Structural analysis of lens epithelial explants induced to differentiate into fibres by fibroblast growth factor (FGF) (original) (raw)
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Fibroblast growth factor receptor signaling is essential for lens fiber cell differentiation
Developmental Biology, 2008
The vertebrate lens provides an excellent model to study the mechanisms that regulate terminal differentiation. Although fibroblast growth factors (FGFs) are thought to be important for lens cell differentiation, it is unclear which FGF receptors mediate these processes during different stages of lens development. Deletion of three FGF receptors (Fgfr1-3) early in lens development demonstrated that expression of only a single allele of Fgfr2 or Fgfr3 was sufficient for grossly normal lens development, while mice possessing only a single Fgfr1 allele developed cataracts and microphthalmia. Profound defects were observed in lenses lacking all three Fgfrs. These included lack of fiber cell elongation, abnormal proliferation in prospective lens fiber cells, reduced expression of the cell cycle inhibitors p27 kip1 and p57 kip2 , increased apoptosis and aberrant or reduced expression of Prox1, Pax6, c-Maf, E-cadherin and α-, βand γ-crystallins. Therefore, while signaling by FGF receptors is essential for lens fiber differentiation, different FGF receptors function redundantly.
Investigative Ophthalmology Amp Visual Science, 1992
Fibroblast growth factor (FGF) is a potent inducer of fiber differentiation in lens epithelial explants from neonatal rats as assessed by the accumulation of fiber-specific proteins (/?-and 7-crystallins) and the progression of cells through a sequence of morphologic events characteristic of fiber differentiation in situ. Because new fibers normally are formed in the lens throughout life, the authors questioned whether epithelial cells from rats of all ages are induced to differentiate into fibers by FGF. Earlier studies have shown that, with the increasing age of the donor rat, the lens epithelial explants had a reduced ability to accumulate 0-and 7-crystallins in response to FGF. To determine if the characteristic morphologic events in fiber differentiation were induced by FGF in explants from rats of different ages, an ultrastructural study was done. Using the time of appearance and level of expression of the following morphologic markers of fiber differentiation: (1) cell elongation, (2) reduction of cytoplasmic organdies, (3) formation of cell processes, and (4) fiber denucleation, the level of fiber differentiation induced by FGF was assessed in explants from 10-, 21-, 100-, and 175-day-old rats. These results showed that, with increasing donor age, epithelial cells showed a gradual decline in responsiveness to FGF. This was manifested by a slower progression through the sequence of fiber-specific structural events as the age of the donor rat increased. At all ages studied, cells in the central region of explants responded more slowly than cells from the peripheral region. The finding that FGF induces events in fiber differentiation, albeit at a slower rate, in explants from mature rats supports the hypothesis that FGF in the eye continues to play a role in inducing lens epithelial cells at the lens equator to differentiate into fibers throughout life.
Overexpression of FGF-2 modulates fiber cell differentiation and survival in the mouse lens
Development (Cambridge, England), 1997
During mammalian embryogenesis, the ocular lens forms through a temporally and spatially regulated pattern of differentiation which is thought to be coordinated at least in part by the FGF-1 and FGF-2 members of the fibroblast growth factor (FGF) family. Previous transgenic experiments in which FGF-1 or dominant negative FGF receptors were overexpressed in the lens indicated that FGF-1 could induce differentiation while differentiated lens cells rely upon FGF signaling for their survival. In this study, we asked if the 17.5 kDa FGF-2 protein was capable of inducing differentiation of lens cells in transgenic mice. Unexpectedly, differentiation was inhibited by lens-specific expression of a transgene encoding a secreted form of the 17.5 kDa bovine FGF-2 protein under the transcriptional control of the murine alphaA-crystallin promoter (alphaAIgFGF-2 transgenic mice). To address the possibility that FGF-2 functions as a modulator of fiber cell survival, alphaAIgFGF-2 transgenic mice w...
Growth Factor Signaling in Vitreous Humor-Induced Lens Fiber Differentiation
Investigative Ophthalmology & Visual Science, 2010
Although some of the factors and signaling pathways that are involved in induction of fiber differentiation have been defined, such as FGF-mediated MAPK/ERK and PI3-K/Akt signaling, the factors in the vitreous that regulate this differentiation process in vivo have yet to be identified. The purpose of this study was to better understand the role of growth factors in vitreous that regulate this process by further characterizing the signaling pathways involved in lens fiber differentiation. METHODS. Rat lens epithelial explants were used to compare the ability of vitreous, IGF-1, PDGF-A, EGF, and FGF-2 to stimulate the phosphorylation of ERK1/2 and Akt leading to fiber differentiation, in the presence or absence of selective receptor tyrosine kinase (RTK) inhibitors. RESULTS. Similar to vitreous, FGF induced a sustained ERK1/2 signaling profile, unlike IGF, PDGF, and EGF, which induced a more transient (shorter) activation of ERK1/2. For Akt activation, IGF was the only factor that induced a profile similar to vitreous. IGF, PDGF, and EGF potentiated the effects of a low dose of FGF on lens fiber differentiation by extending the duration of ERK1/2 phosphorylation. In the presence of selective RTK inhibitors, although the sustained vitreous-induced ERK1/2 signaling profile and subsequent fiber differentiation was perturbed, the results also showed that, although prolonged ERK1/2 phosphorylation was necessary, it was not sufficient for fiber differentiation to proceed. CONCLUSIONS. These results are consistent with FGF's being the key growth factor involved in vitreous-induced signaling leading to lens fiber differentiation; however, they also indicate that other vitreal growth factors such as IGF may be involved in fine-tuning ERK1/2-and Akt-phosphorylation to the level that is necessary for initiation and/or maintenance of lens fiber differentiation in vivo. (Invest Ophthalmol Vis Sci. 2010;51: 3599 -3610)
Experimental Cell Research, 1981
Cultured bovine lens epithelial cells are polygonal in shape. In confluent and multilayer cultures they exhibit elaborate arrays of 6 nm filaments, bundles of intermediate-sized filaments, and a fibrous meshwork of subcellular and intercellular material. Cells grown in the presence of a retinal extract (RE) have a higher growth rate, and are smaller and more regular in shape. In them the 6 nm filaments are mostly aligned in sheets, the intermediate-sized filaments form a fine network, and the cells are closely apposed to the plastic substratum. Some homogeneous material is formed intercellularly in older cultures. Cellular elongation, induced in the former cultures by the addition of RE, is accompanied by an alignment of cytoskeletal elements, including microtubules, parallel to the long axis. Other structural features are similar in all cell types. The response to RE is discussed in terms of shape modulations associated with the restricted expression of structural characteristics acquired in vitro.
FGF Signaling in Chick Lens Development
Developmental Biology, 2001
The prevailing concept has been that an FGF induces epithelial-to-fiber differentiation in the mammalian lens, whereas chick lens cells are unresponsive to FGF and are instead induced to differentiate by IGF/insulin-type factors. We show here that when treated for periods in excess of those used in previous investigations (>5 h), purified recombinant FGFs stimulate proliferation of primary cultures of embryonic chick lens epithelial cells and (at higher concentrations) expression of the fiber differentiation markers ␦-crystallin and CP49. Surprisingly, upregulation of proliferation and ␦-crystallin synthesis by FGF does not require activation of ERK kinases. ERK function is, however, essential for stimulation of ␦-crystallin expression in response to insulin or IGF-1. Vitreous humor, the presumptive source of differentiation-promoting activity in vivo, contains a factor capable of diffusing out of the vitreous body and inducing ␦-crystallin and CP49 expression in chick lens cultures. This factor binds heparin with high affinity and increases ␦-crystallin expression in an ERK-insensitive manner, properties consistent with an FGF but not insulin or IGF. Our findings indicate that differentiation in the chick lens is likely to be mediated by an FGF and provide the first insights into the role of the ERK pathway in growth factor-induced signal transduction in the lens.
Investigative Ophthalmology Amp Visual Science, 1993
Previous research in this laboratory has shown that fibroblast growth factor stimulates lens epithelial explants to proliferate, migrate, and differentiate into fibers in a progressive dose-dependent manner. The lens has distinct compartments where cells proliferate (germinative zone), migrate, or get displaced (equator) and differentiate into fibers (transitional zone). These compartments occur in an anteroposterior spatial sequence and the authors hypothesized that fibroblast growth factor plays a critical role in determining these spatial patterns of lens growth and lens polarity. To investigate this hypothesis the distribution of fibroblast growth factor in the lens was analyzed. Methods. Immunohistochemistry was used to localize acidic fibroblast growth factor and basic fibroblast growth factor in the cells and capsule of lenses from neonatal, weanling, and adult rats. Because of its functional relationship with fibroblast growth factor, heparan sulphate proteoglycan was also localized in the lens, Results. In all ages examined, cytoplasmic acidic fibroblast growth factor is present in the germinative and transitional zones of the lens and both acidic fibroblast growth factor and basic fibroblast growth factor are present in the capsule. A major finding is the co-localization of fibroblast growth factor and heparan sulphate proteoglycan reactivity in the lens capsule in the form of laminae. These laminae become more prominent as the capsule thickens and differences in arrangement of laminae between anterior, equatorial, and posterior regions of the capsule also become apparent. Conclusions. The presence of fibroblast growth factor in lens cells and capsule in neonatal, weanling, and adult rats indicates an important role for fibroblast growth factor in lens cell biology. Moreover, the regional distribution of fibroblast growth factor, particularly in the lens cells, indicates that it may influence determination of lens polarity and growth patterns. Invest Ophthalmol Vis Sci. 1993;34:3355-3365. 1 hroughout development inductive interactions take place between cells and tissues that determine their temporal and spatial differentiation and growth patterns. The lens has a distinctive architecture that, once established in the embryo, is maintained throughout
Experimental Eye Research, 1984
Lenses from 19-day chick embryos are fractionated by a double punch method to obtain the epithelium-annular pad complex (EP), outer fibres (OF), middle fibres (MF) and central fibres (CF). Water-soluble crystallins are characterized by SDS PAGE, isoelectric focusing (IEF) and two-dimensional IEF-SDS PAGE. Crystallins are also characterized by immunoelectrophoresis (IE), rocket IE, IEF-immunoblotting, and quantified by two-dimensional antigen-antibody crossed electrophoresis using antibodies to total 19-day embryonic as well as adult crystallins. In the adult lens, alpha-, beta- and delta-crystallins are 19%, 67% and 14%, respectively, while these are present at concentrations of 9%, 27% and 64%, respectively, in 19-day embryonic lens. In absolute amounts, delta-crystallin increases only by 1.23-fold between 19-day embryonic age and 6 months post-hatching, while total lens protein increases 12.5-fold. The predominance of delta-crystallin in central fibres, located along the optical axis, suggests that this protein is of embryonic origin. delta-Crystallin from fibres is electrofocused as 12 distinct molecular classes (pI 5.2-5.42) which react against anti-delta-crystallin on an immunoblot. Of these, the three most anodal species are not detected in EP. Fibres contain 50 000, 48 000 and 45 000 dalton delta-crystallin subunits while only 50 000 and 48 000 dalton subunits are present in EP.