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Lack of dystrophin protein Dp71 results in progressive cataract formation due to loss of fiber cell organization.

Fort PE, Darche M, Sahel JA, Rendon A, Tadayoni R - Mol. Vis. (2014)

Bottom Line: The role of Dp71 in fiber cells was also suggested by the progressive disorganization of the lens fibers, which was observed in the absence of Dp71 and demonstrated by irregular staining of the actin network and the aqueous channel AQP0.While its role in the retina has been well characterized, this study demonstrates for the first time the role played by Dp71 in a different ocular tissue: the crystalline lens.It primarily demonstrates the role that Dp71 plays in the maintenance of the integrity of the secondary lens fibers.

View Article: PubMed Central - PubMed

Affiliation: Institut de la Vision/INSERM/UPMC, Univ Paris 06/CNRS/CHNO des Quinze-Vingts, Paris, France ; Kellogg Eye Center, Departments of Ophthalmology and Visual Sciences, University of Michigan, Ann Arbor, MI.

ABSTRACT

Purpose: Dp71 is the main product of the Duchenne muscular dystrophy (DMD) gene in the central nervous system. While studying the impact of its absence on retinal functions, we discovered that mice lacking Dp71 also developed a progressive opacification of the crystalline lens. The purpose of this study was to perform a detailed characterization of the cataract formation in Dp71 knockout (KO-Dp71) mice.

Methods: Cataract formations in KO-Dp71 mice and wild-type (wt) littermates were assessed in vivo by slit-lamp examination and ex vivo by histological analysis as a function of aging. The expression and cellular localization of the DMD gene products were monitored by western blot and immunohistochemical analysis. Fiber cell integrity was assessed by analyzing the actin cytoskeleton as well as the expression of aquaporin-0 (AQP0).

Results: As expected, a slit-lamp examination revealed that only one of the 20 tested wt animals presented with a mild opacification of the lens and only at the most advanced age. However, a lack of Dp71 was associated with a 40% incidence of cataracts as early as 2 months of age, which progressively increased to full penetrance by 7 months. A subsequent histological analysis revealed an alteration in the structures of the lenses of KO-Dp71 mice that correlated with the severity of the lens opacity. An analysis of the expression of the different dystrophin gene products revealed that Dp71 was the major DMD gene product expressed in the lens, especially in fiber cells. The role of Dp71 in fiber cells was also suggested by the progressive disorganization of the lens fibers, which was observed in the absence of Dp71 and demonstrated by irregular staining of the actin network and the aqueous channel AQP0.

Conclusions: While its role in the retina has been well characterized, this study demonstrates for the first time the role played by Dp71 in a different ocular tissue: the crystalline lens. It primarily demonstrates the role that Dp71 plays in the maintenance of the integrity of the secondary lens fibers.

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The absence of Dp71 is associated with histological alterations of the crystalline lens. Cross-sections of crystalline lenses from wt and KO-Dp71 mice eyes were stained with hematoxylin and eosin at 2 (A-D) and 7 (E-H) months of age. No major alterations in the histology of the crystalline lenses were observed with aging in wt mice (A and E); alternatively, a progressive disorganization was obvious in lenses from KO-Dp71 mice (B and F). Higher magnification images suggest alterations in the ultrastructure of the lenses in KO-Dp71 mice (D and H, white arrows) along with a disorganized transition zone at both 2 and 7 months of age (C versus D and G versus H, respectively; black arrows; e: epithelial cells; f: fiber cells; tz: transition zone). Scale bars A, B, E, and F: 300 μm; Scale bars C, D, G, and H: 80 μm.
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f2: The absence of Dp71 is associated with histological alterations of the crystalline lens. Cross-sections of crystalline lenses from wt and KO-Dp71 mice eyes were stained with hematoxylin and eosin at 2 (A-D) and 7 (E-H) months of age. No major alterations in the histology of the crystalline lenses were observed with aging in wt mice (A and E); alternatively, a progressive disorganization was obvious in lenses from KO-Dp71 mice (B and F). Higher magnification images suggest alterations in the ultrastructure of the lenses in KO-Dp71 mice (D and H, white arrows) along with a disorganized transition zone at both 2 and 7 months of age (C versus D and G versus H, respectively; black arrows; e: epithelial cells; f: fiber cells; tz: transition zone). Scale bars A, B, E, and F: 300 μm; Scale bars C, D, G, and H: 80 μm.

Mentions: Gross morphological analyses of the isolated lenses of wt and KO-Dp71 mice showed that while lenses from the wt mice showed no morphological defects at the light level, an absence of Dp71 was clearly associated with refractive defects when placed on a 200-mesh EM grid (Figure 1). To better characterize the impact of the lack of Dp71 on the development and maintenance of a functional translucent lens, a histological analysis was performed on lenses from wt and KO-Dp71 mice at different ages. First, we observed that the size of the lens was consistently reduced in KO-Dp71 mice when compared with their wt littermates. This finding was maintained throughout life and represented here at 2 (65.2 mm3 in wt versus 36.8 mm3 in KO-Dp71) and 7 months (91.5 mm3 in wt versus 57.5 mm3 in KO-Dp71) to match the in vivo data (Figure 2). Careful observations of the wt lenses at these ages revealed normal lens development without cellular damage or disorganization in epithelial or fiber cells. While the overall structure and organization of the lenses were normal in the KO-D71 mice, consistent with the in vivo observations, subtle alterations were already detectable in lenses from 2-month-old KO-Dp71 mice, which was exacerbated at 7 months. At 2 months of age, a few instances of structural defects, such as the formation of vacuoles and a loss in the juxtaposition of fiber cells, were observed (Figure 2D, white arrows). Consistent with the slit-lamp observations, those defects seem to accumulate with age (Figure 2H, white arrows). Additionally, the distinct and typical bow-like organization of the cell nuclei observed in the wt mice began to be disrupted in 2-month-old KO-Dp71 mice. In KO-Dp71 animals, cell nuclei could be found much more centrally and deeply inside the fiber cell layers, well beyond the bow region of the equator. This was a phenotypic observation that was exacerbated with age, as demonstrated by images of lenses from 7-month-old KO-Dp71 mice (Figure 2H, arrowhead).


Lack of dystrophin protein Dp71 results in progressive cataract formation due to loss of fiber cell organization.

Fort PE, Darche M, Sahel JA, Rendon A, Tadayoni R - Mol. Vis. (2014)

The absence of Dp71 is associated with histological alterations of the crystalline lens. Cross-sections of crystalline lenses from wt and KO-Dp71 mice eyes were stained with hematoxylin and eosin at 2 (A-D) and 7 (E-H) months of age. No major alterations in the histology of the crystalline lenses were observed with aging in wt mice (A and E); alternatively, a progressive disorganization was obvious in lenses from KO-Dp71 mice (B and F). Higher magnification images suggest alterations in the ultrastructure of the lenses in KO-Dp71 mice (D and H, white arrows) along with a disorganized transition zone at both 2 and 7 months of age (C versus D and G versus H, respectively; black arrows; e: epithelial cells; f: fiber cells; tz: transition zone). Scale bars A, B, E, and F: 300 μm; Scale bars C, D, G, and H: 80 μm.
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Related In: Results  -  Collection

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f2: The absence of Dp71 is associated with histological alterations of the crystalline lens. Cross-sections of crystalline lenses from wt and KO-Dp71 mice eyes were stained with hematoxylin and eosin at 2 (A-D) and 7 (E-H) months of age. No major alterations in the histology of the crystalline lenses were observed with aging in wt mice (A and E); alternatively, a progressive disorganization was obvious in lenses from KO-Dp71 mice (B and F). Higher magnification images suggest alterations in the ultrastructure of the lenses in KO-Dp71 mice (D and H, white arrows) along with a disorganized transition zone at both 2 and 7 months of age (C versus D and G versus H, respectively; black arrows; e: epithelial cells; f: fiber cells; tz: transition zone). Scale bars A, B, E, and F: 300 μm; Scale bars C, D, G, and H: 80 μm.
Mentions: Gross morphological analyses of the isolated lenses of wt and KO-Dp71 mice showed that while lenses from the wt mice showed no morphological defects at the light level, an absence of Dp71 was clearly associated with refractive defects when placed on a 200-mesh EM grid (Figure 1). To better characterize the impact of the lack of Dp71 on the development and maintenance of a functional translucent lens, a histological analysis was performed on lenses from wt and KO-Dp71 mice at different ages. First, we observed that the size of the lens was consistently reduced in KO-Dp71 mice when compared with their wt littermates. This finding was maintained throughout life and represented here at 2 (65.2 mm3 in wt versus 36.8 mm3 in KO-Dp71) and 7 months (91.5 mm3 in wt versus 57.5 mm3 in KO-Dp71) to match the in vivo data (Figure 2). Careful observations of the wt lenses at these ages revealed normal lens development without cellular damage or disorganization in epithelial or fiber cells. While the overall structure and organization of the lenses were normal in the KO-D71 mice, consistent with the in vivo observations, subtle alterations were already detectable in lenses from 2-month-old KO-Dp71 mice, which was exacerbated at 7 months. At 2 months of age, a few instances of structural defects, such as the formation of vacuoles and a loss in the juxtaposition of fiber cells, were observed (Figure 2D, white arrows). Consistent with the slit-lamp observations, those defects seem to accumulate with age (Figure 2H, white arrows). Additionally, the distinct and typical bow-like organization of the cell nuclei observed in the wt mice began to be disrupted in 2-month-old KO-Dp71 mice. In KO-Dp71 animals, cell nuclei could be found much more centrally and deeply inside the fiber cell layers, well beyond the bow region of the equator. This was a phenotypic observation that was exacerbated with age, as demonstrated by images of lenses from 7-month-old KO-Dp71 mice (Figure 2H, arrowhead).

Bottom Line: The role of Dp71 in fiber cells was also suggested by the progressive disorganization of the lens fibers, which was observed in the absence of Dp71 and demonstrated by irregular staining of the actin network and the aqueous channel AQP0.While its role in the retina has been well characterized, this study demonstrates for the first time the role played by Dp71 in a different ocular tissue: the crystalline lens.It primarily demonstrates the role that Dp71 plays in the maintenance of the integrity of the secondary lens fibers.

View Article: PubMed Central - PubMed

Affiliation: Institut de la Vision/INSERM/UPMC, Univ Paris 06/CNRS/CHNO des Quinze-Vingts, Paris, France ; Kellogg Eye Center, Departments of Ophthalmology and Visual Sciences, University of Michigan, Ann Arbor, MI.

ABSTRACT

Purpose: Dp71 is the main product of the Duchenne muscular dystrophy (DMD) gene in the central nervous system. While studying the impact of its absence on retinal functions, we discovered that mice lacking Dp71 also developed a progressive opacification of the crystalline lens. The purpose of this study was to perform a detailed characterization of the cataract formation in Dp71 knockout (KO-Dp71) mice.

Methods: Cataract formations in KO-Dp71 mice and wild-type (wt) littermates were assessed in vivo by slit-lamp examination and ex vivo by histological analysis as a function of aging. The expression and cellular localization of the DMD gene products were monitored by western blot and immunohistochemical analysis. Fiber cell integrity was assessed by analyzing the actin cytoskeleton as well as the expression of aquaporin-0 (AQP0).

Results: As expected, a slit-lamp examination revealed that only one of the 20 tested wt animals presented with a mild opacification of the lens and only at the most advanced age. However, a lack of Dp71 was associated with a 40% incidence of cataracts as early as 2 months of age, which progressively increased to full penetrance by 7 months. A subsequent histological analysis revealed an alteration in the structures of the lenses of KO-Dp71 mice that correlated with the severity of the lens opacity. An analysis of the expression of the different dystrophin gene products revealed that Dp71 was the major DMD gene product expressed in the lens, especially in fiber cells. The role of Dp71 in fiber cells was also suggested by the progressive disorganization of the lens fibers, which was observed in the absence of Dp71 and demonstrated by irregular staining of the actin network and the aqueous channel AQP0.

Conclusions: While its role in the retina has been well characterized, this study demonstrates for the first time the role played by Dp71 in a different ocular tissue: the crystalline lens. It primarily demonstrates the role that Dp71 plays in the maintenance of the integrity of the secondary lens fibers.

Show MeSH
Related in: MedlinePlus