Collagen shield enhancement of topical dexamethasone penetration.

Collagen corneal shields were investigated as a vehicle for enhancing the ocular penetration of topical 0.1% dexamethasone alcohol in rabbit eyes. Four protocols were compared: a single dexamethasone drop, hourly drops, a 24-hour collagen shield presoaked in 0.1% dexamethasone, and a presoaked collagen shield followed by hourly drops. Dexamethasone concentrations in the cornea, aqueous, iris, and vitreous were measured by radioassay at six time intervals, and cumulative drug delivery over 6 hours was calculated for each tissue. Treatment with a presoaked collagen shield plus hourly drops resulted in peak and cumulative drug delivery to the cornea, aqueous, iris, and vitreous that was twofold to fourfold higher than delivery achieved with hourly drops alone. A presoaked shield by itself yielded equivalent or superior peak and cumulative drug delivery compared with a regimen of hourly drops. Collagen shields significantly enhance topical dexamethasone penetration and may be useful for maximizing the intraocular delivery of dexamethasone and for decreasing the required frequency of topical dexamethasone administration.

Detection of herpes simplex viral DNA in the iridocorneal endothelial syndrome.

OBJECTIVE: To test the hypothesis that the iridocorneal endothelial (ICE) syndrome has a viral origin by comparing the incidence of viral DNA in corneal specimens from patients with the ICE syndrome and from controls. DESIGN: Thirty-one corneas obtained from 25 patients with the ICE syndrome and six with chronic herpetic keratitis (n = 31) were compared with 30 control specimens obtained from 15 healthy donors and from 15 patients with other, nonviral chronic corneal diseases. METHODS: Primer pairs and polymerase chain reaction methods were used to identify and amplify either a segment of the DNA polymerase gene in the case of the herpes simplex and zoster viruses or a region of the nuclear antigen gene for the Epstein-Barr virus. The oligonucleotide amplified by polymerase chain reaction was fully characterized with the use of restriction enzyme, hybridization, and sequence analyses to determine that it contained the expected base pair sequence. RESULTS: Sixteen of 25 ICE syndrome specimens and four of six herpetic keratitis specimens were positive for herpes simplex virus (HSV) DNA. All nine ICE syndrome specimens tested were negative for the presence of DNA from the herpes zoster or the Epstein-Barr viruses. Controls were uniformly negative for HSV DNA whether they were obtained from ostensibly normal corneas (n = 15) or from corneas with intestinal keratitis, aphakic bullous keratopathy, or keratoconus (n = 15). Tissue samples cut from positive ICE syndrome specimens yielded negative results when retested after the endothelial layer was removed. These findings indicate that localization of HSV DNA is within the endothelium, the tissue primarily involved in the pathogenesis of the ICE syndrome. CONCLUSIONS: Polymerase chain reaction evidence shows that HSV DNA is present in a substantial percentage of ICE syndrome corneal specimens and that HSV-DNA is absent in normal corneas and in corneas from patients with three other chronic corneal diseases. These results provide direct evidence to support our hypothesis that the ICE syndrome has a viral origin. We discussed clinical implications, including possible therapeutic interventions.

Suppression of interferon-gamma induction of MHC class II and ICAM-1 by a 26-base oligonucleotide composed of deoxyguanosine and deoxythymidine.

Interferon-gamma (IFN-gamma) is an important cytokine released by T lymphocytes and natural killer cells which is able to induce expression of class II MHC and ICAM-1, crucial factors in cellular immune response. HeLa S3, HS 27, and NF-71-1 are cell lines which can be induced to express HLA-DR and HLA-DP by exposure to IFN-gamma. When T2 (5'GGGGTTGGTTGTGTTGGGTGTTGTGTRNH(2)3') oligonucleotide was added at 5-20 microM every other day, cell surface induction of HLA-DR and HLA-DP by IFN-gamma was suppressed in a dose-dependent manner in HeLa S3. T2 suppressive effect on HLA class II was also observed in four different nontransformed human cell lines, HS 27 at passage 18, NF-71-1 at passage 5, human corneal endothelial cell at passage 5, and human retinal pigmented epithelial cell at passage 3. Control oligonucleotides had no suppressive effect. Northern hybridization showed that HLA-DR A mRNA induction by IFN-gamma was blocked by T2 in HeLa S3 and fibroblast 143B. The suppressive effect of T2 was also reversible as continued culture of the treated cells without further addition of the oligonucleotide allowed full re-expression of HLA-DR. Further experiments showed that T2 oligonucleotide was also able to inhibit IFN-gamma enhancement of ICAM-1 (CD54) on human corneal endothelial cell and human retinal pigmented epithelial cell. We conclude that T2 oligonucleotide is effective at suppressing HLA-DR, HLA-DP and ICAM-1 induction by IFN-gamma in transformed and nontransformed cells in vitro.

Corneal allograft rejection after multiple blood transfusions.

We treated two patients with corneal allograft rejections, both of which occurred within a few weeks after administration of multiple blood transfusions. In the first case, a patient with a graft that had been previously clear for 11 years developed severe endothelial rejection six weeks after transfusion of ten units of packed red blood cells. In the second case, a graft that had been clear for 14 months irreversibly rejected two weeks after transfusion of six units of packed red blood cells. These cases suggest that multiple blood transfusions may be a risk factor for corneal allograft rejection.

Combination antibiotic supplementation of corneal storage medium.

Gram-positive cocci frequently contaminate donor corneal tissue and represent the most common cause of postkeratoplasty endophthalmitis. Although gentamicin is currently added to corneal storage medium in an effort to decrease bacterial contamination of donor tissue, it has poor or variable in vitro activity against many strains of streptococci and staphylococci. To investigate whether the antibiotic supplementation of corneal storage media could be improved, we surveyed 11 antibiotics for antimicrobial efficacy under simulated storage conditions against gentamicin-resistant strains of Staphylococcus aureus, S. epidermidis, Streptococcus pneumoniae, and St. viridans. All antibiotics showed markedly reduced activity at 4 C as compared to their predicted activity at 37 C. Bactericidal activity of streptomycin and tobramycin was enhanced by preceding 4 C storage with a three-hour period at room temperature (23 C). Under these conditions, streptomycin showed the best antimicrobial activity of the 11 antibiotics tested. Addition of gentamicin to streptomycin resulted in further improvement of activity against S. aureus and S. epidermidis, whereas the addition of penicillin G to streptomycin enhanced the activity against St. viridans. Optimal antibiotic activity (99% or more killing) against all four isolates of gentamicin-resistant gram-positive cocci was best achieved with the combination of gentamicin, streptomycin, and penicillin G, coupled with a three-hour period at room temperature before 4 C storage.

Objective assessment of transient corneal haze and its relation to visual performance after photorefractive keratectomy.

PURPOSE: Photorefractive keratectomy has the potential to cause transient corneal haze. The purpose of this study was to evaluate the relationship between transient corneal haze as measured by an objective means and high and low contrast visual performance. METHODS: In a prospective study, 44 eyes of 28 patients were examined preoperatively and at 1, 3, 6, and 12 months after photorefractive keratectomy. Five laser in situ keratomileusis and two intrastromal corneal ring segments (Intacs [KeraVision, Fremont, CA]) were included for comparison, because these procedures are not expected to cause haze. Haze was measured using a prototype objective hazemeter, TSPC-3, a modification of the Nidek EAS-1000. Visual performance was measured using high-contrast visual acuity and the Rabin Small Letter Contrast Test. RESULTS: Corneal haze was greatest at the 1-month examination and was consistent with a decrease in visual performance on both tests. Corneal haze resolved in 82% of eyes by 10 +/- 4 months after photorefractive keratectomy. However, visual performance had not returned to preoperative levels in 65% and 81% of these eyes on the high-contrast visual acuity test and the Small Letter Contrast Test, respectively. Eyes that underwent laser in situ keratomileusis and Intacs did not develop corneal haze; however, visual decrements were measured. CONCLUSIONS: As a clinical tool, the TSPC-3 hazemeter objectively measures very subtle changes in haze levels. Corneal haze appears to account for only approximately 50% of visual performance changes in the early healing period after photorefractive keratectomy. Other factors, namely topographic abnormalities, are more likely to be an important cause of persistent visual disturbances.

A phase III, placebo controlled clinical trial of 0.5% levofloxacin ophthalmic solution for the treatment of bacterial conjunctivitis.

AIM: To compare the efficacy and safety of levofloxacin 0.5% ophthalmic solution (Quixin) with placebo for treatment of bacterial conjunctivitis. METHODS: In this prospective, randomised, placebo controlled, double masked, multicentre study, 249 patients with bacterial conjunctivitis received either 0.5% levofloxacin (n = 126) or placebo (n = 123) for 5 days, administered every 2 hours on days 1-2, then every 4 hours on days 3-5. Cultures were obtained and signs/symptoms evaluated at baseline, interim, and final visits. The end point was the last evaluable observation. Primary microbial outcomes were based on culture results; clinical outcomes were based on resolution of cardinal signs. RESULTS: 117 patients (60 levofloxacin, 57 placebo) were evaluated. Microbial eradication rates were significantly greater with levofloxacin at all time points, reaching 90% at end point. In a subgroup analysis, differences in eradication rates at end point were most pronounced in children but were also statistically significant for levofloxacin in adults. Clinical cure rates were significantly greater with levofloxacin at final visit and end point. Statistically significant differences favouring levofloxacin were measured at end point for resolution of conjunctival discharge, bulbar conjunctival injection, palpebral conjunctival injection, burning/stinging, itching, and photophobia. Adverse events were similar between groups. Safety composite scores analysed by age indicated significantly fewer children on levofloxacin experienced worsening symptoms. CONCLUSIONS: Levofloxacin 0.5% ophthalmic solution is safe and effective for treatment of bacterial conjunctivitis.

Bilateral diffuse lamellar keratitis following bilateral simultaneous versus sequential laser in situ keratomileusis.

AIM: To determine the difference in the incidence of bilateral diffuse lamellar keratitis (DLK) in patients undergoing simultaneous versus sequential laser in situ keratomileusis (LASIK) as an indication of intrinsic risk for inflammation. METHODS: A retrospective non-comparative case series of 1632 eyes that had undergone bilateral, simultaneous or sequential LASIK between April 1998 and February 2001 at a university based refractive centre by three surgeons. All cases that developed clinically evident DLK were identified and reviewed. In order to identify isolated cases and exclude those caused by environmental factors, when more than one patient in a given session developed DLK, the session was excluded. The main outcome measure was the incidence of unilateral and bilateral isolated, non-epidemic DLK. RESULTS: Of 1632 eyes, 126 eyes (7.7%) of 107 patients developed at least grade 1 DLK. In six operating sessions, DLK was observed in more than one patient per session, and on this basis 13 patients were excluded. 16 of the 94 remaining patients developed DLK in both eyes (17.0%). Six of 41 patients (14.6%) in the simultaneous group, versus 10 of 53 patients (18.9%) in the sequential group developed bilateral DLK (p >0.5). CONCLUSION: In isolated, non-epidemic bilateral DLK, a similar incidence was observed regardless of whether the surgery was simultaneous or sequential, suggesting an underlying intrinsic cause for DLK.

Identification of urinary metabolites of 2,4,6-trinitrotoluene in rats by liquid chromatography-mass spectometry.

Metabolites of 2,4,6-trinitrotoluene (TNT) were found in the urine of rats fed with TNT. The urine extracts were analysed by liquid chromatography-mass spectrometry (LC-MS). Metabolites found included TNT itself as well as 2-amino-4,6-dinitrotoluene, 4-amino-2,6-dinitrotoluene and 2,4-diamino-6-nitrotoluene, indicating that reduction processes are responsible for the formation of these metabolites.

Flap haze after epithelial debridement and flap hydration for treatment of post-laser in situ keratomileusis striae.

PURPOSE: To report a case of corneal haze after attempts to eliminate post-laser in situ keratomileusis (LASIK) lamellar striae. METHODS: Case report. RESULTS: A 24-year-old woman with visually significant flap striae 2 months after LASIK underwent manual epithelial debridement and flap hydration, refloating, and stretching to eliminate the striae. Three weeks after this intervention, she developed visually significant haze that was confined to the stroma of the flap. The haze slowly improved with use of a topical steroid. CONCLUSION: Stromal haze can develop after treatment of flap striae with epithelial debridement and hypo-osmolar irrigation. We speculate that these maneuvers may have induced cell death of anterior keratocytes and led to haze formation, as can occur after simple epithelial debridement and epithelial scrape-photorefractive keratectomy.

Aspergillus fumigatus keratitis after laser in situ keratomileusis.

PURPOSE: To report a case of Aspergillus fumigatus keratitis after a laser in situ keratomileusis (LASIK) enhancement procedure. METHOD: Case report. RESULTS: A 56-year-old woman developed an ulcer in the flap 13 days after LASIK enhancement. A 4-week course of fortified antibiotics for a presumed bacterial infection followed. The ulcer progressed, causing 60% thinning of the corneal stroma. A biopsy was performed 5 weeks after onset of symptoms, and antifungal agents were initiated. Cultures showed A. fumigatus. Her cornea perforated after the biopsy, requiring cyanoacrylate and lamellar overlay sutures, but the infiltrate resolved on antifungal agents. CONCLUSION: This report is the first description of Aspergillus keratitis after LASIK. We hypothesize that the infection became established on the stromal bed during surgery and led to melting, anteriorly through the flap and posteriorly through the stroma. Diagnosis was made by a corneal biopsy and inoculation of a wide array of media. This case demonstrates the need to consider atypical organisms, including fungi, in the differential diagnosis of post-LASIK infections when there is no response to therapy and highlights the role of corneal biopsy and flap lifting in the diagnosis of this condition.

Identification of antiangiogenic and antiinflammatory proteins in human amniotic membrane.

PURPOSE: To identify the potential antiangiogenic and antiinflammatory proteins expressed in human amniotic membrane tissue. METHODS: Human amniotic epithelial and mesenchymal cells were isolated from human amniotic membranes by sequential trypsin and collagenase digestion. Total RNAs were harvested from freshly obtained human amniotic epithelial and mesenchymal cells. Antiangiogenic and antiinflammatory proteins were detected by the reverse transcriptase-polymerase chain reaction (RT-PCR) technique and further confirmed by DNA sequencing of PCR-amplified transcripts. The distribution of tissue inhibitors of metalloproteinase (TIMPs) were studied further by immunohistochemistry performed on paraffin-embedded amniotic membrane tissue. RESULTS: RT-PCR results showed that both human amniotic epithelial and mesenchymal cells express interleukin-1 receptor antagonist, all four TIMPs, collagen XVIII, and interleukin-10. Thrombospondin-1 was expressed in all of the epithelial cell specimens and in one out of five mesenchymal cell specimens. Furthermore, immunohistochemistry studies performed on freshly prepared amniotic membrane confirmed that all members of the TIMP family were present in epithelial and mesenchymal cells as well as in the compact layer of the amniotic stroma. In cryopreserved amniotic membranes, positive staining was seen in residual amniotic cells and stroma. CONCLUSIONS: Human amniotic membrane epithelial and mesenchymal cells express various antiangiogenic and antiinflammatory proteins. Some of those proteins also were found in amniotic membrane stroma. These findings may explain in part the antiangiogenic and antiinflammatory effects of amniotic membrane transplantation.

In vitro susceptibilities of bacterial ocular isolates to fluoroquinolones.

PURPOSE: To evaluate and compare the in vitro antimicrobial activity of levofloxacin versus ciprofloxacin and ofloxacin against ocular isolates from patients with bacterial conjunctivitis. METHODS: The in vitro antimicrobial susceptibilities of ocular isolates to levofloxacin, ofloxacin, and ciprofloxacin were determined using both the agar disk diffusion and broth dilution methods. RESULTS: Disk diffusion susceptibility testing disclosed that 99% (100 of 101 isolates) of gram-negative isolates and 98% (127 of 129 isolates) of gram-positive isolates were susceptible to levofloxacin; 96% (97 of 101 isolates) of gram-negative isolates and 78% (100 of 129 isolates) of gram-positive isolates were susceptible to ofloxacin; and 94% (95 of 101 isolates) of gram-negative isolates and 61% (79 of 129 isolates) of gram-positive isolates were susceptible to ciprofloxacin. Broth dilution testing disclosed that 99% (72 of 73 isolates) of gram-negative isolates and 98% (111 of 113 isolates) of gram-positive isolates were susceptible to levofloxacin; 96% (70 of 73 isolates) of gram-negative isolates and 92% (104 of 113 isolates) of gram-positive isolates were susceptible to ofloxacin; and 95% (69 of 73 isolates) of gram-negative isolates and 82% (93 of 113 isolates) of gram-positive isolates were susceptible to ciprofloxacin. CONCLUSIONS: In this study, levofloxacin demonstrated superior in vitro activity against human bacterial conjunctival isolates compared with either ofloxacin or ciprofloxacin (levofloxacin > ofloxacin > ciprofloxacin).

The intrastromal corneal ring segments: Intacs.

Although evolution of the intrastromal corneal ring technology will unquestionably continue, the ICRS in its current design is remarkably safe and offers several potential advantages, including preservation of the central cornea, reversibility and adjustability. The approval of this technology by the FDA has added a potentially unique procedure to the refractive surgeon's surgical inventory.

[Development of corneal storage medium--second report. Examination of human cornea].

PURPOSE: To develop and evaluate by histological methods a new corneal storage medium with a simple formula. METHODS: We compared a corneal storage medium which contained minimum essential medium and 2.5% chondroitin sulfate (molecular weight 27,500), pH 7.33, osmolality 320 mOsm/kg with OPTISOL-GS. Paired human donor eyes provided by the Lions Eye Bank of Oregon were stored in a moist chamber until the experiment. A cornea with scleral rim was excised and stored in OPTISOL-GS, and its fellow cornea was stored in the test medium for 5, 10, or 14 days at 4 degrees C. Histological examination of corneal endothelial cells was done by both scanning electron microscopy and transmission electron microscopy. RESULTS: On days 5 and 10, there was no significant difference in histological findings between corneas stored in OPTISOL-GS and those in the test medium. Both corneal groups developed degenerative changes with the increase of storage time, but their histological findings were similar for both storage media. On day 14, corneal endothelial cells showed marked degeneration of intracellular organelles such as a swelling of mitochondria in both media. CONCLUSION: Human corneas stored in the test medium for 14 days maintained their structure as well as those in OPTISOL-GS. This shows that the newly developed corneal storage medium composed can be used for medium-term corneal storage.