Purpose

This study investigated the gene expression of TGFβ isoforms and their receptors in chick retina, retinal pigment epithelium (RPE), and choroid and the effects of short-term imposed optical defocus.

Methods

The expression of TGFβ isoforms (TGF-β1, 2, 3) and TGFβ receptors (TGFBR1, 2, 3) was examined in the retina, RPE, and choroid of young White-Leghorn untreated chicks (19 days-old). The effects on the expression of the same genes of monocular +10 and -10 D defocusing lenses, worn for either 2 or 48 h by age-matched chicks, were also examined by comparing expression in treated and untreated fellow eyes. RNA was purified, characterized and then reverse transcribed to cDNA. Differential gene expression was quantified using real-time PCR.

Results

All 3 isoforms of TGFβ and all 3 receptor subtypes were found to be expressed in all 3 ocular tissues, with apparent tissue-dependent differences in expression profiles. Data are reported as mean normalized expression relative to GAPDH. Sign-dependent optical defocus effects were also observed. Optical defocus did not affect retinal gene expression but in the RPE, TGF-β2 expression was significantly up-regulated with +10 D lenses, worn for either 2 h (349% increase ± 88%, p < 0.01) or 48 h (752% increase ± 166%, p < 0.001), and in the choroid, the expression of TGF-β3 was up-regulated with -10 D lenses, worn for 48 h (147% increase ± 9%, p < 0.01).

Conclusions

The effects of short term exposure to optical defocus on TGFβ gene expression in the RPE and choroid, which were sign-dependent and isoform specific, provide further supporting evidence for important roles of members of the TGFβ family and these two tissues in local signal cascades regulating ocular growth.

The choroid is the richly vascular layer of the eye located between the sclera and Bruch's membrane. Early studies in animals, as well as more recent studies in humans, have demonstrated that the choroid is a dynamic, multifunctional structure, with its thickness directly and indirectly subject to modulation by a variety of physiologic and visual stimuli. In this review, the anatomy and function of the choroid are summarized and links between the choroid, eye growth regulation, and myopia, as demonstrated in animal models, discussed. Methods for quantifying choroidal thickness in the human eye and associated challenges are described, the literature examining choroidal changes in response to various visual stimuli and refractive error-related differences are summarized, and the potential implications of the latter for myopia are considered. This review also allowed for the reexamination of the hypothesis that short-term changes in choroidal thickness induced by pharmacologic, optical, or environmental stimuli are predictive of future long-term changes in axial elongation, and the speculation that short-term choroidal thickening can be used as a biomarker of treatment efficacy for myopia control therapies, with the general conclusion that current evidence is not sufficient.

Leber's hereditary optic neuropathy (LHON) is a maternally inherited eye disease. X-linked nuclear modifiers were proposed to modify the phenotypic manifestation of LHON-associated mitochondrial DNA (mtDNA) mutations. By whole-exome sequencing, we identified the X-linked LHON modifier (c.157C>T, p.Arg53Trp) in PRICKLE3 encoding a mitochondrial protein linked to biogenesis of ATPase in 3 Chinese families. All affected individuals carried both ND4 11778G>A and p.Arg53Trp mutations, while subjects bearing only a single mutation exhibited normal vision. The cells carrying the p.Arg53Trp mutation exhibited defective assembly, stability, and function of ATP synthase, verified by PRICKLE3-knockdown cells. Coimmunoprecipitation indicated the direct interaction of PRICKLE3 with ATP synthase via ATP8. Strikingly, cells bearing both p.Arg53Trp and m.11778G>A mutations displayed greater mitochondrial dysfunction than those carrying only a single mutation. This finding indicated that the p.Arg53Trp mutation acted in synergy with the m.11778G>A mutation and deteriorated mitochondrial dysfunctions necessary for the expression of LHON. Furthermore, we demonstrated that Prickle3-deficient mice exhibited pronounced ATPase deficiencies. Prickle3-knockout mice recapitulated LHON phenotypes with retinal deficiencies, including degeneration of retinal ganglion cells and abnormal vasculature. Our findings provided new insights into the pathophysiology of LHON that were manifested by interaction between mtDNA mutations and X-linked nuclear modifiers.

Refractive errors are common eye disorders of public health importance worldwide. Ocular axial length (AL) is the major determinant of refraction and thus of myopia and hyperopia. We conducted a meta-analysis of genome-wide association studies for AL, combining 12,531 Europeans and 8,216 Asians. We identified eight genome-wide significant loci for AL (RSPO1, C3orf26, LAMA2, GJD2, ZNRF3, CD55, MIP, and ALPPL2) and confirmed one previously reported AL locus (ZC3H11B). Of the nine loci, five (LAMA2, GJD2, CD55, ALPPL2, and ZC3H11B) were associated with refraction in 18 independent cohorts (n = 23,591). Differential gene expression was observed for these loci in minus-lens-induced myopia mouse experiments and human ocular tissues. Two of the AL genes, RSPO1 and ZNRF3, are involved in Wnt signaling, a pathway playing a major role in the regulation of eyeball size. This study provides evidence of shared genes between AL and refraction, but importantly also suggests that these traits may have unique pathways.