Division of Respiratory Medicine, Department of Paediatrics, University Hospital Bern, Bern, Switzerland.
Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland; Institute of Virology and Immunology, Bern, Switzerland; Department of Infectious Diseases and Pathobiology, Vetsuisse Faculty, University of Bern, Bern, Switzerland.
Antiviral Res. 2019 Feb;162:44-50. doi: 10.1016/j.antiviral.2018.12.007. Epub 2018 Dec 11.
Rhinovirus (RV) infection is a major cause of cystic fibrosis (CF) lung morbidity with limited therapeutic options. Various diseases involving chronic inflammatory response and infection are associated with endoplasmic reticulum (ER) stress and subsequent activation of the unfolded protein response (UPR), an adaptive response to maintain cellular homeostasis. Recent evidence suggests impaired ER stress response in CF airway epithelial cells, this might be a reason for recurrent viral infection in CF. Therefore, assuming that ER stress inducing drugs have antiviral properties, we evaluated the activation of the UPR by selected ER stress inducers as an approach to control virus replication in the CF bronchial epithelium.
We assessed the levels of UPR markers, namely the glucose-regulated protein 78 (Grp78) and the C/EBP homologous protein (CHOP), in primary CF and control bronchial epithelial cells and in a CF and control bronchial epithelial cell line before and after infection with RV. The cells were also pretreated with ER stress-inducing drugs and RV replication and shedding was measured by quantitative RT-PCR and by a TCID assay, respectively. Cell death was assessed by a lactate dehydrogenate (LDH) activity test in supernatants.
We observed a significantly impaired induction of Grp78 and CHOP in CF compare to control cells following RV infection. The ER stress response could be significantly induced in CF cells by pharmacological ER stress inducers Brefeldin A, Tunicamycin, and Thapsigargin. The chemical induction of the UPR pathway prior to RV infection of CF and control cells reduced viral replication and shedding by up to two orders of magnitude and protected cells from RV-induced cell death.
RV infection causes an impaired activation of the UPR in CF cells. Rescue of the ER stress response by chemical ER stress inducers reduced significantly RV replication in CF cells. Thus, pharmacological modulation of the UPR might represent a strategy to control respiratory virus replication in the CF bronchial epithelium.
鼻病毒（RV）感染是囊性纤维化（CF）肺部疾病的主要原因，其治疗选择有限。各种涉及慢性炎症反应和感染的疾病都与内质网（ER）应激有关，随后激活未折叠蛋白反应（UPR），这是一种维持细胞内稳态的适应性反应。最近的证据表明，CF 气道上皮细胞中的 ER 应激反应受损，这可能是 CF 中反复发生病毒感染的原因。因此，假设 ER 应激诱导剂具有抗病毒特性，我们评估了选定的 ER 应激诱导剂对 UPR 的激活，作为控制 CF 支气管上皮细胞中病毒复制的一种方法。
我们评估了 UPR 标志物的水平，即葡萄糖调节蛋白 78（Grp78）和 C/EBP 同源蛋白（CHOP），在原发性 CF 和对照支气管上皮细胞以及 CF 和对照支气管上皮细胞系中，在 RV 感染前后。还在用 ER 应激诱导剂预处理细胞，并通过定量 RT-PCR 和 TCID 测定分别测量 RV 复制和脱落。通过上清液中的乳酸脱氢酶（LDH）活性试验评估细胞死亡。
我们观察到，与对照细胞相比，RV 感染后 CF 中的 Grp78 和 CHOP 的诱导明显受损。通过药理学 ER 应激诱导剂布雷菲德菌素 A、衣霉素和他普西龙，可显著诱导 CF 细胞中的 ER 应激反应。在 CF 和对照细胞感染 RV 之前，化学诱导 UPR 途径可将病毒复制和脱落减少多达两个数量级，并防止细胞因 RV 诱导的细胞死亡。
RV 感染导致 CF 细胞中 UPR 的激活受损。通过化学 ER 应激诱导剂挽救 ER 应激反应可显著降低 CF 细胞中的 RV 复制。因此，UPR 的药理学调节可能是控制 CF 支气管上皮细胞中呼吸道病毒复制的一种策略。
