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Conjunctival epitheliopathy induced by topical exposure to bacterial peptidoglycan, muramyl dipeptide
Marlyn P Langford1, Lana Srur1, Thomas B Redens1
1Department of Ophthalmology, Louisiana State University Health Sciences Center Shreveport, LA, 71130, USA.
Abstract:
Noninfectious exudative conjunctivitis can be experimentally produced in rabbits by application of the apoptogenic bacterial cell wall peptidoglycan, muramyl dipeptide (MDP) to the ocular surface. The purpose of this study was to investigate the acute conjunctival cytopathology induced by unilateral ocular surface exposure to MDP. Hematoxylin and eosin staining assessed bilateral tear cytopathology and conjunctival histopathology. The caspases levels in conjunctival tissue and tears were measured in standard assays utilizing p-nitroanaline tagged caspase-specific substrates. Immunofluorescent antibody identified intracellular caspase-3, nuclear factor-κβ (NF-κβ), and oxidative DNA damage (8-OHdG; 8-oxo-2'-deoxyguanosine) in tear and conjunctiva cells. DNA extracted from conjunctival tissues and pooled tear fluids were visualized by ethydium bromide agarose gel electrophoresis. Onset of ipsilateral conjunctivitis was due to an epitheliopathy characterized by loss of conjunctival epithelial cell adherence, exuviation of conjunctival epithelial cells, and neutrophil infiltration. Caspase-3 levels were significantly higher in exuviated cells in ipsilateral than contralateral tear (p's ≤ 0.001) collected at 3-5 h post MDP. Significantly higher caspase-2, -3, -6, -8 and -9 (p's ≤ 0.03) levels were detected in ipsilateral than contralateral conjunctival tissue at 5 h. Polymeric DNA was detected in ipsilateral but not contralateral conjunctival tissue and tears. Caspase-3, NF-κβ, and 8-OHdG positive neutrophils were detected in bilateral conjunctiva and tear. The caspase-3/NF-κβ epithelial cells and polymeric DNA in conjunctival tissue and shedding of caspase positive cells and polymeric DNA into ipsilateral tears support MDP induction of acute programmed cell death in vivo. The results suggest that ipsilateral exudative conjunctivitis is due to acute caspase-mediated conjunctival epitheliopathy induced by topical exposure to the bacterial peptidoglycan MDP.
Insights
Muramyl dipeptide (MDP) causes acute programmed cell death in rabbit eyes, leading to exudative conjunctivitis. This bacterial peptidoglycan triggers caspase-mediated epitheliopathy and DNA damage in conjunctival cells.
Area of Science:
- Ophthalmology
- Cell Biology
- Immunology
Background:
- Noninfectious exudative conjunctivitis can be induced by bacterial cell wall components.
- Muramyl dipeptide (MDP), a peptidoglycan fragment, is a known apoptogenic agent.
- Understanding the cellular mechanisms of MDP-induced conjunctivitis is crucial.
Purpose of the Study:
- To investigate the acute conjunctival cytopathology induced by unilateral ocular surface exposure to MDP.
- To elucidate the role of caspases and programmed cell death in MDP-induced conjunctivitis.
Main Methods:
- Rabbits were exposed to MDP on one ocular surface.
- Tear and conjunctival tissues were analyzed using hematoxylin and eosin staining.
- Caspase levels, NF-κβ, 8-OHdG, and DNA integrity were assessed via assays, immunofluorescence, and gel electrophoresis.
Main Results:
- MDP induced epitheliopathy characterized by cell adherence loss, shedding, and neutrophil infiltration.
- Significantly elevated caspase levels (caspase-2, -3, -6, -8, -9) were found in ipsilateral conjunctival tissue and tears.
- Polymeric DNA and caspase-positive neutrophils were detected in affected tissues and tears, indicating programmed cell death.
Conclusions:
- Topical MDP exposure induces acute caspase-mediated conjunctival epitheliopathy.
- Programmed cell death plays a significant role in the pathogenesis of MDP-induced exudative conjunctivitis.
- MDP acts as a potent inducer of ocular surface inflammation and cell death.
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