Building a hypothesis based upon clinical findings and verifying it through experiments is a real joy for clinician scientists. This review presents the principal work of our laboratory group's search for new concepts.
I. Struggling with corneal infection
The causative microorganisms of infectious keratitis changed over time along with the medical background. In the 1970s, herpes simplex virus (HSV) keratitis exerted an overwhelming influence and we put all our energies into studying the possibility of novel anti-viral therapies as well as subunit vaccines until the introduction of acyclovir in the late 1980s. Thereafter, corneal endotheliitis, a process of progressive corneal endothelial decompensation with mild inflammatory response in the anterior chamber, came to the forefront. HSV DNA was detected in the aqueous humor of one patient and our subsequent study in rabbits suggested that the pathogenesis of this disease may be operative due to anterior chamber-associated immune deviation (ACAID) against viral antigens. More recently, cytomegalovirus corneal endotheliitis has attracted increased attention.
Recent epidemiologic studies of infectious keratitis in Japan revealed that contact lens (CL)-related microbial keratitis is prevalent in the younger generations, underscoring the importance of appropriate contact lens care. The most frequently encountered pathogenic microorganism in CL associated keratitis is Acanthamoeba followed by Pseudomonas aeruginosa. Using a mouse keratitis model, we found that a MucD protease deficient mutant of P. aeruginosa was unable to elicit corneal infection and that MucD protease suppressed macrophage inflammatory protein 2 (MIP-2) during the early stages of the infection and inhibited neutrophil recruitment to the cornea, reconfirming the significance of primary defense mechanism in the pathogenesis of bacterial keratitis. As a new weapon to overcome Acanthamoeba keratitis, we demonstrated that methylene blue mediated photodynamic therapy is effective against Acanthamoeba in vitro and has good synergistic effects with polyhexamethylene biguanide which is topically used by patients.
II. Working to prevent infectious endophthalmitis following cataract surgery
Despite sophisticated surgical instruments and procedures, postoperative endophthalmitis remains a threatening complication in cataract surgery. We focused our research on the process that leads to bacterial contamination of the eye during surgery. Studies with porcine eyes using a visualizing agent demonstrated that administration of ophthalmic viscoelastic substances may be a major risk factor in inducing the contamination of the anterior chamber. Another experiment showed that an anterior hyaloid membrane tear (AHT) could be induced by increased intraocular pressure during hydrodissection in ex vivo porcine eyes and in vivo rabbit eyes. AHT resulted in breakdown of the barrier between the posterior chamber and the vitreous cavity which could allow microbial penetration. Our epidemiologic study of Enterococcus faecalis endophthalmitis demonstrates that visual outcomes can be divided into good and poor groups. Further experiments using a rabbit model showed that extracellular protease of E. faecalis may play a major role in damaging retinal vessels and could determine the visual prognosis.
III. Evaluating of the tear clearance
Tears secreted by the lacrimal glands provide a layer of moisture over the ocular surface and are then drained through the lacrimal duct, forming a flow called the "lacrimal river". To understand the pathogenesis of tear-related disorders, it is essential to evaluate the drainage efficiency as well as the composition of tear fluid. Focusing on the clearance mechanism for excessive tear fluid, we visualized Krehbiel flow by poly (methyl methacrylate) particles suspended in a fluorescein solution, and invented a new method of measuring tear clearance using anterior segment optical coherence tomography (AS-OCT). Our results show that the tear clearance rate is lower in older compared to younger subjects. In another study, we established an absolute quantitative platform of human tear fluid proteome using selected reaction monitoring mass spectrometry and wheat germ cell-free protein synthesis system. Our preliminary results show that tear fluid from older subjects contains multiple proteolytic enzymes and inflammatory products. These results indicate that, in younger subjects tear fluid is constantly being replaced with new fluid, while in older subjects old tear fluid may be recycled, which may influence the pathologic changes of the ocular surface.
IV. Taking a new look at the corneal edema
In a series of clinical cases we found that during the recovery phase from corneal endothelial disturbance corneal edema subsides first from the center and persists late at the periphery and, on the contrary, the initial phase of corneal endothelial dysfunction starts as localized corneal edema at the periphery. To elucidate the above clinical findings, we assumed the existence of water flow from the center to the periphery at the horizontal plane of the cornea and achieved preliminary results supporting this hypothesis using a corneal endothelial dysfunction model in rabbits.
Nippon Ganka Gakkai Zasshi (J Jpn Ophthalmol Soc) 118: 155-188, 2014.