Search

Purpose: To determine whether unilateral acute anterior uveitis (AAU) induces ipsilateral changes in the tear fluid proteome., Methods: Five patients (25-77 years old) with unilateral AAU were included. Tear fluid samples were obtained using Schirmer's test strips. The healthy eye served as control. Proteins were identified by liquid chromatography tandem mass spectrometry., Results: Two hundred forty-two tear fluid sample proteins were identified, of which 75 were present in at least three patients. Nine proteins were at least 1.5-fold increased, whereas eight were at least 1.5-fold decreased in tears from the diseased eye compared with the healthy eye. APOBEC3A was significantly increased (1.43-fold; P = 0.04), whereas TGM2 was significantly decreased (- 1.21-fold; P = 0.03) in tears from the diseased eye relative to the healthy eye. Ingenuity Pathway Analysis identified LXR/RXR (P < 1.02E-4) as a top canonical pathway., Conclusion: Unilateral AAU induced detectable changes in the ipsilateral tear fluid proteome and involvement of the inflammation-associated LXR/RXR pathway.

Purpose: Transplantation of limbal stem cells is a promising therapy for limbal stem cell deficiency. Limbal cells can be harvested from either a healthy part of the patient's eye or the eye of a donor. Small explants are less likely to inflict injury to the donor site. We investigated the effects of limbal explant size on multiple characteristics known to be important for transplant function., Methods: Human limbal epithelial cells were expanded from large versus small explants (3 versus 1 mm of the corneal circumference) for 3 weeks and characterized by light microscopy, immunohistochemistry, and transmission electron microscopy. Epithelial thickness, stratification, outgrowth, ultrastructure and phenotype were assessed., Results: Epithelial thickness and stratification were similar between the groups. Outgrowth size correlated positively with explant size (r = 0.37; P = 0.01), whereas fold growth correlated negatively with explant size (r = -0.55; P < 0.0001). Percentage of cells expressing the limbal epithelial cell marker K19 was higher in cells derived from large explants (99.1±1.2%) compared to cells derived from small explants (93.2±13.6%, P = 0.024). The percentage of cells expressing ABCG2, integrin β1, p63, and p63α that are markers suggestive of an immature phenotype; Keratin 3, Connexin 43, and E-Cadherin that are markers of differentiation; and Ki67 and PCNA that indicate cell proliferation were equal in both groups. Desmosome and hemidesmosome densities were equal between the groups., Conclusion: For donor- and culture conditions used in the present study, large explants are preferable to small in terms of outgrowth area. As regards limbal epithelial cell thickness, stratification, mechanical strength, and the attainment of a predominantly immature phenotype, both large and small explants are sufficient., Competing Interests: The authors have declared that no competing interests exist.

The purpose of this work was to determine whether the morphology of the oral mucosa epithelium (OME) of patients with xerostomia differ from patients without xerostomia. In total, 34 patients with dry eye disease (DED) with or without xerostomia were examined at The Norwegian Dry Eye Disease Clinic with in vivo confocal microscopy of the lower lip. In addition, age- and gender-matched healthy controls (HC) were included. DED patients with xerostomia had a higher superficial to deep backscatter ratio compared with DED patients without xerostomia (p=0.002) and HC (p=0.001). Regression analysis demonstrated that this ratio was related to xerostomia independently of gender and age (p<0.001). Sensitivity and specificity of detecting xerostomia were 0.78 and 0.85, respectively, when using a superficial to deep backscatter ratio cut-off value of 0.995 (p=0.004). The mean nucleus to cytosol backscatter ratio in the superficial OME was lower in patients with xerostomia than in those without xerostomia (p=0.034). In vivo confocal microscopy is a potential tool for evaluating the oral cavity and to assess changes in the OME associated with xerostomia, objectively and quantitatively. The cause of the increased backscatter in the superficial OME in xerostomia, however, remains to be elucidated.

The purpose of the study was to investigate if xerostomia (dry mouth) is associated with symptoms and signs of dry eye disease (DED). At the Norwegian Dry Eye Clinic, patients with symptomatic DED with different etiologies were consecutively included in the study. The patients underwent a comprehensive ophthalmological work-up and completed self-questionnaires on symptoms of ocular dryness (Ocular Surface Disease Index [OSDI] and McMonnies Dry Eye Questionnaire) and the Sjögren's syndrome (SS) questionnaire (SSQ). Three hundred and eighteen patients (52% women and 48% men) with DED were included. Patient demographics were: 0 to 19 years (1%), 20 to 39 (25%), 40 to 59 (34%), 60 to 79 (35%) and 80 to 99 (5%). Xerostomia, defined as "daily symptoms of dry mouth the last three months" (as presented in SSQ) was reported by 23% of the patients. Female sex was more common among patients with xerostomia (81%) than among non-xerostomia patients (44%; P<0.001). Patients with xerostomia (60 ± 15 years) were older than those without xerostomia (51 ± 17; P<0.001). The use of prescription drugs was more prevalent among xerostomia patients (65%) than among non-xerostomia patients (35%; P<0.021; adjusted for age and sex). Patients with xerostomia had a higher OSDI score (19.0 ± 10.0) than those without xerostomia (12.9 ± 8.0; P<0.001). Moreover, xerostomia patients had more pathological meibum expressibility (0.9 ± 0.7) than those without xerostomia (0.7 ± 0.8; P = 0.046). Comparisons of OSDI and ocular signs were performed after controlling for the effects of sex, age and the number of systemic prescription drugs used. In conclusion, xerostomia patients demonstrated a higher DED symptom load and had poorer meibum expressibility than non-xerostomia patients.

Purpose: Storage of cultured human oral keratinocytes (HOK) allows for transportation of cultured transplants to eye clinics worldwide. In a previous study, one-week storage of cultured HOK was found to be superior with regard to viability and morphology at 12°C compared to 4°C and 37°C. To understand more of how storage temperature affects cell phenotype, gene expression of HOK before and after storage at 4°C, 12°C, and 37°C was assessed., Materials and Methods: Cultured HOK were stored in HEPES- and sodium bicarbonate-buffered Minimum Essential Medium at 4°C, 12°C, and 37°C for one week. Total RNA was isolated and the gene expression profile was determined using DNA microarrays and analyzed with Partek Genomics Suite software and Ingenuity Pathway Analysis. Differentially expressed genes (fold change > 1.5 and P < 0.05) were identified by one-way ANOVA. Key genes were validated using qPCR., Results: Gene expression of cultures stored at 4°C and 12°C clustered close to the unstored control cultures. Cultures stored at 37°C displayed substantial change in gene expression compared to the other groups. In comparison with 12°C, 2,981 genes were differentially expressed at 37°C. In contrast, only 67 genes were differentially expressed between the unstored control and the cells stored at 12°C. The 12°C and 37°C culture groups differed most significantly with regard to the expression of differentiation markers. The Hedgehog signaling pathway was significantly downregulated at 37°C compared to 12°C., Conclusion: HOK cultures stored at 37°C showed considerably larger changes in gene expression compared to unstored cells than cultured HOK stored at 4°C and 12°C. The changes observed at 37°C consisted of differentiation of the cells towards a squamous epithelium-specific phenotype. Storing cultured ocular surface transplants at 37°C is therefore not recommended. This is particularly interesting as 37°C is the standard incubation temperature used for cell culture.

Restoration of the retinal pigment epithelial (RPE) cells to prevent further loss of vision in patients with age-related macular degeneration represents a promising novel treatment modality. Development of RPE transplants, however, requires up to 3 months of cell differentiation. We explored whether the silk protein sericin can induce maturation of primary human retinal pigment epithelial (hRPE) cells. Microarray analysis demonstrated that sericin up-regulated RPE-associated transcripts (RPE65 and CRALBP). Upstream analysis identified the NF-κB pathway as one of the top sericin-induced regulators. ELISA confirmed that sericin stimulates the main NF-κB pathway. Increased levels of RPE-associated proteins (RPE65 and the pigment melanin) in the sericin-supplemented cultures were confirmed by western blot, spectrophotometry and transmission electron microscopy. Sericin also increased cell density and reduced cell death following serum starvation in culture. Inclusion of NF-κB agonists and antagonists in the culture medium showed that activation of the NF-κB pathway appears to be necessary, but not sufficient, for sericin-induced RPE pigmentation. We conclude that sericin promotes pigmentation of cultured primary hRPE cells by activating the main NF-κB pathway. Sericin's potential role in culture protocols for rapid differentiation of hRPE cells derived from embryonic or induced pluripotent stem cells should be investigated.

The purpose of the study was to investigate if the number of goblet cells expanded ex vivo from a conjunctival explant is affected by the biopsy harvesting site on the conjunctiva and the distance from the explant. Conjunctival explants from six regions: superior and inferior bulbus, fornix, and tarsus of male Sprague-Dawley rats were grown in RPMI 1640 with 10% fetal bovine serum on coverslips for eight days. Histochemical and immunofluorescent staining of goblet (CK-7/UEA-1/MUC5AC), stratified squamous, non-goblet (CK-4), proliferating (PCNA) and progenitor (ABCG2) cells were analyzed by epifluorescence and laser confocal microscopy. Outgrowth was measured with NIH ImageJ. For statistical analysis the Mann-Whitney test and Spearman's rank-order correlation test were used. Cultures from superior and inferior fornix contained the most goblet cells as indicated by the presence of CK-7+, UEA-1+ and MUC5AC+ cells. Superior and inferior forniceal cultures displayed 60.8% ± 9.2% and 64.7% ± 6.7% CK-7+ cells, respectively, compared to the superior tarsal (26.6% ± 8.4%; P < 0.05), superior bulbar (31.0% ± 4.0%; P < 0.05), inferior bulbar (38.5% ± 9.3%; P < 0.05) and inferior tarsal cultures (27.7% ± 8.3%; P < 0.05). While 28.4% ± 6.3% of CK-7+ goblet cells co-labeled with PCNA, only 7.4% ± 1.6% of UEA-1+ goblet cells did (P < 0.01). CK-7+ goblet cells were located at a lower concentration close to the explant (39.8% ± 3.1%) compared to near the leading edge (58.2% ± 4.5%; P < 0.05). Both markers for goblet cell secretory product (UEA-1 and MUC5AC), however, displayed the opposite pattern with a higher percentage of positive cells close to the explant than near the leading edge (P < 0.05). The percentage of CK-4+ cells was higher near the explant compared to near the leading edge (P < 0.01). The percentage of CK-7+ goblet cells in the cultures did not correlate with the outgrowth size (r(s) = -0.086; P = 0.435). The percentage of UEA-1+ goblet cells correlated negatively with outgrowth size (r(s) = -0.347; P < 0.01), whereas the percentage of CK-4+ cells correlated positively with the outgrowth size (r(s) = 0.473; P < 0.05). We conclude that forniceal explants yield the highest number of goblet cells ex vivo and thereby seem to be optimal for goblet cell transplantation. We also suggest that CK-7+/UEA-1- cells represent highly proliferative immature goblet cells. These cells could be important during conjunctival migration as they are mostly located close to the leading edge and their density does not decrease with increasing outgrowth size., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Purpose: To evaluate the effect of location and size of biopsy on phenotype and proliferative capacity of cultured rat conjunctival epithelial cells., Methods: Pieces of conjunctiva were used from six areas: superior and inferior areas of bulbus, fornix, and tarsus of male Sprague-Dawley rats (n = 6). Explants were grown in RPMI 1640 with 10% fetal bovine serum on coverslips for 8 days or assayed for colony-forming efficiency (n = 9). Analysis included immunofluorescence microscopy and outgrowth measurements with ImageJ software. The Mann-Whitney test and Spearman's rank-order correlation test were used., Results: Superior (23.9 ± 2.9-fold growth) and inferior (22.4 ± 1.2-fold growth) forniceal tissues yielded significantly more outgrowth with respect to explant size than superior bulbar (13.4 ± 1.9-fold growth; P < 0.05 and P < 0.01, respectively), inferior bulbar (13.6 ± 1.6-fold growth; P = 0.01 and P < 0.01, respectively), and inferior tarsal tissues (14.0 ± 1.3-fold growth; P = 0.01). Outgrowth size correlated positively with explant size (r(s) = 0.54; P < 0.001), whereas explant size correlated negatively with fold growth (r(s) = 0.36; P < 0.001). Superior forniceal cells displayed higher colony-forming efficiency (3.6% ± 0.9%) than superior bulbar (1.1% ± 0.3%; P < 0.05) and inferior bulbar cells (1.6% ± 0.8%; P < 0.05). Percentage of p63+ and PCNA+ cells correlated positively with explant and outgrowth size., Conclusions: Small forniceal conjunctival explants grow the most effectively; however, for transplantation purposes, the loss of p63+ and PCNA+ cells with small explants must be considered.

In Norway there is an ongoing outbreak in pigs of infections with pandemic influenza A(H1N1)v virus. The first herd was confirmed positive on 10 October 2009. As of 26 October, a total of 23 herds have been diagnosed as positive. The majority of the herds seem to have been infected by humans. Sequence analysis of pig viruses from the index farm shows that they are identical or virtually identical to human viruses from the same geographical region.

The authors describe two water-borne outbreaks of Campylobacter gastroenteritis that occurred in central Norway in 1994 and 1995. The epidemics were probably caused by contamination of drinking water by the stools of Pink-footed geese on the way from Svalbard to Germany-Netherlands. Campylobacter jejuni from the stools of the geese was transmitted to the population via untreated drinking water, causing disease in 50% of the population. About 1,000 persons suffered from gastroenteritis caused by contaminated drinking water in these two epidemics.

Purpose: Although less than one-third of anti-nuclear antibody (ANA) positive patients with oJIA develop uveitis, ANA positivity is still the most well-known marker for assessing the risk of uveitis in oligoarticular JIA (oJIA). Therefore, novel biomarkers are needed to better assess the risk of developing uveitis. For this purpose, we performed a comparative tear proteome analysis of uveitis patients to reveal the identity of differentially regulated proteins. Design: Tear samples were collected using the Schirmer strips in 7 oJIA and 7 oJIA patients with uveitis (oJIA-U). All oJIA-U patients had developed bilateral anterior uveitis and were inactive and topical treatment-free. Methods: The nHPLC LC–MS/MS system was used for protein identification and label-free proteome comparisons. The PANTHER and STRING analyses were carried out using UniProt accession numbers of the identified proteins. Results: Patient characteristics, e.g., age, gender, disease duration, and treatments were similar. For protein identification, three different databases were searched. Twenty-two, 147, and 258 database searches, respectively. Of these, 15 were common to all three proteome databases. Of these 15 proteins, 10 proteins were upregulated, and 2 were downregulated, based on the twofold regulation criteria. The upregulated proteins were, namely, cystatin-S, secretoglobin family 1D member, opiorphin prepropeptide, mammaglobin-B, lysozyme C, mesothelin, immunoglobulin kappa constant, extracellular glycoprotein lacritin, beta-2-microglobulin, and immunoglobulin J chain. The downregulated proteins were dermcidin and prolactin-inducible protein. Among the differentially regulated proteins, cystatin-S was the most regulated protein with an 18-fold upregulation ratio in tear samples from uveitis patients. Conclusion: Here, the identities and regulation ratios of several proteins were revealed when tear samples from uveitis patients were compared to patients without uveitis. These proteins are putative biomarkers for assessing uveitis risk and require further attention. [ABSTRACT FROM AUTHOR]

Purpose: Although less than one-third of anti-nuclear antibody (ANA) positive patients with oJIA develop uveitis, ANA positivity is still the most well-known marker for assessing the risk of uveitis in oligoarticular JIA (oJIA). Therefore, novel biomarkers are needed to better assess the risk of developing uveitis. For this purpose, we performed a comparative tear proteome analysis of uveitis patients to reveal the identity of differentially regulated proteins., Design: Tear samples were collected using the Schirmer strips in 7 oJIA and 7 oJIA patients with uveitis (oJIA-U). All oJIA-U patients had developed bilateral anterior uveitis and were inactive and topical treatment-free., Methods: The nHPLC LC-MS/MS system was used for protein identification and label-free proteome comparisons. The PANTHER and STRING analyses were carried out using UniProt accession numbers of the identified proteins., Results: Patient characteristics, e.g., age, gender, disease duration, and treatments were similar. For protein identification, three different databases were searched. Twenty-two, 147, and 258 database searches, respectively. Of these, 15 were common to all three proteome databases. Of these 15 proteins, 10 proteins were upregulated, and 2 were downregulated, based on the twofold regulation criteria. The upregulated proteins were, namely, cystatin-S, secretoglobin family 1D member, opiorphin prepropeptide, mammaglobin-B, lysozyme C, mesothelin, immunoglobulin kappa constant, extracellular glycoprotein lacritin, beta-2-microglobulin, and immunoglobulin J chain. The downregulated proteins were dermcidin and prolactin-inducible protein. Among the differentially regulated proteins, cystatin-S was the most regulated protein with an 18-fold upregulation ratio in tear samples from uveitis patients., Conclusion: Here, the identities and regulation ratios of several proteins were revealed when tear samples from uveitis patients were compared to patients without uveitis. These proteins are putative biomarkers for assessing uveitis risk and require further attention., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Retinoid X receptors (RXRs) present a subgroup of the nuclear receptor superfamily with particularly high evolutionary conservation of ligand binding domain. The receptor exists in α, β, and γ isotypes that form homo-/heterodimeric complexes with other permissive and non-permissive receptors. While research has identified the biochemical roles of several nuclear receptor family members, the roles of RXRs in various neurological disorders remain relatively under-investigated. RXR acts as ligand-regulated transcription factor, modulating the expression of genes that plays a critical role in mediating several developmental, metabolic, and biochemical processes. Cumulative evidence indicates that abnormal RXR signalling affects neuronal stress and neuroinflammatory networks in several neuropathological conditions. Protective effects of targeting RXRs through pharmacological ligands have been established in various cell and animal models of neuronal injury including Alzheimer disease, Parkinson disease, glaucoma, multiple sclerosis, and stroke. This review summarises the existing knowledge about the roles of RXR, its interacting partners, and ligands in CNS disorders. Future research will determine the importance of structural and functional heterogeneity amongst various RXR isotypes as well as elucidate functional links between RXR homo- or heterodimers and specific physiological conditions to increase drug targeting efficiency in pathological conditions. [ABSTRACT FROM AUTHOR]

Background: To identify candidate tear fluid biomarkers in patients with unilateral acute anterior uveitis (AAU) that can aid in the differentiation between these patients and patients with bacterial keratitis or healthy controls. Methods: Thirteen patients (40.1 ± 16.2 years of age) with unilateral AAU, seven patients with unilateral bacterial keratitis (40.2 ± 15.3 years of age), and 14 healthy subjects (41.1 ± 11.6 years of age) were included. The tear proteome of affected eyes was compared with that of the unaffected eye or healthy controls. Proteins were identified by liquid chromatography tandem mass spectrometry and enzyme-linked immunosorbent assay. Results: Relative protein ratios were detected and calculated for 272 unique proteins. Compared with healthy controls and the unaffected eye, the top upregulated proteins in AAU eyes were submaxillary gland androgen regulated protein 3B (SMR3B) and SMR3A. Similarly, the top upregulated proteins in bacterial keratitis were S100 calcium-binding protein A9 and orosomucoid 2. The acute phase response protein Serpin Family A Member 3 (SERPINA3) was increased in the healthy eye of AAU patients (P = 0.019) compared with healthy controls. Laser flare measurements in affected eyes of AAU patients showed positive logarithmic correlation with SERPINA3 in tear samples of the unaffected eye (P = 0.022). The use of SERPINA3 as a tear biomarker yielded a sensitivity of 85% and a specificity of 71% in detecting patients with AAU in the study population. Conclusions: The acute phase response protein SERPINA3 was increased in tear samples of unaffected eyes of patients with unilateral AAU compared with healthy controls. This study highlights SERPINA3 as a potential biomarker for AAU. Future research should explore the dynamic properties of SERPINA3 in the tear fluid of active and quiescent uveitis eyes. [ABSTRACT FROM AUTHOR]

Maintaining mature and viable retinal pigment epithelial cells (RPE) in vitro has proven challenging. Investigating compounds that can promote RPE-viability and maturation is motivated by RPE transplantation research, the quest to understand RPE physiology, and a desire to modulate RPE in pathological states. We have previously reported that the silk protein sericin promotes viability, maturation, and pigmentation of human fetal RPE. In the present study, our aim was to uncover whether these effects can be seen in adult retinal pigment epithelial cell line-19 (ARPE-19) and induced pluripotent stem cell-derived RPE (iPSC-RPE). ARPE-19 and iPSC-RPE were cultured with or without 10 mg/mL sericin. After 7 days, viability was assessed with calcein-acetoxymethyl ester (CAM) and ethidium homodimer-1 (EH-1) assays, flow cytometry, and morphometric analysis. Expression levels of RPE65, tyrosinase, and Pmel17 were quantified to compare maturation between the sericin-treated and control cultures. Light microscopy and staining of the tight junction protein zonula occludens protein 1 (ZO-1) were employed to study sericin's effects on RPE morphology. We also measured culture medium pH, glucose, lactate, and extracellular ion content. Sericin-supplemented RPE cultures demonstrated significantly better viability compared to control cultures. Sericin appeared to improve ARPE-19 maturation and morphology in vitro. No effects were seen on RPE pigmentation with the concentration of sericin and duration of cell culture herein reported. This is the first study to demonstrate that supplementing the culture media with sericin promotes the viability of iPSC-RPE and ARPE-19. Sericin's viability-promoting effects may have important implications for retinal therapeutics and regenerative medicine research. [ABSTRACT FROM AUTHOR]

Purpose: Seeking to improve the access to regenerative medicine, this study investigated the structural and transcriptional effects of storage temperature on human oral mucosal epithelial cells (OMECs). Methods: Cells were stored at four different temperatures (4°C, 12°C, 24°C and 37°C) for two weeks. Then, the morphology, cell viability and differential gene expression were examined using light and scanning electron microscopy, trypan blue exclusion test and TaqMan gene expression array cards, respectively. Results: Cells stored at 4°C had the most similar morphology to non-stored controls with the highest viability rate (58%), whereas the 37°C group was most dissimilar with no living cells. The genes involved in stress-induced growth arrest (GADD45B) and cell proliferation inhibition (TGFB2) were upregulated at 12°C and 24°C. Upregulation was also observed in multifunctional genes responsible for morphology, growth, adhesion and motility such as EFEMP1 (12°C) and EPHA4 (4°C–24°C). Among genes used as differentiation markers, PPARA and TP53 (along with its associated gene CDKN1A) were downregulated in all temperature conditions, whereas KRT1 and KRT10 were either unchanged (4°C) or downregulated (24°C and 12°C; and 24°C, respectively), except for upregulation at 12°C for KRT1. Conclusions: Cells stored at 12°C and 24°C were stressed, although the expression levels of some adhesion-, growth- and apoptosis-related genes were favourable. Collectively, this study suggests that 4°C is the optimal storage temperature for maintenance of structure, viability and function of OMECs after two weeks. [ABSTRACT FROM AUTHOR]

To investigate the feasibility of using Optisol-GS as a convenient, xenogeneic-free alternative for storage of cultured human limbal epithelial cells (HLECS) for use in treatment of limbal stem cell deficiency (LSCD). In the present study, we compared storage of cultured HLEC using the conventional hypothermic Optisol-GS storage method at 4°C versus storage at 23°C (room temperature). HLECs were cultured for three weeks on amniotic membrane (AM), transferred to polypropylene containers and stored in Optisol-GS for 4 days at 23°C and 4°C. A calcein-acetoxymethyl ester/ethidium homodimer-1 assay was used to assess viability. Morphology and phenotype were analyzed by light microscopy and immunohistochemistry, respectively. Expression of stem cell and proliferation markers p63, ∆Np63α, ABCG2, K19, K3, Cx43, Ki67, and PCNA was maintained at pre-storage control levels during storage at 23°C. ABCG2 and PCNA expression were both significantly altered during storage at 4°C. HLEC cell sheet viability also significantly declined following storage at 4°C. HLEC sheets stored at 4°C demonstrated extensive detachment of basal cells from the AM in sharp contrast to storage at 23°C, where attachment to the AM was maintained throughout the storage period. The present study demonstrates the feasibility of short-term storage of cultured HLECs in Optisol-GS, which offers a convenient standardized xenogeneic-free storage method. Storage temperature highly affected the results. Maintenance of cell viability, morphology and undifferentiated proliferative phenotype of cultured HLEC sheets favored storage at 23°C. [ABSTRACT FROM AUTHOR]

The diseases affecting the retina or uvea (iris, ciliary body, or choroid) generate changes in the biochemical or protein composition of ocular fluids/tissues due to disruption of blood-retinal barrier. Ocular infections and inflammations are sight-threatening diseases associated with various infectious and non-infectious etiologies. Several etiological entities cause uveitis, a complex intraocular inflammatory disease. These causes of uveitis differ in different populations due to geographical, racial, and socioeconomic variations. While clinical appearance is sufficiently diagnostic in many diseases, some of the uveitic entities manifest nonspecific or atypical clinical presentation. Identification of biomarkers in such diseases is an important aid in their diagnostic armamentarium. Different diseases and their different severity states release varying concentrations of proteins, which can serve as biomarkers. Proteomics is a high throughput technology and a powerful screening tool for serum biomarkers in various diseases that identifies proteins by mass spectrometry and helps to improve the understanding of pathogenesis of a disease. Proteins determine the biological state of a cell. Once identified as biomarkers, they serve as future diagnostic and pharmaceutical targets. With a potential to redirect the diagnosis of idiopathic uveitis, ocular proteomics provide a new insight into the pathophysiology and therapeutics of various ocular inflammatory diseases. Tears, aqueous and vitreous humor represent potential repositories for proteomic biomarkers discovery in uveitis. With an extensive proteomics work done on animal models of uveitis, various types of human uveitis are being subjected to proteome analysis for biomarker discovery in different ocular fluids (vitreous, aqueous, or tears). [ABSTRACT FROM AUTHOR]

Destruction or dysfunction of limbal epithelial stem cells (LESCs) leads to unilateral or bilateral limbal stem cell deficiency (LSCD). Fifteen years have passed since the first transplantation of ex vivo cultivated oral mucosal epithelial cells (COMET) in humans in 2004, which represents the first use of a cultured non-limbal autologous cell type to treat bilateral LSCD. This review summarizes clinical outcomes from COMET studies published from 2004 to 2019 and reviews results with emphasis on the culture methods by which grafted cell sheets were prepared. [ABSTRACT FROM AUTHOR]

Simple limbal epithelial transplantation (SLET) and cultivated limbal epithelial transplantation (CLET) are proven techniques for treating limbal stem cell deficiency (LSCD). However, the precise regions that are most suitable for preparing explants for transplantation have not been identified conclusively. Accordingly, this in vitro study aimed at determining ideal sites to be selected for tissue harvest for limbal stem cell culture and transplantation. We evaluated cell outgrowth potential and the expression of stem cell markers in cultures from 48 limbal explants from five cadaveric donors. The limbal explants were generated from the three specific sites: Lcor (located innermost and adjacent to the cornea), Lm (middle limbus), and Lconj (located outermost adjacent to the conjunctiva). We found that explants from the Lconj and Lm sites exhibited higher growth potential than those from the Lcor site. Transcript encoding the stem cell marker and p63 isoform, ΔNp63, was detected in cells from Lm and Lconj explants; expression levels were slightly, though significantly (p-value < 0.05), higher in Lm than in Lconj, although expression of ΔNp63α protein was similar in cells from all explants. Differential expression of ATP-Binding Cassette Subfamily G Member 2 (ABCG2) did not reach statistical significance. Immunohistochemistry by indirect immunofluorescence analysis of limbus tissue revealed that the basal layer in explant tissue from Lconj and Lm contained markedly more stem cells than found in Lcor explant tissue; these findings correlate with a higher capacity for growth. Collectively, our findings suggest that explants from the Lconj and Lm sites should be selected for limbal cell expansion for both CLET and SLET procedures. These new insights may guide surgeons toward specific limbal sites that are most suitable for stem cell culture and transplantation and may ultimately improve treatment outcomes in the patients with LSCD. [ABSTRACT FROM AUTHOR]

Previous studies have demonstrated that several types of human stem cells of non-dental origin can be induced to differentiate into enamel-secreting ameloblasts after recombined with mouse embryonic dental mesenchyme. However, the successful rate of ameloblastic differentiation is about rather low, which presents a major obstacle for future stem cell-based whole tooth bioengineering. Previous studies have shown that cultures at reduced temperature could improve the differentiation capability of stem cells in tissue engineering. In this study, we systematically investigated the effects of low temperature on the viability, proliferation and stemness of human keratinocytes stem cells (hKSCs) in cell culture and further examined ameloblastic differentiation of the hKSCs in human–mouse recombinant chimeric tooth germs. Our results demonstrated that low temperature indeed reduces growth rate and maintains healthy undifferentiated morphology of hKSCs without any effects on cell viability. Moreover, examination of stemness makers revealed improved stemness of hKSCs cultured at low temperature with increased expression of stemness markers K15, CD29 and p63 and decreased expression differentiation marker K10, as compared to those cultured at 37 °C. These low temperature treated hKSCs, when recombined with mouse embryonic dental mesenchyme, exhibited significantly increased rate (40%) of ameloblastic differentiation, as compared to that (17%) in tissue recombinants with those hKSCs treated at standard temperature. Our studies demonstrate that low temperature cell culture improves the stemness and plasticity of hKSCs, which in turn enhances ameloblastic differentiation capability of the stem cells in bioengineered teeth. [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis (RA) is a chronic inflammatory disease in which numerous cells and mediators affect inflammatory conditions and disease severity. To compare the serum levels of adiponectin, vitamin D, copper, and zinc in patients with RA and to investigate the relationship between these parameters and RA severity. Ninety patients with RA and 30 healthy controls participated in this cross-sectional case-control study between November 2016 and April 2017; according to the ACR/EULAR criteria for RA. Serum levels of adiponectin were determined by ELISA; copper and zinc by colorimetric spectrophotometry; and vitamin D by HPLC. Kruskal-Wallis and Spearman tests were performed using SPSS software and data were depicted by GraphPad Prism software. Compared with healthy controls, the serum level of adiponectin was significantly increased, whereas vitamin D was significantly decreased in patients with RA. Adiponectin and vitamin D levels were inversely correlated in RA subgroups (P < 0.001, r = − 0.410). Adiponectin and vitamin D correlated with RA severity. Furthermore, no significant difference was found in copper and zinc levels between RA groups and controls. The definitive roles of adiponectin, vitamin D, copper, and zinc are not completely determined in RA development. Based on disease activity, these parameters can modulate inflammatory conditions, thus they have the potential to be used as promising therapeutic biomarkers to follow up the severity of disease, as well as the progression and treatment success in patients with RA. [ABSTRACT FROM AUTHOR]

The demand for novel biocompatible materials as surface coating in the field of regenerative medicine is high. We explored molecular layer deposition (MLD) technique for building surface coatings and introduced a new group of substrates consisting of amino acids, or nucleobases, and the biocompatible metal titanium. The substrates were built from titanium tetraisopropoxide (TTIP) with l‐lysine, glycine, l‐aspartic acid, l‐arginine, thymine, uracil, and adenine. Substrates based on zirconium chloride and terephthalic acid were also included. Titanium oxide (TiO2) substrates made by atomic layer deposition and uncoated cover slips served as controls. Rat conjunctival epithelial goblet cells were grown in RPMI 1640 and RT‐PCR, immunofluorescence, cell attachment, proliferation, and viability were analyzed. Cells cultured on MLD and uncoated substrates were proliferating (positive for Ki67). Cell attachment after 3 h of culture on MLD substrates was similar to uncoated coverslips (p > 0.05). Compared to uncoated coverslips, cell proliferation assayed with alamarBlue® after 4 days was significantly higher on all MLD substrates (p < 0.05), whereas terephthalic acid‐containing MLD substrates reduced proliferation (p < 0.01). Viability assessed by LIVE/DEAD® was high (>85%) for all substrates after 5 days. The novel MLD technique is promising for building biocompatible substrates that direct epithelial cell growth. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 3090–3098, 2018. [ABSTRACT FROM AUTHOR]

Dry eye and dry mouth are typical clinical symptoms of primary Sjögren’s syndrome (pSS), yet not considered in the assessment of severity and predictors in China. This study aimed to investigate the prevalence, severity, and potential predictors of dry eye/dry mouth among Chinese pSS patients. A cross-sectional study was conducted from the Affiliated Hospital of Nantong University. A series of questionnaires were applied: Ocular Surface Disease Index (OSDI), EULAR Sjogren’s Syndrome Patient Reported Index (ESSPRI)-dry mouth items, fatigue severity scale (FSS), the 10 cm visual analog scale (VAS). Laboratory examinations were taken to obtain some biochemical indicators (i.e., C-reactive protein, erythrocyte sedimentation rate, anti-SSA/SSB antibody). Stepwise logistic/linear regression model was used to investigate the potential predictors of dry eye/dry mouth, respectively. Statistical analysis was performed using SPSS version 20.0. Two hundred twenty-four pSS patients were included in this study. Among them, 215 (95.98%) patients reported ESSPRI-dry mouth items score > 0, and the mean score was 4.92 ± 2.43. In addition, according to the score of OSDI, 84 (37.5%) subjects reported non-dry eye, whereas 140 (62.5%) subjects reported dry eye (44 mild, 31 moderate, 65 severe), and the mean of the total OSDI score was 25.01 ± 23.58. Then, using logistic regression and linear regression respectively, we found that age and fatigue were the potential predictors of dry eye, whereas dry mouth was predicted by age, fatigue, total pain, and ESR. The results of this study suggested that rheumatologists should pay attention to pSS patients’ dry eye and dry mouth, especially those with older age, higher level of ESR, more severe fatigue, and pain. [ABSTRACT FROM AUTHOR]

Conjunctival injuries are general but intractable ocular surface diseases, the sequelae of which are particularly challenging to treat. A promising therapy for conjunctival injuries is to employ biodegradable scaffolds to deliver conjunctival epithelial cells for repairing damaged or diseased conjunctiva. In the present study, an ultrathin porous nanofibrous scaffold was fabricated by using collagen and poly(L.lactic acid.co.ƒÃ.caprolactone) (PLCL) and displayed a thickness of 20 ƒÊm, with a high porosity and an average fiber diameter of 248.83±26.44 nm. Conjunctival epithelial cells seeded on the scaffolds proliferated well and had a high cell viability. Reverse.transcription quantitative PCR showed the expression of conjunctival epithelial cell.specific genes; in addition, there was no significant difference in the inflammatory gene expression between cells grown on collagen/PLCL scaffolds and tricalcium phosphate scaffolds. After co.culture for 2 weeks in vitro, epithelial cell stratification was observed using hematoxylin and eosin staining, exhibiting three to four epithelial.cell layers. In conclusion, these results suggested that collagen/PLCL scaffolds have potential application for repairing conjunctival epithelial coloboma. [ABSTRACT FROM AUTHOR]

Copyright of Der Ophthalmologe is the property of Springer Nature and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Purpose: The development of a suitable storage method for retinal pigment epithelium (RPE) is necessary in the establishment of future RPE replacement therapy, and storage temperature has proven to be pivotal for cell survival. ARPE-19, a widely used model for RPE, has been shown to yield the greatest number of viable cells when stored at 16°C compared to other storage temperatures. In this study, we analyze the gene expression profile of cultured ARPE-19 cells after seven days of storage at different temperatures in an effort to predict the gene-level consequences of storage of RPE transplants. Materials and Methods: ARPE-19 cells were cultured until confluence and then stored in minimum essential medium at 4°C, 16°C, and 37°C for seven days. The total RNA was isolated and the gene expression profile was determined using DNA microarrays. The Results were validated using qPCR. Results: Principal component and hierarchical clustering analyses show that the gene expression profiles of cell cultures stored at different temperatures cluster into separate groups. Cultures stored at 4°C cluster closest to the control cultures that were not stored and display the least change in gene expression after storage (157 differentially expressed genes). Cultures stored at 16°C and 37°C display a much larger change in differential gene expression (1787 and 1357 differentially expressed genes, respectively). At 16°C, the expression of several genes with proposed tumor suppressor functions was markedly increased. Changes in regulation of several known signaling pathways and of oxidative stress markers were discovered at both 16°C and 37°C, and activation of the angiogenesis marker vascular endothelial growth factor (VEGF) was discovered at 37°C. There was no evidence of the activation of inflammatory processes in stored cell cultures. Conclusion: ARPE-19 cultures stored at 16°C show the greatest propensity to modulate their gene expression profile in a manner that supports cell survival during storage. [ABSTRACT FROM PUBLISHER]

Conjunctival integrity and preservation is indispensable for vision. The self-renewing capacity of conjunctival cells controls conjunctival homeostasis and regeneration; however, the source of conjunctival self-renewal and the underlying mechanism is currently unclear. Here, we characterize the biochemical phenotype and proliferative potential of conjunctival epithelial cells in adult mouse by detecting proliferation-related signatures and conducting clonal analysis. Further, we show that transcription factor 7-like 2 (T-cell-specific transcription factor 4), a DNA binding protein expressed in multiple types of adult stem cells, is highly correlated with proliferative signatures in basal conjunctival epithelia. Clonal studies demonstrated that Transcription factor 7-like 2 (Tcf7l2) was coexpressed with p63α and proliferating cell nuclear antigen (PCNA) in propagative colonies. Furthermore, Tcf7l2 was actively transcribed concurrently with conjunctival epithelial proliferation in vitro. Collectively, we suggest that Tcf7l2 may be involved in maintenance of stem/progenitor cells properties of conjunctival epithelial stem/progenitor cells, and with the fornix as the optimal site to isolate highly proliferative conjunctival epithelial cells in adult mice. [ABSTRACT FROM AUTHOR]

For a searchable version of these abstracts, please visit . Please Note: It may take several minutes for this file to download. [ABSTRACT FROM AUTHOR]

Introduction: Depletion of limbal stem cells leads to a debilitating condition known as limbal stem cell deficiency, characterised by impaired corneal wound healing and poor vision. The aim of this study was to determine whether delivering progenitor cells on a contact lens is a viable and effective alternative to current transplantation techniques, which are complicated by biological and xenogeneic materials. Methods: Sixteen eyes of 16 patients who had total (n = 14) and partial (n = 2) limbal stem cell deficiency (chemical burns, five eyes; iatrogenic causes, four eyes; aniridia, three eyes; trachoma-induced, two eyes; contact lens over-wear, one eye; and cicatrising conjunctivitis, one eye) and who had failed prior therapy were recruited prospectively into the study. Autologous limbal (n = 7) or conjunctival epithelial (n = 9) biopsies were harvested from patients and placed on the concave surface of silicone hydrogel contact lenses. Cells were expanded in culture with autologous serum and transplanted onto the ocular surface. Results: Restoration of a transparent avascular and clinically stable corneal epithelium was attained in 10 of 16 eyes (63%) at a median follow-up time of 2.5 years (range of 0.8 to 5.8 years). Although minor complications occurred in two eyes of two patients because of contact lens insertion or removal, these were not associated with long-term sequelae. Conclusions: This is the first and largest study to evaluate the mid-term outcomes of autologous limbal/conjunctival stem cell transplantation via a US Food and Drug Administration-approved contact lens, demonstrating that delivery of ocular progenitor cells via this procedure offers a viable, effective, and xeno-free alternative to current transplantation methodologies. [ABSTRACT FROM AUTHOR]

Background: Influenza A(H1N1)pdm09 virus infection in Norwegian pigs was largely subclinical. This study tested the hypothesis that the infection causes negligible impact on pigs’ growth performance in terms of feed conversion efficiency, daily feed intake, daily growth, age on reaching 100 kg bodyweight and overall feed intake. A sample of 1955 pigs originating from 43 breeding herds was classified into five infection status groups; seronegative pigs (n = 887); seropositive pigs (n = 874); pigs positive for virus at bodyweight between 33 kg and 60 kg (n = 123); pigs positive for virus at bodyweight between 61 kg and 80 kg (n = 34) and pigs positive for virus at bodyweight between 81 kg and 100 kg (n = 37). Each pig had daily recordings of feed intake and bodyweight from 33 kg to 100 kg. Marginal effects of the virus infection on the outcomes were estimated by multi-level linear regression, which accounted for known fixed effects (breed, birthdate, average daily feed intake and growth phase) and random effects (cluster effects of pig and herd). Results: The seropositive and virus positive pigs had decreased (P value<0.05) growth performance compared to seronegative pigs even though feed intake was not decreased. Reduced feed conversion efficiency led to lower average daily growth, additional feed requirement and longer time needed to reach the 100 kg bodyweight. The effects were more marked (P value<0.03) in pigs infected at a younger age and lasted a longer period. Despite increased feed intake observed, their growth rates were lower and they took more time to reach 100 kg bodyweight compared to the seronegative pigs. Conclusion: Our study rejected the null hypothesis that the virus infection had negligible adverse effects on growth performance of Norwegian pigs. [ABSTRACT FROM AUTHOR]

Background: Following the emergence of the pandemic H1N1 influenza A virus in 2009 in humans, this novel virus spread into the swine population. Pigs represent a potential host for this virus and can serve as a mixing vessel for genetic mutations of the influenza virus. Reassortant viruses eventually emerged from the 2009 pandemic and were reported in swine populations worldwide including Thailand. As a result of the discovery of this emergent disease, pathogenesis studies of this novel virus were conducted in order that future disease protection and control measures in swine and human populations could be enacted. Methods: The pandemic H1N1 2009 virus (pH1N1) and its reassortant virus (rH1N1) isolated from pigs in Thailand were inoculated into 2 separate cohorts of 9, 3-week-old pigs. Cohorts were consisted of one group experimentally infected with pH1N1 and one group with rH1N1. A negative control group consisting of 3 pigs was also included. Clinical signs, viral shedding and pathological lesions were investigated and compared. Later, 3 pigs from viral inoculated groups and 1 pig from the control group were necropsied at 2, 4, and 12 days post inoculation (DPI). Results: The results indicated that pigs infected with both viruses demonstrated typical flu-like clinical signs and histopathological lesions of varying severity. Influenza infected-pigs of both groups had mild to moderate pulmonary signs on 1-4 DPI. Interestingly, pigs in both groups demonstrated viral RNA detection in the nasal swabs until the end of the experiment (12 DPI). Conclusion: The present study demonstrated that both the pH1N1 and rH1N1 influenza viruses, isolated from naturally infected pigs, induced acute respiratory disease in experimentally inoculated nursery pigs. Although animals in the rH1N1-infected cohort demonstrated more severe clinical signs, had higher numbers of pigs shedding the virus, were noted to have increased histopathological severity of lung lesions and increased viral antigen in lung tissue, the findings were not statistically significant in comparison with the pH1N1-infected group. Interestingly, viral genetic material of both viruses could be detected from the nasal swabs until the end of the experiment. Similar to other swine influenza viruses, the clinical signs and pathological lesions in both rH1N1 and pH1N1 were limited to the respiratory tract. [ABSTRACT FROM AUTHOR]

Background: Limbal stem cells (LSC) sustain the corneal integrity and homeostasis. LSC deficiency (LSCD) leads to loss of corneal transparency and blindness. A clinical approach to treat unilateral LSCD comprises autologous cultured limbal epithelial stem cell transplantation (CLET). CLET uses xenobiotic culture systems with potential zoonotic transmission risks, and regulatory guidelines make necessary to find xenofree alternatives. Methods: We compared two xenofree clinical grade media and two feeder layers. We used CnT07, a defined commercial medium for keratinocytes, and a modified xenofree supplemented hormonal epithelial medium with human serum (XSHEM). Optimal formulation was used to compare two feeder layers: the gold standard 3T3 murine fibroblasts and human processed lipoaspirate cells (PLA). We tested the expressions of ΔNp63α and cytokeratin 3 and 12 by qPCR and immunofluorescence. Morphology, viability, clonogenicity, proliferation, and cell growth assays were carried out. We also evaluated interleukin 6 (IL-6) and stromal-derived factor 1 (SDF-1) by qPCR and ELISA. Results: XSHEM maintained better LSC culture viability and morphology than CnT07. Irradiated PLA feeder cells improved the undifferentiated state of LSC and enhanced their growth and clonogenicity stimulating IL-6 secretion and SDF-1 expression, as well as increased proliferation and cell growth when compared with irradiated 3T3 feeder cells. Conclusions: The combination of XSHEM and PLA feeder cells efficiently sustained LSC xenofree cultures for clinical application. Moreover, PLA feeder layers were able to improve the LSC potential characteristics. Our results would have direct clinical application in CLET for advanced therapy. [ABSTRACT FROM AUTHOR]

Human-induced pluripotent stem cells (hiPSCs) provide a personalized approach to study conditions and diseases including those of the eye that lack appropriate animal models to facilitate the development of novel therapeutics. Corneal disease is one of the most common causes of blindness. Hence, significant efforts are made to develop novel therapeutic approaches including stem cell-derived strategies to replace the diseased or damaged corneal tissues, thus restoring the vision. The use of adult limbal stem cells in the management of corneal conditions has been clinically successful. However, its limited availability and phenotypic plasticity necessitate the need for alternative stem cell sources to manage corneal conditions. Mesenchymal and embryonic stem cell-based approaches are being explored; nevertheless, their limited differentiation potential and ethical concerns have posed a significant hurdle in its clinical use. hiPSCs have emerged to fill these technical and ethical gaps to render clinical utility. In this review, we discuss and summarize protocols that have been devised so far to direct differentiation of human pluripotent stem cells (hPSCs) to different corneal cell phenotypes. With the summarization, our review intends to facilitate an understanding which would allow developing efficient and robust protocols to obtain specific corneal cell phenotype from hPSCs for corneal disease modeling and for the clinics to treat corneal diseases and injury. [ABSTRACT FROM AUTHOR]

There are more herbivorous waterfowl (swans and geese) close to humans, livestock and poultry than ever before. This creates widespread conflict with agriculture and other human interests, but also debate about the role of swans and geese as potential vectors of disease of relevance for human and animal health. Using a One Health perspective, we provide the first comprehensive review of the scientific literature about the most relevant viral, bacterial, and unicellular pathogens occurring in wild geese and swans. Research thus far suggests that these birds may play a role in transmission of avian influenza virus,Salmonella, Campylobacter, and antibiotic resistance. On the other hand, at present there is no evidence that geese and swans play a role in transmission of Newcastle disease, duck plague, West Nile virus,Vibrio, Yersinia, Clostridium, Chlamydophila, andBorrelia. Finally, based on present knowledge it is not possible to say if geese and swans play a role in transmission ofEscherichia coli, Pasteurella, Helicobacter, Brachyspira, Cryptosporidium, Giardia, and Microsporidia. This is largely due to changes in classification and taxonomy, rapid development of identification methods and lack of knowledge about host specificity. Previous research tends to overrate the role of geese and swans as disease vectors; we do not find any evidence that they are significant transmitters to humans or livestock of any of the pathogens considered in this review. Nevertheless, it is wise to keep poultry and livestock separated from small volume waters used by many wild waterfowl, but there is no need to discourage livestock grazing in nature reserves or pastures where geese and swans are present. Under some circumstances it is warranted to discourage swans and geese from using wastewater ponds, drinking water reservoirs, and public beaches. Intensified screening of swans and geese for AIV, West Nile virus and anatid herpesvirus is warranted. [ABSTRACT FROM PUBLISHER]