تمایز سلول های بنیادی مزانشیمال با القای ذرات نانو تیوب تیتانیوم و اشعه فرابنفش به سلول های جنسی

نوع مقاله : مقاله پژوهشی

نویسندگان

1 گروه زیست شناسی، واحد علوم تحقیقات، دانشگاه آزاد اسلامی، تهران، ایران

2 گروه آناتومی، دانشگاه علوم پزشکی البرز، کرج، ایران

چکیده

مقدمه و هدف: بسیاری از درمان های ناباروری به علت فقدان گامت های جنسی بیماران کارآمد نیستند. تمایز سلول های بنیادین به سلول های زایا ی جنسی می تواند گزینه مناسبی برای تولید گامت و کاربرد کلینیکی برای درمان ناباروری باشد. 
مواد و روش ها: سلول های بنیادین مزانشیمال به روش فلاشینگ از استخوان ران موشهای بالغ نژاد NMRI تهیه گردید. از ذرات نانو تیوب تیتانیوم titanium nanotube particles (TNT)، پرتو UV با فرکانس (254 ± 20nm)  و رتینوییک اسید برای تمایز استفاده شد. ویژگی ساختاری TNTو فیبرین با میکروسکوب FE- SEM و SEM ارزیابی شد. از مارکر های  CD90,CD31 برای تایید ماهیت سلول ها و برای بیان زایایی سلولهای تمایز یافته جنسی از مارکر MVH استفاده شد.  آزمون های  ANOVA و Mann-Whitney برای تحلیل آماری داده ها  بکارگرفته شد.
نتایج: ساختار یکنواخت TNT با ابعاد 10-15 نانومتر  با کمک  FE- SEM مشخص گردید. تصاویر SEM پوشیده شدن TNT روی فیبرین را نشان داد. در MTT بیشترین میزان بقای سلول ها در غلظت TNT 50 µg/ml دیده شد. ماهیت سلول های بنیادین با بیان مارکر های اختصاصی با روش فلوسایتومتری مشخص گردید. رنگ آمیزی های Alizarin red  رسوب کلسیوم  و Oil red با تایید قطرات چربی ماهیت چند توانی سلول های بنیادی مزانشیمی را اثبات کرد. با روش ایمنوهیستوشیمی بیان مارکر  MVH در سلولهای تمایز یافته در گروه های درمانی نسبت به کنترل تایید گردید.
نتیجه‌گیری: کاربرد TNT  و UV  روی سلول های بنیادی مزانشیمال موجب افزایش معنی دار تمایز سلول های زایای جنسی می شود.

کلیدواژه‌ها


عنوان مقاله [English]

Mesenchymal stem cells differentiation into gamete germ cells by titanium nanotube particles and UV radiation induction

نویسندگان [English]

  • Saman Ebrahimi 1
  • Alireza Shams 2
  • Parvaneh Maghami 1
  • Azadeh Hekmat 1
1 Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran
2 Department of Anatomy, Alborz University of Medical Sciences, Karaj, Iran
چکیده [English]

Background and Objective: Many of traditional infertility treatment methods due to lack of gamete are ineffective.  Differentiation of stem cells to gamete germ cells may be a good choice for producing gametes for treatment of infertility in clinique.
Materials and Methods: Bone marrow mesenchymal stem cells (BMMSc) were obtained from femoral bone of adult NMRI mice by flushing method. Nano tube particles of titanium (TNT), retinoic acid and UV beam 254 ± 20 nm frequencies were used for differentiation of MSc to gamete germ cells. Ultrastructure of TNT was studied with FEM SE electron microscope. Fibrin coating of Nano tubes was evaluated by SEM. Nature and character of stem cells were measured using cluster– differentiation (CD) by flow cytometry confirmed by CD90, and CD31 markers.  Finally, MVH marker was used for assessment of differentiation of MSc to gamete germ cells with immunohistochemistry assay. Statistical data was analyzed by ANOVA and Mann-Whitney tests.
Results: Electron microscopic pictures by FE SEM showed homogenous ultrastructure of titanium nanotubes in 10-15 nm diameters. Coating of TNT by fibrin was confirmed by SEM. MTT assay showed most survival rate of MSc cells in 50 µg/ml concentration of TNT. By CD90 and CD31 markers, nature of the stem cells was controlled by flow cytometry. Finally MVH marker expression showed differentiation of MSc to gamete germ cells in treated as compared to control groups and was confirmed by immunohistochemistry.
Conclusion: Application of TNT fibrin coated and UV beam can increase differentiation rate of BMMSc to gamete germ cells significantly as a clinical protocol.

کلیدواژه‌ها [English]

  • Mesenchymal Stem Cells
  • Differentiation
  • Titanium nanotube
  • Ultraviolet
  • gamete germ cells
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