雙孔鉀離子通道TREK-1對(duì)氧糖剝奪星形膠質(zhì)細(xì)胞和神經(jīng)元功能的調(diào)節(jié)
發(fā)布時(shí)間：2018-08-06 17:06
【摘要】：研究目的：觀察缺氧缺糖培養(yǎng)條件下,雙孔鉀離子通道TREK-1對(duì)大鼠皮質(zhì)星形膠質(zhì)細(xì)胞(Ast)存活率及谷氨酸(Glu)攝取功能的影響；Ast的功能狀態(tài)與神經(jīng)元之間的相互作用及TREK-1通道在其中的作用,探討TREK-1通道是否能成為全新的治療腦缺血疾病的潛在靶點(diǎn)。 研究?jī)?nèi)容：建立氧糖剝奪(oxygen and glucose deprivation, OGD)損傷模型,模擬體外腦缺血環(huán)境,檢測(cè)Ast的存活率及Glu攝取功能,同時(shí),給予花生四烯酸(AA)或甲硫氨酸(Met)改變TREK-1通道的功能,觀察Ast存活率的變化,分析在體外OGD損傷下,Ast TREK-1通道的變化與Glu攝取功能變化間的關(guān)系。建立Ast與神經(jīng)元細(xì)胞共培養(yǎng)體系,觀察體外缺氧缺糖培養(yǎng)條件下,Ast和神經(jīng)元TREK-1通道表達(dá)的變化,神經(jīng)元存活率的變化,Ast和神經(jīng)元谷氨酸受體亞型NR2B和mGluR、谷氨酸轉(zhuǎn)運(yùn)體GLT-1、凋亡通路中ERK和caspase-3的變化。給予AA或Met調(diào)節(jié)TREK-1通道的功能,觀察以上指標(biāo)的變化,以明確體外缺氧缺糖條件下,Ast功能狀態(tài)對(duì)神經(jīng)元的影響以及TREK-1通道在其中所起的作用。 研究結(jié)果：通過(guò)MTT檢測(cè),在生理?xiàng)l件下AA (5μmol/L.10μmol/L.20μmol/L、40μmol/L)和Met (1mmol/L、3mmol/L、10mmol/L、30mmol/L)對(duì)Ast的存活率均無(wú)影響,而缺氧缺糖條件下,AA和Met均可增加Ast的存活率。體外缺氧缺糖后,Ast對(duì)Glu的攝取出現(xiàn)代償性增加,AA對(duì)OGD條件下Ast的Glu攝取有增強(qiáng)作用,而Met可以抑制OGD條件下Ast的Glu攝能力。通過(guò)乳酸脫氫酶(lactate dehydrogenase, LDH)釋放檢測(cè),神經(jīng)元-Ast共培養(yǎng)與神經(jīng)元單獨(dú)培養(yǎng)相比LDH釋放量有所降低,給予Met后LDH釋放量由47.84U/L下降到23.95U/L,說(shuō)明Met可以減少神經(jīng)元的死亡。通過(guò)RT-PCR檢測(cè)顯示,OGD條件下共培養(yǎng)神經(jīng)元中TREK-1和GLT-1mRNA表達(dá)量下調(diào),NR2B和mGluR mRNA表達(dá)量上升,共培養(yǎng)Ast中TREK-1和mGluR mRNA表達(dá)量下調(diào),GLT-1和ERK mRNA表達(dá)量上升,給予AA后可下調(diào)TREK-1mRNA、NR2B mRNA及ERK mRNA的表達(dá)；給予Met后TREK-1mRNA及NR2B mRNA表達(dá)量均受到抑制,對(duì)GLT-1mRNA表達(dá)有雙向調(diào)節(jié)作用。 研究結(jié)論：AA和Met對(duì)生理狀態(tài)下Ast的存活率無(wú)顯著影響,但能夠改善OGD后Ast的存活率。AA與Met有助于增強(qiáng)OGD損傷后共培養(yǎng)神經(jīng)元的存活,推測(cè)[REK-1通道功能的改變影響Ast與神經(jīng)元之間的相互作用,但Met提高Ast的存活率與受體無(wú)明顯聯(lián)系,其促增殖原因有待進(jìn)一步研究。
[Abstract]:Objective: to observe the effects of TREK-1 on the survival rate of (Ast) and the uptake of glutamate (Glu) in rat cortical astrocytes in hypoxic-glucose deficient culture. The functional state of Ast and the interaction between neurons and the role of TREK-1 channel in the interaction, to explore whether the TREK-1 channel can become a new potential target for the treatment of cerebral ischemic diseases. Content: establish oxygen glucose deprivation (oxygen and glucose deprivation, OGD) damage model, simulate the brain ischemic environment in vitro, detect the survival rate of Ast and Glu uptake function, at the same time, give arachidonic acid (AA) or methionine (Met) to change the function of TREK-1 channel. To observe the changes of Ast survival rate and to analyze the relationship between the changes of Ast TREK-1 channel and Glu uptake function under OGD injury in vitro. The co-culture system of Ast and neuronal cells was established to observe the expression of Ast and TREK-1 channels in cultured neurons under hypoxia and glucose deprivation in vitro. The changes of neuronal survival rate were related to the changes of neuronal glutamate receptor subtype NR2B and mGluR, glutamate transporter GLT-1, and ERK and caspase-3 in apoptotic pathway. AA or Met was given to regulate the function of TREK-1 channel, and the changes of the above indexes were observed to clarify the effect of Ast function on neurons under hypoxia and glucose deprivation in vitro and the role of TREK-1 channel in it. Results: under physiological conditions, AA (5 渭 mol/L.10 渭 mol/L.20 渭 mol / L ~ 40 渭 mol/L) and Met (1 mmol / L ~ (3) mmol / L ~ (10) mmol / L ~ (30 mmol / L) had no effect on the survival rate of Ast, but both AA and Met could increase the survival rate of Ast. After hypoxia and glucose deprivation in vitro, the uptake of Glu by Ast in vitro was increased. AA could enhance the Glu uptake of Ast under OGD, while Met could inhibit the Glu uptake of Ast under OGD condition. By detecting the release of lactate dehydrogenase (lactate dehydrogenase, LDH), the release of LDH in neuron Ast co-culture was lower than that in neuron culture alone, and the LDH release decreased from 47.84U/L to 23.95 U / L after Met administration, which indicated that Met could reduce the death of neurons. The expression of TREK-1 and GLT-1mRNA in co-cultured neurons was down-regulated by RT-PCR, and the expression of TREK-1 and mGluR mRNA in co-cultured Ast was down-regulated. The expression of TREK-1 and mGluR mRNA was down-regulated, and the expression of TREK-1mRNA-NR2B mRNA and ERK mRNA was down-regulated after treatment with AA. After Met, the expression of TREK-1mRNA and NR2B mRNA were inhibited, and the expression of GLT-1mRNA was regulated in both directions. Conclusion there is no significant effect on survival rate of Ast in physiological state by Met and% AA, but it can improve the survival rate of Ast after OGD. AA and Met can enhance the survival of co-cultured neurons after OGD injury. It is inferred that the changes of REK-1 channel function affect the interaction between Ast and neurons, but there is no obvious relationship between the increase of Ast survival rate and the receptor by Met, and the reasons for promoting proliferation need to be further studied.
【學(xué)位授予單位】：蘭州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R96
[Abstract]:Objective: to observe the effects of TREK-1 on the survival rate of (Ast) and the uptake of glutamate (Glu) in rat cortical astrocytes in hypoxic-glucose deficient culture. The functional state of Ast and the interaction between neurons and the role of TREK-1 channel in the interaction, to explore whether the TREK-1 channel can become a new potential target for the treatment of cerebral ischemic diseases. Content: establish oxygen glucose deprivation (oxygen and glucose deprivation, OGD) damage model, simulate the brain ischemic environment in vitro, detect the survival rate of Ast and Glu uptake function, at the same time, give arachidonic acid (AA) or methionine (Met) to change the function of TREK-1 channel. To observe the changes of Ast survival rate and to analyze the relationship between the changes of Ast TREK-1 channel and Glu uptake function under OGD injury in vitro. The co-culture system of Ast and neuronal cells was established to observe the expression of Ast and TREK-1 channels in cultured neurons under hypoxia and glucose deprivation in vitro. The changes of neuronal survival rate were related to the changes of neuronal glutamate receptor subtype NR2B and mGluR, glutamate transporter GLT-1, and ERK and caspase-3 in apoptotic pathway. AA or Met was given to regulate the function of TREK-1 channel, and the changes of the above indexes were observed to clarify the effect of Ast function on neurons under hypoxia and glucose deprivation in vitro and the role of TREK-1 channel in it. Results: under physiological conditions, AA (5 渭 mol/L.10 渭 mol/L.20 渭 mol / L ~ 40 渭 mol/L) and Met (1 mmol / L ~ (3) mmol / L ~ (10) mmol / L ~ (30 mmol / L) had no effect on the survival rate of Ast, but both AA and Met could increase the survival rate of Ast. After hypoxia and glucose deprivation in vitro, the uptake of Glu by Ast in vitro was increased. AA could enhance the Glu uptake of Ast under OGD, while Met could inhibit the Glu uptake of Ast under OGD condition. By detecting the release of lactate dehydrogenase (lactate dehydrogenase, LDH), the release of LDH in neuron Ast co-culture was lower than that in neuron culture alone, and the LDH release decreased from 47.84U/L to 23.95 U / L after Met administration, which indicated that Met could reduce the death of neurons. The expression of TREK-1 and GLT-1mRNA in co-cultured neurons was down-regulated by RT-PCR, and the expression of TREK-1 and mGluR mRNA in co-cultured Ast was down-regulated. The expression of TREK-1 and mGluR mRNA was down-regulated, and the expression of TREK-1mRNA-NR2B mRNA and ERK mRNA was down-regulated after treatment with AA. After Met, the expression of TREK-1mRNA and NR2B mRNA were inhibited, and the expression of GLT-1mRNA was regulated in both directions. Conclusion there is no significant effect on survival rate of Ast in physiological state by Met and% AA, but it can improve the survival rate of Ast after OGD. AA and Met can enhance the survival of co-cultured neurons after OGD injury. It is inferred that the changes of REK-1 channel function affect the interaction between Ast and neurons, but there is no obvious relationship between the increase of Ast survival rate and the receptor by Met, and the reasons for promoting proliferation need to be further studied.
【學(xué)位授予單位】：蘭州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R96
【參考文獻(xiàn)】
相關(guān)期刊論文 前7條
1 付振強(qiáng);張博愛(ài);賈延R，
本文編號(hào)：2168389
本文鏈接：http://sikaile.net/yixuelunwen/yiyaoxuelunwen/2168389.html
最近更新
教材專著
