چکیده:
ﻣﻘﺪﻣﻪ : ﻣﺮگ ﺑﺮﻧﺎﻣﻪ رﯾﺰی ﺷﺪه ﺳﻠﻮل ﯾﺎ ﻫﻤﺎن آﭘﻮﭘﺘﻮز ﯾﮑﯽ از ﻣﻬﻤﺘﺮﯾﻦ ﻓﺮآﯾﻨﺪﻫﺎی ﻓﯿﺰﯾﻮﻟﻮژﯾـﮑـﯽ در ﺗﮑﻮﯾﻦ و ﺣﻔﻆ ﻫﻤﻮﺳﺘﺎزی ﻣﻮﺟﻮدات ﭼﻨﺪﺳﻠﻮﻟﯽ ﻣﯽ ﺑﺎﺷﺪ . ﻫﺮ ﮔﻮﻧﻪ اﺧﺘﻼﻟﯽ در ﺗﻨﻈﯿﻢ آﭘﻮﭘﺘﻮز ﻣﻨﺠـﺮ ﺑـﻪ اﯾﺠﺎد اﺧﺘﻼﻻت ﮔﺴﺘﺮده ژﻧﺘﯿﮑﯽ، ﺑﯿﻤﺎریﻫﺎی ﺧﻮد اﯾﻤﻨﯽ و ﺳﺮﻃﺎن ﻣﯽ ﺷﻮد . ﻓﺮآﯾﻨﺪ آﭘﻮﭘﺘﻮز ﻫﻢ در ﺑﺤﺚ ﺳﻼﻣﺖ و ﻫﻢ در ﺑﯿﻤﺎریﻫﺎ ﺿﺮوری ﻣﯽ ﺑﺎﺷﺪ . اﺻﻄﻼح آﭘﻮﭘﺘﻮز ﺑﻪ ﺗﻐﯿﯿﺮات ﻣﻮرﻓﻮﻟﻮژﯾﮑﯽ ﯾﮏ ﺳﻠﻮل ﮐـﻪ درﮔﯿﺮ ﻣﺮگ ﺑﺮﻧﺎﻣﻪ رﯾﺰی ﺷﺪه ﺑﺎﺷﺪ اﻃﻼق ﻣﯽ ﮔﺮدد ﮐﻪ اﯾﻦ ﺗﻐﯿﯿﺮات ﻣﻌﻤﻮﻻ ﺑﻪ ﺻﻮرت ﭼﺮوﮐﯿﺪﮔﯽ ﺳﻠﻮل، ﻣﺘﺮاﮐﻢ ﺷﺪن ﻫﺴﺘﻪ، ﺑﺮآﻣﺪﮔﯽ ﻫﺎی ﺳﻄﺢ ﻏﺸﺎء، ﻗﻄﻌﻪ ﻗﻄﻌﻪ ﺷﺪن ﻏﺸﺎء ﭘﻼﺳﻤﺎﺋﯽ و ﺗﺒﺪﯾﻞ ﺷﺪن آنﻫـﺎ ﺑﻪ اﺟﺴﺎﻣﯽ ﺑﻪ ﻧﺎم اﺟﺴﺎم آﭘﻮﭘﺘﻮزی و ﺗﻐﯿﯿﺮات ﺳﻄﺢ ﻏﺸﺎء ﮐﻪ در ﻧﻬﺎﯾﺖ ﺑﻪ ﻓﺎﮔﻮﺳﯿﺘـﻮز ﺷـﺪن ﺳـﻠـﻮ آﭘﻮﭘﺘﻮزی ﻣﻨﺠﺮ ﻣﯽ ﺷﻮد. ﻣﻮاد و روشﻫﺎ : ﭘﺲ از اﯾﺠﺎد ﺟﻬﺶ ﻫﺪف دار در ژن 1-Apaf ﺑـﻪ ﮐـﻤـﮏ PCR، ﺳـﻠـﻮل ﻫـﺎی HEK293T ﺑﻪ ﮐﻤﮏ ﻟﯿﭙﻮﻓﮑﺘﺎﻣﯿﻦ ﺗﺮاﻧﺴﻔﮑﺖ ﺷﺪﻧﺪ . 24 ﺳﺎﻋﺖ ﺑﻌﺪ از ﺗﺮاﻧﺴﻔﮑﺸﻦ ﺳﻠﻮلﻫﺎ در ﺣﻀﻮر دوﮐﺴﻮروﺑﯿﺴﯿﻦ ﻗﺮار ﮔﺮﻓﺘﻨﺪ و ﻓﻌﺎﻟﯿﺖ ﮐﺎﺳﭙﺎز 3 و 7 آنﻫﺎ در زﻣﺎنﻫﺎی ﻣﺨﺘﻠﻒ ﻣﻮرد ﺳـﻨـﺠـﺶ ﻗـﺮار ﮔﺮﻓﺖ. ﯾﺎﻓﺘﻪﻫﺎ: ﻓ ﻌﺎﻟﯿ ﺖ ﮐﺎ ﺳﭙﺎز 3 و 7 ﺑﻪ ﺧﻮﺑﯽ در ﺳﻠﻮل ﻫﺎی ﺗﺮاﻧﺴﻔﮑﺖ ﺷﺪه ﻣﺸﺎﻫﺪه ﺷﺪ و اﯾﻦ ﻓﻌﺎﻟـﯿـﺖ ﺣﺪود 3 ﺑﺮاﺑﺮ ﺳﻠﻮل ﻫﺎی ﮐﻨﺘﺮل ﺑﻮد ﮐﻪ اﯾﻦ داده ﺑﻪ ﺧﻮﺑﯽ ﻧﺸﺎن دﻫﻨﺪه اﻓﺰاﯾﺶ ﻓﻌﺎﻟﯿﺖ ﮐﺎﺳﭙﺎز 3 و 7 در ﺳﻠﻮل ﻫﺎیHEK293T اﺳﺖ . ﺑﺎ ﺗﻮﺟﻪ ﺑﻪ ﻣﺸﺎﻫﺪه ﺻﻮرت ﮔﺮﻓﺘﻪ اﺣﺘﻤﺎﻻ دوﻣﯿﻦ اﻧﺘﻬﺎﯾـﯽ ﭘـﺮوﺗـﺌـﯿـﻦ 1-Apaf ﻧﻘﺶ ﮐﻨﺘﺮﻟﯽ ﺑﺮ روی آزادﺳﺎزی ﮐﺎﺳﭙﺎز دارد. ﻧﺘﯿﺠﻪ ﮔﯿﺮی : ﻧﺘﺎﯾﺞ اﯾﻦ ﭘﮋوﻫﺶ ﻧﺸﺎن داد دوﻣﯿﻦ اﻧﺘﻬﺎﯾﯽ ﭘﺮوﺗﺌﯿﻦ 1-Apaf ﻓﻌﺎﻟﯿﺖ ﮐﺎﺳﭙﺎزﻫﺎی ﺳﻠﻮ را ﮐﻨﺘﺮل ﻣﯽ ﮐﻨﺪ و اﻣﮑﺎن اﺳﺘﻔﺎده از آن ﺑﻪ ﻋﻨﻮان ﯾﮏ ﺑﯿﻮﺳﻨﺴﻮر ﺑﺮای ﻏﺮﺑﺎﻟﮕﺮی داروﻫﺎی ﺿﺪ ﺳﺮﻃﺎن وﺟﻮد دارد.
Background: Programmed cell death or apoptosis is one of the most important physiological processes involved in development and homeostasis maintenance of multicellular organisms. Every defect in apoptosis regulation leads to wide variety of genetic disorders, auto-immune diseases and cancers. Apoptosis is an essential process in both health and diseases. Apoptosis is defined as morphological changes in a cell undergoing programmed cell death. These changes lead to cell shrinkage, nuclear condensation, membrane intrusion, plasma membrane fractionation and their conversion to apoptotic bodies and changes in cell surface membrane which cause phagocy-tosis of the cell bearing apoptosis.
Materials and Methods: Apaf-1 gene is mutated using PCR and then transfected into HEK293T cells by lipofectamin. 24 h transfected cells exposed to doxorubicin and caspase 3/7 activity was measured over different times.
Result: Caspase 3/7 activity was observed obviousely in transfected cells, which was 3 fold greater than those of controls. This data demonstrates the increase in caspase 3/7 activity in HEK293T cells. Based on our observation, the terminus domain in Apaf-1 is probably play a role in releasing the caspases.
Conclusion: The results of this study indicated that terminal do-main of Apaf-1 has pivotal role in caspase regulation and it is possi-ble to design a biosensor for screening the anticancer drugs by re-moving terminal domain of this protein.