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Y. Du, E. C. Carlson, M. L. Funderburgh, D. E. Birk, W. W. Y. Kao, J. L. Funderburgh; Rescue of the Stromal Phenotype in Lumican Null Mice by Human Corneal Stem Cell Transplantation. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4522.
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Lumican null (Lum-/-) mice exhibit disorganized stromal matrix and corneal haze, thus providing an in vivo model for investigating therapy for corneal opacity. We previously described stem cells from human corneal stroma (CSSC) which adopt characteristics of keratocytes in vitro. In the current study we examine the potential for CSSC to correct the Lum-/- mouse stromal defect in vivo.
50,000 human CSSC were injected directly into corneal stromas of 14-week old Lum-/- mice. Corneal transparency and thickness were quantified using Nidek confocal microscopy. Fluorescent CSSC recovered by FACS from collagenase corneal digests were analyzed for viability with live/dead stain, and for expression of human and mouse genes by RT-PCR. Immunohistology and western blotting demonstrated matrix deposition by CSSC. TEM was used to determine collagen fibril distribution and diameters.
12 weeks after injection, CSSC isolated from corneas were 85% viable. They contained no mouse genomic DNA but expressed human lumican and keratocan mRNA. Lumican was present in the injected Lum-/- stromas. Corneal light scatter and thickness were in the normal range and significantly different from that of age-matched non-injected Lum-/- mice. TEM showed a marked reduction of fused collagen fibrils in posterior stroma compared to Lum-/- mice and a distribution of fibril diameters not significantly different from those of heterozygous Lum-/wt mice.
Xenotransplantation of human CSSC into mouse corneal stroma produced viable differentiated cells assuming keratocytic phenotype. The injected cells actively synthesized and deposited lumican and keratocan in the stromal matrix and remodeled the pre-existing stromal collagen architecture, resulting in restoration of corneal transparency and thickness resembling that of wild type mice. This study shows the Lum-/- mouse to be a useful model for therapy of corneal opacity and demonstrates the potential for clinical application of allogenic stem cells in stromal cell-based therapy.
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