HBV exists in the bloodstream and organs with blood flows. Since the cornea is an avascular area, it has been considered that keratoplasty can not transmit hepatitis B virus. However, in the 1990s, HoftRH et al. reported two cases of hepatitis B surface antigen positive conversion after receiving penetrating keratoplasty from hepatitis B surface antigen positive donors [8]. Ahmad Khalil et al [9], scraped off epithelium from HBsAg positive donor corneas and found HBV DNA in both epithelium and stroma-endothelium complex with ELISA. But it was not clear whether the stroma or the endothelium contained HBV DNA.With the emergence and development of nucleic acid amplification technology, real-time PCR technology was used to detect HBV DNA in donor uveas [10], Compared with ELISA, PCR is more convenient and reliable [11], but the sensitivity and specificity of PCR in detecting virus are satisfactory [12]. The specific location of the virus in the organization cannot be determined.
Since HBV exists in almost all fluid of the human body, including blood, tears, aqueous humor, etc [12, 13]. we thought cells in the cornea would very likely carry HBV DNA. In our study, immunofluorescent staining was used to localize HBsAg in paraffin sections. Our results showed that HBsAg can be detected in the corneal epithelium and/or endothelium in some of the HBsAg seropositive donors’ corneas. Though the presence of HBsAg in cornea epithelium and endothelium cannot prove the infectious mechanism, it laid the foundation for further studies investigating the possibility of using HBsAg positive donor cornea in lamellar keratoplasty. We suppose removing the epithelium and endothelium of donor cornea to prepare cornea lamellar for keratoplasty could be a good way to use HBsAg positive corneas.
Povidone iodine, a broad-spectrum disinfectant, is routinely used in the disinfection of donor corneas. When diluted in the water, povidone iodine releases free iodine, which destroys microorganisms by breaking their membranes and proteins [14]. In our study, HBsAg can still be detected after povidone iodine pretreatment. It is suggested that HBV is resistant to Povidone iodine. Studies showed that chemical agents such as chlorine containing agents, ethylene oxide, glutaraldehyde, peracetic acid, and iodophor can inactivate HBV [15]. However, these chemicals are highly cytotoxic and cannot be used for donor cornea disinfection. According to our results, though povidone iodine has a low cytotoxicity and strong disinfect capability, it cannot inactivate HBV. HBVsAg positive donor corneas disinfected with povidone iodine still have a high risk of HBV infection.
Another finding in the study is that HBVsAg could be detected in the HBsAg positive donor corneas that were preserved in glycerol for a year. Glycerol is a long-term preserving fluid routinely used to preserve donor corneas in the eye bank. This way of inactive preservation removes water from the cornea and inhibits enzyme activity and autolysis in corneal cells [16]. Therefore, it is comprehensible that even preserved in glycerine for a long time, HBVsAg can still be detected from HBsAg positive donor corneas. Though it still has a risk of HBV infection, considering that the HBsAg can be detected in both epithelium and endothelium, we suppose glycerine preserved HBsAg positive corneas can still be used for lamellar keratoplasty. By removing the virus-containing epithelium and endothelium, the risk of HBV infection would be largely reduced. But further studies are needed to verify our conjecture.