بررسی اثر انکپسولهسازی سلولهای بنیادی اسپرماتوگونی در هیدروژل آلژینات در طی انجماد-ذوب
DOI::
https://doi.org/10.22100/jkh.v11i4.1550کلمات کلیدی:
سلولهای بنیادی اسپرماتوگونی، حفاظت سرمایی، آلژیناتچکیده
مقدمه: اسپرماتوژنز پستانداران از سلولهای بنیادی اسپرماتوگونی (SSCs) منشاء میگیرد. درمانهای ضد سرطان، اعم از پرتودرمانی يا شیمیدرمانی، با هدف قرار دادن این سلولهای پرتكثير میتوانند باعث ناباروری شوند. در مردان بالغ مبتلا به سرطان، انجماد اسپرماتوزوئیدها قبل از شروع درمان راه حفظ باروری در طی روند درمان است؛ اما در پسران نابالغ به دلیل نبود اسپرماتوزوئید، انجام این کار مقدور نیست. هدف این مطالعه بررسی اثر آلژینات بر کاهش اثرات سوء انجماد-ذوب بر پتانسیل SSCs و حفظ این سلولها در بچههای سرطانی است که تحت شیمیدرمانی یا رادیوتراپی قرار میگیرند.
مواد و روشها: SSCs از بیضه موشهای 6 روزه نابالغ Balb/C جداسازی شدند. خالصسازی بهوسیله آنتیبادیهای Thy-1 و c-kit به روش MACS انجام گرفت. این سلولها در هیدروژل آلژینات انکپسوله و منجمد شدند. بعد از ذوب سلولها تعیین درصد سلولهای زنده انجام گرفت. پس از استخراج RNA و ساخت cDNA بررسی بیان ژنهای Oct4، Sall4، Plzf، Dazl، Etv5، Bcl6b، Lin28 و Nanog به روش Real time-PCR RT صورت گرفت. برای آنالیز نتایج از نرمافزار آماری SPSS و برای تجزيه و تحليل بیان ژن از نرمافزار Rest استفاده گردید.
نتایج: درصد سلولهای زنده پس از انجماد در گروه سلولهای انکپسوله در آلژینات نسبتبه گروه کنترل کاهش آماری معنیداری را نشان نمیدهد. در گروه انجماد با انکپسولهسازی در آلژینات بیان ژنهای Lin28 و Sall4 افزایشیافته (0001/0P<، 0001/0P<) و بیان ژن Dazl کاهشیافته است (0001/0P<).
نتیجهگیری: این مطالعه نشان میدهد که استفاده از آلژینات بهعنوان یک داربست در حفاظت سرمایی SSCs میتواند برای حفظ پتانسیل بنیادی این سلولها مؤثر باشد.
مراجع
Yang L, Wu W, Qi H. Gene expression profiling revealed specific spermatogonial stem cell genes in mouse. Genesis 2013;51:83-96. doi: 10.1002/dvg.22358
Tagelenbosch RA, de Rooij DG. A quantitative study of spermatogonial multiplication and stem cell renewal in the C3H/101 F 1 hybrid mouse. Mutat Res 1993;290:193-200.
Shalet SM, Tsatsoulis A, Whitehead E, Read G. Vulnerability of the human Leydig cell to radiation damage is dependent upon age. J Endocrinol 1989;120:161-5.
Ginsberg JP. New advances in fertility preservation for pediatric cancer patients. Curr Opin Pediatr 2011;23:9-13. doi: 10.1097/MOP.0b013e3283420fb6
Craft I, Bennett V, Nicholson N. Fertilising ability of testicular spermatozoa. Lancet 1993;342:864.
Hudson MM. Reproductive outcomes for survivors of childhood cancer. Obstet Gynecol 2010;116:1171-83. doi: 10.1097/AOG.0b013e3181f87c4b
Van Casteren N, van Santbrink E, Van Inzen W, Romijn JC, Dohle GR. Use rate and assisted reproduction technologies outcome of cryopreserved semen from 629 cancer patients. Fertil Steril 2008;90:2245-50. doi: 10.1016/j.fertnstert.2007.10.055
Sabanegh ES, Ragheb AM. Male fertility after cancer. Urology 2009;73:225-31. doi: 10.1016/j.urology.2008.08.474
Gouk SS, Loh YF, Kumar SD, Watson PF, Kuleshova LL. Cryopreservation of mouse testicular tissue: prospect for harvesting spermatogonial stem cells for fertility preservation. Fertil Steril 2011;95:2399-403. doi: 10.1016/j.fertnstert.2011.03.035
Loren AW, Mangu PB, Beck LN, Brennan L, Magdalinski AJ, Partridge AH, et al. Fertility preservation for patients with cancer: American Society of Clinical Oncology clinical practice guideline update. J Clin Oncol 2013;31:2500-10. doi: 10.1200/JCO.2013.49.2678
Ginsberg JP, Carlson CA, Lin K, Hobbie WL, Wigo E, Wu X, et al. An experimental protocol for fertility preservation in prepubertal boys recently diagnosed with cancer: a report of acceptability and safety. Hum Reprod 2010;25:37-41. doi: 10.1093/humrep/dep371
Lin N, Lin J, Bo L, Weidong P, Chen S, Xu R. Differentiation of bone marrow‐derived mesenchymal stem cells into hepatocyte‐like cells in an alginate scaffold. Cell Prolif 2010;43:427-34. doi: 10.1111/j.1365-2184.2010.00692.x
Pravdyuk AI, Petrenko YA, Fuller BJ, Petrenko AY. Cryopreservation of alginate encapsulated mesenchymal stromal cells. Cryobiology 2013;66:215-22. doi: 10.1016/j.cryobiol.2013.02. 002
Tomkoria S, Masuda K, Mao J. Nanomechanical properties of alginate-recovered chondrocyte matrices for cartilage regeneration. Proc Inst Mech Eng H 2007;221:467-73.
Aoki T, Koizumi T, Kobayashi Y, Yasuda D, Izumida Y, Jin Z, et al. A novel method of cryopreservation of rat and human hepatocytes by using encapsulation technique and possible use for cell transplantation. Cell Transplant 2005;14:609-20.
Milazzo J, Vaudreuil L, Cauliez B, Gruel E, Massé L, Mousset-Siméon N, et al. Comparison of conditions for cryopreservation of testicular tissue from immature mice. Hum Reprod 2008;23:17-28. doi: 10.1093/humrep/dem355
Oatley JM, Brinster RL. [11]-Spermatogonial Stem Cells. Methods in Enzymology 2006;419:259-82. doi: 10.1016/S0076-6879(06)19011-4
van Waas M, Neggers SJ, Pieters R, van den Heuvel-Eibrink MM. Components of the metabolic syndrome in 500 adult long-term survivors of childhood cancer. Ann Oncol 2010;21:1121-6. doi: 10.1093/annonc/mdp414
Green DM, Kawashima T, Stovall M, Leisenring W, Sklar CA, Mertens AC, et al. Fertility of male survivors of childhood cancer: a report from the childhood cancer survivor study. J Clin Oncol 2010;28:332-9. doi: 10.1200/JCO.2009.24.9037
Thomson AB, Campbell AJ, Irvine DS, Anderson RA, Kelnar CJ, Wallace WHB. Semen quality and spermatozoal DNA integrity in survivors of childhood cancer: a case-control study. Lancet 2002;360:361-7.
Kanatsu-Shinohara M, Ogonuki N, Inoue K, Miki H, Ogura A, Toyokuni S, et al. Long-term proliferation in culture and germline transmission of mouse male germline stem cells. Biol Reprod 2003;69:612-6. doi: 10.1095/biolreprod.103.017012
Liu S, Tang Z, Xiong T, Tang W. Isolation and characterization of human spermatogonial stem cells. Reprod Biol Endocrinol 2011;9:141. doi: 10.1186/1477-7827-9-141
Seandel M, Falciatori I, Rafii S. Pluripotent stem cells from the postnatal testis: unlocking the potential of spermatogonial stem cells. Male Germline Stem Cells: Developmental and Regenerative Potential: Springer; 2011.p.25-47.
Pesce M, Wang X, Wolgemuth DJ, Schöler HR. Differential expression of the Oct-4 transcription factor during mouse germ cell differentiation. Mechanisms of Development 1998;71:89-98. doi: 10.1016/S0925-4773(98)00002-1
Kanatsu-Shinohara M, Lee J, Inoue K, Ogonuki N, Miki H, Toyokuni S, et al. Pluripotency of a single spermatogonial stem cell in mice. Biol Reprod 2008;78:681-7. doi: 10.1095/biolreprod.107.066068
Buaas FW, Kirsh AL, Sharma M, McLean DJ, Morris JL, Griswold MD, et al. Plzf is required in adult male germ cells for stem cell self-renewal. Nat Genet 2004;36:647-52. doi: 10.1038/ng1366
Hartatik T, Okada S, Okabe S, Arima M, Hatano M, Tokuhisa T. Binding of BAZF and Bc16 to STAT6-binding DNA sequences. Biochem Biophys Res Commun 2001;284:26-32. doi: 10.1006/bbrc.2001.4931
Morrow CM, Hostetler CE, Griswold MD, Hofmann MC, Murphy KM, Cooke PS, et al. ETV5 is required for continuous spermatogenesis in adult mice and may mediate blood–testes barrier
Oatley JM, Avarbock MR, Brinster RL. Glial cell line-derived neurotrophic factor regulation of genes essential for self-renewal of mouse spermatogonial stem cells is dependent on Src family kinase signaling. J Biol Chem 2007;282:25842-51. doi: 10.1074/jbc.M703474200
Wu X, Oatley JM, Oatley MJ, Kaucher AV, Avarbock MR, Brinster RL. The POU domain transcription factor POU3F1 is an important intrinsic regulator of GDNF-induced survival and self-renewal of mouse spermatogonial stem cells. Biol Reprod 2010;82:1103-11. doi: 10.1095/biolreprod.109.083097
Lin Y, Page DC. Dazl deficiency leads to embryonic arrest of germ cell development in XY C57BL/6 mice. Dev Biol 2005;288:309-16. doi: 10.1016/j.ydbio.2005.06.032
Schrans-Stassen BH, Saunders PT, Cooke HJ, de Rooij DG. Nature of the spermatogenic arrest in Dazl−/− mice. Biol Reprod 2001;65:771-6.
Chen H-H, Welling M, Bloch DB, Muñoz J, Mientjes E, Chen X, et al. DAZL limits pluripotency, differentiation, and apoptosis in developing primordial germ cells. Stem Cell Reports 2014;3:892-904. doi: 10.1016/j.stemcr.2014.09.003
Chew JL, Loh YH, Zhang W, Chen X, Tam WL, Yeap LS, et al. Reciprocal transcriptional regulation of Pou5f1 and Sox2 via the Oct4/Sox2 complex in embryonic stem cells. Mol Cell Biol 2005;25:6031-46. doi: 10.1128/MCB.25.14.6031-6046.2005
Wang PJ, Page DC, McCarrey JR. Differential expression of sex-linked and autosomal germ-cell-specific genes during spermatogenesis in the mouse. Hum Mol Genet 2005;14:2911-8. doi: 10.1093/hmg/ddi322
Yu J, Vodyanik MA, Smuga-Otto K, Antosiewicz-Bourget J, Frane JL, Tian S, et al. Induced pluripotent stem cell lines derived from human somatic cells. Science 2007;318:1917-20. doi: 10.1126/science.1151526
Zheng K, Wu X, Kaestner KH, Wang PJ. The pluripotency factor LIN28 marks undifferentiated spermatogonia in mouse. BMC Dev Biol 2009;9:38. doi: 10.1186/1471-213X-9-38
Izadyar F, Creemers LB, van Dissel-Emiliani FM, van Pelt AM, de Rooij DG. Spermatogonial stem cell transplantation. Molecular and Cellular Endocrinology 2000;169:21-6. doi: 10.1016/S0303-7207(00)00346-4
Riboldi M, Rubio C, Pellicer A, Gil-Salom M, Simón C. In vitro production of haploid cells after coculture of CD49f+ with Sertoli cells from testicular sperm extraction in nonobstructive azoospermic patients. Fertil Steril 2012;98:580-90. doi: 10.1016/j.fertnstert.2012.05.039
He Z, Kokkinaki M, Jiang J, Dobrinski I, Dym M. Isolation, characterization, and culture of human spermatogonia. Biol Reprod 2010;82:363-72. doi: 10.1095/biolreprod.109.078550
Gang B, Yanfeng L. Isolation and purification of human spermatogenous cells. Acta Acad Med Militaris Tertiae 2005;27:1142-4.
Koruji M, Shahverdi A, Janan A, Piryaei A, Lakpour MR, Sedighi MAG. Proliferation of small number of human spermatogonial stem cells obtained from azoospermic patients. Journal of Assisted Reproduction and Genetics 2012;29:957-67. doi: 10.1007/s10815-012-9817-8
He Z, Kokkinaki M, Jiang J, Zeng W, Dobrinski I, Dym M. Isolation of human male germ-line stem cells using enzymatic digestion and magnetic-activated cell sorting. Methods Mol Biol 2012;825:45-57. doi: 10.1007/978-1-61779-436-0_4
Kanatsu-Shinohara M, Ogonuki N, Miki H, Inoue K, Morimoto H, Takashima S, et al. Genetic influences in mouse spermatogonial stem cell self-renewal. J Reprod Dev 2010;56:145-53.
Conrad S, Renninger M, Hennenlotter J, Wiesner T, Just L, Bonin M, et al. Generation of pluripotent stem cells from adult human testis. Nature 2008;456:344-9.
Shinohara T, Brinster RL. Enrichment and transplantation of spermatogonial stem cells. Int J Androl 2000;23:89-91.
Kubota H, Avarbock MR, Brinster RL. Spermatogonial stem cells share some, but not all, phenotypic and functional characteristics with other stem cells. Proc Natl Acad Sci U S A 2003;100:6487-92. doi: 10.1073/pnas.0631767100
Kubota H, Avarbock MR, Brinster RL. Culture conditions and single growth factors affect fate determination of mouse spermatogonial stem cells. Biol Reprod 2004;71:722-31. doi: 10.1095/biolreprod.104.029207
function and testicular immune privilege. Ann N Y Acad Sci 2007;1120:144-51. doi: 10.1196/annals.1411.005
Stukenborg JB, Wistuba J, Luetjens CM, Elhija MA, Huleihel M, Lunenfeld E, et al. Coculture of spermatogonia with somatic cells in a novel three‐dimensional soft‐agar‐culture‐system. J Androl 2008;29:312-29. doi: 10.2164/jandrol.107.002857
Ogawa T, Dobrinski I, Avarbock MR, Brinster RL. Xenogeneic spermatogenesis following transplantation of hamster germ cells to mouse testes. Biol Reprod 1999;60:515-21.
Dobrinski I, Ogawa T, Avarbock MR, Brinster RL. Computer assisted image analysis to assess colonization of recipient seminiferous tubules by spermatogonial stem cells from transgenic donor mice. Mol Reprod Dev 1999;53:142-8. doi: 10.1002/(SICI)1098-2795(199906)53:2<142::AID-MRD3>3.0.CO;2-O
Kanatsu-Shinohara M, Inoue K, Lee J, Yoshimoto M, Ogonuki N, Miki H, et al. Generation of pluripotent stem cells from neonatal mouse testis. Cell 2004;119:1001-12. doi: 10.1016/j.cell.2004.11.011
چاپ شده
شماره
نوع مقاله
مجوز
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.