在 COPD 痰样本中对候选疫苗基因进行离体 RNA 表达分析。
Ex-vivo RNA expression analysis of vaccine candidate genes in COPD sputum samples.
机构信息
GSK, Via Fiorentina 1, Siena, Italy.
GSK, Rixensart, Belgium.
出版信息
Respir Res. 2023 Oct 5;24(1):243. doi: 10.1186/s12931-023-02525-z.
BACKGROUND
Chronic obstructive pulmonary disease (COPD) is a lung disease characterised by airflow-limiting inflammation and mucus production. Acute exacerbations are a major cause of COPD-related morbidity and mortality and are mostly associated with bacterial or viral infections. A vaccine targeting non-typeable Haemophilus influenzae (NTHi) and Moraxella catarrhalis (Mcat), the main bacteria associated with exacerbations, was tested in a Phase 2 trial. We assessed "ex-vivo" expression of vaccine candidate and housekeeping genes pd, pe, pilA, gapA, ompP6 of NTHi, and uspA2, parE, polA of Mcat in sputum samples of COPD patients and determined whether expression of the vaccine candidate genes pd, pe, pilA (NTHi) and uspA2 (Mcat) differed between stable and exacerbation samples.
METHODS
A single-centre, prospective, observational cohort study was conducted where 123 COPD patients were seen on enrolment, followed monthly for 2 years, and reviewed after onset of acute exacerbations. We selected 69 patients with sputum samples positive for NTHi or Mcat by PCR during at least one stable and one exacerbation visit. mRNA was isolated from the sputum, and expression of NTHi and Mcat genes was analysed with RT-PCR. Statistical analyses compared mRNA concentrations between stable and exacerbation samples and in relationship to COPD severity and exacerbation frequency.
RESULTS
The vaccine candidate genes were variably expressed in sputum samples, suggesting they are expressed in the lung. Absolute and relative expression of all NTHi vaccine candidate genes and Mcat uspA2 were similar between exacerbation and stable samples. Expression of pd and pilA was slightly associated with the number of exacerbations in the year before enrolment, and uspA2 with the disease severity status at enrolment.
CONCLUSIONS
The NTHi-Mcat vaccine candidate genes were expressed in sputum samples, and each gene had a specific level of expression. No statistically significant differences in gene expression were detectable between stable and exacerbation samples. However, the history of COPD exacerbations was slightly associated with the expression of pd, pilA and uspA2. Trial registration NCT01360398 ( https://www.
CLINICALTRIALS
gov ).
背景
慢性阻塞性肺疾病（COPD）是一种以气流受限性炎症和黏液产生为特征的肺部疾病。急性加重是 COPD 相关发病率和死亡率的主要原因，主要与细菌或病毒感染有关。一种针对非分型流感嗜血杆菌（NTHi）和卡他莫拉菌（Mcat）的疫苗已在 2 期试验中进行了测试，这两种细菌是与加重相关的主要细菌。我们评估了 COPD 患者痰液样本中疫苗候选物和管家基因 pd、pe、pilA、gapA、ompP6 的 NTHi 和 uspA2、parE、polA 的“离体”表达，并确定了稳定期和加重期样本中疫苗候选物基因 pd、pe、pilA（NTHi）和 uspA2（Mcat）的表达是否存在差异。
方法
进行了一项单中心、前瞻性、观察性队列研究，其中 123 例 COPD 患者在入组时接受了评估，随后每月评估一次，共 2 年，并在急性加重发作后进行了评估。我们选择了 69 例至少在一次稳定期和一次加重期就诊时 PCR 检测到 NTHi 或 Mcat 阳性的患者进行痰液样本检测。从痰液中提取 mRNA，并通过 RT-PCR 分析 NTHi 和 Mcat 基因的表达。统计分析比较了稳定期和加重期样本之间以及与 COPD 严重程度和加重频率之间的 mRNA 浓度。
结果
疫苗候选物基因在痰液样本中表达不同，表明其在肺部表达。所有 NTHi 疫苗候选物基因和 Mcat uspA2 的绝对和相对表达在加重期和稳定期样本之间相似。Pd 和 pilA 的表达与入组前一年的加重次数略有相关，uspA2 的表达与入组时的疾病严重程度状态相关。
结论
NTHi-Mcat 疫苗候选物基因在痰液样本中表达，每个基因的表达水平都有特异性。在稳定期和加重期样本之间，无法检测到基因表达的统计学显著差异。然而，COPD 加重的历史与 pd、pilA 和 uspA2 的表达略有相关。试验注册 NCT01360398（临床试验.gov）。
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