Myeloid cells in infantile hemangioma

Abstract

Little is known about the pathogenesis of infantile hemangiomas despite the fact that they are relatively common tumors. These benign neoplasms occur in as many as 1 in 10 births, and although rarely life threatening, hemangiomas can pose serious concerns to the cosmetic and psychosocial development of the afflicted child. Ulceration, scarring, and disfigurement are significant problems as are encroachment of the ear and eye, which can threaten hearing and vision. The precise mechanisms controlling the rapid growth observed in the first months of life and the spontaneous involution that follows throughout the course of years remain unknown. In this report we demonstrate the presence of large numbers of hematopoietic cells of the myeloid lineage in proliferating hemangiomas and propose a mechanism for the observed evolution of these lesions that is triggered by hypoxia and involves the participation of myeloid cells. We report the results of experiments using myeloid markers (CD83, CD32, CD14, CD15) that unexpectedly co-labeled hemangioma endothelial cells, providing new evidence that these cells are distinct from normal endothelium.

Figure 1

Hematopoietic cells are abundant in proliferating hemangioma and decrease during involution. a–c: Three independent cases of the indicated ages are shown labeled with an antibody against CD45 (leukocyte common antigen), which is expressed on all cells of hematopoietic origin except erythrocytes. Hematopoietic cells (red) are found throughout the lesions. Nuclei are stained blue. Inset shows negative labeling with an isotype-matched control antibody. Scale bar, 100 μm.

Figure 2

Hemangioma-associated hematopoietic cells are not dividing. a: A proliferating hemangioma was labeled with CD45 (blue) to identify hematopoietic cells, ulex lectin (red) to show endothelium, and Ki67 (green) to show dividing cells. These results demonstrate that cellular proliferation is primarily restricted to hemangioma endothelium. b: In parallel, an involuting case was labeled showing significant reduction in the number of CD45⁺ cells and Ki67 staining, indicating diminished proliferation and demonstrating a correlation between hematopoietic cells and proliferation. Scale bar, 100 μm.

Figure 3

Hemangioma-associated hematopoietic cells are positive for the monocyte marker CD14 and correlate with proliferation. a–h: Sections from hemangiomas in all stages of progression were labeled with anti-CD14 antibody (red). The earliest lesions show the largest number of CD14⁺ cells with dramatic reduction in number as the lesions leave the proliferative phase and begin to involute. Note that e is chronologically out of sequence in this series, but clearly is less involuted than f–h. Nuclei are shown in blue. Percentages indicate estimated content of CD14⁺ cells obtained as described in Materials and Methods. Scale bar, 100 μm.

Figure 4

Hemangioma endothelial cells co-label with a dendritic cell marker, CD83. a and b: Proliferative hemangioma sections were labeled with an antibody against CD83 (green), and either anti-von Willebrand factor (vWF) (a) or Ulex europaeus lectin (ulex) (b) to label endothelial cells (red). Significant, although incomplete, co-localization was observed between CD83 and the endothelium. Each label is shown individually and at higher magnification to more readily visualize co-localization. c: Despite other histopathological similarities, this co-localization was not observed in pyogenic granuloma although a small number of CD83⁺ cells were observed in these lesions (green). Scale bar, 100 μm.

Figure 5

Normal dendritic cells are present in proliferating hemangioma. Using two markers of dendritic cells HLA-DR (a) and DC-SIGN (b), we demonstrated that proliferating lesions contained significant numbers of these cells. Nuclei are shown in blue. Scale bar, 100 μm.

Figure 6

Hemangioma endothelial cells express the myeloid marker CD15. a: Labeling sections of proliferative hemangiomas with anti-CD15 (red) showed an endothelial-like distribution. b: Labeling sections with both anti-CD15 (red) and anti-CD83 (green), which was shown to label endothelial cells (Figure 4), shows that CD15 and CD83 are co-localized on many cells. c: Pyogenic granuloma did not show endothelial staining with anti-CD15, but rather the expected leukocyte staining (red). Scale bars: 100 μm (a); 50 μm (b); 10 μm (c).

Figure 7

GLUT-1-positive hemangioma endothelial cells co-label with the dendritic cell marker, CD83. Sections were triple labeled with a marker for endothelium (ulex lectin) (a), hemangioma-specific endothelium (GLUT-1) (b), and dendritic cells (CD83) (c). As shown in the merged image (d), widespread co-localization is observed. Scale bar, 100 μm.

Figure 8

The myeloid marker CD32 (FcγRII) labels both hemangioma-associated hematopoietic cells and hemangioma endothelium. a: When proliferative hemangioma sections were labeled with antibodies against the monocyte marker CD14 (blue), and the myeloid marker CD32 (red), co-localization is observed on some cells (purple). b: Co-staining sections with CD32 (red) and the endothelial marker CD31 (green) reveals co-expression of these two antigens on a portion of cells within the lesion (yellow). c: When a and b are merged, cells co-labeling with CD14 (blue) and CD32 (red) are seen in purple whereas cells co-expressing CD32 (red) and CD31 (green) are yellow. Note that some CD31⁺ cells (green) do not co-label with CD32. Scale bar, 100 μm.