Tran Hai B, Ahern Jessica, Hodge Greg, Holt Phillip, Dean Melinda M, Reynolds Paul N, Hodge Sandra
Lung Research, Hanson Institute and Department Thoracic Medicine, Royal Adelaide Hospital, Adelaide, South Australia, Australia.
Lung Research, Hanson Institute and Department Thoracic Medicine, Royal Adelaide Hospital, Adelaide, South Australia, Australia; Department of Medicine, University of Adelaide, Adelaide, South Australia, Australia.
PLoS One. 2014 Jun 5;9(6):e98571. doi: 10.1371/journal.pone.0098571. eCollection 2014.
We have previously established that a defect in the ability of alveolar macrophages (AM) to phagocytose apoptotic cells (efferocytosis) and pathogens is a potential therapeutic target in COPD. We further showed that levels of mannose binding lectin (MBL; required for effective macrophage phagocytic function) were reduced in the airways but not circulation of COPD patients. We hypothesized that increased oxidative stress in the airway could be a cause for such disturbances. We therefore studied the effects of oxidation on the structure of the MBL molecule and its functional interactions with macrophages. Oligomeric structure of plasma derived MBL (pdMBL) before and after oxidation (oxMBL) with 2,2'-azobis(2-methylpropionamidine)dihydrochroride (AAPH) was investigated by blue native PAGE. Macrophage function in the presence of pd/oxMBL was assessed by measuring efferocytosis, phagocytosis of non-typeable Haemophilus influenzae (NTHi) and expression of macrophage scavenger receptors. Oxidation disrupted higher order MBL oligomers. This was associated with changed macrophage function evident by a significantly reduced capacity to phagocytose apoptotic cells and NTHi in the presence of oxMBL vs pdMBL (eg, NTHi by 55.9 and 27.0% respectively). Interestingly, oxidation of MBL significantly reduced macrophage phagocytic ability to below control levels. Flow cytometry and immunofluorescence revealed a significant increase in expression of macrophage scavenger receptor (SRA1) in the presence of pdMBL that was abrogated in the presence of oxMBL. We show the pulmonary macrophage dysfunction in COPD may at least partially result from an oxidative stress-induced effect on MBL, and identify a further potential therapeutic strategy for this debilitating disease.
我们之前已经确定，肺泡巨噬细胞（AM）吞噬凋亡细胞（胞葬作用）和病原体的能力缺陷是慢性阻塞性肺疾病（COPD）的一个潜在治疗靶点。我们进一步表明，甘露糖结合凝集素（MBL；有效巨噬细胞吞噬功能所必需）的水平在COPD患者的气道中降低，但在循环系统中未降低。我们推测气道中氧化应激增加可能是导致这种紊乱的原因。因此，我们研究了氧化对MBL分子结构及其与巨噬细胞功能相互作用的影响。通过蓝色非变性聚丙烯酰胺凝胶电泳（blue native PAGE）研究了用2,2'-偶氮二异丁脒二盐酸盐（AAPH）氧化前后血浆来源的MBL（pdMBL）的寡聚体结构（oxMBL）。通过测量胞葬作用、不可分型流感嗜血杆菌（NTHi）的吞噬作用以及巨噬细胞清道夫受体的表达来评估在pd/oxMBL存在下的巨噬细胞功能。氧化破坏了高阶MBL寡聚体。这与巨噬细胞功能改变有关，在存在oxMBL与pdMBL的情况下，吞噬凋亡细胞和NTHi的能力显著降低（例如，NTHi分别降低55.9%和27.0%），这一点很明显。有趣的是，MBL的氧化显著降低了巨噬细胞的吞噬能力，使其低于对照水平。流式细胞术和免疫荧光显示，在存在pdMBL的情况下巨噬细胞清道夫受体（SRA1）的表达显著增加，而在存在oxMBL的情况下这种增加被消除。我们表明，COPD中的肺巨噬细胞功能障碍可能至少部分是由氧化应激对MBL的诱导作用导致的，并为这种使人衰弱的疾病确定了另一种潜在的治疗策略。
