[Battle with herpes for 37 years]

Abstract

Herpes simplex virus type 1(HSV-1) remains latent in the human trigeminal ganglion after primarily infecting the cornea and conjunctiva. Mental stress, heat stimulation, ultraviolet ray and immunosuppression are among the reactivating factors of HSV-1, which can lead to epithelial herpetic keratitis, stromal herpetic keratitis, and other complications. I have been working with HSV-1 for a long time, concentrating especially on its latency and reactivation. I would like to introduce some of the recent research results. 1. Herpetic keratitis cases at the Department of Ophthalmology, Kinki University. There were 129 eyes of 128 patients who visited the Cornea Service in our university hospitals at Osayasayama, Sakai and Nara over 13 years and were diagnosed with herpetic keratitis and followed up for at least one year. They were investigated as to the type of herpetic keratitis at the initial visit and its recurrence. Initial types of herpetic keratitis and number of eyes of each type were: Epithelial type, 65 eyes (50%); Stromal type, 30 eyes (23%); Combined epithelial and stromal types, 18 eyes (14%). Recurrence was seen in 47% of the total 129 eyes. Recurrent cases of the epithelial type were mostly epithelial type. Frequently recurrent cases of the stromal type presented with repeated epithelial, stromal, and combined types. 2. Effects of antiherpetics on mouse epithelial herpetic keratitis. Acyclovir (ACV) eye ointment is usually prescribed for several weeks to treat human epithelial herpetic keratitis. Our question is: Is this long administration really necessary? To find the answer to this question, we investigated time-dependent effects of antiherpetics on mouse epithelial herpetic keratitis. Mouse corneas were infected with HSV-1 and either ACV eye ointment, oral valaciclovir (VACV) or oral famciclovir (FCV) was administered. No virus was detected in the tear fluid examined by viral culture 4 days after start of ACV eye ointment or oral VACV and 6 days after start of oral FCV. Real-time PCR revealed significant decrease of HSV DNA copy number in the eyeball or trigeminal ganglion compared to saline instillation 4 and 6 days after start. These results suggest that antivirals for 5 days could sufficiently decrease the HSV amount in the ocular surface and eyeball. 3. Corneal latency. In order to prove latency of HSV in the human cornea, virological and molecular biological techniques were used to ensure the following 3 prerequisites. 1) Positive HSV DNA in the human cornea. 2) Negative homogenate, positive explant. 3) Only latency-associated transcript (LAT) detected and transcriptional products of other virus genes (α, β, γ) not detected in the cornea. As a result, all the 3 prerequisites have been satisfied in the 3 corneas that had a past history of herpetic keratitis. This result suggests that HSV could remain latent in the human cornea. 4. Detection of HSV-1, HHV-6, and HHV-7 DNA in the anterior segment and aqueous humor using multiplex real-time PCR. Multiplex real-time PCR was applied for the first time ever in opththalmology to human herpes virus 6 (HHV-6) and 7(HHV-7). Samples taken from tear fluid before and 3 days after phacoemulsification and aspiration (PEA) or penetrating keratoplasty (PKP), and aqueous humor aspirated during PEA were used. The results of multiplex real-time PCR showed HSV-1, HHV-6 and HHV-7 DNA present in tear fluid both before and after PEA or PKP. 5. Gene expression when reactivation is suppressed. Nuclear factor-κB (NF-κB) has recently been reported to be involved in reactivation of HSV-1. IkappaB kinase-β (IKK2) inhibitors, which inhibit the activity of NF-κB, were used to examine gene expression during HSV reactivation in a mouse model. Significant decrease of HSV DNA copy number was observed at the trigeminal ganglion with real-time PCR in a group which was given IKK2 inhibitors intraperitoneally. Microarray method demonstrated 2-fold or more increased expression of 1812 probe. By Pathway analysis, eased immunosuppressive effects were observed in the group which was given IKK2 inhibitor intraperitoneally. 6. Immunoresponse involved in herpetic keratitis. Chemokine expression profiles in human corneal herpetic cases and mouse herpetic keratitis were analyzed. The results were similar to previously published reports: Cxcl9, Cxcl10, Ccl5, which are Th1 type chemokines, and Ccl20, a Th17 type chemokine, were observed to increase. On the other hand, Th2 type chemokine did not show an increase. Immunoresponse occurred mainly in the trigeminal ganglion. With these results, we suggest herpetic keratitis could be prevented by actively inducing Th17 type immunoresponse.