| Background and Aims :Limbal stem cell transplantanion(LSCT) is a greatly efficient method applied in treatment of ocular surface stem cells damage to restore the limbal tissue deficiency, supply the limbal stem cells population, treat limbal stem cell deficiency(LSCD) and reestablish the ocular surface structure. The limbal stem cells autografting has a high survival rate without immunological rejection. For patients with the single eye's limbal tissue deficiency, the size of ocular surface stem cells tissue taken from the healthy eye is still investigative to make sure that the donor eye can heal naturally. The fibrin sealant(FS), which is a new type of biomaterial without toxicity, cancerogenesis, mutagenic action and teratogenic action, has been proved to have effects of hemostasis and tissue adhesion. FS also has an optimal applied prospective in ophthalmology field for its good biocompatibility,biodegradation and not causing inflammation, foreign body sensation and tissue necrosis. In this study, we excised different size of rabbit ocular surface stem cells tissue, and determined a safety excision range by observations of ocular surface morphological change, corneal epithelium impression cytology examination,and tissue pathological change. Meanwhile, we explored the feasibility of applying FS in the LSCT instead of nylon sutures by observations of corneal tissue pathology, transmission electron microscope(TEM) and immunohistochemistry, which may provide an experimental base for FS's clinic application.Methods:1. To eatablish the animal model, excise the rabbit limbal stem cell tissues respectively at the ranges of 30°,60°,90°and up to 210°.2. Observe the corneal opacity,epithelium fluorescein stain,corneal neovascularization, by slitlamp, and the corneal epithelial cells'phenotype and pathological change by impression cytology examination and HE stain. Determine the safety excision range of limbal stem cell tissue from the donor eyes.3. Establish the rabbit model of single eye's partial LSCD, and perform a surgery of limbal stem cells autografting(healthy eyes as the donors) applying the FS(group A) or common nylon sutures (group B) randomly. Compare the progress of the corneal epithelium healing between the two groups by fluorescein stain within 3 days after surgery. Compare the corneal opacity and neovascularization between the two groups by slitlamp respectively at 7,14 and 30 days after surgery.4. Observe the donor eyes'corneal opacity, epithelium fluorescein stain and corneal neovascularization of the two groups by slitlamp at 14 and 30 days after surgery.5. Compare the corneal tisuue pathological change and the expression of special corneal epithelial keratin3 and P63 between the two groups at 7,14 and 30 days after surgery by HE stain and immunohistochemistry, and observe the ultramicrostructure change by TEM at 30 days after surgery.Results:1. When the angle of harvesting limbal stem cells autograft exceeded 150°,the corneal opacity, persistent corneal epithelia defect, neovascularization and conjunctival epithelium ingrowth,according to the comparison of indexes scores, the differeces were statistical significance (P=0.000<0.01).2. Impression cytology examination of the cornea which emerge decompensation of limbal stem cell deficiency,showed there were conjunctival caliciform cell which were positive of PAS stain.3. The time of corneal epithelium recover in the FS group was shorter than that in the suture group(P=0.044<0.05). There are significant difference between the FS group and the suture group in restraining neovascularization (P=0.043<0.05).4. On day 7,the expressions of keratin3 in the FS group and the suture group were obviously lower than the normal group(P<0.01),the FS group was higher than the suture group in IOD of keratin3(P<0.05).The FS group and the suture group compared with normal group were not significant difference at 30 days after surgery(P>0.05).5. The quantity of the expression of P63 protein obviously increased in the FS group and the suture group at 7,14and 30days after surgery,The expression of P63 protein which compared with normal group were significant difference(P<0.01). But it is no significant difference between the FS group and the suture group(P>0.05).Conclusions:1. The safety margin of harvesting limbal stem cells autograft was smaller than or equal to 150°.If the angle exceeded 150°,the decompensations of limbal stem cells couldn't be corrected.2. The treatment of autograft for limbal stem cell deficiency with fibrin sealant is feasible. Fibrin sealant is satisfactory biocompatible materials and completely degradation materials. It can relieve the inflammatory reaction, and reduce the growth of blood vessels.3. Fibrin sealant does not influence proliferation of limbal stem cells and growth of limbal stem cell grafts.
|
PDF Full Text Request