Detection of human papillomavirus in branchial cleft cysts

Abstract

High-risk human papillomavirus (HPV) DNA has been reported to be present in branchial cleft cysts, but further information is required to clarify the role of HPV infection in branchial cleft cysts. The presence of HPV, the viral load and the physical statuses in samples from six patients with branchial cleft cysts were investigated using the polymerase chain reaction (PCR), quantitative PCR, in situ hybridization (ISH) using HPV DNA probes and p16^INK4a immunohistochemical analysis. High-risk type HPV-16 DNA was identified in four of the six branchial cleft cysts analyzed. Of the HPV-positive branchial cleft cysts, three exhibited mixed-type integration of HPV. HPV DNA was distributed among the basal-to-granular layers of the cystic wall in ISH analysis, and p16^INK4a was weakly expressed in the nuclei and cytoplasm of the same layers in patients with integration. ISH revealed that one patient with episomal-type infection exhibited HPV DNA in the cyst wall and did not express p16^INK4a. Two patients without evidence of HPV infection exhibited weak p16^INK4a expression in the superficial cyst-lining cells of branchial cleft cysts. These results indicate that infection with high-risk HPV types may be common in branchial cleft cysts. In addition, p16^INK4a is not a reliable surrogate marker for HPV infection in branchial cleft cysts.

Keywords: branchial cleft cyst; human papillomavirus; in situ hybridization; p16INK4a expression; viral integration; viral load.

Figure 1.

Histology results in patient 1 with episomal-type human papillomavirus-16 infection. (A and B) Hematoxylin and eosin staining; (C and D) in situ hybridization (red arrows indicate a diffuse in situ hybridization signal); and (E and F) immunohistochemical staining for p16^INK4a. (A, C and E) Scale bar, 50 µm; and (B, D and F) scale bar, 10 µm.

Figure 2.

Histology results in patient 4 with mixed-type human papillomavirus-16 infection. (A and B) Hematoxylin and eosin staining; (C and D) in situ hybridization at (black and red arrows indicate punctate and diffuse signals, respectively); and (E and F) immunohistochemical staining for p16^INK4a. (A, C and E) Scale bar, 50 µm; and (B, D and F) scale bar, 10 µm.

Figure 3.

Histology results in patient 6, lacking human papillomavirus-16 infection. (A and B) Hematoxylin and eosin staining; (C and D) in situ hybridization; and (E and F) immunohistochemical staining for p16^INK4a. (A, C and E) Scale bar, 50 µm; and (B, D and F) scale bar, 10 µm.

Figure 4.

Histology results in the metastatic lymph node of a patient with HPV-associated oropharyngeal cancer (positive control). (A and B) Hematoxylin and eosin; (C and D) in situ hybridization (black and red arrows indicate punctate and diffuse signals, respectively); and (E and F) immunohistochemical staining for p16^INK4a. HPV, human papillomavirus. (A, C and E) Scale bar, 200 µm; and (B, D and F) scale bar, 10 µm.