کشت سلولی اولیه از بافت آبشش و کلیه ماهی آزاد دریای خزر (Salmo caspius)

نوع مقاله : مقاله کامل علمی - پژوهشی

نویسندگان

1 نویسنده مسئول، دکتری شیلات، دانشکده علوم دریایی، دانشگاه تربیت مدرس، نور، ایران.

2 استاد گروه شیلات، دانشکده علوم دریایی، دانشگاه تربیت مدرس، نور، ایران.

3 استاد گروه میکروبیولوژی، دانشکده علوم پزشکی، دانشگاه تربیت مدرس، تهران، ایران.

چکیده

رده های سلولی ابزار مهم زیستی برای انجام تحقیقات در زمینه فیزیولوژی، ویروس شناسی، داروشناسی، سم شناسی و سرطان است که می‌تواند به عنوان یک مدل جایگزین زیست شناختی حیوان زنده مورد استفاده قرار بگیرد. تولید کشت های سلولی از ماهیان اندمیک و در معرض خطر انقراض مانند ماهی آزاد دریای خزر (Salmo caspius, Kessler, 1877) و ایجاد مدل‌های in vitro از آن، می‌تواند کمک شایانی به مطالعه ابعاد مختلف زیست شناختی آزاد ماهیان بنماید. تعداد 30 عدد ماهی آزاد دریای خزر تهیه شد وکشت سلول اولیه ازبافت کلیه و آبشش از طریق روش کشت ریز بافت صورت گرفته است. بافت‌های مورد نظر در محیط Leibovitz -15 همراه با سرم جنین گاو 20%، 10% و 5% در دمای 15، 18 و 21 درجه سانتی‌گراد کاشته و نگهداری شدند. نتایج نشان داد سلول‌های اولیه تولید شده از ماهی آزاد دریای خزر، دیر رشد هستند و زمان مهاجرت سلول از بافت طولانی است، اما این سلول-ها قابلیت پاساژ‌پذیری نسبتا خوبی دارند و در محیط کشت L-15 با سرم جنین گاوی 10% به خوبی تا پاساژ 5 رشد می‌کنند. در هر دو کشت جمعیت ناهمگن متشکل از سلول‌های فیبروبلاستی- شکل و اپیتلیالی شکل دیده شدند. بررسی روند رشد سلول‌ها در دمای مختلف نشان داد، دمای بهینه رشد برای کشت سلول‌های این گونه 18 درجه سانتی‌گراد است.

کلیدواژه‌ها

موضوعات


عنوان مقاله [English]

Primary cell culture from gill and kidney of Caspian Sea salmon (Salmo caspius, Kessler, 1877)

نویسندگان [English]

  • Zohre Ghodsi 1
  • Mohammad Reza Kalbasi 2
  • Ashraf Mohebati mobarez 3
1 Corresponding Author, Ph.D. in Fisheries, Faculty of Marine Sciences, Tarbiat Modares University, Noor, Iran.
2 Professor, Dept. of Fisheries, Faculty of Marine Sciences, Tarbiat Modares University, Noor, Iran.
3 Professor, Dept. of Microbiology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
چکیده [English]

Cell lines provide an important biological tool for carrying out investigations into physiology, virology, pharmacology, toxicology, cancer which can be used as a biological substitute for living animals. Cell phenomena are studied in controlled conditions in cell culture. Establishment of cell cultures from endemic fish and endangered Such as the Caspian Sea salmon (Salmo caspius, Kessler, 1877). Production in vitro models of it can be of assistance to biological studies, immunology, toxicology, physiology. The purpose of this study is to produce primary cell cultures from the kidney and gill of Caspian salmon in order to create cell lines from these tissues in the next stages. In this study, 30 Caspian Sea salmon (Salmo caspius, Kessler, 1877) prepared and primary cell was produced from kidney and gill tissues through the explant culture method. Kidney and gill tissue pieces were cultured in 6-well plates with L-15 culture medium and 5%, 10% and 20% fetal bovine serum (FBS) at 15, 18° and 21C. The results showed that primary cells produced from Caspian Sea salmon are slow to grow and take a long time for cell migration from the tissue and doubling time. But they have a relatively good subculture and grow well up to passage 5 in L-15 culture medium with 10% fetal bovine serum. The morphological study of the produced cells showed that they are a mixture of fibroblast-like and epithelial-like cells. Investigating the process of cell growth at different temperatures showed that the optimal growth temperature for the cultivation of cells of this species is 18 C.

کلیدواژه‌ها [English]

  • primary cell culture
  • kidney
  • gill
  • Caspian Sea salmon
1.Bols, N. C., Dayeh, V. R., Lee, L. E. J., & Schirmer, K. (2005). Use of fish cell lines in the toxicology and ecotoxicology of fish. Piscine cell lines in environmental toxicology. In: Mommsen, P., Moon, T.W. (Eds.), Biochemistry and Molecular Biology of Fishes. 17, 43-84.2.Wolf, K., & Ahne, W. (1982). Fish Cell culture. In Advances in cell culture Vol. 2 (Ed. Maramorosch K), New York Academic Press, 305-328.3.Goswami, M., Yashwanth, B. S., Trudeau V., & Lakra, W. S. (2022). Role and relevance of fish cell lines in advanced in vitro research. Molecular Biology Reports. 49, 2393-2411.4.Bols, N. C., Barlian, A., Chirino-trejo, M., Caldwell, S. J., & Goegan, P. (1994). Development of a cell line from primary cultures of rainbow trout, Oncorhynchus mykiss (Walbaum), gills Journal of Fish Diseases. 17, 601-611.5.Lakra, W. S., Swaminathan, T. R., & Joy, K. P. (2011). Development, characterization, conservation and storage of fish cell lines: A review. Fish Physiology Biochemical. 37 (1), 1-20.6.Villena, A. J. (2003). Applications and needs of fish and shellfish cell culture for disease control in aquaculture Reviews. Fish Biology and Fisheries. 13, 111-140.7.Lee, J., Park, C., & Park, S. C. (2009). Use of folding modulators to improve heterologous protein production in Escherichia coli Pept. Science. 16, 103-109.8.LaPatra, S. E. (1996). The use of serological techniques for virus surveillance and certification of fish. Annual review Fish Disease. 6, 15-28.9.Menanteau-Ledouble, S., Nöbauer, K., Razzazi-Fazeli, E., & ElMatbouli, M. (2020). Effects of Yersinia ruckeri invasion on the proteome of the Chinook salmon cell line CHSE-214. Sci. Rep.10 (1), 1-9.10.Lakra, W. S., Swaminathan, T. R., & Joy, K. P. (2011). Development, characterization, conservation and storage of fish cell lines: A review. Fish Physiology Biochemical. 37 (1), 1-20.11.Fryer, J., & Lannan, C. (1994). Three decades of fish cell culture: a current listing of cell lines derived from fishes. Methods in Cell Science, 16 (2), 87-94.12.Kazanchev, A. N. (1981). Fishes of Caspian Sea and its watershed area, Iranian Fisheries Organization, 171 p.13.Dorafshan, S., Kalbasi, M. R., Pourkazemi, M., Mojazi, Amiri B., & Soltan Karimi, S. (2008). Effects of triploidy on the caspian salmon (Salmo trutta caspius) haematology, Fish Physiology and Biochemistery,
34, 195-200.14.Kiabi, B. H., Abdoli, A., & Naderi, M. (1999). Status of the fish fauna in the south Caspian Basin of Iran, Zoology in the Middle East, 18, 57-65.15.Jalali, M. A., & Mojazi Amiri, B. (2009). Threatened fishes of the world: Salmo trutta caspius (Kessler, 1877) (Salmoniforms: Salmonidae). Environmental Biology of Fishes. 86 (3), 375-376.16.Nowrozi, K., Kolbasi, M., Farzaneh, P., Shahzad Fazelia, A., Farghdan, M., Nasimian, A., Ashouri, S., Mohammadi, S., Muradmand, Z., Farhang-Nia, M., (2013). Production and evaluation of epithelial cell line from Caspian Sea salmon fin tissue. (Salmo caspius). Journal of Aquatic Physiology and Biotechnology. 2 (3), 69-85. [In Persian]17.Ghodsi, Z., Kolbasi, M., Mohabati Mobarez, A., & Farzane, P. (2018). Antibacterial effects of EC-hepcidin1 polypeptide in inhibiting Streptococcus iniae bacteria in primary cell cultures of rainbow trout Oncorhynchus mykiss. Aquatic Physiology and Biotechnology. 7 (4), 8. [In Persian]18.Ghodsi, Z., Kalbasi, M., Mohabati Mobarez, A., Farzane, P., Beemelmannsd, C., & Amiri Moghaddam, J. (2020). Immunomodulatory function of antimicrobial peptide EC-Hepcidin1 modulates the induction of inflammatory gene expression in primary cells of Caspian Trout (Salmo trutta caspius Kessler, 1877). Fish and Shellfish Immunology. 104, 55-61.19.Wolf, K., & Quimby, M. C. (1976). Primary monolayer culture of fish cells initiated from minced tissues. Tissue Culture Association manual. 2 (4), 445-448.20.McAteer, J. A., & Davis, J. M. (2002). Basic cell culture technique and the maintenance of cell lines. In: Basic Cell Culture. Davis, J. M. (Ed.). (2nd Ed.) The Bath Press, Avon, USA. 135-190.21.Hameed, A. S. et al. (2006). Establishment and characterization of India’s first marine fish cell line(SISK) from the kidney of seabass (Lates calcarifer). Aquaculture, 257 (1-4), 92-103.22.Gjessing, M. C., Aamelfot, M., Batts, W. N., Benestad, S. L., Dale, O. B., & Thoen, E. (2018). Development and characterization of two cell lines from gills of Atlantic salmon. PLoS ONE 13(2), e0191792.23.Hoover, R. L. (1978). Modulations of the cell surface and the effects on cellular interactions. In Cell–Cell Recognition (Curtis, A. S. G., ed.),pp. 221-240. Symposia for the Society for Experimental Biology XXXII. Cambridge: Cambridge University Press.24.Pisam, M., & Repoch, P. (1976). Redistribution of surface macromolecules in dissociated epithelial cells. Journal of Cell Biology. 71, 907-920.25.Wen, C. M. (2016). Development and characterization of a cell line from tilapia head kidney with melanomacrophage characteristics. Fish & Shellfish Immunology. 49, 442-449.26.Grunow, B., Noglick, S., Kruse, M., & Gebert, M. (2011). Isolation of cells from Atlantic sturgeon Acipense oxyrinchus and optimization of culture conditions. Aquatic Biology, 14, 67-75.27.Rathore, G., T Sood, N., & Swaminathan, R. (2001). Primary cell culture from fish gillsand kidney using fish serum. Indian Journal of Experimental Biology, 39, 936-938.28.Kamalendra, J., Kapoor, S., Sharma, M., Goswami, G., & Lakra, W. S. (2011). Development of primary culture from gills of Tortor (Hamilton-buchanan), Indian Journal Animal Science. 81, 1262-1265.29.Parameswaran, V., Shukla, R., Bhonde, R. R., & Hameed, A. S. S. (2006). Development of a pluripotent ES-like cell Line from Asian sea bass (Lates calcarifer) - an oviparous stem cell line mimicking viviparous ES cells. Marine Biotechnology. 9, 766-75.30.Sohana, K. S., George, K. C., Venkat raviE, G., Ittoop, G., & Paulraj, R. (2009). Development of a Cell Culture System from Gill Explants of the Grouper, Epinephelus malabaricus (Bloch and Shneider) Asian Fisheries Science, 22, 1-6.31.Pasquariello, R., Verdile, N., Pavlovic, R., Panseri, S., Schirmer, K., Brevini,
T. A. L., & Gandolfi, F. (2021). New Stable Cell Lines Derived from the Proximal and Distal Intestine of Rainbow Trout (Oncorhynchus mykiss) Retain Several Properties. Vivo. Cells. 10, 1555.32.Lee, L. E. J., Clemons, J. H., Bechtel, D. G., & Caldwell, S. J. H. (1993). Development and characterization of a rainbow trout liver cell line expressing cytochrome P450-dependent monooxygenase activity. Cell Biol. Toxicol. 9, 279-294.33.Yue, Y., Behra, R., Sigg, L., & Schirmer, K. (2016). Silver nanoparticles inhibit fish gill cell proliferation in protein-free culture medium. Nanotoxicology, 10, 1075-1083.34.O’Neill-Mehlenbacher, A., Kilemade, M., Elliott, A. J., Mothersill, C., & Seymour, C. (2007). Comparison of direct and bystander effects induced by ionizing radiation in eight fish cell lines. International Journal of Radiation Biology, 83 (9), 593-602.35.Gstraunthaler, G., Lindl, T., & van der Valk, J. (2013). A plea to reduce or replace fetal bovine serum in cell culture media. Cytotechnology. 65, 791-3.36.Fang, CH. Y., Wu, CH. CH., Fang, CH. L., Chen, W., & Chen, CH. (2017). Long-term growth comparison studies of FBS and FBS alternatives in six head and neck cell lines. PLOS ONE. 1-27.37.Zhou, G. Z., Gui, L., Li, Z. Q., Yuan,X. P., & Zhang, Q. Y. (2008). Establishment of a Chinese sturgeon Acipenser sinensis tail-fin cell line and its susceptibility to frog iridovirus. Journal Fish Biology. 73, 2058-2067.38.Sood, N., Chaudhary, D. K., Pradhan,P. K., Verma, D. K., Swaminathan, T. R., Kushwaha, B., Punia, P., &Jena, J. K. (2015). Establishment and characterization of a continuous cell line from thymus of striped snakehead, Channa striatus (Bloch 1793). In Vitro Cellular & Developmental Biology – Animal. 51 (8), 787-796.39.Swaminathan, T. R., Raj Kumar, P. M. E., Jency, R., Charan, M. U., Syamkrishnan, V. S., Basheer, N., & Sood, J. K. (2016). A new fish cell line derived from the caudal fin of freshwater angelfish Pterophyllum scalare: development and characterization. Journal of Fish Biology. 142, 81-88.40.Swaminathan, R., Thangaraja, B., Ravia, Ch., Kumara, R., Dharmaratnama, A., Saidmuhammeda, V. B., Pradhanb, P. K., & Soodb, N. (2018). Derivation of two tilapia (Oreochromis niloticus) cell lines for efficient propagation of Tilapia Lake Virus (TiLV). Aquaculture. 492, 206-214.41.Soni, P., Pradhan, P. K., Swaminathan, T. R., & Sood, N. (2018). Development, characterization and application of a new epithelial cell line from caudalfin of Pangasianodon hypophthalmus (Sauvage 1878). Acta Tropica. 182, 215-222.42.Zeng, W., Dong, H., Chen, X., Bergmann, S., Yang, Y., Wei, X., & Tong, G. (2022). Establishment and characterization of a permanent heart cell line from largemouth bass Micropterus salmoides and its application to fish virology and immunology. Aquaculture. 547, 737-427.43.Chen, S. L., Ren, G. C., Sha, Z. X., & Shi, C. Y. (2004). Establishment of a continuous embryonic cell line from Japanese flounder Paralichthys olivaceus for virus isolation. Diseases of aquatic organism. 60, 241-246.44.Freshney, R. I. (2000). Culture of Animal Cells, A Manual of Basic Technique, 4th edition, Wiley-Liss, John Wiley and Sons, Inc. Publ. New York, 577 p.45.Chen, S. L., & Qin, Q. W. (2011). Theory and Technology of Fishes Cell Culture, Beijing Science Press, 289 p.46.Ott, T. (2004). Tissue culture of fish cell lines. National Wildlife Fish Health Survey (NWFHS) laboratory procedures manual, Vol 2. In: US Fish & Wildlife Service (Eds.). Handbook of aquatic animal health procedures and protocols, 2nd Edition, Washington DC. 1-16.47.Sayadburani, M., Valipour, A., & Ghasemi, M. (2017). Cultivation of Caspian Sea salmon (Salmo caspius) using Caspian Sea salt water from the fingerling stage to the pre-breeding stage. Journal of Advanced Aquaculture Sciences, 1 (2), 1-14.48.Sayad Borani, M., Maqsoodiyeh, H., Sayad Borani, M., Zahtakash Komleh, A., & Walipour, A. (2011). Investigating the possibility of raising Caspian Sea salmon (Salmo trutta caspius) in different densities using Caspian Sea water Aquaculture Development Magazine. 6 (2), 47-61. 49.Fernandez, R. D., Yoshimizu, M., Ezura, Y., & Kimura, T. (1993). Comparative growth response of fish cell lines in different media, temperature and sodium chloride concentrations. Fish Pathology, 28, 27-34.50.Wolf, K., & Mann, J. A. (1980). Poikilotherm vertebrate cell lines and viruses: a current listing for fishes. In Vitro, 16 (2), 168-179.