Schreiber Adrian, Otto Bjoern, Ju Xinsheng, Zenke Martin, Goebel Ursula, Luft Friedrich C, Kettritz Ralph
HELIOS Klinikum-Berlin, Franz Volhard Clinic and Max Delbrück Center for Molecular Medicine, Medical Faculty of the Charité, Humboldt University of Berlin, Berlin, Germany.
J Am Soc Nephrol. 2005 Jul;16(7):2216-24. doi: 10.1681/ASN.2004070609. Epub 2005 May 18.
A large membrane proteinase 3 (mPR3)-positive neutrophil subset (mPR3high) is a risk for Wegener's granulomatosis (WG). The relationship between mPR3 expression and clinical manifestations was investigated in 81 WG patients and mPR3 expression was studied in CD34+ stem cell-derived human neutrophils. The mPR3high neutrophil percentage correlated with renal function, anemia, and albumin at the time of presentation. The mPR3high neutrophil percentage and renal failure severity correlated directly after 5 yr. For elucidating mechanisms that govern mPR3 expression, studies were conducted to determine whether the genetic information that governs mPR3 expression resides within the neutrophils, even without stimuli possibly related to disease. CD34+ hematopoietic stem cells were differentiated to neutrophils, and their mPR3 expression was determined. A two-step amplification/differentiation protocol was used to differentiate human CD34+ hematopoietic stem cells into neutrophils with G-CSF. The cells progressively expressed the neutrophil surface markers CD66b, CD35, and CD11b. The ferricytochrome C assay demonstrated a strong respiratory burst at day 14 in response to PMA but none at day 0. Intracellular PR3 was detectable from day 4 by Western blotting. An increasing percentage of a mPR3-positive neutrophil subset became detectable by flow cytometry, whereas a second subset remained negative, consistent with a bimodal expression. Finally, human PR3-anti-neutrophil cytoplasmic autoantibodies induced a stronger respiratory burst, compared with human control IgG in stem cell-derived neutrophils. Taken together, these studies underscore the clinical importance of the WG mPR3 phenotype. The surface mPR3 on resting cells is probably genetically determined rather than being dictated by external factors.
一种大的膜蛋白酶3（mPR3）阳性中性粒细胞亚群（mPR3高表达）是韦格纳肉芽肿（WG）的一个危险因素。在81例WG患者中研究了mPR3表达与临床表现之间的关系，并在CD34+干细胞来源的人中性粒细胞中研究了mPR3表达。mPR3高表达中性粒细胞百分比与就诊时的肾功能、贫血和白蛋白相关。5年后，mPR3高表达中性粒细胞百分比与肾衰竭严重程度直接相关。为了阐明调控mPR3表达的机制，进行了研究以确定即使在没有可能与疾病相关的刺激的情况下，调控mPR3表达的遗传信息是否存在于中性粒细胞内。将CD34+造血干细胞分化为中性粒细胞，并测定其mPR3表达。采用两步扩增/分化方案，用粒细胞集落刺激因子（G-CSF）将人CD34+造血干细胞分化为中性粒细胞。细胞逐渐表达中性粒细胞表面标志物CD66b、CD35和CD11b。高铁细胞色素C试验显示，在第14天对佛波酯（PMA）有强烈的呼吸爆发反应，但在第0天没有。通过蛋白质印迹法从第4天开始可检测到细胞内的蛋白酶3（PR3）。通过流式细胞术可检测到mPR3阳性中性粒细胞亚群的百分比不断增加，而另一个亚群仍为阴性，这与双峰表达一致。最后，与干细胞来源的中性粒细胞中的人对照IgG相比，人PR3抗中性粒细胞胞浆自身抗体诱导更强的呼吸爆发。综上所述，这些研究强调了WG的mPR3表型的临床重要性。静息细胞表面的mPR3可能是由基因决定的，而不是由外部因素决定的。
