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Biomedical subjects

J P McCulley

Publications and source records attributed to J P McCulley.

At least 19 recordsLinked to original sources
PubMed1992

The use of conjunctival flaps in the treatment of herpes keratouveitis.

Conjunctival flaps have been used in the treatment of corneal diseases since the 1800s, although in recent years their use has decreased. In selected cases, however, the placement of a conjunctival flap still may be the treatment of choice. We report our experience with the use of conjunctival flaps in patients with herpes keratouveitis who had persistent corneal epithelial defects. Preoperatively, all patients had chronic or recurrent epithelial defects and intraocular inflammation with or without recurrent live viral infection requiring frequent medicine application and office visits. Postoperatively, all patients had an intact, healthy ocular surface and a noninflamed eye requiring few to no medications and infrequent office visits. No patient has had recurrent live viral activity.

Conjunctiva↗
PubMed1992

Characterization and pathogenic potential of a soil isolate and an ocular isolate of Acanthamoeba castellanii in relation to Acanthamoeba keratitis.

Acanthamoeba castellanii, one isolate from the eye and one from the soil, were compared on the basis of: (a) pathogenic potential; (b) plasminogen activator activity; (c) chemotactic activity; (d) cytopathic effects; (e) collagenolytic activity; (f) binding ability to contact lenses; and (g) and binding ability to corneal buttons. The ocular isolate of A. castellanii was found to be pathogenic based on its ability to produce corneal infections in Chinese hamsters. By contrast, the soil isolate produced only mild lesions in a single Chinese hamster. Amoebae from the ocular isolate bound to corneal epithelium in greater numbers than the soil isolate counterparts. Moreover, ocular isolate organisms displayed plasminogen activator activity that was not detected in cultures from soil isolates of A. castellanii. Although neither the soil isolate nor the ocular isolate amoebae responded chemotactically to epithelial or stromal components, the ocular isolate displayed a curious and reproducible positive chemotactic response to endothelial extracts. Both A. castellanii isolates produced cytopathic effects on pig corneal epithelium, however the cytotoxicity from the ocular isolate was significantly greater than that of the soil isolate. The results indicate that the pathogenic potential of A. castellanii is correlated with the parasite's capacity to bind to corneal epithelium, respond chemotactically to corneal endothelial extracts, elaborate plasminogen activators, and produce cytopathic effects on corneal epithelium.

Acanthamoeba↗
PubMed1992

Chemotactic response of macrophages to Acanthamoeba castellanii antigen and antibody-dependent macrophage-mediated killing of the parasite.

The chemotactic potential of antigens of Acanthamoeba castellanii for macrophages and the ability of naive and immune rat peritoneal macrophages to kill A. castellanii in vitro were assessed. The amoebolytic capacity of immune rat serum and complement was also examined. No parasite was killed in the presence of heat-inactivated naive rat serum. Low numbers of parasites were lysed in the presence of heat-inactivated immune rat serum, whereas significantly greater numbers of parasites were lysed in the presence of nonheat-inactivated naive and immune rat serum. Macrophages from naive rats were capable of lysing some parasites. However, the amoebolytic capability of these cells was significantly increased in the presence of serum from immune rats. Regardless of the source of serum used, macrophages from immune rats demonstrated about twice the amoebolytic proficiency of cells from naive rats. Macrophages from naive rats showed their highest capacity for lysing amoebae when incubated in the presence of gamma interferon and immune rat serum. The greatest overall proficiency in lysing parasites was displayed by cells from immune rats incubated with A. castellanii in the presence of gamma interferon and nonheat-inactivated serum from immune rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Acanthamoeba↗
PubMed1992

Susceptibility of corneas from various animal species to in vitro binding and invasion by Acanthamoeba castellanii [corrected].

A crucial requirement for establishing corneal infection by the extracellular protozoal parasite, Acanthamoeba, is the ability of the parasite to bind to the corneal surface. In a series of in vitro studies, we examined the ability of Acanthamoeba castellanii [corrected] to adhere, invade, and damage normal, intact corneas of 11 mammalian and one avian species. A. castellanii [corrected] (80-90% trophozoites and 10-20% cysts) were incubated with corneas for 24 hours in vitro and examined by scanning electron microscopy (SEM). Results of several independent SEM experiments revealed that parasites not only failed to produce cytopathic effects but did not even bind to the corneal epithelium of mice, rats, cotton rats, horses, guinea pigs, cows, chickens, dogs, and rabbits. However, parasites adhered, invaded, and produced severe damage to human, pig, and Chinese hamster corneas during the 24-hour in vitro incubation period. Additional in vitro experiments quantified the binding of A. castellanii [corrected] to the corneas of selected susceptible and nonsusceptible species. In vitro binding assays revealed scant binding of parasites to mouse, rat, and rabbit (range = 5-20 parasites/7.07 mm2 corneal button). In contrast, extensive binding was observed on Chinese hamster, pig, and human corneas (range = 100-200 parasites/7.07 mm2 button). The results indicate that A. castellanii [corrected] exercises rigid host specificity at the host cell surface.

Acanthamoeba↗
PubMed1992

A pig model of Acanthamoeba keratitis: transmission via contaminated contact lenses.

A model of contact lens-induced Acanthamoeba keratitis was developed in Yucatan micropigs. Pigs fitted with parasite-laden soft contact lenses developed corneal infections that clinically and histopathologically mimicked the human counterpart. Three distinct stages of disease became apparent and were categorized as: acute, condensed infiltrate, and resolution stages. Viable parasites were isolated from corneal scrapings and smears were taken during the acute and condensed infiltrate stages. In addition, cysts could be identified deep within the stroma of histological specimens taken during the resolution stages. The characteristic dense, white ring-like infiltrates, stroma edema, keratic precipitates, and the chronic nature of the infections were similar to those observed in human Acanthamoeba keratitis. Histopathological examination of infected corneas revealed extensive neutrophilic infiltrates, stromal necrosis, and disorganization of the collagen lamellae. The strong correlation between the clinical and histopathologic features of contact lens-induced Acanthamoeba keratitis in the pig as well as the anatomical similarity of the pig eye with the human eye make the porcine model a valuable tool for investigations of the immunology, cell biology, and therapy for Acanthamoeba keratitis.

Acanthamoeba↗
PubMed1991

Effect of contact lens preservatives on Acanthamoeba.

Single preservatives used in contact lens solutions were evaluated for their effectiveness in killing Acanthamoeba castellanii and Acanthamoeba polyphaga trophozoites and cysts. Preservatives were tested against amoebae at intervals varying from 30 minutes to 24 hours. The preservatives were tested with axenically and nonaxenically grown organisms. Chlorhexidine (0.001% and 0.005%), polyaminopropyl biguanide (0.0015%), benzalkonium chloride (0.001% and 0.004%), and hydrogen peroxide (3%) were very effective preservatives. Lower concentrations of these same preservatives were less effective. Thimerosal (0.001% and 0.004%), sorbic acid (0.1%), potassium sorbate (0.13%), EDTA (0.1%), polyaminopropyl biguanide (0.00005%), and polyquaternium-1 (0.001%) were not effective as tested. However, thimerosal 0.004% when combined in solution with EDTA was effective. Preservatives were more effective when tested against axenically prepared organisms than when tested against co-cultured organisms.

Acanthamoeba↗
PubMed1991

In vitro penetration of human corneal epithelium by Acanthamoeba castellanii: a scanning and transmission electron microscopy study.

Human corneal buttons were exposed to Acanthamoeba castellanii trophozoites and cysts for 12 hours at 35 degrees C. The buttons examined by light microscopy and scanning and transmission electron microscopy had severe epithelial ulceration and penetration by trophozoites. Observations on trophozoites below the surface suggest that penetration is accomplished by both secreted cytolytic enzymes and phagocytosis. It is likely that the secretion of one or more enzymes constitutes the initial step in preparing the host tissue for endocytosis or that the secretory mechanism is used by the amebas to move through the outer squamous layer to the basement epithelium where phagocytosis occurs. Based on this study and a previous study, it appears that entry into the cornea is a two-step process involving adherence and penetration by trophozoites.

Acanthamoeba↗
PubMed1991

In vitro intercellular adherence of Acanthamoeba castellanii: a scanning and transmission electron microscopy study.

Human corneal buttons were exposed to trophozoites and cysts of Acanthamoeba castellanii for 12 hours. Examination of the buttons by scanning electron microscopy showed numerous trophozoites on the surface of the epithelium. Trophozoites examined by transmission electron microscopy had limited regions of attachment to the epithelium but extensive regions of attachment to each other. Attachment regions were characterized as plaque-like maculae of an incomplete desmosome junction. Firm attachment mechanisms may explain how penetration of the human cornea occurs.

Acanthamoeba↗
PubMed1991

In vitro transfer of rabbit corneal epithelium from carriers to denuded corneas or cryolathed lenticules.

Rabbit corneal basal epithelial cells seeded onto fixed gelatin membranes or commercial collagen shields formed 3 to 5 cell layers after 1 to 3 weeks of culture at 35 degrees C in nutrient medium. The cells grew better, by comparison, in the collagen shields and eventually formed a multilayered tissue that resembled the stratified morphology of native epithelium. Transfer of multilayered cultures (prior to stratification) from these carriers in vitro to denuded corneal buttons or cryolathed lenticules resulted in complete adhesion of the grafted tissue to the underlying recipient buttons after 24- to 48-h incubations. After mechanically removing the carriers, most of the epithelial cells remained attached to the stromal surface. Our experimental findings indicated that both kinds of carriers may be suitable for epithelial transplantation, although the collagen shield is probably superior because of its better biocompatibility and physical characteristics.

Animals↗
PubMed1991

Differential effect of chelation on the pH tolerance of corneal epithelium in tissue culture.

The impact of chelation on the biocompatibility of various vehicles was determined after 1- to 32-min exposures using stratified rabbit corneal epithelial cultures. Each formulation was tested at either pH 5.0 or 7.5 to mimic the specifications of most commercial ophthalmic preparations. Adding ethylenediamine tetraacetic acid (EDTA) to balanced salt solutions was unremarkable, and all of the formulations were essentially nontoxic. In contrast, EDTA moderately reduced the toxicity of the acidic acetate/citrate-buffered vehicles, but enhanced the toxicity of the alkaline solutions. The preservative, on the other hand, had little impact on the biotolerance of acidic unbuffered and phosphate-buffered vehicles, although it also increased the toxicity of the alkaline formulations. These findings are interpreted in terms of the different pKa values of the chelating agents, and an unsuspected interaction between citrate and the preservative. We believe that this type of testing can be used to develop guidelines for the use of buffers, excipients and preservatives in commercial formulations.

Animals↗
PubMed1991

Growing human corneal epithelium on collagen shield and subsequent transfer to denuded cornea in vitro.

Three fundamental in vitro experiments have been done in the present report: 1) comparison of three different nutrient media on their abilities to culture and passage the human corneal epithelial cells; 2) evaluation of the ability of extracellular matrix material to promote the growth of cultured human corneal epithelium on collagen corneal shields; and 3) determination of the feasibility of the shield to serve as a carrier for the transfer of cultured cells to allogeneic, denuded corneal surface in vitro. Primary cultures of human corneal epithelium were established from explants which were obtained from limbal and peripheral corneal tissue by three different nutrient media respectively: KGM (Keratinocyte Growth Medium), SHEM (Supplemental Hormonal Epithelial Medium), and one combination of the two media (KGM/SHEM). We found the KGM/SHEM combination to be more favorable because morphology was better preserved, the proliferation rate increased five-fold over the 14 days observed time course, and we were able to subculture the tissue for at least three passages. With this combined medium, a suspension of cultured corneal epithelial cells (5 x 10(5)/ml) was seeded onto either the concave surface of collagen corneal shields or onto shields which had been coated with extracellular matrix materials (Matrigel or type IV collagen). The cells attached readily to all the coated shields (20/20) but to only a few of the uncoated shields (3/10), and formed a stratified tissue (2 to 3 layers) within seven days once the cells attached. However, the cells on the shields coated with Matrigel failed to become confluent under these conditions. The stratified tissue on type IV collagen coated shields could then be subsequently transferred to denuded human corneal stroma in organ culture by placing them together and incubating for 2-7 days. After that, histologic examinations showed that the epithelial cells had attached tightly to the recipient stromal surface, even after the removal of the collagen shield.

Biological Dressings↗
PubMed1991

The role of tetracycline in chronic blepharitis. Inhibition of lipase production in staphylococci.

Tetracycline administered in low doses can be effective in the long-term management of patients with meibomian keratoconjunctivitis (MKC). However, the mechanism of action does not appear to be a reduction of bacteria. Seventy-five percent of the ocular staphylococci in such patients are resistant to tetracycline. An alternative mechanism of action could be the inhibition of production of extracellular enzymes by the ocular flora. Inhibition of lipase production could result in lowered levels of toxic hydrolysis products (free fatty acids), which may exacerbate the disease process. The authors tested this hypothesis by examining the differential effect of tetracycline on growth and lipase production in a tetracycline-resistant and tetracycline-sensitive strain of Staphylococcus epidermidis and S. aureus isolated from patients with MKC and Staphylococcal blepharitis. Tetracycline caused significant decreases in the production of lipase in the sensitive and resistant strains of S. epidermidis without concominant decreases in growth. In contrast, S. aureus strains showed parallel decreases in both lipase production and inhibition of growth. The authors propose that the sensitivity of lipase production to tetracycline, in tetracycline-resistant S. epidermidis, may partially explain the clinical improvement observed in MKC patients.

Blepharitis↗
PubMed1991

The role of wax and sterol ester fatty acids in chronic blepharitis.

The authors analyzed the long-chain fatty acids derived from the wax and sterol ester fractions of meibomian secretions from patients with chronic blepharitis and normal individuals. Meibomian secretions were partitioned into separate lipid classes by thin-layer chromatography (TLC). Wax and sterol esters were eluted and transesterified. The liberated fatty acid methyl esters (FAME) were analyzed by gas liquid chromatography. Equivalent chain lengths (ECL) were determined for the 58 peaks found. Thirty-three peaks were positively identified by standards. Peaks were quantified by area normalization. Percentage compositions were computed for each individual and tabulated by group; each fatty acid was analyzed by analysis of variance, and each clinical group was compared with normal subjects. The authors found increases in the series of monounsaturated fatty acids from patient wax/sterol esters compared with normal subjects (P less than 0.05). The authors also found differences in some members of the series of normal, straight, and branched saturated moieties. These differences between normal groups and the blepharitic groups represent a biologically significant pattern that may relate to the disease process. Discriminant analysis provided a 73% probability of correct classification into clinical groups based strictly on FAME analysis (P less than 0.05).

Blepharitis↗
PubMed1991

The role of cholesterol in chronic blepharitis.

Chronic blepharitis has been a difficult disease to define either microbiologically or biochemically. Sterols from meibomian secretions of normal subjects and patients were analyzed, and important differences were observed. Based on analyses of these secretions, two significantly different (P less than 0.001) types of normal subjects were found, those with and those without cholesterol esters [Norm(CP) and Norm(CA), respectively]. All patients' secretions contained cholesterol esters. Evidence was obtained which suggests that oxysterols may control the ester cholesterol accumulation. Furthermore, only when cholesterol esters were present did wax and sterol esters containing unsaturated fatty acids accumulate. Over 90% of these unsaturated fatty acids were normal (unbranched); the rest were iso-fatty acids. Preliminary results also suggest that the ester fatty alcohols are much more complex than previously reported; seven alcohols were common to all samples analyzed. Additionally, highly oxygenated alcohols were detected, especially in the meibomian keratoconjunctivitis (MKC) disease group. The MKC samples also contained an alcohol (mass, M/Z 378) not present in any of the other samples analyzed. Based on analysis of variance and linear-regression models, it was determined that the long-chain (C20-28) fatty acids were more important in determining disease signs. Furthermore, in the MKC group, the ratio of unsaturated C18 fatty acids to cholesterol in the wax and sterol esters was significantly different (P less than 0.05) from the Norm(CP) group. The authors discuss the fact that rabbit meibomian secretions are stable, despite containing a very high percentage of ester sterols, and relate this to their high percentage of branched-chain fatty acids and low percentage of unsaturated fatty acids.(ABSTRACT TRUNCATED AT 250 WORDS)

Blepharitis↗
PubMed1990

The effect of currently available contact lens disinfection systems on Acanthamoeba castellanii and Acanthamoeba polyphaga.

Contact lens disinfection systems were evaluated for their effectiveness in killing Acanthamoeba castellanii and Acanthamoeba polyphaga trophozoites and cysts. Amoebae were inoculated into commercially available contact lens cleaning and soaking solutions. At intervals varying from 30 minutes to 24 hours, solutions were filtered. The filters were removed and cultured for Acanthamoeba organisms. Striking differences were observed in the abilities of the different disinfecting solutions to kill the organisms. Solutions containing chlorhexidine were effective at very short exposure times. Solutions containing benzalkonium chloride required slightly longer exposure times but were faster than solutions containing only thimerosal. Solutions containing sorbate, polyaminopropyl biguanide, or polyquaternium-1 were not effective at killing Acanthamoeba organisms in the time allotted for the experiment. Solutions containing hydrogen peroxide were quite effective if the agent was not prematurely catalyzed. A. polyphaga generally required longer exposure to disinfectants than did A. castellanii for complete inhibition to occur.

Acanthamoeba↗
PubMed1990

Ocular hydrofluoric acid burns: animal model, mechanism of injury and therapy.

A series of ocular HF burns was produced in rabbits in order to clarify the nature of the injury and to provide a description of the animal model. Burned eyes were evaluated clinically and allowed to progress for up to 65 days before histologic examination. The mechanism of HF toxicity was investigated through the study of burns produced by chemicals chosen to mimic its pH effects, osmotic effects, and effects of the free fluoride ion alone. The severe progressive caustic effect of HF on the eyes was found to depend on the combination of pH and the toxic effects of the free fluoride ion, together causing extensive dose-related damage to superficial and deep structures of the eye. Mild burns caused reversible ocular injury; whereas more severe burns lead to corneal stromal scarring, vascularization, edema, formation of calcific band keratopathy plus iris and ciliary body fibrosis. An investigation was made of potential treatments for experimental ocular HF burns in rabbits. Topical ointments containing MgO or MgSO4 and irrigations with or subconjunctival injections of H2O or solutions containing NaCl, MgCl2, CaCl2, LaCl3, hyamine, zephiran, calcium gluconate or a mixture of divalent metal ions were tested for toxicity and for therapeutic value in ocular HF burns. Immediate single irrigation with H2O, NaCl or MgCl2 solution was most effective. Other therapeutic agents commonly used in HF skin burn therapy were either too toxic in normal eyes or caused additive damage to burned eyes.

Animals↗
PubMed1990

In vivo and in vitro collagenolytic activity of Acanthamoeba castellanii.

Axenic cultures of Acanthamoeba castellanii contained a collagenolytic enzyme that digested collagen shields and purified collagen in vitro. Specificity of biologic activity was determined by the addition of selected enzyme inhibitors to the assays and revealed that the parasite-conditioned medium contained both collagenase and lower concentrations of other proteolytic enzymes. However, most of the collagenolytic and pathogenic activity was directly attributable to specific collagenase. Intrastromal injection of sterile, Acanthamoeba-conditioned culture medium into naive Lewis rats produced corneal lesions clinically similar to and closely resembling those found in biopsy specimens of human patients diagnosed with acanthamoebic keratitis. Histopathologic analysis revealed moderate-to-severe neutrophil infiltration, disruption of stromal lamellae, and edema. Identical pathologic sequelae were produced by intrastromal injection of purified collagenase (25 units/ml). The pathogenicity of the soluble parasite-derived product was removed by passage over affinity columns armed with antibody specific for collagenase. These results indicated that soluble parasite-derived factors were capable of producing lesions characteristic of acanthamoebic keratitis and that the pathogenicity of these factors was either directly or indirectly attributable to specific collagenase activity.

Acanthamoeba↗