Neural stem cells from Persian sturgeon (Acipenser persicus): Determining optimum temperature, long-term culture and immunocytochemistry

Document Type : scientific research article

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Abstract

Cell culture from fish brain have high potential in investigation of different responses to toxins, nutrients and growth factors. In the present study, to investigate the possibility of preparing long-term culture from Persian surgeon brain, characterization of neural stem cells and determining optimum temperature for the growth of cells, the front (olfactory lobe), middle (optic lobe) and rear (cerebellum) regions of Persian sturgeon brain were placed separately in the medium of DMEM/F12 with 15% FBS, penicillin-streptomycin and amphotrisin in incubator (250C, 5%Co2). The obtained cells were mostly fusiform. Cells from rear part of the brain are still alive after more than 8 months and 9 passages. To determination of optimum temperature for the growth of Persian sturgeon brain cells, cells were exposed to temperatures 20, 22, 25 and 280C and the number of cells in each temperature treatment was investigated for surveying growth condition. The highest growth and proliferation of cells were observed at 250C. To maintain cell stocks, the cultures were cryopreserved in liquid nitrogen. To characterization of neural stem cells, reactivity of cells from rear part of the brain with anti-nestin marker was investigated, that 11% of cells were immunopositive. According to the high stability of cultures from rear part of the brain, it can be used in different research areas such as virus characterization, ecotoxicology and gene expression.

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Alexander, K., Yang, H.S., and Hinds, P.W. 2004. Cellular senescence requires CDK5 repression of Rac1 activity. Molecular Cell Biology, 24: 2808–2819.
Bani, A., Zadbagher, F., and Jadidi, N. 2008. Brain morphology in Acipenser stellatus and Acipenser persicus brain morphology, in persian, Iranian Journal of Biology, 21(2): 341-350.
Bols, N.C., Dayeh, V.R., Lee, L.E.J., and Schirmer, K. 2005. Use of fish cell lines in the toxicology and ecotoxicology of fish, Piscine cell lines in environmental toxicology, Enviromental toxicology,
6: 43-84.
Bols, N.C., and Lee, L.E.J. 1991. Technology and uses of cell cultures from the tissues and organs of bony fish, Cyto technology, 6: 163-187.
Chi, S.C., Wu, Y.C., and Cheng, T.M. 2005. Persistent infection of betanodavirus in a novel cell line derived from the brain tissue of barramundi Lates calcalifer, Disease of aquatic organisms, 65(2): 91-98.
Grandel, H., Kaslin, J., Ganz, J., Wenzel, I., and Brand, M. 2006. Neural stem cells and neurogenesis in the adult zebrafish brain: origin, proliferation dynamics, migration and cell fate, Development Biology, 295(1): 263–277.
Gregg, C.T., Chojnacki, A.K.,and Weiss, S. 2002. Radial glial cellsas neural precursors: the next generation, Journal of neuroscience research, 69(6): 708-713.
Grunow, B., Noglick, C., Kruse, C., and Gebert, M. 2011. Isolation of cells from Atlantic sturgeon Acipenser oxyrinchus and optimization of culture conditions, Aquatic Biology, 14: 67-75.
Hinsch, K., and Zupanc, G.K. 2006. Isolation, cultivation, and differentiation of neural stem cells from adult fish brain, Journal of Neuroscience Methods, 158: 75–88.  
Kaslin, J., Ganz, J., and Brand, M. 2008. Proliferation, neurogenesis and regeneration in the non-mammalian vertebrate brain, Philos. Trans. R. Soc. Lond., B Biology Science, 363(1489): 101–122.
 Khabbaz, M., Shirazi, S.H., Eric Aghaji, H., and Khabaz, M. 2008. Investigation of economic importance of Persian sturgeon and it,s conservations strategies in Iranian waters, In Persian, 11th Iraniam veterinary congress, Tehran, Iranian veterinary society, http://www.civilica.com/Paper-THVC15-THVC15_249.html.
Lakra, W.S., Swaminathan, T.R., and Joy, K.P. 2011. Development, characterization, conservation and storage of fish cell lines: a review, Fish physiology and biochemistry, 37(1): 1-20.
 Moles, G., Carrillo, M., Mañanos, E., Mylonas C.C., and Zanuy, S. 2007. Temporal profile of brain and pituitary GnRHs, GnRH-R and gonadotropin mRNA expression and content
during early development in European sea bass (Dicentrarchus labrax L[p1] .), Gene Comparative Endocrinology, 150(1): 75–86.
 Neuhaus, J., and Fedoroff, S. 1994. Development of microglia in mouse neopallial cell cultures, Glia, 11(1): 11-17.
Ott, T. 2004. Tissue culture of fish cell lines; National wildlife fish health survey laboratory procedures and protocols, 2th edition, US fish and wildlife service: Washington DC, p: 1-16.
Parameswaran, V., Shukla, R., Bhonde, R.R., and Sahul Hameed, A.S. 2006. Splenic cell line from sea
bass, Lates calcalifer[p2] : establishment and characterization, Aquaculture, 261: 43-53.
 Pitchford, S., De Moor, K., and Glaeser, B.S. 1995. Nerve growth factor stimulates rapid metabolic responses in PC12 cells, American Journal of physiology, 268: 936-943.
 Saha, K., Keung, A., Irwin, E., Li, Y., Little, L., Schaffer, D., and Healy, K.E. 2008. Substrate modulus directs neural stem cell behavior, Biophysics Journal. 95: 4426-4438.
 Rizk, T., Montero-Menei, C., Jollivet, C., Benoit, J.P., and Menei, P. 2004. Pitfalls in the detection of lipid vectors in neural cell culture and in brain tissue, Journal of biomedical materials research, 68(2): 360-364.
Servilia, A., Bufalinoa, M.R., Nishikawac, R., Sanchez de Melod, I., Munoz-Cuetob, J., and Lea, L. 2008. Establishment of long term cultures of neural stem cells from adult sea bass, Dicentrarchus labrax, Comparative biochemistry and physiology Part A: Molecular and integrative physiology, 62(3): 32-36.
 Tang, H.W., Yan, H.L., Hu, X.H., Liang, Y.X., and Shen, X.Y. 1996. Lead cytotoxicity in primary cultured rat astrocytes and Schwann cells, Journal of applied toxicology, 16(3): 187-196.
Veronesi, B. 1992. In vitro screening batteries for neurotoxicants, Neurotoxicology, 13(1): 185-195.
 Wen, C.M., Cheng, Y.H., Huang, Y.F., and Wang, C.S. 2008. Isolation and characterization of a neural progenitor cell line from tilapia brain, Comparative Biochemistry and Physiology Part A: Molecular and integrative Physiology, 149(2): 167-180.
 Yang, H.Y., Lieska, N., Shao, D., Kriho, V., and Pappas, G.D. 1993. Immunotyping of radial glia and their glia derivatives during development of the rat spinal cord, Journal of neurocytology, 22(7): 558-571.
 Zupanc, G.K., and Clint, S.C. 2003. Potential role of radial glia in
adult neurogenesis of teleost fish, Glia, 43(1): 77–86.
Zupanc, G.K., and Horschke, I. 1995. Proliferation zones in the brain of adult gymnotiform fish: a quantitative mapping study, Journal of Comparative Neurology, 353(2): 213–233.
Zupanc, G.K., and Ott, R. 1999. Cell proliferation after lesions in the cerebellum of adult teleost fish: time course, origin, and type of new cells produced, Experimental Neurology, 160(1): 78–87.