Figure 2
Obliterative bronchiolitis (OB) patients had higher amounts of fibrocytes in lung tissue. Fibrocytes were identified by fluorescence microscopy of lung tissues, by combining detection of one of the haematopoetic markers (CXCR4, CD45R0, or CD34) with detection of prolyl 4-hydroxylase. Panels a-e show a fibrocyte situated in the lumen of the alveolar structure. Panels a, b, and c show the separate markers for CXCR4, prolyl 4-hydroxylase, and nuclear staining, while panel d represents a merged image of the fibrocyte and panel e shows a differential interference contrast image. Panels f-j show a fibrocyte identified in parenchymal tissue: panels f, g and h show the separate markers for CD45R0, prolyl 4-hydroxylase, and nuclear staining, while panel i is a merged image of the fibrocyte and panel j is a differential interference-contrast image. Panels k-o show two fibrocytes (indicated with arrows), which were situated 32 μm and 45 μm from the endothelial layer. Panels k-m show the separate markers for CD34, prolyl 4-hydroxylase, and nuclear staining, while panel n is a merged picture of the fibrocyte and panel o is a differential interference-contrast image. Controls for unspecific binding of antibodies and background fluorescence were included in all experiments. Scale bars (10 μm) are indicated on the differential interference-contrast images. Original magnification: 40×. Panel p represents the numbers of fibrocytes identified in lung tissue from obliterative bronchiolitis patients and healthy controls. All three staining combinations showed significantly higher levels of fibrocytes in the obliterative bronchiolitis group as compared to the controls. In the obliterative bronchiolitis group, significantly more fibrocytes were stained with the detection of CXCR4 and prolyl 4-hydroxylase than with the detection of CD34 and prolyl 4-hydroxylase. Lumen of vessel is indicated as LV. Prolyl 4-hydroxylase is indicated as P4-OH. Lung transplanted, closed circle; bone marrow transplanted, open circle; control, circle with dot.