Date of Award
Medical Doctor (MD)
Nils A. Loewen
Purpose: To establish a rat in vivo model of trabecular meshwork (TM) ablation and regeneration. Healthy TM cells are vital to regulate pressure. Ablation may be a useful strategy in glaucoma by reducing outflow resistance and possibly allowing dormant stem cells from adjacent structures to repopulate this structure.
Methods: Inducible, trackable, cytotoxic feline immunodeficiency viral vectors HSVtkiG were produced that expressed herpes simplex virus 1 thymidine kinase (HSVtk) and IRES-mediated eGFP, while the control vector GINSIN expressed cap-dependent eGFP. Filtered vectors were first used to transduce CrFK, GTM3, and NTM5 cells in vitro to confirm that ablation was feasible. Rats then received an intracameral vector into into one eye (GINSIN n=10, HSVtkiG n=13), followed by ganciclovir (GCV) administration to trigger ablation. Intraocular pressure (IOP), central corneal thickness (CCT) and slit lamp exams were performed daily. Successful transduction was confirmed by direct gonioscopic visualization of eGFP expression and its disappearance following induction of ablation. Anterior segment histology was obtained at different time points.
Results: Durable and high-grade transgene expression in the TM was achieved both in vitro and in vivo in the rat model, followed by effective removal of the transduced TM cells using ganciclovir. Ablation resulted in an IOP decrease of 25% in HSVtkiG injected eyes 2 days after GCV. No change was observed in the GINSIN controls and non-injected eyes (n=11, P<0.05). The effect persisted for 6 weeks and then regressed. Decreased cellularity was noted at the time of lowest IOP.
Conclusion: We demonstrated successful ablation of TM in vitro and in vivo using a conditionally cytotoxic FIV vector. Selective ablation of TM cells in the rat lead to a statistically significant decrease in IOP without damaging surrounding tissue. This model may be useful to further study the TM, its stem cells and cell migration patterns.
Zhang, Ze, "Outflow Tract Ablation Using A Conditionally Cytotoxic Feline Immunodeficiency Viral Vector" (2013). Yale Medicine Thesis Digital Library. 1851.