Data are mean and S.
Yet, there have been few reports of ROS-generating enzymes in stem cell biology [16, 32]. In this study, we investigated the role of ROS and a ROS-generating enzyme in SVZ NSCs dissected from post- natal mouse brain. Consistent with previous studies, ROS appeared to be a key part of neuronal differentiation of SVZ NSCs.
제안 방법
These results sup- port the notion that an increase in Nox4 expression during differentiation of SVZ NSCs occurs mainly in neurons. Further, we examined ROS generation in differentiated NSCs by staining them with DCFDA. Neuronal-like cells with processes were positive for DCFDA fluorescence (Fig.
To confirm the qRT-PCR data, immunocytochemistry was performed to check the levels of the Nox4 protein. At 5 days of differentiation, Nox4 was detected in neuronal-specific tu- bulin Tuj1-positive cells (Fig.
대상 데이터
Both viral particles were purchased from Dharma- con (Lafayette, CO, USA). For Nox4 shRNA, clone ID was VGM5520 and for negative control shRNA, RHS4348 was purchased. After transduction, cells were passaged several times and then transferred to laminin-coated 8-well Lab-Tek CC2 chamber slide (Nunc, ThermoFisher, Waltham, MA, USA).
성능/효과
Recently, ROS have been added to the list. The results of this study suggest that Nox4 might be a major ROS-generat- ing enzyme in neurogenesis in postnatal SVZ NSC cultures. Still, more work needs to be done to fully identify the molec- ular mechanisms of Nox4 gene regulation and the players downstream of ROS during neuronal differentiation.
참고문헌 (34)
Alfadda, A. A. and Sallam, R. M. 2012. Reactive oxygen species in health and disease. J. Biomed. Biotechnol. 2012, 936486.
Bigarella, C. L., Liang, R. and Ghaffari, S. 2014. Stem cells and the impact of ros signaling. Development 141, 4206-4218.
Chuikov, S., Levi, B. P., Smith, M. L. and Morrison, S. J. 2010. Prdm16 promotes stem cell maintenance in multiple tissues, partly by regulating oxidative stress. Nat. Cell Biol. 12, 999-1006.
Dickinson, B. C., Peltier, J., Stone, D., Schaffer, D. V. and Chang, C. J. 2011. Nox2 redox signaling maintains essential cell populations in the brain. Nat. Chem. Biol. 7, 106-112.
Doetsch, F., Garcia-Verdugo, J. M. and Alvarez-Buylla, A. 1997. Cellular composition and three-dimensional organization of the subventricular germinal zone in the adult mammalian brain. J. Neurosci. 17, 5046-5061.
Domenis, R., Bergamin, N., Gianfranceschi, G., Vascotto, C., Romanello, M., Rigo, S., Vagnarelli, G., Faggiani, M., Parodi, P., Kelley, M. R., Beltrami, C. A., Cesselli, D., Tell, G. and Beltrami, A. P. 2014. The redox function of ape1 is involved in the differentiation process of stem cells toward a neuronal cell fate. PLoS One 9, e89232.
Forsberg, K. and Di Giovanni, S. 2014. Cross talk between cellular redox status, metabolism, and p53 in neural stem cell biology. Neuroscientist 20, 326-342.
Haack, F., Lemcke, H., Ewald, R., Rharass, T. and Uhrmacher, A. M. 2015. Spatio-temporal model of endogenous ros and raft-dependent wnt/beta-catenin signaling driving cell fate commitment in human neural progenitor cells. PLoS Comput. Biol. 11, e1004106.
Hou, Y., Ouyang, X., Wan, R., Cheng, H., Mattson, M. P. and Cheng, A. 2012. Mitochondrial superoxide production negatively regulates neural progenitor proliferation and cerebral cortical development. Stem Cells 30, 2535-2547.
Hwang, W. W., Salinas, R. D., Siu, J. J., Kelley, K. W., Delgado, R. N., Paredes, M. F., Alvarez-Buylla, A., Oldham, M. C. and Lim, D. A. 2014. Distinct and separable roles for ezh2 in neurogenic astroglia. Elife 3, e02439.
Infanger, D. W., Sharma, R. V. and Davisson, R. L. 2006. Nadph oxidases of the brain: Distribution, regulation, and function. Antioxid. Redox Signal. 8, 1583-1596.
Kanda, Y., Hinata, T., Kang, S. W. and Watanabe, Y. 2011. Reactive oxygen species mediate adipocyte differentiation in mesenchymal stem cells. Life Sci. 89, 250-258.
Kim, J. and Wong, P. K. 2009. Loss of atm impairs proliferation of neural stem cells through oxidative stress-mediated p38 mapk signaling. Stem Cells 27, 1987-1998.
Kim, S. R., Kim, D. I., Kim, S. H., Lee, H., Lee, K. S., Cho, S. H. and Lee, Y. C. Nlrp3 inflammasome activation by mitochondrial ros in bronchial epithelial cells is required for allergic inflammation. Cell Death Dis. 5, e1498.
Le Belle, J. E., Orozco, N. M., Paucar, A. A., Saxe, J. P., Mottahedeh, J., Pyle, A. D., Wu, H. and Kornblum, H. I. 2011. Proliferative neural stem cells have high endogenous ros levels that regulate self-renewal and neurogenesis in a pi3k/akt-dependant manner. Cell Stem Cell 8, 59-71.
Lee, J. E., Cho, K. E., Lee, K. E., Kim, J. and Bae, Y. S. 2014. Nox4-mediated cell signaling regulates differentiation and survival of neural crest stem cells. Mol. Cells 37, 907-911.
Lim, D. A., Huang, Y. C., Swigut, T., Mirick, A. L., Garcia-Verdugo, J. M., Wysocka, J., Ernst, P. and Alvarez-Buylla, A. 2009. Chromatin remodelling factor mll1 is essential for neurogenesis from postnatal neural stem cells. Nature 458, 529-533.
Liu, J., Cao, L., Chen, J., Song, S., Lee, I. H., Quijano, C., Liu, H., Keyvanfar, K., Chen, H., Cao, L. Y., Ahn, B. H., Kumar, N. G., Rovira, II., Xu, X. L., van Lohuizen, M., Motoyama, N., Deng, C. X. and Finkel, T. 2009. Bmi1 regulates mitochondrial function and the DNA damage response pathway. Nature 459, 387-392.
Menn, B., Garcia-Verdugo, J. M., Yaschine, C., Gonzalez-Perez, O., Rowitch, D. and Alvarez-Buylla, A. 2006. Origin of oligodendrocytes in the subventricular zone of the adult brain. J. Neurosci. 26, 7907-7918.
Park, D. H., Hong, S. J., Salinas, R. D., Liu, S. J., Sun, S. W., Sgualdino, J., Testa, G., Matzuk, M. M., Iwamori, N. and Lim, D. A. 2014. Activation of neuronal gene expression by the jmjd3 demethylase is required for postnatal and adult brain neurogenesis. Cell Rep. 8, 1290-1299.
Park, J. E., Seo, Y. K., Yoon, H. H., Kim, C. W., Park, J. K. and Jeon, S. 2013. Electromagnetic fields induce neural differentiation of human bone marrow derived mesenchymal stem cells via ros mediated egfr activation. Neurochem. Int. 62, 418-424.
Price, J. D., Park, K. Y., Chen, J., Salinas, R. D., Cho, M. J., Kriegstein, A. R. and Lim, D. A. 2014. The ink4a/arf locus is a barrier to direct neuronal transdifferentiation. J. Neurosci. 34, 12560-12567.
Sanders, Y. Y., Liu, H., Liu, G. and Thannickal, V. J. 2015. Epigenetic mechanisms regulate nadph oxidase-4 expression in cellular senescence. Free Radic. Biol. Med. 79, 197-205.
Serrander, L., Cartier, L., Bedard, K., Banfi, B., Lardy, B., Plastre, O., Sienkiewicz, A., Forro, L., Schlegel, W. and Krause, K. H. 2007. Nox4 activity is determined by mrna levels and reveals a unique pattern of ros generation. Biochem. J. 406, 105-114.
Shen, Q., Wang, Y., Kokovay, E., Lin, G., Chuang, S. M., Goderie, S. K., Roysam, B. and Temple, S. 2008. Adult svz stem cells lie in a vascular niche: A quantitative analysis of niche cell-cell interactions. Cell Stem Cell 3, 289-300.
Topchiy, E., Panzhinskiy, E., Griffin, W. S., Barger, S. W., Das, M. and Zawada, W. M. 2013. Nox4-generated superoxide drives angiotensin ii-induced neural stem cell proliferation. Dev. Neurosci. 35, 293-305.
Vieira, H. L., Alves, P. M. and Vercelli, A. 2011. Modulation of neuronal stem cell differentiation by hypoxia and reactive oxygen species. Prog. Neurobiol. 93, 444-455.
Walton, N. M., Shin, R., Tajinda, K., Heusner, C. L., Kogan, J. H., Miyake, S., Chen, Q., Tamura, K. and Matsumoto, M. 2012. Adult neurogenesis transiently generates oxidative stress. PLoS One 7, e35264.
Wei, W., Lu, Y., Hao, B., Zhang, K., Wang, Q., Miller, A. L., Zhang, L. R., Zhang, L. H. and Yue, J. 2015. Cd38 is required for neural differentiation of mouse embryonic stem cells by modulating reactive oxygen species (ros). Stem Cells. 33, DOI: 10.1002/stem.2057.
Xiao, Q., Luo, Z., Pepe, A. E., Margariti, A., Zeng, L. and Xu, Q. 2009. Embryonic stem cell differentiation into smooth muscle cells is mediated by nox4-produced h2o2. Am. J. Physiol. Cell Physiol. 296, C711-723.
Yoneyama, M., Kawada, K., Gotoh, Y., Shiba, T. and Ogita, K. 2010. Endogenous reactive oxygen species are essential for proliferation of neural stem/progenitor cells. Neurochem. Int. 56, 740-746.
Zelko, I. N. and Folz, R. J. 2015. Regulation of oxidative stress in pulmonary artery endothelium: Modulation of ec-sod and nox4 expression using hdac class i inhibitors. Am. J. Respir. Cell Mol. Biol. 53, 513-524.
※ AI-Helper는 부적절한 답변을 할 수 있습니다.