Human papilloma virus in neoplastic and non-neoplastic conditions of the external eye

Abstract

Aim: Human papilloma virus (HPV) types 16 and 18 have been associated with neoplastic conditions of the conjuctiva. However, the presence of this virus has not been reported in non-neoplastic disorders of the external eye nor has it been studied in normal conjunctival tissues.

Methods: Ninety six paraffin embedded tissue specimens with neoplastic and non-neoplastic lesions and 19 conjunctiva samples free from overt disease were studied for HPV types 16 and 18 positivity with the polymerase chain reaction.

Results: HPV types 16 and 18 DNA were identified in 57% of in situ squamous cell carcinoma, in 55% of invasive squamous cell carcinoma, in 20% of climatic droplet keratopathy, in 35% of scarred corneas, and in 32% of normal conjunctival tissue obtained during routine cataract extractions.

Conclusion: These findings indicate that HPV types 16 and 18 are detectable with the polymerase chain reaction not only in epithelial neoplasms of the ocular mucous membrane but also in non-neoplastic lesions as well as in apparently healthy conjunctiva.

Figure 1

Detection of β globin gene. Ethidium bromide stained 2% agarose gel showing β globin gene amplification (110 bp product) following PCR amplification using PCO3/PCO4 primer and 1 µg of DNA as a template from the following samples: lane 1, positive control (normal blood lymphocytes); lane 2, invasive squamous cell carcinoma (SCC); lane 3, in situ SCC; lane 4, corneal scar; lane 5, conjunctiva; lane 6, negative control (no DNA); M, GelMarker; M₁, PhiX 174 RF Hae III marker.

Figure 2

First PCR amplification of HPV DNA: consensus primer for E6 ORF of HPV 16 and 18 was used as outer primer along with 1 µg of DNA as a template (307 bp product) from the following samples: lane 1, positive control for HPV 16 (CaSki cell DNA); lane 2, positive control for HPV 18 (HeLa cell DNA); lane 3, conjunctiva; lane 4, invasive squamous cell carcinoma (SCC); lane 5, in situ SCC; lane 6, negative control (no DNA); M, GelMarker; M₁, PhiX 174 RF Hae III marker.

Figure 3

Detection of HPV 16 DNA. HPV 16 specific inner primer was used along with 2 µl of the first PCR reaction mixture as a template. (A) Analysis of the amplification DNA products on 2% agarose gel stained with ethidium bromide (124 bp product). (B) Southern blot analysis of the amplification DNA products hybridised with non-radioactive labelled HPV 16 specific oligonucleotide probe for the following samples: lane 1, positive control (CaSki cell DNA); lane 2, conjunctiva; lane 3, conjunctiva; lane 4, invasive squamous cell carcinoma (SCC); lane 5, in situ SCC; lane 6, negative control (no DNA); M, GelMarker; M₁, PhiX 174 RF Hae III marker.

Figure 4

Detection of HPV 18 DNA. HPV 18 specific inner primer was used along with 2 µl of the first PCR reaction mixture as a template. (A) Analysis of the amplification DNA products on 2% agarose gel stained with ethidium bromide (188 bp product). (B) Southern blot of the amplification products hybridised with non-radioactive labelled HPV 18 specific oligonucleotide probe for the following samples: lane 1, positive control for HPV 16 (HeLa cell DNA); lane 2, conjunctiva; lane 3, invasive squamous cell carcinoma (SCC); lane 4, invasive SCC; lane 5, in situ SCC; lane 6, negative control (no DNA); M, GelMarker; M₁, PhiX 174 RF Hae III marker.