Etiology Of Keratoconus Research Articles

Expression of Tissue Inhibitors of Metalloproteinases (TIMP-1, TIMP-2, TIMP-3, TIMP-4) in Blood Serum of Patients with Keratoconus.

The etiology of keratoconus is unclear. Current evidence suggests that inflammatory and systemic mechanisms might play a role in its pathophysiology. The proper interaction of proteolytic enzymes-matrix metalloproteinases-and their specific tissue inhibitors (TIMPs) within the cornea is essential in maintaining its structure, transparency and healing processes. The aim of the study was to determine the concentration of the TIMPs TIMP-1, TIMP-2, TIMP-3, and TIMP-4 in the blood serum samples of patients with keratoconus compared to the control group. The study encompassed 132 patients, of which 83 people constituted the study group and 49 the control group. The concentration of selected TIMPs was determined using the Human Magnetic Luminex® Performance Assay method. In the study group, the concentrations of TIMP-1 and TIMP-3 were statistically significantly reduced, and TIMP-2 and TIMP-4 increased compared to the control group. The analysis of individual TIMPs in terms of their usefulness as potential predictors of keratoconus showed high results of diagnostic sensitivity and specificity for all TIMPs, in particular for TIMP-1 and TIMP-2. The above results may indicate systemic disturbances in the TIMPs regulation among keratoconus patients. High diagnostic sensitivity and specificity of all TIMPs, in particular TIMP-1 and TIMP-2, may confirm their participation in the etiopathogenesis of this disease.

Association between Polymorphism rs61876744 in PNPLA2 Gene and Keratoconus in a Saudi Cohort.

The genetic etiology of Keratoconus (KC) in Middle Eastern Arabs of Saudi origin is still unclear. A recent genome-wide study identified two significant loci in the region of PNPLA2 (rs61876744) and CSNK1E (rs138380) for KC that may be associated with KC in the Saudi population. In addition, polymorphisms in the apolipoprotein E (APOE) gene, namely, rs429358 and rs7412, responsible for APOE allelic variants ε2, ε3, and ε4, may influence KC via oxidative stress mechanism(s). Thus, we investigated the possible association of polymorphisms rs61876744, rs138380, rs429358, rs7412, and APOE genotypes in KC patients of the Saudi population. This study included 98 KC cases and 167 controls. Polymorphisms rs6187644 and rs138380 were genotyped using TaqMan assays, and rs429358 and rs7412 were genotyped via Sanger sequencing. Although the allele frequency of rs61876744(T) in PNPLA2 was a protective effect against KC (odds ratio (OR) = 0.64, 95% confidence interval (CI) = 0.44-0.93), the p-value (p = 0.020) was not significant for multiple testing correction (p = 0.05/4 = 0.015). However, rs6187644 genotype showed a modestly significant protective effect in the dominant model (OR = 0.53, 95% CI = 0.32-0.88, p = 0.013). Polymorphisms rs138380, rs429358, and rs7412 showed no significant allelic or genotype association with KC. However, the ε2-carriers (ε2/ε2 and ε2/ε3 genotypes) exhibited a greater than 5-fold increased risk of KC, albeit non-significantly (p = 0.055). Regression analysis showed no significant effect of age, gender, and the four polymorphisms on KC. Our results suggest that polymorphism rs6187644 in PNPLA2 might be associated with KC in the Middle Eastern Arabs of Saudi origin but warrant a large-scale association analysis at this locus.

Screening and identification of genes related to ferroptosis in keratoconus

Corneal keratoconus (KC) is a dilated (ectatic) corneal disease characterized by a central thinning of the cornea, which causes protrusion into a conical shape that seriously affects vision. However, due to the complex etiology of keratoconus, its entire mechanism remains unclear and there is no mechanism-directed treatment method. Ferroptosis is a novel programmed cell death mechanism related to lipid peroxidation, stress, and amino acid metabolism, which plays a crucial role in various diseases. This study aimed to explore the relationship between keratoconus and ferroptosis, to provide new insights into the mechanism of keratoconus development, and potential treatment options based on further elucidation of this mechanism. The corresponding mRNA microarray expression matrix data of KC patients were obtained from GEO database (GSE204791). Weighted co-expression network analysis (WGCNA) and support vector machine recursive feature elimination (SVM-RFE) were selected to screen hub genes, which were overlapped with ferroptosis genes (FRGs) from FerrDb. GO and GSEA were performed to analyze differential pathways, ssGSEA was used to determine immune status, and then, feasible drugs were predicted by gene-drug network. Additionally, we predicted the miRNA and IncRNA of hub genes to identify the underlying mechanism of disease so as to predict treatment for the disease. The epithelial transcriptome from keratoconus tissue mRNA microarray data (GSE204791) was extracted for the main analysis, including eight epithelial cells and eight epithelial control cells. The differential genes that were overlapped by WGCAN, SVM-RFE and FRGs were mainly related to oxidative stress, immune regulation, cellular inflammation, and metal ion transport. Through further analysis, aldo–keto reductase family 1 member C3 (AKR1C3) was selected, and negatively correlated with mature CD56 natural killer (NK) cells and macrophages. Then, gene-drug interaction network analysis and miRNA prediction were performed through the website. It was concluded that four immune-related drugs (INDOMETHACIN, DAUNORUBICIN, DOXORUBICIN, DOCETAXEL) and a miRNA (has-miR-184) were screened to predict potential drugs and targets for disease treatment. To our knowledge, this was the first report of KC being associated with ferroptosis and prompted search for differential genes to predict drug targets of gene immunotherapy. Our findings provided insight and a solid basis for the analysis and treatment of KC.

Non-allergic eye rubbing is a major behavioral risk factor for keratoconus.

Since the environmental, behavioral, and socioeconomic factors in the etiology of keratoconus (KTCN) remain poorly understood, we characterized them as features influencing KTCN phenotype, and especially affecting the corneal epithelium (CE). In this case-control study, 118 KTCN patients and 73 controls were clinically examined and the Questionnaire covering the aforementioned aspects was completed and then statistically elaborated. Selected KTCN-specific findings were correlated with the outcomes of the RNA-seq assessment of the CE samples. Male sex, eye rubbing, time of using a computer after work, and dust in the working environment, were the substantial KTCN risk factors identified in multivariate analysis, with ORs of 8.66, 7.36, 2.35, and 5.25, respectively. Analyses for genes whose expression in the CE was correlated with the eye rubbing manner showed the enrichment in apoptosis (TP53, BCL2L1), chaperon-related (TLN1, CTDSP2, SRPRA), unfolded protein response (NFYA, TLN1, CTDSP2, SRPRA), cell adhesion (TGFBI, PTPN1, PDPK1), and cellular stress (TFDP1, SRPRA, CAPZB) pathways. Genes whose expression was extrapolated to the allergy status didn't contribute to IgE-related or other inflammatory pathways. Presented findings support the hypothesis of chronic mechanical corneal trauma in KTCN. Eye-rubbing causes CE damage and triggers cellular stress which through its influence on cell apoptosis, migration, and adhesion affects the KTCN phenotype.

RISK PROFILE OF KERATOCONUS: A TERTIARY CARE HOSPITAL BASED STUDY

Background: The study aimed to assess the risk factors implicated in the causation of keratoconus and the demographic profile of the patients diagnosed as keratoconus. Materials and Methods: This retrospective, observational, hospital-based study included 376 patients presenting to the ophthalmology outpatient department of a tertiary care hospital in Kashmir from January 2022 to December 2022. Written informed consent was obtained from all participants after explaining the nature of the study.A well-designed questionnaire was used to collect the data of the recruited patients.The demographic characteristics like age, gender, residence were noted and the role of certain risk factors associated with causation of keratoconus was noted. Results: The mean age of participants was 21.8 ± 4.7 years (range, 9-32). The study participants were 58.6% (n = 220/376) males and 41.4% (n = 156/376) females. Majority of the participants belonged to urban areas(55.60%) than rural areas(44.4%). The maximum association of keratoconus was found with eye rubbing(61.2%) followed by male gender (58.6%) and age group 21-30 years (51%). Conclusion: We conclude that eye rubbing is the most significant risk factor, followed by exposure to sunlight and ocular allergy. The findings of our study corroborate that the etiology of keratoconus is multifactorial.

Biomechanics of keratoconus: Two numerical studies.

The steep cornea in keratoconus can greatly impair eyesight. The etiology of keratoconus remains unclear but early injury that weakens the corneal stromal architecture has been implicated. To explore keratoconus mechanics, we conducted two numerical simulation studies. A finite-element model describing the five corneal layers and the heterogeneous mechanical behaviors of the ground substance and lamellar collagen-fiber architecture in the anterior and posterior stroma was developed using the Holzapfel-Gasser-Ogden constitutive model. The geometry was from a healthy subject. Its stroma was divided into anterior, middle, and posterior layers to assess the effect of changing regional mechanical parameters on corneal displacement and maximum principal stress under intraocular pressure. Specifically, the effect of softening an inferocentral corneal button, the collagen-based tissues throughout the whole cornea, or specific stromal layers in the button was examined. The effect of simply disorganizing the orthogonally-oriented posterior stromal fibers in the button was also assessed. The healthy cornea was also subjected to eye rubbing-like loading to identify the corneal layer(s) that experienced the most tensional stress. Conical deformation and corneal thinning emerged when the corneal button or the mid-posterior stroma of the button underwent gradual softening or when the collagen fibers in the mid-posterior stroma of the button were dispersed. Softening the anterior layers of the button or the whole cornea did not evoke conical deformation. Button softening greatly increased and disrupted the stress on Bowman's membrane while mid-posterior stromal softening increased stress in the anterior layers. Eye rubbing profoundly stressed the deep posterior stroma while other layers were negligibly affected. These observations suggest that keratoconus could be initiated, at least partly, by mechanical instability/damage in the mid-posterior stroma that then imposes stress on the anterior layers. This may explain why subclinical keratoconus is marked by posterior but not anterior elevation on videokeratoscopy.

Risk Profile of Keratoconus among Secondary School Students in the West Region of Cameroon.

This study determined the risk factors of keratoconus (KC) among secondary school students in the West Region of Cameroon. A stratified, random sampling technique was used to select the 3015 secondary school students, 8 years and older, within the West Region of Cameroon. Selected school students completed the validated Keratoconus Risk Investigative Survey (KRIS) and a structured demographic questionnaire to determine the risk profile of KC. Descriptive analysis, logistic regression and p-values were used to provide an overview of the demographic findings and the risk factors of KC. Estimates were made as the proportion of affected school students and presented with a 95% confidence interval (CI). Multivariate logistic regression analysis was performed to explore the association between KC and the independent predictors that were found significant in the univariate analysis. The ages of the majority (93.2%) of students ranged from eight years to 18 years (mean = 13.18 ± years) and were mostly female (59.7%). Gender (OR 2.024, p < 0.001), eye rubbing (OR 3.615, p < 0.001), exposure to sunlight (OR 2.735, p < 0.001), blood relations with KC (OR 41.819, p < 0.001) and allergic experience (OR 1.070, p < 0.001) were considered. Eye rubbing was the most significant risk factor of keratoconus followed by refractive error, allergic experiences and sunlight exposure. These findings support the evidence that the etiology of KC is multifactorial, with eye rubbing being the most significant factor in this cohort. There is a need to address eye rubbing among students to minimize the risk of KC. Furthermore, 34.46% of students in Cameroon were at risk of developing KC. Hence the risk profile is that one engages in eye rubbing, has a family member with KC, spends more than eight hours per week in the sun and is prone to allergies. It will therefore be prudent for these risk factors for keratoconus to be included in the school health education programs.

Phosphoprotein network analysis of corneal epithelium of keratoconus patients.

Keratoconus (KC) is non-inflammatory, bilateral progressive corneal ectasia, and a disease of established biomechanical instability. The etiology of KC is believed to be multifactorial. Although previous studies gained insight into the understanding of the disease, little is known thus far on global protein phosphorylation changes in keratoconus. We performed phosphoproteome analysis of corneal epithelium from control (N=5) and KC patients. Tandem mass tag (TMT) multiplexing technology along with immobilized metal affinity chromatography (IMAC) were used for the phosphopeptides enrichment and quantitation. Enriched peptides were analyzed on Orbitrap Fusion Tribrid mass spectrometer. This leads to the identification of 2939 unique phosphopeptides derived from 1270 proteins. We observed significant differential phosphorylation of 591 phosphopeptides corresponding to 375 proteins. Our results provide first phosphoproteomic signature of the keratoconus disease and identified dysregulated signaling pathways that can be targeted for therapy in future studies.

Fourteen years follow-up of a stable unilateral Keratoconus: unique case report of clinical, tomographical and biomechanical stability

BackgroundKeratoconus (KC) is a noninflammatory corneal ectatic disorder. In 2015, the Global Consensus on Keratoconus and Ectatic Diseases agreed that the pathophysiology of KC includes environmental, biomechanical, genetic, and biochemical disorders on one hand, and that true unilateral KC does not exist on the other hand. However, with the increasingly advancements in detection methods, we report the first case of a stable unilateral keratoconus with the longest follow up period of 14 years (2006–2020). We used topographic, tomographic, and biomechanical values for both eyes over the years to confirm the diagnosis, which has never been done before. Our study focuses on a single patient therefore it illustrates the mere possibility that unilateral keratoconus exists.Case presentationWe present the case of a 19-year-old male with no previous ocular or general health conditions who presented to our clinic in November 2006 for incidental finding of decreased vision of the right eye (OD) on a routine examination. Topographies, tomographies, and biomechanical analysis of both eyes were obtained and showed a unilateral right keratoconus at the time. Patient admitted to unilateral right eye rubbing. Although we cannot prove that previous eye rubbing alone led to these initial symptoms, he was advised to stop rubbing and was followed up without any intervention for fourteen years during which topographic, tomographic, and biomechanical values for both eyes remained stable, proving for the first time that unilateral KC could exist.ConclusionWe think that the data we are presenting is important because acknowledging that true unilateral keratoconus exists questions the genetic or primary biomechanical etiology of keratoconus versus the secondary biomechanical etiologies like eye rubbing. Our report also shows the importance of corneal biomechanics in detecting early changes. This is important to detect early, prevent progression, and tailor treatment.

Nutritional and Metabolic Imbalance in Keratoconus.

Keratoconus (KC) is a progressive corneal degeneration characterized by structural changes consisting of progressive thinning and steepening of the cornea. These alterations result in biomechanical weakening and, clinically, in vision loss. While the etiology of KC has been the object of study for over a century, no single agent has been found. Recent reviews suggest that KC is a multifactorial disease that is associated with a wide variety of genetic and environmental factors. While KC is typically considered a disease of the cornea, associations with systemic conditions have been well described over the years. In particular, nutritional and metabolic imbalance, such as the redox status, hormones, metabolites, and micronutrients (vitamins and metal ions), can deeply influence KC initiation and progression. In this paper, we comprehensively review the different nutritional (vitamins and minerals) and metabolic (hormones and metabolites) factors that are altered in KC, discussing their possible implication in the pathophysiology of the disease.

Impact of Hypothyroidism on Tomography and Biomechanics in Keratoconus - Cross-Sectional and Longitudinal Assessment within the Homburg Keratoconus Center at the Time of Inclusion and after 1 Year.

The etiology of keratoconus (KC) is probably multifactorial but remains essentially unknown. Previous scientific observations have suggested that hypothyroidism might play a role in the development and progression of KC. The purpose of this study was to analyze the tomographic and biomechanical parameters in KC patients with or without hypothyroidism. Twenty-eight patients with KC and hypothyroidism (HT group) and fifty-six KC patients without thyroid dysfunction (WHT group) with matching gender and age were analyzed. Mean age was 40.3 years (range 14 - 57) in the HT group and 40.3 years (range 14 - 57) in the WHT group. Routine ophthalmic examinations consisted of corneal tomography and biomechanical parameters. We extracted the following KC parameters from the Pentacam (Pentacam HR, Oculus, Wetzlar, Germany): Keratoconus Index (KI), maximum keratometry (Kmax), astigmatism, and thinnest pachymetry (TP). From the ocular response analyzer (ORA, Reichert Ophthalmic Instruments, Depew, NY, USA), we extracted corneal hysteresis (CH), corneal resistance factor (CRF), and KC match index (KMI). The comparison of the tomographic and biomechanical values from cross-sectional and longitudinal analyses showed no significant differences between the HT and WHT groups. The severity of KC based on tomographical and biomechanical parameters does not seem to depend on the presence of hypothyroidism.

Assessing the role of systemic inflammation in the etiopathogenesis of advanced stage keratoconus.

Purpose:It was aimed to compare the levels of inflammation-related parameters, such as neutrophil/lymphocyte ratio (NLR) and platelet/lymphocyte ratio (PLR), in patients with advanced keratoconus (KC) and healthy controls. Also, we evaluated the relationships between these parameters and several corneal topography values used in the KC diagnostic index.Methods:Forty patients with advanced and 40 healthy volunteers were included in this study. In the KC group, 20 patients were nonprogressive KC and 20 patients were progressive KC. In all participating individuals, we evaluated detailed ophthalmologic examination findings and complete blood count data, while corneal topographic measurements were also recorded in patients with KC.Results:The mean NLR value was 2.3 ± 1.19 in the progressive KC group; nonprogressive KC values were 1.99 ± 1.69 and 1.81 ± 0.72 in the control group. Mean PLR value was 113.24 ± 48.44 in the progressive KC group, nonprogressive KC values were 96.47 ± 31.04 and 104.09 ± 35.14 in the control group. No statistically significant difference was found between patients with progressive KC, nonprogressive KC, and healthy volunteers in terms of mean NLR and PLR values (P > 0.05). NLR values were found to demonstrate significant positive correlations with the corneal topography parameters, Symmetry Index front (r = 0.278, P = 0.025), KC Vertex front (r = 0.247, P = 0.048), and Baiocchi Calossi Versaci front (r = 0.273, P = 0.028); there was no significant relationship between corneal topography parameters and PLR values.Conclusion:Although there was no significant difference between the progressive KC, nonprogressive KC, and control groups in inflammation parameters such as NLR and PLR, a positive correlation was observed between the NLR value and some corneal topography findings used in the diagnosis of KC. The role of inflammation in the etiology of KC can be better understood by clinical studies and laboratory tests conducted with prospective studies involving a higher number of patients.

The Enigma of Environmental Factors in Keratoconus.

Keratoconus is an ectatic disorder with highly complex and varied causes including genetic variations and environmental factors. Its prevalence varies widely between regions and countries. Many environmental factors have been proposed to be associated with keratoconus, but the interpretation of their individual contributions is difficult due to the presence of many confounding variables. The current literature was reviewed to evaluate the strength of the associations and the causative effects of environmental factors on keratoconus. Ethnicity and consanguinity have been revealed as important determinants for geographical variations in keratoconus prevalence. Eye rubbing, atopy, floppy eyelid syndrome, contact lens wear, pregnancy, and thyroid hormone disturbances are likely associated with keratoconus. The first 4 factors can induce ocular surface inflammation, matrix metalloproteinase release, and keratocyte apoptosis, consistent with the postulated etiology of keratoconus. The associations of keratoconus with UV exposure, cigarette smoking, personality, and sex were less convincing once confounding factors were considered. Future studies powered for multivariate analysis of factors discussed will hopefully shed light on what is truly important in the development and progression of keratoconus.

Metabolomic Signature Discriminates Normal Human Cornea from Keratoconus-A Pilot GC/MS Study.

The molecular etiology of keratoconus (KC), a pathological condition of the human cornea, remains unclear. The aim of this work was to perform profiling of metabolites and identification of features discriminating this pathology from the normal cornea. The combination of gas chromatography and mass spectrometry (GC/MS) techniques has been applied for profiling and identification of metabolites in corneal buttons from 6 healthy controls and 7 KC patients. An untargeted GC/MS-based approach allowed the detection of 377 compounds, including 46 identified unique metabolites, whose levels enabled the separation of compared groups of samples in unsupervised hierarchical cluster analysis. There were 13 identified metabolites whose levels differentiated between groups of samples. Downregulation of several carboxylic acids, fatty acids, and steroids was observed in KC when compared to the normal cornea. Metabolic pathways associated with compounds that discriminated both groups were involved in energy production, lipid metabolism, and amino acid metabolism. An observed signature may reflect cellular processes involved in the development of KC pathology, including oxidative stress and inflammation.

Serum 25-hydroxyvitamin D, Selenium, Zinc and Copper in patients with keratoconus

PurposeTo assess the possible association between keratoconus (KC) and serum levels of 25-hydroxyvitamin D (25OHD), Selenium (Se), Zinc (Zn), and Copper (Cu) and to compare it with age-matched healthy subjects. MethodsOne hundred patients with KC and 100 normal subjects were included. The two groups were compared for serum 25OHD and serum levels of three trace elements: Se, Zn, and Cu. These factors were also compared between groups with different KC stages. ResultsSerum levels of vitamin D, Zn, Cu, and Se were significantly different between the KC and normal groups (P = 0.006, P = 0.015, P = 0.004, and P = 0.038, respectively). Although a lower level of 25OHD was found in severe stages of KC, it was not significantly different among different KC groups (P = 0.441). KC stage groups were not significantly different for mean serum Zn, Cu, and Se (P = 0.130, P = 0.98, P = 0.113, respectively). Although the Cu/Zn ratio was higher in cases than in controls, there was no significant difference between the two groups and between KC stages (P = 0.168, P = 0.143, respectively). ConclusionLower serum 25OHD, Cu, Zn, and Se were found in the KC group compared to the control group. The results of this study suggest that a lower antioxidative activity may be involved in the possible etiology of KC.

Keratoconus at a Molecular Level: A Review.

Keratoconus is the most common ectatic disease of the cornea. The disease is usually detected between ages 15 and 25. Incidence is estimated at one out of every 2000 individuals, with some specific ethnic groups being more at risk. Keratoconus manifests itself as a progressive stromal thinning and deformation of the corneal tissue into a conical shape. The etiology of keratoconus is uncertain, although several studies have associated the disease to environmental factors, behavioral conditions and certain genetic disorders. In an effort to better understand how the corneal stroma becomes compromised, multiple experiments have been conducted over the last few years looking at the cells themselves and the factors they produce. The secretion pathways and levels of inflammatory molecules, growth factors, digestive enzymes, and apoptotic factors are all relevant to keratoconus. This review describes the current knowledge of keratoconic pathological signaling pathways within the cornea that may help future developments in disease prevention, treatment and modeling. Anat Rec, 2019. © 2019 Wiley Periodicals, Inc.

An analytical enrichment-based review of structural genetic studies on keratoconus.

Keratoconus is a progressive bilateral corneal protrusion that leads to irregular astigmatism and impairment of vision. Keratoconus is an etiologically heterogeneous corneal dystrophy and both environmental and genetic factors play a role in its etiopathogenesis. In this analytical review, we have studied all the genes that are structurally associated with keratoconus and have tried to explain the function of each gene and its association with other eye disorders in a concise way. In addition, using gene set enrichment analysis, it was attempted to find the most important impaired metabolic pathways in keratoconus. Several genetic studies have been carried out on keratoconus and several genes have been identified as risk factors involved in the etiology of the disease. In the current study, 16 studies, including nine association studies, five genome-wide association studies, one linkage study, and one meta-analysis, were reviewed and based on the 19 genes found, enrichment was performed and the most important metabolic pathways involved in the disease were identified. The enrichment results indicated that the two pathways, interleukin 1 processing and assembly of collagen fibrils, are significantly associated with the disease. Obviously, the results of this study, in addition to providing information about the genes involved in the disease, can provide an integrated insight into the gene-based etiology of keratoconus and therapeutic opportunities thereof.

Overexpression of Tear Inflammatory Cytokines as Additional Finding in Keratoconus Patients and Their First Degree Family Members.

Keratoconus is a progressive corneal ectasia that may lead to severe visual impairment due to the irregular astigmatism caused by corneal thinning. In addition to its association with atopy, eye rubbing, or genetic component, late reports suggest the involvement of inflammation in the pathogenesis of the disease. Our aim was to determine the concentration of IL-4, IL-6, IL-10, RANTES, IFN gamma, and TNF alpha in the tear film of patients with keratoconus and their first degree family members. We analyzed forty-eight participants in an observational cross-sectional study. The diagnosis of keratoconus had to be confirmed in addition to a minimum of 47 D corneal refractive power by corneal topography readings provided by a Placido-based topography system and analysis of the pattern: irregular astigmatism with an asymmetric “bow-tie.” As for the other groups, the most important diagnosis criteria were a normal topographic pattern with a regular astigmatism. 17 keratoconus patients, 16 relatives, and 15 controls were recruited after clinical assessment as part of the research. The cytokine's mean values were similar in the keratoconus group and the relatives' samples but significantly higher compared to the controls. Important differences were found in IL-4 levels between keratoconus patients and relatives and between relatives and controls (mean difference of 302.42, p < 0.0016 and 219.16, p < 0.033, Tukey's HSD procedure). In the keratoconus group, using the CORR procedure, we found statistically strong correlations of IL-6 lacrimal concentrations with the disease stage (r = 0.56, p < 0.01), keratometry (r = 0.55, p < 0.02), pachymetry (r = −0.64, p < 0.048), and corneal hysteresis (r = −0.53, p < 0.02). Cytokine overexpression may be relevant for the inflammatory etiology of keratoconus. In conclusion, in the case of some first degree family members, the elevated tear biomarkers may represent a supplementary risk factor.

Collagen synthesis disruption and downregulation of core elements of TGF-β, Hippo, and Wnt pathways in keratoconus corneas.

To understand better the factors contributing to keratoconus (KTCN), we performed comprehensive transcriptome profiling of human KTCN corneas for the first time using an RNA-Seq approach. Twenty-five KTCN and 25 non-KTCN corneas were enrolled in this study. After RNA extraction, total RNA libraries were prepared and sequenced. The discovery RNA-Seq analysis (in eight KTCN and eight non-KTCN corneas) was conducted first, after which the replication RNA-Seq experiment was performed on a second set of samples (17 KTCN and 17 non-KTCN corneas). Over 82% of the genes and almost 75% of the transcripts detected as differentially expressed in KTCN and non-KTCN corneas were confirmed in the replication study using another set of samples. We used these differentially expressed genes to generate a network of KTCN-deregulated genes. We found an extensive disruption of collagen synthesis and maturation pathways, as well as downregulation of the core elements of the TGF-β, Hippo, and Wnt signaling pathways influencing corneal organization. This first comprehensive transcriptome profiling of human KTCN corneas points further to a complex etiology of KTCN.

Brittle cornea syndrome ZNF469 mutation carrier phenotype and segregation analysis of rare ZNF469 variants in familial keratoconus.

Brittle cornea syndrome 1 (BCS1) is a rare recessive condition characterized by extreme thinning of the cornea and sclera, caused by mutations in ZNF469. Keratoconus is a relatively common disease characterized by progressive thinning and ectasia of the cornea. The etiology of keratoconus is complex and not yet understood, but rare ZNF469 variants have recently been associated with disease. We investigated the phenotype of BCS1 carriers with known pathogenic ZNF469 mutations, and recruited families in which aggregation of keratoconus was observed to establish if rare variants in ZNF469 segregated with disease. Patients and family members were recruited and underwent comprehensive anterior segment examination, including corneal topography. Blood samples were donated and genomic DNA was extracted. The coding sequence and splice sites of ZNF469 were PCR amplified and Sanger sequenced. Four carriers of three BCS1-associated ZNF469 loss-of-function mutations (p.[Glu1392Ter], p.[Gln1930Argfs*6], p.[Gln1930fs*133]) were examined and none had keratoconus. One carrier had partially penetrant features of BCS1, including joint hypermobility. ZNF469 sequencing in 11 keratoconus families identified 9 rare (minor allele frequency [MAF] ≤ 0.025) variants predicted to be potentially damaging. However, in each instance the rare variant(s) identified, including two previously reported as potentially keratoconus-associated, did not segregate with the disease. The presence of heterozygous loss-of-function alleles in the ZNF469 gene did not cause keratoconus in the individuals examined. None of the rare nonsynonymous ZNF469 variants identified in the familial cohort conferred a high risk of keratoconus; therefore, genetic variants contributing to disease pathogenesis in these 11 families remain to be identified.