[Partial lamellar grafting of a desiccated cornea in prolonged suppurative diseases and ulcers of the cornea].
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Corneal Transplantation↗
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[Experimental transplantation of the human cornea into the rabbit cornea].
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Animals↗
[Artificial scleral sinus with covering of recipient's cornea with cadaveric cornea for treating acute corneal staphyloma and keratoconus (author's transl)].
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Acute Disease↗
Altered expression of aquaporins in bullous keratopathy and Fuchs' dystrophy corneas.
Corneas with edema-related diseases lose transparency, which causes significant vision loss. This study analyzed seven aquaporins (AQPs) in normal corneas, pseudophakic/aphakic bullous keratopathy (PBK/ABK) corneas, Fuchs' dystrophy corneas, keratoconus corneas, post-cataract surgery (PCS) corneas, and normal organ-cultured corneas. RNA levels for AQP1, AQP4, and beta2-microglobulin were measured by RT-PCR. AQP1 antibody localized to stromal cells of all corneas. PBK/ABK and Fuchs' dystrophy corneas had decreased endothelial cell staining compared with normal. AQP1 mRNA was found in whole corneas and cultured stromal fibroblasts but not in isolated epithelial cells. AQP3 staining was found in basal epithelial cells of the normal, Fuchs' dystrophy, and keratoconus corneas but throughout the entire epithelium of PBK/ABK corneas. AQP4 antibody localized to endothelial cells of all corneas and in stromal cells of PBK/ABK corneas. AQP4 mRNA was identified in whole human corneas. AQP5 was found in epithelial cells of all corneas. AQP0, AQP2, and AQP9 were not found in any corneas. Normal AQP distributions were found in PCS and organ-cultured corneas, although they showed signs of swelling. Our study demonstrates that AQP abnormalities are found in PBK/ABK corneas (decreased AQP1, increased AQP3 and AQP4) and Fuchs' dystrophy corneas (decreased AQP1). Although both have vision-disrupting corneal edema, the mechanisms of fluid accumulation may be different in each disease.
Aquaporin 1↗
Expression of collagen I, smooth muscle alpha-actin, and vimentin during the healing of alkali-burned and lacerated corneas.
PURPOSES: Alkali-burned corneas can seldom heal properly to restore corneal transparency. To provide a better understanding of this devastating corneal injury, we compared the expression of collagen I, smooth muscle alpha-actin (alpha-SMA), and vimentin in lacerated and alkali-burned rabbit corneas. METHODS: A radiolabeled cDNA probe of alpha 1(I) chain was used in slot-blot hybridization to determine the levels of alpha 1(I) mRNA in alkali-burned corneas. In situ hybridization was used to identify the cell types that express the alpha 1(I) chain. Antibodies against collagen I, alpha-SMA, and vimentin were used in immunohistochemical studies to determine the tissue distribution of collagen I and to identify cells expressing alpha-SMA and vimentin. RESULTS: The levels of alpha 1(I) mRNA in alkali-burned corneas increased steadily after the alkali burn and reached a plateau within 2 weeks. One day after alkali burn, specific in situ hybridization signals were detected in stromal cells immediately surrounding the edge of the corneal injury. As the healing proceeded, the fibroblastic cells migrated into the injured stroma, and they showed positive reactions by in situ hybridization and by immunostaining with anti-collagen I probes. In alkali-burned corneas, retrocorneal membranes were formed 1 week after injury. This fibrillar membrane was stained by anti-collagen I antibody, and the fibroblastic cells in the membrane were hybridized by the 3H-labeled alpha 1(I) cDNA probe. No retrocorneal membrane was formed in the lacerated corneas, even after the injured corneas were allowed to heal for 3 weeks. The epithelial cells in the epithelial plug of lacerated corneas were positive by in situ hybridization, whereas the epithelial cells in the regenerated epithelium of alkali-burned cornea was not. Antibodies against alpha-SMA reacted with the migrating fibroblastic cells but did not react with epithelial cells or endothelial cells in the injured corneas. Anti-vimentin antibody reacted with fibroblastic cells, endothelial cells, and keratocytes in normal and injured corneas, and with the basal epithelial cells of injured corneas. CONCLUSIONS: During wound healing, the keratocytes that migrate to injured stroma transform into myofibroblasts. These myofibroblasts express high levels of alpha 1(I) mRNA, alpha-SMA, and vimentin. The healing of alkali-burned corneas differ from that of lacerated corneas in that the retrocorneal membranes are formed in the former but not in the latter. In addition, the epithelial cells of alkali-burned corneas lack alpha 1(I) mRNA, whereas it is found in the epithelium of lacerated corneas. These differences may result from the persistence of inflammatory cells in the alkali-burned corneas.
Actins↗
Ultraviolet light-induced changes in the glucose-6-phosphate dehydrogenase activity of porcine corneas.
PURPOSE: Corneas have the high activity of the pentose phosphate pathway. To clarify the relevance of the pentose phosphate pathway to their antioxidant defense, we examined the activity of glucose-6-phosphate dehydrogenase (G6PDH), the rate-determining enzyme of the pathway, in corneas exposed to ultraviolet light (UV). METHODS: Fresh porcine eye globes were exposed to ultraviolet light A (UVA) or ultraviolet light C (UVC), and the tear film-removed eye globes to UVA. After UV exposure, the corneas were dissected from the eye globes and extracted with a saline solution, and the G6PDH activity in the extract was assayed. The G6PDH activity of unilateral opaque corneas, their paired, transparent corneas, and normal corneas, all of which were obtained from fresh, UV-unexposed eye globes, and the L-lactate dehydrogenase (LDH) activity of these opaque and normal corneas also were assayed. RESULTS: The G6PDH activity of corneas increased with UVA exposure, and decreased with long-term UVC exposure, although it increased with short-term UVC exposure. A UV-blocking contact lens screened corneas from the UVA-induced increase. Removal of the tear film enhanced the UVC-induced decrease. The G6PDH activity of unilateral opaque corneas was lower than that of paired, transparent corneas or normal corneas, all of which were obtained from fresh, UV-unexposed eye globes. The LDH activity of the opaque corneas was much higher than that of the normal corneas. CONCLUSION: Exposure of corneas to UVA or a small dose of UVC enhances the G6PDH activity, i.e., the pentose phosphate pathway. This activity enhancement may play an important role in corneal antioxidant defense against UV-induced oxidative stress. However, exposure of corneas to large doses of UVC appears to damage the pathway.
Animals↗
Stress-strain measurements of human and porcine corneas after riboflavin-ultraviolet-A-induced cross-linking.
PURPOSE: To evaluate the biomechanical effect of combined riboflavin-ultraviolet A (UVA) treatment on porcine and human corneas. SETTING: Department of Ophthalmology, Technical University of Dresden, Dresden, Germany. METHODS: Corneal strips from 5 human enucleated eyes and 20 porcine cadaver corneas were treated with the photosensitizer riboflavin and irradiated with 2 double UVA diodes (370 nm, irradiance = 3 mW/cm2) for 30 minutes. After cross-linking, static stress-strain measurements of the treated and untreated corneas were performed using a microcomputer-controlled biomaterial tester with a prestress of 5 x 10(3) Pa. RESULTS: There was a significant increase in corneal rigidity after cross-linking, indicated by a rise in stress in treated porcine corneas (by 71.9%) and human corneas (by 328.9%) and in Young's modulus by the factor 1.8 in porcine corneas and 4.5 in human corneas. The mean central corneal thickness was 850 microm +/- 70 (SD) in porcine corneas and 550 +/- 40 microm in human corneas. CONCLUSIONS: Riboflavin-UVA-induced collagen cross-linking led to an increase in mechanical rigidity in porcine corneas and an even greater increase in human corneas. As collagen cross-linking is maximal in the anterior 300 microm of the cornea, the greater stiffening effect in human corneas can be explained by the relatively larger portion of the cornea being cross-linked in the overall thinner human cornea.
Animals↗
Generation of complement-derived anaphylatoxins in normal human donor corneas.
Complement-derived anaphylatoxins (C3a, C4a, and C5a) are potent, stable mediators of acute inflammation. Because human corneas contain functional complement, the authors subjected normal human donor corneas to various forms of immunologic or chemical injury to determine if the complement system could be activated and anaphylatoxins generated. The experimental cornea of each donor pair was injected with lipopolysaccharide (LPS) or immune complexes or injured by application of acid or alkali. The remaining cornea of each donor pair served as a control. After incubation of corneas in tissue culture media for 6 hours and elution in phosphate-buffered saline for 24 hours, C3a, C4a, and C5a were measured in corneal eluates by radioimmunoassay. Compared with control corneas, C3a levels were significantly increased in corneas injected with LPS or immune complexes and in corneas injured with acid or alkali. C4a levels were significantly elevated in corneas injected with immune complexes and in corneas injured with acid or alkali but not in corneas injected with LPS. C5a levels were detectable only in corneas injured with acid or alkali. These results suggest that immunologic reactions in the human cornea may activate the classic or alternative complement pathways and generate anaphylatoxins. Additionally, chemical injuries with acid or alkali generate anaphylatoxins in the cornea. Anaphylatoxins may participate in the acute inflammatory response of the human cornea to chemical or immunologic injury.
Acids↗
Angiogenic activity in injured rat corneas as assayed on the chick chorioallantoic membrane.
The temporal appearance of an angiogenic effect in chemically cauterized rat corneas was determined by studying the responses that they induced in the vessels of the chick chorioallantoic membrane (CAM). Injured rat corneas were grafted to the CAM from 90 minutes to 7 days after cautery. As controls, uninjured rat corneas and corneas of healthy rats cauterized immediately after death were also grafted. The vascular responses to the grafts were graded in a masked fashion by stereoscopic biomicroscopy on a five-tiered scale, by evaluations of projected colored photographs on the same scale, and by histologic examination of the grafts. Separate coefficients of angiogenesis were determined for the stereoscopic and photographic evaluations. We detected significant differences between corneas of healthy rats that were uninjured or cauterized chemically immediately after death and those that were cauterized in the living rat. Uninjured corneas and corneas cauterized postmortem elicited a mild vascular response in the CAM, as reflected by low coefficients of angiogenesis. Whereas blood vessels were not detected in corneas injured postmortem, some normal corneas vascularized but only after being on the CAM for at least 7 days. The coefficients of angiogenesis of corneas that were cauterized during life were significantly higher than those of both control groups prior to grafting after comparable times on the CAM. Corneas vascularized on the CAM included those that were cauterized as soon as 90 minutes prior to grafting. The strongest vascular responses, as reflected by coefficient of angiogenesis and the frequency of histologically confirmed nucleated avian erythrocytes within intracorneal blood vessels, were found with corneas that were grafted to the CAM 3 days after chemical cauterization. Corneas that vascularized on the CAM were associated with a prominent leukocytic infiltrate suggestively derived from the chick embryo. The results suggest that chemically cauterized rat corneas contain a chemoattractant for polymorphonuclear leukocytes within 90 minutes of injury and that such polymorphonuclear leukocytes or other components of the injured corneas possess the ability to stimulate angiogenesis on the CAM.
Allantois↗
[Evaluation of the quality of a donor cornea before transplantation].
UNLABELLED: Transplantation of the cornea is often the only possible treatment of many ophthalmological diseases and injuries. The quality of the donor cornea determines in a major way the success of transplantation and therefore the demands on its quality must be high. OBJECTIVE: Assess how the vitality of the endothelium changes in relation to the time which elapsed since the donor's death after storage of the cornea in the medium (death/preservation time, DPT). If the vitality of the endothelium is poor, the oedema of the cornea persists at room temperature. METHODS: The endothelium of the donor cornea was examined before transplantation under a specular microscope Bio Optics (USA). According to DPT the corneae were divided into three groups: 1st group--22 corneae where DPT is shorter than 5 hours 2nd group--33 corneae where DPT is 6-10 hours 3rd group--46 corneae where DPT was longer than 10 hours. RESULTS: In the first group the corneae got rid of the oedema after 20 minutes at room temperature and endothelial cells could be observed in detail. In the second group the corneae got rid of the oedema after 20 minutes but there were rare oedematous sites where the endothelial function was not adequate. In the third group even after long time at room temperature the cornea did not get rid of the oedema, and on examination it was very difficult to find a group of endothelial cells for evaluation. CONCLUSION: Corneae from group 1 have a totally vital endothelium, in the second group there are sites on the cornea where the vitality of the endothelium is already impaired, in the third group the vitality of the entire endothelium is impaired. For transplantation corneae with a DPT shorter than 10 hours are suitable. With a DPT longer than 10 hours they are less suitable or unsuitable for transplantation.
Cadaver↗
The expression of tenascin and fibronectin in keratoconus, scarred and normal human cornea.
BACKGROUND: The etiology and pathogenesis of keratoconus remain unclear, and therefore we decided to study the distribution of different isoforms of tenascin (Tn) and fibronectin (Fn) in normal human corneas and in those obtained from penetrating keratoplasty for keratoconus and corneal scarring. METHODS: Frozen sections of human cornea and conjunctiva were stained by immunohistochemical methods with a panel of monoclonal antibodies (MAbs) against different isoforms of Tn and Fn. RESULTS: In the normal human eye, Tn was found in the limbal and conjunctival basement membrane region, in the conjunctival blood vessels and at the junction of the cornea and sclera, but no immunoreaction was seen in the normal cornea. In the corneas from the keratoconus patients, a clear immunoreaction for Tn was seen in the defects of Bowman's membrane as well as in the distorted stroma beneath the defects. In some of the keratoconus corneas, basement membrane adjacent to the defects also showed reactivity for Tn, and in clinically and histologically scarred keratoconus corneas the scars expressed Tn. In the scarred corneas, only blood vessels in the posterior portion of the cornea showed immunoreactivity for Tn, while no Tn was noted in the scar area or in Bowman's membrane. No major differences were noticed in the reactivity of different MAbs against Tn isoforms. Fn, extradomain A Fn (EDA-Fn) and oncofetal Fn (onc-Fn) were found in the basement membrane of the central cornea of the normal eye. In keratoconus corneas, the defects and clinical and histological scars bound MAbs against Fn, EDA-Fn and onc-Fn, but in the scarred corneas no enhancement in the expression of Fns was noted. Extradomain B cellular Fn (EDB-Fn) was not expressed in any of the eyes studied. CONCLUSIONS: The results suggest that the anterior portion of the cornea is involved in the pathogenesis of keratoconus. Furthermore, it seems that the expression of Tn and Fns in the clinically scarred keratoconus corneas is due to a process in which both repairing and scar-forming mechanisms operate at the same time. However, the origin of the defects in Bowman's membrane seen in keratoconus still remains unclear. They may be minor scars due to the disease or primary defects in the process leading to keratoconus.
Adolescent↗
Does the human cornea contain silicon?
BACKGROUND: Our study investigated the presence, type and quantity of silicon in the human cornea. We report the results of silicon measurements in the corneas of silicotic individuals, bricklayers and apparently normal human individuals and offer a hypothesis for the mechanism of silicon deposition in the human cornea. METHODS: We examined corneas from 13 decreased subjects who suffered from silicosis, 2 bricklayers and 6 apparently healthy subjects. Cornea samples were examined by energy-dispersive x-ray analysis (EDXA) under calibrated conditions in a scanning electron microscope (SEM). The EDXA detector was a silicon-free germanium crystal. Five distinct layers (epithelium, Bowman's membrane, central stroma. Descemet's membrane and endothelium) were analyzed in each cornea. The method allows simultaneous semiquantitative analysis of, among other elements, silicon, calcium and oxygen. We measured amorphous silicon and visible particles of silicon. RESULTS: We found amorphous silicon in low concentrations in 38% of the silicotic corneas and in very low concentrations in 29% of the healthy corneas. Bricklayers showed high concentrations of amorphous silicon. These accumulations of silicon were predominantly located in Descemet's membrane. Silicotic corneas showed significantly more silicon-containing particles than corneas of healthy controls (chi 2-test, P < 0.01). CONCLUSION: Normal corneas contain very low amounts of silicon. Longterm exposure to inhalative silicon dusts results in only very slightly increased levels of amorphous silicon in the cornea. However, silicon-containing particles accumulate in the cornea of silicotic individuals. Bricklayers incorporate more amorphous silicon into the cornea.
Aged↗
Lectin binding in normal, scarred, and keratoconus corneas.
PURPOSE: Keratoconus is characterized by central thinning and an increased curvature of cornea. Scarring of cornea accompanies the disease as it proceeds, and the ultimate treatment is keratoplasty. The pathogenesis of keratoconus is still unknown, and therefore we studied normal, keratoconus, and scarred corneas by a panel of lectin conjugates. METHODS: The following lectin conjugates were used: concanavalin A (Con A), Dolichos biflorus agglutinin (DBA), peanut agglutinin (PNA), Ulex europaeus agglutinin (UEA-I), wheat-germ agglutinin (WGA), Helix aspersa agglutinin (HAA), Psophocarpus tetragonolobus agglutinin (PTA), soybean agglutinin (SBA), Griffonia simplicifolia agglutinin-I-B4 (GSA-I-B4), Ricinus communis agglutinin (RCA-I), and Lotus tetragonolobus agglutin (LTA) conjugates, differently recognizing saccharide residues in glycoconjugates. RESULTS: One of the keratoconus corneas was badly scarred, whereas no scars were seen on histopathologic examination in other keratoconus corneas. In all keratoconus corneas, discontinuities or breaks (defects) were seen in the Bowman's layer. DBA and HAA conjugates, both blood group A reactive, reacted with corneal and conjunctival epithelium, blood vessels, and endothelium in blood groups A and AB. The blood group B-reactive GSA-I-B4 conjugate reacted with corneal and conjunctival epithelium, blood vessels, and endothelium only in specimens with blood group B. The blood group O-reactive UEA-I conjugate revealed the apical epithelial cells and blood vessels in all blood groups, whereas LTA conjugate, also blood group O reactive, reacted with all structures in cornea and conjunctiva despite blood groups. HAA and DBA conjugates reacted with defects in keratoconus corneas, SBA and RCA-I conjugates with defect area in keratoconus and scar region in scarred corneas, and PNA and WGA conjugates with the scar region of scarred keratoconus and scarred corneas. No other differences in the binding of lectins between normal, scarred, and keratoconus corneas were seen. CONCLUSIONS: The results indicate that the blood group status influences lectin binding in cornea. Furthermore, the difference in lectin binding to defects in keratoconus corneas (HAA and DBA conjugates) and to scar regions in scarred corneas (PNA and WGA conjugates) suggests that the defects in keratoconus are not solely the results of scarring.
Adolescent↗
[Organ culture for preservation of the cornea: human umbilical cord serum versus fetal bovine serum].
OBJECTIVE: To evaluate the changes of porcine corneal endothelium, the morphology, histology, ultrastructure, enzymes activity and metabolism of the cornea induced by organ culture with two different media containing fetal bovine serum (FBS) or human umbilical cord serum (HCS). METHODS: Fifty pairs of porcine corneas were preserved at 31 degrees C for 7, 14, 21, 28 days. One cornea of each pair was cultivated in medium I containing 10% FBS (group 1); the other one was stored in medium II containing 10% HCS (group 2). Thirteen fresh porcine corneas served as controls. All stored corneas were dehydrated for 24 hours. Twelve corneas from each group were evaluated each week, including the morphology, histology and enzyme histochemical staining of the cornea. Scanning electron microscopy was performed on one cornea from each group at 14 and 28 days and compared with the fresh cornea. pH value, glucose and lactate concentration of the culture media before and after culture were examined. Microbiological evaluation was also performed. RESULTS: Endothelium evaluation did not differ statistically between the two groups of porcine corneas. The morphological endothelium study showed some alterations such as pleomorphism. After 28 days of cultivation, the mean cell losses of endothelium were 10.98% and 10.85% in medium I and medium II stored corneas, respectively. There were no statistical differences of the histology, ultrastructure and enzymes activity of corneas between the two groups. The histological study showed corneal swelling and epithelial sloughing after preservation. Scanning electron microscopy showed an intact endothelial layer in all corneas. Enzyme histochemical staining showed vigorous enzyme activity in the corneal epithelium and endothelium. Enzyme activity in stroma decreased with preservation time. Corneas showed good glucose metabolism. Incidence of contamination was 6% for storage medium. CONCLUSIONS: The corneal endothelium can maintain a good viability for 4 weeks in these two organ culture media. HCS can replace FBS in the organ culture medium.
Animals↗
Modification of cornea-evoked reflex blinks in rats.
Although maintaining the tear film on the cornea is the most important role of blinking, information about the organization and modification of cornea-evoked blinks is sparse. This study characterizes cornea-evoked blinks and their modification in urethane-anesthetized rats. Cornea-evoked blinks typically begin 16.2 ms after an electrical stimulus to the cornea and last an average of 50.2 ms. In anesthetized rats, the blink only occurs ipsilateral to the stimulus. In response to cornea stimulation, the orbicularis oculi EMG activity typically exhibits two bursts that correlate with the arrival of A delta and C-fiber inputs to the spinal trigeminal complex. In the paired-stimulus paradigm, suppression of the blink evoked by the second cornea stimulus occurs for interstimulus intervals less than 300 ms and is exclusively unilateral. Stimulation of the contralateral cornea does not affect subsequent blinks evoked from stimulation of the ipsilateral cornea. To determine whether activation of cornea-related neurons in the border region between the spinal trigeminal caudalis subdivision and the C1 spinal cord (Vc/C1) inhibits the second blink in the paired-stimulus paradigm, we examine the suppression of cornea-evoked blinks caused by microstimulation in this region. This suppression of orbicularis oculi EMG activity begins 8.3 ms after Vc/C1 stimulation. Activation of this region, however, is unlike suppression in the paired-stimulus paradigm because Vc/C1 activation bilaterally inhibits cornea-evoked blinks. Thus, activation of Vc/C1 is a previously unidentified mechanism for modulating cornea-evoked blinks.
Afferent Pathways↗
Effect of benzalkonium chloride/EDTA on the ocular bioavailability of ketorolac tromethamine following ocular instillation to normal and de-epithelialized corneas of rabbits.
This study was designed to examine the effect of benzalkonium chloride/ethylenediaminetetraacetic acid (BAK/EDTA) on the ocular bioavailability (Focular) of ketorolac tromethamine after ocular instillation to normal and de-epithelialized corneas of rabbits both in vitro and in vivo. The in vitro Focular of the formulations was measured in flow-through perfusion chambers. For in vivo studies, a 35 microL dose of 0.5% ketorolac tromethamine with or without BAK/EDTA was instilled into rabbit eyes with intact or de-epithelialized corneas. At 0.5, 1, 2, 4, 6, and 8 h postdose, rabbits were euthanized, and the corneas and aqueous humor were collected from both eyes. The ketorolac concentrations from both in vivo and in vitro samples were quantified by reversed-phase high-performance liquid chromatography. The in vitro study results indicated that BAK/EDTA statistically significantly increased the Focular of ketorolac through de-epithelialized corneas but not through intact corneas. The in vivo study results showed that BAK/EDTA had no effect on the Focular of ketorolac in rabbits with intact corneas, based on the values of the area under the aqueous humor concentration versus time curves (AUC0-6h) of ketorolac. As expected, de-epithelialization of the corneas produced a faster and greater ocular absorption of ketorolac as evidenced by the smaller Tmax and larger AUC values compared to those for the intact corneas in vivo. However, BAK/EDTA decreased the ocular absorption of ketorolac in rabbits with de-epithelialized corneas. The half-lives (t 1/2) of ketorolac in corneal tissue and aqueous humor were longer in rabbits with intact corneas than those in rabbits with de-epithelialized corneas. In conclusion, the in vivo Focular of ketorolac was not altered by BAK/EDTA in rabbits with intact corneas, but it was decreased by BAK/EDTA in rabbits with de-epithelialized corneas. Therefore, the formulation with ketorolac alone may be better as a post-operative ocular analgesic.
Animals↗
Annulus of collagen fibrils in mouse cornea and structural matrix alterations in a murine-specific keratopathy.
PURPOSE: Mouse corneas were investigated to see whether a limbal annulus of corneal collagen exists as in humans. Mice with corneas predisposed to topographical changes (the SKC strain) were also examined, to establish the size and spacing of stromal collagen fibrils and the integrity of the annulus. METHODS: X-ray diffraction was used to measure collagen fibril spacing and diameter in normal (the BALB/c strain; four male, two female) and SKC (six male and six female) corneas and to identify the degree of preferred collagen orientation at 200- microm intervals across two BALB/c and four SKC corneas. RESULTS: The average collagen fibril diameter measured 35.5 nm in 3-month-old BALB/c corneas, and 36.9 nm and 37.0 nm, respectively, in corneas of age-matched male and female SKC mice. In male and female SKC corneas, average collagen interfibrillar Bragg spacing was significantly higher (64.5 and 59.9 nm, respectively) than in corneas of BALB/c mice (49.7 nm). Circumferentially aligned collagen, indicative of a limbal annulus of fibrillar collagen 2.2 mm in diameter, was identified in mouse cornea. On occasion, this was disturbed in the SKC phenotype. CONCLUSIONS: Collagen fibrils are marginally larger in the corneas of SKC mice than in the corneas of BALB/c mice and are considerably more widely spaced. An annulus of fibrillar collagen probably exists near the limbus of the normal mouse cornea that may help promote biomechanical stability and maintain corneal shape. A loss of structural integrity in the annulus of some SKC mice may predispose the corneas to biomechanical instability and shape changes.
Animals↗
Accumulation of mitochondrial DNA damage in keratoconus corneas.
PURPOSE: To determine whether keratoconus (KC) corneas have more mitochondrial (mt)DNA damage than do normal corneas. METHODS: Thirty-three normal corneas and 34 KC corneas were studied. Immunohistochemistry for mitochondria-encoded cytochrome c oxidase (complex IV) subunit 1 (CO-Iota) and porins was performed. Total DNA was isolated and mtDNA genome amplified by either long-extension-polymerase chain reaction (LX-PCR) or short-extension-PCR (SX-PCR). LX-PCR mtDNA was digested with restriction enzymes to confirm full-length mtDNA amplicon. SX-PCR mtDNA was probed by Southern blot analysis. The T414G mutation was analyzed by peptide nucleic acid directed clamping PCR. Real-time PCR measured the ratio of mtDNA to nuclear (n)DNA. RESULTS: KC corneas had decreased CO-Iota in areas of corneal thinning. LX-PCR mtDNA digested with restriction enzymes showed expected size bands except for PstI, which showed two additional bands in some KC corneas (2/18). By both LX-PCR (7.4 +/- 3.8 vs. 4.3 +/- 2.7, P < 0.04) and SX-PCR (5.5 +/- 0.55 vs. 2.4 +/- 2.0, P < 0.006), KC corneas had an increased number of smaller-sized bands (representing mtDNA deletions/mutations) compared with normal corneas. Southern blot analysis of SX-PCR products confirmed their mtDNA origin. The T414G mutation was not detected in either KC or normal corneas. KC corneas showed a trend of lower mtDNA-to-nDNA ratio (26%, P < 0.7) than did normal corneas. CONCLUSIONS: KC corneas exhibit more mtDNA damage than do normal corneas. The previously reported increased oxidative stress and altered integrity of mtDNA may be related to each other and may be important in KC pathogenesis.
Adolescent↗