本文選題：柔嫩艾美耳球蟲 + 盲腸上皮細(xì)胞��； 參考：《山西農(nóng)業(yè)大學(xué)》2015年碩士論文

【摘要】：為探討死亡受體凋亡通路在雞柔嫩艾美耳球蟲(E.tenella)誘導(dǎo)的宿主細(xì)胞凋亡中的作用以及感染E.tenella的細(xì)胞培養(yǎng)上清對未感染細(xì)胞凋亡的影響,采用E.tenella體外盲腸培養(yǎng)技術(shù)、流式細(xì)胞術(shù)、原位末端標(biāo)記技術(shù)、酶聯(lián)免疫分析法、HE染色等技術(shù)等,對Caspase-8抑制劑(Z-IETD-FMK)處理組和非處理組E.tenella宿主細(xì)胞凋亡率、DNA損傷情況、Caspase-8、Caspase-3、sFas、sFasL、mFas、mFasL的活性和蟲體感染率進(jìn)行動態(tài)測定；對不同時間段感染E.tenella與未感染E.tenella的細(xì)胞培養(yǎng)上清培養(yǎng)的盲腸上皮細(xì)胞凋亡率、Caspase-8、Caspase-9活性進(jìn)行動態(tài)測定,結(jié)果表明：(1)感染E.tenalla 4h后,宿主細(xì)胞的早期凋亡率、晚期凋亡和壞死率低于空白對照組,但差異不顯著(P0.05), DNA損傷率顯著(P0.05)低于空白對照組；感染E.tenalla24h-72h、120h后,宿主細(xì)胞的早期凋亡率、晚期凋亡和壞死率極顯著(P0.01)高于空白對照組；感染E.tenalla 24h-96h后,宿主細(xì)胞DNA損傷率極顯著(P0.01)高于空白對照組；宿主細(xì)胞凋亡率隨著E.tenalla感染時間的延長而增加。表明感染E.tenella早期,能抑制宿主細(xì)胞凋亡,后期能加快宿主細(xì)胞的凋亡。(2) Caspase-8抑制劑(Z-IETD-FMK)組在感染E.tenella早期(4h),宿主細(xì)胞早期凋亡率、晚期凋亡和壞死率、DNA損傷率以及蟲體感染率高于接種不加藥組,但差異不顯著(P0.05)；Caspase-8抑制劑組在48h-96h,宿主細(xì)胞的早期凋亡率顯著(P0.05)或極顯著(P0.01)低于接種不加藥組；在24h-120h,Caspase-8抑制劑組宿主細(xì)胞的晚期凋亡和壞死率均低于接種不加藥組,且在72h、120h差異極顯著(P0.01)；在24h-96h,Caspase-8抑制劑組宿主細(xì)胞的DNA損傷率均極顯著(P0.01)或顯著(P0.05)低于接種不加藥組；在24h-120h,Caspase-8抑制劑組宿主細(xì)胞的蟲體感染率均極顯著(P0.01)高于接種不加藥組。表明死亡受體通路參與了E.tenella引起的宿主細(xì)胞凋亡,抑制死亡受體通路有助于E.tenella子孢子的發(fā)育。(3)感染E.tenella早期(4h),宿主細(xì)胞Caspase-3的表達(dá)極顯著(P0.01)或顯著(P0.05)低于空白對照組；感染E.tenella后的24h-120h,宿主細(xì)胞Caspase-3、Caspase-8的表達(dá)極顯著(P0.01)高于空白對照組；感染E.tenella早期(4h),接種并添加Caspase-8抑制劑(Z-IETD-FMK)組宿主細(xì)胞Caspase-8的表達(dá)顯著(P0.05)高于接種不加藥組；感染E.tenella后的24h-120h,接種并添加Caspase-8抑制劑組宿主細(xì)胞Caspase-3、Caspase-8的表達(dá)極顯著(P0.01)低于接種不加藥組。表明E.tenella可通過激活Caspase-8和Caspase-3的活性來調(diào)節(jié)宿主細(xì)胞的死亡受體通路。(4)接種E.tenella組,除48h外細(xì)胞培養(yǎng)上清中sFas表達(dá)均顯著(P0.05)或極顯著(P0.01)低于空白對照組,而細(xì)胞mFas表達(dá)均顯著(P0.05)或極顯著(P0.01)高于空白對照組；接種E.tenella組細(xì)胞培養(yǎng)上清中sFasL表達(dá)均顯著(P0.05)或極顯著(P0.01)低于空白對照組,除48h外細(xì)胞mFasL表達(dá)均顯著(P0.05)或極顯著(P0.01)高于空白對照組。表明Fas/FasL表達(dá)升高是E.tenalla引起的宿主細(xì)胞的凋亡機(jī)制之一。(5)含48h感染E.tenella的細(xì)胞培養(yǎng)上清液組培養(yǎng)的宿主細(xì)胞的早期凋亡率顯著(P0.05)高于含48h不感染E.tenella的細(xì)胞培養(yǎng)上清液組；含12h感染E.tenella的細(xì)胞培養(yǎng)上清液組培養(yǎng)的宿主細(xì)胞的晚期凋亡和壞死率極顯著(P0.01)低于含12h不感染E.tenella的細(xì)胞培養(yǎng)上清液組,含4h、24h-120h感染E.tenella的細(xì)胞培養(yǎng)上清液組培養(yǎng)的宿主細(xì)胞的晚期凋亡和壞死率高于含4h、24h-120h不感染E.tenella的細(xì)胞培養(yǎng)上清液組,且在24h、120h時差異極顯著(P0.01)或顯著(P0.05)。含感染E.tenella的細(xì)胞培養(yǎng)上清液培養(yǎng)的宿主細(xì)胞的Caspase-8的表達(dá)高于不感染E.tenella的細(xì)胞培養(yǎng)上清液培養(yǎng)組,但差異不顯著(P0.05)；含4h、24h感染E.tenella的細(xì)胞培養(yǎng)上清液培養(yǎng)的宿主細(xì)胞的Caspase-9的表達(dá)顯著(P0.05)高于含4h、24h不感染E.tenell的細(xì)胞培養(yǎng)上清液組,其余時間段差異不顯著(P0.05)。表明宿主細(xì)胞感染E.tenella后,球蟲的分泌物對未感染細(xì)胞的凋亡有影響,且主要通過線粒體通路引起宿主細(xì)胞的凋亡。
[Abstract]:In order to investigate the role of death receptor apoptotic pathway in the apoptosis of host cells induced by E.tenella and the effect of culture supernatant on apoptosis of non - infected cells , the apoptosis rate , DNA damage , Caspase - 8 , Caspase - 3 , sFas , sFasL , mFas , mFasL and the rate of infection of E . tenella host cells were measured by E . tenella .
The apoptosis rate , Caspase - 8 and Caspase - 9 activity of the cultured epithelial cells of E . tenella infected with E . tenella in different time periods were measured dynamically . The results showed that : ( 1 ) The early apoptotic rate , late apoptosis and necrosis rate of the host cells were lower than those in the blank control group after infection of E . tenella 4h , but the difference was not significant ( P0.05 ) .
The early apoptotic rate , late apoptosis and necrosis rate of the host cells were significantly higher than those in the blank control group after infection of E . tenalla24h - 72h , 120h .
After infection of E . tenalla 24h - 96h , the DNA damage rate of host cells was significantly higher than that in blank control group ( P0.01 ) .
( 2 ) Caspase - 8 inhibitor ( Z - IETD - FMK ) could inhibit the apoptosis of host cells in the early stage of infection .
The apoptosis rate of Caspase - 8 inhibitor was significantly higher ( P0.05 ) or very significant ( P0.01 ) .
In 24h - 120h , the apoptosis and necrosis rate of Caspase - 8 inhibitor group were significantly lower than those in the dose - free group , and the difference was significant at 72 h and 120 h ( P0.01 ) .
The DNA damage rate of the host cells of the caspase - 8 inhibitor group was significantly higher ( P0.01 ) or significant ( P0.05 ) at 24h - 96h , but lower than that in the administration group .
The results showed that the death receptor pathway was involved in the development of E . tenella . ( 3 ) The expression of Caspase - 3 in the host cell was significantly higher than that in the blank control group ( P0.05 ) .
The expression of Caspase - 3 and Caspase - 8 in the host cells was significantly higher than that in the blank control group ( P0.01 ) .
The expression of Caspase - 8 was significantly higher in the host cells infected with E . tenella ( 4h ) , and Caspase - 8 inhibitor ( Z - IETD - FMK ) was inoculated and added ( P0.05 ) .
The expression of Caspase - 3 and Caspase - 8 was significantly higher than that in control group ( P0.05 ) .
The expression of sFasL in culture supernatant of E . tenella group was significantly higher than that in blank control group ( P0.05 ) or very significant ( P0.01 ) .
The apoptosis and necrosis rate of the host cells cultured in the culture supernatant of E . tenella infected with E . tenella were significantly higher than those in the culture supernatant of E . tenella group ( P0.05 ) .
The expression of Caspase - 9 in the culture supernatant of E . tenella infected with E . tenella was significantly higher than that in culture supernatant ( P0.05 ) .
【學(xué)位授予單位】：山西農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2015
【分類號】：S852.7

【參考文獻(xiàn)】

相關(guān)期刊論文 前10條

1 安健,汪明,孔繁瑤,殷佩云;柔嫩艾美耳球蟲(Eimeria tenella)小配子發(fā)育超微結(jié)構(gòu)的觀察[J];北京農(nóng)學(xué)院學(xué)報;2000年04期

2 鄧家儉;王黎霞;安健;;雞柔嫩艾美耳球蟲對宿主細(xì)胞凋亡的影響[J];北京農(nóng)學(xué)院學(xué)報;2011年02期

3 王英,王曉麗,張錫林;寄生蟲與細(xì)胞凋亡[J];重慶醫(yī)學(xué);2005年09期

4 謝明權(quán),吳惠賢,謝宏料,魏文康,彭新宇,張健，

本文編號：2113860