Effect of Extracellular Matrix on Differentiation of Mouse Embryonic Stem Cells into Insulin-Producing Cells

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Effect of Extracellular Matrix on Differentiation of

Mouse Embryonic Stem Cells into Insulin-Producing Cells

A. Farrokhi, M.Sc.1,2_{, E. Elahi, Ph.D.}2_{, M.Pirouz, M.Sc.}1_{, H. Jafary, M.Sc.}1_{, H. Baharvand, Ph.D.}1, 3_*

1. Stem Cells Department, Cell Science Research Center, Royan Institute, , ACECR, Tehran, Iran 2. School of Biology, Tehran University, Tehran, I.R. Iran

3. Developmental Biology Department, University of Sciences and Culture, Tehran, Iran

* Corresponding Address: P.O.Box: 19395-4644, Stem Cells Department, Cell Sciences Researh Center, Royan Institute, ACECR, Tehran, Iran

Email: Baharvand@royaninstitute.org

Abstract

Received: 14/Nov/2007, Accepted: 25/May/2008

Objective: Evaluation of extracellular matrics (ECMs) effect on differentiation of embryonic stem cells (ESCs) to pancreatic β-cell.

Materials and Methods: Mouse ESC line, Royan B1, was subjected to differ-entiation into β-like cells in a three-step method: generation of embryoid bodies (EBs), spontaneous differentiation and induction by Nicotinamide onto different matrices including poly L-ornithine/laminin, gelatin, and two different dilution of matrigel (1:30, 1:100) and control group (no ECM). At the final step, differentiated cells were analyzed for expression of some pancreas-specific genes using "semi-quantitative RT-PCR ", for detection of insulin and C-peptide presence in cells us-ing "immunocytochemistry" and for the evaluation of the amount of secretd insulin in response to glucose Using "insulin secretion assay".

Results: The semi-quantitative RT-PCR analysis of differentiated cells on 1:30 matrigel coated-plates showed consistent higher expression of β-cell specific markers including Insulin I, Insulin II, Slc2a2 in comparison to the other ECMs. The results of immunostainig for C-peptide showed no significant differences be-tween the experimental groups and finally insulin secretion assay revealed that differentiated cells on 1:30 matrigel coated-plates secreted more insulin in re-sponse to glucose in comparison to the other ECMs.

Conclusion: Our results suggest that type of ECM may influence ESC differentia-tion into insulin-secreting cells and 1:30 matrigel was more effective. However, the success rate of differentiation needs further investigations using other appropri-ate ECMs.

Keywords: Embryonic Stem Cell, Differentiation, β-Cell, Extracellular Matrix Yakhteh Medical Journal, Vol 10, No 4, Winter 2009, Pages: 250-259

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ﻯﺎﻫ

ﻝﻮﻠﺳ

ﺯﺍ

ﻦﻴﻟﻮﺴﻧﺍ

ﺪﻟﻮﻣ

ﻯﺎﻫ

ﻝﻮﻠﺳ

ﺰﻳﺎﻤﺗ

ﺮﺑ

ﻰﻟﻮﻠﺳ

ﻥﻭﺮﺑ

ﻪﻨﻴﻣﺯ

ﻩﺩﺎﻣ

ﺮﻴﺛﺎﺗ

ﺵﻮﻣ

ﻰﻨﻴﻨﺟ

ﻯﺩﺎﻴﻨﺑ

*3 ،1_Ph.D.ﺪﻧﻭﺭﺎﻬﺑﻦﻴﺴﺣ،1_M.Sc.ﻯﺮﻔﻌﺟﻪﻴﻧﺎﻫ،1_M.Sc.ﺯﻭﺮﻴﭘﻯﺪﻬﻣ،2_Ph.D.ﻰﻬﻟﺍﻪﻬﻟﺍ،2 ،1_M.Sc.ﻰﺧﺮﻓﻰﻠﻋ ﻥﺍﺮﻳﺍ،ﻥﺍﺮﻬﺗ،ﻯﺩﺎﻴﻨﺑﻯﺎﻫﺩﻝﻮﻠﺳﻩﻭﺮﮔ،ﻰﻫﺎﮕﺸﻧﺍﺩﺩﺎﻬﺟﻰﻟﻮﻠﺳﻡﻮﻠﻋﺕﺎﻘﻴﻘﺤﺗﺰﻛﺮﻣ،ﻥﺎﻳﻭﺭﻩﺪﻜﺸﻫﻭﮋﭘ .1

ﻥﺍﺮﻳﺍ،ﻥﺍﺮﻬﺗ،ﻰﺳﺎﻨﺷﺖﺴﻳﺯﻩﺪﻜﺸﻧﺍﺩ،ﻥﺍﺮﻬﺗﻩﺎﮕﺸﻧﺍﺩ .2 ﻥﺍﺮﻳﺍ،ﻥﺍﺮﻬﺗ،ﻰﻨﻳﻮﻜﺗﻰﺳﺎﻨﺷﺖﺴﻳﺯﻩﻭﺮﮔ،ﮓﻨﻫﺮﻓﻭﻢﻠﻋﻩﺎﮕﺸﻧﺍﺩ .3

،ﻰﻫﺎﮕﺸﻧﺍﺩﺩﺎﻬﺟﻰﻟﻮﻠﺳﻡﻮﻠﻋﺕﺎﻘﻴﻘﺤﺗﺰﻛﺮﻣ،ﻥﺎﻳﻭﺭﻩﺪﻜﺸﻫﻭﮋﭘ،19395-4644 :ﻰﺘﺴﭘﻕﻭﺪﻨﺻ،ﻥﺍﺮﻬﺗ،ﻥﺍﺮﻳﺍ :ﻝﻮﺌﺴﻣﻩﺪﻨﺴﻳﻮﻧﺱﺭﺩﺁ * ﻯﺩﺎﻴﻨﺑﻯﺎﻫﺩﻝﻮﻠﺳﻩﻭﺮﮔ

Email: Baharvand@royaninstitute.org:ﻚﻴﻧﻭﺮﺘﻜﻟﺍﺖﺴﭘ

ﻩﺪﻴﻜﭼ

87/3/5 :_{ﻪﻟﺎﻘﻣ}_{ﺵﺮﻳﺬﭘ}_،86/8/23 :_{ﻪﻟﺎﻘﻣ}_{ﺖﻓﺎﻳﺭﺩ}

ﻩﺩﺯﺎﺑﺶﻳﺍﺰﻓﺍﺮﺑ (ﻝﮋﻳﺮﺗﺎﻣﻭﻦﻴﺗﻻژ،ﻦﻴﻨﻴﻣﻻ/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍ-ﻲﻠﭘ) ﻒﻠﺘﺨﻣﻲﻟﻮﻠﺳﻥﻭﺮﺑﻪﻨﻴﻣﺯﻯﺎﻫﻩﺩﺎﻣﺮﻴﺛﺎﺗﻲﺳﺭﺮﺑ_:ﻑﺪﻫ_*

ﺵﻮﻣﻲﻨﻴﻨﺟﻱﺎﻫﻪﺘﺧﺎﻳﻦﺑﺯﺍﻦﻴﻟﻮﺴﻧﺍﺪﻟﻮﻣﻱﺎﻫﻝﻮﻠﺳﺰﻳﺎﻤﺗ

.ﺪﺷﻡﺎﺠﻧﺍRoyan B1ﻰﻟﻮﻠﺳﻩﺩﺭﺵﻮﻣﻰﻨﻴﻨﺟﻯﺩﺎﻴﻨﺑﻯﺎﻫﻝﻮﻠﺳﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑﺰﻳﺎﻤﺗﻪﻌﻟﺎﻄﻣﻦﻳﺍﺭﺩ_:ﺎﻫﺵﻭﺭﻭﺩﺍﻮﻣ_*

ﻥﻭﺮﺑﻪﻨﻴﻣﺯ ﻯﺎﻫﻩﺩﺎﻣﻯﻭﺭﻪﺑ،ﻯﺩﻮﺧﻪﺑﺩﻮﺧﺰﻳﺎﻤﺗﺲﭙﺳﻭﻰﻨﻴﻨﺟﻪﺒﺷﻡﺎﺴﺟﺍﻪﺑﻥﺪﺷﻞﻳﺪﺒﺗﺯﺍﺲﭘﻰﻨﻴﻨﺟﻯﺩﺎﻴﻨﺑﻯﺎﻫﻝﻮﻠﺳ

ﺭﺍﺩﺖﻬﺟﺰﻳﺎﻤﺗﻯﺎﻘﻟﺍﺖﺤﺗﻭﺪﻧﺪﺷﻞﻘﺘﻨﻣ 1/100 ﻭ 1/30 ﺖﻈﻠﻏﻭﺩﺎﺑﻝﮋﻳﺮﺗﺎﻣﻭﻦﻴﻨﻴﻣﻻ/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍ-ﻲﻠﭘ،ﻦﻴﺗﻻژﻲﻟﻮﻠﺳ

ﻭ (Ins) ﻦﻴﻟﻮﺴﻧﺍﻪﻠﻤﺟﺯﺍﻰﺳﺍﺮﻜﻧﺎﭘﻯﺎﻫﻝﻮﻠﺳﻰﺻﺎﺼﺘﺧﺍﻯﺎﻫﻥژﺯﺍﻯﺩﺍﺪﻌﺗﻥﺎﻴﺑ .ﺪﻨﺘﻓﺮﮔﺭﺍﺮﻗﻦﻴﻟﻮﺴﻧﺍﺪﻟﻮﻣﻯﺎﻫﻝﻮﻠﺳﻪﺑ

ﻦﻴﻟﻮﺴﻧﺍﺪﺿﻯﺎﻫﻯﺩﺎﺑﻰﺘﻧﺁﺎﺑﻪﺘﻓﺎﻳﺰﻳﺎﻤﺗﻯﺎﻫﻝﻮﻠﺳ .ﺪﺷﻰﺳﺭﺮﺑRT-PCRﺵﻭﺭﺎﺑﻪﺘﻓﺎﻳﺰﻳﺎﻤﺗﻯﺎﻫﻝﻮﻠﺳﺭﺩﺯﺍﺮﻔﺴﻧﺍﺮﺗﺰﻛﻮﻠﮔ

ﻦﻴﻨﭽﻤﻫ .ﺪﺷﻪﺒﺳﺎﺤﻣﺰﻴﻧﺖﺒﺜﻣ_C-ﺪﻴﺘﭙﭘﻭﺖﺒﺜﻣﻦﻴﻟﻮﺴﻧﺍﻯﺎﻫﻝﻮﻠﺳﺪﺻﺭﺩﻭﻯﺰﻴﻣﺁﮓﻧﺭﻰﻤﻴﺷﻮﺘﻴﺳﻮﻨﻤﻳﺍﺵﻭﺭﻪﺑ_C-ﺪﻴﺘﭙﭘﻭ

.ﺪﻨﺘﻓﺮﮔﺭﺍﺮﻗﻰﺑﺎﻳﺯﺭﺍﺩﺭﻮﻣﺰﻛﻮﻠﮔﺮﺘﻣﻭﺮﻜﻴﻣ 22 ﻭﺮﺘﻣﻭﺮﻜﻴﻣ 3 ﻯﺎﻫﺖﻈﻠﻏﻪﺑﺦﺳﺎﭘﺭﺩﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗﻥﺍﺰﻴﻣﺮﻈﻧﺯﺍﺎﻫﻝﻮﻠﺳ

ﻥﺍﺰﻴﻣﻦﻳﺮﺘﺸﻴﺑﺎﻫﻩﻭﺮﮔ ﺮﻳﺎﺳﺎﺑﻪﺴﻳﺎﻘﻣﺭﺩ،ﺪﻧﺩﻮﺑ ﻪﺘﻓﺎﻳﺰﻳﺎﻤﺗ 1/30 ﺖﻗﺭﺎﺑﻝﮋﻳﺮﺗﺎﻣﻯﻭﺭﻪﻛ ﻰﻳﺎﻫﻝﻮﻠﺳ_:ﺎﻫﻪﺘﻓﺎﻳ_*

ﻩﺮﻴﺠﻧﺯﺶﻨﻛﺍﻭﺱﻮﻜﻌﻣﻱﺭﺍﺩﺮﺑﻪﺨﺴﻧﻰﻤﻛﻪﻤﻴﻧﻰﺳﺭﺮﺑﺞﻳﺎﺘﻧﻦﻴﻨﭽﻤﻫﺪﻧﺩﻮﺑﺍﺭﺍﺩﺰﻛﻮﻠﮔﻪﺑﺦﺳﺎﭘﺭﺩﺍﺭﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗ

ﺞﻳﺎﺘﻧ .ﺩﺍﺩﻰﻣﻥﺎﺸﻧﺍﺭﻩﻭﺮﮔﻦﻳﺍﺭﺩIns1, Ins2, Slc2a2،ﺎﺘﺑﻯﺎﻫﻝﻮﻠﺳﻰﺻﺎﺼﺘﺧﺍﻯﺎﻫﻥژﻥﺎﻴﺑﺶﻳﺍﺰﻓﺍﺯﺍﺮﻣﻲﻠﭘﻱﺍ

ﻯﺍﺮﺑ ﻪﺘﻓﺮﮔ ﮓﻧﺭ ﻯﺎﻫﻝﻮﻠﺳ ﺩﺍﺪﻌﺗ ﺮﻈﻧ ﺯﺍ ﻯﺭﺍﺩﻰﻨﻌﻣ ﺕﻭﺎﻔﺗ C- ﺪﻴﺘﭙﭘﺪﺿ ﻯﺩﺎﺑﻰﺘﻧﺁ ﺎﺑ ﻰﻤﻴﺷﻮﺘﻴﺳﻮﻨﻤﻳﺍ ﻯﺰﻴﻣﺁﮓﻧﺭ

.ﺩﺍﺪﻧﻥﺎﺸﻧﻝﮋﻳﺮﺗﺎﻣﻭﻦﻴﺗﻻژ،ﻦﻴﻨﻴﻣﻻ/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍ-ﻲﻠﭘﻩﻭﺮﮔﻪﺳﺮﻫﻯﻭﺭﻪﺘﻓﺎﻳﺰﻳﺎﻤﺗﻯﺎﻫﻝﻮﻠﺳ

ﻥﺎﻴﺑﻝﮋﻳﺮﺗﺎﻣﻰﻟﻮﻠﺳﻥﻭﺮﺑﻪﻨﻴﻣﺯﻩﺩﺎﻣﻯﻭﺭﻦﻴﻟﻮﺴﻧﺍﺪﻟﻮﻣﻱﺎﻫﻝﻮﻠﺳﻪﺑﺵﻮﻣﻲﻨﻴﻨﺟﻱﺎﻫﻪﺘﺧﺎﻳﻦﺑﺰﻳﺎﻤﺗﻯﺎﻘﻟﺍ:ﻱﺮﻴﮔﻪﺠﻴﺘﻧ *

.ﺩﺭﺍﺩﺖﺒﺜﻣﺮﻴﺛﺎﺗﺰﻛﻮﻠﮔﻪﺑﺦﺳﺎﭘﺭﺩﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗﻥﺍﺰﻴﻣﺮﺑﻭﺪﻨﻛﻰﻣﻚﻳﺮﺤﺗﺍﺭﺎﺘﺑﻝﻮﻠﺳﺺﺧﺎﺷﻱﺎﻫﻥژ

ﻰﻟﻮﻠﺳﻥﻭﺮﺑﺲﻜﻳﺮﺗﺎﻣ،ﺎﺘﺑﻝﻮﻠﺳ،ﺰﻳﺎﻤﺗ،ﻰﻨﻴﻨﺟﻯﺩﺎﻴﻨﺑﻝﻮﻠﺳ:ﻥﺎﮔژﺍﻭﺪﻴﻠﻛ *

250-259 :ﺕﺎﺤﻔﺻ،87 ﻥﺎﺘﺴﻣﺯ،4 ﻩﺭﺎﻤﺷ،ﻢﻫﺩﻝﺎﺳ،ﻪﺘﺧﺎﻳﻰﻜﺷﺰﭘﻪﻣﺎﻨﻠﺼﻓ

ﻪﻣﺪﻘﻣ

ﻲﺑﺮﭼ،ﺕﺍﺭﺪﻴﻫﻮﺑﺮﻛﻢﺴﻴﻟﻮﺑﺎﺘﻣﻝﻼﺘﺧﺍﻡﺭﺪﻨﺳﻚﻳﻦﻳﺮﻴﺷﺖﺑﺎﻳﺩ

ﻊﻗﺍﻭﺭﺩ .ﺪﻨﺘﺴﻫﻼﺘﺒﻣﻥﺁﻪﺑﺎﻴﻧﺩﻡﺩﺮﻣﺯﺍﺮﻔﻧﺎﻫﻥﻮﻴﻠﻴﻣﻪﻛﺖﺳﺍﻦﻴﺌﺗﻭﺮﭘﻭ

ﻲﻌﻴﺒﻃ ﺪﺣ ﺯﺍﺮﺘﺸﻴﺑ ﻥﻮﺧﺪﻨﻗﺖﻈﻠﻏﻥﺁﺭﺩ ﻪﻛﺖﺳﺍﻱﺭﺎﻤﻴﺑ ﻲﻋﻮﻧ

ﻭ 1 ﻉﻮﻧ :ﺩﺮﻛ ﻢﻴﺴﻘﺗﻰﻠﺻﺍ ﻩﻭﺮﮔﻭﺩ ﻪﺑ ﻥﺍﻮﺗﻲﻣﺍﺭﻦﻳﺮﻴﺷﺖﺑﺎﻳﺩ .ﺖﺳﺍ

ﻦﻳﺍ .ﺪﻨﻣﺎﻧﻲﻣﻦﻴﻟﻮﺴﻧﺍﻪﺑﻪﺘﺴﺑﺍﻭﺖﺑﺎﻳﺩﺍﺭ 1 ﻉﻮﻧﻦﻳﺮﻴﺷﺖﺑﺎﻳﺩ .2 ﻉﻮﻧ

ﺩﺎﺠﻳﺍﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗﻡﺪﻋﺮﺛﺍﺭﺩﻭﺖﺳﺍﻥﻻﺎﺳﻢﻛﺖﺑﺎﻳﺩ،ﺖﺑﺎﻳﺩﻉﻮﻧ

ﻦﻴﻟﻮﺴﻧﺍﺪﻟﻮﻣ ﻱﺎﻫﻝﻮﻠﺳ ﻦﻤﻳﺍﺩﻮﺧ ﺐﻳﺮﺨﺗﺎﺑ ﻪﻛ ﺖﺳﺍ ﻲﻟﻼﺘﺧﺍ .ﺩﻮﺷﻲﻣ

ﻪﺑﻪﺘﺴﺑﺍﻭﺮﻴﻏﺖﺑﺎﻳﺩﻪﻛﺍﺭ 2 ﻉﻮﻧ ﺖﺑﺎﻳﺩ .ﺩﻮﺷﻲﻣ ﺺﺨﺸﻣ ﺱﺍﺮﻜﻧﺎﭘﺭﺩ

ﺕﺍﺮﺛﺍﻪﺑ ﻑﺪﻫﻱﺎﻫﺖﻓﺎﺑﺖﻴﺳﺎﺴﺣﺶﻫﺎﻛﺮﺛﺍ ﺮﺑﺪﻨﻣﺎﻧﻲﻣﻦﻴﻟﻮﺴﻧﺍ

ﺖﻠﻋﻪﺑ ﺪﻧﺍﻮﺗﻲﻣ 2 ﻉﻮﻧ ﺖﺑﺎﻳﺩ .ﺪﻳﺁﻲﻣﺩﻮﺟﻭﻪﺑﻦﻴﻟﻮﺴﻧﺍﻚﻴﻟﻮﺑﺎﺘﻣ

ﺩﺮﻜﻠﻤﻋﺎﺗ ﻦﻴﻟﻮﺴﻧﺍ ﺕﺍﺮﺛﺍ ﻪﺑ ﻲﻄﻴﺤﻣ ﻱﺎﻫﺖﻓﺎﺑ ﺖﻣﻭﺎﻘﻣﺯﺍﺩﺪﻌﺘﻣﻞﻣﺍﻮﻋ

ﺪﺷﺎﺑ ﻲﺳﺍﺮﻜﻧﺎﭘﻯﺎﺘﺑ ﻱﺎﻫﻝﻮﻠﺳ گﺮﻣﺯﺍﻰﺷﺎﻧﺎﺘﺑ ﻱﺎﻫﻝﻮﻠﺳ ﺢﻴﺤﺻﺎﻧ .(1-3)

ﻪﻫﺩ ﻦﻳﺪﻨﭼ ﻲﻧﺎﻣﺭﺩ ﻑﺍﺪﻫ ﺍﺯﺍﻲﺳﺍﺮﻜﻧﺎﭘﻯﺎﺘﺑ ﻱﺎﻫﻝﻮﻠﺳ ﻲﻨﻳﺰﮕﻳﺎﺟ . ﺖﺳﺍ ﻩﺩﻮﺑ 1 ﻉﻮﻧﺖﺑﺎﻳﺩﺖﻓﺮﺸﻴﭘﻭﺮﻴﻣﻭگﺮﻣﻥﺍﺰﻴﻣﺶﻫﺎﻛﻱﺍﺮﺑ

ﺮﻳﺍﺰﺟﺎﻳ ﻞﻣﺎﻛ ﺱﺍﺮﻜﻧﺎﭘ ﻞﻜﺷ ﻪﺑ ﻦﻴﻟﻮﺴﻧﺍﺪﻟﻮﻣﻯﺎﺘﺑ ﻱﺎﻫﻝﻮﻠﺳﺪﻧﻮﻴﭘ

ﻦﻳﺍ ﻥﺎﻣﺭﺩ ﻱﺍﺮﺑ ﻱﺍﻩﺪﻨﻨﻛﺭﺍﻭﺪﻴﻣﺍ ﻲﻧﺎﻣﺭﺩ ﻪﻨﻳﺰﮔ ،ﻩﺪﺷﺍﺪﺟ ﺲﻧﺎﻫﺮﮕﻧﻻ

ﻭﻚﻳژﻮﻟﻮﻧﻮﻤﻳﺍﺩﺭﺎﻫﺵﻭﺭﻦﻳﺍﻰﻠﺻﺍﺕﻼﻜﺸﻣﺯﺍﺎﻣﺍ . ﺖﺳﺍ ﻱﺭﺎﻤﻴﺑ

ﻩﺪﺷﺍﺪﺟ ﺲﻧﺎﻫﺮﮕﻧﻻﺮﻳﺍﺰﺟﺎﻳ ﻞﻣﺎﻛ ﺱﺍﺮﻜﻧﺎﭘﻊﺑﺎﻨﻣﻦﻴﻣﺎﺗﺭﺩﺖﻳﺩﻭﺪﺤﻣ .ﺖﺳﺍﺪﻧﻮﻴﭘﻯﺍﺮﺑﻰﻧﺎﺴﻧﺍ

ﺎﺘﺑ ﻱﺎﻫﻝﻮﻠﺳ ﺯﺍ ﻱﺪﻳﺪﺟ ﻊﺑﺎﻨﻣ ﻝﺎﺒﻧﺩ ﻪﺑ ﻥﺎﻘﻘﺤﻣ ﻞﻴﻟﺩ ﻦﻴﻤﻫ ﻪﺑ

ﻩﺪﻨﻨﻛﺪﻴﻟﻮﺗ ﺖﻓﺎﺑ ﺪﻴﻟﻮﺗ ﻱﺍﺮﺑ ﻯﺩﺎﻳﺯ ﻲﺗﺎﻘﻴﻘﺤﺗ ﻱﺎﻫﺵﻼﺗ ﻭﺪﻨﺘﺴﻫ .ﺖﺳﺍ ﻩﺪﺷﺰﻛﺮﻤﺘﻣ ﻥﺎﺴﻧﺍﻭ ﻰﻧﺍﻮﻴﺣﻯﺎﻫﻝﺪﻣﺭﺩﺪﻧﻮﻴﭘﻯﺍﺮﺑ ﻦﻴﻟﻮﺴﻧﺍ

ﻱﺎﻫﻝﻮﻠﺳﺯﺍﻲﻌﺒﻨﻣﺯﺍﻩﺩﺎﻔﺘﺳﺍ،ﺖﻓﺎﺑﺎﻳﻮﻀﻋﺪﻧﻮﻴﭘﻱﺍﺮﺑﻦﻳﺰﮕﻳﺎﺟﻚﻳ

ﺎﻳ ﻥﻻﺎﺴﮔﺭﺰﺑﺯﺎﺳﺶﻴﭘ-ﻯﺩﺎﻴﻨﺑ ﻱﺎﻫﻝﻮﻠﺳﺰﻳﺎﻤﺗﻭﺖﺳﺍﻩﺪﻧﻮﺷﺪﻳﺪﺠﺗ ﺎﻫﺵﻭﺭﻦﻳﺍﺯﺍﻰﻜﻳﻦﻴﻟﻮﺴﻧﺍ ﺪﻟﻮﻣ ﻲﺳﺍﺮﻜﻧﺎﭘﻯﺎﺘﺑ ﻱﺎﻫﻝﻮﻠﺳﻪﺑﻲﻨﻴﻨﺟ

ﺪﻧﺭﺩﺎﻗﻪﻛﺪﻨﺘﺴﻫﺍﺮﻴﻣﺎﻧﻲﻳﺎﻫﻝﻮﻠﺳ،ﻲﻨﻴﻨﺟﻱﺩﺎﻴﻨﺑﻱﺎﻫﻝﻮﻠﺳ . (4) ﺖﺳﺍ

ﻦﻳﺍﺰﻳﺎﻤﺗﻪﻨﻴﻣﺯﺭﺩﻯﺩﺎﻳﺯﺕﺎﻌﻟﺎﻄﻣ .ﺪﻨﺑﺎﻳﺰﻳﺎﻤﺗﻰﻟﻮﻠﺳﻥﺎﻣﺩﻭﺩﻦﻳﺪﻨﭼﻪﺑ

ﻪﺘﻓﺮﮔﻡﺎﺠﻧﺍﻦﻴﻟﻮﺴﻧﺍﺪﻟﻮﻣﻯﺎﻫﻝﻮﻠﺳﻪﺑﻲﻫﺎﮕﺸﻳﺎﻣﺯﺁﻂﻳﺍﺮﺷﺭﺩﺎﻫﻝﻮﻠﺳ

ﺖﺑﺎﻳﺩﻥﺎﻣﺭﺩﺭﺩﻩﻮﻘﻟﺎﺑﻱﺭﺎﻜﻫﺍﺭﻥﺍﻮﻨﻋﻪﺑﻯﺩﺎﻴﻨﺑﻯﺎﻫﻝﻮﻠﺳﺯﺍﻩﺩﺎﻔﺘﺳﺍﻭ

.ﺖﺳﺍﻪﺘﻓﺮﮔﺭﺍﺮﻗﻪﺟﻮﺗﺩﺭﻮﻣ 1 ﻉﻮﻧ

(3)

.(5-8) ﺖﺳﺍ ﻩﺪﺷ ﻪﺋﺍﺭﺍ ﻲﺷﻮﻣ ﻭ ﻲﻧﺎﺴﻧﺍ ﻲﻨﻴﻨﺟ ﻱﺩﺎﻴﻨﺑ ﻱﺎﻫﻝﻮﻠﺳ

ﺮﺛﺍ ﻲﻨﻴﻨﺟ ﻱﺎﻫﻪﺘﺧﺎﻳﻦﺑ ﺭﺍﺩﺖﻬﺟ ﺰﻳﺎﻤﺗ ﻱﻭﺭ ﻲﻔﻠﺘﺨﻣ ﻱﺎﻫﺭﻮﺘﻛﺎﻓ

ﻲﻟﻮﻠﺳ ﻥﻭﺮﺑ ﻪﻨﻴﻣﺯ ﻩﺩﺎﻣ ﻪﺑ ﻥﺍﻮﺗﻲﻣ ﻪﻠﻤﺟ ﻥﺁ ﺯﺍ ﻪﻛ ﺪﻧﺭﺍﺬﮔﻲﻣ

ﺲﻜﻳﺮﺗﺎﻣ .ﺩﺮﻛ ﻩﺭﺎﺷﺍ (Extra Cellular Matrix: ECM)

،ﺎﻫﻥﺎﻜﻴﻠﮔﻮﺌﺗﻭﺮﭘ ﺯﺍ ﻞﻜﺸﺘﻣ ﻱﺍﻪﺘﻓﺎﻳ ﻥﺎﻣﺯﺎﺳ ﻪﻜﺒﺷ ،ﻲﻟﻮﻠﺳ ﻥﻭﺮﺑ

،ﺎﻫﻦﻴﻨﻴﻣﻻﻪﻠﻤﺟﺯﺍﺩﺪﻌﺘﻣﻱﺎﻫﻦﻴﺌﺗﻭﺮﭘﻮﻜﻴﻠﮔﻭ ﻥﺎﻜﻴﻠﮔﻮﻨﻴﻣﺁﺯﻮﻜﻴﻠﮔ

ﻭ ﻯﺭﺎﺘﺧﺎﺳﺏﻮﭼﺭﺎﭼ ،ﻪﻋﻮﻤﺠﻣ ﻦﻳﺍ .ﺖﺳﺎﻫﻥژﻼﻛ ﻭ ﺎﻫﻦﻴﺘﻜﻧﻭﺮﺒﻴﻓ

ﻭﺪﻧﺭﻭﺁﻰﻣﻢﻫﺍﺮﻓﺪﻧﺍﻪﺘﻓﺮﮔﺮﺑﺭﺩﻪﻛﻰﻳﺎﻫﻝﻮﻠﺳﻯﺍﺮﺑﺍﺭﻰﺘﺴﻳﺯﻢﻳﻼﻋ

ﻦﻴﻨﺟﺭﺩﻰﻟﻮﻠﺳﻥﻭﺮﺑﻪﻨﻴﻣﺯﻩﺩﺎﻣ .ﺪﻨﻨﻛﻰﻣﻢﻴﻈﻨﺗﺍﺭﻰﻟﻮﻠﺳﺰﻳﺎﻤﺗﺪﻨﻳﺍﺮﻓ

ﻯﺍﺰﺟﺍﺐﻴﻛﺮﺗﺪﻨﭼﺮﻫ،ﺪﺑﺎﻳﻰﻣﺮﻫﺎﻈﺗﻰﻟﻮﻠﺳﻭﺩﻪﻠﺣﺮﻣﺯﺍﻥﺍﺭﺍﺪﻧﺎﺘﺴﭘ

ﻪﺑﺖﺒﺴﻧﻥﺁﻰﻳﺎﻀﻓ ﺖﻴﻌﻗﻮﻣﻭﻯﺮﻴﮔﺭﺍﺮﻗ ﺯﺮﻃﻭﻥﺁﻩﺪﻨﻫﺩ ﻞﻴﻜﺸﺗ

ﺖﺳﺍﻦﻳﺍﺮﺑﻝﺍﺩﺪﻫﺍﻮﺷ .(9) ﺩﺭﺍﺩﺕﻭﺎﻔﺗﻒﻠﺘﺨﻣﻯﺎﻫﺖﻓﺎﺑﺭﺩﻝﻮﻠﺳ

ﻝﻮﻠﺳﺶﻨﻛﺭﺪﻧﺍ،ﺪﻧﺮﻴﮔﻲﻣﻞﻜﺷﻲﺳﺍﺮﻜﻧﺎﭘﺮﻳﺍﺰﺟﻱﺎﻫﻝﻮﻠﺳﻲﺘﻗﻭﻪﻛ

ﻝﻮﻠﺳ-ﻝﻮﻠﺳ ﻱﺎﻫﺶﻨﻛﺭﺪﻧﺍ ﻭ ﻲﻟﻮﻠﺳﻥﻭﺮﺑ ﻪﻨﻴﻣﺯ ﻩﺩﺎﻣﺎﺑ ﻱﺍ ﻩﺮﻳﺰﺟ

.ﺪﻨﻨﻛﻲﻣ ﻢﻴﻈﻨﺗ ﺍﺭ ﻱﺍﻩﺮﻳﺰﺟ ﻱﺎﻫﻝﻮﻠﺳ ﻱﺪﻌﺑ ﻱﺪﻨﺑﻢﻴﺴﻘﺗ ﻭ ﻊﻤﺠﺗ

ﻲﻨﻳﺮﻛﻭﺪﻧﺍﻱﺎﻫﻝﻮﻠﺳﻉﺍﻮﻧﺍﻲﻫﺪﻧﺎﻣﺯﺎﺳﻪﻛﻩﺩﺍﺩﻥﺎﺸﻧﺮﻴﺧﺍﺕﺎﻌﻟﺎﻄﻣ

،«ﺎﻫﻦﻳﺮﮕﺘﻨﻳﺍ» ﻪﻠﻴﺳﻭ ﻪﺑ ﺪﻧﺍﻮﺗﻲﻣ ،ﺐﺳﺎﻨﻣ ﻱﺍﻩﺮﻳﺰﺟ ﻱﺎﻫﺭﺎﺘﺧﺎﺳ ﻪﺑ

ﻥﺎﻴﺑﺎﺘﺑﻱﺎﻫﻝﻮﻠﺳ ﻭ ﻱﺍﻩﺮﻳﺰﺟﻱﺎﻫﻝﻮﻠﺳﺭﺩ ﻪﻛﻲﻳﺎﺸﻏﻱﺎﻫﻩﺪﻧﺮﻴﮔ

ﻢﻴﻈﻨﺗ،ﺪﻧﻮﺷﻲﻣﻞﺼﺘﻣﻲﻟﻮﻠﺳﻥﻭﺮﺑﻪﻨﻴﻣﺯﻩﺩﺎﻣﻱﺎﻫﻦﻴﺌﺗﻭﺮﭘﻪﺑﻭﻩﺪﺷ

ﻲﻟﻮﻠﺳ ﻥﻭﺮﺑ ﻪﻨﻴﻣﺯ ﻩﺩﺎﻣ-ﻝﻮﻠﺳ ﺶﻨﻛﺭﺪﻧﺍ ﻱﺩﺮﻜﻠﻤﻋ ﺖﻴﻤﻫﺍ .ﺩﻮﺷ

،ﻩﺪﺷ ﻡﺎﺠﻧﺍ ﻱﺍﻩﺮﻳﺰﺟ ﻱﺎﻫﻝﻮﻠﺳ ﺖﺸﻛ ﻱﻭﺭ ﺮﺑ ﻪﻛ ﻲﺗﺎﻌﻟﺎﻄﻣ ﻲﻃ

ﻱﺯﺎﺳﺍﺪﺟﻱﺍﻩﺮﻳﺰﺟﻱﺎﻫﻝﻮﻠﺳﺕﺎﻌﻟﺎﻄﻣﻦﻳﺍﺭﺩ .ﺖﺳﺍﻩﺪﺷﻩﺩﺍﺩﻥﺎﺸﻧ

ﺢﺷﺮﺗ ﻲﻳﺎﻧﺍﻮﺗ ﺲﻜﻳﺮﺗﺎﻣ ﻥﻭﺪﺑ ﻲﻜﻴﺘﺳﻼﭘ ﺡﻮﻄﺳ ﻱﻭﺭ ﺮﺑ ،ﻩﺪﺷ

ﺖﺳﺩﺯﺍﺍﺭﺰﻛﻮﻠﮔﻚﻳژﻮﻟﻮﻳﺰﻴﻓﻱﺎﻫﺖﻈﻠﻏﺎﺑﻚﻳﺮﺤﺗﺯﺍﺪﻌﺑﻦﻴﻟﻮﺴﻧﺍ

ﻥﻭﺮﺑﺲﻜﻳﺮﺗﺎﻣﻱﻭﺭﺮﺑﻱﺍﻩﺮﻳﺰﺟﻱﺎﻫﻝﻮﻠﺳﻲﺘﻗﻭﻪﻛﻲﻟﺎﺣﺭﺩ .ﺪﻨﻫﺩﻲﻣ

.ﺪﻨﺑﺎﻳﻲﻣﺯﺎﺑﺍﺭﺰﻛﻮﻠﮔﻪﺑﺦﺳﺎﭘﺩﺮﻜﻠﻤﻋ،ﺪﻧﻮﺷﻲﻣﻩﺩﺍﺩﺖﺸﻛﻲﻟﻮﻠﺳ

ﻰﻤﻬﻣﺶﻘﻧﻰﻟﻮﻠﺳﻥﻭﺮﺑﺲﻜﻳﺮﺗﺎﻣﻪﻛﺪﻧﺍﻩﺩﺍﺩﻥﺎﺸﻧﻒﻠﺘﺨﻣﺕﺎﻌﻟﺎﻄﻣ

(13) ﺎﻫﺖﻴﺳﻮﻴﻣﻮﻳﺩﺭﺎﻛ (10-12) ﺎﺘﺑﻯﺎﻫﻝﻮﻠﺳﻰﻫﺎﮕﺸﻳﺎﻣﺯﺁﺰﻳﺎﻤﺗﺭﺩ

ﺎﻬﻨﺗﻪﻧﺐﺳﺎﻨﻣﺮﺘﺴﺑﻚﻳﻥﺩﺭﻭﺁﻢﻫﺍﺮﻓﺎﺑﻰﻟﻮﻠﺳﻥﻭﺮﺑﺲﻜﻳﺮﺗﺎﻣ .ﺩﺭﺍﺩ

ﻝﻮﻠﺳﺰﻳﺎﻤﺗﻭﺮﻴﺜﻜﺗ،ﺎﻘﺑﺮﺑﻪﻜﻠﺑﺩﻮﺷﻰﻣﺎﻫﻝﻮﻠﺳﺮﺘﻬﺑﻝﺎﺼﺗﺍﺐﺟﻮﻣ

ﻪﻛﻰﺴﻜﻳﺮﺗﺎﻣﻉﻮﻧﺪﺳﺭﻰﻣﺮﻈﻧﻪﺑ ﻭﺭﻦﻳﺍﺯﺍ .ﺖﺳﺍﺭﺍﺬﮔﺮﻴﺛﺎﺗﺰﻴﻧﺎﻫ

ﻢﻫﺍﺮﻓ ﺭﺩ ﻰﻤﻬﻣ ﺶﻘﻧ ﺪﻧﺍﻮﺗﻰﻣ ﺪﻧﻮﺷﻰﻣ ﺖﺸﻛ ﻥﺁ ﻯﻭﺭ ﺎﻫﻝﻮﻠﺳ

ﻪﺑﻱﺰﻳﺎﻤﺗﺦﺳﺎﭘ .ﺪﺷﺎﺑﻪﺘﺷﺍﺩﺎﻬﻧﺁ ﺐﺳﺎﻨﻣﻯﺰﻳﺎﻤﺗﻂﻴﺤﻣﺰﻳﺭﻥﺩﺭﻭﺁ

ﺎﻳﻥﻭﺭﺩ،ﺎﻫﻝﻮﻠﺳ .ﺖﺳﺍﻝﻮﻠﺳﻉﻮﻧﻪﺑﻪﺘﺴﺑﺍﻭ،ﻒﻠﺘﺨﻣﻱﺎﻫﺲﻜﻳﺮﺗﺎﻣ

ﺕﺎﻗﻭﺍﺯﺍﻱﺭﺎﻴﺴﺑﻭﺪﻨﻨﻛﻰﻣﺭﺍﺮﻗﺮﺑﻁﺎﺒﺗﺭﺍﺮﮕﻳﺪﻜﻳﺎﺑﺲﻜﻳﺮﺗﺎﻣﻱﻭﺭ

ﻦﻳﺍﺭﺩ .ﺪﻨﻨﻛﻲﻣﺍﺪﻴﭘ ﻥﺎﺷﺩﻮﺧ ﺄﺸﻨﻣﺖﻓﺎﺑ ﺎﺑﻪﺑﺎﺸﻣ ﻱﺪﻌﺑ ﻪﺳﺖﻟﺎﺣ

ﻦﻴﻟﻮﺴﻧﺍﺪﻟﻮﻣﻱﺎﻫﻝﻮﻠﺳﺰﻳﺎﻤﺗﺮﺑﻲﻟﻮﻠﺳﻥﻭﺮﺑﻪﻨﻴﻣﺯ ﻩﺩﺎﻣﺮﻴﺛﺎﺗﻪﻌﻟﺎﻄﻣ

.ﺖﺳﺍﻪﺘﻓﺮﮔ ﺭﺍﺮﻗ ﻰﺑﺎﻳﺯﺭﺍ ﺩﺭﻮﻣ ،ﺵﻮﻣ ﻲﻨﻴﻨﺟﻯﺩﺎﻴﻨﺑ ﻱﺎﻫﻝﻮﻠﺳ ﺯﺍ

/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍﻰﻠﭘ ﻭ ﻦﻴﺗﻻژ ،ﻝﮋﻳﺮﺗﺎﻣ ،ﻩﺩﺎﻔﺘﺳﺍ ﺩﺭﻮﻣ ﻯﺎﻫﺲﻜﻳﺮﺗﺎﻣ

.ﺪﻧﺍﻩﺩﻮﺑﻦﻴﻨﻴﻣﻻ

ﺎﻫ

ﺵﻭﺭ

ﻭ

ﺩﺍﻮﻣ

B1ﻥﺎﻳﻭﺭﻩﺩﺭﺵﻮﻣﻲﻨﻴﻨﺟﻱﺩﺎﻴﻨﺑﻱﺎﻫﻝﻮﻠﺳﺯﺍﻪﻌﻟﺎﻄﻣﻦﻳﺍﺭﺩ

.(14) ﺪﺷﻩﺩﺎﻔﺘﺳﺍ ﻲﻨﻴﻨﺟﻱﺩﺎﻴﻨﺑﻱﺎﻫﻝﻮﻠﺳﺯﺍﻦﻴﻟﻮﺴﻧﺍﺪﻟﻮﻣﻱﺎﻫﻝﻮﻠﺳﺰﻳﺎﻤﺗ

ﻲﺷﻮﻣﻲﻨﻴﻨﺟﻱﺩﺎﻴﻨﺑﻱﺎﻫﻝﻮﻠﺳﺖﺸﻛ

ﻲﻛﺪﻧﺍﺎﺑ (15)ﺶﻧﺍﺭﺎﻜﻤﻫﻭﻙﻭﺰﻴﻠﺑﺵﻭﺭﺱﺎﺳﺍﺮﺑﺰﻳﺎﻤﺗﻞﺣﺍﺮﻣ

.ﺖﺳﺍﻩﺪﺷﻩﺩﺍﺩﻥﺎﺸﻧ 1 ﻞﻜﺷﺭﺩﻪﺻﻼﺧﺭﻮﻃﻪﺑﻪﻛﺪﺷﻡﺎﺠﻧﺍﺕﺍﺮﻴﻴﻴﻐﺗ

ﺭﺎﻛﻞﺣﺍﺮﻣﺯﺍﻚﻴﺗﺎﻤﺷﻯﺮﻳﻮﺼﺗ : 1ﻞﻜﺷ

ﻪﺑ ﻥﺩﺮﻛ ﻪﻨﻴﺴﭙﻳﺮﺗﻪﻠﻴﺳﻭ ﻪﺑ ﻱﺩﺎﻴﻨﺑ ﻱﺎﻫﻝﻮﻠﺳ ،ﺰﻳﺎﻤﺗ ﺯﺎﻏﺁ ﻱﺍﺮﺑ

ﺖﺳﻼﺑﻭﺮﺒﻴﻓ ﻩﺪﻨﻨﻛ ﻪﻳﺬﻐﺗ ﻱﺎﻫﻝﻮﻠﺳﺯﺍ ﺰﻴﻧ ﻭ ﺪﻧﺪﻣﺁﺭﺩ ﻰﻜﺗ ﺕﺭﻮﺻ

ﻲﻨﻴﻨﺟ ﻪﺒﺷ ﻡﺎﺴﺟﺍﻞﻴﻜﺸﺗ ﻱﺍﺮﺑ ﺵﺭﺎﻤﺷ ﺯﺍﺲﭘ .ﺪﻧﺪﺷ ﺍﺪﺟ ﻲﺷﻮﻣ

ﺖﺸﻛﻂﻴﺤﻣﺭﺩﺎﻫﻝﻮﻠﺳﺯﺍﻲﻧﻮﻴﺴﻧﺎﭙﺳﻮﺳ (Embryoid Body; EB)

Leucemia Inhibitory Factor (LIF)ﺪﻗﺎﻓ ﻯﺩﺎﻴﻨﺑﻯﺎﻫﻝﻮﻠﺳﺹﻮﺼﺨﻣ

ﻲﺘﻧﺎﺳ 10 ﺖﻴﻠﭘﻩﺪﺷﻥﻮﮔژﺍﻭﺏﺭﺩﻲﻠﺧﺍﺩﺢﻄﺳﻱﻭﺭﻯﺭﺍﺬﮔﻩﺮﻄﻗ .ﺪﺷﻪﻴﻬﺗ

ﺖﻴﻠﭘﻭﺪﺷﻩﺪﻧﺍﺩﺮﮔﺮﺑﺖﻴﻠﭘﻱﻭﺭﻲﻣﺍﺭﺁﻪﺑﺏﺭﺩﺲﭙﺳ .ﺪﺷﻡﺎﺠﻧﺍﻱﺮﺘﻣ

20) ﻩﺮﻄﻗﺮﻫﺭﺩﻪﻛﻯﺭﻮﻃﻪﺑ .ﺪﺷﻩﺩﺍﺩﺭﺍﺮﻗﺭﻮﺗﺎﺑﻮﻜﻧﺍﻥﻭﺭﺩﻱﺮﺘﻣﻲﺘﻧﺎﺳ 10

،ﺯﻭﺭﻭﺩﺖﺷﺬﮔﺯﺍﺪﻌﺑ .ﺪﺷﺎﺑﻪﺘﺷﺍﺩﺩﻮﺟﻭﻝﻮﻠﺳ 500 ﺩﺍﺪﻌﺗ (ﻯﺮﺘﻴﻟﻭﺮﻜﻴﻣ 6 ﻲﻳﺎﻳﺮﺘﻛﺎﺑﺖﻴﻠﭘﻪﺑﻮﻳﺮﺘﺳﺍپﻮﻜﺳﻭﺮﻜﻴﻣﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑﻲﻨﻴﻨﺟﻪﺒﺷﻡﺎﺴﺟﺍ

ﻯﺩﺎﻴﻨﺑﻯﺎﻫﻝﻮﻠﺳﺹﻮﺼﺨﻣﺖﺸﻛﻂﻴﺤﻣﺮﺘﻴﻟﻰﻠﻴﻣ 4 ﻱﻭﺎﺣﻱﺮﺘﻣﻲﺘﻧﺎﺳ

.ﺪﻧﺪﺷﻩﺩﺍﺩﺖﺸﻛﺮﮕﻳﺩﺯﻭﺭﻭﺩﺕﺪﻣﻪﺑﻭﻞﻘﺘﻨﻣ،LIFﺪﻗﺎﻓ

ﻪﻧﺎﺧ 6 ﺖﺸﻛﻑﻭﺮﻇﻪﺑﻩﺯﻭﺭ 4 ﻲﻨﻴﻨﺟﻪﺒﺷﻡﺎﺴﺟﺍﺯﺍﺩﺪﻋ 30 ﺩﺍﺪﻌﺗ ﻯﺩﺎﻴﻨﺑﻯﺎﻫﻝﻮﻠﺳﺹﻮﺼﺨﻣﺖﺸﻛﻂﻴﺤﻣﺮﺘﻴﻟﻰﻠﻴﻣ 2 ﻭﺪﻧﺪﺷﻞﻘﺘﻨﻣ (TPP)

ﺖﺸﻛﻂﻴﺤﻣ،ﻥﻮﻴﺳﺎﺑﻮﻜﻧﺍﺯﻭﺭﻚﻳﺯﺍﺪﻌﺑ .ﺪﺷﻪﻓﺎﺿﺍﻪﻧﺎﺧﺮﻫﻪﺑLIFﺪﻗﺎﻓ

ﺮﻫﻪﺑ (1ﻝﻭﺪﺟ) ﻱﺩﻮﺧﻪﺑﺩﻮﺧﺰﻳﺎﻤﺗﺖﺸﻛﻂﻴﺤﻣﺯﺍﺮﺘﻴﻟﻰﻠﻴﻣ 2 ﻭﻑﺬﺣ

ﺭﺎﺑﻚﻳﺯﻭﺭﻭﺩﺮﻫﺖﺸﻛﻂﻴﺤﻣﻩﺯﻭﺭ 8 ﻪﻠﺣﺮﻣﻦﻳﺍﺭﺩ .ﺪﺷﻪﻓﺎﺿﺍﻪﻧﺎﺧ

.ﺪﺷﻲﻣﺾﻳﻮﻌﺗ

/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍﻰﻠﭘ :ﺪﺷﻩﺩﺎﻔﺘﺳﺍﺲﻜﻳﺮﺗﺎﻣﻉﻮﻧﻪﺳﺯﺍﻪﻌﻟﺎﻄﻣﻦﻳﺍﺭﺩ

ﺯﺍﻝﺮﺘﻨﻛﻩﻭﺮﮔﺭﺩ .(ﺕﻭﺎﻔﺘﻣﺖﻈﻠﻏﻭﺩﺎﺑ) ﻝﮋﻳﺮﺗﺎﻣﻭﻦﻴﺗﻻژ،ﻦﻴﻨﻴﻣﻻ

.ﺪﺸﻧﻩﺩﺎﻔﺘﺳﺍﻰﺸﺷﻮﭘﭻﻴﻫ

ﺖﺸﻛﻑﻭﺮﻇﻪﻧﺎﺧﺮﻫﻪﺑﻦﻴﻨﻴﻣﻻ/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍﻰﻠﭘﺎﺑﺶﺷﻮﭘﻱﺍﺮﺑ

ﻡﺮﮔﻭﺮﻜﻴﻣ 16/6 ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍ ﻰﻠﭘ ﻝﻮﻠﺤﻣ ﺮﺘﻴﻟﻭﺮﻜﻴﻣ 300 ،ﻪﻧﺎﺧ 24 .ﺪﺷﻪﺑﻮﻜﻧﺍﺭﻮﺗﺎﺑﻮﻜﻧﺍﺭﺩﺖﻋﺎﺳ 1 ﺕﺪﻣﻪﺑﻭﺪﺷﻪﻓﺎﺿﺍﺮﺘﻴﻟﻭﺮﻜﻴﻣ ﺮﺑ

ﺮﺘﻴﻟﻭﺮﻜﻴﻣ 300 ﺯﺍ .ﺪﺷﻊﻤﺟﻪﻧﺎﺧﺢﻄﺳﻱﻭﺭﺯﺍﺮﻈﻧﺩﺭﻮﻣﻝﻮﻠﺤﻣﺲﭙﺳ

ﺖﻋﺎﺳ 1 ﺕﺪﻣﻪﺑﻭﺪﺷﻩﺩﺎﻔﺘﺳﺍﺮﺘﻴﻟﻭﺮﻜﻴﻣﺮﺑﻡﺮﮔﻭﺮﻜﻴﻣ 5 ﻦﻴﻨﻴﻣﻻ ﻝﻮﻠﺤﻣ

ﺮﺘﻴﻟﻭﺮﻜﻴﻣ 300 ﻦﻴﺗﻻژﺎﺑﺶﺷﻮﭘﻱﺍﺮﺑ .ﺪﺷﻪﺑﻮﻜﻧﺍﺭﻮﺗﺎﺑﻮﻜﻧﺍﺭﺩﺮﮕﻳﺩ ،ﻝﮋﻳﺮﺗﺎﻣ ﺎﺑ ﺶﺷﻮﭘ ﻱﺍﺮﺑ ﻭ ﺮﺘﻴﻟﻰﻠﻴﻣ ﺮﺑ ﻡﺮﮔﻭﺮﻜﻴﻣ 1 ﻦﻴﺗﻻژ ﻝﻮﻠﺤﻣ

ﻂﻴﺤﻣﺭﺩ) ﺮﺑﺍﺮﺑ 30 ﻭ 100 ﻩﺩﺎﻔﺘﺳﺍﺩﺭﻮﻣ (Sigma, E 1270) ﻝﮋﻳﺮﺗﺎﻣ .ﺪﺷﻪﺑﻮﻜﻧﺍﺭﻮﺗﺎﺑﻮﻜﻧﺍﺭﺩﺖﻋﺎﺳ 3 ﺕﺪﻣﻪﺑﻭﺪﺷﻖﻴﻗﺭ (ﻲﻳﺎﻬﻧﺰﻳﺎﻤﺗﺖﺸﻛ

ﺎﺘﺑﻝﻮﻠﺳﺰﻳﺎﻤﺗﻭﻰﻟﻮﻠﺳﻥﻭﺮﺑﻪﻨﻴﻣﺯﻩﺩﺎﻣ

ﻯﺩﺎﻴﻨﺑ ﻯﺎﻫﻝﻮﻠﺳ

ﻰﻨﻴﻨﺟ ﻪﻴﺒﺷ ﻡﺎﺴﺟﺍ ﻞﻴﻜﺸﺗ

ﻪﻧﺎﺧ 6 ﻑﻭﺮﻇ ﻪﺑ ﻰﻨﻴﻨﺟ ﻪﺒﺷ ﻡﺎﺴﺟﺍ ﻝﺎﻘﺘﻧﺍ

ﻯﺩﻮﺧ ﻪﺑ ﺩﻮﺧ ﺰﻳﺎﻤﺗ

DMEM+Pen/Strp+βME+20% ES-FCS

ﻩﺪﻴﺷﻮﭘ ﻪﻧﺎﺧ 24 ﻑﻭﺮﻇ ﺭﺩ ﻥﺎﺷﺩﺪﺠﻣ ﺖﺸﻛ ﻭ ﺎﻫﻝﻮﻠﺳ ﻥﺩﺮﻛ ﻪﻨﻴﺴﭙﻳﺮﺗ ﻦﻴﻨﻴﻣﻻ /ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍ ﻰﻠﭘ ﻭ ﻦﻴﺗﻻژ ،ﻝﮋﻳﺮﺗﺎﻣ ﻯﺍ ﻪﻨﻴﻣﺯ ﺩﺍﻮﻣ ﺎﺑ

ﺰﻳﺎﻤﺗ ءﺎﻘﻟﺍ

(4)

ﻱﻭﺭﺯﺍﺮﻈﻧﺩﺭﻮﻣﻝﻮﻠﺤﻣ،ﻡﺍﺪﻛﺮﻫﻯﺍﺮﺑﻩﺪﺷﺩﺎﻳﻯﺎﻫﻥﺎﻣﺯﺯﺍﺲﭘ

ﻞﺑﺎﻗﻩﺪﺷﻩﺪﻧﺎﺷﻮﭘﺖﺸﻛﻑﻭﺮﻇﻪﻠﺣﺮﻣﻦﻳﺍﺭﺩ .ﺪﺷﻊﻤﺟﻪﻧﺎﺧﺢﻄﺳ

.ﺪﻧﺩﻮﺑﻩﺩﺎﻔﺘﺳﺍ

ﺯﺍ ﻩﺩﺎﻔﺘﺳﺍ ﺎﺑ ﺎﻫﻝﻮﻠﺳ ،ﻱﺩﻮﺧﻪﺑﺩﻮﺧ ﺰﻳﺎﻤﺗ ﻪﻠﺣﺮﻣ ﻥﺎﻳﺎﭘ ﺭﺩ

/ﻦﻴﺴﭙﻳﺮﺗ ﺪﺻﺭﺩ 0/05 ﻢﻳﺰﻧﺁ ﻝﻮﻠﺤﻣﺯﺍﻩﺩﺎﻔﺘﺳﺍ ﺎﺑ ﻭ ﻰﻤﻳﺰﻧﺁ ﺵﻭﺭ

ﺕﺭﻮﺻﻪﺑﻥﺎﻜﻣﻻﺍﻲﺘﺣﻥﺩﺮﻛژﺎﺘﭙﻴﭘﻭ (Gibco,25300) EDTA

ﻭﺪﺷﻡﺎﺠﻧﺍﻲﻟﻮﻠﺳﺵﺭﺎﻤﺷﺲﭙﺳ .ﺪﻧﺪﻣﺁﺭﺩ (Single Cell) ﻲﻜﺗ

ﻱﺎﻫﺲﻜﻳﺮﺗﺎﻣﻪﻠﻴﺳﻭ ﻪﺑ ﻼﺒﻗﻪﻛﻪﻧﺎﺧ 24 ﺖﺸﻛﻑﻭﺮﻇﻪﻧﺎﺧ ﺮﻫﻪﺑ

ﺎﻫﻝﻮﻠﺳ .ﺪﺷﻞﻘﺘﻨﻣ ﻝﻮﻠﺳ 105ﺩﺍﺪﻌﺗ،ﺩﻮﺑﻩﺪﺷ ﻩﺩﺍﺩﺶﺷﻮﭘ ﻒﻠﺘﺨﻣ

ﻝﻮﻠﺳﺪﻌﺑﺯﻭﺭ .ﺪﻧﺪﺷﻩﺩﺍﺩﺖﺸﻛ،ﻂﻴﺤﻣﻦﻴﻤﻫﺭﺩﺯﻭﺭﻚﻳﺕﺪﻣﻪﺑ

(ﺩﻮﺟﻮﻣﻱﺎﻫﺲﻜﻳﺮﺗﺎﻣﻱﻭﺭﺖﻘﻴﻘﺣﺭﺩﻭ) ﺖﺸﻛﻑﻭﺮﻇﻒﻛﻪﺑﺎﻫ

(1 ﻝﻭﺪﺟ) ﻲﻳﺎﻬﻧﺰﻳﺎﻤﺗﻂﻴﺤﻣﺎﺑﻰﻠﺒﻗﻂﻴﺤﻣﺯﻭﺭﻦﻳﺍﺭﺩ .ﺪﻧﺩﻮﺑﻩﺪﻴﺒﺴﭼ

ﺕﺪﻣﻦﻳﺍﺭﺩ .ﺪﻴﻣﺎﺠﻧﺍﻝﻮﻃﻪﺑﺯﻭﺭ 18 ﻲﻳﺎﻬﻧﺰﻳﺎﻤﺗﻪﻠﺣﺮﻣ .ﺪﺷﻦﻳﺰﮕﻳﺎﺟ

ﺰﻳﺎﻤﺗﻂﻴﺤﻣﺎﺑﺎﻫﻝﻮﻠﺳﺖﺸﻛﻂﻴﺤﻣﻡﻮﺳﻭﺩ،ﺭﺎﺑﻚﻳﺯﻭﺭ 2 ﻪﻠﺻﺎﻓﻪﺑﻭ

ﻱﺎﻫﺵﻭﺭﻪﺑﺎﻫﻝﻮﻠﺳﻪﻠﺣﺮﻣﻦﻳﺍﻥﺎﻳﺎﭘﺭﺩ .ﺪﺷﻦﻳﺰﮕﻳﺎﺟﺪﻳﺪﺟﻲﻳﺎﻬﻧ

.ﺪﻨﺘﻓﺮﮔﺭﺍﺮﻗﻲﺳﺭﺮﺑﺩﺭﻮﻣﻒﻠﺘﺨﻣ

ﻪﻠﻴﺳﻭ ﻪﺑ ﻩﺪﺷ ﮓﻧﺭ ﻯﺎﻫﻝﻮﻠﺳ ﺵﺭﺎﻤﺷ ﻭ ﻲﻤﻴﺷﻮﺘﻴﺳﻮﻨﻤﻳﺍ ﺎﻫﻱﺩﺎﺑﻲﺘﻧﺁ

ﺎﺑ ﺭﺎﺑ 2 ﻩﺎﮕﺸﻳﺎﻣﺯﺁ ﻱﺎﻣﺩ ﺭﺩ ﻩﺪﺷ ﺖﺸﻛ ﻱﺎﻫﻝﻮﻠﺳ ﺍﺪﺘﺑﺍ ﺭﺩ

Phosphate Buffered Salin (PBS)+0.05% Tween 20

ﺖﻴﺒﺜﺗ ﺲﭙﺳ .ﺪﻧﺪﺷ ﻩﺩﺍﺩ ﻮﺷﻭﺖﺴﺷ ﻪﻘﻴﻗﺩ 5 ﺕﺪﻣ ﻪﺑ ﺭﺎﺑ ﺮﻫ ﻭ

ﻪﻘﻴﻗﺩ 20 ﺕﺪﻣ ﻪﺑ ﺪﺻﺭﺩ4 ﺪﻴﻫﺪﻟﺎﻣﺮﻓﺍﺭﺎﭘ ﺯﺍ ﻩﺩﺎﻔﺘﺳﺍ ﺎﺑ ﺎﻫﻝﻮﻠﺳ

ﻝﻮﻠﺤﻣ ﺎﺑ ﺎﻫﻝﻮﻠﺳ ﻯﺮﻳﺬﭘﺫﻮﻔﻧ ﺶﻳﺍﺰﻓﺍ ﻭ ﻩﺎﮕﺸﻳﺎﻣﺯﺁ ﻱﺎﻣﺩ ﺭﺩ

ﻡﺎﺠﻧﺍ ،ﺭﻮﺗﺎﺑﻮﻜﻧﺍ ﺭﺩ ﻪﻘﻴﻗﺩ 54 ﺕﺪﻣ ﻪﺑ Triton X-100 0.2%

ﺪﺻﺭﺩ10 ﻡﺮﺳﺎﺑﺖﻋﺎﺳ 1 ﺕﺪﻣﻪﺑ ﻰﺻﺎﺼﺘﺧﺍﺮﻴﻏﻯﺎﻫﻥژﻰﺘﻧﺁ .ﺪﺷ

ﺲﭙﺳ .ﺪﻧﺪﺷ ﻩﺪﻧﺎﺷﻮﭘ (Goat Whole Serum) ﻱﺰﺑ ﻞﻣﺎﻛ

ﺭﺩ ﻭ ﻪﻴﻬﺗ ﺪﺻﺭﺩ0/5 BSA ﺭﺩ ﻩﺪﺷ ﻖﻴﻗﺭ ﻪﻴﻟﻭﺍ ﻱﺩﺎﺑﻲﺘﻧﺁ ﻝﻮﻠﺤﻣ

ﺩﺍﺮﮕﻴﺘﻧﺎﺳﻪﺟﺭﺩ 37 ﺭﻮﺗﺎﺑﻮﻜﻧﺍﺭﺩﺖﻋﺎﺳ 1 ﺕﺪﻣﻪﺑﻭﺪﺷﻪﻓﺎﺿﺍ PBS

ﻩﺰﻴﻧﻮﻳﺩﺏﺁﺭﺩﻩﺪﺷﻖﻴﻗﺭﻪﻳﻮﻧﺎﺛﻱﺩﺎﺑﻲﺘﻧﺁﺖﻳﺎﻬﻧﺭﺩ .ﺪﺷﻯﺭﺍﺪﻬﮕﻧ

.ﺪﺷﻯﺭﺍﺪﻬﮕﻧﺭﻮﺗﺎﺑﻮﻜﻧﺍﺭﺩ ﻪﻘﻴﻗﺩ 30 ﺕﺪﻣﻪﺑ ﻭ ﻪﻓﺎﺿﺍﻲﻜﻳﺭﺎﺗ ﺭﺩ

ﻰﺷﻮﻣ ﻯﺩﺎﺑﻰﺘﻧﺁ ﺯﺍﺪﻧﺩﻮﺑ ﺕﺭﺎﺒﻋﻩﺩﺎﻔﺘﺳﺍﺩﺭﻮﻣ ﻪﻴﻟﻭﺍﻯﺎﻫﻯﺩﺎﺑﻰﺘﻧﺁ

ﻰﺘﻧﺁ ﻭ (Chemicon, CBL94) C- ﺪﻴﺘﭙﭘ ﻪﻴﻠﻋ ﻝﺎﻧﻮﻠﻛﻮﻧﻮﻣ

ﻰﺘﻧﺁ.(Sigma, I 2018) ﻰﺷﻮﻣ ﻦﻴﻟﻮﺴﻧﺍ ﻪﻴﻠﻋ ﻝﺎﻧﻮﻠﻛﻮﻧﻮﻣ ﻯﺩﺎﺑ

Fluorescence IsothiocyanateﻪﺑﻞﺼﺘﻣﻰﺷﻮﻣﺪﺿIgGﻯﺩﺎﺑ

ﻪﻳﻮﻧﺎﺛ ﻯﺩﺎﺑﻰﺘﻧﺁ ﻥﺍﻮﻨﻋ ﻪﺑ ﺰﻴﻧ ﺩﻮﺑ ﻩﺪﺷ ﺪﻴﻟﻮﺗ ﺰﺑ ﺭﺩ ﻪﻛ (FITC)C

ﺰﻴﻧﺎﻫﻪﺘﺴﻫﻦﺘﻓﺮﮔﮓﻧﺭﺭﻮﻈﻨﻣﻪﺑ .ﺪﺷﻪﺘﻓﺮﮔﺭﺎﻛﻪﺑ ( Sigma, I 9259)

ﻭ ﺪﺷ ﻪﻓﺎﺿﺍ ﺎﻫﻝﻮﻠﺳ ﻪﺑ (Propidium Iodide) PI ﮓﻧﺭ

.ﺪﻧﺪﺷ ﻪﻌﻟﺎﻄﻣ ﺖﻨﺳﺭﻮﻠﻓ پﻮﻜﺳﻭﺮﻜﻴﻣ ﻚﻤﻛ ﻪﺑ ﺎﻫﻪﻧﻮﻤﻧ ﺲﭙﺳ

ﻱﺩﺎﺑﻲﺘﻧﺁ ﻪﻠﻴﺳﻭ ﻪﺑ ﻩﺪﺷ ﮓﻧﺭ ﻱﺎﻫﻝﻮﻠﺳ ﺩﺍﺪﻌﺗ ﺵﺭﺎﻤﺷ ﺭﻮﻈﻨﻣ ﻪﺑ

ﻑﺮﻇ ﻱﺎﻫﺖﻤﺴﻗ ﻡﺎﻤﺗ ﺯﺍ ﻲﻔﻠﺘﺨﻣ ﻯﺎﻫﻪﻨﻴﻣﺯ ﻲﻓﺩﺎﺼﺗ ﺭﻮﻃ ﻪﺑ ،ﺎﻫ

ﻪﺘﻓﺮﻳﺬﭘ ﮓﻧﺭﻱﺎﻫﻝﻮﻠﺳﺪﺻﺭﺩ ﻭﺪﺷﺏﺎﺨﺘﻧﺍﺎﻫﻝﻮﻠﺳ ﻱﻭﺎﺣﺖﺸﻛ

.ﺪﺷﺵﺭﺍﺰﮔﻪﻨﻴﻣﺯﻱﺎﻫﻝﻮﻠﺳﻞﻛﻪﺑﺖﺒﺴﻧ (Positive)

Ditizoneﺎﺑﻱﺰﻴﻣﺁﮓﻧﺭ

ﺭﺩ ﺍﺭ (Ditizone) DTZ ﻩﺩﺎﻣ ﺯﺍ ﻡﺮﮔﻲﻠﻴﻣ 50 ﺍﺪﺘﺑﺍ

ﺕﺪﻣﻪﺑﻭﻞﺣ(Dimethyl Sulphoxide) DMSOﺮﺘﻴﻟﻰﻠﻴﻣ 50

ﺲﭙﺳ .ﺪﺷﻩﺩﺍﺩﺭﺍﺮﻗﺩﺍﺮﮔﻰﺘﻧﺎﺳﻪﺟﺭﺩ 15 ﻯﺎﻬﻨﻣﻯﺎﻣﺩﺭﺩﻪﻘﻴﻗﺩﺪﻨﭼ

ﻪﻘﻴﻗﺩ 15 ﺕﺪﻣ ﻪﺑ ﺭﻻﻮﻣﻭﺮﻜﻴﻣ 10 ﮓﻧﺭﻲﻳﺎﻬﻧ ﺖﻈﻠﻏﺭﺩ ﺎﻫﻝﻮﻠﺳ

ﻦﻳﺍﻪﺠﻴﺘﻧﺭﺩﻪﻛﻲﻳﺎﻫﻝﻮﻠﺳ .ﺪﻧﺪﺷﻪﺑﻮﻜﻧﺍ ﺩﺍﺮﮔﻲﺘﻧﺎﺳﻪﺟﺭﺩ 37 ﺭﺩ

ﺯﺍﺲﭘ .ﺪﻧﺍﻦﻴﻟﻮﺴﻧﺍﻩﺪﻨﻨﻛﺢﺷﺮﺗ،ﺪﻨﻳﺁﺭﺩﻰﻧﺍﻮﻏﺭﺍﮓﻧﺭﻪﺑﻱﺰﻴﻣﺁﮓﻧﺭ

ﻂﻴﺤﻣﺎﻬﻧﺁﻪﺑﻭﺪﻧﺪﺷﻩﺩﺍﺩﻮﺸﺘﺴﺷ PBS-ﺎﺑﺭﺎﺑ₃ﺎﻫﻝﻮﻠﺳ،ﻱﺰﻴﻣﺁﮓﻧﺭ

.ﺪﺷﻪﻓﺎﺿﺍﻲﻳﺎﻬﻧﺰﻳﺎﻤﺗﺖﺸﻛ

ﺶﻨﻛﺍﻭ ﺱﻮﻜﻌﻣ ﻱﺭﺍﺩﺮﺑﻪﺨﺴﻧ ﺵﻭﺭ ﻪﺑ ﺎﻫﻥژ ﻥﺎﻴﺑ ﻲﺳﺭﺮﺑ ﻥﺁ ﻰﻤﻛ ﻪﻤﻴﻧ ﺶﺠﻨﺳ ﻭ (RT-PCR) ﺯﺍﺮﻣﻲﻠﭘ ﻱﺍﻩﺮﻴﺠﻧﺯ (Semi-quantitative RT-PCR)

ﺯﺍﺮﻣﻲﻠﭘ ﻱﺍﻩﺮﻴﺠﻧﺯ ﺶﻨﻛﺍﻭ ﺱﻮﻜﻌﻣ ﻱﺭﺍﺩﺮﺑﻪﺨﺴﻧ ﺵﻭﺭ ﺯﺍ

ﺹﺎﺧ (Transcripts) ﻱﺎﻫﺖﺷﻮﻧﻭﺭﻥﺎﻴﺑﻲﺑﺎﻳﺯﺭﺍﻱﺍﺮﺑ (RT-PCR)

ﻥﺎﻴﺑﻥﺍﺰﻴﻣﺮﻴﻴﻐﺗﻪﻜﻧﺁﻯﺍﺮﺑﻦﻴﻨﭽﻤﻫ .ﺪﺷﻩﺩﺎﻔﺘﺳﺍﻲﺳﺍﺮﻜﻧﺎﭘﻭﻲﻣﺭﺩﻭﺪﻧﺍ

ﺵﻭﺭﺯﺍﺩﺮﻴﮔﺭﺍﺮﻗﻰﺑﺎﻳﺯﺭﺍﺩﺭﻮﻣﺰﻴﻧﻒﻠﺘﺨﻣﻯﺎﻫﻩﻭﺮﮔﺭﺩﻢﻬﻣﻥژﺪﻨﭼ

ﻥﺎﺸﻧﺮﻈﻧﺩﺭﻮﻣﻱﺎﻫﻥژﻡﺎﻧ 2 ﻝﻭﺪﺟﺭﺩ .ﺪﺷﻩﺩﺎﻔﺘﺳﺍﻰّﻤﻛﻪﻤﻴﻧﺶﺠﻨﺳ .ﺖﺳﺍﻩﺪﺷﻩﺩﺍﺩ Nucleospin RNA II kit (740955;)ﺖﻴﻛﺯﺍ ﻩﺩﺎﻔﺘﺳﺍﺎﺑﺍﺪﺘﺑﺍﺭﺩ

ﺯﺍ RNA ﻱﺩﻮﺟﻮﻣ ﻞﻛ Macherey-Nagel, Germany)y

ﻱﺎﻫﻲﮔﺩﻮﻟﺁ ﻑﺬﺣﺭﻮﻈﻨﻣ ﻪﺑ ﻭ ﺪﺷ ﺝﺍﺮﺨﺘﺳﺍ ﻪﺘﻓﺎﻳ ﺰﻳﺎﻤﺗ ﻱﺎﻫﻝﻮﻠﺳ

ﺹﻮﻠﺧ ﺲﭙﺳ .ﺪﻨﺘﻓﺮﮔﺭﺍﺮﻗDnase I ﺎﺑﺭﺎﻤﻴﺗ ﺖﺤﺗ،ﻚﻴﻣﻮﻧژ DNA .ﺪﺷ ﻦﻴﻴﻌﺗ ﻱﺮﺘﻣﻮﺗﻮﻓﻭﺮﺘﻜﭙﺳﺍ ﺵﻭﺭﺎﺑ ﻩﺪﺷ ﺝﺍﺮﺨﺘﺳﺍ RNA ﺖﻈﻠﻏﻭ

ﺮﻤﻳﺍﺮﭘ ﺯﺍ ﻩﺩﺎﻔﺘﺳﺍ ﺎﺑ ﻩﺪﺷ ﺝﺍﺮﺨﺘﺳﺍ RNA ﺯﺍ ﻡﺮﮔﻭﺮﻜﻴﻣ 1

ﺖﻴﻛﻭRandom Hexamer (K 1622 Fermentas) Revert AidTM_{H-Minus First} Strand cDNA Synthesis

ﻩﺩﺎﻔﺘﺳﺍﺩﺭﻮﻣﺖﺸﻛﻯﺎﻫﻂﻴﺤﻣ :1 ﻝﻭﺪﺟ

ﺕﺎﻳﻮﺘﺤﻣ

ﺖﺸﻛﻂﻴﺤﻣ

KODMEM (10829-018)+2mM L-glutamin (25030-024)+0.1mM Non-essential amino acids (11140-035)+0.1mMβ-mercaptoethanol (sigma, M7522)+13.5%

Fetal Calf Serum (10270-106)+50u_/ml_Penicillin+₅₀μ_g/ml_{Streptomycine} (15070-063)+1000u/ml Leukemia Inhibitory Factor (Chemicon, ESG1107)

ﻯﺩﺎﻴﻨﺑﻯﺎﻫﻝﻮﻠﺳﺖﺸﻛﻂﻴﺤﻣ

DMEM (12800-116)+2mM L-glutamin (25030-024)+0.1mM Non-essential ami-no acids (11140-035)+0.1mMβ-mercaptoethanol (sigma, M7522)+20% Fetal Calf Serum (10270-106)+50u/ml Penicillin+50μg/ml Streptomycine (15070-063)+44mM NaHCO3

ﻱﺩﻮﺧﻪﺑﺩﻮﺧﺰﻳﺎﻤﺗﺖﺸﻛﻂﻴﺤﻣ

DEMEM/F12 (12456-018)+2mM L-glutamin (25030-024)+1% N2 (17502-048) + 2% B27 (17504-044)+10mM Nicotin amide (sigma, N0636)+50u/ml Penicillin +50μg/ml Streptomycine (15070-063)

ﻰﻳﺎﻬﻧﺰﻳﺎﻤﺗﺖﺸﻛﻂﻴﺤﻣ

(5)

ﻡﺯﻻﺩﺍﻮﻣﻡﺎﻤﺗﻪﻛﻱﺍﻪﻧﻮﻤﻧﺮﺑﻩﻭﻼﻋﺎﺠﻨﻳﺍﺭﺩ .ﺪﺷ ﺱﻮﻜﻌﻣ ﻱﺭﺍﺩﺮﺑﻪﺨﺴﻧ RTﻢﻳﺰﻧﺁﺪﻗﺎﻓﻪﻛ (RT-₎ﻲﻔﻨﻣﻝﺮﺘﻨﻛﻪﻧﻮﻤﻧﺯﺍﺩﻮﺑﺍﺭﺍﺩﺍﺭﺶﻨﻛﺍﻭﻯﺍﺮﺑ ﺭﺍﺮﻗﻲﺑﺎﻳﺯﺭﺍﺩﺭﻮﻣDNAﻪﺑﻪﻧﻮﻤﻧﻰﻟﺎﻤﺘﺣﺍﻲﮔﺩﻮﻟﺁﺎﺗﺪﺷﻩﺩﺎﻔﺘﺳﺍﺰﻴﻧﺩﻮﺑ .ﺪﺷﻩﺩﺎﻔﺘﺳﺍPCRﻱﺍﺮﺑﻮﮕﻟﺍﻥﺍﻮﻨﻋﻪﺑ ﻩﺪﺷﺪﻴﻟﻮﺗcDNAﺯﺍ ﺲﭙﺳ .ﺩﺮﻴﮔ ﻱﺎﻫﺮﻤﻳﺍﺮﭘﺯﺍ ﻖﻴﻘﺤﺗ ﻦﻳﺍﺭﺩ ﻪﻌﻟﺎﻄﻣﺩﺭﻮﻣ ﻱﺎﻫﻥژ ﻥﺎﻴﺑ ﻪﺑ ﻥﺩﺮﺑﻲﭘ ﻱﺍﺮﺑ

ﺍﺪﺟﺪﺻﺭﺩ1/7 ﺯﺭﺎﮔﺁﻝژ ﻱﻭﺭPCR ﺕﻻﻮﺼﺤﻣ .ﺪﺷ ﻩﺩﺎﻔﺘﺳﺍ 1 ﻝﻭﺪﺟ

ﺭﻮﺗﻮﻨﻴﻣﻮﻟﺲﻧﺍﺮﺗﻩﺎﮕﺘﺳﺩﺎﺑﺲﭙﺳ .ﺪﻧﺪﺷ ﺖﻳﺅﺭ ﻞﺑﺎﻗﺪﻳﺎﻣﻭﺮﺑ ﻡﻮﻳﺪﻴﺗﺍﺎﺑﻭ

.ﺖﻓﺮﮔﺭﺍﺮﻗﻲﺑﺎﻳﺯﺭﺍﺩﺭﻮﻣ (Uvidoc, UK) UV

β-Tubulinﻪﻛﺮﻈﻧﺩﺭﻮﻣﻯﺎﻫﻥژﻰّﻤﻛﻪﻤﻴﻧﺶﺠﻨﺳﻡﺎﺠﻧﺍﻯﺍﺮﺑ

ﺪﻧﺩﻮﺑInsulin1, Insulin2, Slc2a2, IAPP, Glucagonﻭ

ﻦﻴﻴﻌﺗﺎﻫﻥژﻦﻳﺍﻯﺍﺮﺑﻞﺒﻗﺯﺍﻪﻛﻰﻄﻳﺍﺮﺷﺎﺑﺯﺍﺮﻣﻲﻠﭘﻱﺍﻩﺮﻴﺠﻧﺯﺶﻨﻛﺍﻭ

ﻰﻓﺮﺼﻣﺮﻤﻳﺍﺮﭘ ﺭﺍﺪﻘﻣﻅﺎﺤﻟ ﺯﺍ ﻂﻳﺍﺮﺷﻦﻳﺍ . ﺖﻓﺮﮔﻡﺎﺠﻧﺍ ﺩﻮﺑﻩﺪﺷ

ﻪﻛﺩﻮﺑ ﻯﺍﻪﻧﻮﮔ ﻪﺑ ﺯﺍﺮﻣﻲﻠﭘ ﻱﺍﻩﺮﻴﺠﻧﺯ ﺶﻨﻛﺍﻭ ﻯﺎﻫﻞﻜﻴﺳ ﺩﺍﺪﻌﺗ ﻭ

ﺲﭘ .ﺪﻨﺷﺎﺑﻰﻳﺎﻤﻧﺯﺎﻓﺭﺩﻭﻉﺎﺒﺷﺍﺯﺎﻓﺯﺍﻞﺒﻗﻚﻴﺘﻴﻨﻛﻅﺎﺤﻟﺯﺍﺎﻫﺶﻨﻛﺍﻭ

ﺕﻻﻮﺼﺤﻣﺯﺭﻮﻓﻭﺮﺘﻜﻟﺍﻥﺁﺯﺍﺪﻌﺑﻭﺯﺍﺮﻣﻲﻠﭘﻱﺍﻩﺮﻴﺠﻧﺯﺶﻨﻛﺍﻭﻡﺎﺠﻧﺍﺯﺍ

،Uvi Docﻝژﺯﺍﻯﺭﺍﺩﺮﺑﺲﻜﻋﻢﺘﺴﻴﺳﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑ،ﻝژﻯﺰﻴﻣﺁﮓﻧﺭﻭ

ﻯﻭﺭﺮﺑﺮﻳﻭﺎﺼﺗﻦﻳﺍ .ﺪﺷﻪﺘﻓﺮﮔﺲﻜﻋﺕﻻﻮﺼﺤﻣﻯﺎﻫﺪﻧﺎﺑﻭﻝژﺯﺍ

ﻁﻮﺑﺮﻣ ﻯﺎﻫﺪﻧﺎﺑ .ﺪﻧﺪﺷ ﻩﺩﺍﺩ ﻝﺎﻘﺘﻧﺍ ﺮﺗﻮﻴﭙﻣﺎﻛ ﻪﺑ ﻭ ﻩﺮﻴﺧﺫ ﺖﻜﺴﻳﺩ

Uvi Band Map ﺭﺍﺰﻓﺍ ﻡﺮﻧ ﺯﺍ ﻩﺩﺎﻔﺘﺳﺍﺎﺑ ﺎﻫﻪﻧﻮﻤﻧ ﺯﺍ ﻚﻳﺮﻫﻪﺑ

ﺕﻻﻮﺼﺤﻣ ﻥﺍﺰﻴﻣ ﻩﺪﻣﺁﺖﺳﺩﻪﺑ ﺮﻳﺩﺎﻘﻣ .ﺪﻧﺪﺷ ﺰﻴﻟﺎﻧﺁ ﻭ ﻯﺮﺘﻣﻮﺘﻴﺴﻧﺍﺩ

ﻪﻧﻮﻤﻧ ﺮﻫ ﺩﺭﻮﻣ ﺭﺩ .ﺪﻧﺩﺍﺩﻰﻣ ﻥﺎﺸﻧ ﺍﺭ ﺎﻫﻪﻧﻮﻤﻧ ﺯﺍ ﻚﻳ ﺮﻫ PCR

ﻯﺍﺮﺑﻩﺪﻣﺁﺖﺳﺩﻪﺑﺭﺍﺪﻘﻣﺮﺑﻢﻴﺴﻘﺗ ،ﻥژﺮﻫﻯﺍﺮﺑﻩﺪﻣﺁﺖﺳﺩﻪﺑ ﺭﺍﺪﻘﻣ

ﺮﻳﺩﺎﻘﻣ ﻥﻮﻴﺳﺍﺰﻴﻟﺎﻣﺮﻧ ﺎﺗ ،ﺪﺷ ﻪﻧﻮﻤﻧ ﻥﺎﻤﻫ ﻯﺍﺮﺑ β-Tubulin ﻥژ

Housekeeping ﻯﺎﻫﻥژ ﻩﺮﻣﺯ ﺭﺩ β-Tubulin .ﺩﺮﻴﮔ ﺕﺭﻮﺻ

ﺮﻈﻧﺭﺩﺖﺑﺎﺛﻰﻟﻮﻠﺳﺭﺎﻤﻴﺗﻒﻠﺘﺨﻣﻂﻳﺍﺮﺷﺭﺩﻥﺁﻥﺎﻴﺑﻥﺍﺰﻴﻣﻪﻛﺖﺳﺍ

ﻦﻳﺍﻪﺑ .ﺩﺮﻴﮔپﻰﻣﺭﺍﺮﻗﻩﺩﺎﻔﺘﺳﺍﺩﺭﻮﻣﻰﻠﺧﺍﺩﻝﺮﺘﻨﻛﻥﺍﻮﻨﻋﻪﺑﻭﻩﺪﺷﻪﺘﻓﺮﮔ

ﻭﻯﺭﻭﺁﻊﻤﺟﺎﻫﻪﻧﻮﻤﻧ،ﻰﻟﻮﻠﺳﻯﺎﻫﺶﻳﺎﻣﺯﺁﻞﻘﺘﺴﻣﺭﺍﺮﻜﺗﻪﺳﺯﺍﺐﻴﺗﺮﺗ

ﻯﺍﺮﺑﺎﻫﻥژﻩﺪﺷﻥﺎﻴﺑﺮﻳﺩﺎﻘﻣ،ﻯﺭﺎﻣﺁﺰﻴﻟﺎﻧﺁﺎﺑﺖﻳﺎﻬﻧﺭﺩﻭﺪﻧﺪﺷﻰﺑﺎﻳﺯﺭﺍ

.ﺪﻧﺪﺷﻪﺴﻳﺎﻘﻣﻢﻫﺎﺑﻒﻠﺘﺨﻣﻯﺎﻫﻪﻧﻮﻤﻧ

ﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗﻥﺍﺰﻴﻣﻲﺳﺭﺮﺑ

،(Nacl)ﺮﺘﻣﻰﻠﻴﻣ 120 ﻞﻣﺎﺷ ﻪﻛﺮﮕﻨﻳﺭ- ﺲﺑﺮﻛﺮﻓﺎﺑﺎﺑﺎﻫﻝﻮﻠﺳﺍﺪﺘﺑﺍ

NaHCO₃ Buff-ﺮﺘﻣﻰﻠﻴﻣ 25،(Cacl₂)ﺮﺘﻣﻰﻠﻴﻣ 2/5،(Kcl)ﺮﺘﻣﻰﻠﻴﻣ 5

ﻱﻭﺎﺣﺮﻓﺎﺑ ﺲﭙﺳﻭﺪﻧﺪﺷﻮﺷﻭﺖﺴﺷﺭﺎﺑﻭﺩ،ﺖﺳﺍ BSAﺪﺻﺭﺩ10ﻭ er

ﻱﺎﻣﺩﺭﺩﻪﻘﻴﻗﺩ 60 ﺕﺪﻣﻪﺑﺲﭙﺳ .ﺪﺷﻪﻓﺎﺿﺍﺮﺘﻣﻰﻠﻴﻣ 3 ﺖﻈﻠﻏﺎﺑﺰﻛﻮﻠﮔ

ﻝﻮﻠﺳﻱﻭﺭﻂﻴﺤﻣ،ﺕﺪﻣﻦﻳﺍﺯﺍﺲﭘ .ﺪﻧﺪﺷﺭﺎﻤﻴﺗﺩﺍﺮﮔﻰﺘﻧﺎﺳﻪﺟﺭﺩ 37

ﺭﺩ،ﻦﻴﻟﻮﺴﻧﺍﻥﺍﺰﻴﻣﻲﺳﺭﺮﺑﻥﺎﻣﺯﺎﺗﻭﺪﺷﻪﺘﺷﺍﺩﺮﺑ (Supernatant) ﺎﻫ

ﺎﺑﺭﺎﺑﻭﺩﺎﻫﻝﻮﻠﺳﻥﺁﺯﺍﺲﭘ .ﺪﺷﻱﺭﺍﺪﻬﮕﻧﺩﺍﺮﮔﻰﺘﻧﺎﺳﻪﺟﺭﺩ -20 ﻱﺎﻣﺩ

ﻝﻮﻠﺳﺭﺎﺑﻦﻳﺍ،ﺪﻌﺑﻪﻠﺣﺮﻣﺭﺩ .ﺪﻧﺪﺷﻩﺩﺍﺩﻮﺷﻭﺖﺴﺷﺮﮕﻨﻳﺭـ ﺲﺑﺮﻛﺮﻓﺎﺑ

ﻱﺎﻣﺩﺭﺩﻪﻘﻴﻗﺩ 60 ﺕﺪﻣﺮﺘﻣﻰﻠﻴﻣ 22 ﺖﻈﻠﻏﺎﺑﺰﻛﻮﻠﮔﻱﻭﺎﺣﺮﻓﺎﺑﺎﺑﺎﻫ .ﺩﺮﻛﺍﺪﻴﭘﻪﻣﺍﺩﺍﻞﺒﻗﺪﻨﻧﺎﻣ،ﻞﺣﺍﺮﻣﻭﺪﻧﺪﺷﺭﺎﻤﻴﺗﺩﺍﺮﮔﻰﺘﻧﺎﺳﻪﺟﺭﺩ 37

ﺶﺠﻨﺳﺖﻴﻛﺯﺍﺎﻫﻝﻮﻠﺳﻂﺳﻮﺗﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗﻥﺍﺰﻴﻣﻲﺳﺭﺮﺑﺭﻮﻈﻨﻣﻪﺑ

(DRG، EIL-3440ﺖﻛﺮﺷﺯﺍ) ELISAﺵﻭﺭ ﻪﺑﻲﺷﻮﻣﻦﻴﻟﻮﺴﻧﺍ

.ﺪﺷﻩﺩﺎﻔﺳﺍﺖﻴﻛﻞﻤﻌﻟﺍﺭﻮﺘﺳﺩﺎﺑﻖﺑﺎﻄﻣ

ﻯﺭﺎﻣﺁﺵﻭﺭ

ﻯﺭﺎﻣﺁﺖﺴﺗﺯﺍﻩﺩﺎﻔﺘﺳﺍﺎﺑﻭSPSS 13ﺭﺍﺰﻓﺍﻡﺮﻧﺎﺑﻞﺻﺎﺣﺞﻳﺎﺘﻧ

ﻰﺳﺭﺮﺑ p<0/05 ﻯﺭﺍﺩﻰﻨﻌﻣﺢﻄﺳﺭﺩ ﻭ ﻪﻓﺮﻃﻚﻳﺲﻧﺎﻳﺭﺍﻭﺰﻴﻟﺎﻧﺁ

ﻯﺎﻄﺧ ±ﻦﻴﮕﻧﺎﻴﻣﺕﺭﻮﺻﻪﺑﺎﻫﻩﺩﺍﺩﻦﻳﺍ،ﻰّﻤﻛﻯﺎﻫﻩﺩﺍﺩﺩﺭﻮﻣﺭﺩ .ﺪﻧﺪﺷ

(RT- PCR₎ﺯﺍﺮﻤﻴﻠﭘﻱﺍﻩﺮﻴﺠﻧﺯﺶﻨﻛﺍﻭﺱﻮﻜﻌﻣﻱﺭﺍﺩﺮﺑﻪﺨﺴﻧﺵﻭﺭﺭﺩﻩﺩﺎﻔﺘﺳﺍﺩﺭﻮﻣﻱﺎﻫﺮﻤﻳﺍﺮﭘ_{: 2}ﻝﻭﺪﺟ

Genes Primer sequences (5´-3´) Annealing Tem-preture (°C)

Size (bp) Gene bank Code

Ins 1 (Insulin I)

ATC AGC AAG CAG GTC ATT GTT TAG TTC TCC AGC TGG TAG AGG G

58 343 NM_008386

Ins 2 (Insulin II)

CCT GCT GGC CCT GCT CTT AGG TCT GAA GGT CAC CTG CT

58 212 NM_008387

Sst

(somatostatin )

TGC CTG AGG ACCTGCGACTA TCT CTG TCT GGT TGG GCT CG

60 257 NM_009215

Gcg/ GLP-1 (Glucagon)

CAG AGG AGA ACC CCA GAT CA TCA TGA CGT TTG GCA ATG T

58 205 NM_008100

Slc2a2/ Glut2 (glucose transporter)

GAG TTA CGT GAG CAT GAC TG TCA CAC ACT CTC TGA AGA CG

58 388 NM_031197

Pcsk1/ PC1

(Proprotein convertase 1)

GAA GGA AGG ATT GTG AAC TG AGC AGC CTG TCA TCT CTA TG

58 485 NM_013628

Gata4 CTC TAT CAC AAG ATG AAC GGC A

CTG CTG TGC CCA TAG TGA GA

58 413 NM_008092

Nestin TCG AGC AGG AAG TGG TAG G

TTG GGA CCA GGG ACT GTT A

58 352 NM_016701

Iapp/ amylin

(Islet amyloid polypeptide)

CTC TCT GTG GCA CTG AAC CA GAT TCC CTA TTT GGA TCC CC

58 221 NM_010491

Isl1 ( islet 1) TAT CCA GGG GAT GAC AGG AAC

GCT GTT GGG TGT ATC TGG GAG

64 256 NM_021459

Cpa1

(Carboxypeptidase A1)

CTG CTC AGA AAC TTA CCG TG CCA CTG GCT TGA TAG ATC GT

56 276 NM_025350

β-Tubulin GGA ACA TAG CCG TAA ACT GC

TCA CTG TGC CTG AAC TTA CC

60 318 NM_011655

(6)

.ﺪﻧﺪﺷﺵﺭﺍﺰﮔ (Mean±SEM) ﻦﻴﮕﻧﺎﻴﻣﺯﺍﺭﺎﻴﻌﻣ

ﺎﻫ

ﻪﺘﻓﺎﻳ

ﻲﻳﺎﻫﻝﻮﻠﺳ ﺭﺩSlc2a2ﻭ Ins2, Ins1ﻯﺎﻫﻥژﻥﺎﻴﺑﺶﻳﺍﺰﻓﺍ

ﻥﺎﻴﺑ ،ﺪﻨﺘﻓﺎﻳ ﺰﻳﺎﻤﺗﻝﮋﻳﺮﺗﺎﻣ ﺎﺑ ﻩﺪﺷﻩﺩﺍﺩ ﺶﺷﻮﭘﻑﻭﺮﻇﻱﻭﺭ ﺮﺑﻪﻛ

ﻲﺳﺍﺮﻜﻧﺎﭘﺮﮕﻳﺩﻱﺎﻫﻝﻮﻠﺳ ﻭﺎﺘﺑﻱﺎﻫﻝﻮﻠﺳﺺﺧﺎﺷ ﻒﻠﺘﺨﻣﻱﺎﻫﻥژ

ﻪﻛﻲﻳﺎﻫﻥژ .(3 ﻭ 2 ﻞﻜﺷ) ﺪﺷﻲﺳﺭﺮﺑﻲﻳﺎﻬﻧﺰﻳﺎﻤﺗﻪﻠﺣﺮﻣﻥﺎﻳﺎﭘﺭﺩ

ﺎﺘﺑﻝﻮﻠﺳ ﺺﺧﺎﺷﻱﺎﻫﻥژﻞﻣﺎﺷﺖﻓﺮﮔﺭﺍﺮﻗ ﻲﺳﺭﺮﺑﺩﺭﻮﻣﺎﻬﻧﺁﻥﺎﻴﺑ

ﻲﻳﺎﺸﻏ ﻱﺎﻫﻦﻴﺌﺗﻭﺮﭘ ﻩﺪﻨﻨﻛﺰﻣﺭ ﻥژ ﻭ (IAPP ﻭ Ins I, Ins II)

Slc2a2 or) ﺪﻧﺯﺎﺳﻲﻣﺍﺭﺎﺸﻏﺯﺍﺰﻛﻮﻠﮔﺭﻮﺒﻋﻝﺎﻧﺎﻛ ﻪﻛﺎﺘﺑﻝﻮﻠﺳ

ﻩﺎﮕﺘﺳﺩﺭﺩﻦﻴﻟﻮﺴﻧﺍﻝﻮﻜﻟﻮﻣﺵﺯﺍﺩﺮﭘﺮﻴﺴﻣﻱﺩﺮﻜﻠﻤﻋﻥژ،(Glut2

،(Cpt1) ﺱﺍﺮﻜﻧﺎﭘﺰﻳﺭﻥﻭﺮﺑﺶﺨﺑﺺﺧﺎﺷﻥژ،(PC1/3) ﻱﮋﻠﮔ

،ﺖﺳﺎﺘﺑﻯﺎﻫﻝﻮﻠﺳ ﻯﺍﺮﺑﻡﺯﻻﻰﺴﻳﻮﻧﻭﺭ ﻯﺎﻫﺭﻮﺘﻛﺎﻓﺯﺍﻪﻛ Isl1ﻥژ

ﻰﺳﺍﺮﻜﻧﺎﭘ ﻯﺎﺘﻟﺩﻭ (Glucagon) ﺎﻔﻟﺁﻱﺎﻫﻝﻮﻠﺳ ﺺﺧﺎﺷ ﻯﺎﻫﻥژ

ﻥﺍﻮﻨﻋ ﻪﺑ ﺰﻴﻧ Nestin ﻭ Gta4 ﻥژ ﻭﺩ ﺯﺍ ﻭ (somatostatin)

ﺎﺗ ﺪﺷ ﻩﺩﺎﻔﺘﺳﺍ ﻰﻣﺭﺩﻮﺘﻛﺍ ﻭ ﻰﻣﺭﺩﻭﺪﻧﺍﻭﺰﻣ ﻯﺎﻫﻝﻮﻠﺳ ﻯﺎﻫﺺﺧﺎﺷ

ﺩﺭﻮﻣﺰﻴﻧﺎﻫﻝﻮﻠﺳﻉﺍﻮﻧﺍﻪﻴﻘﺑﻥﺩﻮﺑﺍﺭﺍﺩﻅﺎﺤﻟﺯﺍﻰﻳﺎﻬﻧﻰﻟﻮﻠﺳﺖﻴﻌﻤﺟ

.ﺪﻧﺮﻴﮔﺭﺍﺮﻗﻰﺑﺎﻳﺯﺭﺍ

Sst ،PC1/3 ،Cpa1 ،Isl 1 ،Gata4،Nestinﻱﺎﻫﻥژﻥﺎﻴﺑ_{: 2}ﻞﻜﺷ 1: poly-l ornithin/laminin, 2: Gelatin, 3: matrigel 1/30, 4: atrigel 1/100, 5: uncoated, M) DNA ladder

ﻒﻠﺘﺨﻣﻯﺎﻫﻩﻭﺮﮔﻦﻴﺑ Ins1, Ins2, IAPP, Slc2a2, Glucagon ﻯﺎﻫﻥژﻰّﻤﻛﻪﻤﻴﻧﻥﺎﻴﺑﻰﺳﺭﺮﺑ : 3ﻞﻜﺷ

1: poly-l ornithin /laminin(O/L), 2: Gelatin(gel), 3: matrigel 1/30(mat1/30), 4: matrigel 1/100(mat1/100), 5: uncoated, M- DNA ladder,

.ﺪﻧﺍﻩﺪﺷﻪﺋﺍﺭﺍp≤0.05, Mean±SEMﺕﺭﻮﺻﻪﺑﺎﻫﻩﺩﺍﺩ

1 2 3 4 5 1 2 3 4 5

120 100 80 60 40 20 0

Insulin 1

1 2 3 4 5

ﻥژ

ﻰﺒﺴﻧ

ﻥﺎﻴﺑ

1 2 3 4 5

160 140 120 100 80 60 40 20 0

ﻥژ

ﻰﺒﺴﻧ

ﻥﺎﻴﺑ

Slc2a2

1 2 3 4 5

ﻥژ

ﻰﺒﺴﻧ

ﻥﺎﻴﺑ

Glucagon 140

120 100 80 60 40 20 0

ﻥژ

ﻰﺒﺴﻧ

ﻥﺎﻴﺑ

Insulin 2

1 2 3 4 5

180 160 140 120 100 80 60 40 20 0

1 2 3 4 5 Insulin 1

Insulin 2

IAPP

Slc2a2

Glucagon

β-Tubulin

Iapp

1 2 3 4 5 200

180 160 140 120 100 80 60 40 20 0

ﻥژ

ﻰﺒﺴﻧ

(7)

ﺰﻴﻧﻭﻦﻴﻨﻴﻣﻻ/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍﻰﻠﭘﻭﻦﻴﺗﻻژﻱﻭﺭﻪﻛﻲﻳﺎﻫﻝﻮﻠﺳﺭﺩ

،ﺪﻨﺘﻓﺎﻳﺰﻳﺎﻤﺗ (Un-coatedﻩﻭﺮﮔ) ﻩﺪﺸﻧﻩﺩﺍﺩﺶﺷﻮﭘﺖﺸﻛﻑﻭﺮﻇ

،Slc2a2،PCl/3،IAPPﻪﻠﻤﺟﺯﺍﻲﻣﻮﻤﻋﺺﺧﺎﺷﻱﺎﻫﻥژﻥﺎﻴﺑ

ﻰﻨﻌﻳﺎﺘﺑﻯﺎﻫﻝﻮﻠﺳﻰﺻﺎﺼﺘﺧﺍﻥژﻥﺎﻴﺑﺎﻣﺍ .ﺪﺷﻩﺩﺍﺩﻥﺎﺸﻧIsl1،Cpt1

.ﺪﺷﻑﺬﺣﻰﻟﻮﻠﺳﻯﺎﻫﺭﺍﺮﻜﺗﺮﺜﻛﺍﺭﺩIns I

ﻝﮋﻳﺮﺗﺎﻣﺎﺑ ﻩﺪﺷﻩﺩﺍﺩﺶﺷﻮﭘ ﻑﻭﺮﻇﻱﻭﺭ ﺮﺑﻪﻛﻲﻳﺎﻫﻝﻮﻠﺳﺭﺩ

ﺯﺍﻪﻛﺪﺷﻥﺎﻴﺑﻪﻌﻟﺎﻄﻣﺩﺭﻮﻣﺺﺧﺎﺷﻱﺎﻫﻥژﻲﮕﻤﻫﺪﻧﺩﻮﺑﻪﺘﻓﺎﻳﺰﻳﺎﻤﺗ

ﻝﻮﻠﺳﻲﻠﺻﺍﻱﺎﻫﺺﺧﺎﺷ،Ins IIﻭIns Iﻱﺎﻫﻥژﺯﺭﺎﺑﻥﺎﻴﺑﺮﺗﻢﻬﻣﻪﻤﻫ

ﻩﻭﺮﮔﺭﺩﻪﻛﺩﺍﺩﻥﺎﺸﻧﺰﻴﻧ _ﻰّﻤﻛﻪﻤﻴﻧﻥﺎﻴﺑﻰﺳﺭﺮﺑﺞﻳﺎﺘﻧ .ﺖﺳﺎﺘﺑﻱﺎﻫ

Ins2, ﻯﺎﻫﻥژ ﻥﺎﻴﺑ (_p<0/05 ) ﻯﺭﺍﺩﻰﻨﻌﻣ ﺢﻄﺳ ﺭﺩ Mat1/30

ﺭﺩﻰﺘﺣ .ﺖﺳﺍﻪﺘﻓﺎﻳﺶﻳﺍﺰﻓﺍﺎﻫﻩﻭﺮﮔﻪﻴﻘﺑﻪﺑﺖﺒﺴﻧSlc2a2ﻭ Ins1

ﻰﺳﺍﺮﻜﻧﺎﭘﻯﺎﻔﻟﺁ ﻯﺎﻫﻝﻮﻠﺳﻪﺼﺧﺎﺷ ﻪﻛﺰﻴﻧ Glucagonﻥژ ﺩﺭﻮﻣ

.ﺩﻮﺑﺐﻴﺗﺮﺗﻦﻴﻤﻫﻪﺑﺖﻴﻌﺿﻭ،ﺖﺳﺍ

ﻪﺑﺖﺒﺴﻧMat1/30ﻩﻭﺮﮔﺭﺩﻥژﻥﺎﻴﺑﻰﺳﺭﺮﺑﺮﺘﻬﺑﺞﻳﺎﺘﻧﻞﻴﻟﺩﻪﺑ

ﻪﺑMat1/30ﻩﻭﺮﮔﺮﺘﺸﻴﺑﻯﺎﻫﻰﺳﺭﺮﺑﻭﺭﺎﻛﻪﻣﺍﺩﺍﻯﺍﺮﺑMat1/100 .ﺪﺷﻩﺩﺎﻔﺘﺳﺍﻝﮋﻳﺮﺗﺎﻣﻪﻨﻴﻣﺯﻩﺩﺎﻣﻩﺪﻨﻳﺎﻤﻧﻩﻭﺮﮔﻥﺍﻮﻨﻋ

ﻱﺩﺎﺑﻲﺘﻧﺁﻪﻠﻴﺳﻭﻪﺑﺎﻫﻝﻮﻠﺳﻲﻤﻴﺷﻮﺘﻴﺳﻮﻨﻤﻳﺍﻱﺰﻴﻣﺁﮓﻧﺭ

C-ﺪﻴﺘﭙﭘﻭﻦﻴﻟﻮﺴﻧﺍﻪﻴﻠﻋ

ﻪﺘﺴﻫﻩﺪﻨﻫﺩﻥﺎﺸﻧﻪﻛﺪﻧﺩﻮﺑﻩﺪﺷﻯﺰﻴﻣﺁﮓﻧﺭPIﺎﺑﺎﻫﻝﻮﻠﺳﻪﻴﻠﻛ

ﺭﺩ،(ﻪﺘﺴﻫﺖﻴﻌﻗﻮﻣ) ﺰﻣﺮﻗﮓﻧﺭﺮﺑﻩﻭﻼﻋﺖﺒﺜﻣﻱﺎﻫﻝﻮﻠﺳ .ﺩﻮﺑﺎﻬﻧﺁ

ﺖﻴﻌﻗﻮﻣﻩﺪﻨﻫﺩﻥﺎﺸﻧﻪﻛﺪﻧﺩﻮﺑﻪﺘﻓﺮﻳﺬﭘﺍﺭﺰﺒﺳﮓﻧﺭﺩﻮﺧﻢﺳﻼﭘﻮﺘﻴﺳ

.ﺩﻮﺑ (4A, B, Cﻞﻜﺷ) C-ﺪﻴﺘﭙﭘﻭﻦﻴﻟﻮﺴﻧﺍﻲﻤﺳﻼﭘﻮﺘﻴﺳ

ﻭ ﺮﻴﻴﻐﺗ C-ﺪﻴﺘﭙﭘ ﻯﺍﺮﺑﺖﺒﺜﻣ ﻯﺎﻫﻝﻮﻠﺳ ﺩﺍﺪﻌﺗﺵﺭﺎﻤﺷ ﺯﺍ ﺲﭘ

ﺭﺩﻪﻛﻯﺍﻪﻧﻮﮔﻪﺑﺪﺸﻧﻩﺪﻳﺩﻒﻠﺘﺨﻣﻯﺎﻫﻩﻭﺮﮔﻦﻴﺑﻯﺭﺍﺩﻰﻨﻌﻣﺶﻳﺍﺰﻓﺍ

ﻰﻠﭘ ﻩﻭﺮﮔﻭ 0/82_±0/04 ﻝﮋﻳﺮﺗﺎﻣﻩﻭﺮﮔ،0/66_±0/11 ﻦﻴﺗﻻژ ﻩﻭﺮﮔ

ﺖﺒﺜﻣC-ﺪﻴﺘﭙﭘﺎﻫﻝﻮﻠﺳﻞﻛﺪﺻﺭﺩ 0/73_±0/05 ﻦﻴﻨﻴﻣﻻ/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍ

ﻪﻛﺩﻮﺑﻦﻴﻨﻴﻣﻻ/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍﻰﻠﭘﻩﻭﺮﮔﺎﻬﻨﺗ،ﻦﻴﻟﻮﺴﻧﺍﺩﺭﻮﻣﺭﺩ .ﺪﻧﺩﻮﺑ

ﻯﺎﻫﻝﻮﻠﺳ ﺪﺻﺭﺩ 1/68_±0/12 ﺎﺑ (p<0/05) ﻯﺭﺍﺩﻰﻨﻌﻣ ﺕﺭﻮﺻ ﻪﺑ 1/25_±0/07 ﻦﻴﺗﻻژﻩﻭﺮﮔ .ﺩﻮﺑﺎﻫﻩﻭﺮﮔﺮﻳﺎﺳ ﺯﺍﺮﺘﺸﻴﺑﺖﺒﺜﻣ ﻦﻴﻟﻮﺴﻧﺍ

ﻪﺑ .ﺪﻨﺘﺷﺍﺩ ﺖﺒﺜﻣﻦﻴﻟﻮﺴﻧﺍ ﻯﺎﻫﻝﻮﻠﺳ ،ﺪﺻﺭﺩ 1/37_±0/04 ﻝﮋﻳﺮﺗﺎﻣﻭ 30000 ﺯﺍﺶﻴﺑ،ﻩﻭﺮﮔﺮﻫﻯﺍﺮﺑﺖﺒﺜﻣﻱﺎﻫﻝﻮﻠﺳﺪﺻﺭﺩﺵﺭﺎﻤﺷﺭﻮﻈﻨﻣ

ﺕﺭﻮﺻ ﻪﺑ ﺎﻫﻩﺩﺍﺩ) ﺪﺷ ﺵﺭﺎﻤﺷ ﻢﻫ ﺯﺍ ﻞﻘﺘﺴﻣ ﺭﺍﺮﻜﺗ ﻪﺳ ﺯﺍ ﻝﻮﻠﺳ

.(ﺪﻧﺍﻩﺪﺷﻪﺋﺍﺭﺍMean±SEM

DTZﻪﻠﻴﺳﻭﻪﺑﺖﺸﻛﺭﺩﻦﻴﻟﻮﺴﻧﺍﻱﻭﺎﺣﻱﺎﻫﻝﻮﻠﺳﻱﺰﻴﻣﺁﮓﻧﺭ

ﻦﻴﻟﻮﺴﻧﺍﻝﻮﻜﻟﻮﻣﺭﺩﺩﻮﺟﻮﻣﻱﻭﺭﻥﻮﻳﻪﺑ ﻪﻛﺖﺳﺍﻲﮕﻧﺭDTZ

ﺭﺩﺰﻣﺮﻗﮓﻧﺭﻪﺑﺍﺭﺎﻬﻧﺁﻭ (Chelate) ﺩﻮﺷﻰﻣﻞﺼﺘﻣﻝﻮﻠﺳﺭﺩﺩﻮﺟﻮﻣ

ﻱﺰﻴﻣﺁﮓﻧﺭDTZﺎﺑﺎﻫﻝﻮﻠﺳ،ﻲﻳﺎﻬﻧﺰﻳﺎﻤﺗﻪﻠﺣﺮﻣﻥﺎﻳﺎﭘﺭﺩ .(8) ﺩﺭﻭﺁﻲﻣ

ﻪﺑﻦﻴﻟﻮﺴﻧﺍﻱﻭﺎﺣﻱﺎﻫﻝﻮﻠﺳ،ﺖﺸﻛﺭﺩﺩﻮﺟﻮﻣﻱﺎﻫﻝﻮﻠﺳﻦﻴﺑﺭﺩ .ﺪﻧﺪﺷ

.(4Dﻞﻜﺷ) ﺪﻨﺘﻓﺮﮔﮓﻧﺭ،ﻲﺧﺮﺳﻲﻟﻮﻠﺳﺕﺎﻋﺎﻤﺘﺟﺍﺕﺭﻮﺻ

ﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗﻥﺍﺰﻴﻣﻲﺳﺭﺮﺑ

ﺭﺩ ﺎﺘﺑ ﻱﺎﻫﻝﻮﻠﺳ (Functional) ﻱﺩﺮﻜﻠﻤﻋ ﻲﮔﮋﻳﻭ ﻦﻳﺮﺘﻤﻬﻣ

ﻢﻴﻈﻨﺗﺢﺷﺮﺗ .ﺖﺳﺍﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗﻖﻳﺮﻃﺯﺍﻥﻮﺧﺪﻨﻗﻥﺍﺰﻴﻣﻢﻴﻈﻨﺗ،ﻥﺪﺑ

ﺭﺩ .ﺖﺳﺍﻡﺯﻻﺍﺬﻏﺯﺍﺪﻌﺑﻦﻴﻟﻮﺴﻧﺍﺩﺎﻳﺯﺮﻳﺩﺎﻘﻣﻱﺯﺎﺳﺩﺍﺯﺁﻱﺍﺮﺑﻩﺪﺷ

ﻱﺎﻫﻝﻮﻠﺳ ﻲﻳﺎﻧﺍﻮﺗ ﻲﺳﺭﺮﺑ ﺭﻮﻈﻨﻣ ﻪﺑ ﻱﺩﺎﻴﻨﺑ ﻱﺎﻫﻝﻮﻠﺳ ﺰﻳﺎﻤﺗ ﻥﺎﻳﺎﭘ

ﻲﺤﺷﺮﺗﻦﻴﻟﻮﺴﻧﺍﻥﺍﺰﻴﻣﺶﺠﻨﺳﻥﻮﻣﺯﺁ،ﺎﻫﻝﻮﻠﺳﻦﻳﺍﺯﺍﻪﺘﻓﺎﻳﺰﻳﺎﻤﺗﻱﺎﺘﺑﻪﺒﺷ ﺯﺍ،ﺰﻛﻮﻠﮔ (ﺭﻻﻮﻣﻰﻠﻴﻣ 22 ،ﺭﻻﻮﻣﻰﻠﻴﻣ 3) ﻒﻠﺘﺨﻣﺖﻈﻠﻏﻭﺩﻪﺑﺦﺳﺎﭘﺭﺩ .ﺪﻣﺁﻞﻤﻋﻪﺑﻲﻳﺎﻬﻧﺰﻳﺎﻤﺗﺯﺎﻓﻱﺎﻫﻝﻮﻠﺳ

/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍﻰﻠﭘ:C،ﻝﮋﻳﺮﺗﺎﻣ:B،ﻦﻴﺗﻻژﻱﻭﺭﺮﺑﺎﻫﻝﻮﻠﺳ:A؛C-ﺪﻴﺘﭙﭘﻪﻴﻠﻋﺮﺑﻱﺩﺎﺑﻲﺘﻧﺁﻪﻠﻴﺳﻭﻪﺑﺎﻫﻝﻮﻠﺳﻲﻤﻴﺷﻮﺘﻴﺳﻮﻨﻤﻳﺍﻱﺰﻴﻣﺁﮓﻧﺭ :4ﻞﻜﺷ

ﻦﻴﻟﻮﺴﻧﺍﻱﻭﺎﺣﻱﺎﻫﻝﻮﻠﺳﻯﺍﺭﺍﺩ،DTZﺎﺑﻪﺘﻓﺮﻳﺬﭘﮓﻧﺭﻲﻟﻮﻠﺳﻉﺎﻤﺘﺟﺍ:D .ﺪﻧﺍﻩﺪﺷﻩﺩﺍﺩﺖﺸﻛﻦﻴﻨﻴﻣﻻ

(8)

ﻲﺗﻭﺎﻔﺘﻣﺦﺳﺎﭘﻒﻠﺘﺨﻣﻱﺎﻫﺲﻜﻳﺮﺗﺎﻣﻱﻭﺭﻪﺘﻓﺎﻳﺰﻳﺎﻤﺗﻱﺎﻫﻝﻮﻠﺳ

ﻱﻭﺭﻩﺪﺷﺖﺸﻛﻱﺎﻫﻝﻮﻠﺳﻪﻛﻱﺭﻮﻃﻪﺑﺪﻧﺩﺍﺩﻥﺎﺸﻧﻂﻴﺤﻣﺰﻛﻮﻠﮔﻪﺑ

ﺖﺸﻛﻱﺎﻫﻝﻮﻠﺳﺎﺑﻪﺴﻳﺎﻘﻣﺭﺩﺰﻛﻮﻠﮔﺭﻻﻮﻣﻰﻠﻴﻣ 22 ﺖﻈﻠﻏﺭﺩ،ﻝﮋﻳﺮﺗﺎﻣ

ﺭﻻﻮﻣﻰﻠﻴﻣ 3 ﺖﻈﻠﻏﺭﺩﻭﻦﻴﻨﻴﻣﻻ/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍﻰﻠﭘﻭﻦﻴﺗﻻژﻱﻭﺭﻩﺪﺷ

/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍ ﻰﻠﭘﻱﻭﺭ ﻩﺪﺷ ﺖﺸﻛ ﻱﺎﻫﻝﻮﻠﺳ ﺎﺑ ﻪﺴﻳﺎﻘﻣ ﺭﺩ ﺰﻛﻮﻠﮔ

ﻥﺎﺸﻧﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗﻥﺍﺰﻴﻣﺭﺩ (_p≤0/05 ) ﻱﺭﺍﺩﻲﻨﻌﻣﺶﻳﺍﺰﻓﺍﻦﻴﻨﻴﻣﻻ .(1 ﺭﺍﺩﻮﻤﻧ) ﺪﻧﺩﺍﺩ

ﺦﺳﺎﭘﺭﺩﻒﻠﺘﺨﻣﻯﺎﻫﻩﻭﺮﮔﻦﻴﺑﺭﺩﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗﻥﺍﺰﻴﻣ:A :1ﺭﺍﺩﻮﻤﻧ

ﺰﻛﻮﻠﮔ (ﺭﻻﻮﻣﻰﻠﻴﻣ 22 ،ﺭﻻﻮﻣﻰﻠﻴﻣ 3) ﻒﻠﺘﺨﻣﺖﻈﻠﻏﻭﺩﻪﺑ .ﺪﻧﺍﻩﺪﺷﻪﺋﺍﺭﺍMean±SEMp< 0/05 ,ﺕﺭﻮﺻﻪﺑﺎﻫﻩﺩﺍﺩ *

ﺚﺤﺑ

ﻚﻳﻪﻄﺳﺍﻭﻪﺑ ﻞﻗﺍﺪﺣ،ﻒﻠﺘﺨﻣﻯﺎﻫﺖﻓﺎﺑﺭﺩﺎﻫﻝﻮﻠﺳﺯﺍﻯﺭﺎﻴﺴﺑ

ﺭﺎﺘﺧﺎﺳﻦﻴﻟﻭﺍﺎﺒﻳﺮﻘﺗﻭﺪﻨﺘﺴﻫﻁﺎﺒﺗﺭﺍﺭﺩﻰﻟﻮﻠﺳﻥﻭﺮﺑﻪﻨﻴﻣﺯﻩﺩﺎﻣﺎﺑﺢﻄﺳ ﻰﻣﺮﻈﻧﻪﺑﻭﺪﻧﺭﺍﺩﺶﻨﻛﻢﻫﺮﺑﺎﻬﻧﺁﺎﺑﻯﺩﺎﻴﻨﺑﻯﺎﻫﻝﻮﻠﺳﻪﻛﺖﺳﺍﻰﻟﻮﻜﻟﻮﻣ

.(16) ﺪﺷﺎﺑﺖﻴﻤﻫﺍﺰﺋﺎﺣﺎﻬﻧﺁﺰﻳﺎﻤﺗﻯﺰﻳﺭﻪﻣﺎﻧﺮﺑﻯﺍﺮﺑﻪﻛﺪﺳﺭ

ﻪﻃﺎﺣﺍﺍﺭﻰﺳﺍﺮﻜﻧﺎﭘﺮﻳﺍﺰﺟﻰﻟﻮﻠﺳﻥﻭﺮﺑﻪﻨﻴﻣﺯﻩﺩﺎﻣ،ﻰﻌﻴﺒﻃﺖﻟﺎﺣﺭﺩ

ﺯﺍﺍﺭﻰﻨﻳﺮﻛﻭﺪﻧﺍﻯﺎﻫﻝﻮﻠﺳﻪﻛﺪﻫﺩﻰﻣﻞﻴﻜﺸﺗﺍﺭﻯﺍﻪﻳﺎﭘﻯﺎﺸﻏﻭﺪﻨﻛﻰﻣ

ﻪﻳﺎﭘﻯﺎﺸﻏﻦﻳﺍﻰﻟﻮﻜﻟﻮﻣﻯﺍﺰﺟﺍﻪﭼﺮﮔﺍ .ﺪﻨﻛﻰﻣﺍﺪﺟﻰﺳﺍﺮﻜﻧﺎﭘﻯﺭﺎﺠﻣ

ﺪﻫﺩﻰﻣﻥﺎﺸﻧﻪﻛﺩﺭﺍﺩﺩﻮﺟﻭﻯﺪﻫﺍﻮﺷﺎﻣﺍ،ﻩﺪﺸﻧﻪﻌﻟﺎﻄﻣﻞﻣﺎﻛﺭﻮﻃﻪﺑﺯﻮﻨﻫ

ﺪﻧﻮﺷﻰﻣﺖﻓﺎﻳﻰﻟﺎﻴﻠﺘﻴﭘﺍﻪﻳﺎﭘﻯﺎﺸﻏﺭﺩﺎﻣﻮﻤﻋﻪﻛﻰﻳﺎﻫﻝﻮﻜﻟﻮﻣﺯﺍﻰﻀﻌﺑ

.(17) ﺪﻨﻨﻛﻰﻣﺎﻔﻳﺍﺶﻘﻧﺎﺘﺑﻯﺎﻫﻝﻮﻠﺳﺰﻳﺎﻤﺗﻭﺪﺷﺭﺭﺩ

،ﻝﮋﻳﺮﺗﺎﻣﻲﻟﻮﻠﺳﻥﻭﺮﺑﻪﻨﻴﻣﺯ ﻩﺩﺎﻣ ﻉﻮﻧﻪﺳﺮﻴﺛﺎﺗ،ﻪﻌﻟﺎﻄﻣﻦﻳﺍﺭﺩ

ﺯﺍﻦﻴﻟﻮﺴﻧﺍﺪﻟﻮﻣﻱﺎﻫﻝﻮﻠﺳﺰﻳﺎﻤﺗﺮﺑﻦﻴﻨﻴﻣﻻ/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍﻰﻠﭘﻭﻦﻴﺗﻻژ

ﻝﻮﻠﺤﻣﻲﻨﻴﺌﺗﻭﺮﭘﻩﺭﺎﺼﻋ،ﻝﮋﻳﺮﺗﺎﻣ .ﺪﺷﻰﺳﺭﺮﺑﺵﻮﻣﻲﻨﻴﻨﺟﻱﺎﻫﻪﺘﺧﺎﻳﻦﺑ

ﺵﻮﻣ ﻯﺎﻣﻮﻛﺭﺎﺳ ﻱﺎﻫﻝﻮﻠﺳ ﻪﻳﺎﭘ ﻱﺎﺸﻏ ﺯﺍ ﻩﺪﺷ ﻪﺘﻓﺮﮔ ﻞﻳﺮﺘﺳﺍ ﻭ

ﻥﺁﻰﻠﺻﺍﻯﺍﺰﺟﺍﻪﻛﺖﺳﺍEngelbreth-Holm-Swarm (EHS)

ﻥﺍﺭﺎﭙﻫﻯﺎﻫﻥﺎﻜﻴﻠﮔﻮﺌﺗﻭﺮﭘ،(ﺪﺻﺭﺩ 30) ﻥژﻼﻛ،(ﺪﺻﺭﺩ 60) ﻦﻴﻨﻴﻣﻻ

ﻯﻭﺎﺣﻦﻴﻨﭽﻤﻫ .ﺖﺳﺍ (ﺪﺻﺭﺩ 1) ﻥژﻭﺪﻴﻧﺎﻳﻦﻴﻛﺎﺘﻧﺍﻭ (ﺪﺻﺭﺩ 8) ﺕﺎﻔﻟﻮﺳ TGF-β, FGF-β, EGFﺪﻨﻧﺎﻣﺪﺷﺭﻯﺎﻫﺭﻮﺘﻛﺎﻓﺯﺍﻰﻔﻠﺘﺨﻣﻉﺍﻮﻧﺍ

ﻱﺪﻴﺳﺍﺵﺯﺍﺩﺮﭘﺭﺩﻥژﻼﻛﻩﺪﺷﺖﺷﺮﺳﺍﻭﻡﺮﻓ،ﻦﻴﺗﻻژ .(18) ﺖﺳﺍ ...ﻭ

ﻪﭼﺮﮔﺍﻭﺖﺳﺍ (Alkaline Process) ﻱﺯﺎﺑﺎﻳ (Acidic Process)

ﺎﻣﺍﺖﺳﺍﻥژﻼﻛﻢﻈﻨﻣﻩﺪﻴﻨﺗﻢﻫﺭﺩﻥﺎﻤﺘﺧﺎﺳﺪﻗﺎﻓﻥﺪﺷﻪﺘﺴﺑﻡﺎﮕﻨﻫﻪﺑ

ﻥﺎﺸﻧﻰﻠﺒﻗﺕﺎﻌﻟﺎﻄﻣ .ﺖﺳﺍﻥژﻼﻛﺎﺑﻪﺑﺎﺸﻣﻱﺍﻪﻨﻴﻣﺁﺪﻴﺳﺍﻲﻟﺍﻮﺗﻅﺎﺤﻟﺯﺍ

ﺭﻮﻀﺣﺭﺩﻪﺘﺒﻟﺍ،ﺵﻮﻣﻦﻴﻨﺟﺱﺍﺮﻜﻧﺎﭘﻰﻟﺎﻴﻠﺘﻴﭘﺍﻯﺎﻫﻝﻮﻠﺳﺖﺸﻛﻪﻛﻩﺩﺍﺩ

ﻰﻨﻳﺮﻛﻭﺪﻧﺍﻯﺎﻫﻝﻮﻠﺳﻪﺑ ﺎﻬﻧﺁ ﺰﻳﺎﻤﺗﺚﻋﺎﺑ 1- ﻥژﻼﻛﻯﻭﺭ ،ﻢﻴﺸﻧﺍﺰﻣ

ﻭﻮﻧﺎﺴﺘﻧﻮﻣ .(19) ﺪﻨﻫﺩﻰﻣﻞﻜﺷﺍﺭﻯﺪﻨﻧﺎﻣﻩﺮﻳﺰﺟﻯﺎﻫﺭﺎﺘﺧﺎﺳﻭﺩﻮﺷﻰﻣ

ﺱﺍﺮﻜﻧﺎﭘﻰﻨﻳﺮﻛﻭﺪﻧﺍﻯﺎﻫﻝﻮﻠﺳﺖﺸﻛﻪﻛﺪﻧﺍﻩﺩﺍﺩﻥﺎﺸﻧﺰﻴﻧﺶﻧﺍﺭﺎﻜﻤﻫ

ﺪﻨﻧﺎﻣﻩﺮﻳﺰﺟﻯﺎﻫﺭﺎﺘﺧﺎﺳﻯﺮﻴﮔﻞﻜﺷﺚﻋﺎﺑﻥژﻼﻛﺲﻜﻳﺮﺗﺎﻣﺭﺩﺕﺭ

ﻯﺎﻫﻝﻮﻜﻟﻮﻣﺯﺍﻯﺍﻪﻜﺒﺷ،ﺰﻴﻧﻦﻴﻨﻴﻣﻻ /ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍﻰﻠﭘ .(20) ﺖﺳﺍﻩﺪﺷ

ﻪﻳﺎﭘﻯﺎﺸﻏﻰﻠﺻﺍﻯﺍﺰﺟﺍﺯﺍﻦﻴﻨﻴﻣﻻ .ﺖﺳﺍﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍﻰﻠﭘﻭﻦﻴﻨﻴﻣﻻ

.(21) ﺖﺳﺍﻰﺳﺍﺮﻜﻧﺎﭘﺮﻳﺍﺰﺟ

ﻰﻤﻴﺷﻮﺘﻴﺳﻮﻧﻮﻤﻳﺍ_{،ﻰّﻤﻛ}ﻪﻤﻴﻧRT-PCRﺵﻭﺭﻪﺳﺯﺍﻰﺳﺭﺮﺑﻯﺍﺮﺑ

ﺐﻴﺗﺮﺗﻪﺑﺍﺭﻪﻌﻟﺎﻄﻣﻥﺎﻜﻣﺍﻪﻛﺪﺷﻩﺩﺎﻔﺘﺳﺍﻦﻴﻟﻮﺴﻧﺍﺢﺷﺮﺗﻥﺍﺰﻴﻣﺶﺠﻨﺳﻭ

ﻭﻰﺻﺎﺼﺘﺧﺍﻯﺎﻫﻥژ (ﻪﻤﺟﺮﺗ) ﻦﻴﺌﺗﻭﺮﭘ،(ﻰﺴﻳﻮﻧﻭﺭ) RNAﺢﻄﺳﻪﺳﺭﺩ .ﺩﺮﻛﻰﻣﻢﻫﺍﺮﻓﻪﺘﻓﺎﻳﺰﻳﺎﻤﺗﻯﺎﻫﻝﻮﻠﺳﺩﺮﻜﻠﻤﻋ

ﻥﺎﻴﺑ،ﺖﻓﺮﮔﺭﺍﺮﻗﻲﻠﺻﺍﺭﺎﻴﻌﻣﻲﻧژﻥﺎﻴﺑﺰﻴﻟﺎﻧﺁﺞﻳﺎﺘﻧﺮﻴﺴﻔﺗﺭﺩﻪﭽﻧﺁ ﻱﺯﺎﺘﺳﻮﺋﻮﻤﻫﻱﺭﺍﺮﻗﺮﺑﺭﺩﻢﻬﻣﻲﻧﻮﻣﺭﻮﻫﻦﻴﻟﻮﺴﻧﺍ .ﺩﻮﺑﻦﻴﻟﻮﺴﻧﺍﻱﺪﻴﻠﻛﻥژ

ﺢﺷﺮﺗ ﻭ ﻩﺮﻴﺧﺫ ،ﺰﺘﻨﺳﻲﺳﺍﺮﻜﻧﺎﭘ ﺮﻳﺍﺰﺟ ﻱﺎﺘﺑ ﻱﺎﻫﻝﻮﻠﺳ ﺭﺩ ﻪﻛ ﺖﺳﺍ

ﻦﻴﻟﻮﺴﻧﺍﻭﺮﭘﻩﺮﭘ ،ﺯﺎﺳﺶﻴﭘ ﻚﻳ ﺕﺭﻮﺻ ﻪﺑ ﻦﻴﻟﻮﺴﻧﺍ ﻦﻴﺌﺗﻭﺮﭘ .ﺩﻮﺷﻲﻣ

ﺖﺳﺍ ﻩﺪﺷ ﻆﻔﺣ ﺭﺎﻴﺴﺑ ﻱﺭﻮﻧﺎﺟ ﻱﺎﻫﻪﻧﻮﮔ ﻦﻴﺑ ﺭﺩ ﻪﻛ ﺩﻮﺷﻲﻣ ﺰﺘﻨﺳ

،ﺕﺭﻭﺵﻮﻣﺭﺩ،ﻥﺍﺭﺍﺪﻧﺎﺘﺴﭘﺐﻠﻏﺍﻑﻼﺧﺮﺑ .(Highly Conserved)

ﺰﻣﺭﺍﺭ 2 ﻭ 1 ﻱﺎﻫﻦﻴﻟﻮﺴﻧﺍﻭﺮﭘﻪﻛﻞﻟﺁﺮﻴﻏﻲﻟﻭﻪﺑﺎﺸﻣﻥژﻭﺩﺯﺍﻦﻴﻟﻮﺴﻧﺍ

،ﻦﻴﻟﻮﺴﻧﺍﻭﺮﭘﻥژﻭﺩﻪﻛﺖﺳﺍﻩﺩﺍﺩﻥﺎﺸﻧﺕﺎﻌﻟﺎﻄﻣ .ﺪﻳﺁﻲﻣﺩﻮﺟﻭﻪﺑﺪﻨﻨﻛﻲﻣ

ﺭﺩﺎﻬﻨﺗIns Iﻥژ .ﺪﻧﻮﺷﻲﻣﻢﻴﻈﻨﺗﻞﻘﺘﺴﻣﺕﺭﻮﺻﻪﺑ،ﻱﺍﻩﺯﺍﺪﻧﺍﺎﺗﻞﻗﺍﻻ

ﻝﻮﻠﺳﺭﻮﻀﺣﺪﻳﻮﻣﻥﺁﻥﺎﻴﺑﻭﺩﻮﺷﻲﻣﻥﺎﻴﺑﺵﻮﻣﻲﺳﺍﺮﻜﻧﺎﭘﻱﺎﺘﺑﻱﺎﻫﻝﻮﻠﺳ

ﺭﺩﻢﻫﻭﺩﻮﺷﻲﻣﻥﺎﻴﺑﺎﺘﺑﻝﻮﻠﺳﺭﺩﻢﻫIns IIﻥژ .ﺖﺳﺍﺖﺸﻛﺭﺩﺎﺘﺑﻪﺒﺷ

ﻪﻜﻧﺁﺩﻮﺟﻭﺎﺑﻭﺭﻦﻳﺍﺯﺍ .(22-24) ﻲﺳﺍﺮﻜﻧﺎﭘﺮﻴﻏﻱﺎﻫﻝﻮﻠﺳﺯﺍﻯﺩﺍﺪﻌﺗ

ﺍﺭﻥﺁﻥﺎﻴﺑﻝﺎﺣﻦﻳﺍﺎﺑﺖﺳﺍﺪﻨﻤﺷﺯﺭﺍﻥژﻦﻳﺍﻥﺎﻴﺑﻡﺪﻋﺎﻳﻥﺎﻴﺑﻲﺳﺭﺮﺑ

.ﺩﺍﺩﺖﺒﺴﻧﺎﺘﺑﻱﺎﻫﻝﻮﻠﺳﻪﺑﻖﻠﻄﻣﺭﻮﻃﻪﺑﻥﺍﻮﺗﻲﻤﻧ

ﺭﻮﻀﺣ ﺮﮕﻧﺎﻴﺑ ﺪﻣﺁ ﺖﺳﺩ ﻪﺑ RT-PCR ﻯﺎﻫﻩﺩﺍﺩ ﺯﺍ ﻪﻛ ﻪﭽﻧﺁ

ﻩﻭﺮﮔﺭﺩ .ﺩﻮﺑ ﻝﮋﻳﺮﺗﺎﻣﻯﺎﻫﻩﻭﺮﮔﺭﺩﺎﻬﻨﺗ 1 ﻦﻴﻟﻮﺴﻧﺍ ﻥژﻯﺎﻫﺖﺷﻮﻧﻭﺭ Glucagonﻯﺎﻫﻥژﻥﺎﻴﺑ (p<0/05) ﻯﺭﺍﺩﻰﻨﻌﻣﺢﻄﺳﺭﺩ،1/30 ﻝﮋﻳﺮﺗﺎﻣ

ﺯﺍ .ﺩﻮﺑﻪﺘﻓﺎﻳﺶﻳﺍﺰﻓﺍﺎﻫﻩﻭﺮﮔﻪﻴﻘﺑﻪﺑﺖﺒﺴﻧIns1،Slc2a2،Ins2ﻭ

ﺖﻴﻌﺿﻭﺭﺩﻢﻫﺪﺻﺭﺩ1 ﻝﮋﻳﺮﺗﺎﻣﻩﻭﺮﮔﻪﺑﺖﺒﺴﻧ 1/30 ﻝﮋﻳﺮﺗﺎﻣﻩﻭﺮﮔﻰﻓﺮﻃ

ﻯﺍﺮﺑﻝﮋﻳﺮﺗﺎﻣﺯﺍﺖﻈﻠﻏﻦﻳﺍﻥﺩﻮﺑﺐﺳﺎﻨﻣﻩﺪﻨﻫﺩﻥﺎﺸﻧﻪﻛﺖﺷﺍﺩﺭﺍﺮﻗﻯﺮﺘﻬﺑ

. ﺩﻮﺑﺖﺸﻛﻑﻭﺮﻇﻒﻛﻥﺪﻧﺎﺷﻮﭘ

ﻦﻴﺌﺗﻭﺮﭘﺩﻮﺟﻭﻥﺩﺍﺩﻥﺎﺸﻧﻯﺍﺮﺑﻪﻛﻰﻤﻴﺷﻮﺘﻴﺳﻮﻧﻮﻤﻳﺍﻯﺎﻫﻰﺳﺭﺮﺑﺭﺩ

/ﻦﻴﺘﻴﻧﺭﻭﺍ-ﻝﺍ ﻰﻠﭘﻩﻭﺮﮔ ﺪﺷﻡﺎﺠﻧﺍ ﻪﺘﻓﺎﻳ ﺰﻳﺎﻤﺗ ﻯﺎﻫﻝﻮﻠﺳ ﺭﺩ ﻦﻴﻟﻮﺴﻧﺍ

ﻪﺘﻜﻧﻦﻳﺍ ﺎﻣﺍ .ﺩﺍﺩﻰﻣﻥﺎﺸﻧ ﺍﺭﺖﺒﺜﻣ ﻯﺎﻫﻝﻮﻠﺳ ﻥﺍﺰﻴﻣ ﻦﻳﺮﺘﺸﻴﺑﻦﻴﻨﻴﻣﻻ

ﺰﻳﺎﻤﺗ ﺭﺩ ﻝﻭﺍﺪﺘﻣﻩﺩﺎﻔﺘﺳﺍ ﺩﺭﻮﻣﺖﺸﻛﻂﻴﺤﻣ ﻪﻛﺩﻮﺷﺵﻮﻣﺍﺮﻓﺪﻳﺎﺒﻧ

ﻲﻳﻻﺎﺑﺖﻈﻠﻏﺩﻮﺧﺭﺩﻪﻛﺖﺳﺍN2ﻲﻧﺩﻭﺰﻓﺍﻱﻭﺎﺣ،ﻱﺩﺎﻴﻨﺑﻱﺎﻫﻝﻮﻠﺳ

ﻭ (ﺎﻫﺵﻭﺭﻭﺩﺍﻮﻣ .ﻙ.ﺭ) ﺩﺭﺍﺩﻦﻴﻟﻮﺴﻧﺍﺯﺍ (ﺮﺘﻴﻟﻰﻠﻴﻣﺮﺑﻡﺮﮔﻰﻠﻴﻣ 500)

ﻂﺳﻮﺗﺖﺸﻛﻂﻴﺤﻣﺯﺍﻦﻴﻟﻮﺴﻧﺍ (Uptake) ﺏﺬﺟﻝﺎﻤﺘﺣﺍﻭﺭﻦﻳﺍﺯﺍ

.ﺩﺯﺎﺳﻲﻣﻪﺟﺍﻮﻣﺖﻳﺩﻭﺪﺤﻣﺎﺑ ﺍﺭﺞﻳﺎﺘﻧﺮﻴﺴﻔﺗﻪﻛﺩﺭﺍﺩﺩﻮﺟﻭﺎﻫﻝﻮﻠﺳ

ﻩﺩﺎﻔﺘﺳﺍﻰﻳﺎﻫﻯﺩﺎﺑﻰﺘﻧﺁﺯﺍﻥﺍﻮﺗﻲﻣﺖﻳﺩﻭﺪﺤﻣﻦﻳﺍﻦﺘﺧﺎﺳﻊﻔﺗﺮﻣ ﻱﺍﺮﺑ

ﻦﻴﻟﻮﺴﻧﺍﻪﻜﻧﺁ ﺎﻳ ﻭﺪﻨﻨﻛﻲﻣﻲﻳﺎﺳﺎﻨﺷﺍﺭ C-ﺪﻴﺘﭙﭘﺎﺻﺎﺼﺘﺧﺍﻪﻛﺩﺮﻛ

ﻝﻮﻠﺳﺩﻮﺧﻪﻛﻲﻨﻴﻟﻮﺴﻧﺍﺯﺍﺎﺗﺩﺮﻛﺭﺍﺪﻧﺎﺸﻧﺍﺭﺖﺸﻛﺭﺩﻩﺩﺎﻔﺘﺳﺍﺩﺭﻮﻣ

ﻥﺩﺮﻛ ﺰﻳﺎﻤﺘﻣ ﻯﺍﺮﺑ ،ﻪﻌﻟﺎﻄﻣ ﻦﻳﺍ ﺭﺩ .ﺪﺷﺎﺑ ﺺﻴﺨﺸﺗ ﻞﺑﺎﻗ ﺩﺯﺎﺳﻲﻣ

ﺪﻴﺘﭙﭘﺪﺿﻯﺎﻫﻯﺩﺎﺑﻰﺘﻧﺁﺯﺍﻰﺑﺬﺟﻯﺎﻫﻦﻴﻟﻮﺴﻧﺍﺯﺍﻰﺤﺷﺮﺗﻯﺎﻫﻦﻴﻟﻮﺴﻧﺍ

ﺎﻫﻩﻭﺮﮔﻪﻤﻫ،ﺖﺒﺜﻣC-ﺪﻴﺘﭙﭘﻯﺎﻫﻝﻮﻠﺳﺵﺭﺎﻤﺷﺎﺑ .ﺪﺷﻩﺩﺎﻔﺘﺳﺍﺰﻴﻧ

C-ﻯﺎﻫﻩﻭﺮﮔ ﻦﻴﺑ ﻯﺭﺍﺩﻰﻨﻌﻣﺕﻭﺎﻔﺗ ﺎﻣﺍ ﺪﻨﺘﺷﺍﺩ ﻪﺘﻓﺮﮔ ﮓﻧﺭﻯﺎﻫﻝﻮﻠﺳ

ﺮﮕﻧﺎﻴﺑﺰﻴﻧDTZﺎﺑﻯﺰﻴﻣﺁﮓﻧﺭﺯﺍﻞﺻﺎﺣﻯﺎﻫﻩﺩﺍﺩ .ﺪﺸﻧﻩﺪﻳﺩﻒﻠﺘﺨﻣ

.(4Dﻞﻜﺷ) ﺩﻮﺑﻩﻭﺮﮔﻪﺳﺮﻫﺭﺩﻪﺘﻓﺮﮔﮓﻧﺭﺕﺎﻌﻤﺠﺗﺭﻮﻀﺣ

ﻪﻛ ﺩﺍﺩﻥﺎﺸﻧ ﺰﻴﻧ ﻦﻴﻟﻮﺴﻧﺍ ﺢﺷﺮﺗﻥﺍﺰﻴﻣ ﻲﺳﺭﺮﺑ ﺯﺍﻞﺻﺎﺣ ﺞﻳﺎﺘﻧ

ﺦﺳﺎﭘﺭﺩ،ﺎﻫﻩﻭﺮﮔﻪﻴﻘﺑﻪﺑﺖﺒﺴﻧﻝﮋﻳﺮﺗﺎﻣﻯﻭﺭﻩﺪﺷﺖﺸﻛﻯﺎﻫﻝﻮﻠﺳ

،ﺭﺎﻈﺘﻧﺍ ﻑﻼﺧﺮﺑ ﻪﺘﺒﻟﺍ .ﺪﻧﺍﻩﺩﺮﻛ ﺢﺷﺮﺗ ﻯﺮﺘﺸﻴﺑ ﻦﻴﻟﻮﺴﻧﺍ ،ﺰﻛﻮﻠﮔ ﻪﺑ

ﺯﺍ ﺮﺘﺸﻴﺑ ﺰﻛﻮﻠﮔ ﺮﺘﻣﻰﻠﻴﻣ 22 ﺖﻈﻠﻏ ﺭﺩ ﻰﺤﺷﺮﺗ ﻦﻴﻟﻮﺴﻧﺍ ﺭﺍﺪﻘﻣ

ﻪﺘﻓﺎﻳﺰﻳﺎﻤﺗﻯﺎﻫﻝﻮﻠﺳﻪﻛﺖﺳﺍﻦﻳﺍﺮﮕﻧﺎﻴﺑﺎﻳﻪﻠﺌﺴﻣﻦﻳﺍﻪﻛﺩﻮﺒﻧﺮﺘﻣﻰﻠﻴﻣ 3

ﻒﻠﺘﺨﻣﻯﺎﻫﺖﻈﻠﻏﺎﺑﺐﺳﺎﻨﺘﻣﺪﻨﻧﺍﻮﺗﻰﻤﻧﻭﺪﻨﺘﺴﻴﻧﻰﻌﻴﺒﻃﺩﺮﻜﻠﻤﻋﻯﺍﺭﺍﺩ

*

* *

18 16 14 12 10 8 6 4 2 0

(

ﺮﺘﻴﻟ

ﺮﺑ

ﻡﺮﮔﻭﺮﻜﻴﻣ

)

ﻦﻴﻟﻮﺴﻧﺍ

ﺖﻈﻠﻏ

ﺭﻻﻮﻣﻰﻠﻴﻣ 3 ﺭﻻﻮﻣﻰﻠﻴﻣ 22 poly-l ornithin/laminin Gelatin