The role of Staphylococcal enterotoxin in atopic keratoconjunctivitis and corneal ulceration.

BACKGROUND: Patients with atopic eczema frequently experience colonization with Staphylococcus aureus that is directly correlated with the eczema severity. We hypothesized that S. aureus-secreted enterotoxins (SE) are involved in the pathophysiology of atopic keratoconjunctivitis (AKC). METHODS: A total of 45 subjects (18 with AKC, nine vernal keratoconjunctivitis (VKC), eight seasonal allergic conjunctivitis (SAC), and ten healthy volunteers) were enrolled. Slit lamp examinations, including fluorescein staining, were performed. Scraped samples were collected from the upper tarsal conjunctiva, lower conjunctival sacs, and the skin around the eyelid margins. Superantigen (SAg) genes were detected using polymerase chain reaction (PCR). RESULTS: Among 45 cases, S. aureus was detected significantly more in AKC patients than VKC patients (P = 0.026), SAC patients (P = 0.0003), and healthy volunteers (P = 0.0001). SAg genes were detected in 11 patients. SEB (2/11), SEG (8/11), and SEI (8/11) were detected, but no other SE. There was a significant difference in SE detection between AKC and SAC patients (P = 0.03). In severe types of ocular allergic disease such as AKC and VKC (N = 27), SE was detected in six of ten patients with corneal ulcers and two of 17 patients without corneal ulcers. SE was detected in significantly more patients with corneal ulcers (P = 0.025). CONCLUSIONS: In patients with AKC, S. aureus and SE were detected more frequently compared with other patients and healthy volunteers, especially in association with corneal ulceration suggesting a role of SE. So far, it is unknown whether SE leads to tissue damage of the cornea by initiating an immune response or has direct toxic effects.

Alterations of the ocular surface epithelial MUC16 and goblet cell MUC5AC in patients with atopic keratoconjunctivitis.

BACKGROUND: An increased understanding of the ocular surface at cellular level in the conjunctiva and the cornea may help explain the pathogenesis and the subsequent clinical appearance of atopic ocular allergies, which may be potentially blinding. PURPOSE: To investigate the MUC16 and MUC5AC alterations, tear function and the ocular surface disorder in patients with atopic keratoconjunctivitis (AKC). METHODS: Thirty-six eyes of 18 AKC patients as well as 28 eyes of 14 age- and sex-matched normal subjects were studied. The subjects underwent corneal sensitivity measurements, Schirmer test, tear film break-up time (BUT), fluorescein and Rose-Bengal staining of the ocular surface, conjunctival impression cytology and brush cytology. Impression cytology samples underwent periodic acid schiff and immunohistochemical staining with MUC16 and MUC5AC antibodies. Brush cytology specimens underwent evaluation for inflammatory cell numbers and quantitative real-time polymerase chain reaction (PCR) for MUC16 and MUC5AC mRNA expression. RESULTS: The mean corneal sensitivity and BUT values were significantly lower in patients with AKC, compared with controls (P < 0.001). Brush cytology specimens from AKC patients revealed significantly higher numbers of inflammatory cells (P < 0.001). Specimens from patient eyes showed positive staining for MUC5AC and MUC16. MUC16 mRNA expression was significantly upregulated with significant downregulation of MUC5AC mRNA expression in eyes with AKC compared with the eyes of control subjects. CONCLUSION: Ocular surface inflammation, decline in corneal sensitivity, tear film instability, changes in conjunctival epithelial MUC5AC and MUC16 mRNA expressions were thought to be important in the pathogenesis of atopic ocular surface disease.

Ocular surface and tear functions after topical cyclosporine treatment in dry eye patients with chronic graft-versus-host disease.

We investigated the effect of 0.05% topical cyclosporine (Cys) on the ocular surface and tear functions in dry eye patients with chronic GVHD (cGVHD) in a prospective comparative study. Thirty eyes of 15 patients refractory to baseline treatment were recruited and the patients assigned for topical Cys treatment group (14 eyes of 7 patients) and control group (12 eyes of 6 patients) respectively. Two patients dropped out because of intolerable irritation while using topical Cys eye drops. Visual analog scale symptom scores, corneal sensitivity, Schirmer I test value, tear film break-up time (TBUT), tear evaporation rate and ocular surface vital staining scores were recorded at baseline and at the end of the following one month. Conjunctival impression and brush cytology were performed before and after the treatment. After topical Cys treatment, significant improvements were found in symptom scores, corneal sensitivity, tear evaporation rate, TBUT, vital staining scores, goblet cells density, conjunctival squamous metaplasia grade, inflammatory cell numbers and the MUC5AC expression. Our study suggests that 0.05% topical Cys may be an effective treatment for dry eye patients with cGVHD. The improvements in the ocular surface and tear functions resulted presumably from the decreased inflammation, increased goblet cell density and MUC5AC mRNA expression. Bone Marrow Transplantation (2008) 41, 293-302; doi:10.1038/sj.bmt.1705900; published online 5 November 2007.

The differences of tear function and ocular surface findings in patients with atopic keratoconjunctivitis and vernal keratoconjunctivitis.

BACKGROUND: The pathogenesis of the ocular surface disease in atopic keratoconjunctivitis (AKC) and vernal keratoconjunctivitis (VKC) has not been fully understood. We tried to clarify the differences in the ocular surface status in patients with AKC, VKC, and healthy control subjects. METHODS: Twenty-four eyes of 12 AKC patients, 12 eyes of six VKC patients, and 20 eyes of 10 normal control subjects were studied. The subjects underwent corneal sensitivity measurements, Schirmer test, tear film break-up time (BUT), vital staining of the ocular surface, conjunctival impression and brush cytology. Impression cytology samples underwent periodic acid Schiff staining for goblet cell density, squamous metaplasia grading, and immunohistochemical staining for MUC1, 2, 4, and 5AC. Brush cytology specimens underwent staining for inflammatory cell counting and Real Time PCR for MUC1, 2, 4, and 5AC mRNA expression. RESULTS: The mean BUT, corneal sensitivity, and conjunctival goblet cell density values in AKC patients were significantly lower compared with VKC patients and control subjects. The squamous metaplasia grades in eyes with AKC were significantly higher compared to eyes with VKC and controls. The inflammatory cell response in brush cytology specimens was different between patients with AKC and VKC. Eyes with AKC showed significantly higher MUC1, 2 and 4 and lower MUC5AC mRNA expression compared to eyes with VKC. CONCLUSIONS: Differences of the infiltrates, higher level of tear instability, lower corneal sensitivity, up-regulation of MUC1, 2, and 4, and down regulation of MUC5AC were important differential features of the ocular surface disease in AKC compared with VKC.

Alterations of the ocular surface epithelial mucins 1, 2, 4 and the tear functions in patients with atopic keratoconjunctivitis.

BACKGROUND: An increased understanding of the ocular surface alterations at the cellular level in the conjunctiva and the cornea, may help explain the pathogenesis and the subsequent clinical appearance of atopic ocular allergies, which may be potentially blinding. PURPOSE: To investigate MUC 1, 2 and 4 alterations, tear function and the ocular surface disorder in patients with atopic keratoconjunctivitis. METHODS: Twenty-eight eyes of 14 atopic keratoconjunctivitis patients as well as 22 eyes of 11 age-and sex-matched normal subjects were studied. The subjects underwent corneal sensitivity measurements, Schirmer's test, tear film break-up time (BUT), fluorescein and Rose Bengal staining of the ocular surface, conjunctival impression cytology and brush cytology. Impression cytology samples underwent periodic acid-Schiff and immunohistochemical staining with MUC 1, 2 and 4 antibodies. Brush cytology specimens underwent evaluation for inflammatory cell numbers and quantitative real-time-PCR for MUC 1, 2 and 4 mRNA expression. Patient eyes with fluorescein and Rose Bengal scores greater than four points were regarded to have significant epithelial disease in this study. RESULTS: The mean corneal sensitivity and BUT values were significantly lower in atopic patients with significant epithelial disease, compared with patients with insignificant epithelial disease and controls (P < 0.01). Brush cytology specimens from patients with significant epithelial disease revealed significantly higher numbers of inflammatory cells (P < 0.01). Specimens from patient eyes showed positive staining for MUC 1, 2 and 4. MUC 1, 2 and 4 mRNA expressions were significantly higher in eyes with significant epithelial disease compared with eyes with insignificant epithelial disease and eyes of control subjects. CONCLUSION: Ocular surface inflammation, decline in corneal sensitivity, tear film instability, changes in conjunctival epithelial MUC 1, 2 and 4 mRNA expressions were thought to be important in the pathogenesis of atopic ocular surface disease.

Inflammatory cells in brush cytology samples correlate with the severity of corneal lesions in atopic keratoconjunctivitis.

BACKGROUND: Inflammatory cells infiltrating to the tarsal conjunctiva are thought to be involved in the pathogenesis of corneal lesions in severe allergic conjunctival diseases. The relation between such cells and the severity of corneal lesions was studied. METHODS: Six patients with atopic keratoconjunctivitis (AKC) were enrolled in this study. Tarsal brush cytology findings and the severity of corneal damage at that point were recorded and analysed for correlation. RESULTS: Four out of six patients exhibited correlation between eosinophils and corneal damage. Three out of six patients exhibited correlation between neutrophils and corneal damage. Two out of six patients exhibited correlation between both eosinophils and neutrophils and corneal damage. Analysis of all data from all patients taken together revealed that both eosinophils and neutrophils in brush cytology samples significantly correlated with corneal damage. CONCLUSIONS: Inflammatory cells in brush cytology samples correlated with corneal damage. Evaluation of the relative percentages of inflammatory cells in brush cytology samples is a useful method of assessing disease activity in allergic conjunctival disease.

Improvement of corneal fluorescein staining in post cataract surgery of diabetic patients by an oral aldose reductase inhibitor, ONO-2235.

AIM: While the mechanism in the pathogenesis of diabetic corneal disease is unclear, aldose reductase has been implicated in corneal disease. The effects of an oral aldose reductase inhibitor (ARI) on the ocular surface of diabetic patients after cataract surgery were studied. METHODS: This clinical trial was designed to be randomised, double blinded, and placebo controlled. Pseudophakic patients with diabetes were randomly assigned to treatment with either oral ARI (ONO-2235) (n=12) or placebo (n=9) for 12 weeks. The vital staining of the ocular surface, tear production and clearance, break up time in tears (BUT), corneal and conjunctival sensation, and symptom score before treatments were examined as well as 4, 8, 12 weeks after the administration. Specular microscopic evaluation was also performed. RESULTS: After a 12 week period of oral ARI administration, fluorescein staining scores (from 2.04 (SD 1.12) to 1.46 (1.18); p=0.016), conjunctival sensation (from 1.15 (0.37) to 1.36 (0.31); p=0.0006), and symptom scores (from 5.38 (1.932) to 4.00 (2.07); p=0.0002) recovered significantly. Fluorescein staining of oral ARI administration also decreased compared with placebo (p=0.017). Rose bengal staining, tear clearance, and corneal sensation were improved although this increase was minor. Tear production, BUT, and specular microscopic evaluation of the corneal epithelium and endothelium did not demonstrate a significant change. CONCLUSION: Oral ARI opposes the ocular surface changes caused by diabetes, by recovery of ocular surface sensitivity as demonstrated through an improvement in vital staining.

kdsA mutations affect FtsZ-ring formation in Escherichia coli K-12.

No one has, as yet, addressed the relationship between the nature of the outer membrane and cell division. kdsA encodes 3-deoxy-D-manno-octulosonic acid (KDO) 8-phosphate synthetase which catalyses the first step in the synthesis of KDO, the linker between lipid A and oligosaccharide of lipopolysaccharide (LPS). Seven temperature-sensitive mutants containing missense mutations in kdsA were affected in the production of KDO and all mutants stopped dividing at 41 degrees C and formed filaments with either one or no FtsZ ring. All observed defects were reversed by the plasmid-borne wild-type kdsA gene. Western blotting analysis, however, demonstrated that the amount of FtsZ protein was not affected by the mutation. The mutants were more susceptible to various hydrophobic materials, such as novobiocin, eosin Y and SDS at 36 degrees C. Methylene blue, however, restored kdsA mutant growth. Plasmid-borne wild-type msbA, encoding a lipid A transporter in the ABC family, partially suppressed kdsA mutation. A mutation of lpxA, functioning at the first stage in lipid A biosynthesis, inhibited both cell division and growth, producing short filaments. These results indicate that the instability of the outer membrane, caused by the defect in KDO biosynthesis, affects FtsZ-ring formation.

Interleukin-8 concentrations in conjunctival epithelium brush cytology samples correlate with neutrophil, eosinophil infiltration, and corneal damage.

PURPOSE: To evaluate whether interleukin-8 (IL-8) and RANTES (regulated on activation, normal T-cell expressed and secreted) concentrations in the supernatants of conjunctival epithelial samples from patients with vernal keratoconjunctivitis (VKC) correlate with the number of infiltrating eosinophils or neutrophils and with the severity of corneal lesions. METHODS: Thirty-four patients with VKC, 5 patients with seasonal allergic conjunctivitis, and 10 volunteers without allergic diseases were enrolled in this study. Conjunctival epithelial cells were collected by brush cytology and the number of inflammatory cells was counted. The chemokine expression in the cells was investigated by immunocytochemistry and the chemokine concentrations of the cell suspensions were measured by enzyme-linked immunosorbent assay. RESULTS: The percentages of eosinophils and neutrophils in cell suspensions from VKC patients with corneal erosion or ulcer were higher than those from subjects with clear corneas or superficial punctate keratopathy. IL-8 concentrations in the supernatant of samples correlated significantly with the percentages of neutrophils and eosinophils in paired cell suspensions. No correlation was observed between RANTES and the percentages of eosinophils. Positive staining for IL-8 was observed in the cytosol of conjunctival epithelial cells. CONCLUSION: IL-8 in the extracellular space of the conjunctival epithelium may play a role in the recruitment of neutrophils and possibly eosinophils and in the pathogenesis of corneal damage in severe allergic diseases.

Antimalarial and cytotoxic activities of bicycl.

Biological evaluations of bicyclo[6.4.0]dodecenone derivatives on antimalarial activity in vitro against Plasmodium falciparum and cytotoxicity against human KB cells were made. (+/-)-(1R*,4S*,7R*,8S*)-4-tert-Butyl-dimethylsiloxy-5,5-dimethyl-1-methyl-9-methylene-7-phenylsulfonylbicyclo[6.4.0]dodec-2,11-dien-10-one (15) exhibited potent antimalarial activity, whereas (+/-)-(1R*,7R*,8S*)-1-methyl-9-methylene-7-phenylsulfonylbicyclo[6.4.0]dodec-2,11-dien-10-one (14) showed significant cytotoxic activity in human KB cells. Both 14 and 15 possess, as a structural character, the exo-methylene moiety in their 6-membered ring of the 8-6 fused ring system.

A novel approach to the taxane BC ring system through formation of alpha-ketol by oxidative removal of the phenylsulfonyl group with subsequent in situ oxidation.

Cis-fused bicyclo[6.4.0]dodecene 11 was converted into taxane BC ring system 21 in three steps; transformation of the phenylsulfonyl group to an alpha-hydroxy carbonyl group by the treatment with potassium hexamethyldisilazide (KHMDS) and triethyl phosphite under oxygen atmosphere, followed by reductive elimination of the hydroxyl group of alpha-ketol moiety, and inversion of ring juncture. Epimerization of the sulfonyl group of 11 was indispensable for the first oxidation process (17 --> 18) and the second oxidation of 12 leading to hydroxylation at the alpha-position of the carbonyl group proceeded with high regio- and stereoselectivity to give 13. On the other hand, reaction of the cross-conjugated compound 5 with KHMDS at 0 degrees C brought about a complete reorganization of molecular framework to provide the compound 7 in which the five-membered ring and the conjugated seven-membered ring were connected through a single bond.

Quantitative evaluation of atopic blepharitis by scoring of eyelid conditions and measuring the water content of the skin and evaporation from the eyelid surface.

PURPOSE: To quantitatively evaluate the condition of the eyelid skin of patients with atopic blepharitis, their symptoms were scored and the water content of the skin and evaporation from the skin were measured. METHODS: Forty patients with atopic blepharitis were examined. The condition of eyelid skin (erythema, edema/papulation/oozing/crust, excoriation/lichenification) was scored from 0 to 3 points. Water content and water evaporation were measured with a Moisture Checker and an evaporimeter, respectively. Eleven age-matched volunteers without atopic disorders were recruited as normal controls. RESULTS: The Moisture Checker values and water evaporation from lid skin were significantly correlated (r = -0.44, p = 0.006). The Moisture Checker values of the patients with atopic blepharitis was 35.5+/-8.2% (44.7 +/-10.6% in the normal controls, p = 0.009), and water evaporation from their lid skin was 3.6+/-0.9 g/cm2 per second (2.0+/-0.3 g/cm2 per second, p < 0.001); then, the patients were divided into four groups, from "asymptomatic" to "severe," according to the sum of their blepharitis scores. Patients with lower blepharitis scores tended to have higher Moisture Checker values and lower water evaporation values. CONCLUSION: Scoring of eyelid condition enabled us to objectively estimate the severity of atopic blepharitis. Measurements of the water content of lid skin and water evaporation from lid skin are useful in evaluation of the severity of this disease.

Immunologic rejection of the central graft after limbal allograft transplantation combined with penetrating keratoplasty.

PURPOSE: To study the incidence and prognosis of immunologic rejection of the central graft after limbal allograft transplantation (keratolimbal allograft transplantation [KLAT]) combined with penetrating keratoplasty (PKP). METHODS: Endothelial rejection in central penetrating graft after simultaneous KLAT and PKP using the same donor cornea was retrospectively studied. Incidence, reversibility, prognosis, and changes in limbal grafts were examined. RESULTS: Forty-five eyes underwent simultaneous PKP and KLAT. Endothelial rejection of the central graft was found in 16 eyes (35.6%). At last examination, 10 grafts (62.5%) restored clarity after immunosuppressive therapy. During rejection episodes, four eyes showed engorgement of vessels in limbal grafts, which existed before the episodes. Only one eye developed neovascularization with mild edema of the limbal grafts; however, no other limbal grafts showed abnormalities on biomicroscopy. No epithelial changes were noted, and 14 grafts (87.5%) were covered by corneal epithelium after the rejection. CONCLUSION: Approximately one third of eyes had endothelial rejection in the central graft after simultaneous KLAT and PKP. Abnormalities suggestive of rejection in the limbal grafts were seldom observed in these eyes, suggesting that immunologic response was different in central and limbal grafts.

Combined medical and surgical treatment of severe vernal keratoconjunctivitis.

PURPOSE: We report the efficacy of an alternative method of treatment for vernal keratoconjunctivitis (VKC) that consists of excision of the palpebral conjunctiva followed by supratarsal injection of corticosteroid and five times daily topical application of 0.05% cyclosporine A (CsA) and cromolyn sodium. METHODS: We evaluated 10 patients with severe treat-resistant VKC with corneal complications. The patients were evaluated for symptoms and for signs, including conjunctival changes, corneal limbal infiltrates, vascularization, reduction of epitheliopathy, meibomitis, visual acuity, intraocular pressure, and pathologic evaluation, before and after treatment. RESULTS: All patients showed marked improvement after 2 weeks of treatment. The symptoms (P <.01), signs (P <.02), and the visual acuity of all patients (P <.01) had significantly improved following treatment. Histological examination showed significant inflammatory cell decreases 4 weeks after surgery (P <. 05). CONCLUSION: Surgery plus topical drug therapy may be useful in treating patients with very severe VKC.

Association of preoperative tear function with surgical outcome in severe Stevens-Johnson syndrome.

OBJECTIVE: To retrospectively study the surgical outcome in severe Stevens-Johnson syndrome (SJS). DESIGN: Retrospective noncomparative case series. PARTICIPANTS: Twenty-nine eyes of patients with severe SJS associated with total conjunctivalization were examined. INTERVENTION: Preserved human amniotic membrane grafting and keratolimbal allograft transplantation was performed. Intensive immunosuppression and epithelial management were continued postoperatively. MAIN OUTCOME MEASURES: Successful ocular surface reconstruction was determined by epithelialization with corneal epithelium. The association between surgical outcome and preoperative tear function was studied. RESULTS: The ocular surface was successfully covered by corneal epithelium in 13 eyes (44.8%). The mean corrected visual acuity recovered from 0.0039 to 0.017. Persistent epithelial defect was the most common complication. Eyes with successful ocular surface reconstruction had significantly better preoperative Schirmer's test values (P = 0.025) and tear clearance rates (P = 0.043) than those that failed. CONCLUSIONS: In patients with severe SJS, preoperative tear function significantly influenced surgical outcome. Eyes with Schirmer's test value of >10 mm have a greater chance of successful ocular surface reconstruction.

Trabeculectomy with mitomycin C for post-keratoplasty glaucoma.

AIM: To investigate the effect of trabeculectomy with and without mitomycin C in post-keratoplasty glaucoma. METHODS: A retrospective study was performed on patients who underwent trabeculectomy for glaucoma after penetrating keratoplasty. 34 eyes of 32 patients were included in this study. 26 eyes received trabeculectomy with mitomycin C and eight eyes without mitomycin C. The procedure was deemed successful if the intraocular pressure was maintained below 21 mm Hg with or without use of additional antiglaucoma medication (mean follow up time 22.3 (SD 10.3) months). RESULTS: At the last examination trabeculectomy was successful in 19 of 26 eyes (73.0%) with mitomycin C (+) and two of eight (25.0%) without (p=0.0219). When the prognosis was analysed by Kaplan-Meier curve, the mitomycin C (+) group showed a better prognosis (p=0.0182). Mean intraocular pressure and average number of glaucoma medications improved in the group with mitomycin C without severe side effects on the graft. Graft rejection after trabeculectomy was seen in two eyes in the mitomycin C group. Final graft clarity rate was 69.2% (18/26) in the mitomycin C (+) group and 37.5% (3/8) in the mitomycin C (-) group. Complications such as persistent epithelial defect, cystoid macular oedema, choroidal detachment, leakage from bleb were seen in four eyes in the mitomycin C (+) group and in one eye in the mitomycin C (-) group. CONCLUSIONS: Trabeculectomy with mitomycin C showed better results for glaucoma following keratoplasty.

Pregnanes and pregnane glycosides from Hoya carnosa.

Eleven pregnanes were isolated from the hydrolysate of the CHCl3 extract fractionated from the caules of Hoya carnosa. Among these, six pregnanes, including 19-acetoxydigipurpurogenin II, were new, and their structures were elucidated. The structures of twenty new pregnane tetraosides and pentaosides, named hoyacarnosides A-T, besides three known ones from the CHCl3 extract, were determined.

Cytosolic phospholipase A2 is essential for both the immediate and the delayed phases of eicosanoid generation in mouse bone marrow-derived mast cells.

We have used mice in which the gene for cytosolic phospholipase A2 (cPLA2) has been disrupted to demonstrate the absolute requirement for cPLA2 in both the immediate and the delayed phases of eicosanoid generation by bone marrow-derived mast cells. For the immediate phase, quantitative analysis of the products of the 5-lipoxygenase pathway showed that gene disruption of cPLA2 prevented the provision of arachidonic acid substrate for biosynthesis of proximal intermediates. By analogy, we conclude that arachidonic acid substrate was also not available to prostaglandin endoperoxide synthase 1 in the immediate phase of prostaglandin (PG) D2 generation. These defects occurred with two distinct stimuli, stem cell factor and IgE/antigen, which were, however, sufficient for signal transduction defined by exocytosis of beta-hexosaminidase. Whereas cPLA2 is essential for immediate eicosanoid generation by providing arachidonic acid, its role in delayed-phase PGD2 generation is more complex and involves the activation-dependent induction of prostaglandin endoperoxide synthase 2 and the supply of arachidonic acid for metabolism to PGD2.

v-src induces cisplatin resistance by increasing the repair of cisplatin-DNA interstrand cross-links in human gallbladder adenocarcinoma cells.

Activation of Src, which has an intrinsic protein tyrosine kinase (PTK) activity, has been demonstrated in human solid tumors, such as colorectal and breast cancers. To investigate the role of activated Src in drug resistance, we evaluated the effect of v-src on the resistance to various anti-cancer drugs using v-src-transfected HAG-1 human gallbladder adenocarcinoma cells. Compared with parental or mock-transfected HAG-1 cells, v-src-transfected HAG/src3-1 cells showed a 3.5-fold resistance to cis-diamminedichloroplatinum (II) (CDDP) but not to doxorubicin, etoposide or 5-fluorouracil. By contrast, activated H-ras, which acts downstream of src, failed to induce resistance to either of these drugs. Furthermore, wortmannin, a phosphatidylinositol (PI) 3-kinase inhibitor, and H7, a protein kinase C (PKC) inhibitor, did not alter CDDP resistance. Evaluation of the kinetics of the removal of DNA interstrand cross-links (ICLs), measured by alkaline elution, showed a significant increase in this removal in HAG/src3-1 cells as compared with mock-transfected cells, though no differences were found in the formation of DNA ICLs between these cell lines. CDDP resistance in v-src-transfected cells was reversed, if not completely, by either herbimycin A or radicicol, specific inhibitors of Src-family PTKs, suggesting that Src tyrosine kinase activity induces CDDP resistance. Moreover, significant reduction in the repair of CDDP-induced DNA ICLs was observed upon treatment with radicicol. The intracellular glutathione content and mRNA expression of topoisomerase II and metallothionein were virtually identical between these cell lines, except for topoisomerase I mRNA. Our data strongly suggest that the ability of activated src, but not ras, to induce CDDP resistance is mediated by augmentation of DNA repair through Src to downstream signal-transduction pathways distinct from either the Ras, PI 3-kinase or PKC pathway.