Danish heritable retinoblastoma survivors' perspectives on reproductive choices: "It's important for me, not to pass on this condition".

Despite reporting an overall normal life, survivors of heritable retinoblastoma face numerous physical and psychosocial issues. In particular, reproductive decision-making is often complex and difficult. This study aims to examine survivors' reflections on passing on heritable retinoblastoma to their children, how survivors approach their reproductive choices, and how the healthcare system can optimize counseling and support. Semi-structured interviews with Danish adult survivors of heritable retinoblastoma were qualitatively analyzed to explore their experiences. Participants were recruited from the Retinoblastoma Survivorship Clinic, Aarhus University Hospital, Denmark. Thematic data analysis was conducted followed by a condensing process specifically for the subthemes relating to reproductive choices. A common subtheme for all participants was a strong wish to avoid passing on retinoblastoma to their children. The participants emphasized the various medical, practical, emotional, and moral issues impacting their final reproductive choice in the process of family planning to conceive a child unaffected by retinoblastoma. Some had no option other than to conceive naturally and hope for an unaffected baby; while others weighed the pros and cons of choosing natural conception with prenatal testing and then considering termination of pregnancy (in case of an affected fetus) versus choosing fertility treatment with preimplantation genetic testing to achieve an unaffected pregnancy. Several participants underlined the complexity of their decisions, and also expressed feelings of guilt, both toward their affected child, and guilt for putting their partner through many difficult decisions and obstacles due to their genetic condition. Our findings demonstrate how one family-planning decision is not unequivocally "better" or easier than another. Healthcare professionals must provide the necessary information and tools to support the individual's unique decision-making process. Survivors' autonomy and individual needs, as well as the numerous and diverse aspects of heritable retinoblastoma, should be carefully considered.

[Bisphosphonate-induced ocular and orbital inflammation].

The inflammatory side effects of bisphosphonates are well-known. We report a case of orbital and ocular inflammation secondary to the use of intravenous zoledronic acid and a case of scleritis secondary to oral alendronate. Bisphosphonate-induced inflammation can present as uveitis, (epi)scleritis, and orbital inflammation. The course is typically self-limiting after cessation of bisphosphonate therapy, but resolution can be further promoted by steroid therapy. The ocular side effects of bisphosphonates should be duly considered.

Genetic testing in adult survivors of retinoblastoma in Denmark: A study of the experience and impact of genetic testing many years after initial diagnosis.

BACKGROUND: Survivors with heritable retinoblastoma (RB) face a high risk for second primary cancer and RB in their children. Knowledge of heredity can support second cancer surveillance, convey reproductive options or early diagnosis of RB in their offspring. Currently, all newly diagnosed Danish patients with RB are offered genetic testing, as opposed to a minority of survivors diagnosed before available DNA testing. OBJECTIVE: To examine RB survivors' response to unsolicited contact, uptake of genetic testing, and RB1 variant detection rate, and to qualitatively evaluate the experience and overall impact of genetic testing for heritable RB. METHODS: Genetically untested adult RB survivors were invited to receive genetic counseling, undergo genetic testing for heritable RB and complete an eye examination. The number of responses, uptake of genetic testing and genetic results are descriptively reported. Additionally, responding survivors participated in a qualitative interview study of the perceived impact of genetic testing. Interviews were audio-recorded, transcribed verbatim and thematically analyzed. RESULTS: Among invited RB survivors, 58% responded. Of these, 88% opted for genetic counseling and genetic testing. A diagnosis of heritable RB was established in 23% of RB survivors. Interestingly, all of these survivors were unilaterally affected. Analysis of data from the interviews revealed three recurring themes regarding the impact of genetic counseling and testing several years after initial diagnosis: 'Risk of what?', 'Knowledge is important' and 'Impact of the result'. The possible risk ofsecond cancer and RB in their children was new knowledge for several participants; however, in general, the participants appreciated receiving genetic information and certainty about heredity. Accordingly, the impact of genetic counseling and testing was perceived in a positive way. CONCLUSION: Overall, RB survivors valued the opportunity to receive genetic counseling and undergo genetic testing many years after diagnosis. Responding RB survivors appreciated the invitation to test, felt well-informed and described little decisional conflict regarding their decision-making, valuing the genetic information and certainty. Heritable RB was confirmed in 23% of the previously untested RB survivors. These individuals emphasized the value of knowing and being proactive regarding both reproduction and cancer risk.

Metachronous, non-pineal, trilateral retinoblastoma in a patient with a seemingly reduced-expressivity RB1 germline deletion.

The clinical course of trilateral retinoblastoma can be unpredictable, and expressivity of germline RB1 variants may vary during development. We describe an unexpected fatal case of trilateral retinoblastoma with an intracranial tumor in an unusual location and discuss genetic copy number analyses as a useful diagnostic tool with therapeutic potential.

Genomic Alterations in Human Papillomavirus-Positive and -Negative Conjunctival Squamous Cell Carcinomas.

Purpose: The genomic alterations contributing to the pathogenesis of conjunctival squamous cell carcinomas (SCCs) and their precursor lesions are poorly understood and hamper our ability to develop molecular therapies to reduce the recurrence rates and treatment-related morbidities of this disease. We aimed to characterize the somatic DNA alterations in human papillomavirus (HPV)-positive and HPV-negative conjunctival SCC. Methods: Patients diagnosed with conjunctival SCC in situ or SCC treated in ocular oncology referral centers in Denmark were included. HPV detection (HPV DNA PCR, p16 immunohistochemistry, and mRNA in situ hybridization) and targeted capture-based next-generation sequencing of 523 genes frequently involved in cancer were performed to describe the mutational profile based on HPV status. Results: Tumor tissue was available in 33 cases (n = 8 conjunctival SCCs in situ, n = 25 conjunctival SCCs), constituting 25 male and 8 female patients. Nine cases were HPV positive. The HPV-positive SCCs in situ and SCCs were characterized by transcriptionally active high-risk HPV (types 16 and 39) within the tumor cells, frequent mutations in PIK3CA (n = 5/9), and wild-type TP53, CDKN2A, and RB1, while the HPV-negative counterparts harbored frequent mutations in TP53 (n = 21/24), CDKN2A (n = 7/24), and RB1 (n = 6/24). Conclusions: Our findings have delineated two potentially distinct distributions of somatic mutations in conjunctival SCC based on HPV status-pointing to different biological mechanisms of carcinogenesis. The present findings support a causal role of HPV in a subset of conjunctival SCC.

Living with heritable retinoblastoma and the perceived role of regular follow-up at a retinoblastoma survivorship clinic: 'That is exactly what I have been missing'.

OBJECTIVE: To explore living with heritable retinoblastoma, specifically survivors' perceived role of regular follow-up at a retinoblastoma survivorship clinic. METHODS AND ANALYSIS: Adult survivors of heritable retinoblastoma were recruited from the Retinoblastoma Survivorship Clinic, Aarhus University Hospital. Ten survivors participated in individual explorative, semistructured interviews. Thematic data analysis was conducted. RESULTS: Five key themes relating to vision, social life, family, second cancer risk and the healthcare system were identified. Subthemes relating to the Retinoblastoma Survivorship Clinic included the retinoblastoma coordinator, cancer risk, psychosocial support and genetic knowledge. The retinoblastoma-related physical and psychosocial issues influenced survivors' everyday living; however, the opportunity to live a normal life varied considerably, with the majority experiencing no major limitations. The need for specialised management and a coordinator was emphasised to be the main value of the Retinoblastoma Survivorship Clinic. CONCLUSION: Despite reporting an overall normal life and no major limitations in daily living activities, our data confirm that heritable retinoblastoma impacts several aspects of daily living. Uniquely, this study demonstrates that the main value of the Retinoblastoma Survivorship Clinic was a specialised contact person and coordinator in the healthcare system, providing continuous and necessary management and guidance after retinoblastoma treatment, and for all aspects of health related to heritable retinoblastoma. The needs of heritable retinoblastoma survivors are complex and extensive, and the specific role of the healthcare system to support survivorship should be prioritised, specialised and multidisciplinary.

Conjunctival intraepithelial neoplasia and carcinoma: distinct clinical and histological features in relation to human papilloma virus status.

AIMS: Human papillomavirus (HPV) is considered a causative agent for the development of a broad range of human carcinomas. The role of HPV in the development of conjunctival intraepithelial neoplasia (CIN) and carcinoma (cSCC) remains unclear. The purpose of the present study was to investigate the HPV prevalence in a nationwide cohort and to describe clinical and histopathological features in relation to HPV status. METHODS: All cases of CIN and cSCC in Denmark from 1980 to 2016 were included. We combined p16 immunohistochemistry (IHC), RNA in situ hybridisation (RNA ISH) and HPV DNA PCR to detect HPV. The results were correlated to clinical and histopathological parameters. RESULTS: One hundred twelve primary tumours and 33 recurrent tumours were included for HPV analysis. Twenty-four (21%) of the primary tumours were HPV positive by PCR. Eighteen of out 19 HPV-positive tumours were positive by RNA ISH. HPV16 was the most prevalent genotype (n=18, 75%). The patients with HPV-positive tumours were significantly younger (mean difference 11.5 years, 95% CI 5.2 to 17.9, p=0.0005) and had a higher recurrence compared with patients with HPV-negative tumours (HR 2.30, 95% CI 1.02 to 5.21, p=0.046). The HPV-positive tumours were associated with a positive p16 IHC and a non-keratinising morphology. CONCLUSION: We describe distinct clinical and histopathological features associated with HPV status in cSCC. The finding of transcriptionally active HPV in this material lends support to a causal role of HPV in a subset of cSCC.

Human papillomavirus infection plays a role in conjunctival squamous cell carcinoma: a systematic review and meta-analysis of observational studies.

PURPOSE: We aimed to study the prevalence of human papillomavirus (HPV) in conjunctival intraepithelial neoplasia and carcinoma. Furthermore, we aimed to explore whether geographical differences or different detection modalities are associated with the conflicting information regarding HPV and the development of the disease. METHODS: We searched the MEDLINE, EMBASE and Scopus databases for studies reporting on HPV and conjunctival intraepithelial neoplasia or carcinoma. The pooled prevalence proportions, odds ratio (OR) and corresponding 95% confidence intervals were calculated assuming a random-effects model. Subgroup analyses and meta-regression explored possible sources of heterogeneity. RESULTS: A total of 39 studies were included in the systematic review. The pooled prevalence of HPV in conjunctival intraepithelial neoplasia and carcinoma was 26%, with HPV16, 18, and 33 being the most frequently reported genotypes. Human papillomavirus (HPV) infection was associated with an increased risk of conjunctival intraepithelial neoplasia and carcinoma (OR 8.4, 95% confidence interval (CI) 3.7-19.1); lower in studies from African countries (OR 1.7, 95% CI 0.9-3.5) than other countries (OR 16.1, 95% CI 5.8-44.3), p = 0.013. CONCLUSION: Human papillomavirus infection increases the odds of conjunctival intraepithelial neoplasia and carcinoma by 8.4 compared to healthy conjunctival mucosa or other ocular surface diseases. There seem to be geographical differences regarding HPV in conjunctival intraepithelial neoplasia and carcinoma. HPV16 was the most prevalent genotype, followed by HPV18 and HPV33, meaning that most of the HPV-related conjunctival intraepithelial neoplasia and carcinoma may be prevented by the HPV vaccines that are currently available.

Incidence and Mortality of Second Primary Cancers in Danish Patients With Retinoblastoma, 1943-2013.

Importance: In heritable retinoblastoma, there is a significantly increased risk of second primary cancers (SPCs). Improved knowledge about the incidence and influence of heritability and treatment is important during therapy for patients with retinoblastoma. Objective: To assess the incidence of SPC in patients diagnosed with retinoblastoma in Denmark from 1943 to 2013 with a focus on heritability and the association of external radiotherapy with mortality. Design, Setting, and Participants: In this retrospective cohort study, data were extracted from the Danish Ocular Oncology Group Database containing complete data on all patients diagnosed with retinoblastoma , and obtained from the Danish Cancer Registry, which includes information on all patients with cancer from 1943 to December 31, 2013. Data analysis was conducted from December 1, 2017, to October 1, 2019. Data on 323 patients were included. Exposures: Heritability and retinoblastoma treatment. Main Outcomes and Measures: Standardized incidence rate, excess absolute risk, cumulative incidence of SPC, and mortality from SPC. Association of heritability and treatment with outcomes was estimated. Results: Of the 323 patients included in the analysis, 181 were men (56%), 133 had heritable retinoblastoma (41%), and 190 had nonheritable retinoblastoma (59%). The median age at diagnosis of SPC was 32.4 (interquartile range, 15.4-43.9) years in patients with heritable retinoblastoma and 38.6 (interquartile range, 20.5-49.4) years in those with nonheritable retinoblastoma. Twenty-five SPCs were identified in patients with heritable retinoblastoma vs 14 in patients with nonheritable retinoblastoma. Standardized incidence rate (SIR) of SPC in patients with heritable retinoblastoma was 11.39 (95% CI, 7.37-16.81) with an excess absolute risk of 70 cases per 10 000 person-years; the highest SIRs were for sarcoma (181.13; 95% CI, 98.94-303.92) and malignant melanoma (26.78; 95% CI, 9.78-58.30). The SIR for SPC in patients with nonheritable retinoblastoma was 1.52 (95% CI, 0.81-2.60). The cumulative incidence of SPCs at age 60 years was significantly higher in patients with heritable retinoblastoma (51%) compared with those with nonheritable retinoblastoma (13%) (P < .001) (hazard ratio, 5.0; 95% CI, 2.5-10.3). No significant differences were identified in overall risk of SPC in patients with heritable retinoblastoma treated with 3 different modalities: external radiotherapy, plaque (but no external) radiotherapy, and enucleation only, but an increased proportion of sarcomas was noted in the irradiated field. Mortality due to SPC was also higher in survivors of heritable retinoblastoma compared with those with nonheritable retinoblastoma (cumulative mortality, 34% vs 12% at age 60 years; P = .03). Conclusions and Relevance: The findings of this study suggest that the incidence and mortality associated with SPC were significantly higher in patients with heritable retinoblastoma vs patients with nonheritable retinoblastoma. The largest increases in risk were noted for sarcoma and malignant melanoma. External radiotherapy did not appear to increase the risk. These findings are relevant when treating patients with retinoblastoma to manage the risk for SPC.

A multidimensional cohort study of late toxicity after intensity modulated radiotherapy for sinonasal cancer.

PURPOSE: To evaluate the occurrence of late toxicity after curatively intended intensity modulated radiotherapy (IMRT) for sinonasal cancer and assess dose-response associations. METHODS: Patients treated with IMRT in 2008-2016 were included. Cross sectional examinations of toxicity from the optic pathway, the brain, the pituitary gland and the nose were performed along with quality of life - (QoL) and dose-response analyses. RESULTS: Twenty-seven patients were enrolled; median age was 67 years (range 47-83). Five patients (19%) had radiation-related ocular toxicity. The risk of visual acuity impairment increased with increasing dose (grade 2 odds ration (OR) 1.12, p = 0.01; grade 3 OR 1.14, p = 0.02) and dose constraint violations (grade 2, OR = 21, p < 0.01; grade 3, OR = 41, p < 0.01). Six patients (22%) exhibited evidence of radiation-related hypopituitarism, but no dose-response association was detected. Seventeen patients (63%) had impaired olfactory function. The risk of olfactory impairment increased with higher stage (OR = 3.32, p = 0.03). Three patients (11%) had structural abnormalities in irradiated areas of the brain, and impaired cognitive function was present in 17 patients (63%). Cognitive, physical, role functioning as well as fatigue and insomnia were affected the most in QOL analyses. Fifteen patients (56%) had grade 2 radiation-related impairment in at least one organ. Grade 3 toxicity was only present in patients with toxicities in >3 organs and in patients initially treated for T4 tumours. Three patients (11%) had radiation-related impaired function in all examined OARs. CONCLUSION: Late toxicity after radiotherapy was substantial in all examined organs, with dose-response associations between visual acuity impairment and the optic nerve. The results have led to changed praxis for follow-up examinations in Denmark.

Danish retinoblastoma patients 1943-2013 - genetic testing and clinical implications.

BACKGROUND: In heritable retinoblastoma there is a 50% risk of transmitting the RB1 mutation, and offspring carriers have more than 90% risk of developing retinoblastoma. Today, all newly diagnosed retinoblastoma patients in Denmark are screened for mutations in RB1, as opposed to only a minority of patients diagnosed before DNA testing was offered. Knowledge of heredity increases the chance of early diagnosis in offspring, leading to improved prognosis. We present data from the Danish retinoblastoma patients that emphasize the need for genetic counseling and RB1 screening in all untested retinoblastoma survivors. MATERIAL AND METHODS: Data are extracted from The Danish Ocular Oncology Group Database, a national population database containing data on all Danish retinoblastoma patients since 1943. RESULTS: In total 323 retinoblastoma patients have been diagnosed between 1943 and 2013. Since 1963, the rate has been stable around 1 per 14 000 live births with 95% of the patients surviving their retinoblastoma. Stratifying data on the time of diagnosis and status of genetic testing, the number of screened patients gradually increased from 5% in the beginning of the period to 96% in the last five-year period. A cohort of 181 retinoblastoma survivors with sporadic disease (15% heritable) did not receive genetic testing. Since the introduction of routine testing, one of 14 sporadic unilateral patients tested (7%) has been identified with a germline mutation. Before routine testing, five additional sporadic unilateral patients have been identified as heritable. CONCLUSION: Only a minority of Danish retinoblastoma patients diagnosed before routine genetic testing was offered have been RB1 screened. To counsel the remaining untested patients and their families sufficiently regarding the risk to offspring and elevated risk of second primary cancers, we recommend information and access to genetic counseling and RB1 screening. This has ethical, psychological and possible economic consequences, and should be handled with caution.

Ray-tracing analysis of intraocular lens power in situ.

PURPOSE: To describe a method for back-solving the power of an intraocular lens (IOL) in situ based on laser biometry and ray-tracing analysis of the pseudophakic eye. SETTING: University Eye Clinic, Aarhus Hospital, Aarhus, Denmark. DESIGN: Evaluation of diagnostic test or technology. METHODS: This study comprised pseudophakic eyes with an IOL power ranging from -2.00 to +36.00 diopters (D). Preoperatively, the corneal radius was measured with conventional autokeratometry and the axial length (AL) with optical biometry. After surgery, the position of the IOL was recorded using laser interferometry. Based on the postoperative refraction and the biometric measurements, a ray-tracing analysis was performed back-solving for the power of the IOL in situ. The analysis was performed assuming pupil diameters from 0.0 to 8.0 mm with and without correction for the Stiles-Crawford effect. RESULTS: The study evaluated 767 pseudophakic eyes (583 patients). Assuming a 3.0 mm pupil, the mean prediction error between the labeled and the calculated IOL power (± 1 standard deviation [SD]) was -0.26 D ± 0.65 (SD) (range -2.4 to +1.8 D). The prediction error showed no bias with IOL power or with AL. The calculated IOL power depended on the assumed pupil size and the Stiles-Crawford effect. However, the latter had a modulatory effect on the prediction error for large pupil diameters (>5.0 mm) only. CONCLUSION: The optics of the pseudophakic eye can be accurately described using exact ray tracing and modern biometric techniques.

Simultaneous quantification of 17 immune mediators in aqueous humour from patients with corneal rejection.

PURPOSE: To simultaneously quantitate and compare the concentrations of 17 immune mediators: (1) the cytokines interleukin-1beta, IL-2, IL-4, IL-5, IL-6, IL-7, IL-10, IL-12p70, IL-13, IL-17, tumour necrosis factor-alpha, interferon-gamma; (2) the growth factors granulocyte-monocyte colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF), and (3) the chemokines CXCL-8, monocyte chemoattractant protein-1, and macrophage inflammatory protein-1beta in aqueous humour from patients with corneal rejection and patients with a non-inflammatory condition in the anterior chamber. METHODS: Aqueous humour was obtained by paracentesis of the anterior chamber in 14 patients with corneal rejection, three patients with cataract and six patients with Fuchs' endothelial dystrophy. Simultaneous quantitation of 17 mediators in 25 micro l aqueous humour from each patient was performed by employing a highly sensitive Luminex 100 multiplex array assay. RESULTS: All 17 immune mediators were detected in aqueous humour from rejection patients. The ranges of the immune mediators were determined. The immune mediators were significantly increased in aqueous humour from rejection patients compared with that from other patients. CONCLUSIONS: The Luminex 100 multiplex array assay is very efficient in simultaneous quantitation of multiple immune mediators in small volumes of aqueous humour. A total of 17 immune mediators were increased in aqueous humour from rejection patients. This underlines the complex immunological interactions of the rejection process.

The congenital "ant-egg" cataract phenotype is caused by a missense mutation in connexin46.

PURPOSE: "Ant-egg" cataract is a rare, distinct variety of congenital/infantile cataract that was reported in a large Danish family in 1967. This cataract phenotype is characterized by ant-egg-like bodies embedded in the lens in a laminar configuration and is inherited as an autosomal dominant trait. We retrieved the family and performed linkage analysis to determine the disease locus and identify the mutated gene. METHODS: The family (CC00103) was identified in a National Register of Hereditary Eye Diseases and updated based on The Danish Civil Register System. Genome wide linkage analysis and haplotyping using STS marker systems were carried out to achieve a LOD score above 3. The disease-causing candidate gene was sequenced and the mutation was identified and verified by restriction enzyme digestion of genomic DNA from all individuals in family CC00103 and 60 healthy controls. RESULTS: Linkage analysis resulted in a LOD score of 3.91 for marker D13S1275 located close to the known cataract gene GJA3. A novel missense mutation c.32T > C (L11S), was found by sequencing DNA from two affected members. The mutation was present in all affected individuals and was neither found in unaffected family members nor in 60 healthy individuals by restriction enzyme digests. CONCLUSIONS: The congenital "ant-egg" cataract phenotype is caused by a L11S mutation in connexin46 (Cx46) located in the signal peptide domain. Further studies are needed to unravel the mechanism leading to the formation of the "ant-eggs".

Alpha-1--antitrypsin in aqueous humour from patients with corneal allograft rejection.

PURPOSE: To investigate the presence and concentration of alpha1-antitrypsin in aqueous humour at the time of corneal rejection and to compare results obtained from patients with reversible and irreversible rejection. METHODS: Samples of aqueous humour were obtained from 17 patients with acute corneal endothelial allograft rejection. The presence of alpha1-antitrypsin in aqueous humour was confirmed by immunoblotting and measured employing a sandwich ELISA. Total protein concentrations in aqueous humour were measured using Bradford's method. The outcome of corneal rejection episodes was determined 1 month after diagnosing corneal rejection and described as reversible or irreversible rejection. RESULTS: alpha1-antitrypsin was detected in aqueous humour. Patients with reversible rejection had significantly higher alpha1-antitrypsin concentration than patients with irreversible rejection (p = 0.044). There was no significant difference in total protein concentrations (p = 0.745), and no correlation was found between alpha1-antitrypsin and total protein concentrations (p = 0.368). CONCLUSIONS: alpha1-antitrypsin in aqueous humour seems to signal a favourable outcome of corneal rejection. The possible mechanism is discussed.

Soluble CD163 and interleukin-6 are increased in aqueous humour from patients with endothelial rejection of corneal grafts.

PURPOSE: To evaluate soluble CD163 (sCD163) as a new marker of macrophage activity in aqueous humour from patients with corneal rejection and to investigate correlations between sCD163, the CD163 inducing interleukin-6 (IL-6), and albumin; to investigate whether increases in sCD163 and IL-6 levels in aqueous humour were results of intra- or extraocular production, and to investigate the impact of sCD163 and IL-6 levels on graft outcome. METHODS: Aqueous humour was obtained from 19 patients with endothelial rejection of corneal grafts, seven cataract patients and five cataract patients with uncomplicated corneal grafts. The presence of sCD163 was investigated by immunoprecipitation and immunoblotting. The concentrations of sCD163, IL-6 and albumin were measured by ELISA. RESULTS: Soluble CD163 was detected in aqueous humour from patients with corneal rejection. Soluble CD163, IL-6 and albumin were significantly increased in aqueous humour from patients with corneal rejection when compared with control patients. When normalizing sCD163 and IL-6 with albumin, the differences between rejection patients and control groups became insignificant for sCD163 but remained significant for IL-6. Neither IL-6 nor sCD163 were related to the outcome of the corneal rejection. CONCLUSIONS: Both sCD163 and IL-6 are present in high levels in aqueous humour from patients with rejection of corneal grafts. Our data suggest that IL-6 is produced locally, whereas sCD163 is at least partly derived from influx from plasma.

Proteomic analysis of aqueous humour from patients with acute corneal rejection.

PURPOSE: To compare the basic proteomic composition of aqueous humour (AH) from patients with corneal rejection (patients) with AH from patients with cataract (controls). METHODS: Aqueous humour was analysed for total protein concentration using Bradford's method and for protein composition using two-dimensional (2D) gel electrophoresis. Image analysis was used to detect protein spots in 2D gels that were increased by more than factor 2 in patients as compared with controls. Increased spots were identified by immunoblotting and mass spectrometry. RESULTS: Aqueous humour from patients contained significantly higher total protein concentration than did AH from controls. A total of 31 spots were significantly increased in 2D gels from patients. The spots were derived from albumin, alpha1-antitrypsin, apolipoprotein J, cytokeratin type II, serin proteinase inhibitor and transthyretin. After correction of spot volumes by total protein concentrations, 10 spots derived from albumin, cytokeratin type II and alpha1-antitrypsin remained significantly increased. CONCLUSION: The proteomic composition of AH differed significantly between patients and controls. The identified proteins suggest that the changes in AH are due to at least three different mechanisms: breakdown of the aqueous-blood barrier, enzymatic degradation, and liberation of locally synthesized proteins.