epidermal-growth-factor and Eye-Burns

20 eyes of 11 patients suffering from most severe eye burns of grade 4 or worse were treated in addition to various other drugs with epiderm growth factor (EGF) or placebo in a double-blind test. Among 10 eyes treated with EGF, 6 achieved complete regeneration of the corneal epithelium, and 1 additional case, of the conjunctival epithelium. Among the eyes receiving placebo, only 3 out of 10 healed; all of them were also treated with fibronectin. Although in these very difficult cases EGF could not be investigated as a single therapy, and various other undefined factors may have been effective, in the extended course of the disease the period of EGF treatment was marked by a significant better regeneration of the epithelium.

Topics: Burns, Chemical; Clinical Trials as Topic; Double-Blind Method; Epidermal Growth Factor; Eye Burns; Fibronectins; Humans

Epidermal growth factor (EGF) is used to treat alkali-burned corneas. However, EGF-induced corneal angiogenesis, which is currently untreatable, is a side effect of this therapy. We therefore explored the role of the intermediate-conductance Ca(2+)-activated K(+) channel (KCa3.1) in EGF-induced angiogenesis and tested whether KCa3.1 blockade can suppress EGF-induced corneal angiogenesis. The proliferation, migration and tube formation of HUVECs (human umbilical vein endothelial cells) in response to EGF, the MEK inhibitor PD98059 and the KCa3.1 inhibitor TRAM-34 were analyzed in vitro via MTT, cell counting, scratch and tube formation assays. The protein and mRNA levels of KCa3.1, phosphorylated-ERK (P-ERK), total-ERK (T-ERK), cyclin-dependent kinase 4 (CDK4), vimentin and MMP-2 were assessed via western blotting and RT-PCR. KCa3.1 and vimentin expression were also detected through immunofluorescence staining. Flow cytometry was performed to examine the cell cycle. Further, an in vivo murine alkali-burned cornea model was developed and treated with EGF and TRAM-34 eye drops to analyze the effect of these treatments on corneal healing and angiogenesis. The corneas were also analyzed by histological staining. The in vitro results showed that EGF induces the upregulation of KCa3.1 and P-ERK in HUVECs and that this upregulation is suppressed by PD98059. EGF stimulates proliferation, migration and tube formation in HUVECs, and this effect can be suppressed by TRAM-34. TRAM-34 also arrests HUVECs in the G1 phase of the cell cycle and downregulates CDK4, vimentin and MMP-2 in these cells. The in vivo results indicated that TRAM-34 suppresses EGF-induced corneal angiogenesis without affecting EGF-induced corneal wound healing. In summary, the upregulation of KCa3.1 may be crucial for EGF-induced angiogenesis through the MAPK/ERK signaling pathway. Thus, KCa3.1 may be a potential target for the treatment of EGF-induced corneal angiogenesis.

Topics: Animals; Blotting, Western; Burns, Chemical; Cell Movement; Cell Proliferation; Cornea; Corneal Neovascularization; Cyclin-Dependent Kinase 4; Disease Models, Animal; Epidermal Growth Factor; Eye Burns; Flavonoids; Flow Cytometry; Human Umbilical Vein Endothelial Cells; Humans; Intermediate-Conductance Calcium-Activated Potassium Channels; Male; Matrix Metalloproteinase 2; Mice; Mice, Inbred BALB C; Mitogen-Activated Protein Kinases; Phosphorylation; Protein Kinase Inhibitors; Pyrazoles; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sodium Hydroxide; Up-Regulation; Vimentin; Wound Healing

By using both in vivo and in vitro (organ-cultured) systems, the optimal concentrations of hEGF to enhance epithelial healing after alkali wounds were evaluated in the rabbit cornea.. Alkali-injured corneas (pi = 5.5 mm, 1 N NaOH, 60 s) were treated with 0.01, 0.1, 1.0, 10 and 100 ng/ml hEGF for the in vitro study. The healing of epithelium and endothelium was determined at 1, 2, 3, 4, and 6 days after treatment. For the in vivo experiment, the eyes were treated with 2, 5, 10, and 50 microg/ml hEGF 3 times per day. The measurement of epithelial healing rate, transmission electron microscopy and immunohistochemical observation were performed after 7 days treatment.. In in vitro tests, hEGF enhanced the epithelial healing rates, showing a maximum enhancement at the concentration of 1.0 ng/ml, and endothelial healing was increased at 100 ng/ml. In in vivo studies, no significant difference was observed in the rates of epithelial healing between control and each hEGF-treated group. Among the tested concentrations, 5 microg/ml hEGF induced the most active proliferation of basal cells and 50 microg/ml hEGF remarkably produced a vascular ingrowth to the central wound area. The thickness of re-surfaced epithelium was increased by hEGF in a concentration-dependent manner.. The results of the present study indicate that a low concentration of hEGF may selectively enhance epithelial healing without affecting endothelial healing. The optimal concentration of hEGF for the stimulation of epithelial healing appears to be 5 microg/ml in rabbit corneal alkali wounds.

Topics: Animals; Burns, Chemical; Cell Count; Corneal Diseases; Dose-Response Relationship, Drug; Endothelium, Corneal; Epidermal Growth Factor; Epithelium, Corneal; Eye Burns; Humans; Immunoenzyme Techniques; Organ Culture Techniques; Proliferating Cell Nuclear Antigen; Rabbits; Sodium Hydroxide; Time Factors; Wound Healing

The local treatment effects of EGF forms on alkali burned mice corneal wounds were identified. The corneal wounds were induced by 0.5 M NaOH solution on the corneal surfaces of the mice. The local epidermal growth factor solutions (100 ng/ml) and gel form in 0.2 per cent w/w carbopol 940 (100 ng/ml) were dropped in 5 microliters aliquots into the eye twice a day. The corneal wounds were measured for 15 days at 7-day intervals and examined histologically at the end of 15th day of the experimental period. The results indicated that topical epidermal growth factor treatment in solution improved the healing of alkali burned corneal wounds when compared with epidermal growth factor delivered in a polymer system.

Topics: Administration, Topical; Animals; Burns, Chemical; Cornea; Drug Carriers; Epidermal Growth Factor; Eye Burns; Female; Male; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Solutions; Wound Healing

Healing of corneal alkali injuries remains a severe clinical challenge. The authors evaluated the effect of a new synthetic inhibitor of matrix metalloproteinases (GM6001 or N-[2(R)-2-(hydroxamido carbonylmethyl)-4-methylpentanoyl]-L-tryptophane methylamide) on preventing ulceration of rabbit corneas after alkali injury. Topical treatment of corneas with severe alkali injuries with 400 micrograms/ml or 40 micrograms/ml GM6001 alone prevented ulceration for 28 days, although 8 of 10 corneas treated with vehicle perforated. Corneas treated with 4 micrograms/ml GM6001 had midstromal depth ulcers. Corneas treated with 400 micrograms/ml of GM6001 contained very few inflammatory cells and had significantly reduced vessel ingrowth compared with vehicle-treated corneas. Epithelial regeneration after moderate alkali injuries also was investigated. Persistent epithelial defects developed 4 days after moderate alkali injury in rabbit corneas treated with vehicle and progressively increased to an average of 20% of the original 6 mm diameter wound by 27 days after moderate alkali injury. By contrast, epithelial regeneration was complete and persisted for 21 days for corneas treated with a formulation containing GM6001 (400 micrograms/ml), epidermal growth factor (10 micrograms/ml), fibronectin (500 micrograms/ml), and aprotinin (400 micrograms/ml). Sporadic punctate staining developed in 20% of the corneas treated with the combination of agents between days 21-28 after moderate alkali injury. These results demonstrate that topical application of GM6001 prevented corneal ulceration after severe alkali injury and that a combination containing GM6001, epidermal growth factor, fibronectin, and aprotinin promoted stable regeneration of corneal epithelium after moderate alkali injury.

Topics: Alkalies; Animals; Aprotinin; Burns, Chemical; Cornea; Corneal Injuries; Corneal Ulcer; Dipeptides; Dose-Response Relationship, Drug; Epidermal Growth Factor; Extracellular Matrix; Eye Burns; Fibronectins; Metalloendopeptidases; Rabbits; Regeneration

Human recombinant epidermal growth factor (hEGF) was evaluated in various corneal wound healing models in the rabbit. Human EGF accelerated epithelial wound healing in corneal reepithelialization, anterior-keratectomy, and alkali-burn models at concentrations of 10-500 micrograms/ml given four times daily (qid). In the corneal reepithelialization model, 100 micrograms/ml of hEGF qid produced a 45% increase in the wound-healing rate compared with control (0.13 versus 0.09 mm/hr) with a similar response at 500 micrograms/ml qid. In the anterior-keratectomy model, 500 micrograms/ml of hEGF qid accelerated healing by 40% (0.07 versus 0.05 mm/hr), although the 100 micrograms/ml dose was not active in this model, and 1 microgram/ml of hEGF actually slowed the healing rate. In the alkali-burn model, 10 and 100 micrograms/ml of hEGF qid for 32 days appeared to produce faster initial healing of the wound compared with control, although the wound recurred in both hEGF and control groups. These results suggest that hEGF may be helpful in some epithelial disorders in humans, although considerations of dose response and optimal dosing regimens must be addressed.

Topics: Animals; Burns, Chemical; Cornea; Corneal Injuries; Dose-Response Relationship, Drug; Drug Evaluation; Epidermal Growth Factor; Epithelium; Eye Burns; Humans; Rabbits; Recombinant Proteins; Sodium Hydroxide; Wound Healing

The proliferation and collagen synthesis of keratocytes during corneal wound healing after alkali-burn were investigated using 3H-thymidine or 3H-proline autoradiography. The effect of epidermal growth factor (EGF) on keratocytes was also examined. On day 1 after burn, the wounded stroma lacked keratocytes, and keratocytes at the periphery of the wound started to proliferate from the endothelial side. On day 7 and day 14, the population of keratocytes returned to normal. Collagen synthesis activity of the keratocytes was observed from day 7 through day 56, with the highest activity on around day 21. After day 14, keratocytes at the site of epithelial reopening or ulceration started to proliferate again. In the early stage of wound healing more keratocytes incorporated H-thymidine in eyes treated with EGF than those treated without EGF, suggesting that EGF may stimulate the proliferation of keratocytes in the early stage of wound healing.

Topics: Alkalies; Animals; Burns, Chemical; Cell Division; Cornea; Corneal Injuries; Epidermal Growth Factor; Eye Burns; Rabbits; Wound Healing

Epidermal growth factor (EGF) is known to promote corneal epithelial wound healing in experimental scrape and keratectomy models. We studied its efficacy in treating alkali-burned epithelial ulceration. EGF induced hyperplasia and cell proliferation to resurface the denuded and denatured corneal stroma, but it did not prevent recurrent erosions. A combination of EGF with fibronectin (Fn) was noted to enhance epithelial defect closure after alkali burns of the cornea and was seen to prevent recurrent erosions. Histopathologic examination of these corneas revealed marked leukocytic infiltration of the alkali-burned corneal stroma. To find ways to retard this inflammatory response, we studied the role of topical steroids and their efficacy when used with EGF, Fn, and laminin (Ln) in the management of alkali-burned corneas. Use of steroids decreased the incidence of recurrent erosions and corneal perforations. Histologically, steroids markedly decreased the leukocytic infiltration of stromal tissue, thereby retarding the collagenolysis. Topical steroids used with EGF and fibronectin were seen to promote epithelial wound closure and to prevent recurrent erosions in alkali-burned corneas. Combinations of EGF, fibronectin, and steroids may have a place in the treatment of clinical corneal alkali burns.

Topics: Adrenal Cortex Hormones; Animals; Cell Division; Corneal Injuries; Corneal Ulcer; Epidermal Growth Factor; Epithelium; Eye Burns; Fibronectins; Ophthalmic Solutions; Rabbits; Recurrence; Sodium Hydroxide; Wound Healing

Regeneration of corneal epithelium following injury is essential for visual rehabilitation. A limited number of approaches are available for treating patients who fail to heal epithelial injuries adequately. The presence of specific receptors for epidermal growth factor (EGF) on epithelial cells suggests that this potent mitogen may play a role in normal epithelial wound healing. Topical application of biosynthetic human EGF significantly accelerated epithelial regeneration in primates following epikeratophakia surgery. Epidermal growth factor alone and with fibronectin accelerated epithelial regeneration of rabbits following mild alkali burns. Since prolonged exposure of cells to EGF is necessary to induce mitosis, the dynamics of EGF in the eye and with various lenses was studied. When applied in methylcellulose-based eye drops, 90% of the EGF was lost from tear film within 10 min, while a small amount (10%) remained associated with conjuctival tissue. Soft contact lenses or epikeratophakia lenticles took up substantial amounts of EGF (50 micrograms) and released 85% of the EGF within 24 h, with a half-life of 4 h in vitro. Epidermal growth factor did not diffuse through corneas or lenticles and did not promote epithelial downgrowth along sutures in primate corneas. These results suggest that biosynthetic growth factors may be useful in the treatment of some epithelial injuries.

Topics: Alkalies; Animals; Burns, Chemical; Contact Lenses, Hydrophilic; Cornea; Corneal Injuries; Corneal Transplantation; Epidermal Growth Factor; Epithelium; Eye Burns; Iodine Radioisotopes; Regeneration

Ten rabbits with the right eye burnt by 1 N NaOH were treated 5 times daily with epidermal growth factor (EGF) eyedrops or placebo. The epithelium seemed to heal better under EGF treatment than with placebo. Also, ulceration and secondary calcification of the cornea were reduced in the EGF-treated group. The vascularisation was slightly diminished by EGF. The lactate dehydrogenase and N-acetylglucose aminidase activities and the lactate and glucose levels in the corneae were not different between the EGF and the placebo groups. The results showed that there was a beneficial effect of EGF in these experiments with most severe eye burn disease. But the improved regeneration of the epithelium seemed to be counteracted and partly abolished by the severe inflammatory reaction in these eyes.

Topics: Animals; Blood Vessels; Burns, Chemical; Cornea; Corneal Ulcer; Epidermal Growth Factor; Epithelium; Eye Burns; Microscopy, Electron, Scanning; Rabbits; Regeneration; Sodium Hydroxide

We conducted a double-masked study to evaluate the effect of epidermal growth factor on epithelial wound healing and recurrent erosions in alkali-burned rabbit corneas. Epithelial wounds 10 mm in diameter healed completely under the influence of topical epidermal growth factor, whereas the control corneas did not resurface in the center. On reversal of treatment, the previously nonhealing epithelial defects healed when treated with topical epidermal growth factor eyedrops. Conversely, the epidermal growth factor-treated and resurfaced corneas developed epithelial defects when treatment was discontinued. Histopathologic examination disclosed hyperplastic epithelium growing over the damaged stroma laden with polymorphonuclear leukocytes when treated with epidermal growth factor eyedrops, but it did not adhere to the underlying tissue. Hydropic changes were seen intracellularly as well as between the epithelial cells and the stroma.

Topics: Alkalies; Animals; Burns, Chemical; Corneal Injuries; Double-Blind Method; Epidermal Growth Factor; Eye Burns; Kinetics; Rabbits; Time Factors; Wound Healing