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Franz H Grus - One of the best experts on this subject based on the ideXlab platform.
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proteomic analysis of Tear fluid reveals disease specific patterns in patients with parkinson s disease a pilot study
Parkinsonism & Related Disorders, 2019Co-Authors: Matthias Boerger, Sebastian Funke, Andreas Leha, Annaelisa Roser, Annkatrin Wuestemann, Fabian Maass, Mathias Bahr, Franz H GrusAbstract:Abstract Background The diagnosis of Parkinson's disease (PD) is still challenging and biomarkers could contribute to an improved diagnostic accuracy. Tear fluid (TF) is an easily accessible body fluid reflecting pathophysiological changes in systemic and ocular diseases and is already used as a biomarker source for several ophthalmological disorders. Here, we analyzed the TF of patients with PD and controls (CTR) to describe disease-related changes in TF and identify putative biomarkers for the diagnosis of PD. Methods Unstimulated TF samples of a pilot cohort with 36 PD patients and 18 CTR were collected via Schirmer Tear test strips and then analyzed via a Bottom-up liquid chromatography electrospray ionization tandem mass spectrometry (BULCMS) workflow, followed by functional analysis encompassing Protein-Protein interaction as well as cellular component and pathway analysis. Results BULCMS analysis lead to the identification of 571 Tear Proteins (false discovery rate, FDR Conclusions To our best knowledge, this is the first description of TF proteome in PD patients. Tear Protein level alterations suggest the contribution of different disease-related mechanisms in ocular pathology in PD and propose candidate Proteins to be validated as potential biomarkers in larger cohorts.
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analysis of the effects of preservative free tafluprost on the Tear proteome
American Journal of Translational Research, 2016Co-Authors: Sebastian Funke, Norbert Pfeiffer, Dominik Wolters, Katrin Lorenz, Sabine Beck, Marion Kotterer, Natarajan Perumal, Franz H GrusAbstract:The purpose of the present study was to assess the ocular surface health status in primary open angle glaucoma (POAG) patients switching from topical application of preserved latanoprost (LT) to preservative-free tafluprost (PFT) by Tear proteomic monitoring. Tear fluid of POAG patients showing dry eye symptoms, using LT and switching to PFT as well as Tear fluid of healthy controls has been examined. Tear proteome dynamics was monitored over 24 weeks in a first mass spectrometric explorative analysis in a small POAG patient cohort (N = 3). Longitudinal responses of candidate Proteins as well as cytokines were comparatively analyzed by microarray in a larger cohort of POAG patients (N = 16) and healthy controls (N = 15). Clinical parameters including Tear breakup time (TBUT) and basal Schirmer test (BST) were recorded. Distinct post-switch level alterations could be documented in POAG Tear Proteins (> 1000). Cellular leakage Proteins, dry eye related candidates and cytokines showed predominantly level diminishment in POAG patients and approximation to the Tear Protein level of healthy controls in response to PFT. Tear Proteins like pyruvate kinase isozymes M1/M2 or galectin 7 displayed linear Tear film level decline in POAG patients (R2≥0.9; P < 0.05) distinctly converging the healthy level. Proteomic outcome fit well with improved clinical parameters, TBUT and BST. In conclusion, Tear proteomic alterations indicated ocular surface recovery regarding epithelia leakage and inflammation recession. Together with improved clinical parameters the study output proposes beneficial effects of PFT glaucoma therapy.
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effect of contact lenses on the Protein composition in Tear film a Proteinchip study
Graefes Archive for Clinical and Experimental Ophthalmology, 2011Co-Authors: Christina Kramann, Norbert Pfeiffer, Nils Boehm, Katrin Lorenz, Nelli Wehrwein, B M Stoffelns, Franz H GrusAbstract:The aim of this study was to analyze and compare the effects of rigid gas permeable and soft contact lenses on the Protein composition in the Tear film of contact lens wearers. Wearers of soft contact lenses (CL_S, n = 13) and rigid gas permeable contact lenses (CL_H, n = 13) were recruited for this study. Thirteen non-contact lens wearers were also included as the control. Tears were collected using Schirmer strips and frozen until use. The Tears were eluted and analyzed on ProteinChips SELDI-TOF (surface-enhanced laser desorption and ionization in time of flight mass spectrometry; Bio-Rad, USA) with different chromatographic surfaces (cationic and anionic exchanger and reversed phase surface). The SELDI spectra were analyzed by multivariate statistical analysis and artificial neural networks in order to find a biomarker panel which differentiates best between the groups. In order to identify Protein/peptide peaks from SELDI spectra which showed a significant difference between groups, fractionated Tear samples were analyzed using MALDI-TOF MS (matrix-assisted laser desorption/ionization time-of-flight mass spectrometry). For validation of biomarkers, we used an antibody microarray approach. Complex patterns of Tear Proteins and peptides were detected in the control group and in both contact lens groups. The Tear Protein composition in both wearers of rigid gas permeable (CL_H) and soft contact lenses (CL_S) differed significantly from Protein composition in non-contact lens wearers (p < 0.01). The identification of biomarkers revealed an increase of Protein S100 A8 in the group of wearers of soft contact lenses (CL_S) and a decrease of a main Tear Protein, lysozyme, in both contact lens groups. The identified biomarker cystatin was upregulated in the group of rigid gas permeable lens wearers (CL_H), whereas the Protein intensity of secretoglobin was significantly reduced in this group. Using the microarray approach, detected alterations could be confirmed. Contact lens wear alters the Protein profiles in a complex manner. This study demonstrates that significant changes can be found in wearers of soft contact lenses (CL_S) and rigid gas permeable contact lenses (CL_H). Some biomarker intensities are significantly altered only in the group of rigid gas permeable lens wearers (CL_H).
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effects of multipurpose contact lens solutions on the Protein composition of the Tear film
Contact Lens and Anterior Eye, 2005Co-Authors: Franz H Grus, Christina Kramann, N Bozkurt, N Wiegel, Kai Bruns, N Lackner, Norbert PfeifferAbstract:Abstract Objectives: To analyze the influence of multipurpose contact lens cleaning solutions on Tear Proteins. Changes in Tear film Protein profiles of contact lens wearers who used several marketed brands of multipurpose contact lens care solutions, were assessed by ProteinChip analysis. Methods: Three studies were conducted. Study I was a comparison of Complete and OptiFree multipurpose solutions. Study II was a study with Complete Moisture Plus solution, Study II was a comparison of Renu and Solocare contact lens solutions. Wearers of soft contact lenses were assigned to use the contact lens care solutions for 4 weeks. Non-contact lens wearing patients were used as controls. Tear samples of each participant were analyzed with the ProteinChip (SELDI–TOF) system. Multivariate statistical analysis and artificial neural networks were used to determine the Tear Protein profiles of each study group. Results: Before starting the use of the solutions, the Tear Protein composition in all contact lens wearers deviated from the Tear composition of the non-contact lens wearing controls. After 4 weeks of using the different care regimens, the Tear Protein composition of the patients using Complete or Complete Moisture Plus solutions tended to move toward that of the non-contact lens wearing controls. The Tear Protein composition of patients using the OptiFree, Renu or Solocare solutions did not undergo a measureable change in the Protein level. Conclusions: The ProteinChip system can analyze Protein profiles for large-scale applications as in clinical studies. Two multipurpose solutions, Complete and Complete Mositure Plus, demonstrated a beneficial effect on the Tear Proteins in contact lens wearers.
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changes in the Tear Protein patterns of diabetic patients using two dimensional electrophoresis
Advances in Experimental Medicine and Biology, 2002Co-Authors: Simone Herber, Franz H Grus, Perihan Sabuncuo, Albert J AugustinAbstract:Four to five million Germans suffer from diabetes mellitus, and worldwide there are about 100 million people that suffer with the disease. In diabetic patients, dry eye and other ocular surface diseases occur more often than in healthy subjects. Very little is known about the alterations in Tears caused by diabetes mellitus, or its influence on the pathogenesis of e.g., the dry eye disease. Recent studies from our group showed that there are differences in the one-dimensional electrophoretic Tear Protein separations between diabetic patients and healthy volunteers.1,2 The aim of this study is to analyze the Tear Protein patterns of patients suffering from diabetes (DIA) (Fig.l), and to compare them to the patterns of healthy subjects (CTRL) using two-dimensional electrophoretic techniques.
Lei Zhou - One of the best experts on this subject based on the ideXlab platform.
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Tear Proteins Calcium binding Protein A4 (S100A4) and Prolactin Induced Protein (PIP) are Potential Biomarkers for Thyroid Eye Disease
Nature Publishing Group, 2018Co-Authors: Chiaw-ling Chng, Lay Leng Seah, Morgan Yang, Sunny Yu Shen, Siew Kwan Koh, Yan Gao, Lu Deng, Louis Tong, Roger Wilmer Beuerman, Lei ZhouAbstract:Abstract There are no reliable biomarkers to predict thyroid eye disease (TED) in patients with autoimmune thyroid disease (AITD) currently. Several evidences support the involvement of the lacrimal gland in TED. The aim of our study was to quantitatively correlate the changes in Tear Protein profile with increasing severity of TED. Tear samples were collected from four groups of patients; AITD without TED (AITD), AITD with mild TED (mild TED), AITD with severe TED (severe TED) and normal controls. A total of 72 patients were recruited for the study. In discovery phase, isobaric tags for relative and absolute quantification (iTRAQ) 4-plex was used for quantitative proteomics analysis. For verification of results from discovery phase, sequential window acquisition of all theoretical fragment ion spectra (SWATH) was used to analyze an independent cohort from normal controls, AITD, mild TED and severe TED. Two Proteins, S100A4 and PIP showed consistent dysregulation trends in the discovery and validation phase experiments. Our study demonstrated the differences in Tear proteome across the spectrum of different severity and activity of TED in patients with AITD. Two Tear Proteins, S100A4 and PIP may serve as potential biomarkers to predict progression to severe TED in patients with AITD
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proteomic analysis revealed the altered Tear Protein profile in a rabbit model of sjogren s syndrome associated dry eye
Proteomics, 2013Co-Authors: Lei Zhou, Siew Kwan Koh, Roger W Beuerman, Ruihua Wei, Ping Zhao, Chuanqing DingAbstract:Sjogren's syndrome (SS) is an autoimmune disease that results in pathological dryness of mouth and eye. The diagnosis of SS depends on both clinical evaluation and specific antibodies. The goal of this study was to use quantitative proteomics to investigate changes in Tear Proteins in a rabbit model of SS-associated dry eye, induced autoimmune dacryoadenitis (IAD). Proteomic analysis was performed by iTRAQ and nano LC-MS/MS on Tears collected from the ocular surface, and specific Proteins were verified by high resolution MRM. It was found that in the Tears of IAD rabbits at 2 and 4 weeks after induction, S100 A6, S100 A9, and serum albumin were upregulated, whereas serotransferrin (TF), prolactin-inducible Protein (PIP), polymeric immunoglobulin receptor (pIgR), and Ig gamma chain C region were downregulated. High resolution MRM with mTRAQ labeling verified the changes in S100 A6, TF, PIP, and pIgR. Our results indicated significant changes of Tear Proteins in IAD rabbits, suggesting these Proteins could potentially be used as biomarkers for the diagnosis and prognosis of dry eye. Several of these Proteins were also found in the Tears of non-SS dry eye patients indicating a common basis of ocular surface pathology, however, pIgR appears to be unique to SS.
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in depth analysis of the human Tear proteome
Journal of Proteomics, 2012Co-Authors: Lei Zhou, Shao Zhen Zhao, Liyan Chen, Vivek Tanavde, X Li, Roger W BeuermanAbstract:The Tears, a critical body fluid of the surface of the eye, contain an unknown number of molecules including Proteins/peptides, lipids, small molecule metabolites, and electrolytes. There have been continued efforts for exploring the human Tear proteome to develop biomarkers of disease. In this study, we used the high speed TripleTOF 5600 system as the platform to analyze the human Tear proteome from healthy subjects (3 females and 1 male, average age: 36±14). We have identified 1543 Proteins in the Tears with less than 1% false discovery rate, which represents the largest number of human Tear Proteins reported to date. The data set was analyzed for gene ontology (GO) and compared with the human plasma proteome, NEIBank lacrimal gland gene dataset and NEIBank cornea gene dataset. This comprehensive Tear Protein list may serve as a reference list of human Tear proteome for biomarker research of ocular diseases or establishment of MRM (Multiple Reaction Monitoring) assays for targeted analysis. Tear fluid is a useful and an accessible source not only for evaluating ocular surface tissues (cornea and conjunctiva), inflammation, lacrimal gland function and a number of disease conditions, such as dry eye as well as response to treatment.
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proteomic profiling of inflammatory signaling molecules in the Tears of patients on chronic glaucoma medication
Investigative Ophthalmology & Visual Science, 2011Co-Authors: Tina T Wong, Siew Kwan Koh, Louis Tong, Lei Zhou, Shao Zhen Zhao, Roger Wilmer BeuermanAbstract:PURPOSE To identify the Tear Proteins associated with the long-term use of glaucoma medication by using proteomic analysis and to compare these Proteins to those previously reported in primary dry eye disease. METHODS Eighteen patients treated with topical antiglaucoma medications and 10 normal age-matched subjects with no prior topical treatment were recruited for the study. Tears were collected by using Schirmer's strip and analyzed by iTRAQ (isobaric tag for relative and absolute quantitation) for Tear Proteins by mass spectrometry. Conjunctival samples were collected and RNA expression determined by PCR. RESULTS Of the 124 identified Tear Proteins (99% confidence, ProtScore ≥ 2.0), we found that the Tear levels of S100-A8, S100-A9, mammaglobin B, and 14-3-3 ζ/δ were significantly increased in the medicated group compared with levels in the nonmedicated group (P < 0.05). For S100-A9, mammaglobin B, and 14-3-3 ζ/δ, use of topical medication for less than 1 year did not reach statistical significance compared with that in the nonmedicated group. Eyes on topical medication for less than 1 year showed a decrease in proline-rich 4 Protein Tear level (P = 0.0049) compared to nonmedicated group. The Tear Proteins detected in the medicated group differed from those in the primary dry eye group. CONCLUSIONS Treatment with topical antiglaucoma medications for longer than 1 year may start to induce ocular surface inflammation. The inflammatory Tear Protein profile present in chronically medicated glaucomatous eyes appears to be different from that found in primary dry eye. Identification of Tear Proteins specific to medicated glaucomatous eyes will help to specifically develop targeted screening modalities and therapeutic agents different from current conventional dry eye management.
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elevation of human α defensins and s100 calcium binding Proteins a8 and a9 in Tear fluid of patients with pterygium
Investigative Ophthalmology & Visual Science, 2009Co-Authors: Lei Zhou, Roger W Beuerman, Fook Tim Chew, Leonard P K Ang, Choi Mun Chan, Donald T H TanAbstract:PURPOSE The pathogenesis of pterygia is still not well understood. Recent studies suggest that it may be associated with inflammation and progressive proliferation triggered by ultraviolet radiation. In this study the authors determined that the inflammatory nature of pterygium is reflected in the Protein components of Tears. METHODS Consent for this study was obtained from 12 patients (average age, 57 years; eight men, four women) with unilateral pterygium. Tears were collected from diseased eyes and contralateral healthy control eyes with the use of fire-polished 10-microL calibrated glass pipettes before pterygium surgery. Tear Protein profiles obtained from diseased and control eyes (seven patient samples) were compared using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) technology. Tears from another five patients with pterygium were used for subsequent Protein identification experiments with nano-LC coupled with nano-electrospray tandem mass spectrometry (nano-ESI-MS/MS). RESULTS SELDI mass spectra showed that one peptide cluster with a molecular weight of 3.4 kDa and two Proteins with molecular weights of 10.8 kDa and 12.7 kDa were elevated in Tears of eyes with pterygium. Proteins of interest were identified by nano-LC-ESI-MS/MS as human alpha-defensins (human neutrophil peptide [HNP]-1, HNP-2, HNP-3) and S100 calcium-binding Proteins A8 and A9. Mean concentrations (n = 7) of alpha-defensins were 1.33 +/- 0.47 microg/mL (HNP-1, P < 0.015) and 0.61 +/- 0.23 microg/mL (HNP-2, P < 0.012) for pterygium eyes and 0.17 +/- 0.12 microg/mL (HNP-1) and 0.02 +/- 0.02 microg/mL (HNP-2) for fellow control eyes. Compared with Tears from eyes without pterygium or other abnormalities, the level of S100 A8 increased 1.4- to 13.4-fold (average fold change, 4.5) and S100 A9 increased 1.5- to 4.0-fold (average fold change, 2.3) in 4 of 7 patients. CONCLUSIONS The upregulated expression of human alpha-defensins and S100 A8 and A9 in Tear fluids of patients with pterygium indicates that they may be part of the response of the ocular surface to the formation of this fibrovascular tissue or the accompanying inflammation. They may also serve as a useful indicator for predicting recurrent pterygium.
Albert J Augustin - One of the best experts on this subject based on the ideXlab platform.
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changes in the Tear Protein patterns of diabetic patients using two dimensional electrophoresis
Advances in Experimental Medicine and Biology, 2002Co-Authors: Simone Herber, Franz H Grus, Perihan Sabuncuo, Albert J AugustinAbstract:Four to five million Germans suffer from diabetes mellitus, and worldwide there are about 100 million people that suffer with the disease. In diabetic patients, dry eye and other ocular surface diseases occur more often than in healthy subjects. Very little is known about the alterations in Tears caused by diabetes mellitus, or its influence on the pathogenesis of e.g., the dry eye disease. Recent studies from our group showed that there are differences in the one-dimensional electrophoretic Tear Protein separations between diabetic patients and healthy volunteers.1,2 The aim of this study is to analyze the Tear Protein patterns of patients suffering from diabetes (DIA) (Fig.l), and to compare them to the patterns of healthy subjects (CTRL) using two-dimensional electrophoretic techniques.
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two dimensional analysis of Tear Protein patterns of diabetic patients
Electrophoresis, 2001Co-Authors: Simone Herber, Franz H Grus, Perihan Sabuncuo, Albert J AugustinAbstract:In diabetic patients, dry eye and other ocular surface diseases occur more often than in healthy subjects. The aim of this study was to analyze the Tear Protein patterns of patients suffering from diabetes mellitus type II (dia) and to compare them to the patterns of healthy volunteers (ctrl). Tear Proteins of nonstimulated Tears of 20 patients (ctrl n = 10, dia n = 10) were separated using two-dimensional electrophoretic techniques. The Protein patterns of each group were analyzed by a multivariate analysis of discriminance. Furthermore, for all spots of each gel, a 50×50 variables pH/Mr (molecular weight) array was generated and subsequently analyzed by a multivariate analysis of discriminance. Additionally, an artificial neural network was trained using the matrix data as input and a sensitivity analysis was performed to figure out, which spots were the most important to differentiate between the Tear Protein patterns. In both groups a complex staining pattern could be obtained. In diabetic patients significantly more spots were detected compared to the control group (P<0.02). The analysis of discriminance found a highly significant difference between dia and ctrl (P<0.00001). Using the matrix data, the analysis of discriminance showed a significant difference between the two groups, too (P<0.0001). The sensitivity analysis by means of the artificial neural network revealed several spots that were more expressed or more frequently present in the diabetic group. Our findings reveal that the composition of Tear Proteins of diabetic patients is different from that of healthy subjects. The use of the two-dimensional electrophoretic technique could give more insight into the diabetic-related changes in the Tear film composition.
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analysis of Tear Protein patterns of dry eye patients using fluorescent staining dyes and two dimensional quantification algorithms
Electrophoresis, 2001Co-Authors: Franz H Grus, Perihan Sabuncuo, Albert J AugustinAbstract:Tear Proteins of nonstimulated Tears of 29 patients (healthy subjects, n = 8; dry-eye syndrome patients, n = 12; diabetic dry-eye patients, n = 9) were electrophoretically separated and stained by SYPRO Orange, followed by Coomassie blue staining. Both, the fluorescent and the Coomassie stains were subsequently analyzed by an automated two-dimensional algorithm for finding and quantification of peaks and by a discriminant analysis. Using SYPRO Orange, an average number of peaks/sample between three (at 200 ms) and 15 (at 3000 ms) could be found. In comparison, Coomassie staining resulted only in an average number of six peaks/sample. This corresponds to a sensitivity obtained at approx. 400-600 ms exposure time of SYPRO Orange stained gels. For all exposure times, the Protein patterns of the three clinical groups were statistically significantly different from each other (P < 0.05). Only at 200 ms the distances between the groups decreased slightly. The Coomassie-stained gels revealed only a mid range discrimination power similar to that of 200-400 ms exposure in the fluorescing gels. The use of SYPRO Orange provides faster results than those obtained by Coomassie staining. In addition, the sensitivity of staining can be varied even in the same gel by changing the exposure time. The use of the two-dimensional algorithm allows to distinguish between the three clinical groups in accordance to earlier studies using one-dimensional densitographic raw data. Thus, the high speed of evaluation and the more sensitive results as compared to earlier studies could be a step further in the use of Tear Protein patterns in the diagnosis of DRY.
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high performance liquid chromatography analysis of Tear Protein patterns in diabetic and non diabetic dry eye patients
European Journal of Ophthalmology, 2001Co-Authors: Franz H Grus, Albert J AugustinAbstract:PurposeTo analyze and compare the high performance liquid chromatography (HPLC) runs of Tear Proteins from diabetic (DIDRY) and non-diabetic (DRY) dry-eye patients, and healthy subjects (CTRL). The...
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analysis of Tear Protein patterns by a neural network as a diagnostical tool for the detection of dry eyes
Electrophoresis, 1999Co-Authors: F H Grus, Albert J AugustinAbstract:The electrophoretic patterns of Tears from patients with dry-eye disease (n = 43) and from healthy subjects (n = 17) were analyzed by means of multivariate statistical methods and an artificial neural network (ANN), following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). From each electrophoretic pattern a data set was created, randomly divided into test (unknown samples) and training patterns (known samples), with ANN training by one of these sets. After training, the performance of the ANN was checked by presenting the test data set to the ANN. Furthermore, the data was classified using multivariate analysis of discriminance. The groups were significantly different from each other (P < 0.05). The statistical procedure yielded 97% (known samples) and 71% (unknown samples) correct classifications. The ANN revealed 89% of correct classifications using the test set (unknown samples). The use of pruning algorithms (optimization procedure which automatically eliminates small weighted neurons) or genetic algorithms (optimization procedure which performs genetically induced changes of the neural net) resulted in a slight decrease of correct classifications compared to those of the nonoptimized neural network. The results reveal significant differences between the two groups. Using the ANN we were able to classify the electrophoretic Tear Protein pattern for diagnostic purposes.
Roger W Beuerman - One of the best experts on this subject based on the ideXlab platform.
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proteomic analysis revealed the altered Tear Protein profile in a rabbit model of sjogren s syndrome associated dry eye
Proteomics, 2013Co-Authors: Lei Zhou, Siew Kwan Koh, Roger W Beuerman, Ruihua Wei, Ping Zhao, Chuanqing DingAbstract:Sjogren's syndrome (SS) is an autoimmune disease that results in pathological dryness of mouth and eye. The diagnosis of SS depends on both clinical evaluation and specific antibodies. The goal of this study was to use quantitative proteomics to investigate changes in Tear Proteins in a rabbit model of SS-associated dry eye, induced autoimmune dacryoadenitis (IAD). Proteomic analysis was performed by iTRAQ and nano LC-MS/MS on Tears collected from the ocular surface, and specific Proteins were verified by high resolution MRM. It was found that in the Tears of IAD rabbits at 2 and 4 weeks after induction, S100 A6, S100 A9, and serum albumin were upregulated, whereas serotransferrin (TF), prolactin-inducible Protein (PIP), polymeric immunoglobulin receptor (pIgR), and Ig gamma chain C region were downregulated. High resolution MRM with mTRAQ labeling verified the changes in S100 A6, TF, PIP, and pIgR. Our results indicated significant changes of Tear Proteins in IAD rabbits, suggesting these Proteins could potentially be used as biomarkers for the diagnosis and prognosis of dry eye. Several of these Proteins were also found in the Tears of non-SS dry eye patients indicating a common basis of ocular surface pathology, however, pIgR appears to be unique to SS.
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in depth analysis of the human Tear proteome
Journal of Proteomics, 2012Co-Authors: Lei Zhou, Shao Zhen Zhao, Liyan Chen, Vivek Tanavde, X Li, Roger W BeuermanAbstract:The Tears, a critical body fluid of the surface of the eye, contain an unknown number of molecules including Proteins/peptides, lipids, small molecule metabolites, and electrolytes. There have been continued efforts for exploring the human Tear proteome to develop biomarkers of disease. In this study, we used the high speed TripleTOF 5600 system as the platform to analyze the human Tear proteome from healthy subjects (3 females and 1 male, average age: 36±14). We have identified 1543 Proteins in the Tears with less than 1% false discovery rate, which represents the largest number of human Tear Proteins reported to date. The data set was analyzed for gene ontology (GO) and compared with the human plasma proteome, NEIBank lacrimal gland gene dataset and NEIBank cornea gene dataset. This comprehensive Tear Protein list may serve as a reference list of human Tear proteome for biomarker research of ocular diseases or establishment of MRM (Multiple Reaction Monitoring) assays for targeted analysis. Tear fluid is a useful and an accessible source not only for evaluating ocular surface tissues (cornea and conjunctiva), inflammation, lacrimal gland function and a number of disease conditions, such as dry eye as well as response to treatment.
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elevation of human α defensins and s100 calcium binding Proteins a8 and a9 in Tear fluid of patients with pterygium
Investigative Ophthalmology & Visual Science, 2009Co-Authors: Lei Zhou, Roger W Beuerman, Fook Tim Chew, Leonard P K Ang, Choi Mun Chan, Donald T H TanAbstract:PURPOSE The pathogenesis of pterygia is still not well understood. Recent studies suggest that it may be associated with inflammation and progressive proliferation triggered by ultraviolet radiation. In this study the authors determined that the inflammatory nature of pterygium is reflected in the Protein components of Tears. METHODS Consent for this study was obtained from 12 patients (average age, 57 years; eight men, four women) with unilateral pterygium. Tears were collected from diseased eyes and contralateral healthy control eyes with the use of fire-polished 10-microL calibrated glass pipettes before pterygium surgery. Tear Protein profiles obtained from diseased and control eyes (seven patient samples) were compared using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) technology. Tears from another five patients with pterygium were used for subsequent Protein identification experiments with nano-LC coupled with nano-electrospray tandem mass spectrometry (nano-ESI-MS/MS). RESULTS SELDI mass spectra showed that one peptide cluster with a molecular weight of 3.4 kDa and two Proteins with molecular weights of 10.8 kDa and 12.7 kDa were elevated in Tears of eyes with pterygium. Proteins of interest were identified by nano-LC-ESI-MS/MS as human alpha-defensins (human neutrophil peptide [HNP]-1, HNP-2, HNP-3) and S100 calcium-binding Proteins A8 and A9. Mean concentrations (n = 7) of alpha-defensins were 1.33 +/- 0.47 microg/mL (HNP-1, P < 0.015) and 0.61 +/- 0.23 microg/mL (HNP-2, P < 0.012) for pterygium eyes and 0.17 +/- 0.12 microg/mL (HNP-1) and 0.02 +/- 0.02 microg/mL (HNP-2) for fellow control eyes. Compared with Tears from eyes without pterygium or other abnormalities, the level of S100 A8 increased 1.4- to 13.4-fold (average fold change, 4.5) and S100 A9 increased 1.5- to 4.0-fold (average fold change, 2.3) in 4 of 7 patients. CONCLUSIONS The upregulated expression of human alpha-defensins and S100 A8 and A9 in Tear fluids of patients with pterygium indicates that they may be part of the response of the ocular surface to the formation of this fibrovascular tissue or the accompanying inflammation. They may also serve as a useful indicator for predicting recurrent pterygium.
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proteomic analysis of rabbit Tear fluid defensin levels after an experimental corneal wound are correlated to wound closure
Proteomics, 2007Co-Authors: Lei Zhou, Roger W Beuerman, Liqun Huang, Amutha Barathi, Yong Hwee Foo, Fook Tim Chew, Donald T H TanAbstract:The cornea is the major refracting optical element of the eye and therefore critical for forming a retinal image. The exposed surface of the eye is protected from pathogens by the innate immune system whose components include defensins, naturally occurring peptides with antimicrobial properties, and the physical barrier formed by the outer epithelial layer of the cornea. The proteomic approach has revealed that Tear levels of defensins are correlated with the course of healing of an experimental corneal wound. Tears were collected from New Zealand White rabbits prior to (day 0) and daily for 5 days (days 1-5) following a standard unilateral 6 mm diameter corneal epithelial abrasion. Tear Protein profiles obtained from wounded and contra-lateral control eyes were compared using SELDI ProteinChip technology. Peptides and Proteins of interest were purified by RP-HPLC and characterized by nanoESI-MS/MS. Mass spectra of Tears on post-wound day 1, revealed 13 peaks whose level decreased and five that increased. During wound healing the Tear Protein profile correlated with wound closure. An important finding was that the levels of rabbit defensins (NP-1 and NP-2), which were elevated after wounding returned to normal levels by the time the corneal abrasion healed. Relative quantification of NP-2 in Tear fluid prior to (day 0) and after corneal wounding (days 1- 3) was determined using iTRAQ technology. A corneal wound eliminates the barrier function of innate immunity and puts the cornea at risk from microbial attack until the epithelial cells restore the surface barrier. The increased availability of defensins in the Tears during healing suggests that these peptides could protect the cornea from microbial attack during a period of increased vulnerability.
Marlyn P Langford - One of the best experts on this subject based on the ideXlab platform.
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bilateral acute pyogenic conjunctivitis with iritis induced by unilateral topical application of bacterial peptidoglycan muramyl dipeptide in adult rabbits
Experimental Eye Research, 2013Co-Authors: Marlyn P Langford, Bridgett Foreman, Lana Srur, J P Ganley, Thomas B RedensAbstract:The factors responsible for the conjunctivitis and iritis associated with acute ocular infection and post enteric inflammatory disease are not fully known. The pro-inflammatory activity of unilateral topical application of muramyl dipeptide (MDP; the smallest bio-active Gram-positive and Gram-negative bacterial cell wall component) was investigated in adult rabbits. The resultant bilateral conjunctivitis/iritis and pyogenic responses were characterized. Bilateral symptoms were graded by slit lamp examinations; Tear fluid, Schirmer tests (Tear production), blood and aqueous humor (AH) samples were obtained from MDP-treated and untreated rabbits. MDP concentration, gamma-glutamyltranspeptidase activity (GGT; key enzyme in glutathione recapture, xenobiotic detoxification, eicosanoid synthesis and neutrophil function), Protein concentration, and Tear cell density, cytology, and immunofluorescent antibody reactivity to GGT and calreticulin (CRT; MDP-binding Protein) were determined. MDP was cleared from ipsilateral Tears and serum by 6 h, but was undetected in mock-treated contralateral Tears. Bilateral signs of acute transient pyogenic conjunctivitis, characterized by Tearing, lid edema, conjunctival hyperemia, chemosis and leukocytic infiltrate with iritis (erythema and aqueous flare) were detected. Milder symptoms occurred in the mock-treated contralateral eyes. Bilateral symptoms, Tear production, Tear Protein, GGT activity, and mucopurulent discharge (containing up to 2.5–5.0 × 106 cells/mL) were elevated 4–8 h post MDP and resolved to near pre-treatment levels by 24 h. Tear GGT activity and Protein levels were higher in MDP-treated and mock-treated contralateral eyes than in eyes of untreated adult rabbits (p's < 0.001). Elevated Tear GGT activity was associated with histopathology and increased vascular and epithelial permeability to serum Protein, GGT-positive epithelia cells, macrophages and heterophils. Repeat MDP applications induced recurrent induction and resolution patterns of bilateral conjunctivitis/iritis and Tear GGT activity, but ipsilateral GGT responses were lower. The results suggest unilateral topical MDP application to adult rabbit eyes induces a bilateral acute pyogenic conjunctivitis/iritis (PCI) characterized by increased vascular and epithelial permeability similar to acute bacterial conjunctivitis in man. The detection of CRT/GGT positive heterophils in Tears suggests efferocytosis (phagocytosis of dead/dying cells). Tear GGT activity may be a useful means to quantify MDP-induced toxicity and extraocular inflammation.
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analysis of neutralizing antibodies to enterovirus 70 and coxsackievirus a24 variant levels of immunoglobulins and total Protein in Tears of patients with acute hemorrhagic conjunctivitis
Ocular Immunology and Inflammation, 1995Co-Authors: Marlyn P Langford, Jeffrey B Robertson, Rogelio OrillacAbstract:Tear samples were collected from 37 residents of the Dominican Republic 3.5 U/ml) and anti-Coxsackievirus A24 variant (CA24v) TNA (10<1-3 U/ml), but no anti-poliovirus (PV) TNA was detected. The anti-EV70 TNA in pooled Tear samples sedimented in sucrose density gradient fractions that corresponded to 19-7S serum anti-PV immunoglobulin (1g). Anti-CA24v TNA sedimented as 7S1g. 1gG levels (mean, 3.13±4.2mg/ml) were higher than 1gA levels (mean, 0.92±0.98 mg/ml) in 21 of 27 Tear samples. 1gG levels in Tears from six patients with bilateral AHC were associated with total Tear Protein (p=0.003), but not with the levels of TNA or interferon (IFN). The total Protein in AHC Tears (5.13±1.72 mg/ml) was two-fold less than the total Protein in normal Tears (11.2±3.25 mg/ml). 1gA levels...
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analysis of neutralizing antibodies to enterovirus 70 and coxsackievirus a24 variant levels of immunoglobulins and total Protein in Tears of patients with acute hemorrhagic conjunctivitis
Ocular Immunology and Inflammation, 1995Co-Authors: Marlyn P Langford, Jeffrey B Robertson, Rogelio OrillacAbstract:Tear samples were collected from 37 residents of the Dominican Republic 3.5) U/ml) and anti-Coxsackievirus A24 variant (CA24v) TNA (10(<1-3) U/ml), but no anti-poliovirus (PV) TNA was detected. The anti-EV70 TNA in pooled Tear samples sedimented in sucrose density gradient fractions that corresponded to 19-7S serum anti-PV immunoglobulin (1g). Anti-CA24v TNA sedimented as 7S1g. 1gG levels (mean, 3.13±4.2mg/ml) were higher than 1gA levels (mean, 0.92±0.98 mg/ml) in 21 of 27 Tear samples. 1gG levels in Tears from six patients with bilateral AHC were associated with total Tear Protein (p=0.003), but not with the levels of TNA or interferon (IFN). The total Protein in AHC Tears (5.13±1.72 mg/ml) was two-fold less than the total Protein in normal Tears (11.2±3.25 mg/ml). 1gA levels increased from 0.31±.3 to 1.34±1.28 mg/ml in Tears collected up to 3 d p.o. of AHC. 1gM was not detected (<0.01 mg/ml). EV70 was isolated from the Tears of one patient. Taken together, our results suggest that EV70 and CA24v are endemic in the Dominican Republic and that the 1992 epidemic of AHC was due to EV70. The detection of 19S (IgM) and high levels of 7S (IgG) TNA to EV70<1 d p.o. of AHC indicate a rapid ocular immune response to EV70 and suggests that virus-specific TNAs inhibit AHC virus infection.
