Aref Salah, Abdullah Doaa, Fouda Manal, El Menshawy Nadia, Azmy Emaad, Bassam Ansaf, Menessy Aymen, El Refaei Mohammed
Indian J Hematol Blood Transfus. 2011 Jun;27(2):81-7. doi: 10.1007/s12288-011-0067-1. Epub 2011 Jun 8.
Patients with chronic HCV infection are prone to increased susceptibility bacterial infection due to neutropenia complicating the course of this disease. Neutropenia in those patients may stem from enhanced neutrophil apoptosis. However, the molecular mechanism of neutrophil apoptosis has not been clearly defined. Neutrophils harvested from 26 neutropenic patients with hepatitis C infection and nine age and sex-matched healthy control subjects were examined for the degree of apoptosis. Neutrophil apoptosis was quantified by flow cytometry through determination of annexin-V expression at 0 time (fresh neutrophil), and 24 h culture. Neutrophils from healthy subjects were also incubated with either 10% heterologous normal or neutropenic sera, with and without 10 µg GM-CSF. Caspases 3, 10 were assessed colormetrically in neutrophils at 0 times and after 24 h culture. At 0 time culture the neutrophil apoptosis of the HCV patients was in significantly higher as compared to that of normal control (P = 0.059). At 24 h culture patients neutrophils cultured with neutropenic patients own sera showed neutrophil apoptosis significantly increased as compared to that at 0 time culture and this effect was significantly attenuated in similar culture with addition of GM-CSF (P < 0.001). On the other hand patient's neutrophil cultured with normal sera showed insignificantly increased neutrophil apoptosis at 24 h culture as compared to that at 0 time culture. Caspases 3 and 10 activities were significantly higher in patients neutrophil after 24 h cultured with patients own sera as compared to 0 time culture (P < 0.001 for both). Addition of GM-CSF to the neutrophil culture down regulates the caspases 3 and 10 activities. The correlation study between annexin-V expression and caspases activities revealed a borderline positive correlation between annexin-V and caspase 3 (r = 0.376, P = 0.058), and significant positive correlation with caspase 10 activity (r = 0.494, P = 0.01). In conclusion, these findings suggest that enhanced neutrophil apoptosis demonstrated in neutropenic patients with HCV infection might be induced through activation of caspase 10 and is attenuated by GM-CSF.
慢性丙型肝炎病毒（HCV）感染患者由于中性粒细胞减少并发于该疾病过程中，容易出现细菌感染易感性增加。这些患者的中性粒细胞减少可能源于中性粒细胞凋亡增强。然而，中性粒细胞凋亡的分子机制尚未明确界定。对26例丙型肝炎感染伴中性粒细胞减少患者以及9例年龄和性别匹配的健康对照者采集的中性粒细胞进行凋亡程度检测。通过流式细胞术在0小时（新鲜中性粒细胞）和培养24小时时测定膜联蛋白-V表达来量化中性粒细胞凋亡。健康受试者的中性粒细胞也分别与10%异种正常血清或中性粒细胞减少血清一起孵育，同时添加或不添加10μg粒细胞-巨噬细胞集落刺激因子（GM-CSF）。在0小时和培养24小时后，用比色法评估中性粒细胞中的半胱天冬酶3、10。在0小时培养时，HCV患者的中性粒细胞凋亡明显高于正常对照（P = 0.059）。在24小时培养时，用中性粒细胞减少患者自身血清培养的患者中性粒细胞显示，与0小时培养相比，中性粒细胞凋亡显著增加，并且在添加GM-CSF的类似培养中这种效应显著减弱（P < 0.001）。另一方面，用正常血清培养的患者中性粒细胞在24小时培养时与0小时培养相比，中性粒细胞凋亡增加不显著。与0小时培养相比，并在患者自身血清中培养24小时后，患者中性粒细胞中的半胱天冬酶3和10活性显著更高（两者P均< 0.001）。向中性粒细胞培养物中添加GM-CSF可下调半胱天冬酶3和10的活性。膜联蛋白-V表达与半胱天冬酶活性之间的相关性研究显示，膜联蛋白-V与半胱天冬酶3之间存在临界正相关（r = 0.376，P = 0.058），与半胱天冬酶10活性存在显著正相关（r = 0.494，P = 0.01）。总之，这些发现表明，HCV感染伴中性粒细胞减少患者中显示的中性粒细胞凋亡增强可能是通过半胱天冬酶10的激活诱导的，并且被GM-CSF减弱。
