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Two skin cell lines from wild-type and albino Japanese flounder (Paralichthys olivaceus): establishment, characterization, virus susceptibility, efficient transfection, and application to albinism study

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Abstract

In order to provide an applicable cell platform to study fish pathology and skin pigmentation, two cell lines derived from skin tissues of wild-type and albino Japanese flounder were established and named JFSK_wt and JFSK_alb, respectively. These two cell lines were cultured for 45 passages within approximately 300 days. JFSK_wt and JFSK_alb cells were maintained in Dulbecco’s Modified Eagle’s Medium and Ham’s F-12 Nutrient Mixture (DMEM/F12) supplemented with antibiotics, fetal bovine serum (FBS), 2-mercaptoethanol (2-Me), N-2-hydroxyethylpiperazine-N′-2-ethanesulfonic acid (HEPES), and basic fibroblast growth factor (bFGF). The optimal growth temperature for JFSK_wt and JFSK_alb cells was 24 °C, and microscopically, the two cell lines were composed of fibroblast-like cells. Chromosomal analysis revealed that JFSK_wt and JFSK_alb cells had an identical diploid karyotype with 2n = 48t. Results of viral inoculation assays revealed that both cell lines shared similar patterns of viral susceptibility to nervous necrosis virus (NNV). High transfection efficiency was observed in JFSK_wt and JFSK_alb cells transfected with a pEGFP-N3 reporter plasmid and Cy3-siRNA. The detection of dermal marker Dermo-1 showed that these two cells were both derived from the dermis. Finally, three genes involved in the melanogenesis pathway, including adenylate cyclase type 5 (adcy5), microphthalmia-associated transcription factor (mitf), and endothelin B receptor (ednrb), were downregulated in JFSK_alb versus JFSK_wt cells. Thus, the two cell lines, sampled from skin tissue of wild-type and albino Japanese flounder will be not only helpful for fish pathogen research but also beneficial for albinism-related gene function studies.

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References

  • Baynash AG, Hosoda K, Giaid A, Richardson JA, Emoto N, Hammer RE, Yanagisawa M (1994) Interaction of endothelin-3 with endothelin-B receptor is essential for development of epidermal melanocytes and enteric neurons. Cell 79:1277–1285

    Article  CAS  PubMed  Google Scholar 

  • Bolker JA, Hill CR (2000) Pigmentation development in hatchery-reared flatfishes. J Fish Biol 56:1029–1052

    Article  Google Scholar 

  • Bols NC, Dayeh VR, Lee LEJ, Schirmer K (2005) Chapter 2 Use of fish cell lines in the toxicology and ecotoxicology of fish. Piscine cell lines in environmental toxicology. Biochem Mol Biol Fishes 6:43–84

    Article  CAS  Google Scholar 

  • Chen SL, Ren GC, Sha ZX, Shi CY (2004) Establishment of a continuous embryonic cell line from flounder (Paralichthys olivaceus) for virus isolation. Dis Aquat Org 60:241–246

    Article  PubMed  Google Scholar 

  • Chen SL, Ren GC, Sha ZX, Hong Y (2005) Development and characterization of a continuous embryonic cell line from turbot (Scophthalmus maximus). Aquaculture 249:63–68

    Article  CAS  Google Scholar 

  • Clem LW, Ely IE, Wilson M, Chinchar VG, Stuge T, Barker K (1996) Fish immunology: the utility of immortalized lymphoid cells-a mini review. Vet Immunol lmmunopathol 54:137–144

    Article  CAS  Google Scholar 

  • Curtis PA, Drawbridge M, Iwamoto T, Nakai T, Hedrick RP, Gendron AP (2001) Nodavirus infection of juvenile white seabass, Atractoscion nobilis, cultured in southern California: first record of viral nervous necrosis (VNN) in North America. J Fish Dis 24:263–271

    Article  Google Scholar 

  • Dou S, Masuda R, Tanaka M, Tsukamoto K (2003) Identification of factors affecting the growth and survival of the settling Japanese flounder larvae, Paralichthys olivaceus. Aquaculture 218:309–327

    Article  Google Scholar 

  • Dumont E, Rallière C, Rescan PY (2008) Identification of novel genes including Dermo-1, a marker of dermal differentiation, expressed in trout somitic external cells. J Exp Biol 211:1163–1168

    Article  CAS  PubMed  Google Scholar 

  • Fan LC, Collodi P (2002) Progress towards cell-mediated gene transfer in zebrafish. Briefing Funct Genomics Proteomics 1:131–138

    Article  Google Scholar 

  • Fonds M, Tanaka M, Van der Veer HW (1995) Feeding and growth of juvenile Japanese flounder Paralichthys olivaceus in relation to temperature and food supply. Neth J Sea Res 34:111–118

    Article  Google Scholar 

  • Fujiwara A, Fujiwara M, Nishida-Umehara C, Abe S, Masaoka T (2007) Characterization of Japanese flounder karyotype by chromosome bandings and fluorescence in situ hybridization with DNA markers. Genetica 131:267–274

    Article  PubMed  Google Scholar 

  • Goswami M, Sharma BS, Tripathi AK, Yadav K, Bahuguna SN, Nagpure NS, Lakra WS, Jena JK (2012) Development and characterization of cell culture systems from Puntius (Tor) chelynoides, (McClelland). Gene 500:140–147

    Article  CAS  PubMed  Google Scholar 

  • Gui L, Zhang P, Liang X, Su M, Wu D, Zhang J (2016) Adaptive responses to osmotic stress in kidney-derived cell lines from Scatophagus argus, a euryhaline fish. Gene 583:134–140

    Article  CAS  PubMed  Google Scholar 

  • Higaki S, Koyama Y, Shirai E, Yokota T, Fujioka Y, Sakai N, Takada T (2013) Establishment of testicular and ovarian cell lines from Honmoroko (Gnathopogon caerulescens). Fish Physiol Biochem 39:701–711

    Article  CAS  PubMed  Google Scholar 

  • Kang MS, Oh MJ, Kim YJ, Kawai K, Jung SJ (2003) Establishment and characterization of two new cell lines derived from flounder, Paralichthys olivaceus, (Temminck & Schlegel). J Fish Dis 26:657–665

    Article  CAS  PubMed  Google Scholar 

  • Lakra WS, Swaminathan TR, Joy KP (2011) Development, characterization, conservation and storage of fish cell lines a review. Fish Physiol Biochem 37:1–20

    Article  CAS  PubMed  Google Scholar 

  • Lei XY, Chen ZY, He LB, Pei C, Yuan XP, Zhang QY (2012) Characterization and virus susceptibility of a skin cell line from red-spotted grouper (Epinephelus akaara). Fish Physiol Biochem 38:1175–1182

    Article  CAS  PubMed  Google Scholar 

  • Li H, Tong SL, Hetick FM (1997) The tissue culture of Paralichthys olivaceus and the derivation of gill cell line. J Fish China 21:193–196

    Google Scholar 

  • Lu MW, Chao YM, Guo TC, Santi N, Evensen O, Kasani SK, Hong JR, Wu JL (2008) The interferon response is involved in nervous necrosis virus acute and persistent infection in zebrafish infection model. Mol Immunol 45:1146–1152

    Article  CAS  PubMed  Google Scholar 

  • Ma J, Sun S, Zeng L, Lu Y (2013) Establishment, characterization and viral susceptibility of two cell lines derived from leopard wrasse Macropharyngodon geoffroy. J Fish Biol 83:560–573

    Article  CAS  PubMed  Google Scholar 

  • Medzon EL, Gedies A (1971) Substitution of 4-(2-hydroxyethyl)-1-piperazineethane sulfonic acid (HEPES) for bicarbonate in protein-free animal cell culture medium: application to vaccinia virus quantitation and fluorogenic acetylesterase assay in living LM cells. Can J Microbiol 17(5):651–653

    Article  CAS  PubMed  Google Scholar 

  • Munday BL, Kwang J, Moody N (2002) Betanodavirus infections of teleost fish: a review. J Fish Dis 25:127–142

    Article  Google Scholar 

  • Muroga K (2001) Viral and bacterial diseases of marine fish and shellfish in Japanese hatcheries. Aquaculture 202:23–44

    Article  Google Scholar 

  • Nguyen HD, Mekuchi T, Imura K, Nakai T, Nishizawa T, Muroga K (1994) Occurrence of viral nervous necrosis (VNN) in hatchery-reared juvenile Japanese flounder Paralichthys olivaceus. Fisheries Sci 5:551–554

  • Nguyen HD, Nakai T, Muroga K (1996) Progression of striped jack nervous necrosis virus (SJNNV) infection in naturally and experimentally infected striped jack Pseudocaranx dentex larvae. Dis Aquat Organ 24:99–105

  • Nishio T, Usami M, Awaji M, Shinohara S, Sato K (2016) Dual effects of acetylsalicylic acid on ERK signaling and mitf transcription lead to inhibition of melanogenesis. Mol Cell Biochem 412:101–110

    Article  CAS  PubMed  Google Scholar 

  • Ogata HY, Oku H, Murai T (2002) Growth, feed efficiency and feed intake of offspring from selected and wild Japanese flounder (Paralichthys olivaceus). Aquaculture 211:183–193

    Article  Google Scholar 

  • Oh YB, Lee JS, Park EH (2001) Fish cell line as an in vitro test system for analyzing chromosome aberrations. Bull Environ Contam Toxicol 67:6–11

    CAS  PubMed  Google Scholar 

  • Ren GC, Chen SL, Sha ZX (2007) Development and characterization of a continuous embryonic cell line from southern flounder (Paralichthys lethostigma). J Fish Sci China 14:579–583

    Google Scholar 

  • Rode M, Berg T, GjØen T (1997) Effect of temperature on endocytosis and intracellular transport in the cell line SHK-1 derived from salmon head kidney. Comp Biochem Physiol A 117:531–537

    Article  Google Scholar 

  • Sato GH, Sato JD, Okamoto T, Mckeehan WL, Barnes DW (2010) Tissue culture: the unlimited potential. In Vitro Cell Dev Biol: Anim 46:590–594

    Article  Google Scholar 

  • Sun A, Fan T, Yang X, Xu B, Xu X, Guo X (2011) Establishment of a spleen cell line from large yellow croaker Pseudosciaena crocea and its primitive application in foreign gene transfection. J Ocean Univ China 10:397–401

    Article  CAS  Google Scholar 

  • Sun A, Chen SL, Gao FT, Li HL, Liu XF, Wang N, Sha ZX (2015) Establishment and characterization of a gonad cell line from half-smooth tongue sole Cynoglossus semilaevis pseudomale. Fish Physiol Biochem 41:673–683

    Article  CAS  PubMed  Google Scholar 

  • Tabata K (1991) Genetic factors on the albinism of hirame Paralichthys olivaceus. Aquacult Sci 39:29–35

    Google Scholar 

  • Takahashi Y (1994) Influence of stocking density and food at late phase of larval period on hypermalanosis on the blind body side in juvenile Japanese flounder. Nippon Suisan Gakkaishi 60:593–598

    Article  Google Scholar 

  • Tanaka S, Kuriyama I, Nakai T, Miyazaki T (2003) Susceptibility of cultured juveniles of several marine fish to the seven band grouper nervous necrosis virus. J Fish Dis 26:109–115

    Article  CAS  PubMed  Google Scholar 

  • Tong SL, Li H, Miao HZ (1997) The establishment and partial characterization of a continuous fish cell line FG-9307 from the gill of flounder Paralichthys olivaceus. Aquaculture 156:327–333

    Article  Google Scholar 

  • Tu J, Chen W, Fu X, Lin Q, Chang O, Zhao L, Lan J, Li N, Lin L (2016) Susceptibility of Chinese perch brain (CPB) cell and mandarin fish to red-spotted grouper nervous necrosis virus (RGNNV) infection. Int J Mol Sci 17:740

    Article  PubMed Central  Google Scholar 

  • Wang N, Wang XL, Sha ZX, Tian YS, Chen SL (2010) Development and characterization of a new marine fish cell line from turbot (Scophthalmus maximus). Fish Physiol Biochem 36:1227–1234

    Article  CAS  PubMed  Google Scholar 

  • Wang N, Wang XL, Sha ZX, Tian YS, Chen SL (2011) Establishment, characterization and virus susceptibility of a kidney-derived cell line from southern flounder, Paralichthys lethostigma Jordan Gilbert. J Fish Dis 34:81–85

    Article  CAS  PubMed  Google Scholar 

  • Wei YB, Fan TJ, Jiang GJ, Sun A, Xu XH, Wang J (2009) Establishment of a novel fin cell line from Brown-marbled grouper, Epinephelus fuscoguttatus (Forsskål), and evaluation of its viral susceptibility. Aquac Res 40:1523–1531

    Article  CAS  Google Scholar 

  • Whitear M (1970) The skin surface of bony fishes. J Zool 160:437–454

    Article  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  • Ye HQ, Chen SL, Sha ZX, Xu MY (2006) Development and characterization of five cell lines from heart, liver, spleen and head kidney of sea perch (Lateolabrax japonicus). J Fish Biol 69:115–126

    Article  CAS  Google Scholar 

  • Yoshikoshi K, Inoue K (1990) Viral nervous necrosis in hatchery-reared larvae and juveniles of Japanese parrotfish, Oplegnathus fasciatus (Temminck & Schlegel). J Fish Dis 13:69–77

    Article  Google Scholar 

  • Zhao Z, Montgomery-Brock D, Lee CS, Lu Y (2003) Establishment, characterization and viral susceptibility of 3 new cell lines from snakehead, Channa striatus (Blooch). Methods Cell Sci 25:155–166

    Article  PubMed  Google Scholar 

  • Zheng Y, Wang N, Xie MS, Sha ZX, Chen SL (2012) Establishment and characterization of a new fish cell line from head kidney of half-smooth tongue sole (Cynoglossus semilaevis). Fish Physiol Biochem 38:1635–1643

    Article  CAS  PubMed  Google Scholar 

  • Zheng Y, Peng LM, You F, Zou YX, Zhang PJ, Chen SL (2015) Establishment and characterization of a fish-cell line from the brain of Japanese flounder Paralichthys olivaceus. J Fish Biol 87:115–122

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

This work was supported by grants from the National Natural Science Foundation of China (31472273), and the Taishan Scholar Climb Project of Shandong Province.

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Authors and Affiliations

  1. Key Laboratory for Sustainable Development of Marine Fisheries, Ministry of Agriculture, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, 106 Nanjing Road, Qingdao, 266071, China

    Ruoqing Wang, Nianwei Zhang, Renkai Wang, Shengpeng Wang & Na Wang

  2. Laboratory for Marine Fisheries Science and Food Production Processes, Qingdao National Laboratory for Marine Science and Technology, Qingdao, 266235, China

    Na Wang

  3. College of Fisheries and Life Science, Shanghai Ocean University, Shanghai, 201306, China

    Ruoqing Wang, Nianwei Zhang & Renkai Wang

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  1. Ruoqing Wang
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  2. Nianwei Zhang
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  4. Shengpeng Wang
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  5. Na Wang
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Contributions

NW conceived and designed the experiments. NW and RQW analyzed the experimental results and wrote the paper. NWZ conducted the primary cell culture. NW, RQW, and RKW subcultured JFSK_wt and JFSK_alb cell lines. RQW and NW conducted chromosome analysis, virus infection experiments, reporter gene and siRNA transfection assay, and qRT-PCR experiments. SPW provided the NNV infected grouper. All authors have read and approved the final manuscript.

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Correspondence to Na Wang.

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Wang, R., Zhang, N., Wang, R. et al. Two skin cell lines from wild-type and albino Japanese flounder (Paralichthys olivaceus): establishment, characterization, virus susceptibility, efficient transfection, and application to albinism study. Fish Physiol Biochem 43, 1477–1486 (2017). https://doi.org/10.1007/s10695-017-0386-8

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