Vernooy Juanita H J, Möller Gertrude M, van Suylen Robert J, van Spijk Marlies P, Cloots Roy H E, Hoet Peter H, Pennings Herman Jan, Wouters Emiel F M
Nutrition and Toxicology Research Institute Maastricht, Department of Respiratory Medicine, University Hospital Maastricht, P.O. Box 5800, 6202 AZ Maastricht, The Netherlands.
Am J Respir Crit Care Med. 2007 Mar 1;175(5):464-72. doi: 10.1164/rccm.200602-169OC. Epub 2006 Nov 30.
Chronic obstructive pulmonary disease (COPD) is associated with increased numbers of CD8(+) cytotoxic T lymphocytes (CTLs) in the lung, but the functional activity of CTLs remains unknown. Granzyme A (GrA) and B (GrB) are serine proteases considered to be important effector molecules of CTLs and natural killer cells.
To investigate protein and mRNA expression of GrA and GrB in peripheral lung tissue from patients with COPD and control subjects with normal lung function.
Paraffin-embedded sections of surgical lung specimens from 22 patients with COPD (FEV(1), 22% predicted; GOLD stage 4) and 15 control subjects (FEV(1), 108% predicted) were immunostained for GrA and GrB, and semiquantified on a 3-point scale. Messenger RNA expression in total lung, specific cell types enriched for by laser capture microdissection, and freshly isolated primary cells were determined by reverse transcriptase-polymerase chain reaction.
GrA and GrB immunoreactivity was observed in CD8(+) CTLs and CD57(+) natural killer cells, but also in type II pneumocytes and alveolar macrophages in both groups. Bronchiolar epithelium stained positive for GrA, but negative for GrB. These observations were confirmed by reverse transcriptase-polymerase chain reaction on total lung, laser capture microdissection-enriched specific cell types and freshly isolated primary type II pneumocytes. The scores of GrA-expressing type II pneumocytes were significantly higher in patients with COPD versus control subjects.
GrA and GrB mRNA and protein are detectable in human lung tissue. GrA expression is increased in type II pneumocytes of patients with very severe COPD. These results indicate that GrA may be important in the development of COPD.
慢性阻塞性肺疾病（COPD）与肺内CD8(+) 细胞毒性T淋巴细胞（CTLs）数量增加有关，但CTLs的功能活性尚不清楚。颗粒酶A（GrA）和颗粒酶B（GrB）是丝氨酸蛋白酶，被认为是CTLs和自然杀伤细胞的重要效应分子。
研究COPD患者和肺功能正常的对照者外周肺组织中GrA和GrB的蛋白及mRNA表达。
对22例COPD患者（第1秒用力呼气容积[FEV(1)]为预计值的22%；全球慢性阻塞性肺疾病倡议[GOLD]4期）和15例对照者（FEV(1)为预计值的108%）手术肺标本的石蜡包埋切片进行GrA和GrB免疫染色，并采用3分制进行半定量。通过逆转录聚合酶链反应测定全肺、经激光捕获显微切割富集的特定细胞类型以及新鲜分离的原代细胞中的信使RNA表达。
在两组的CD8(+) CTLs和CD57(+) 自然杀伤细胞中均观察到GrA和GrB免疫反应性，在II型肺细胞和肺泡巨噬细胞中也有观察到。细支气管上皮GrA染色呈阳性，但GrB染色呈阴性。这些观察结果通过对全肺、经激光捕获显微切割富集的特定细胞类型以及新鲜分离的原代II型肺细胞进行逆转录聚合酶链反应得到证实。与对照者相比，COPD患者中表达GrA的II型肺细胞评分显著更高。
在人肺组织中可检测到GrA和GrB的mRNA及蛋白。在极重度COPD患者的II型肺细胞中GrA表达增加。这些结果表明GrA可能在COPD的发生发展中起重要作用。
