| In order to explore the relationship between E. tenella infection and chicken embryos cecum epithelial cell apoptosis and the role of PI3K/Akt signaling pathway in E. tenella early infection, we employed actinomycin D (Act D) as apoptosis inducers and use the primary chick embryo cecum epithelial cells as host cells, hematoxylin and eosin staining was used to determine infection rates, the activity of Caspase-3 was measured through spectrophotometry, apoptosis was determined by flow cytometry to studied the relationship between E. tenella infection and chicken embryos cecum epithelial cell apoptosis. We studied whether PI3K/Akt signaling pathways are involved in E.tenella-induced apoptosis of chick embryo cecal epithelial cells employed PI3K/Akt signaling pathways inhibitor (LY294002), by detecting the apoptosis, the activity of Caspase-3 and Caspase-9, the opening of mitochondrial permeability transition (MPTP), p-Akt, Bad, and NF-kB of E. tenella host cell in infected and uninfected groups, the results showed that:(1) The apoptosis rate of the infected group significantly (P< 0.05) lower than the control group at 4h after inoculating sporozoits, and significantly (P< 0.05) higher than the control group at 12-20 h; the apoptosis rate and the activity of Caspase-3 in the treated group (infected and treat with Act D) are significant (P<0.01) higher than the untreated group (infected with E.tenella sporozoits) at 4-20h after inoculating sporozoits.It shows that the invasion of E.tenella sporozoite inhibited host cell apoptosis in the early, and then promoted. At the same time, it shows that infected with E. tenella can suppress the host cell apoptosis which induced by actinomycin D.(2) The early apoptosis rate, the late apoptosis and necrosis of treated groups were significant higher (P<0.01) than the untreated group at 4-20 h, but the infection rate was significantly (P< 0.01) lower than untreated group, shows that the host cell apoptosis prevented E.tenella sporozoite further invasion.(3) The level of p-Akt in infected group significantly (p< 0.01) higher than the control group at 4h after infection, and had no significant difference (P>0.05) between this two groups at 8-20h after infection; The level of p-Akt in treated group (infected and treated with LY294002) were lower then untreated group at 4?20h after infection, at the same time, the early apoptosis rate, the late apoptosis and necrosis were significantly (p< 0.01) higher than the untreated group, shows that E.tenella sporozoite invade intochicken embryos cecum epithelial cells can activated PI3K/Akt signal pathway; and inhibited PI3K/Akt signaling pathway can promote E.tenella host cell apoptosis, indicated that PI3K/Akt signaling pathways are involved in E.tenella-induced apoptosis of chick embryo cecal epithelial cells.(4)The degrees of MPTP openness in untreated group were significantly (P< 0.01) higher than the control group at 4?20h after infection; The degrees of MPTP openness in treated group (infected and treated with LY294002) were significantly (P< 0.01) higher than the untreated group at 4h after infection, but had no significant difference (P>0.05) between this two groups at 12?20h after infection; indicated that the invasion of E.tenella sporozoite can lead to MPTP opens, PI3K/Akt signaling pathways can inhibite E.tenella-induced apoptosis of chick embryo cecal epithelial cells through limiting the openness of MPTP.(5) The level of Bad, the activity of Caspase-3 and Caspase-9 of untreated group lower (P< 0.05) then the control group, the level of NF-kB was significantly (P< 0.05) higher then the control group at 4?12h after infection; The level of Bad, the activity of Caspase-3 and Caspase-9 and NF-kB in treated group (infected and treated with LY294002) were significantly (P< 0.01) higher than the untreated group, showed that PI3K/Akt signaling pathway can involved in E.tenella early infection of the host cell apoptosis through Bad, Caspase-9, Caspase-3 and NF-kB. |
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