Biomedical subjects

P J Pisella

Publications and source records attributed to P J Pisella.

At least 19 recordsLinked to original sources

PubMed2003

[Flow cytometry in impression cytology during keratoconjunctivitis sicca: effects of topical cyclosporin A on HLA DR expression].

PURPOSE: Immune-based inflammation has been observed as a common mechanism of keratoconjunctivitis sicca (KCS). In KCS-affected eyes, up-regulated expression of HLA DR by conjunctival epithelial cells has been demonstrated in impression cytology (IC) specimens using a technique of flow cytometry. The purpose of this study was to monitor the effects of topical cyclosporin A on the expression of this marker over a 12-month period of treatment. METHODS: Patients with moderate-to-severe KCS included in a large European multicenter clinical trial (Cyclosporin Dry Eye Study, Allergan, Irvine, CA) underwent collection of IC specimens at baseline, month 3, month 6, and month 12. They randomly received 0.05% or 0.1% cyclosporin A or vehicle. Patients randomized to receive vehicle received 0.1% cyclosporin A from month 6 onwards. Specimens were processed and analyzed in a masked manner by flow cytometry, using monoclonal antibodies directed to HLA DR. RESULTS: We included 169 patients in this study. HLA DR expression, both in percentage of positive cells and level of expression, was highly significantly reduced after 0.05% and 0.1% cyclosporin A treatment at months 3, 6, and 12 compared with baseline values, whereas vehicle did not induce any change in HLA DR expression over time. The 0.05% and 0.1% cyclosporin emulsions were significantly more effective than the vehicle in reducing HLA DR at months 3 and 6 (0.05%) and at month 6 (0.1%). CONCLUSIONS: Topical cyclosporin A strikingly reduced HLA DR, whereas the vehicle, used as a control tear substitute, had almost no effect. This study confirms that cyclosporin A may be effective in reducing conjunctival inflammation in moderate-to-severe KCS and is consistent with clinical results in this indication.

Administration, Topical↗

PubMed2003

[Effect of preservatives on the conjunctiva: a comparative study of beta-blocker eye drops with and without preservatives in glaucoma patients].

PURPOSE: To compare the effect of timolol with or without preservatives on the conjunctival epithelium of glaucoma patients. METHODS: A retrospective study using impression cytology (IC) was conducted on patients treated with 0.5% timolol with or without 0.01% benzalkonium chloride (BAC). Fifteen eyes from 15 patients treated with timolol, BAC+ and 17 eyes from 17 patients treated with timolol, BAC- were included in two groups comparable for age and duration of treatment lasting at least 1 year. Specimens were analyzed by flow cytometry for inflammatory profile (using antibodies directed against HLA DR and ICAM-1) and mucin detection (anti-M1/MUC5AC antibody). RESULTS: IC analyses showed a significant increase in the expression of the two inflammatory markers, HLA DR and ICAM-1 in the timolol, BAC+ group and also a significant decrease in goblet cell density in the same group as compared to the timolol, BAC- group. CONCLUSION: The use of long-term preserved beta-blocker in glaucoma patients is associated with a direct subclinical epithelial toxicity in the conjunctiva, as already demonstrated by previous experimental studies.

Adolescent↗

PubMed2002

Follow up by colour Doppler imaging of 102 patients with retinal vein occlusion over 1 year.

BACKGROUND/AIM: Retinal vein occlusion (RVO) is one of the most frequent ocular vascular diseases and leads to severe vision impairment. Colour Doppler imaging (CDI) is the first method which allows distinct evaluation of arterial and venous velocities in RVO. CDI is valuable for diagnosis of RVO and shows the effects of isovolaemic haemodilution. Patients with RVO were monitored by CDI for 1 year in order to clarify venous and arterial involvement in the pathogenesis of this disease. METHODS: Patients with RVO were monitored prospectively for 1 year with clinical examinations, fluorescein angiography, and CDI every 3 months. 102 adults referred for RVO for less than 2 months were enrolled. Unaffected eyes were used as control. The maximum systolic and diastolic flow velocities and the resistance index (RI) were measured in the central retinal artery (CRA) and the maximum and minimum blood flow velocities in the central retinal vein (CRV). RESULTS: During the year of observation, branch retinal vein occlusion (BRVO), ischaemic central retinal vein occlusion (CRVO), and non-ischaemic CRVO had a distinct pattern of venous velocity changes. BRVO had a similar profile to that observed in controls. Venous velocities were continuously lower in central forms, with the lowest values in ischaemic occlusion. In contrast, a brief decrease in arterial diastolic velocity was observed in ischaemic CRVO at presentation, correlated with arteriovenous passage time on fluorescein angiography, but with rapid normalisation. CONCLUSIONS: CDI findings were correlated with the type of RVO at all times during follow up. CDI showed persistent impairment of central venous velocity in CRVO whereas there was a fast initial values recovery of the arterial velocity. These results using CDI show strong evidence of a primary venous mechanism in RVO.

Adult↗

PubMed2002

Prevalence of ocular symptoms and signs with preserved and preservative free glaucoma medication.

AIM: To determine the incidence of ocular toxicity of preservatives with glaucoma medications. METHODS: A prospective epidemiological survey was carried out in 1999 by 249 ophthalmologists on 4107 patients. Ocular symptoms, conjunctiva, cornea, and eyelids were assessed. A chi(2) test was used for differences between preserved eye drops (P) and preservative free eye drops (PF). RESULTS: 84% patients used P, 13% received PF, and 3% a combination of P and PF eye drops. All symptoms were more prevalent with P than with PF drops (p<0.001): discomfort upon instillation (43% versus 17%), and symptoms between instillations such as burning-stinging (40% versus 22%), foreign body sensation (31% versus 14%), dry eye sensation (23% versus 14%), tearing (21% versus 14%), and eyelid itching (18% versus 10%). An increased incidence (>2 times) of ocular signs was seen with P eye drops. The prevalence of signs and symptoms was dose dependent, increasing with the number of P drops. A reduction in the symptoms and signs was observed when patients changed from P to PF eye drops (p<0.001). CONCLUSIONS: Symptoms and signs are less prevalent when PF drops are used. Moreover, most of the adverse reactions induced by P glaucoma medication are reversible after removing preservatives.

Adolescent↗

PubMed2002

[Corneal stromal changes after LASIK].

PURPOSE: To assess stromal modifications after laser in situ keratomileusis (LASIK) for myopia using in vivo confocal microscopy. METHODS: Thirteen eyes from 10 patients were examined before surgery and at days 8 and 30 after surgery using an in vivo confocal microscope coupled with a Z-Scan system. Stromal morphological changes, keratocyte density, flap thickness, and subclinical haze were evaluated and compared at different time points. RESULTS: Microfolds at the Bowman's layer were found in 55%, eyes as well as variable reflectivity particles located at the interface level in all postoperative examined eyes. The mean flap and activated-cell area thicknesses were respectively, 101+/-28 micrometer and 50.5+/-14 micrometer with a significant negative correlation (r=-0.89, p=0.01). The intensity of the added peak (47.3+/-8.6% scattered light), corresponding to the subclinical haze, as measured by Z-Scan, was also negatively correlated with the flap thickness (r=-0.89, p=0.01). CONCLUSION: This study confirms the presence of microfolds and particles at the interface level as subclinical complications. Evaluating the keratocytic activation by confocal microscopy can lead to a better understanding of corneal wound healing after LASIK and can help to improve the techniques. The flap thickness seems to be involved in the cellular activation induced by LASIK.

Adult↗

PubMed2002

[Influence of corneal refractive surgery on the lacrymal film].

Functional and clinical signs of ocular surface involvement are observed after corneal refractive surgery (LASIK or PRK) and are evocative of dry eye syndrome. The surgical process induces a total or partial section or abrasion of the corneal-sensitive nerves. This anatomical consequence of surgery is responsible for a lachrymal dysregulation with reduced tear production. Neither technique is currently able to demonstrate its superiority in preserving the lachrymal secretion. These postoperative findings should be clearly explained to patients before surgery and discomfort should be prevented with artificial tears and/or punctual plugs for 1 to 6 months after surgery.

Dry Eye Syndromes↗

PubMed2001

Evaluation of corneal stromal changes in vivo after laser in situ keratomileusis with confocal microscopy.

PURPOSE: To assess by in vivo confocal microscopy the modifications of the corneal stroma after laser in situ keratomileusis (LASIK) for myopia. DESIGN: Nonrandomized comparative (self-controlled) trial. PARTICIPANTS: Sixteen eyes of 13 patients were examined before surgery and at days 8, 30, and 90, and 9 eyes were examined at 6 months postoperatively using an in vivo confocal microscope. TESTING/INTERVENTION: Stromal morphologic changes, keratocyte density, flap thickness, and subclinical haze were evaluated and compared at different time points. LASIK was performed with a Flapmaker microkeratome (Solan Ophthalmic products, Jacksonville, FL) and a Lasersight LSX excimer laser (LaserSight Technologies Inc., Winter Park, FL). MAIN OUTCOME MEASURE: Confocal microscopy results. RESULTS: Microfolds at the Bowman's layer were found in most eyes, as well as variable reflectivity particles (pa) located at the interface level in all eyes examined postoperatively. The density of these particles significantly decreased with time with, respectively, 504 +/- 101 pa/mm2 at day 8 and 380 +/- 111 pa/mm2 at day 30 (P = 0.003), 332 +/- 100 pa/mm2 at month 3 and 312 +/- 40 pa/mm2 at month 6. The mean flap and the activated-cells area thicknesses were, respectively, 102 +/- 26 microm and 61 +/- 19 microm and showed significant negative correlation (P < 0.0001). The intensity of the added peak (47.3 microm 8.6%), corresponding to the subclinical haze, realized by Z-scan measure, was also negatively correlated with flap thickness (P = 0.01). Keratocyte (k) density quantified in the posterior stroma significantly increased from day 0 (480 +/- 67 k/mm2) to day 8 (701 +/- 41 k/mm2, P < 0.0001 compared with day 0) and day 30 (917 +/- 143 k/mm2, P = 0.0006, compared with day 0) but significantly decreased at 3 months postoperatively (597 +/- 56 k/mm2, P < 0.0001 compared with day 30) to reach the initial level at month 6 (502 +/- 41 k/mm2, nonsignificant compared with day 0). There was no correlation between preoperative or postoperative spherical equivalent and the density of particles, keratocytes, and the haze intensity. CONCLUSIONS: This study confirms the presence of microfolds and particles at the interface level, as well as subclinical impairment. Evaluation of keratocyte density constitutes a major contribution of confocal microscopy toward an understanding of the keratocyte response to corneal wound healing after corneal refractive surgery. Moreover, flap thickness seems to be involved in the postoperative cellular activation with a higher response when thin.

Adult↗

PubMed2001

Ocular surface changes induced by contact lens wear.

PURPOSE: To evaluate subclinical inflammation and mucus production of the conjunctiva in asymptomatic contact lens (CL) wearers, and to obtain an estimation of the chronologic variations in each group. METHODS: Eighteen eyes fitted with rigid CL (RCL) and 28 eyes with soft CL (SCL) worn daily were compared with 10 eyes from five healthy non-CL wearers. Impression cytology (IC) specimens were collected after clinical examination and were analyzed by flow cytometry using antibodies directed to HLA DR and intercellular adhesion molecule type 1 (ICAM-1) (CD 54), as inflammatory markers, and to the peptidic core of the conjunctival mucin (M1/MUC5AC) for mucus and goblet cell detection. The percentage of positive cells was calculated, and levels of fluorescence expression were quantified and compared between each group. RESULTS: A significant increase of HLA DR and ICAM-1 was observed in the SCL group in comparison with the control group. The two inflammatory markers were highly positively correlated with each other. Mucin detection with M1/MUC5AC did not find a significant difference between each group in terms of percentage of positive cells, but analyses of mean levels of fluorescence showed a significant decrease in the two CL groups. Evolution in time was different for each group, with a regular low level of inflammation in the RCL group in the first 10 years in comparison with the SCL group. In the SCL group, inflammation seemed to be higher before 2 years and after 10 years of wear. Mucin expression was variable in time, but without significant difference at any time. CONCLUSION: This study confirms difference in expression of subclinical conjunctival inflammation in asymptomatic CL wearers, with lower levels for RCL than SCL wearers with daily or extended wear. The mucin system is also modified by this low but chronic aggression of the ocular surface, with a tendency to decrease with time in the RCL and SCL groups.

Adult↗

PubMed2001

Flow cytometric analysis of inflammatory markers in KCS: 6-month treatment with topical cyclosporin A.

PURPOSE: Immune-based inflammation has been observed as a common mechanism of keratoconjunctivitis sicca (KCS). In KCS-affected eyes, upregulated expression of HLA DR and various immune- or apoptosis-related markers by conjunctival epithelial cells has been demonstrated in an earlier study, by a technique of flow cytometry in impression cytology (IC) specimens. The purpose of this study was to monitor the effects of topical cyclosporin A on the expression of these markers throughout a 6-month period of treatment. METHODS: Patients with moderate to severe KCS included in a large European multicenter clinical trial (Cyclosporin Dry Eye Study, Allergan, Irvine, CA) underwent collection of IC specimens at baseline, month 3, and month 6. For 6 months, they randomly received 0.05% or 0.1% cyclosporin A or vehicle. Specimens were processed and analyzed in a masked manner by flow cytometry, using monoclonal antibodies directed to HLA DR, CD40, CD40 ligand, Fas, and the apoptotic marker APO2.7. Percentages of positive cells were calculated and levels of expression quantified after conversion into standardized units of fluorescence. RESULTS: One hundred fifty-eight patients had at least two IC specimens available for flow cytometry analysis. HLA DR expression, both in percentage of positive cells and level of expression, was highly significantly reduced after 0.05% and 0.1% cyclosporin A treatment at months 3 and 6 compared with baseline values, whereas vehicle did not induce any change in HLA DR expression over time. The 0.05% and 0.1% cyclosporin emulsions were significantly more effective than the vehicle in reducing HLA DR at months 3 and 6 (0.05%), and at month 6 (0.1%). CD40 expression was significantly reduced at month 3 and partially at month 6, compared with baseline, with no reduction in patients who received the vehicle. CD40 ligand expression also decreased at months 3 and 6 in patients taking both concentrations of cyclosporin A. APO2.7 expression was significantly increased in all three groups, whereas percentage of Fas-positive cells decreased only in patients treated with 0.05% cyclosporin A at months 3 and 6. CONCLUSIONS: Flow cytometry provided an objective technique to monitor the effects of topical cyclosporin A on immune- and apoptosis-related markers in the conjunctival epithelium of patients with KCS enrolled in a large multicenter trial. Topical cyclosporin A strikingly reduced HLA DR and to a lesser extent, other inflammatory and apoptotic markers, whereas the vehicle, used as a control tear substitute, had almost no effect. This study confirms that cyclosporin A may be efficient in reducing conjunctival inflammation in moderate to severe KCS and is consistent with clinical results in this indication.

Administration, Topical↗

PubMed2001

Quaternary ammoniums and other preservatives' contribution in oxidative stress and apoptosis on Chang conjunctival cells.

PURPOSE: To investigate some of the toxicity mechanisms of 10 preservatives currently used in ophthalmic solutions in vitro. METHODS: A continuous human conjunctival cell line was treated with different concentrations of various preservatives for 15 minutes and for 15 minutes followed by 24 hours of cell recovery: three benzalkonium chlorides (BACs) with different hydrocarbon chain length, benzododecinium bromide (BOB), cetrimide (Cet), phenylmercuric nitrate (PM), thimerosal (thi), methyl parahydroxybenzoate (MPHB), chlorobutanol (clb), and EDTA. An inhibition study was then conducted using a 1-hour vitamin E pretreatment followed by a 15-minute BAC treatment. Membrane integrity was assessed using a neutral red test and chromatin condensation with a Hoechst 33342 test. Reactive oxygen species were measured using dichlorofluorescein diacetate test for H2O2 production and hydroethidine test for O2.- production. These tests were performed using microplate cold light cytofluorometry. Cell size and DNA content were also analyzed using flow cytometry. Confocal microscopy was used to explore morphologic changes. RESULTS: A significant decrease of membrane integrity with chromatin condensation was observed with all the quaternary ammoniums tested at concentrations of 0.005% and higher. The effect was amplified after 24 hours of cell recovery. The other preservatives tested did not decrease membrane integrity. H2O2 production was observed with all the preservatives, whereas O2.- production was significantly higher with the quaternary ammoniums at 0.005% and 0.01%, compared with the other preservatives. Flow cytometry results confirmed the cytotoxicity observed with cold light cytofluorometry. CONCLUSIONS: The quaternary ammoniums tested (BAC, BOB, and Cet) were the most cytotoxic preservatives in the current model. An apoptotic mechanism appeared to be present at low concentrations of quaternary ammoniums, whereas a necrotic process appeared at higher concentrations. Superoxide anions may play an important role in tissue damage induced by preservatives in ocular surface disorders.

Apoptosis↗

PubMed2001

[Antioxidant effects of mast cell inhibitors in a human conjunctival cell line].

PURPOSE: The aim of this study was to evaluate in vitro antioxidant effects of two mast cells inhibitors. METHODS: Cytotoxicity tests were done on a continuous human conjunctival cell line using microplate cold light cytofluorimetry. Membrane integrity (neutral red test), DNA condensation (Hoechst 33342 test), and reactive oxygen species (ROS) production (dichlorofluoresceine diacetate and hydroethidine tests) were evaluated on living cells treated with sodium cromoglycate and N-acetyl-aspartyl glutamic acid (NAAGA) preserved (benzalkonium chloride: BAC at 0.01%) and unpreserved after 60 minutes of treatment or 60 minutes and 24 hours of cell recovery. They were tested pure and at a 1/10 dilution. ROS production was also evaluated after a 60 minute pretreatment with antiallergic drugs and a 15-minute treatment with BAC, according to previous experiments performed on BAC showing its ROS production properties. RESULTS: No cytotoxicity was observed with the unpreserved formulations of antiallergic drugs. An apoptotic phenomenom was suggested with preserved drugs after a 1-hour treatment, whereas a necrotic mechanism appeared after a 24-hour cell recovery period. A ROS production decrease was observed with the two preserved and unpreserved drugs tested (p<0.001 compared to BAC) even if it was significantly higher with cromoglycate formulations. A ROS production decrease also was detected after a pretreatment with antiallergic drugs and treatment with BAC (p<0.001 compared to BAC alone). CONCLUSION: In vitro, no cytotoxicity was found with the two unpreserved mast cell inhibitors tested. An antioxidant effect also was observed with these two molecules; sodium cromoglycate appeared to be the best free radical scavenger.

Anti-Allergic Agents↗

PubMed2001

[The lipid layer of the lacrimal tear film: physiology and pathology].

The preocular tear film (POTF) is composed of a deep aqueous-mucin phase that supports a thin superficial lipid phase. The tear lipid layer (TLL), although thin, stabilizes the POTF providing a 25% surface-tension decrease and a 90-95% aqueous evaporation reduction. TLL is formed from lipids secreted by tarsal meibomian glands and spread onto the ocular surface by blinking. The TLL itself is composed of two phases. A thin and deep polar phase, adjacent to the aqueous-mucin layer, has a surfactant role. A thicker and superficial nonpolar phase has antievaporative properties. At the same time, tear lipocalins help the TLL spread and stabilize the lipid-aqueous interface. For clinical examination, TLL is directly observed with the Tearscope. POTF stability and the evaporation rate depend on the lipid layer pattern. When chronic, POTF qualitative trouble (evaporative syndrome) due to a TLL anomaly, leads to secondary ocular surface impairment with increased tear instability and self-propagation of ocular dryness. Meibomian gland dysfunction (MGD) results from local pathology, dermatologic disease (ocular rosacea) or iatrogenic etiology. Cosmetic use is the other principal cause of TLL destabilization. Lid hygiene is the mainstay of MGD treatment. Systemic antibiotics (cyclins) can be associated in cases of severe symptoms. Topical treatment is useful if there is marginal lid inflammation or infection.

Humans↗

PubMed2001

Mitochondrial activity and glutathione injury in apoptosis induced by unpreserved and preserved beta-blockers on Chang conjunctival cells.

PURPOSE: Quaternary ammonium ions have been demonstrated to induce apoptosis correlated with superoxide anion production in vitro. The purpose of this study was to further investigate the mechanisms of benzalkonium chloride (BAC), unpreserved and preserved beta-blocker eye-drops-induced programmed cell death, with special attention to the roles of mitochondrial transmembrane potential and intracellular reduced glutathione. METHODS: Chang conjunctival cells were incubated with different concentrations of unpreserved or preserved timolol (0.1%, 0.25%, and 0.4%), or carteolol (1% and 2%), or BAC (0.0001% to 0.01%) for 15 minutes, or for 15 minutes with a 24-hour recovery period in normal medium. Cellular viability (neutral red test), mitochondrial activity (rhodamine 123 test), intracellular reduced glutathione (monochlorobimane test), DNA condensation (Hoechst 33342 test), and reactive oxygen species (ROS) production (dichlorofluorescein diacetate and hydroethidine tests) were evaluated using microplate cold-light cytofluorometry. RESULTS: A significant, concentration-dependent decrease in cellular viability was found with preserved beta-blockers and with BAC alone, whereas unpreserved preparations did not show any toxicity. Only preserved beta-blockers induced chromatin condensation associated with an alteration of mitochondrial activity and a decrease of glutathione, suggesting an apoptotic phenomenon. BAC increased glutathione after 15 minutes, whereas a decrease was observed after a recovery period. ROS production was found with preserved formulations at significantly higher levels than those observed with unpreserved drugs. CONCLUSIONS: This in vitro study demonstrates that oxidative stress, evidenced by enhanced ROS production and mitochondrial injury rather than by cellular glutathione depletion, is a mechanism involved in apoptosis induced by preservative-containing eye-drops.

Adrenergic beta-Antagonists↗

PubMed2001

[Human onchocerciasis and "sowda" in the Republic of Yemen].

The geophysics of the north Yemen, associating a north-south directed mountainous fish bone (rising in more of 2,000 meters), to numerous rivers or "wadis" is convenient to the development of simulium shelters, main vectors for cutaneous filariasis to Onchocerca sp. Following several missions of bio-clinical and epidemiological evaluations in neighbouring villages of wadis, it has been possible to study different clinical aspects: one reminding the classical african onchocerciasis with generalized and diffused dermatitis, and, on an other hand, a hyperreactive dermatitis on one side of the body and associated with a collateral lymphatic ganglion. This disease is well known for local populations as "aswad" meaning "black" or "sowda". Clinically whatever the studied focus, coexists the two types of onchodermatitis (uni or bilateral). Yhe sowda patients are proportionally less numerous than those touched by the generalized type. Frequent eye lesions of the West African onchocerciasis are not found in sowda cases. In classical optical microscopy, microfilaria is morphologically indifferenciable between sowda and onchocerciasis clinical aspects. Skin snips were carried out on patients of both groups. Identification of microfilaria by molecular biology through the study of the DNA genome was done out of 5 skin snips. Microfilaria was kept dry between laminas and the DNA extracted from rehydrated microfilaria. DNA was intensified with specific primers of Onchocerca type (O150PCR). This phase was followed by hybridisation of amplification products by PCR to specific stains: OVS-2 for Onchocerca volvulus species, OCH for Onchocerca ochengi, PFS1 and PSS1-BT respectively for the forest strain and the savannah strain of Onchocerca volvulus as described previously. We can distinguish 2 kinds of answers based on the clinical origin of the snip-tests: the first one concern 3 patients with numerous dermal microfilariae but without any clinical sowda and corresponding to microfilaria O. volvulus type but different from the forest or savannah strains found in sub-Saharan Africa. The second one corresponds to 2 patients with less than 5 microfilaria in their snip-test. They show the typical clinical picture of sowda. They are identified as microfilaria type Onchocerca but they do not belong to species volvulus, or to species ochengi. It seems quite probable that the clinical picture of sowda be the result of developing onchocerciasis of animal origin and not identified as to day. The ivermectin, therapeutic of choice for African onchocerciasis in annual unique cure seems less effective in the coverage of sowda. In that case rehearsal of cures every 3 months would be necessary for mass campaigns to limit the transmission of this filariasis.

Animals↗

PubMed2000

Flow cytometric analysis of conjunctival epithelium in ocular rosacea and keratoconjunctivitis sicca.

PURPOSE: To investigate by flow cytometry and impression cytology (IC) specimens the inflammatory status of the conjunctival epithelium and goblet cell density in two series of patients with rosacea and dry eye syndrome compared with a population of healthy subjects. DESIGN: Nonrandomized, prospective, comparative case series. PARTICIPANTS: Twenty-six eyes of 13 patients with rosacea, 26 eyes of 13 patients with dry eye syndrome, and 24 eyes of 12 control subjects were included in this study. METHODS: IC specimens were collected after clinical examination of the ocular surface and analyzed by flow cytometry, using antibodies directed to human lymphocyte antigen-DR (HLA-DR), intercellular adhesion molecule-1 (ICAM-1) (CD 54), and the peptidic core of the conjunctival mucin (M1/MUC5AC). The percentage of positive cells was calculated and levels of fluorescence expression quantified and compared with those obtained in a series of 12 healthy subjects. MAIN OUTCOME MEASURES: Tear break-up time (TBUT), Schirmer test, fluorescein and lissamin green stainings, and IC were realized in this study. RESULTS: A significant increase of HLA-DR and ICAM-1 expressions by epithelial cells was consistently found in the two pathologic groups compared with levels calculated in normal eyes. The two markers were well correlated with each other and inversely with TBUT and Schirmer test. The percentage of goblet cells was significantly decreased in rosacea patients and in dry eye patients compared with the normal group with a significant negative correlation with both HLA DR and ICAM-1 markers. CONCLUSIONS: Ocular rosacea and keratoconjunctivitis sicca were associated with severe ocular surface changes, such as an overexpression of inflammatory markers and a significant decrease in the number of goblet cells.

Adult↗

PubMed2000

[Evaluation of free radical production by conjunctival impression cytology of patients treated with long-term antiglaucoma drugs or of contact lens wearers].

PURPOSE: To investigate free radical production in impression cytology specimens using microplate cold light cytofluorimetry. METHODS: 60 impression cytology specimens (IC) were harvested in 30 patients aged from 20 to 90 years. IC were taken in 10 healthy subjects, 10 glaucoma patients receiving longterm treatment for glaucoma and 10 contact lens wearers for more than five years. A complete ophthalmologic examination (BUT, corneal staining) was performed. Evaluation of free radical production was done using a dichlorofluoresceine diacetate (DCFHDA) test for H(2)O(2) evaluation and a hydroethidine test for O(2)(.)- measurement. RESULTS: The BUT values in the two pathological groups were significantly lower than those of controls (p<0.001). A free radical production was found in both contact lens wearers (p<0.001 compared to control values) and glaucoma patients (p<0.001 compared to control values). Both hydrogen peroxyde and superoxide anion productions were significantly correlated to the decrease of BUT values and the increase of superficial punctuate keratitis. CONCLUSION: The results of this preliminary study confirm that conjunctival epithelial cells may abnormally produce oxygen free radicals in chronic ocular surface disorders. This may participate to the cellular alterations observed in inflammatory diseases of ocular surface.

Adult↗

PubMed2000

Flow cytometric analysis of inflammatory markers in conjunctival epithelial cells of patients with dry eyes.

PURPOSE: To investigate in impression cytology (IC) specimens the expression of inflammatory and apoptosis-related markers by conjunctival epithelial cells from patients with dry eye as a rationale for treatment with topical cyclosporine. METHODS: Immunologic anomalies were identified at baseline, before treatment with the masked medication, in a homogeneous series of patients with dry eye syndrome, who were enrolled in a large European multicenter clinical trial (Cyclosporin A Dry Eye Study; Allergan, Irvine, CA). IC specimens were collected in 243 patients with moderate to severe keratoconjunctivitis sicca (KCS), with or without Sjogren's syndrome (SS). Fifty normal subjects were separately examined to provide normal control values. Specimens were analyzed in a masked manner by flow cytometry, using antibodies directed to markers of the immune system and/or apoptotic pathway: HLA DR, CD40, CD40 ligand, Fas, and APO2.7. Levels of expression were quantified, and results were compared with those obtained in the 50 normal patients. RESULTS: One hundred sixty-nine specimens were successfully interpreted at baseline, including 41% from patients with SS. A highly significant increase of HLA DR expression by conjunctival cells was found in KCS-affected eyes compared with normal eyes, which did not express this marker or did so very weakly. HLA DR expression in eyes with SS was significantly higher than in KCS-affected eyes without SS. Fas and APO2.7 were found at low levels in all normal and KCS-affected eyes. CD40 and CD40 ligand expressions were significantly increased in eyes with KCS compared with normal eyes. HLA DR, CD40 and Fas were found at significantly higher levels in the SS group than in the non-SS group. CONCLUSIONS. Conjunctival cells from patients with dry eye with moderate to severe KCS, with or without SS, overexpress inflammatory and apoptosis-related markers. Whether inflammation is a primary phenomenon in KCS or is the consequence of repetitive abrasion of the ocular surface after tear film deficiency remains to be determined. These data, nevertheless, support the use of immunomodulatory and/or anti-inflammatory drugs in the treatment of patients with KCS.

Adolescent↗

PubMed2000

[Cytotoxicity evaluation of three tear substitutes used in the treatment of dry eye syndromes].

PURPOSE: To investigate cell tolerance of three tear substitutes used in the treatment of dry eye syndromes. METHODS: Cytotoxicity tests were done on a continuous human conjunctival cell line using microplate cold light cytofluorimetry. Membrane integrity, DNA condensation and reactive oxygen species (ROS) production (hydrogen peroxyde and superoxide anion) were evaluated after 15 minutes of treatment or 15 minutes and 24 hours of cell recovery. Hyaluronic acid, hydroxypropyl methylcellulose associated with sodium perborate (HPMC) and carbomer 934P were tested at their commercial concentrations (respectively 0.18%, 0.3% and 0.3%) and after a 1/10 dilution. RESULTS: Cell viability and chromatin condensation were not altered by hyaluronic acid for all concentrations and times tested. A decrease in membrane integrity was observed with 0.3% carbomer 934P after 24 hours of cell recovery and with 0.3% HPMC after 15 minutes. This decrease was amplified after 24 hours and associated with an apoptotic phenomenon. A H(2)O(2) production was observed with HPMC associated with sodium perborate and an O(2) production was found with hyaluronic acid diluted at 1/10. CONCLUSION: Hyaluronic acid, carbomer and HPMC are in vitro well-tolerated even if HPMC induces a more important decrease of cell viability compared to the other drugs. Hyaluronic acid, with its rheologic properties, with no in vitro toxicity, seems to be efficient for dry eye patients.

Borates↗