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Proteoglycan synthesis by human corneal explants submitted to laser in situ keratomileusis (LASIK).

Martins SA, Campos MQ, Vidal BC, Berto AG, Aguiar JA, Michelacci YM - Mol. Vis. (2007)

Bottom Line: After 24 h incubation, PGs were extracted and identified by a combination of agarose gel electrophoresis and enzymatic degradation with protease and specific glycosaminoglycan lyases.Histopathological and birefringence analysis were performed in fixed tissue slices.Laser application led to decreased PG biosynthesis in human corneal explants, with marked changes in the collagen fibril organization, as revealed by changes in the tissue birefringence.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Oftalmologia, Universidade Federal de São Paulo, Escola Paulista de Medicina, São Paulo, Brazil. suyreboucas@uol.com.br <suyreboucas@uol.com.br>

ABSTRACT

Purpose: To evaluate the acute effects of laser in situ keratomileusis (LASIK) upon the synthesis of proteoglycans (PGs) and collagen fibril organization in human corneal explants.

Methods: Human corneas that had been rejected for transplants were obtained at Banco de Olhos of Hospital São Paulo. For each eye pair, one cornea was submitted to refractive surgery, and the other was used as its matched control. After surgery, the corneas were excised from the eyes and immediately placed in a Ham F-12 nutrient mixture containing (35)S-sulfate for the metabolic labeling of PGs. After 24 h incubation, PGs were extracted and identified by a combination of agarose gel electrophoresis and enzymatic degradation with protease and specific glycosaminoglycan lyases. Histopathological and birefringence analysis were performed in fixed tissue slices.

Results: A marked decrease in (35)S-sulfate incorporation in PGs was observed in corneal explants that received LASIK, especially concerning dermatan sulfate-PGs, with keratan sulfate- and heparan sulfate-PG synthesis reduced to a lower degree. Only low molecular weight PGs were present in the corneas, both before and 24 h after LASIK. No sign of wound healing processes were observed, but a marked change in corneal birefringence was seen following LASIK treatment.

Conclusions: Laser application led to decreased PG biosynthesis in human corneal explants, with marked changes in the collagen fibril organization, as revealed by changes in the tissue birefringence.

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Related in: MedlinePlus

Control and LASIK human cornea birefringence images. The experiment was performed as describe in Methods, with (C,D) and without (A,B) compensation, for control (A,C) and LASIK-submitted (B,D) corneal slices. The samples are positioned 45° from the polarizer azimuth. Ep: epithelium; St: stroma; End: endothelium. The scale bar is equal to 20 μm.
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f7: Control and LASIK human cornea birefringence images. The experiment was performed as describe in Methods, with (C,D) and without (A,B) compensation, for control (A,C) and LASIK-submitted (B,D) corneal slices. The samples are positioned 45° from the polarizer azimuth. Ep: epithelium; St: stroma; End: endothelium. The scale bar is equal to 20 μm.

Mentions: Figure 7 illustrates typical birefringence images of a normal (Figure 7A,C) and a LASIK-treated cornea (Figure 7B,D), with dark and bright shadows and also some visible irregularities in the intensity. The corneal collagen fibrils were oriented 45° from the polarizer azimuths, allowing maximum shining of the fibrils. Sample Figure 7A represents the control cornea section without compensation. In this human corneal section, collagen fibrils are seen as either parallel aligned linear structures or as hexagonal arranged structures. Figure 7C represents the same corneal micrograph with compensation to allow the measurements of optical retardation values. In birefringence images of human corneal sections after LASIK (Figure 7B,D), only minor differences were observed in collagen fibril alignments when compared to the control cornea. These differences are evident after obtaining statistical analysis of the quantitative data of optical retardation (Table 2). Eighty-two measurements were performed for each corneal group (Control and LASIK) in different regions of corneal slices. Note that there is a significant decrease in optical values after LASIK, in comparison to the control group.


Proteoglycan synthesis by human corneal explants submitted to laser in situ keratomileusis (LASIK).

Martins SA, Campos MQ, Vidal BC, Berto AG, Aguiar JA, Michelacci YM - Mol. Vis. (2007)

Control and LASIK human cornea birefringence images. The experiment was performed as describe in Methods, with (C,D) and without (A,B) compensation, for control (A,C) and LASIK-submitted (B,D) corneal slices. The samples are positioned 45° from the polarizer azimuth. Ep: epithelium; St: stroma; End: endothelium. The scale bar is equal to 20 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2533033&req=5

f7: Control and LASIK human cornea birefringence images. The experiment was performed as describe in Methods, with (C,D) and without (A,B) compensation, for control (A,C) and LASIK-submitted (B,D) corneal slices. The samples are positioned 45° from the polarizer azimuth. Ep: epithelium; St: stroma; End: endothelium. The scale bar is equal to 20 μm.
Mentions: Figure 7 illustrates typical birefringence images of a normal (Figure 7A,C) and a LASIK-treated cornea (Figure 7B,D), with dark and bright shadows and also some visible irregularities in the intensity. The corneal collagen fibrils were oriented 45° from the polarizer azimuths, allowing maximum shining of the fibrils. Sample Figure 7A represents the control cornea section without compensation. In this human corneal section, collagen fibrils are seen as either parallel aligned linear structures or as hexagonal arranged structures. Figure 7C represents the same corneal micrograph with compensation to allow the measurements of optical retardation values. In birefringence images of human corneal sections after LASIK (Figure 7B,D), only minor differences were observed in collagen fibril alignments when compared to the control cornea. These differences are evident after obtaining statistical analysis of the quantitative data of optical retardation (Table 2). Eighty-two measurements were performed for each corneal group (Control and LASIK) in different regions of corneal slices. Note that there is a significant decrease in optical values after LASIK, in comparison to the control group.

Bottom Line: After 24 h incubation, PGs were extracted and identified by a combination of agarose gel electrophoresis and enzymatic degradation with protease and specific glycosaminoglycan lyases.Histopathological and birefringence analysis were performed in fixed tissue slices.Laser application led to decreased PG biosynthesis in human corneal explants, with marked changes in the collagen fibril organization, as revealed by changes in the tissue birefringence.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Oftalmologia, Universidade Federal de São Paulo, Escola Paulista de Medicina, São Paulo, Brazil. suyreboucas@uol.com.br <suyreboucas@uol.com.br>

ABSTRACT

Purpose: To evaluate the acute effects of laser in situ keratomileusis (LASIK) upon the synthesis of proteoglycans (PGs) and collagen fibril organization in human corneal explants.

Methods: Human corneas that had been rejected for transplants were obtained at Banco de Olhos of Hospital São Paulo. For each eye pair, one cornea was submitted to refractive surgery, and the other was used as its matched control. After surgery, the corneas were excised from the eyes and immediately placed in a Ham F-12 nutrient mixture containing (35)S-sulfate for the metabolic labeling of PGs. After 24 h incubation, PGs were extracted and identified by a combination of agarose gel electrophoresis and enzymatic degradation with protease and specific glycosaminoglycan lyases. Histopathological and birefringence analysis were performed in fixed tissue slices.

Results: A marked decrease in (35)S-sulfate incorporation in PGs was observed in corneal explants that received LASIK, especially concerning dermatan sulfate-PGs, with keratan sulfate- and heparan sulfate-PG synthesis reduced to a lower degree. Only low molecular weight PGs were present in the corneas, both before and 24 h after LASIK. No sign of wound healing processes were observed, but a marked change in corneal birefringence was seen following LASIK treatment.

Conclusions: Laser application led to decreased PG biosynthesis in human corneal explants, with marked changes in the collagen fibril organization, as revealed by changes in the tissue birefringence.

Show MeSH
Related in: MedlinePlus