Laboratório de Neuroquímica e Biologia Celular, Instituto de Ciências da Saúde, Universidade Federal da Bahia - UFBA, Av. Reitor Miguel Calmon s/n, Vale do Canela, CEP 41100-100 Salvador, Bahia, Brazil.
Exp Parasitol. 2013 Mar;133(3):269-74. doi: 10.1016/j.exppara.2012.11.016. Epub 2012 Dec 19.
Neospora caninum causes abortion in cattle and neurological disorders in dogs. The immunological response to this parasite has been described as predominantly of the Th1 type. However, infected primary glial cell cultures release IL-10 and IL-6 but not IFN-γ. This suggests a rather protective response of the glia to avoid inflammatory damage of the nervous tissue. In this study, we investigated the effects of pro-inflammatory cytokines in primary mixed cultures of rat astrocytes and microglia infected with N. caninum. The cells were treated with either IFN-γ, TNF-α, anti-IL-10 or anti-TGF-β antibodies and were infected with parasite tachyzoites 24h later. Trypan Blue exclusion and MTT assays were performed to test cell viability. It was observed that cytokines, antibody treatment and in vitro infection did not reveal significant cell death in the various culture conditions. Treatment with 50, 150 and 300 IU/mL of either IFN-γ or TNF-α reduced tachyzoites numbers in cultures by 36.7%, 54.8% and 63.8% for IFN-γ and by 27.6%, 38.4% and 29.7% for TNF-α, respectively. In the absence of IL-10 and TGF-β, tachyzoite numbers were reduced by 52.8% and 41.5%, respectively. While IFN-γ (150 and 300 IU/mL) increased the nitrite levels in uninfected cells, parasite infection seemed to reduce the nitrite levels, and this reduction was more expressive in IFN-γ-infected cells, thereby suggesting an inhibitory effect on its production. However, TNF-α, IL-10 and TGF-β did not affect the nitrite levels. Basal PGE(2) levels also increased by 17% and 25%; 78% and 13% in uninfected and infected cells treated with IFN-γ or anti-TGF-β, respectively. Nevertheless, the antibody neutralization of IL-10 reduced PGE(2) release significantly. These results highlight the possibility of a combined effect between the IFN-γ and parasite evasion strategies and show that the IFN-γ, TNF-α, IL-10 and TGF-β cytokines participate in parasite proliferation control mechanisms.
刚地弓形虫可引起牛流产和犬神经紊乱。该寄生虫的免疫反应主要为 Th1 型。然而，感染的原代神经胶质细胞培养物释放 IL-10 和 IL-6，但不释放 IFN-γ。这表明神经胶质细胞对避免神经组织的炎症损伤具有较强的保护作用。在这项研究中，我们调查了促炎细胞因子对感染刚地弓形虫的大鼠星形胶质细胞和小胶质细胞混合原代培养物的影响。细胞用 IFN-γ、TNF-α、抗 IL-10 或抗 TGF-β 抗体处理，24 小时后用寄生虫速殖子感染。采用台盼蓝排除法和 MTT 法检测细胞活力。结果表明，细胞因子、抗体处理和体外感染在各种培养条件下均未导致显著的细胞死亡。用 50、150 和 300 IU/mL 的 IFN-γ或 TNF-α处理，分别使培养物中的速殖子数量减少 36.7%、54.8%和 63.8%，而用 TNF-α处理，速殖子数量分别减少 27.6%、38.4%和 29.7%。在没有 IL-10 和 TGF-β的情况下，速殖子数量分别减少 52.8%和 41.5%。IFN-γ（150 和 300 IU/mL）增加未感染细胞中的亚硝酸盐水平，而寄生虫感染似乎降低了亚硝酸盐水平，IFN-γ 感染细胞中的降低更为明显，这表明对其产生具有抑制作用。然而，TNF-α、IL-10 和 TGF-β并不影响亚硝酸盐水平。基础 PGE（2）水平也分别增加 17%和 25%；未感染和感染细胞用 IFN-γ或抗 TGF-β处理后，分别增加 78%和 13%。然而，IL-10 的抗体中和显著降低 PGE（2）的释放。这些结果突出了 IFN-γ 和寄生虫逃避策略之间可能存在的协同作用，并表明 IFN-γ、TNF-α、IL-10 和 TGF-β 细胞因子参与寄生虫增殖控制机制。
