Wang Xuanding, Liu Fuguang, Bewig Burkhard
Department of Pulmonology, Second Affiliated Hospital of Medical School, Zhejiang University, Hangzhou 310009, China.
Chin Med J (Engl). 2002 Jan;115(1):76-80. doi: 10.3901/jme.2002.supp.076.
To investigate the expression of the granulocyte-macrophage colony-stimulating factor (GM-CSF) and GM-CSF/IL-3/IL-5 receptor common beta chain (beta c receptor) in an adult patient with idiopathic pulmonary alveolar proteinosis (PAP), so as to demonstrate the possible association of the GM-CSF and beta c receptor with the pathogenesis of human PAP.
The GM-CSF levels were measured with a commercial ELISA kit (sensitivity 5 pg/ml) and the beta c receptor expression on the cell surface was detected by flow cytometry analysis. Reverse transcription polymerase chain reaction (RT-PCR) analysis was employed to detect the expression of the GM-CSF mRNA and the beta c receptor mRNA in peripheral blood mononuclear cells and alveolar macrophages. The entire coding regions of the GM-CSF cDNA and the beta c receptor cDNA were sequenced by the Sanger dideoxy-mediated chain termination method to detect possible mutations.
The patient with PAP failed to release the GM-CSF protein either from circulating mononuclear cells or from alveolar macrophages. The expression of the GM-CSF mRNA was normal after the stimulation of lipopolysaccharide, whereas a point mutation at position 382 of the GM-CSF cDNA from "T" to "C" was revealed by cDNA sequencing, which caused a change in amino acid 117 of the protein from isoleucine to threonine. The beta c receptor expression on the cell surface was normal, and the beta c receptor mRNA expression and the sequence of the entire coding region of the beta c receptor were also normal.
The decreased GM-CSF production is associated with the pathogenesis of human PAP. A point mutation of the GM-CSF cDNA may contribute to the decreased GM-CSF production in our adult PAP patient. The mutation of the beta c receptor in some of paediatric patients with PAP may not be a common problem in adult patients.
研究粒细胞-巨噬细胞集落刺激因子（GM-CSF）及GM-CSF/IL-3/IL-5受体共同β链（βc受体）在一名成年特发性肺泡蛋白沉积症（PAP）患者中的表达，以证明GM-CSF和βc受体与人类PAP发病机制之间的可能关联。
使用商用ELISA试剂盒（灵敏度5 pg/ml）测量GM-CSF水平，并通过流式细胞术分析检测细胞表面βc受体的表达。采用逆转录聚合酶链反应（RT-PCR）分析检测外周血单核细胞和肺泡巨噬细胞中GM-CSF mRNA和βc受体mRNA的表达。通过桑格双脱氧介导的链终止法对GM-CSF cDNA和βc受体cDNA的整个编码区进行测序，以检测可能的突变。
PAP患者无论是循环单核细胞还是肺泡巨噬细胞均未能释放GM-CSF蛋白。脂多糖刺激后GM-CSF mRNA的表达正常，但cDNA测序显示GM-CSF cDNA第382位的点突变，由“T”变为“C”，导致蛋白质第117位氨基酸由异亮氨酸变为苏氨酸。细胞表面βc受体的表达正常，βc受体mRNA的表达及βc受体整个编码区的序列也正常。
GM-CSF产生减少与人类PAP的发病机制有关。GM-CSF cDNA的点突变可能导致我们的成年PAP患者GM-CSF产生减少。βc受体突变在一些儿童PAP患者中可能存在，但在成年患者中并非常见问题。
