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Evaluation of Various Tissues of the Caerulean Damsel, Pomacentrus caeruleus for Initiating In Vitro Cell Culture Systems

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Abstract

Explantation and trypsinisation methods for tissue dissociation were attempted for the establishment of primary cell cultures from the caerulean damsel, Pomacentrus caeruleus. Among the tissues taken, fin, liver and caudal peduncle showed good attachment with emergence of cells. The cells were best suited to grow in Leibovitz’s L-15 basal medium supplemented with foetal bovine serum (initially 20 % which was later reduced to 5–10 % during subsequent passages) at an ambient temperature of 28 ± 2 °C and pH 7.2 ± 0.2. These cultures persisted at temperatures from 17 to 32 °C, and proliferated at temperatures from 24 to 30 °C. The cells have been cryopreserved successfully with a survival rate of 80 %. Results suggest that fin, caudal peduncle and liver cell cultures have potential for development into cell lines.

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References

  1. Silas EG, Gopalakrishnan A, Ramachandran A, Anna Mercy TV, Kripan Sarkar, Pushpangadan KR, Anil Kumar P, Ram Mohan MK, Anikuttan KK (2011) Guidelines for green certification of freshwater ornamental fish. The Marine Products Export Development Authority, Kochi, India. xii+106p

  2. FAO (2008) http://www.fao.org/fi/statist/FISOFT/FISHPLUS.asp. Accessed 10 Jul 2015

  3. Gopakumar G, Madhu K, Madhu R, Ignatius B, Krishnan L, Mathew G (2009) Broodstock development, breeding and seed production of selected marine food fishes and ornamental fishes. Mar Fish Info Serv T&E Ser 201:1–9

    Google Scholar 

  4. Lakra WS, Swaminathan TR, Joy KP (2010) Development, characterization, conservation and storage of fish cell lines. Fish Physiol Biochem 37:1–20. doi:10.1007/s10695-010-9411-x

    Article  PubMed  Google Scholar 

  5. Wolf K, Mann JA (1980) Poikilotherm vertebrate cell lines and viruses: a current listing for fishes. In Vitro 16(2):168–179. doi:10.1007/BF02831507

    Article  CAS  PubMed  Google Scholar 

  6. Hightower LE, Renfro JL (1988) Recent applications of fish cell culture to biomedical research. J Exp Zool 248:290–302

    Article  CAS  PubMed  Google Scholar 

  7. Bols NC, Lee LEJ (1991) Technology and uses of cell cultures from tissues and organs of bony fish. Cytotechnol 6(3):163–187. doi:10.1007/BF00624756

    Article  CAS  Google Scholar 

  8. Burkhart JG (2000) Fishing for mutations. Nat Biotechnol 18:21–22. doi:10.1038/71869

    Article  CAS  PubMed  Google Scholar 

  9. Hackett PB, Alvarez MC (2000) The molecular genetics of transgenic fish. In: Fingerman M, Nagabhushanam R (eds) Recent advances in marine biotechnology: vol. 4. Aquaculture: part B fishes. Science Publishers, New York, pp 77–145

    Google Scholar 

  10. Gallardo-Gálvez JB, Méndez T, Béjar J, Alvarez MC (2011) Endogenous transposases affect differently Sleeping Beauty and Frog Prince transposons in fish cells. Mar Biotechnol 13:695–705. doi:10.1007/s10126-010-9331-x

    Article  PubMed  Google Scholar 

  11. Molina A, Carpeaux R, Martial JA, Muller M (2002) A transformed fish cell line expressing a green fluorescent protein-luciferase fusion gene responding to cellular stress. Toxicol In Vitro 16(2):201–207

    Article  CAS  PubMed  Google Scholar 

  12. Kawakami K, Abe G, Asada T, Asakawa K, Fukuda R, Ito A, Lal P, Mouri N, Muto A, Suster ML, Takakubo H, Urasaki A, Wada H, Yoshida M (2010) zTrap: zebrafish gene trap and enhancer trap database. BMC Dev Biol 10:105. doi:10.1186/1471-213X-10-105

    Article  PubMed  PubMed Central  Google Scholar 

  13. Crespo B, Zanuy S, Gómez A (2013) Development of an in vitro system for functional studies of ovarian follicular cells in European sea bass (Dicentrarchus labrax). Cytotechnol 65(2):273–286. doi:10.1007/s10616-012-9484-8

    Article  CAS  Google Scholar 

  14. Kaltenbach JP, Kaltenbach MH, Lyons WB (1958) Nigrosin as a dye for differentiating live and dead ascites cells. Exp Cell Res 15:112–117

    Article  CAS  PubMed  Google Scholar 

  15. Nicholson BL (1989) Fish cell culture: an update. Adv in Cell Culture 7:1–18

    Article  CAS  Google Scholar 

  16. Bols NC, Lee LEJ (1991) Technology and uses of cell cultures from tissues and organs of bony fish. Cytotechnol 6(3):163–187. doi:10.1007/BF00624756

    Article  CAS  Google Scholar 

  17. Lakra WS, Goswami M, Yadav K, Gopalakrishnan A, Patiyal RS, Singh M (2011) Development and characterization of two cell lines PDF and PDH from Puntius denisonii (Day 1865). In Vitro Cell Dev Biol Anim 47:89–94. doi:10.1007/s11626-010-9374-3

    Article  PubMed  Google Scholar 

  18. Lakra WS, Goswami M (2011) Development and characterization of a continuous cell line PSCF from Puntius sophore. J Fish Biol 78:987–1001. doi:10.1111/j.1095-8649.2010.02891.x

    Article  CAS  PubMed  Google Scholar 

  19. 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 34:757–768. doi:10.1111/j.1365-2761.2011.01292.x

    Article  CAS  PubMed  Google Scholar 

  20. Swaminathan TR, Lakra WS, Gopalakrishnan A, Basheer VS, Kushwaha B, Sajeela KA (2012) Development and characterization of a fibroblastic-like cell line from caudal fin of the red-line torpedo, Puntius denisonii (Day) (Teleostei: Cyprinidae). Aquac Res 43:498–508. doi:10.1111/j.1365-2109.2011.02854.x

    Article  Google Scholar 

  21. Sobhana KS, George KC, Ravi GV, Ittoop G, Paulraj R (2009) Development of a cell culture system from gill explants of the grouper, Epinephelus malabaricus (Bloch and Shneider). Asian Fish Sci 22:1–6

    Google Scholar 

  22. 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. doi:10.1111/j.1365-2109.2009.02253.x

    Article  CAS  Google Scholar 

  23. Faisal M, Rutan BJ, Sami-Demmerle S (1995) Development of continuous liver cell cultures from the marine teleost, spot (Leiostomus xanthurus, Pisces: Sciaenidae). Aquaculture 132:59–72. doi:10.1016/0044-8486(94)00382-X

    Article  Google Scholar 

  24. 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 and Schlegel), and partial characterization of grouper iridovirus. J Fish Dis 23:379–388. doi:10.1046/j.1365-2761.2000.00247.x

    Article  Google Scholar 

  25. Williams LM, Crane MSJ, Gudkovs N (2003) Development and characterisation of pilchard (Sardinops sagax neopilchardus) cell lines derived from liver and heart tissues. Methods Cell Sci 25:105–113. doi:10.1007/s11022-004-9801-5

    Article  PubMed  Google Scholar 

  26. Ye HQ, Chen SL, Sha ZX, Xu MY (2006) Development and characterization of cell lines from heart, liver, spleen and head kidney of sea perch, Lateolabrax japonicas. J Fish Biol 69(Supplement A):115–126. doi:10.1111/j.1095-8649.2006.01155.x

    Article  CAS  Google Scholar 

  27. Lee LEJ, Clemons JH, Bechtel DG, Caldwell SJ, Han KB, Pasitschniak-Arts M, Mosser DD, Bols NC (1993) Development and characterization of a rainbow trout liver cell line expressing cytochrome P450-dependent monooxygenase activity. Cell Biol Toxicol 9(3):279–294. doi:10.1007/BF00755606

    Article  CAS  PubMed  Google Scholar 

  28. Karunasagar I, Millar SD, Frerichs GN (1995) A new cell line from Puntius schwanenfeldi sensitive to snakehead fish cell line C-type retrovirus. Asian Fish Sci 8:151–157

    Google Scholar 

  29. Bradford CS, Sun L, Collodi P, Barnes DW (1994) Cell cultures from zebrafish embryos and adult tissues. Methods Cell Sci 16:99–107. doi:10.1007/BF01404818

    Google Scholar 

  30. Imajoh M, Ikawa T, Oshima SI (2007) Characterization of a new fibroblast cell line from a tail fin of red sea bream, Pagrus major, and phylogenetic relationships of a recent RSIV isolate in Japan. Virus Res 126:45–52. doi:10.1016/j.virusres.2006.12.020

    Article  CAS  PubMed  Google Scholar 

  31. Sobhana KS, George KC, Ravi GV, Ittoop G, Paulraj R (2008) A cell culture system developed from heart tissue of the greasy grouper, Epinephelus tauvina (Forsskål 1775) by enzymatic dissociation. Indian J Fish 55(3):251–256

    Google Scholar 

  32. 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. doi:10.1007/s10695-012-9603-7

    Article  CAS  PubMed  Google Scholar 

  33. Pärt P, Bergström E (1995) Primary cultures of teleost branchial epithelial cells. In: Wood CM, Shuttleworth TJ (eds) Fish Physiology, volume 14: cellular and molecular approaches to fish ionic regulation. Academic Press, New York, pp 207–227. ISBN 0123504384

    Chapter  Google Scholar 

  34. Kumar GS, Singh ISB, Rosamma P, Raveendranath M, Shanmugam J (1998) Efficacy of fish and prawn muscle extracts as supplements to development of a primary cell culture system from larval tissue of aquarium fish Poecilia reticulata. Indian J Exp Biol 36:91–94

    Google Scholar 

  35. Kumar GS, Singh ISB, Philip R (2001) Development of a cell culture system from the ovarian tissue of African catfish (Clarias gariepinus). Aquaculture 194:51–62. doi:10.1016/S0044-8486(00)00509-3

    Article  CAS  Google Scholar 

  36. Lakra WS, Sivakumar N, Goswami M, Bhonde RR (2006) Development of two cell culture systems from Asian seabass Lates calcarifer (Bloch). Aquac Res 37(1):18–24. doi:10.1111/j.1365-2109.2005.01387

    Article  Google Scholar 

  37. Clem LW, Moewus L, Siegel MM (1961) Studies with cells from marine fish in tissue culture. Proc Soc Exp Biol Med 108:762–766

    Article  CAS  PubMed  Google Scholar 

  38. Fryer JL (1964) Methods for the in vitro cultivation of cells from the tissues of salmonid fishes. Doctoral Dissertation, Oregon State University

  39. Fryer JL, Yusha A, Pilcher KS (1965) The in vitro cultivation of tissue and cells of Pacific salmon and steelhead trout. Ann NY Acad Sci 126(1):566–586. doi:10.1111/j.1749-6632.1965.tb14303.x

    Article  CAS  PubMed  Google Scholar 

  40. Collodi P, Barnes DW (1990) Mitogenic activity from trout embryos. Proc Natl Acad Sci USA 87(9):3498–3502. doi:10.1073/pnas.87.9.3498

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Pärt P, Norrgren L, Bergström E, Sjöberg P (1993) Primary cultures of epithelial cells from rainbow trout gills. J Exp Biol 175:219–232

    Google Scholar 

  42. Avella M, Berhaut J, Payan P (1994) Primary culture of gill epithelial cells from the sea bass Dicentrarchus labrax. In Vitro Cell Dev Biol Anim 30:41–49. doi:10.1007/BF02631417

    Article  Google Scholar 

  43. Diago ML, Lòpez-Fierro MP, Razquin B, Villena A (1993) Long-term myelopoietic cultures from the renal hematopoietic tissue of the rainbow trout, Oncorhychus mykiss W. Exp Hematol 21(9):1277–1287

    CAS  PubMed  Google Scholar 

  44. Moritomo T, Watanabe T (1994) Colony growth of carp hematopoietic cells in vitro. In: Stolen JS, Fletcher TC, Rowley AF, Zeliko AF, Kaattari SL, Smith SA (eds) Techniques in fish immunology-3. SOS Publications, Fair Haven, pp 35–44

    Google Scholar 

  45. Prashanth RG, Debala Devi CH, Usha Anandhi D (2012) Growth rate studies of marine ornamental fish Pomacentrus caeruleus in artificial conditions. J Environ Biol 33:929–932

    CAS  PubMed  Google Scholar 

  46. Middlebrooks BL, Ellender RD, Wharton JH (1979) Fish cell culture: a new cell line from Cynoscion nebulosus. In Vitro 15(2):109–111

    Article  Google Scholar 

  47. Grunow B, Noglick S, Kruse C, Gebert M (2011) Isolation of cells from Atlantic sturgeon Acipenser oxyrinchus oxyrinchus and optimization of culture conditions. Aquat Biol 14:67–75. doi:10.3354/ab00383

    Article  Google Scholar 

  48. Bain PA, Hutchinson RG, Marks AB, Crane MSJ, Schuller KA (2013) Establishment of a continuous cell line from southern bluefin tuna (Thunnus maccoyii). Aquaculture 376–379:59–63. doi:10.1016/j.aquaculture.2012.11.008

    Article  Google Scholar 

  49. Tong SL, Lee 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. doi:10.1016/S0044-8486(97)00070-7

    Article  Google Scholar 

  50. Freshney RI (2010) Culture of animal cells—a manual of basic technique and specialized applications, 6th edn. Wiley-Blackwell, NJ, pp 317–334

    Book  Google Scholar 

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Acknowledgments

The authors would like to express their deepest gratitude to the Director and staff of the Central Marine Fisheries Research Institute, Kochi for the facilities provided to carry out the study. This work was supported by grants from the Department of Biotechnology, New Delhi.

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George, G.A., Sobhana, K.S., Sunny, S.M. et al. Evaluation of Various Tissues of the Caerulean Damsel, Pomacentrus caeruleus for Initiating In Vitro Cell Culture Systems. Proc. Natl. Acad. Sci., India, Sect. B Biol. Sci. 88, 293–303 (2018). https://doi.org/10.1007/s40011-016-0751-x

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