Herpes Simplex Virus Type-1 Infection of Corneal Epithelial Cells Induces Apoptosis of the Underlying Keratocytes

doi:10.1006/exer.1996.0266

Experimental Eye Research

Volume 64, Issue 5, May 1997, Pages 775-779

https://doi.org/10.1006/exer.1996.0266 Get rights and content

Abstract

The purpose of this study was to determine whether primary corneal infection with Herpes simplex virus (HSV)-1 induces keratocyte apoptosis in the rabbit. New Zealand white rabbit eyes were inoculated with HSV-1 strain 17 Syn⁺. Rabbits that developed slit lamp signs of epithelial infection were killed between 12 and 120 hr post infection. One cornea of each animal was fresh-frozen for TUNEL assay to detect DNA fragmentation in situ. The other cornea was fixed for transmission electron microscopy (TEM). Mechanical scrape wounded rabbit corneas were included as positive controls. DNA fragmentation consistent with apoptosis was detected in anterior keratocytes of corneas at 18, 24, and 48 hr after primary infection with HSV-1 and 2 hr after an epithelial scrape, but not in unwounded control corneas. Electron microscopic evidence of keratocyte apoptosis that included chromatin condensation, chromatin fragmentation, and cellular blebbing with formation of membrane bound cell fragments was detected in mechanical scrape wounded corneas and infected rabbit corneas at 12, 18, 24, 48, and 120 hr after infection, but not in unwounded control corneas. This study suggests that anterior stromal keratocyte apoptosis occurs following primary HSV-1 infection of the corneal epithelium. Previous studies have demonstrated that corneal epithelial scrape wounds induce apoptosis in the underlying keratocyte cells. We hypothesize that soluble mediators released by epithelial injury mediate anterior keratocyte apoptosis and that one of the physiologic functions of this epithelial-stromal apoptosis system is to limit viral extension.

References (0)

Cited by (87)

Corneal wound healing
2020, Experimental Eye Research
Citation Excerpt :
Viruses such as herpes simplex virus (HSV) or smallpox virus have the potential for this type of spreading from surface epithelium to other structures in the eye and brain. This hypothesis was supported by experiments in rabbits performed with the late virologist James M. Hill, PhD where the corneal epithelium was infected with herpes simplex virus-1 strain 17 Syn + without scarification and the underlying keratocytes underwent apoptosis detected with the TUNEL assay and TEM (Wilson et al., 1997). This apoptosis system was subsequently shown in collaboration with George R. Stark, PhD to be defective in STAT1 null mice (Mohan et al., 2000) and when mice had a cornea infected with HSV more than 70% of the STAT1 knockout mice, but 0% of the control mice, died within a few days from HSV encephalitis (Hill JM, Wilson SE, unpublished data, 2001).
The corneal wound healing response is typically initiated by injuries to the epithelium and/or endothelium that may also involve the stroma. However, it can also be triggered by immune or infectious processes that enter the stroma via the limbal blood vessels. For mild injuries or infections, such as epithelial abrasions or mild controlled microbial infections, limited keratocyte apoptosis occurs and the epithelium or endothelium regenerates, the epithelial basement membrane (EBM) and/or Descemet's basement membrane (DBM) is repaired, and keratocyte- or fibrocyte-derived myofibroblast precursors either undergo apoptosis or revert to the parent cell types. For more severe injuries with extensive damage to EBM and/or DBM, delayed regeneration of the basement membranes leads to ongoing penetration of the pro-fibrotic cytokines transforming growth factor (TGF) β1, TGFβ2 and platelet-derived growth factor (PDGF) that drive the development of mature alpha-smooth muscle actin (SMA)+ myofibroblasts that secrete large amounts of disordered extracellular matrix (ECM) components to produce scarring stromal fibrosis. Fibrosis is dynamic with ongoing mitosis and development of SMA + myofibroblasts and continued autocrine-or paracrine interleukin (IL)-1-mediated apoptosis of myofibroblasts and their precursors. Eventual repair of the EBM and/or DBM can lead to at least partial resolution of scarring fibrosis.
Posterior stromal cell apoptosis triggered by mechanical endothelial injury and basement membrane component nidogen-1 production in the cornea
2018, Experimental Eye Research
Citation Excerpt :
This finding had an enormous impact on research on the corneal wound healing response and stromal changes that ensue following injuries, surgeries, infections and diseases of the cornea that include damage to the corneal epithelium (Wilson and Kim, 1998; Wilson et al., 2001; Ljubimov and Saghizadeh, 2015; Torricelli et al., 2016). Subsequent studies demonstrated this is likely a system that evolved as an immediate cellular response to retard the extension of viral infections, such as herpes simplex virus keratitis, to the stroma and into the eye prior to mobilization of the immune system to fight the threatening infection (Wilson et al., 1997). Subsequent work also demonstrated this programmed cell death is likely modulated by the Fas/Fas ligand system triggered by interleukin (IL)-1, and possibly other cytokines, released from the injured corneal epithelium (Inoue, 2014).
This study was performed to determine whether cells in the posterior stroma undergo apoptosis in response to endothelial cell injury and to determine whether basement membrane component nidogen-1 was present in the cornea. New Zealand White rabbits had an olive tip cannula inserted into the anterior chamber to mechanically injure corneal endothelial cells over an 8 mm diameter area of central cornea with minimal injury to Descemet's membrane. At 1 h (6 rabbits) and 4 h (6 rabbits) after injury, three corneas at each time point were cryopreserved in OCT for terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and immunohistochemistry (IHC) for vimentin and nidogen-1, and three corneas at each time point were fixed for transmission electron microscopy (TEM). Uninjured corneas were controls. Stromal cells over approximately the posterior 25% of the stroma overlying to the site of corneal endothelial injury underwent apoptosis detected by the TUNEL assay. Many of these apoptotic cells were vimentin+, suggesting they were likely keratocytes or corneal fibroblasts. Stromal cells peripheral to the site of endothelial injury and more anterior stromal cells overlying the site of endothelial injury did not undergo apoptosis. Stromal cell death was confirmed to be apoptosis by TEM. No apoptosis of stromal cells was detected in control, uninjured corneas. Nidogen-1 was detected in the stroma of unwounded corneas, with higher nidogen-1 in the posterior stroma than the anterior stroma. After endothelial scrape injury, concentrations of nidogen-1 appeared to be in the extracellular matrix of the posterior stroma and, possibly, within apoptotic bodies of stromal cells. Thus, posterior stromal cells, likely including keratocytes, undergo apoptosis in response to corneal endothelial injury, analogous to anterior keratocytes undergoing apoptosis in response to epithelial injury.
Heat shock induces apoptosis through reactive oxygen species involving mitochondrial and death receptor pathways in corneal cells
2011, Experimental Eye Research
Although many studies have been performed to elucidate the molecular consequences of ultraviolet irradiation, little is known about the effect of infrared radiation on ocular disease. In addition to photons, heat is generated as a consequence of infrared irradiation, and heat shock is widely considered to be an environmental stressor. Here, we are the first to investigate the biological effect of heat shock on Statens Seruminstitut Rabbit Cornea (SIRC) cells. Our results indicate that heat shock exhibits effective cell proliferation inhibition by inducing apoptosis. Heat shock triggers the mitochondrial apoptotic pathway indicated by a change in Bax/Bcl-2 ratios, resulting in caspase-9 activity. In addition, heat shock triggered the death receptor apoptotic pathway indicated by a change in Fas ligand expression, resulting in caspase-8 activity. Furthermore, we also found that generation of reactive oxygen species (ROS) is a critical mediator in heat shock-induced apoptosis. In addition, the antioxidant vitamin C significantly decreased heat shock-mediated apoptosis. Taken together, these findings suggest a critical role for ROS involving mitochondrial and death receptor pathways in heat shock-mediated apoptosis of cornea cells.
In vivo confocal microscopic evaluation of morphologic changes and dendritic cell distribution in pterygium
2010, American Journal of Ophthalmology
Citation Excerpt :
Keratocyte apoptosis has been reported to be the earliest stromal response following epithelial injury.28 This has been reported in viral infection29 and in various corneal surgical procedures such as photorefractive keratectomy (PRK) and laser in situ keratomeliusis (LASIK).30 Interleukin-1 (IL-1) is considered to be the key modulator of keratocyte apoptosis.31
To observe morphologic changes and the distribution of dendritic cells in pterygium using in vivo laser scanning confocal microscopy (LSCM).
Prospective comparative study.
Twenty-six eyes of 26 patients with pterygium and 17 eyes of 17 healthy subjects were recruited. Using LSCM, in vivo images of the pterygium and adjacent clear cornea were captured. The density of basal corneal epithelial cells and keratocytes in the anterior and posterior stroma and the density of dendritic cells in the pterygium and adjacent clear cornea were determined. In the controls, the central cornea and nasal bulbar conjunctiva were imaged. The density of basal corneal epithelial cells, keratocytes, and dendritic cells was evaluated.
Morphologic alterations of the sub-basal nerve plexus were observed in pterygium. The density of basal corneal epithelial cells and anterior keratocytes in pterygium was 5359.0 ± 543.1 cells/mm² and 407.4 ± 188.7 cells/mm² respectively, which was significantly lower than that in the controls (P < .001). The density of dendritic cells in the clear corneas of pterygia was 60.3 ± 25.5 cells/mm², which was significantly higher than the 23.6 ± 11.1 cells/mm² in the central corneas of controls (P < .001). The dendritic cell density in the pterygium was significantly higher than the density in the nasal bulbar conjunctiva of controls (P < .001).
Histopathologic alterations and increased dendritic cells were evident in pterygium and the adjacent clear cornea by in vivo LSCM. In vivo LSCM was found to be an effective method of observing the morphologic alterations of pterygium.
Wound healing after laser in situ keratomileusis and photorefractive keratectomy
2010, Ocular Disease: Mechanisms and Management Expert Consult
Wound healing after laser in situ keratomileusis and photorefractive keratectomy
2010, Ocular Disease: Mechanisms and Management

View all citing articles on Scopus

^f1: For correspondence at: Eye Institute/A31, The Cleveland Clinic Foundation, 9500 Euclid Ave., Cleveland, OH 44195, U.S.A.

View full text

Regular articleHerpes Simplex Virus Type-1 Infection of Corneal Epithelial Cells Induces Apoptosis of the Underlying Keratocytes

Abstract

Regular article
Herpes Simplex Virus Type-1 Infection of Corneal Epithelial Cells Induces Apoptosis of the Underlying Keratocytes