نوع مقاله : مقاله پژوهشی
نویسندگان
1 گروه پاتوبیولوژی، دانشکده دامپزشکی، دانشگاه فردوسی مشهد، مشهد، ایران
2 گروه میکروب شناسی و ایمنی شناسی، دانشکده دامپزشکی، دانشگاه تهران، تهران ایران تلفن: 0098-21-61117053
3 انستیتوبیوتکنولوژی، دانشگاه فردوسی مشهد، مشهد، ایران
4 گروه جراحی، دانشگاه علوم پزشکی مشهد، مشهد، ایران
5 گروه بیوشیمی، دانشگاه تربیت مدرس، تهران، ایران
چکیده
زمینه سرطان کولون از سرطان های رایج در جهان می‌باشد. نقش IL-17A در سرطان از مراحل اولیه تومورزایی شروع می شود و به نظر می‌رسد با ایجاد التهاب نقش مهمی در تومورزایی دارد. هدف از مطالعه حاضر بررسی میزان بیان ژن های KI67 و P53 در سلولهای اپیتلیوم کولونHT-29 تیمار شده با IL-17A می باشد.
روش کار IL-17A خریداری شد و به میزان 50 ng/ml ، به محیط کشت HT-29، اضافه شد و پس از 24 ساعت سلول ها از محیط کشت جدا شد و نکروز سلولی با MTT بررسی گردید. سپس استخراج RNA صورت گرفت و میزان بیان P53 و KI67، با استفاده از پرایمر های جدید طراحی شده از روش Reverse transcription (RT) qPCR انجام و با نرم افزار GeniX6، مورد آنالیز قرار گرفت.
یافته ها آزمایش MTT نشان داد که غلظت 150 ng/mlبیشترین کشندگی طی 24 ساعت را دارد. پس از 24 ساعت انکوباسیون با IL-17A ، میزان بیان ژنهای KI67 (P=0.003) و P53 (P=0.001) در سلولهای HT-29در گروه تیمار شده با IL-17A بترتیب، کاهش و افزایش یافت. همچنین تعداد سلول های مورد بررسی (HT-29)در گروه تحت تیمار نسبت به گروه کنترل کاهش معنی داری را نشان داد(P < 0.05).
نتیجه گیری نتایج حاصل نشان داد IL17a می تواند با افزایش بیان P53 و کاهش KI67 موجب جلوگیری از تکثیر سلول های HT-29 در شرایط آزمایشگاهی شود. این می‌تواند زمینه برای استفاده از IL17a به عنوان عاملی برای ایمونوتراپی و پاتوژنز سرطان روده بزرگ باشد.
کلیدواژه‌ها
موضوعات
عنوان مقاله [English]
The study of expression of KI67 and P53 in co-culturing of colon epithelial cell line HT-29 with IL-17A
نویسندگان [English]
- Ali Ghorbani Ranjbary 1
- Jalil Mehrzad 2
- Hesam Dehghani 3
- Abbas Abdollahi 4
- Saman Hosseinkhani 5
1 Department of Pathobiology, Section Biotechnology, Faculty of Veterinary Medicine, and Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran.
2 Department of Microbiology and Immunology, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran Tel: +98-21-61117053; Fax: +98-21-66933222
3 3. Stem Cells and Regenerative Medicine Research Group, Research Institute of Biotechnology, Ferdowsi University of Mashhad, Mashhad, Iran.
4 Surgical Oncology Research Center, Mashhad University of Medical Sciences, Mashhad, Iran.
5 Department of Biochemistry, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
چکیده [English]
Background Colorectal cancer (CRC) is one of the most common cancers in the world. The role of IL-17A in cancer begins in the early stages of tumorigenesis and appears to play an important role in tumorigenesis by causing inflammation. The present study was conducted to investigate the expression of KI67 and P53 genes in HT-29 colon epithelial cells with IL-17A.
Materials and Methods IL-17A was purchased and 50 ng/ml was added to HT-29 culture medium and after 24 hours the cells were isolated from the culture medium and cell necrosis was examined by MTT. Then RNA was extracted and the expression levels of P53 and KI67 were analyzed using newly designed primers by Reverse transcription (RT) qPCR method and GeniX6 software.
Results The MTT test showed that a concentration of 150 ng/ml for 24 hours had maximal necrosis rate in HT-29. After 24 hours of IL-17A incubation, the expression of KI67 (P = 0.003) and P53 (P = 0.001) genes in HT-29 cells in the IL-17A exposed group decreased and increased, respectively. Also, compared to the control group the number of examined HT-29 cells in the IL-17A treated group showed a significant decrease (P
کلیدواژه‌ها [English]
- KI67
- P53
- HT-29
- IL17a
- qPCR
- cell culture
- Squamous cell carcinoma
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