Demonstration of Kaposi's sarcoma-associated herpes virus cyclin D homolog in cutaneous Kaposi's sarcoma by colorimetric in situ hybridization using a catalyzed signal amplification system.

Kaposi's sarcoma-associated herpes virus (KSHV)/human herpes virus 8 (HHV8) DNA sequences have been demonstrated in Kaposi's sarcoma (KS), as well as in some acquired immunodeficiency syndrome (AIDS)-related non-Hodgkin's lymphomas (NHL) and in multicentric Castleman's disease. Although KSHV DNA generally is abundant in KSHV-associated lymphomas, few copies of the virus are present in KS, a property that confounds detection by in situ methods. Previous in situ studies, which identified KSHV in lesions of KS, relied on the use of polymerase chain reaction (PCR) to amplify target DNA sequences before in situ hybridization (ISH) for localization or used ISH with radioactively-labeled probes to obtain adequate levels of detection sensitivity. In this study, a novel nonisotopic nucleic acid ISH method using catalyzed signal amplification and colorimetric detection without PCR-dependent target amplification was used to identify KSHV-specific sequences. The level of sensitivity was increased further by using a probe that detects viral cyclin D homolog transcripts, which are expressed at significant levels during latent viral infection. Thirty cutaneous lesions of KS (25 AIDS-related and five classical European type) were evaluated. AIDS-related NHL and cell lines derived from patients with AIDS-related NHL, all of which were known to harbor KSHV by Southern blot analysis, were used as positive controls. NHL and benign cutaneous vascular lesions not associated with AIDS were used as negative controls. For each of the 30 KS lesions studied, hybridization signals were detected in most of the spindle cells surrounding the atypical slit-like vascular channels and also were detected in some endothelial cells in well-formed blood vessels in the perilesional dermis. Plaque and nodular lesions generally contained more labeled cells than did early patch lesions. All AIDS-related NHL and cell lines contained KSHV-specific sequences; however, the non-AIDS-related NHLs and benign vascular lesions were negative. These results confirm the presence of KSHV sequences in cutaneous KS and provide in situ evidence of infection by this virus in early patch-stage lesions. This study also defines the in situ expression of the KSHV cyclin D homolog viral oncogene in cutaneous KS. The use of this sensitive nonisotopic ISH method should allow detection of other KSHV-specific gene products, further defining the pathobiology of this virus.