Departamento de Parasitologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, SP, Brazil.
National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rocky Mountain Laboratories, Hamilton, MT, USA.
Parasit Vectors. 2023 Mar 10;16(1):96. doi: 10.1186/s13071-023-05701-8.
The tick Amblyomma sculptum is the major vector of Rickettsia rickettsii, the causative agent of the highly lethal Brazilian spotted fever. It has been shown that R. rickettsii inhibits apoptosis in both human endothelial cells and tick cells. Apoptosis is regulated by different factors, among which inhibitors of apoptosis proteins (IAPs) play a central role. In the study reported here, we selected an IAP of A. sculptum that has not yet been characterized to assess its role in cell death and to determine the effects of its gene silencing on tick fitness and R. rickettsii infection.
An A. sculptum cell line (IBU/ASE-16) was treated with specific double-stranded RNA (dsRNA) for either IAP (dsIAP) or green fluorescent protein (dsGFP; as a control). The activity of caspase-3 and the exposure of phosphatidylserine were determined in both groups. In addition, unfed adult ticks, infected or not infected with R. rickettsii, were treated with either dsIAP or dsGFP and allowed to feed on noninfected rabbits. In parallel, noninfected ticks were allowed to feed on an R. rickettsii-infected rabbit. Ticks (infected or not with R. rickettsii) that remained unfed were used as a control.
Caspase-3 activity and the externalization of phosphatidylserine were significantly higher in IBU/ASE-16 cells treated with dsIAP than in those treated with dsGFP. The mortality rates of ticks in the dsIAP group were much higher than those in the dsGFP group when they were allowed to feed on rabbits, independent of the presence of R. rickettsii. Conversely, lower mortality rates were recorded in unfed ticks.
Our results show that IAP negatively regulates apoptosis in A. sculptum cells. Moreover, IAP-silenced ticks experienced higher mortality rates following the acquisition of a blood meal, suggesting that feeding may trigger the activation of apoptosis in the absence of this physiological regulator. These findings indicate that IAP is a potential antigen for an anti-tick vaccine.
硬蜱属的纹皮硬蜱是导致高度致命的巴西斑疹热的病原体立氏立克次体的主要传播媒介。已证实，立氏立克次体抑制人内皮细胞和蜱细胞的细胞凋亡。细胞凋亡由不同的因素调节，其中凋亡抑制蛋白（IAPs）起着核心作用。在本报告中，我们选择了一种尚未被表征的纹皮硬蜱 IAP，以评估其在细胞死亡中的作用，并确定其基因沉默对蜱的适应力和立氏立克次体感染的影响。
用特异性双链 RNA（dsRNA）处理纹皮硬蜱细胞系（IBU/ASE-16），用于 IAP（dsIAP）或绿色荧光蛋白（dsGFP；作为对照）。在两组中均测定了 caspase-3 的活性和磷脂酰丝氨酸的暴露情况。此外，未喂食的成年蜱，无论是否感染立氏立克次体，均用 dsIAP 或 dsGFP 处理，并允许其叮咬未感染的兔子。同时，允许非感染的蜱叮咬感染立氏立克次体的兔子。未喂食的蜱（无论是否感染立氏立克次体）作为对照。
dsIAP 处理的 IBU/ASE-16 细胞中的 caspase-3 活性和磷脂酰丝氨酸的外化明显高于 dsGFP 处理的细胞。当允许 dsIAP 组的蜱叮咬兔子时，其死亡率明显高于 dsGFP 组，无论是否存在立氏立克次体。相反，未喂食的蜱的死亡率较低。
我们的结果表明，IAP 负调节纹皮硬蜱细胞中的细胞凋亡。此外，在获得血液餐食后，沉默 IAP 的蜱的死亡率更高，这表明在没有这种生理调节剂的情况下，进食可能会引发细胞凋亡的激活。这些发现表明 IAP 是一种潜在的抗蜱疫苗抗原。
