Exposure to lipopolysaccharide (LPS) reduces contractile response of small airways from GSTCD-/- mice

IntroductionGenome-Wide Association Studies suggest glutathione S transferase C terminal domain (GSTCD) may play a role in development of Chronic Obstructive Pulmonary Disease. We aimed to define the potential role of GSTCD in airway inflammation and contraction using precision cut lung slice (PCLS) from wild-type (GSTCD+/+) and GSTCD knockout mice (GSTCD-/-).MethodsPCLS from age and gender matched GSTCD+/+ and GSTCD-/- mice were prepared using a microtome. Contraction was studied after applying either a single dose of Methacholine (Mch) (1 μM) or different doses of Mch (0.001 to 100 μM). Each slice was then treated with lipopolysaccharide (LPS) or vehicle (PBS) for 24 hours. PCLS contraction in the same airway was repeated before and after stimulation. Levels of TNFα production was also measured.ResultsThere were no differences in contraction of PCLS from GSTCD+/+ and GSTCD-/- mice in response to Mch (EC50 of GSTCD+/+ vs GSTCD-/- animals: 100.0±20.7 vs 107.7±24.5 nM, p = 0.855, n = 6 animals/group). However, after LPS treatment, there was a 31.6% reduction in contraction in the GSTCD-/- group (p = 0.023, n = 6 animals). There was no significant difference between PBS and LPS treatment groups in GSTCD+/+ animals. We observed a significant increase in TNFα production induced by LPS in GSTCD-/- lung slices compared to the GSTCD+/+ LPS treated slices.ConclusionGSTCD knockout mice showed an increased responsiveness to LPS (as determined by TNFα production) that was accompanied by a reduced contraction of small airways in PCLS. These data highlight an unrecognised potential function of GSTCD in mediating inflammatory signals that affect airway responses.引言 全基因组关联研究提示，谷胱甘肽S转移酶C端结构域（glutathione S transferase C terminal domain, GSTCD）可能与慢性阻塞性肺疾病的发生发展相关。本研究旨在利用野生型（GSTCD+/+）及GSTCD敲除小鼠（GSTCD-/-）的精密切割肺切片（precision cut lung slice, PCLS），明确GSTCD在气道炎症与气道收缩中的潜在作用。 方法 采用切片机制备年龄与性别匹配的GSTCD+/+及GSTCD-/-小鼠的精密切割肺切片。分别给予单剂量乙酰甲胆碱（Methacholine, Mch，1 μM）或梯度剂量乙酰甲胆碱（0.001至100 μM），检测气道收缩反应。随后将每份切片分别用脂多糖（lipopolysaccharide, LPS）或溶媒对照（磷酸盐缓冲液，PBS）处理24小时。在刺激前后，于同一气道内重复检测肺切片的收缩反应，并测定肿瘤坏死因子α（TNFα）的分泌水平。 结果 在乙酰甲胆碱刺激下，GSTCD+/+与GSTCD-/-小鼠的精密切割肺切片收缩反应无显著差异（GSTCD+/+组与GSTCD-/-组的半最大效应浓度EC50分别为100.0±20.7 nM与107.7±24.5 nM，P=0.855，每组n=6只动物）。但经脂多糖处理后，GSTCD-/-组的气道收缩反应降低了31.6%（P=0.023，n=6只动物）；而GSTCD+/+小鼠的脂多糖处理组与磷酸盐缓冲液处理组之间无显著差异。与GSTCD+/+的脂多糖处理切片相比，GSTCD-/-肺切片中脂多糖诱导的TNFα分泌水平显著升高。 结论 GSTCD敲除小鼠对脂多糖的反应性增强（以TNFα分泌水平为评价指标），同时其精密切割肺切片的小气道收缩反应降低。本研究结果揭示了GSTCD在介导影响气道反应的炎症信号通路中一项未被认知的潜在功能。