| 1. |
- Björkblom, Benny, et al.
(författare)
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Xenobiotic- and Serum-Free Culture of Oral Mucosal Epithelial Cells on Contact Lenses
- 2016
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Ingår i: Current Eye Research. - : Informa UK Limited. - 0271-3683 .- 1460-2202. ; 41:1, s. 20-27
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Tidskriftsartikel (refereegranskat)abstract
- Purpose/aim: Cultured autologous oral mucosal epithelial cells (OMECs) have proven useful in the treatment of ocular surface disorders. This study is the first to investigate the potential of expanding OMEC in a xenobiotic- and serum-free medium using therapeutic contact lenses (CLs) as a substrate and carrier.Materials and methods: Porcine OMEC were seeded on laminin-coated lotrafilcon A therapeutic CLs with the density of 8x10(4)cells/lens and cultured in a defined serum and xenobiotic-free medium. Confocal immunofluorescence microscopy was used to analyze the following: (1) cellular morphology by using rhodamine-phalloidin staining of F-actin, (2) phenotype by applying antibodies against the progenitor cell marker p63 and the putative stem cell marker ABCG2 and (3) cell viability by using propidium iodide and Hoechst 33342 dual staining.Results: Porcine OMEC attached well to the CLs, and cell-to-cell contacts were evident. After three days in culture, the OMEC displayed a confluent monolayer with uniform cobblestone morphology, whereas stratified cultures with 2-3 layers were formed after six days. No significant difference in expression of p63 was observed after three-day culture (79.414.8%) compared with six-day culture (60.3 +/- 18.9%). ABCG2 expression in the basal cell layer was 6.3 +/- 1.0% and 4.8 +/- 1.8% after three- and six-day culture, respectively. The basal layer viability of cultured OMECs was 99.3 +/- 0.2% and 82.8 +/- 1.1% after three and six days culture, respectively.Conclusions: The use of therapeutic CLs has potential as a substrate and carrier for OMEC cultured in a xenobiotic- and serum-free culture system.
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| 2. |
- Fostad, Ida G., et al.
(författare)
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Dry Eye Disease Patients with Xerostomia Report Higher Symptom Load and Have Poorer Meibum Expressibility
- 2016
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Ingår i: PLOS ONE. - : PUBLIC LIBRARY SCIENCE. - 1932-6203. ; 11:5, s. e0155214-
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Tidskriftsartikel (refereegranskat)abstract
- 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 Sjogrens 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%; Pamp;lt; 0.001). Patients with xerostomia (60 +/- 15 years) were older than those without xerostomia (51 +/- 17; Pamp;lt; 0.001). The use of prescription drugs was more prevalent among xerostomia patients (65%) than among non-xerostomia patients (35%; Pamp;lt; 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; Pamp;lt; 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.
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| 3. |
- Fostad, Ida G., et al.
(författare)
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Identification of Objective Morphometric Markers of Xerostomia in the Oral Mucosa Epithelium with In Vivo Confocal Microscopy
- 2017
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Ingår i: Microscopy and Microanalysis. - : CAMBRIDGE UNIV PRESS. - 1431-9276 .- 1435-8115. ; 23:1, s. 88-96
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Tidskriftsartikel (refereegranskat)abstract
- 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 (pamp;lt;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.
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| 4. |
- Islam, Rakibul, et al.
(författare)
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Effect of Storage Temperature on Structure and Function of Cultured Human Oral Keratinocytes
- 2015
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Ingår i: PLOS ONE. - : Public Library of Science. - 1932-6203. ; 10:6, s. e0128306-
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Tidskriftsartikel (refereegranskat)abstract
- Purpose/Aims To assess the effect of storage temperature on the viability, phenotype, metabolism, and morphology of cultured human oral keratinocytes (HOK). Materials and Methods Cultured HOK cells were stored in HEPES- and sodium bicarbonate-buffered Minimum Essential Medium (MEM) at nine temperatures in approximately 4 degrees C increments from 4 degrees C to 37 degrees C for seven days. Cells were characterized for viability by calcein fluorescence, phenotype retention by immunocytochemistry, metabolic parameters (pH, glucose, lactate, and O-2) within the storage medium by blood gas analysis, and morphology by scanning electron microscopy and light microscopy. Results Relative to the cultured, but non-stored control cells, a high percentage of viable cells were retained only in the 12 degrees C and 16 degrees C storage groups (85%+/- 13% and 68%+/- 10%, respectively). Expression of ABCG2, Bmi1, C/EBP delta, PCNA, cytokeratin 18, and caspase-3 were preserved after storage in the 5 groups between 4 degrees C and 20 degrees C, compared to the non-stored control. Glucose, pH and pO(2) in the storage medium declined, whereas lactate increased with increasing storage temperature. Morphology was best preserved following storage of the three groups between 12 degrees C, 16 degrees C, and 20 degrees C. Conclusion We conclude that storage temperatures of 12 degrees C and 16 degrees C were optimal for maintenance of cell viability, phenotype, and morphology of cultured HOK. The storage method described in the present study may be applicable for other cell types and tissues; thus its significance may extend beyond HOK and the field of ophthalmology.
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| 5. |
- Jackson, Catherine, et al.
(författare)
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Effect of Storage Temperature on Cultured Epidermal Cell Sheets Stored in Xenobiotic-Free Medium
- 2014
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 9:8, s. e105808-
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Tidskriftsartikel (refereegranskat)abstract
- Cultured epidermal cell sheets (CECS) are used in regenerative medicine in patients with burns, and have potential to treat limbal stem cell deficiency (LSCD), as demonstrated in animal models. Despite widespread use, short-term storage options for CECS are limited. Advantages of storage include: flexibility in scheduling surgery, reserve sheets for repeat operations, more opportunity for quality control, and improved transportation to allow wider distribution. Studies on storage of CECS have thus far focused on cryopreservation, whereas refrigeration is a convenient method commonly used for whole skin graft storage in burns clinics. It has been shown that preservation of viable cells using these methods is variable. This study evaluated the effect of different temperatures spanning 4 degrees C to 37 degrees C, on the cell viability, morphology, proliferation and metabolic status of CECS stored over a two week period in a xenobiotic-free system. Compared to non-stored control, best cell viability was obtained at 24 degrees C (95.2 +/- 9.9%); reduced cell viability, at approximately 60%, was demonstrated at several of the temperatures (12 degrees C, 28 degrees C, 32 degrees C and 37 degrees C). Metabolic activity was significantly higher between 24 degrees C and 37 degrees C, where glucose, lactate, lactate/glucose ratios, and oxygen tension indicated increased activation of the glycolytic pathway under aerobic conditions. Preservation of morphology as shown by phase contrast and scanning electron micrographs was best at 12 degrees C and 16 degrees C. PCNA immunocytochemistry indicated that only 12 degrees C and 20 degrees C allowed maintenance of proliferative function at a similar level to non-stored control. In conclusion, results indicate that 12 degrees C and 24 degrees C merit further investigation as the prospective optimum temperature for short-term storage of cultured epidermal cell sheets.
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| 6. |
- Paaske Utheim, Tor, et al.
(författare)
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Storage Temperature Alters the Expression of Differentiation-Related Genes in Cultured Oral Keratinocytes
- 2016
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Ingår i: PLOS ONE. - : PUBLIC LIBRARY SCIENCE. - 1932-6203. ; 11:3, s. e0152526-
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Tidskriftsartikel (refereegranskat)abstract
- 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 degrees C compared to 4 degrees C and 37 degrees C. To understand more of how storage temperature affects cell phenotype, gene expression of HOK before and after storage at 4 degrees C, 12 degrees C, and 37 degrees C was assessed. Materials and Methods Cultured HOK were stored in HEPES-and sodium bicarbonate-buffered Minimum Essential Medium at 4 degrees C, 12 degrees C, and 37 degrees 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 degrees C and 12 degrees C clustered close to the unstored control cultures. Cultures stored at 37 degrees C displayed substantial change in gene expression compared to the other groups. In comparison with 12 degrees C, 2,981 genes were differentially expressed at 37 degrees C. In contrast, only 67 genes were differentially expressed between the unstored control and the cells stored at 12 degrees C. The 12 degrees C and 37 degrees C culture groups differed most significantly with regard to the expression of differentiation markers. The Hedgehog signaling pathway was significantly downregulated at 37 degrees C compared to 12 degrees C. Conclusion HOK cultures stored at 37 degrees C showed considerably larger changes in gene expression compared to unstored cells than cultured HOK stored at 4 degrees C and 12 degrees C. The changes observed at 37 degrees C consisted of differentiation of the cells towards a squamous epithelium-specific phenotype. Storing cultured ocular surface transplants at 37 degrees C is therefore not recommended. This is particularly interesting as 37 degrees C is the standard incubation temperature used for cell culture.
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