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Establishment and characterization of a fin cell line from blunt snout bream, Megalobrama amblycephala

Fish Physiology and Biochemistry volume 39, pages 1399–1410 (2013)Cite this article

Abstract

This study established and characterized a new cell line (MAF) from the fin of blunt snout bream (Megalobrama amblycephala), a freshwater fish cultivated in China. MAF cells proliferated well in medium 199 supplemented with 10 % fetal bovine serum at 28 °C and have been subcultured more than 95 times in almost a year. MAF cells were revived at 90–95 % viability after 3–6 months of storage in liquid nitrogen. Karyotyping indicated that the modal chromosome number of MAF cells was 48. The MAF cell line consisted predominantly of fibroblastic and epithelial-like cells from M. amblycephala, which was confirmed by immunofluorescence and mitochondrial 12s rRNA sequencing. Viral susceptibility tests showed that MAF cells were susceptible to infection by snakehead rhabdovirus, spring viremia carp virus, and channel catfish virus, which was demonstrated by the presence of cytopathic effect, high viral titers, and PCR products. Bacterial cytotoxicity studies showed that extracellular products from Aeromonas hydrophila were toxic to MAF cells. Cu2+ was also cytotoxic to MAF cells, and the 24-h IC50 value was 144.48 μmol/l. When MAF cells were transfected with pEGFP-N1 plasmid, bright fluorescent signals were observed, and the transfection efficiency reached up to 5 %. These results suggest that the MAF cell line may provide a valuable tool for studying virus pathogenesis, as well as cytotoxicity testing and genetic manipulation studies.

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References

  • Babu VS, Nambi KSN, Chandra V, Ishaq Ahmed VP, Bhonde R, Sahul Hameed AS (2011) Establishment and characterization of a fin cell line from Indian walking catfish, Clarias batrachus (L.). J Fish Dis 34(4):355–364

    PubMed  Article  CAS  Google Scholar 

  • Bai JJ, Lao HH, Ye X, Li YH, Luo JR (2001) Molecular cloning and sequence analysis of insulin-like growth factor-I cDNA from bluntnose bream (Megalobrama amblycephala). Zool Res 22(6):502–506

    CAS  Google Scholar 

  • Balebona MC, Moriñigo MA, Borrego JJ (1995) Role of extracellular products in the pathogenicity of Vibrio strains on cultured gilt-head seabream (Sparus auarata). Microbiologia 11(4):439–446

    PubMed  CAS  Google Scholar 

  • Bejar J, Borrego JJ, Alvarez MC (1997) A continuous cell line from the cultured marine fish gilt-head seabream (Sparus aurata L.). Aquaculture 150(1–2):143–153

    Article  Google Scholar 

  • Bowser PR, Plumb JA (1980) Fish cell lines establishment of a line from ovaries of channel catfish. In Vitro 16:365–368

    PubMed  Article  CAS  Google Scholar 

  • CAFS (Chinese Academy of Fishery Sciences) (2001) Fishery statistic data. Chinese Academy of Fishery Sciences, Beijing. (In Chinese)

  • Chen QL, Luo Z, Liu X, Song YF, Liu CX, Zheng JL, Zhao YH (2013) Effects of waterborne chronic copper exposure on hepatic lipid metabolism and metal-element composition in synechogobius hasta. Arch Environ Contam Toxicol 64(2):301–315

    PubMed  Article  CAS  Google Scholar 

  • Chi SC, Hu WW, Lo BJ (1999) Establishment and characterization of a continuous cell line (GF-1) derived from grouper, Epinephelus coioides (Hamilton): a cell line susceptible to grouper nervous necrosis virus (GNNV). J Fish Dis 22(3):173–182

    Article  Google Scholar 

  • Du YD, Chen XX, Wu ZX, Wang M, Liu Q, Xia J, Wang SY, Li LJ (2009) Development of a PCR—ELISA assay for the detection of the channel catfish virus. J Huazhong Agr Univ 28(5):604–608 (Chinese with English abstract)

    CAS  Google Scholar 

  • Du ZY, Ma T, Winterthun S, Kristiansen K, Froyland L, Madsen L (2010) Beta-oxidation modulates metabolic competition between eicosapentaenoic acid and arachidonic acid regulating prostaglandin E(2) synthesis in rat hepatocytes-Kupffer cells. Biochim Biophys Acta 1801(4):526–536

    PubMed  Article  CAS  Google Scholar 

  • Earley EM (1975) Chromosome preparations from monolayer cell cultures. Methods Cell Sci 1:31–35

    CAS  Google Scholar 

  • Fan Tj, Ren BX, Gang XF, Yu QT, Wang LY (2010) Establishment of a turbot fin cell line and its susceptibility to turbot reddish body iridovirus. Cytotechnology 62(3):217–223

    PubMed  Article  Google Scholar 

  • Fijan N, Sulimanovic D, Bearzotti M, Muzinic D, Zwillenberg LO, Chilmonczyk S, Vautherot JF, De Kinkelin P (1983) Some properties of the Epithelioma papulosum cyprinid (EPC) cell line from carp Cyprinus carpio. Ann Virol 134E:207–220

    Google Scholar 

  • Forest D, Nishikawa R, Kobayashi H, Parton A, Bayne CJ, Barnes DW (2007) RNA expression in a cartilaginous fish cell line reveals ancient 3′ noncoding regions highly conserved in vertebrates. PNAS 104(4):1224–1229

    PubMed  Article  CAS  Google Scholar 

  • Gao ZX (2010) Study on sex control, sex determination mechanism and sex related markers in bluegill sunfish Lepomis macrochirus. Dissertation, Huazhong agricultural university (Chinese with English abstract)

  • Huang XH, Huang YH, Ouyang ZL, Qin QW (2011) Establishment of a cell line from the brain of grouper (Epinephelus akaara) for cytotoxicity testing and virus pathogenesis. Aquaculture 311(1):65–73

    Article  CAS  Google Scholar 

  • Ishaq Ahmed VP, Chandra V, Parameswaran V, Venkatesan C, Shukla R, Bhonde R, Sahul Hameed AS (2008) A new epithelial-like cell line from eye muscle of catla Catla catla (Hamilton): development and characterization. J Fish Biol 72(8):2026–2038

    Article  Google Scholar 

  • Ishaq Ahmed VP, Babu VS, Chandra V, Nambi KS, Thomas J, Bhonde R, Sahul Hameed AS (2009a) A new fibroblastic-like cell line from heart muscle of the Indian major carp (Catla catla): development and characterization. Aquaculture 293(3–4):180–186

    Article  Google Scholar 

  • Ishaq Ahmed VP, Chandra V, Sudhakaran R, Rajesh Kumar S, Sarathi M, Sarath Babu V, Ramesh B, Sahul Hameed AS (2009b) Development and characterization of cell lines derived from rohu, Labeo rohita (Hamilton), and catla, Catla catla (Hamilton). J Fish Dis 32(3):211–218

    Article  Google Scholar 

  • Ke H (1965) The artificial reproduction and culture experiment of Megalobrama amblycephala. Acta Hydrobiol Sin 5:282–283

    Google Scholar 

  • Ku CC, Teng YC, Wang CS, Lu CH (2009) Establishment and characterization of three cell lines derived from the rock fish grouper Epinephelus quoyanus: use for transgenic studies and cytotoxicity testing. Aquaculture 294(1–2):147–151

    Article  CAS  Google Scholar 

  • Lai YS, Murali S, Ju HY, Wu MF, Guo IC, Chen SC, Fang K, Chang CY (2000) Two iridovirus-susceptible cell lines established from kidney and liver of grouper, Epinephelus awoara (Temminck & Schlegel), and partial characterization of grouper iridovirus. J Fish Dis 23(6):379–388

    Article  Google Scholar 

  • Liu B, Ge XP, Xie J, Xu P, He YJ, Cui YT, Ming JH, Zhou QL, Pan LK (2012) Effects of anthraquinone extract from Rheum officinale Bail on the physiological responses and HSP70 gene expression of Megalobrama amblycephala under Aeromonas hydrophila infection. Fish Shellfish Immun 32(1):1–7

    Article  Google Scholar 

  • Lu Y, Aguirre AA, Braun RC, Loh PC (1998) Establishment of monk seal cell lines. In Vitro Cell Dev Biol-Animal 34:367–369

    Article  CAS  Google Scholar 

  • Mauger PE, Labbé C, Bobe J, Cauty C, Leguen I, Baffet G, Le Bail PY (2009) Characterization of goldfish fin cells in culture: some evidence of an epithelial cell profile. Comp Biochem Phys B 152(3):205–215

    Article  Google Scholar 

  • Nicholson BL, Danner DJ, Wu JL (1987) Three new continuous cell lines from marine fishes of Asia. In Vitro Cell Dev Biol 23(3):199–204

    Article  Google Scholar 

  • Ou-yang ZL, Huang XH, Huang EY, Huang YH, Gong J, Sun JJ, Qin QW (2010) Establishment and characterization of a new marine fish cell line derived from red-spotted grouper Epinephelus akaara. J Fish Biol 77(5):1083–1095

    PubMed  Article  CAS  Google Scholar 

  • Parameswaran V, Shukla R, Bhonde RR, Sahul Hameed AS (2006) Splenic cell line from sea bass, Lates calcarifer: establishment and characterization. Aquaculture 261(6):43–53

    Article  CAS  Google Scholar 

  • Phelan PE, Pressley ME, Witten PE, Mellon MT, Blake S, Kim CH (2005) Characterization of snakehead rhabdovirus infection in zebrafish (Danio rerio). J Virol 79(3):1842–1852

    PubMed  Article  CAS  Google Scholar 

  • Reed LJ, Muench H (1938) A simple method of estimating fifty percent endpoints. Am J Epidemiol 27(3):493–497

    Google Scholar 

  • Rougée L, Ostrander GK, Richmond RH, Lu YA (2007) Establishment, characterization, and viral susceptibility of two cell lines derived from goldfish Carassius auratus muscle and swim bladder. Dis Aquat Org 77(2):127–135

    PubMed  Article  Google Scholar 

  • Sahul Hameed AS, Parameswaran V, Shukla R, Bright Singh IS, Thirunavukkarasu AR, Bhonde RR (2006) Establishment and characterization of India’s first marine fish cell line (SISK) from the kidney of sea bass (Lates calcarifer). Aquaculture 257:92–103

    Article  Google Scholar 

  • Song YL, Liu L, Shen HX, You J, Luo YK (2011) Effect of sodium alginate-based edible coating containing different anti-oxidants on quality and shelf life of refrigerated bream (Megalobrama amblycephala). Food Control 22(3–4):608–615

    Article  CAS  Google Scholar 

  • Stone DM, Ahne W, Denham KL, Dixon PF, Liu CTY, Sheppard AM, Taylor GR, Way K (2003) Nucleotide sequence analysis of the glycoprotein gene of putative spring viraemia of carp virus and pike fry rhabdovirus isolates reveals four genogroups. Dis Aquat Organ 53:203–210

    PubMed  Article  CAS  Google Scholar 

  • Swaminathan TR, Lakra WS, Gopalakrishnan A, Basheer VS, Khushwaha B, Sajeela KA (2010) Development and characterization of a new epithelial cell line PSF from caudal fin of Green chromide, Etroplus suratensis (Bloch, 1790). In Vitro Cell Dev Biol Anim 46(8):647–656

    PubMed  Article  CAS  Google Scholar 

  • Tan F, Wang M, Wang W, Lu Y (2008) Comparative evaluative cytotoxicity of six fish cell lines to four heavy metals. Toxicol In Vitro 22(1):164–170

    PubMed  Article  CAS  Google Scholar 

  • Tong SL, Li H, Miao HZ (1997) Four continuous cell lines of three marine fish species were established. Prog Biot 17(3):324–328 (Chinese with English abstract)

    Google Scholar 

  • Wang WW, Zhou ZG, He SX, Liu YC, Cao YN, Shi PJ, Yao B, Ring E (2010) Identification of the adherent microbiota on the gills and skin of poly-cultured gibel carp (Carassius auratus gibelio) and bluntnose black bream (Megalobrama amblycephala Yih). Aquac Res 41(9):e72–e83

    Article  CAS  Google Scholar 

  • Wang XL, Wang N, Sha ZX, Chen SL (2011) Establishment and characterization of a testicular cell line from the half-smooth tongue sole, Cynoglossus semilaevis. Int J Biol Sci 7(4):452–459

    PubMed  Google Scholar 

  • Wolf K, Quimby MC (1962) Established eurythermic line of fish cells in vitro. Science 135(3508):1065–1066

    PubMed  Article  CAS  Google Scholar 

  • Xiao Q, Li DZ, Liu HY (2011) A flounder (Paralichthys olivaceus) gill cell line as in vitro acute assay system of nonylphenol cytotoxicity. Environ Monit Assess 175(1–4):315–319

    PubMed  Article  CAS  Google Scholar 

  • Xu WY, Zhao YJ, Zhang H (2004) The high yields of channel catfish (Ictalurus punctatus) and Chinese bream (Megalobrama amblycephala) in polyculture ponds. Fish Sci 23(10):22–24

    Google Scholar 

  • Yan W, Nie P, Lu Y (2011) Establishment, characterization and viral susceptibility of a new cell line derived from goldfish, Carassius auratus (L.), tail fin. J Fish Dis 34(10):757–768

    PubMed  Article  CAS  Google Scholar 

  • Yin HB, Fan ZT, Sun ZW (1995) The Karyotype and DNA content analysis of blunt snout bream (Megalobrama amblycephala). Chinese J Fish 8(1):22–26 (Chinese with English abstract)

    Google Scholar 

  • Zeng C, Cao XJ, Luo W, Liu XL, Gao ZX, Qian XQ, Wang WM (2011) Analysis of effects of morphometric traits on body weight of Megalobrama amblycephala. Chinese Agr Sci Bull 27(32):66–71 (Chinese with English abstract)

    Google Scholar 

  • Zhang B, Chen SL (2011) Decades of researching progresses of fish cell culture and its application prospects. Mar Sci 35(7):113–121 (Chinese with English abstract)

    Google Scholar 

  • Zhang GY, Wang Y, Peng KM, Wang C, Tian G, Liu HZ (2009) Strategic localization of toll-like receptor 4 in the digestive tract of blunt snout bream (Megalobrama amblycephala). Anat Histol Embryol 38(6):401–405

    PubMed  Article  CAS  Google Scholar 

  • Zhang LL, Luo Q, Fang Q, Wang YP (2010) An improved RT-PCR assay for rapid and sensitive detection of grass carp reovirus. J Virol Methods 169:28–33

    Google Scholar 

  • Zhao YH, Gul Y, Li S, Wang WM (2011) Cloning, identification and accurate normalization expression analysis of PPARa gene by GeNorm in Megalobrama amblycephala. Fish Shellfish Immunol 31(3):462–468

    PubMed  Article  CAS  Google Scholar 

  • Zhou GZ, Li ZQ, Yuan XP, Zhang QY (2007) Establishment, characterization, and virus susceptibility of a new marine cell line from red spotted grouper (Epinephelus akaara). Mar Biotechnol 9(3):370–376

    PubMed  Article  CAS  Google Scholar 

  • Zhou GZ, Gui L, Li ZQ, Yuan XP, Zhang QY (2008) Establishment of a Chinese sturgeon Acipenser sinensis tail-fin cell line and its susceptibility of rogiridovirus. J Fish Biol 73(8):2058–2067

    Article  CAS  Google Scholar 

  • Zhu DM, Song W, Yang K, Cao XJ, Gul Y, Wang WM (2012) Flow cytometric determination of genome size for eight commercially important fish species in China. In Vitro Cell Dev Biol Anim 48:507–517

    PubMed  Article  Google Scholar 

  • Zou SM, Li SF, Cai WQ, Zhao JL, Yang HY (2004) Establishment of fertile tetraploid population of blunt snout bream (Megalobrama amblycephala). Aquaculture 238(1–4):155–164

    Article  Google Scholar 

  • Zou SM, Jiang XY, He ZZ, Yuan J, Yuan XN, Li SF (2007) Hox gene clusters in blunt snout bream, Megalobrama amblycephala and comparison with those of zebrafish, fugu and medaka genomes. Gene 400(1–2):60–70

    PubMed  Article  CAS  Google Scholar 

  • Zuo WG, Qian HX, Xu YF, Du SY, Yang XL (1986) A cell line derived from the kidney of grass carp (Ctenopharyncodon idellus). J Fish China 10(1):11–17 (Chinese with English abstract)

    Google Scholar 

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Acknowledgments

This study was funded by the earmarked fund for Modern Agro-industry Technology Research System entitled “Staple Freshwater Fishery Industry Technology System” (No. CARS-46-05) and Fundamental Research Funds for the Central Universities (No. 2011PY023).

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  1. Key Laboratory of Agricultural Animal Genetics, College of Fisheries, Breeding and Reproduction of Ministry of Education, Huazhong Agricultural University, Wuhan, 430070, People’s Republic of China

    Dong-Mei Zhu, Kun Yang, Wei-Min Wang & Wen Song

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  1. Dong-Mei Zhu
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  2. Kun Yang
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  3. Wei-Min Wang
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  4. Wen Song
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Correspondence to Wei-Min Wang.

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Zhu, DM., Yang, K., Wang, WM. et al. Establishment and characterization of a fin cell line from blunt snout bream, Megalobrama amblycephala . Fish Physiol Biochem 39, 1399–1410 (2013). https://doi.org/10.1007/s10695-013-9794-6

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  • DOI: https://doi.org/10.1007/s10695-013-9794-6

Keywords

  • Megalobrama amblycephala
  • Fin
  • Cell line
  • Cytotoxicity
  • Transfection