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Purification and in vitro cultivation of archaeocytes (stem cells) of the marine sponge Hymeniacidon perleve (Demospongiae)

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Abstract

Marine sponges (Porifera) are the best source of marine bioactive metabolites for drug discovery and development, although the sustainable production of most sponge-derived metabolites remains a difficult task. In vitro cultivation of sponge cells in bioreactors has been proposed as a promising technology. However, no continuous cell line has as yet been developed. Archaeocytes are considered to be toti/multipotent stem cells in sponges and, when purified, may allow the development of continuous sponge cell lines. As a prerequisite, we have developed a novel four-step protocol for the purification of archaeocytes from a marine sponge, Hymeniacidon perleve: (1) differential centrifugation to separate large sponge cells including archaeocytes; (2) selective agglomeration in low-Ca2+/Mg2+ artificial seawater in which living archaeocytes form small loose aggregates with some pinacocytes and collencytes; (3) differential adherence to remove anchorage-dependent pinacocytes, collencytes and other mesohyl cells; (4) Ficoll-Vrografin density gradient centrifugation to purify archaeocytes. The final purity of archaeocytes is greater than 80%. The proliferation potential of the archaeocytes has been demonstrated by high levels of BrdU incorporation, PCNA expression and telomerase activity. In 4-day primary cultures, the purified archaeocytes show a 2.5-fold increase in total cell number. This study opens an important avenue towards developing sponge cell cultures for the commercial exploitation of sponge-derived drugs.

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References

  • Almendral JM, Huebsch D, Blundell PA, Macdonald-Bravo H, Bravo R (1987) Cloning and sequence of the human nuclear protein cyclin: homology with DNA-binding proteins. Proc Natl Acad Sci USA 84:1575–1579

    Article  PubMed  CAS  Google Scholar 

  • Bauer GA, Burgers PMJ (1988) The yeast analog of mammalian cyclin/proliferating-cell nuclear antigen interacts with mammalian DNA polymerase. Proc Natl Acad Sci USA 85:7506–7510

    Article  PubMed  CAS  Google Scholar 

  • Bayne CJ (1998) Invertebrate cell culture considerations: insects, ticks, shellfish, and worms. In: Mather JP, Barnes D (eds) Methods in cell biology, vol 57. Academic Press, New York, pp 187–201

    Google Scholar 

  • Bergquist PR (1978) Sponges. University of California Press, Berkeley

    Google Scholar 

  • Berry L (2003) Soaking up the limelight. Nature 421:791

    Article  CAS  Google Scholar 

  • Blunt JW, Copp BR, Munro MHG, Northcote PT, Prinsep MR (2005) Marine natural products. Nat Prod Rep 22:15–61

    Article  PubMed  CAS  Google Scholar 

  • Boury-Esnault N (1977) A cell type in sponge involved in the metabolism of glycogen: the gray cells. Cell Tissue Res 175:523–539

    Article  PubMed  CAS  Google Scholar 

  • Bravo R (1986) Synthesis of the nuclear protein cyclin (PCNA) and its relationship with DNA replication. Exp Cell Res 163:287–293

    Article  PubMed  CAS  Google Scholar 

  • Celis JE, Madsen P, Celis A, Nielsen HV, Gesser B (1987) Cyclin (PCNA, auxiliary protein of DNA polymerase δ) is a central component of the pathway(s) leading to DNA replication and cell division. FEBS Lett 220:1–7

    Article  PubMed  CAS  Google Scholar 

  • Corredor JE, Wilkinson CR, Vicente VP, Morell JM, Otero E (1988) Nitrate release by Caribbean reef sponges. Limnol Oceanogr 33:114–120

    Article  CAS  Google Scholar 

  • Custódio MR, Prokic I, Steffen R, Koziol C, Borojevic R, Brummer F, Nickel M, Müller WEG (1998) Primmorphs generated from dissociated cells of the sponge Suberites domuncula: a model system for studies of cell proliferation and cell death. Mech Ageing Dev 105:45–59

    Article  PubMed  Google Scholar 

  • Custódio MR, Hajdu E, Muricy G (2004) Cellular dynamics of in vitro allogeneic reactions of Hymeniacidon heliophila (Demospongiae: Halichondrida). Mar Biol 144:999–1010

    Article  Google Scholar 

  • Daidoji H, Takasaki Y, Nakane PK (1992) Proliferating cell nuclear antigen (PCNA/cyclin) in plant proliferating cells: immunohistochemical and quantitative analysis using autoantibody and murine monoclonal antibodies to PCNA. Cell Biochem Funct 10:123–132

    Article  PubMed  CAS  Google Scholar 

  • De Rosa S, De Caro S, Iodice C, Tommonaro G, Stefanov K, Popov S (2003) Development in primary cell culture of Demosponges. J Biotechnol 100:119–125

    Article  PubMed  Google Scholar 

  • De Sutter D, Van de Vyver G (1977) Aggregative properties of different cell types of the fresh-water sponge Ephydatia fluviatilis isolated on Ficoll gradients. Rouxs Arch 181:151–161

    Article  Google Scholar 

  • Faulkner DJ (1998) Marine natural products. Nat Prod Rep 15:113–158

    Article  PubMed  CAS  Google Scholar 

  • Flowers AE, Garson MJ, Webb RI, Dumdei EJ, Charan RD (1998) Cellular origin of chlorinated diketopiperazines in the dictyoceratid sponge Dysidea herbacea (Keller). Cell Tissue Res 292:597–607

    Article  PubMed  CAS  Google Scholar 

  • Fusetani N (2000) Drugs from the sea. Karger, Basel

    Book  Google Scholar 

  • Garson MJ, Flowers AE, Webb RI, Charan RD, McCaffrey EJ (1998) A sponge/dinoflagellate association in the haplosclerid sponge Haliclona sp.: cellular origin of cytotoxic alkaloids by Percoll density gradient fractionation. Cell Tissue Res 293:365–373

    Article  PubMed  CAS  Google Scholar 

  • Gratzner HG (1982) Monoclonal antibody to 5-bromo and 5-iododeoxyuridine: a new reagent for detection of DNA replication. Science 218:474–475

    Article  PubMed  CAS  Google Scholar 

  • Ilan M, Contini H, Carmeli S, Rinkevich B (1996) Progress towards cell cultures from a marine sponge that produces bioactive compounds. J Mar Biotechnol 4:145–149

    Google Scholar 

  • Imsiecke G, Steffen R, Custodio MR, Borojevic R, Müller WEG (1995) Formation of spicules by sclerocytes from the freshwater sponge Ephydatia muelleri in short-term cultures in vitro. In Vitro Cell Dev Biol 31:528–535

    CAS  Google Scholar 

  • Kim NW, Piatyszek MA, Prowse KR, Harley CB, West MD, Ho PL (1994) Specific association of human telomerase activity with immortal cells and cancer. Science 266:2011–2015

    Article  PubMed  CAS  Google Scholar 

  • Koziol C, Borojevic R, Steffen R, Müller WEG (1998) Sponges (Porifera) model systems to study the shift from immortal to senescent somatic cells: the telomerase activity in somatic cells. Mech Ageing Dev 100:107–120

    Article  PubMed  CAS  Google Scholar 

  • Le Pennec G, Perovic S, Ammar MSA, Grebenjuk VA, Steffen R, Brümmer F, Müller WEG (2003) Cultivation of primmorphs from the marine sponge Suberites domuncula: morphogenetic potential of silicon and iron. J Biotechnol 100:93–108

    Article  PubMed  Google Scholar 

  • Matsumoto K, Moriuchi T, Koji T, Nakane PK (1987) Molecular cloning of cDNA coding for rat proliferating cell nuclear antigen (PCNA)/cyclin. EMBO J 6:637–642

    PubMed  CAS  Google Scholar 

  • Moriuchi T, Matsumoto K, Koji T, Nakane PK (1986) Molecular cloning and nucleotide sequence analysis of rat PCNA/cyclin cDNA. Nucleic Acids Symp Ser 17:117–120

    PubMed  CAS  Google Scholar 

  • Müller WEG, Schäcke H (1996) Characterization of the receptor protein-tyrosine kinase genes from the marine sponge Geodia cydonium. Prog Mol Subcell Biol 17:183–208

    PubMed  Google Scholar 

  • Müller WEG, Wiens M, Batel R, Steffen R, Schröder HC, Borojevic R, Custódio MR (1999) Establishment of a primary cell culture from a sponge: primmorphs from Suberites domuncula. Mar Ecol Prog Ser 178:205–219

    Google Scholar 

  • Müller WEG, Böhm M, Batel R, De Rosa S, Tommonaro G, Müller IM, Schröder HC (2000) Application of cell culture for the production of bioactive compounds from sponges: synthesis of avarol by primmorphs from Dysidea avara. J Nat Prod 63:1077–1081

    Article  PubMed  CAS  Google Scholar 

  • Naganuma T, Degan BM, Horikoshi K, Morse DE (1994) Myogenesis in primary cell cultures from larvae of the abalone, Haliotis rufescens. Mol Mar Biol Biotech 3:131–140

    CAS  Google Scholar 

  • Osinga R, Tramper J, Wijffels RH (1999) Cultivation of marine sponges. Mar Biotechnol 1:509–532

    Article  PubMed  CAS  Google Scholar 

  • Paunesku T, Mittal S, Protic M, Oryhon J, Korolev SV, Joachimiak A, Woloschak GE (2001) Proliferating cell nuclear antigen (PCNA): ringmaster of the genome. Int J Radiat Biol 77:1007–1021

    Article  PubMed  CAS  Google Scholar 

  • Piatyszek MA, Kim NW, Weinrich SL, Hiyama K, Hiyama E, Wright WE, Shay JW (1995) Detection of telomerase activity in human cells and tumors by a telomeric repeat amplification protocol (TRAP). Methods Cell Sci 17:1–15

    Article  Google Scholar 

  • Pomponi SA, Willoughby R (1994) Sponge cell culture for production of bioactive metabolites. In: van Soest SR, Balkema AA (eds) Sponges in time and space. Rotterdam, Brookfield, pp 395–400

    Google Scholar 

  • Prelich G, Tan CK, Kostura M, Mathews MB, So AG, Downey KM, Stillman B (1987) Functional identify of proliferating cell nuclear antigen and a DNA polymerase-δ auxiliary protein. Nature 326:517–520

    Article  PubMed  CAS  Google Scholar 

  • Rinkevich B (1999) Cell cultures from marine invertebrates: obstacles, new approaches and recent improvements. J Biotechnol 70:133–153

    Article  CAS  Google Scholar 

  • Rinkevich B (2005) Marine invertebrate cell cultures: new millennium trends. Mar Biotechnol 7:429–439

    Article  PubMed  CAS  Google Scholar 

  • Rinkevich B, Rabinowitz C (1994) Acquiring embryo-derived cell cultures and aseptic metamorphosis of larvae from the colonial protochordate Botryllus schlosseri. Invertebr Reprod Dev 25:59–72

    Google Scholar 

  • Rosenfield A, Kern FG, Keller BJ (1994) Invertebrate neoplasia: initiation and promotion mechanisms. NOAA Technical Memorandum NMFS-NE-107:31

    Google Scholar 

  • Salomon CE, Deerinck T, Ellisman MH, Faulkner DJ (2001) The cellular localization of dercitamide in the Palauan sponge Oceanapia sagittaria. Mar Biol 139:313–319

    Article  CAS  Google Scholar 

  • Simpson TL (1984) The cell biology of sponges. Springer, Berlin Heidelberg New York

    Google Scholar 

  • Suzuka I, Daidoji H, Matsuoka M, Kadowaki K, Takasaki Y, Nakane PK, Moriuchi T (1989) Gene for proliferating-cell nuclear antigen (DNA polymerase delta auxiliary protein) is present in both mammalian and higher plant genomes. Proc Natl Acad Sci USA 86:3189–3193

    Article  PubMed  CAS  Google Scholar 

  • Uriz MJ, Turon X, Galera J, Tur JM (1996) New light on the cell location of avarol within the sponge Dysidea avara (Dendroceratida). Cell Tissue Res 285:519–527

    Article  Google Scholar 

  • Willoughby R, Pomponi SA (2000) Quantitative assessment of marine sponge cells in vitro: development of improved growth medium. In Vitro Cell Dev Biol Animal 36:194–200

    Article  CAS  Google Scholar 

  • Wilkinson CR (1987) Interocean differences in size and nutrition of coral reef sponge populations. Science 236:1654–1657

    Article  PubMed  Google Scholar 

  • Yin CQ, Humphreys T (1996) Acute cytotoxic allogeneic histoincompatibility reactions involving gray cells in the marine sponge, Callyspongeia diffusa. Biol Bull 191:159–167

    Article  CAS  Google Scholar 

  • Zhang W, Zhang XY, Cao XP, Xu JY, Zhao QY, Yu XJ, Jin MF, Deng MC (2003a) Optimizing the formation of in vitro sponge primmorphs from the Chinese sponge Stylotella agminata (Ridley). J Biotechnol 100:161–168

    Article  PubMed  CAS  Google Scholar 

  • Zhang XY, Cao XP, Zhang W, Yu XJ, Jin MF (2003b) Primmorphs from archaeocytes—dominant cell population of the sponge Hymeniacidon perleve: improved cell proliferation and spiculogenesis. Biotechnol Bioeng 84:583–590

    Article  CAS  Google Scholar 

  • Zhao QY, Jin MF, Müller WEG, Zhang W, Yu XJ, Deng MC (2003) Attachment of marine sponge cells of Hymeniacidon perleve on microcarriers. Biotechnol Prog 19:1569–1573

    Article  PubMed  CAS  Google Scholar 

  • Zhao QY, Zhang W, Jin MF, Yu XJ, Deng MC (2005) Formulation of a basal medium for primary cell culture of the marine sponge Hymeniacidon perleve. Biotechnol Prog 21:1008–1012

    Article  PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Marine Bioproducts Engineering Group,Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China

    Liming Sun, Yuefan Song, Yi Qu, Xingju Yu & Wei Zhang

  2. Graduate School, Chinese Academy of Sciences, Beijing, 100039, China

    Liming Sun, Yuefan Song & Yi Qu

  3. Department of Medical Biotechnology, School of Medicine, Flinders University, Adelaide, SA 5042, Australia

    Wei Zhang

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  1. Liming Sun
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  2. Yuefan Song
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  3. Yi Qu
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  4. Xingju Yu
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  5. Wei Zhang
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Correspondence to Wei Zhang.

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The authors are grateful for the financial support of the Chinese Academy of Sciences under the “100 Talent Project”, the “Innovation Fund” from the Dalian Institute of Chemical Physics, the “Hi-Tech Research and Development Program of China” (2001AA620404), and the European Commission (project: Silicon Biotechnology).

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Sun, L., Song, Y., Qu, Y. et al. Purification and in vitro cultivation of archaeocytes (stem cells) of the marine sponge Hymeniacidon perleve (Demospongiae). Cell Tissue Res 328, 223–237 (2007). https://doi.org/10.1007/s00441-006-0342-x

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  • DOI: https://doi.org/10.1007/s00441-006-0342-x

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