ﺗﺎﺛﯿﺮ ورﺗﻤﺎﻧﯿﻦ ﺑﺮ ﺑﯿﺎن ژن ﻫﺎي ﺳﯿﺘﻮﮐﯿﻦ ﻫﺎي ﻣﺤﺎﻓﻈﺖ ﮐﻨﻨﺪه ﻧﻮروﻧﯽ در آﺳﺘﺮوﺳﯿﺖ ﻫﺎي ﮐﺸﺖ ﺷﺪه در ﺷﺮاﯾﻂ آزﻣﺎﯾﺸﮕﺎﻫﯽ

Effect of wortmanin on the expression of BDNF and NT3 neuroprotective cytokines in cultured Astrocyt

ﺑﺮرﺳﯽ ﮔﯿﺎﻫﺎن زراﻋﯽ در ﺷﺮاﯾﻂ ﺗﻨﺶ ﻣﯽﺗﻮاﻧﺪ ﻣﻨﺠﺮ ﺑﻪ ﺑﻬﺒﻮد ﺻﻔﺎت ﻣﺮﺗﺒﻂ ﺑﺎ ﺗﺤﻤﻞ و اﻓﺰاﯾﺶ رﺷﺪ و ﻋﻤﻠﮑﺮد آﻧﻬﺎ در ﺷﺮاﯾﻂ ﮐﻢآﺑﯽ ﺷﻮد. ﺑﻨﺎﺑﺮاﯾﻦ ﻧﺤﻮه و ﻧﻮع روش آﺑﯿﺎري ﺑﺮ ﮔﯿﺎﻫﺎن اﻫﻤﯿﺖ دارد. ﺑﻪ ﻣﻨﻈﻮر ﺑﺮرﺳﯽ ﺗﺄﺛﯿﺮ ﻣﻘﺎدﯾﺮ ﻣﺨﺘﻠﻒ آﺑﯿﺎري ﺑﺮ ﮐﯿﻔﯿﺖ اﻟﯿﺎف و ﻋﻤﻠﮑﺮد ﭘﻨﺒﻪ رﻗﻢ ﮔﻠﺴﺘﺎن ﺑﺎ ﻫﺪف ﺗﮑﺜﯿﺮ ﺑﺬر ﺑﺮاي ﻣﺰارع دﯾﻢ، آزﻣﺎﯾﺸﯽ ﺑﺮ اﺳﺎس ﺗﺎﻣﯿﻦ ﻧﯿﺎز رﻃﻮﺑﺘﯽ ﺧﺎك در 4 ﺳﻄﺢ )دﯾﻢ )ﺑﺪون آﺑﯿﺎري(، 33، 66 و100 درﺻﺪ( ﺑﻪ ﺻﻮرت اﺳﭙﻠﯿﺖ ﭘﻼت در ﻗﺎﻟﺐ ﻃﺮح ﺑﻠﻮك ﮐﺎﻣﻞ ﺗﺼﺎدﻓﯽ در ﺳﻪ ﺗﮑﺮار ﻃﯽ ﺳﺎل ﻫﺎي زراﻋﯽ 1395-98 اﺟﺮا ﺷﺪ. ﭘﺲ از رﺳﯿﺪﮔﯽ ﻓﯿﺰﯾﻮﻟﻮژﯾﮑﯽ، ﻋﻤﻠﮑﺮد و ﺻﻔﺎت ﮐﯿﻔﯽ اﻟﯿﺎف ارزﯾﺎﺑﯽ ﺷﺪﻧﺪ. در اﯾﻦ ﭘﮋوﻫﺶ وﯾﮋﮔﯽ ﻫﺎي ﭘﺮوﺗﺌﯿﻨﯽ و ﻓﯿﻠﻮژﻧﺘﯿﮑﯽ آﻧﺰﯾﻢ CesA) cellulose synthase A( و endotransglucosylase (XET1) ﺗﻮﺳﻂ اﺑﺰارﻫﺎي ﺑﯿﻮاﻧﻔﻮرﻣﺎﺗﯿﮏ ﻣﻮرد ارزﯾﺎﺑﯽ ﻗﺮارﮔﺮﻓﺖ. ﻧﺘﺎﯾﺞ ﻧﺸﺎن داد ﮐﻪ ﺑﯿﺸﺘﺮﯾﻦ ﻋﻤﻠﮑﺮد ﭼﯿﻦ اول، در ﺗﯿﻤﺎر آﺑﯿﺎري 66 درﺻﺪ ﻣﺸﺎﻫﺪه ﺷﺪ. ﺑﯿﺸﺘﺮﯾﻦ ﻣﻘﺪار وزن اﻟﯿﺎف ﭼﯿﻦ اول در ﺗﯿﻤﺎر آﺑﯽ 66 درﺻﺪ و ﮐﻤﺘﺮﯾﻦ ﻣﻘﺪار در ﺷﺮاﯾﻂ دﯾﻢ ﻣﺸﺎﻫﺪه ﺷﺪ. ﺑﺬرﻫﺎﯾﯽ ﮐﻪ در ﺷﺮاﯾﻂ 66 درﺻﺪ آﺑﯿﺎري رﺷﺪ ﮐﺮدﻧﺪ، ﺑﺎﻋﺚ ﺗﻮﻟﯿﺪ ﺑﺬر ﺑﺎﻟﻘﻮه ﺑﺎ اﻟﯿﺎف ﺑﺎﮐﯿﻔﯿﺖ ﺷﺪﻧﺪ. ﻫﻤﭽﻨﯿﻦ آﻧﺎﻟﯿﺰ ﺑﯿﻮاﻧﻔﻮرﻣﺎﺗﯿﮑﯽ ﻣﺸﺨﺺ ﮐﺮد ﮐﻪ ﻣﺤﻞ درونﺳﻠﻮﻟﯽ آﻧﺰﯾﻢﻫﺎي CesA و XET1 ﺑﻪ ﺗﺮﺗﯿﺐ ﻏﺸﺎي ﭘﻼﺳﻤﺎﯾﯽ و دﯾﻮاره ﺳﻠﻮﻟﯽ ﻣﯽﺑﺎﺷﻨﺪ. آﻧﺰﯾﻢ CesA و XET1 ﺑﻪ ﺗﺮﺗﯿﺐ از ﺧﺎﻧﻮادهي ﭘﺮوﺗﺌﯿﻨﯽ ﺗﺮﻧﺴﻔﺮازﻫﺎ و ﻫﯿﺪروﻻزﻫﺎ ﭘﯿﺸﺒﯿﻨﯽ ﺷﺪ. در ﺑﺮرﺳﯽ درﺧﺖ ﻓﯿﻠﻮژﻧﺘﯿﮑﯽ ﻣﺸﺨﺺ ﺷﺪ ﮐﻪ ﺗﻮاﻟﯽ ﻫﺮ دو آﻧﺰﯾﻢ در ﮔﯿﺎه ﭘﻨﺒﻪ ﺑﻪ ﻫﻤﺮاه ﮔﯿﺎﻫﺎن ﻫﻢﺧﺎﻧﻮاده، در ﺷﺎﺧﻪﻫﺎي ﯾﮏ ﮐﻼد ﻗﺮار ﮔﺮﻓﺘﻨﺪ. ﺑﻪ ﻃﻮر ﮐﻠﯽ، ﺗﮑﺜﯿﺮ ﺑﺬر ﺑﺎ ﺗﺎﻣﯿﻦ 66 درﺻﺪ ﻧﯿﺎز آﺑﯽ، ﺿﻤﻦ دﺳﺘﯿﺎﺑﯽ ﺑﻪ ﻋﻤﻠﮑﺮد ﻣﻄﻠﻮب ﻣﯽ ﺗﻮاﻧﺪ ﺑﺎ اﻟﻘﺎ ﻣﮑﺎﻧﯿﺴﻢ ﻫﺎي ﺗﺤﻤﻞ ﻧﺴﺒﺖ ﺑﻪ ﺗﻨﺶ ﮐﻢ آﺑﯽ ﭘﺎﯾﺪارﺗﺮ ﺑﺎش زﻣﯿﻨﻪ و ﻫﺪف: ﺑﯿﻤﺎري ﻫﺎي ﻧﻮرودژﻧﺮاﺗﯿﻮ ﺑﺎ ﺗﺎﺛﯿﺮ ﺑﺮ روي ﺳﯿﺴﺘﻢ ﻋﺼﺒﯽ ﻣﺮﮐﺰي ﺑﺎﻋﺚ اﻟﻘﺎ آﭘﻮﭘﺘﻮزﯾﺲ در ﻧﻮرون ﻫﺎ ﻣﯽ ﮔﺮدد. ﺳﻠﻮل ﻫﺎي ﮔﻠﯿﺎ ﻧﻘﺶ ﺑﺴﯿﺎر ﻣﻬﻤﯽ در ﺟﻠﻮﮔﯿﺮي از ﭘﯿﺸﺮﻓﺖ اﯾﻦ ﺑﯿﻤﺎري ﻫﺎ و اﻓﺰاﯾﺶ ﺑﻘﺎي ﻧﻮروﻧﯽ دارﻧﺪ. ﻣﻬﻤﺘﺮﯾﻦ ﻋﻤﻠﮑﺮد ﺳﻠﻮل ﻫﺎي ﮔﻠﯿﺎ در ﺑﻘﺎي ﻧﻮرون ﻫﺎ ﺗﺮﺷﺢ ﻓﺎﮐﺘﻮر ﻫﺎي ﻣﺤﺎﻓﻈﺖ ﮐﻨﻨﺪه ﻧﻮروﻧﯽ ﻣﯽ ﺑﺎﺷﺪ. از ﻓﺎﮐﺘﻮرﻫﺎي ﻣﺤﺎﻓﻈﺖ ﮐﻨﻨﺪه ﻧﻮروﻧﯽ ﮐﻪ ﺗﻮﺳﻂ آﺳﺘﺮوﺳﯿﺖ ﻫﺎ ﺗﺮﺷﺢ ﻣﯽ ﮔﺮدد. ﻣﯽ ﺗﻮان ﺑﻪ BDNF،GDNF،TGFβ2،TGFβ1 و NT3 اﺷﺎره ﮐﺮد. در اﯾﻦ ﻣﻄﺎﻟﻌﻪ ﻧﻘﺶ اﺣﺘﻤﺎﻟﯽ ورﺗﻤﺎﻧﯿﻦ در ﺑﯿﺎن ﻓﺎﮐﺘﻮرﻫﺎي ﻣﺤﺎﻓﻈﺖ ﮐﻨﻨﺪه ﻧﻮروﻧﯽBDNF و NT3 ﻣﺘﺮﺷﺤﻪ از ﻣﯿﮑﺮوﮔﻠﯿﺎﻫﺎ و اﻟﯿﮕﻮدﻧﺪروﺳﯿﺖ ﻫﺎ درﻣﺤﯿﻂ in vitro ﻣﻮرد ﺑﺮرﺳﯽ ﻗﺮار ﮔﺮﻓﺖ. ﺑﺮاي ﮐﺸﺖ ﺳﻠﻮل ﻫﺎي آﺳﺘﺮوﺳﯿﺖ از ﻧﻮزاد 2-1 روزه ﻣﻮش ﺻﺤﺮاﯾﯽ Wistar اﺳﺘﻔﺎده ﺷﺪ. ﺑﺪﯾﻦ ﺗﺮﺗﯿﺐ ﭘﺲ از ﺿﺪ ﻋﻔﻮﻧﯽ ﭘﻮﺳﺖ ﺑﺪن و دﺳﺖ ﻫﺎي ﻧﻮزادان ﺑﺎ ﭘﻨﺒﻪ آﻏﺸﺘﻪ ﺑﻪ اﻟﮑﻞ 70 درﺻﺪ، ﺑﺎ ﻗﯿﭽﯽ ﭘﻮﺳﺖ روي ﺳﺮ ﮐﺎﻣﻼً ﺑﺮداﺷﺘﻪ و ﺟﻤﺠﻤﻪ از ﻧﺎﺣﯿﻪ ﮔﺮدن ﺑﻪ ﻃﺮف ﭘﯿﺸﺎﻧﯽ ﺷﮑﺎﻓﺘﻪ ﺷﺪ ﺗﺎ ﮐﻮرﺗﮑﺲ ﻣﻐﺰ آﺷﮑﺎر ﮔﺮدد. ﺳﭙﺲ ﻣﻐﺰ را از ﺟﻤﺠﻤﻪ ﺟﺪا و در ﺑﺎﻓﺮ ﻫﻨﮑﺲ ﻗﺮار داده ﺷﺪ.ﻏﻠﻈﺖ ﻫﺎي 16 ،12 ،8 ﻣﻮل ﺑﺮ ﻟﯿﺘﺮ ورﺗﻤﺎﻧﯿﻦ ﺑﻪ وﺳﯿﻠﻪ ي ﺳﻤﭙﻠﺮ ﺑﻪ ﺳﻠﻮل ﻫﺎي آﺳﺘﺮوﺳﯿﺖ ﮐﺸﺖ داده ﺷﺪه در ﭘﻠﯿﺖ ﻫﺎي 6-well اﺿﺎﻓﻪ ﺷﺪ و ﻣﯿﺰان ﺑﯿﺎن ﻓﺎﮐﺘﻮرﻫﺎي ﻣﺤﺎﻓﻈﺖ ﮐﻨﻨﺪه ﻧﻮروﻧﯽ از ﻃﺮﯾﻖ R-T PCR اﻧﺪازه ﮔﯿﺮي ﺷﺪ. ﺳﻠﻮل ﻫﺎي آﺳﺘﺮوﺳﯿﺖ ﺗﯿﻤﺎر ﺷﺪه ﺑﻮﺳﯿﻠﻪ ورﺗﻤﺎﻧﯿﻦ از ﻟﺤﺎظ ﻣﻮرﻓﻮﻟﻮژي ﺗﻐﯿﯿﺮات ﻗﺎﺑﻞ ﻣﻼﺣﻈﻪ اي را ﻧﺸﺎن دادﻧﺪ. اﯾﻦ در ﺣﺎﻟﯽ ﺑﻮد ﮐﻪ ﺗﻐﯿﯿﺮات ﻣﻮرﻓﻮﻟﻮژﯾﮑﯽ در ﺳﻠﻮل ﻫﺎي ﮐﻨﺘﺮل در ﻣﻘﺎﯾﺴﻪ ﺑﺎ ﺳﻠﻮل ﻫﺎي ﺗﯿﻤﺎر ﺷﺪه ﺑﺴﯿﺎر ﮐﻤﺘﺮ ﻣﺸﺎﻫﺪه ﺷﺪ.

Background and Aim: Neurodegenerative diseases induce apoptosis in neurons by affecting the central nervous system. Glia cells play an important role in preventing the progression of these diseases and increasing neuronal survival. The most important function of glia cells after neuronal survival is the secretion of neuroprotective factors. Neuroprotective factors secreted by astrocytes include TGFβ1, TGFβ2, GDNF, BDNF, and NT3. In this study, we will investigate the possible role of vortmanine in the expression of neuroprotective factors BDNF and NT3 secreted by microglia and oligodendrocytes in vitro. For this purpose, cultured microglia and oligodendrocytes were treated with concentrations of 8, 12, 16 μmol / L for 72 hours. A 1-2-day-old Wistar rat was used to culture astrocyte cells. Thus, after disinfecting the skin of the body and hands of infants with 70% alcohol-soaked cotton, the infant's head was cut with scissors, the scalp was completely removed and the skull was cut from the neck to the forehead. To reveal the cerebral cortex. The brain was then removed from the skull and placed in Hanks buffer. Concentrations of 8, 12, and 16 mol / L of vortmanine are added by sampler to microglia and oligodendrocyte cells cultured in 6-well plates and The expression of neuroprotective factors was measured by R-T PCR. Vertmanine-treated astrocyte cells showed significant morphological changes.However, morphological changes were much less observed in control cells compared to treated cells.