Journal: PLoS ONE

Article Title: Ex vivo model of herpes simplex virus type I dendritic and geographic keratitis using a corneal active storage machine

doi: 10.1371/journal.pone.0236183

Figure Lengend Snippet: (A) Chronology of the development of herpetic lesions on the corneas stored in the active storage machine (ASM) versus organ culture (OC). Bright field macroscopic observation (x1 and x6.3 zooms) after scratching (D0) and infection at D3, D4 and D5. In the OC cornea, the red arrows show the initial scratch; and the yellow arrows, the "buds" that appeared at the edges of the infected area. Note that the OC cornea presented a characteristic tessellated pattern (highlighted by retro illumination) corresponding to the numerous deep posterior folds caused by stromal edema. On the contrary, the ASM-stored cornea was thinner, presented no folds and thus appeared smooth. (B) Anti HSV-gB immunolabeling on unfixed whole human corneas. Observation with a fluorescence macro-zoom microscope (x1 and x6.3 zooms) of the corneal epithelium infected with HSV-1 in the ASM or OC and labeled with anti-HSV-gB and Hoechst after 5 days' incubation in the ASM or OC. Yellow asterisks show full-thickness epithelial ulcers where nuclei of Hoechst-positive keratocytes were visible through transparency. Yellow arrows show the rounded infected cells demonstrating the cytopathogenic effect (CPE). Note that the corneal limbus and the scleral rim were strongly autofluorescent on this wavelength, but this did not prevent corneal imaging. Scale bar: 1000 μm for low magnification images and 200 μm for high magnification images.

Article Snippet: After infection by HSV-1, corneas were monitored daily for 3 to 6 days (depending on the rate of lesion development) by imaging the epithelium under a macro zoom microscope with retro illumination (MVX-10 macroscope run by the CellSens Dimension imaging software, Olympus, Tokyo, Japan) ( ).

Techniques: Organ Culture, Infection, Immunolabeling, Fluorescence, Microscopy, Labeling, Incubation, Imaging