Histochemical Comparison of Human and Rat Lacrimal Glands: Implications for Bio-Engineering Studies.

Purpose: The purpose of this study was to determine whether rodent lacrimal glands (LGs) represent a suitable surrogate for human tissue in bio-engineering research, we undertook a meticulous histological and histochemical comparison of these two tissues. Methods: Histological techniques and immunohistochemistry were used to compare the structure of adult human and rat LG tissues and the expression of key functional tissue elements. Results: Compared with humans, the rat LG is comprised of much more densely packed acini which are devoid of an obvious central lumen. Myoepithelial, fibroblasts, dendritic cells, T cells, and putative progenitor cells are present in both tissues. However, human LG is replete with epithelium expressing cytokeratins 8 and 18, whereas rat LG epithelium does not express cytokeratin 8. Furthermore, human LG expresses aquaporins (AQPs) 1, 3, and 5, whereas rat LG expresses AQPs 1, 4, and 5. Additionally, mast cells were identified in the rat but not the human LGs and large numbers of plasma cells were detected in the human LGs but only limited numbers were present in the rat LGs. Conclusions: The cellular composition of the human and rat LGs is similar, although there is a marked difference in the actual histo-architectural arrangement of the tissue. Further variances in the epithelial cytokeratin profile, in tissue expression of AQPs and in mast cell and plasma cell infiltration, may prove significant. Translational Relevance: The rat LG can serve as a useful surrogate for the human equivalent, but there exist specific tissue differences meaning that caution must be observed when translating results to patients.

Histological and histochemical characteristics of lacrimal glands in beluga whales (Delphinapterus leucas).

The objective of this study was to describe the histological and histochemical characteristics of the lacrimal glands of beluga whales. The study was carried out on the formalin-fixed ocular globes from 96 carcasses of beluga whales found stranded in the St. Lawrence estuary in Quebec, Canada. Hematoxylin and eosin (H&E) stained slides from the eyes of each whale were examined for lacrimal glands. Histological description was done with H&E and Masson Trichrome (MT) stains. Period Acid-Schiff (PAS), Alcian blue (AB) pH 1.0 and 2.5, and High Iron Diamine (HID) stains were used for histochemical characterization of glycoproteins. Thirteen ocular samples from animals ranging from neonate to 48 y included sections of lacrimal glands. The H&E stain revealed a tubuloalveolar gland architecture, separated into lobules by dense connective tissue. Each lobule contained a mixture of acini and tubules with ductules. Small and large acini were composed of low and tall columnar cells, respectively. Acinar cells contained basophilic cytoplasmic granules. The ductules were lined with a bi-layered cuboidal-to-squamous epithelium. The MT stain highlighted the connective tissue separating ductules and acini. Large acini were positive for PAS and some small acini had patchy uptake. Positive staining for AB pH 1.0 and 2.5 was mainly seen in tall columnar cells as compared to small acini that had faint to no stain uptake. High Iron Diamine stain revealed 90% staining of all acinar cells, with 10% exhibiting a mixed blue-black tinge. It was concluded that the lacrimal glands of beluga whales have similar histological and histochemical findings to those of artiodactyla and carnivora orders.

L'objectif de cette étude était de décrire les caractéristiques histologiques et histochimiques des glandes lacrymales des bélugas. L'étude a été réalisée sur les globes oculaires fixés au formol de 96 carcasses de bélugas trouvées échouées dans l'estuaire du Saint-Laurent au Québec, Canada. Des lames colorées à l'hématoxyline et à l'éosine (H&E) des yeux de chaque baleine ont été examinées pour la présence de glandes lacrymales. La description histologique a été réalisée avec des colorations H&E et trichrome de Masson (MT). Les colorations Periodic acid-Schiff (PAS), au bleu Alcian (AB) pH 1,0 et 2,5, et diamine à haute teneur en fer (HID) ont été utilisées pour la caractérisation histochimique des glycoprotéines. Treize échantillons oculaires provenant d'animaux allant du nouveau-né à 48 ans comprenaient des sections de glandes lacrymales. La coloration H&E a révélé une architecture de glande tubulo-alvéolaire, séparée en lobules par un tissu conjonctif dense. Chaque lobule contenait un mélange d'acini et de tubules avec des ductules. Les petits et les grands acini étaient respectivement composés de cellules cylindriques basses et hautes. Les cellules acinaires contenaient des granules cytoplasmiques basophiles. Les canaux étaient tapissés d'un épithélium cuboïde à squameux bicouche. La coloration MT a mis en évidence le tissu conjonctif séparant les canaux et les acini. Les grands acini étaient positifs pour le PAS et certains petits acini avaient une absorption inégale. Une coloration positive pour AB pH 1,0 et 2,5 a été principalement observée dans les cellules cylindriques hautes par rapport aux petits acini qui avaient une absorption de coloration faible ou nulle. La coloration HDI a révélé une coloration de 90 % de toutes les cellules acinaires, 10 % présentant une teinte mixte bleu-noir. Il a été conclu que les glandes lacrymales des bélugas présentent des résultats histologiques et histochimiques similaires à ceux des ordres des artiodactyles et des carnivores.(Traduit par Docteur Serge Messier).

Binding of metals in dacryoliths by means of biomineralization.

Dacryoliths are organomineral aggregates released from human lacrimal ducts, and are known to include a 1 to 3 % (by volume) of a mineral (inorganic) component This study was based on a collection of approximately 800 dacryoliths collected from a patient in Siberia. The shapes and sizes of mineral grains, their composition and their distribution in the organic matrix are very diverse; our detailed investigation identifies a number of inorganic phases that were not previously recognized. These were imaged on a scanning electron microscope (Tescan VEGA II LMU), and the chemical composition of the inorganic component was determined by energy dispersive X-Ray spectroscopy (INCA Energy350, Oxford Instrument), and X-ray analysis (X-ray diffractometer XPert PRO (PANalytical). For the first time, we identified the following minerals which can be divided into three groups: 1) oxide Si, pyrite, calcium oxalate, barite, which are often present in the tissues of the living organism and are also known in geological processes; 2) abundant complex mineral phases, consisting of Al, Si, K and Ti or Fe (titanium and iron aluminosilicates), which are also present in organism but have no analogues in the abiogenic environment; and 3) oxygen-containing compounds of Ni and Pb-Cr (possibly nickel oxide and lead chromate, respectively) which are only found in the abiogenic environment. Those mineral phases of biogenic origin are due to an endogenous (pathological) process in the body caused by internal factors (hypoergosis and metabolic disorders). This biomineralization may have a role in the removal of toxic and / or excess ions of Fe, Ti, Cr, Pb, Ni, Al, and Si from tissues.

Longitudinal analysis of tear cathepsin S activity levels in male non-obese diabetic mice suggests its potential as an early stage biomarker of Sjögren's Syndrome.

CONTEXT: Cathepsin S (CTSS) activity is elevated in Sjögren's Syndrome (SS) patient tears. OBJECTIVE: To evaluate longitudinal expression of tear and tissue CTSS activity relative to other disease indicators in Non-Obese Diabetic (NOD) mice. METHODS: CTSS activity was measured in tears and lacrimal glands (LG) from male 1-6 month (M) NOD and 1 and 6 M BALB/c mice. Lymphocytic infiltration was quantified by histopathology, while disease-related proteins (Rab3D, CTSS, collagen 1) were quantified using q-PCR and immunofluorescence. RESULTS: In NOD LG, lymphocytic infiltration was noted by 2 M and established by 3 M (p < 0.01). IFN-É£, TNF-α, and MHC II expression were increased by 2 M (p < 0.01). Tear CTSS activity was significantly elevated at 2 M (p < 0.001) to a maximum of 10.1-fold by 6 M (p < 0.001). CTSS activity in LG lysates was significantly elevated by 2 M (p < 0.001) to a maximum of 14-fold by 3 M (p < 0.001). CTSS and Rab3D immunofluorescence were significantly increased and decreased maximally in LG acini by 3 M and 2 M, respectively. Comparable changes were not detected between 1 and 6 M BALB/c mouse LG, although Collagen 1 was decreased by 6 M in LG of both strains. CONCLUSION: Tear CTSS activity is elevated with other early disease indicators, suggesting potential as an early stage biomarker for SS.

Human postmortem lacrimal and submandibular glands stored in RNAlater are suitable for molecular, biochemical, and cell biological studies.

PURPOSE: Gene expression and protein analysis studies require high-quality human tissue which is a challenge and difficult to obtain through live human biopsies. Human postmortem lacrimal gland (LG) and submandibular gland (SMG) tissues have the potential to provide an invaluable source for studying the mechanisms involved in LG and SMG dysfunction. Therefore, we aimed to test the suitability of post-mortem LG and SMG for molecular, biochemical, and cell biological studies. METHODS: LG and SMG tissue from healthy donors was collected and immediately placed in RNAlater solution and then shipped overnight at 4 °C. After receipt, each gland was divided into three pieces for RNA, protein, and histological analysis, respectively. Total RNA isolated from each LG and SMG was analyzed for RNA integrity using an Agilent Bioanalyzer and reverse transcription-PCR (RT-PCR). For histology, tissues were embedded in paraffin and stained with hematoxylin and eosin. For protein analysis, lysates were prepared and processed for sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and western blotting. RESULTS: When the LG and SMG samples were preserved in RNAlater, the RNA integrity number (RIN) values from the LG and SMG were >7.0 from all three donors, while the RNAs from tissue not preserved in RNAlater were of poorer quality. The gene and/or protein expression of E-cadherin, aquaporin 5, alpha-smooth muscle actin (α-SMA), ß-actin, and GAPDH was preserved in all samples. In addition, histological analyses showed normal tubuloacinar structures of all glands with serous and mucous producing acini within lobules interspersed with adipose fat. CONCLUSIONS: In this study, we determined that RNA, protein, and histological sections obtained from postmortem human LG and SMG tissue preserved in RNAlater were of high quality. This would provide a viable source of human LG and SMG tissue suitable for studies of diseases that affect these glands, such as Sjögren's syndrome.

Comparative proteomic analysis of tear fluid in Graves' disease with and without orbitopathy.

BACKGROUND: Graves' orbitopathy (GO) is a severe organ-specific autoimmune inflammatory ocular complication most often associated with Graves' disease (GD). Besides the cosmetic problems these patients develop, GO may also cause severe, sight-threatening complications. Additionally, GO complicates the treatment of patients with GD, making the identification of Graves patients at risk for eye disease before they develop symptoms a critical step in the clinical management and quality of life of these patients. The high concentration of proteins in tear fluid makes it an important source for studying potential protein biomarkers for GO. PATIENTS AND METHODS: The aim of this study was to quantitatively compare tear fluid from GD patients with moderate/severe GO (GO) and patients with GD without GO (controls) using untargeted quantitative proteomics based on dimethyl labelling in combination with two-dimensional liquid chromatography-mass spectrometry. RESULTS: Among the 1212 proteins identified, 16 showed significant alterations in abundance between the two groups. Thus, in this study, we reveal a number of novel dysregulated proteins in GO which may contribute to a better understanding of the disease. In particular, upregulation of lacrimal gland proteins such as lysozyme C, lacritin, antileukoproteinase and zinc-alpha-2-glycoprotein 1 suggests involvement of the lacrimal gland in the pathogenesis of GO. CONCLUSIONS: It remains to be elucidated whether some of these proteins can be used as markers for patients at risk for developing GO as well as useful indicators for disease activity.

Undefined role of mucus as a barrier in ocular drug delivery.

Mucus layer covers the ocular surface, and soluble mucins are also present in the tear fluid. After topical ocular drug administration, the drugs and formulations may interact with mucus layer that may act as a barrier in ocular drug delivery. In this mini-review, we illustrate the mucin composition of the ocular surface and discuss the influence of mucus layer on ocular drug absorption. Based on the current knowledge the role of mucus barrier in drug delivery is still undefined. Furthermore, interactions with mucus may prolong the retention of drug formulations on the ocular surface. Mucus may decrease or increase ocular bioavailability depending on the magnitude of its role as barrier or retention site, respectively. Mechanistic studies are needed to clarify the role of mucin in ocular drug delivery.

Endoscopic dacryocystorhinostomy for acquired nasolacrimal duct obstruction: long-term results in 91 procedures.

BACKGROUND: The purpose of the present study was to evaluate long-term outcomes of endoscopic dacryocystorhinostomy (DCR) using a drill without the use of mucosal flaps. Ninety one procedures in eighty seven patients were reviewed. All patients showed epiphora, caused by primary or secondary nasolacrimal obstruction. METHODOLOGY: All patients underwent preoperative evaluation (irrigation and probing of the lacrimal drainage system, fluorescein tests, computerized tomography scan of the paranasal sinuses, dacryocystography and endoscopic examination of the nasal cavity). In 19 patients further intranasal procedures were conducted simultaneously with DCR (10 FESS, 2 septoplasties, 5 functional endoscopic sinus surgery (FESS) and septoplasties, 2 septoplasties and turbinoplasties). Stents were placed intraoperatively and removed 4 to 12 weeks, postoperatively. Postoperative follow-up ranged between 12 and 24 months. RESULTS: Long-term success was achieved in 87/91 procedures. No major complications were observed. Failure was caused by granulation tissue formation in three patients and inappropriate stent removal in one patient. CONCLUSION: The success rate achieved is comparable to success rates of external DCR.

Changes of lacrimal gland and tear inflammatory cytokines in thyroid-associated ophthalmopathy.

PURPOSE: To explore changes in lacrimal gland and tear inflammatory cytokines in thyroid-associated ophthalmopathy (TAO) patients. METHODS: Patients with TAO were divided into active and inactive TAO groups. These two TAO groups and the control completed the Ocular Surface Disease Index (OSDI), underwent thorough ophthalmologic examinations, and underwent orbital magnetic resonance scan to measure the size of the lacrimal gland. Basal tears, reflex tears induced by nasal stimulation and serum samples, were collected to analyze the concentrations of interleukin (IL)-1ß, IL-6, IL-7, IL-17A, interferon γ, and tumor necrosis factor α by multiplex bead analysis. RESULTS: The coronal lacrimal gland area was significantly larger in active (P < 0.000) and inactive TAO (P = 0.002) than in the control, and the axial lacrimal gland area was significantly larger in active (P < 0.000) and inactive TAO (P = 0.001) than in the control. The coronal lacrimal gland width was significantly greater in active (P < 0.000) and inactive TAO (P = 0.001) than in the control, and axial lacrimal gland width was significantly greater in active (P < 0.000) and inactive TAO (P = 0.035) than in the control. In TAO patients, the axial area was positively correlated with IL-1ß and IL-17A concentrations in tears (r = 0.357, P = 0.013; r = 0.359, P = 0.012), and both coronal and axial areas were positively correlated with IL-6 concentrations in tears (r = 0.346, P = 0.016; r = 0.340, P = 0.018). CONCLUSIONS: Increased inflammatory cytokines play an important role in ocular surface damages, and might be associated with the inflammatory involvement of the lacrimal gland.

Development and validation of UHPLC method for the determination of cyclosporine A in biological samples.

The aim of the study was to develop and validate a simple and rapid method for the determination of cyclosporine A (CsA) in ocular rabbit tissues using reversed-phase ultra-high-performance liquid chromatography (UHPLC) with UV detection. Previous publications on chromatographic methods of CsA determination in ocular tissues involved only reversed-phase HPLC separation, usually in combination with such detection techniques as radio-immunoassay and mass spectrometry. The application of the UHPLC technique allowed us to significantly decrease the analysis time. Cyclosporine D (CsD) was applied as the internal standard. Satisfactory separation was achieved on an XB-C18 Kinetex column at 60°C with the use of gradient elution mode. The retention times of CsA and CsD were found to be 4.5 and 5.1 min, respectively. The developed assay is specific, sensitive (limit of detection = 6 ng/mL and limit of quantitation = 18 ng/mL) and linear within the analyte concentration range of 0.018-5 µg/mL, with a correlation coefficient of 0.999. High sensitivity, low injection volume (10 µL), short time of analysis (6.5 min) and simplicity make this method useful for the fast analysis of CsA in rabbit ocular tissues and fluids: lacrimal fluid, aqueous humor, cornea, conjunctiva and eye globe.

In-depth analysis of the human tear proteome.

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.

Detection of BrdU-label retaining cells in the lacrimal gland: implications for tissue repair.

The purpose of the present study was to determine if the lacrimal gland contains 5-bromo-2'-deoxyuridine (BrdU)-label retaining cells and if they are involved in tissue repair. Animals were pulsed daily with BrdU injections for 7 consecutive days. After a chase period of 2, 4, or 12 weeks, the animals were sacrificed and the lacrimal glands were removed and processed for BrdU immunostaining. In another series of experiments, the lacrimal glands of 12-week chased animals were either left untreated or were injected with interleukin 1 (IL-1) to induce injury. Two and half days post-injection, the lacrimal glands were removed and processed for BrdU immunostaining. After 2 and 4 weeks of chase period, a substantial number of lacrimal gland cells were BrdU(+) (11.98 ± 1.84 and 7.95 ± 1.83 BrdU(+) cells/mm(2), respectively). After 12 weeks of chase, there was a 97% decline in the number of BrdU(+) cells (0.38 ± 0.06 BrdU(+) cells/mm(2)), suggesting that these BrdU-label retaining cells may represent slow-cycling adult stem/progenitor cells. In support of this hypothesis, the number of BrdU labeled cells increased over 7-fold during repair of the lacrimal gland (control: 0.41 ± 0.09 BrdU(+) cells/mm(2); injured: 2.91 ± 0.62 BrdU(+) cells/mm(2)). Furthermore, during repair, among BrdU(+) cells 58.2 ± 3.6 % were acinar cells, 26.4 ± 4.1% were myoepithelial cells, 0.4 ± 0.4% were ductal cells and 15.0 ± 3.0% were stromal cells. We conclude that the murine lacrimal gland contains BrdU-label retaining cells that are mobilized following injury to generate acinar, myoepithelial and ductal cells.

Porphyrins are not present in feline ocular tissues or corneal sequestra.

OBJECTIVE: To determine if feline lacrimal glands, glands of the third eyelid, corneas, and corneal sequestra contain porphyrins, which could be responsible for the brown/amber discoloration of corneal sequestra and tears in affected cats. PROCEDURES: Samples of grossly normal cornea, lacrimal gland, gland of the third eyelid, and sequestra obtained via keratectomy were collected. Porphyrin concentrations of the homogenate were determined by spectrofluorometry with protoporphyrin IX and coproporphyrin III dihydrochloride used as standards. A hamster harderian gland was used as a positive control. RESULTS: Normal tissues were harvested from one eye each of 14 nonclient owned, adult, mixed-breed, short-hair cats euthanized for reasons not associated with this study. Eighteen sequestra were acquired from cats undergoing unilateral lamellar keratectomies. Breeds of the affected cats included eight Himalayan, five domestic shorthair, and one each of four other breeds. Only the positive control and standards contained levels of porphyrins above background. All feline samples examined were histologically normal with no evidence of porphyrins. CONCLUSIONS: Porphyrins are absent in normal feline lacrimal glands, corneas, and corneal sequestra. Porphyrins do not appear to be the cause of the brown/amber color of feline corneal sequestra.

Lipidomics of human Meibomian gland secretions: Chemistry, biophysics, and physiological role of Meibomian lipids.

Human Meibomian gland secretions (MGS) are a complex mixture of diverse lipids that are produced by Meibomian glands that are located in the upper and the lower eyelids. During blinking, MGS are excreted onto the ocular surface, spread and mix with aqueous tears that are produced by lachrymal glands, and form an outermost part of an ocular structure called "the tear film" (TF). The main physiological role of TF is to protect delicate ocular structures (such as cornea and conjunctiva) from desiccating. Lipids that are produced by Meibomian glands are believed to "seal" the aqueous portion of TF by creating a hydrophobic barrier and, thus, retard evaporation of water from the ocular surface, which enhances the protective properties of TF. As lipids of MGS are interacting with underlying aqueous sublayer of TF, the chemical composition of MGS is critical for maintaining the overall stability of TF. There is a consensus that a small, but important part of Meibomian lipids, namely polar, or amphiphilic lipids, is of especial importance as it forms an intermediate layer between the aqueous layer of TF and its upper (and much thicker) lipid layer formed mostly of very nonpolar lipids, such as wax esters and cholesteryl esters. The purpose of this review is to summarize the current knowledge on the lipidomics of human MGS, including the discussions of the most effective modern analytical techniques, chemical composition of MGS, biophysical properties of Meibomian lipid films, and their relevance for the physiology of TF. Previously published results obtained in numerous laboratories, as well as novel data generated in the author's laboratory, are discussed. It is concluded that despite a substantial progress in the area of Meibomian glands lipidomics, there are large areas of uncertainty that need to be addressed in future experiments.

Changes of aquaporins in the lacrimal glands of a rabbit model of Sjögren's syndrome.

AIMS: To test the hypothesis that the expression of aquaporins (AQPs) 4 and 5 is altered in the lacrimal glands (LG) of rabbits with induced autoimmune dacryoadenitis (IAD). MATERIALS AND METHODS: LGs were obtained from adult female rabbits with IAD, and age-matched female control rabbits. LGs were processed for laser capture microdissection (LCM), real time RT-PCR, Western blot, and immunofluorescence for the detection and quantification of protein and mRNAs of AQP4 and AQP5 in whole LGs, and purified acinar cells and duct cells from specific duct segments. RESULTS: In rabbits with IAD, abundances of mRNAs for AQP4 and AQP5 from whole LGs were significantly lower than controls. Levels of mRNA for AQP4 were lower in most duct segments from rabbits with IAD. However, the mRNA abundance for AQP5 was significantly lower in acini from rabbits with IAD, while its abundance was higher in each duct segment. Western blot showed that the expression of AQP4 in LGs from rabbits with IAD was 36% more abundant than normal controls, whereas AQP5 was 72% less abundant. Immunofluorescence indicated that AQP4 immunoreactivity (AQP4-IR) was present on the basolateral membranes of acinar and ductal cells in control and diseased LGs, with ductal cells showing stronger AQP4-IR than acinar cells. AQP5-IR was found on apical and basolateral membranes of acinar cells, and showed a "mosaic" pattern, i.e., with some acini and/or acinar cells showing stronger AQP5-IR than others. Minimal AQP5-IR was detected in ductal cells from control animals, while its intensity was significantly increased in rabbits with IAD. CONCLUSIONS: These data strongly support our hypothesis that expressions of AQPs are altered in rabbits with IAD, and that specific ductal segment play important roles in lacrimal secretion.

Tear osmolarity in the diagnosis and management of dry eye disease.

PURPOSE: To evaluate the use of tear osmolarity in the diagnosis of dry eye disease. DESIGN: A prospective, observational case series to determine the clinical usefulness of tear osmolarity and commonly used objective tests to diagnose dry eye disease. METHODS: A multicenter, 10-site study consisting of 314 consecutive subjects between 18 and 82 years of age. Bilateral tear osmolarity, tear film break-up time (TBUT), corneal staining, conjunctival staining, Schirmer test, and meibomian gland grading were performed. Diagnostic performance was measured against a composite index of objective measurements that classified subjects as having normal, mild or moderate, or severe dry eye. The main outcome measures were sensitivity, specificity, area under the receiver operating characteristic curve, and intereye variability. RESULTS: Of the 6 tests, tear osmolarity was found to have superior diagnostic performance. The most sensitive threshold between normal and mild or moderate subjects was found to be 308 mOsms/L, whereas the most specific was found at 315 mOsms/L. At a cutoff of 312 mOsms/L, tear hyperosmolarity exhibited 73% sensitivity and 92% specificity. By contrast, the other common tests exhibited either poor sensitivity (corneal staining, 54%; conjunctival staining, 60%; meibomian gland grading, 61%) or poor specificity (tear film break-up time, 45%; Schirmer test, 51%). Tear osmolarity also had the highest area under the receiver operating characteristic curve (0.89). Intereye differences in osmolarity were found to correlate with increasing disease severity (r(2) = 0.32). CONCLUSIONS: Tear osmolarity is the best single metric both to diagnose and classify dry eye disease. Intereye variability is a characteristic of dry eye not seen in normal subjects.

Cloning, overexpression, purification, crystallization and preliminary X-ray analysis of a female-specific lipocalin (FLP) expressed in the lacrimal glands of Syrian hamsters.

Proteins belonging to the lipocalin superfamily are usually secretory proteins of molecular mass approximately 20 kDa with a hydrophobic pocket for the binding and transport of diverse small ligands. Various lipocalins have been associated with many biological processes, e.g. immunomodulation, odorant transport, pheromonal activity, retinoid transport, cancer-cell interactions etc. However, the exact functions of many lipocalins and the ligands bound by them are unclear. Previously, the cDNA of a 20 kDa lipocalin (FLP) which is female-specifically expressed in the lacrimal glands of Syrian (golden) hamsters and secreted in the tears of females has been identified and cloned. His-tagged recombinant FLP (rFLP) has now been cloned, overexpressed in Escherichia coli as a soluble protein and purified to homogeneity using Ni-affinity followed by size-exclusion chromatography. Purified rFLP was crystallized using the sitting-drop vapour-diffusion method. The crystals tested belonged to space group P2(1)2(1)2(1) and diffracted to beyond 1.86 A resolution. Solvent-content analysis indicated the presence of one monomer in the asymmetric unit.

Proteomic approaches to Sjögren's syndrome: a clue to interpret the pathophysiology and organ involvement of the disease.

Sjögren's syndrome (SS) is a chronic, inflammatory, autoimmune disease characterized by lymphocytic infiltration of the exocrine glands leading to qualitatively altered and diminished or absent salivary and lachrymal secretion, and by marked B-cell hyperreactivity. Many efforts have been made to define a panel of salivary and lachrymal markers helpful to design diagnostic tests able to replace blood tests and tissue biopsies for the diagnosis of primary and secondary SS. Several proteomic-based studies have indicated that a number of proteins and peptides can be considered SS biomarkers, being 2-3-fold up- or down-regulated compared to normal subject or having an exclusive presence in the saliva or tears of SS patients. Unfortunately, several factors make it difficult to define a comprehensive salivary and lachrymal panel of markers of SS, as the lack of a comprehensive proteomic analysis of human tears and saliva of healthy subjects, the lack of uniform protocols to collect and treat these samples, and the high grade of posttranslational modification of the proteins in these fluids.

Argirose ocular e do sistema lacrimal excretor: relato de caso / Eye's and excretory lacrimal apparatus' argyrosis: case report

No período de março a novembro de 2001, foi diagnosticado um caso de argirose ocular baseando-se na história clínica, no exame biomicroscópico e histopatologia das estruturas oculares. No exame biomicoscópio, observou-se depósitos de pigmentos pardos no estroma corneano e conjuntiva. Na biópsia da conjuntiva, cristalino, canalículo lacrimal superior e saco lacrimal foram encontrados depósitos de prata.

From March to November 2001, a case of eye's argyrosis was diagnosed based on clinical history, eye's biomicroscopy and histopathology. In the biomicroscopy exam, there were gray deposits of pigments in corneal stroma and conjunctiva. In the incisional biopsy of conjuctiva, lens, upper lacrimal canaliculi and lacrimal sac deposits of silver were found.