AIM： To analyze the concentration-dependent effects of autologous serum （AS） and fetal bovine serum （FBS） on human corneal epithelial cell （HCEC） viability, migration and proliferation. METHODS： AS was prepar...AIM： To analyze the concentration-dependent effects of autologous serum （AS） and fetal bovine serum （FBS） on human corneal epithelial cell （HCEC） viability, migration and proliferation. METHODS： AS was prepared from 13 patients with non- healing epithelial defects Dulbecco＇s modified eagle medium/ Ham＇s F12 （DMEM/F12） with 5% FBS, 0.5% dimethyl sulphoxide （DMSO）, 10 ng/mL human epidermal growth factor, 1% insulin-transferrin-selenium, then were incubated in serum media： DMEM/F12 supplemented by 5%, 10%, 15% or 30% AS or FBS. HCEC viability was analyzed using cell proliferation kit XTI＇, migration using a wound healing assay, proliferation by the cell proliferation enzyme-linked immunosorbent assay （ELISA） BrdU kit. Statistical analysis was performed using the generalized linear model, the values at 30% AS or 30% FBS were used as the baselines. RESULTS： HCEC viability was the highest at 30% AS or 15% FBS and the lowest at 10% AS or 30% FBS application. HCEC migration was the quickest through 30% AS or 30% FBS and the slowest through 5% AS or 5% FBS concentrations. Proliferation was the most increased through 15% AS or 5% FBS and the least increased through 30% AS or 30% FBS concentrations. HCEC viability at 10% and 15% AS was significantly worse （P=0.001, P=0.023） compared to baseline and significantly better at 15% FBS （P=0.003） concentrations. HCEC migration was significantly worse （P〈0.007） and HCEC proliferation significantly better （P〈0.001） in all concentration groups compared to baseline. CONCLUSION： For the best viability of HCEC 30% AS or 15% FBS, for HCEC migration 30% AS or 30% FBS, for proliferation 15% AS or 5% FBS should be used. Therefore, we suggest the use of 30% AS in clinical practice.展开更多
Riboflavin-UVA photodynamic inactivation is a potential treatment altemative in therapy resistant infectious keratitis. The purpose of our study was to determine the impact of riboflavin-UVA photodynamic inactivation ...Riboflavin-UVA photodynamic inactivation is a potential treatment altemative in therapy resistant infectious keratitis. The purpose of our study was to determine the impact of riboflavin-UVA photodynamic inactivation on viability, apop- tosis and activation of human keratocytes in vitro. Primary human keratocytes were isolated from human corneal buttons and cultured in DMEM/Ham＇s F12 medium supplemented with 10% fetal calf serum. Keratocytes underwent UVA light illumination （375 nm） for 4.10 minutes （2 J/cm2） during exposure to different concentrations of riboflavin. Twenty-four hours after treatment, cell viability was evaluated photometrically, whereas apoptosis, CD34 and alpha-smooth muscle actin （α-SMA） expression were assessed using flow cytometry. We did not detect significant changes in cell viability, apoptosis, CD34 and α-SMA expression in groups only treated with riboflavin or UVA light. In the group treated with riboflavin-UVA-photodynamic inactivation, viability of keratocytes decreased significantly at 0.1% riboflavin （P〈0.01） while the percentage of CD34 （P〈0.01 for both 0.05% and 0.1% riboflavin） and alpha-SMA positive keratocytes （P〈0.01 and P〈0.05 for 0.05% and 0.1% riboflavin, respectively） increased significantly compared to the controls. There was no significant change in the percentage of apoptotic keratocytes compared to controls at any of the used ribo- flavin concentrations （P=0.09 and P=0.13）. We concluded that riboflavin-UVA-photodynamic-inactivation decreases viability of myofibroblastic transformation and multipotent haematopoietic stem cell transformation; however, it does not have an impact on apoptosis of human keratocytes in vitro.展开更多
AIM： To determine the impact of biometric characteristics on the biomechanical properties of the human cornea using the ocular response analyzer （ORA） and standard comprehensive ophthalmic examinations before and a...AIM： To determine the impact of biometric characteristics on the biomechanical properties of the human cornea using the ocular response analyzer （ORA） and standard comprehensive ophthalmic examinations before and after standard phacoemulsificaUon. METHODS： This study comprised 54 eyes with cataract with significant lens opacification in stages I or II that underwent phacoemulsification （2.8 mm incision）. Corneal hysteresis （CH）, corneal resistance factor （CRF）, Goldmann-correlated intraocular pressure （IOPg）, and corneal-compensated intraocular pressure （IOPcc） were measured by ORA preoperatively and at lmo postoperatively. Biometric characteristics were derived from corneal topography [TMS-5, anterior equivalent （EQTMS） and cylindric （CYLTMS） power], corneal tomography [Casia, anterior and posterior equivalent （EQaCASC, EQpCASIA） and cylindric （CYLaCASIA, CYLpCASIA） power], keratometry [IOLMaster, anterior equivalent （EQIOL） and cylindric （CYL,oL） power] and autorefractor [anterior equivalent （EQAR）]. Results from ORA were analyzed and correlated with those from all other examinations taken at the same time point. RESULTS： Preoperatively, CH correlated with EQpCASIA and CYLpCASIA only （P=0.001, P=0.002）. Postoperatively, IOPg and IOPcc correlated with all equivalent powers （EQTMS, EQIOL, EQAR, EQaCASIA, and EQpCASIA）（P=0.001, P=0.007, P=0.001, P= 0.015, P=0.03 for IOPg and P〈0.001, ,0=0.003, P〈0.001, P= 0.009, P=0.014 for IOPcc）. CH correlated postoperatively with EQaCASIA and EQpCASIA only （P=0.021, P=0.022）. CONCLUSION： Biometric characteristics may significantly affect biomechanical properties of the cornea in terms of CH, IOPcc and IOPg before, but even more after cataract surgery.展开更多
We analysed histologically two Acanthamoeba keratitis(AK) eyes with anterior and posterior segment inflammation and blindness. Two enucleated eyes of 2 patients(age 45 and 51y) with AK(PCR of epithelial abrasion posit...We analysed histologically two Acanthamoeba keratitis(AK) eyes with anterior and posterior segment inflammation and blindness. Two enucleated eyes of 2 patients(age 45 and 51y) with AK(PCR of epithelial abrasion positive) were analysed. Histological analysis was performed using hematoxylin-eosin, periodic acid-Schiff and G?m?rimethenamine silver staining. We could not observe Acanthamoeba trophozoites or cysts neither in the cornea nor in other ocular tissues. Meanwhile, we found uveitis, retinal vasculitis and scleritis in these eyes, due to the long-standing, recalcitrant AK. So in this stage of AK, systemic immune suppression may be necessary for a longer time period.展开更多
AIM:To analyse the macular vascularisation changes analysed using optical coherence tomography angiography(OCTA)after successful rhegmatogenous retinal detachment(RRD)repair by comparing gas vs silicone oil and macula...AIM:To analyse the macular vascularisation changes analysed using optical coherence tomography angiography(OCTA)after successful rhegmatogenous retinal detachment(RRD)repair by comparing gas vs silicone oil and macula-on vs macula-off.METHODS:This retrospective data collection included 77 eyes with RRD that underwent pars plana vitrectomy(PPV)and gas or silicone oil tamponade.We performed an OCTA during the postoperative control between 6 and 24mo after the last surgery and evaluated the main parameters measured by OCTA:foveal avascular zone(FAZ)and parafoveolar vascular density(PVD)in the superficial capillary plexus.The patients were divided into four groups:RRD with macular involvement treated with gas tamponade,RRD without macular involvement treated with gas tamponade,RRD with macular involvement treated with silicone oil tamponade and RRD without macular involvement treated with silicone oil tamponade.A one-way ANOVA test combined with post hoc Bonferroni corrections compared FAZ sizes and PVD in all four groups.RESULTS:The FAZ size was statistically significantly larger in eyes with RRD involving the macula than in those not involving it(P=0.005).There was no statistically significant difference in the FAZ sizes of the eyes treated with silicone oil tamponade compared to those treated with gas tamponade(P=0.54).There was no statistically significant difference in the PVD comparing all four groups.CONCLUSION:Despite the known risks associated with silicone oil,our findings suggest that the type of tamponade used during PPV to treat an RRD has no significant effect on the future integrity of the PVD or the size of the FAZ in the superficial capillary plexus as measured by OCTA.展开更多
AIM： To evaluate the anterior and posterior corneal surfaces using scanning-slit topography and to determine the diagnostic ability of the measured corneal parameters in keratoconus.METHODS： Orbscan II measurements ...AIM： To evaluate the anterior and posterior corneal surfaces using scanning-slit topography and to determine the diagnostic ability of the measured corneal parameters in keratoconus.METHODS： Orbscan II measurements were taken in 39 keratoconic corneas previously diagnosed by corneal topography and in 39 healthy eyes. The central minimum, maximum, and astigmatic simulated keratometry（K） and anterior axial power values were determined. Spherical and cylindrical mean power diopters were obtained at the central and at the steepest point of the cornea both on anterior and on posterior mean power maps. Pachymetry evaluations were taken at the center and paracentrally in the 3 mm zone from the center at a location of every 45 degrees. Receiver operating characteristic（ROC） analysis was used to determine the best cut-off values and to evaluate the utility of the measured parameters in identifying patients with keratoconus.RESULTS： The minimum, maximum and astigmatic simulated K readings were 44.80±3.06 D, 47.17±3.67 D and 2.42±1.84 D respectively in keratoconus patients and these values differed significantly（P〈0.0001 for all comparisons） from healthy subjects. For all pachymetry measurements and for anterior and posterior mean power values significant differences were found between the two groups. Moreover, anterior central cylindrical power had the best discrimination ability（area under the ROC curve=0.948）.CONCLUSION： The results suggest that scanning-slit topography and pachymetry are accurate methods both for keratoconus screening and for confirmation of the diagnosis.展开更多
High-quality DNA extraction is a crucial step in metagenomic studies.Bias by different isolation kits impairs the comparison across datasets.A trending topic is,however,the analysis of multiple metagenomes from the sa...High-quality DNA extraction is a crucial step in metagenomic studies.Bias by different isolation kits impairs the comparison across datasets.A trending topic is,however,the analysis of multiple metagenomes from the same patients to draw a holistic picture of microbiota associated with diseases.We thus collected bile,stool,saliva,plaque,sputum,and conjunctival swab samples and performed DNA extraction with three commercial kits.For each combination of the specimen type and DNA extraction kit,20-gigabase(Gb)metagenomic data were generated using short-read sequencing.While profiles of the specimen types showed close proximity to each other,we observed notable differences in the alpha diversity and composition of the microbiota depending on the DNA extraction kits.No kit outperformed all selected kits on every specimen.We reached consistently good results using the Qiagen QiAamp DNA Microbiome Kit.Depending on the specimen,our data indicate that over 10 Gb of sequencing data are required to achieve sufficient resolution,but DNA-based identification is superior to identification by mass spectrometry.Finally,longread nanopore sequencing confirmed the results(correlation coefficient>0.98).Our results thus suggest using a strategy with only one kit for studies aiming for a direct comparison of multiple microbiotas from the same patients.展开更多
文摘AIM： To analyze the concentration-dependent effects of autologous serum （AS） and fetal bovine serum （FBS） on human corneal epithelial cell （HCEC） viability, migration and proliferation. METHODS： AS was prepared from 13 patients with non- healing epithelial defects Dulbecco＇s modified eagle medium/ Ham＇s F12 （DMEM/F12） with 5% FBS, 0.5% dimethyl sulphoxide （DMSO）, 10 ng/mL human epidermal growth factor, 1% insulin-transferrin-selenium, then were incubated in serum media： DMEM/F12 supplemented by 5%, 10%, 15% or 30% AS or FBS. HCEC viability was analyzed using cell proliferation kit XTI＇, migration using a wound healing assay, proliferation by the cell proliferation enzyme-linked immunosorbent assay （ELISA） BrdU kit. Statistical analysis was performed using the generalized linear model, the values at 30% AS or 30% FBS were used as the baselines. RESULTS： HCEC viability was the highest at 30% AS or 15% FBS and the lowest at 10% AS or 30% FBS application. HCEC migration was the quickest through 30% AS or 30% FBS and the slowest through 5% AS or 5% FBS concentrations. Proliferation was the most increased through 15% AS or 5% FBS and the least increased through 30% AS or 30% FBS concentrations. HCEC viability at 10% and 15% AS was significantly worse （P=0.001, P=0.023） compared to baseline and significantly better at 15% FBS （P=0.003） concentrations. HCEC migration was significantly worse （P〈0.007） and HCEC proliferation significantly better （P〈0.001） in all concentration groups compared to baseline. CONCLUSION： For the best viability of HCEC 30% AS or 15% FBS, for HCEC migration 30% AS or 30% FBS, for proliferation 15% AS or 5% FBS should be used. Therefore, we suggest the use of 30% AS in clinical practice.
基金supports of the China Scholarship Council(CSC)for the author's study(J Wang and X Song)the Alexander von Humboldt Foundation for supporting the work of Dr.N.Szentmary at the Department of Ophthalmology of Saarland University,Homburg/Saar,Germanysupported by"Zentrales Innovationsprogram Mittelstand(ZIM)"of the German Federal Ministry of Economics and Technology(Project number:KF2152004MD0)
文摘Riboflavin-UVA photodynamic inactivation is a potential treatment altemative in therapy resistant infectious keratitis. The purpose of our study was to determine the impact of riboflavin-UVA photodynamic inactivation on viability, apop- tosis and activation of human keratocytes in vitro. Primary human keratocytes were isolated from human corneal buttons and cultured in DMEM/Ham＇s F12 medium supplemented with 10% fetal calf serum. Keratocytes underwent UVA light illumination （375 nm） for 4.10 minutes （2 J/cm2） during exposure to different concentrations of riboflavin. Twenty-four hours after treatment, cell viability was evaluated photometrically, whereas apoptosis, CD34 and alpha-smooth muscle actin （α-SMA） expression were assessed using flow cytometry. We did not detect significant changes in cell viability, apoptosis, CD34 and α-SMA expression in groups only treated with riboflavin or UVA light. In the group treated with riboflavin-UVA-photodynamic inactivation, viability of keratocytes decreased significantly at 0.1% riboflavin （P〈0.01） while the percentage of CD34 （P〈0.01 for both 0.05% and 0.1% riboflavin） and alpha-SMA positive keratocytes （P〈0.01 and P〈0.05 for 0.05% and 0.1% riboflavin, respectively） increased significantly compared to the controls. There was no significant change in the percentage of apoptotic keratocytes compared to controls at any of the used ribo- flavin concentrations （P=0.09 and P=0.13）. We concluded that riboflavin-UVA-photodynamic-inactivation decreases viability of myofibroblastic transformation and multipotent haematopoietic stem cell transformation; however, it does not have an impact on apoptosis of human keratocytes in vitro.
文摘AIM： To determine the impact of biometric characteristics on the biomechanical properties of the human cornea using the ocular response analyzer （ORA） and standard comprehensive ophthalmic examinations before and after standard phacoemulsificaUon. METHODS： This study comprised 54 eyes with cataract with significant lens opacification in stages I or II that underwent phacoemulsification （2.8 mm incision）. Corneal hysteresis （CH）, corneal resistance factor （CRF）, Goldmann-correlated intraocular pressure （IOPg）, and corneal-compensated intraocular pressure （IOPcc） were measured by ORA preoperatively and at lmo postoperatively. Biometric characteristics were derived from corneal topography [TMS-5, anterior equivalent （EQTMS） and cylindric （CYLTMS） power], corneal tomography [Casia, anterior and posterior equivalent （EQaCASC, EQpCASIA） and cylindric （CYLaCASIA, CYLpCASIA） power], keratometry [IOLMaster, anterior equivalent （EQIOL） and cylindric （CYL,oL） power] and autorefractor [anterior equivalent （EQAR）]. Results from ORA were analyzed and correlated with those from all other examinations taken at the same time point. RESULTS： Preoperatively, CH correlated with EQpCASIA and CYLpCASIA only （P=0.001, P=0.002）. Postoperatively, IOPg and IOPcc correlated with all equivalent powers （EQTMS, EQIOL, EQAR, EQaCASIA, and EQpCASIA）（P=0.001, P=0.007, P=0.001, P= 0.015, P=0.03 for IOPg and P〈0.001, ,0=0.003, P〈0.001, P= 0.009, P=0.014 for IOPcc）. CH correlated postoperatively with EQaCASIA and EQpCASIA only （P=0.021, P=0.022）. CONCLUSION： Biometric characteristics may significantly affect biomechanical properties of the cornea in terms of CH, IOPcc and IOPg before, but even more after cataract surgery.
文摘We analysed histologically two Acanthamoeba keratitis(AK) eyes with anterior and posterior segment inflammation and blindness. Two enucleated eyes of 2 patients(age 45 and 51y) with AK(PCR of epithelial abrasion positive) were analysed. Histological analysis was performed using hematoxylin-eosin, periodic acid-Schiff and G?m?rimethenamine silver staining. We could not observe Acanthamoeba trophozoites or cysts neither in the cornea nor in other ocular tissues. Meanwhile, we found uveitis, retinal vasculitis and scleritis in these eyes, due to the long-standing, recalcitrant AK. So in this stage of AK, systemic immune suppression may be necessary for a longer time period.
文摘AIM:To analyse the macular vascularisation changes analysed using optical coherence tomography angiography(OCTA)after successful rhegmatogenous retinal detachment(RRD)repair by comparing gas vs silicone oil and macula-on vs macula-off.METHODS:This retrospective data collection included 77 eyes with RRD that underwent pars plana vitrectomy(PPV)and gas or silicone oil tamponade.We performed an OCTA during the postoperative control between 6 and 24mo after the last surgery and evaluated the main parameters measured by OCTA:foveal avascular zone(FAZ)and parafoveolar vascular density(PVD)in the superficial capillary plexus.The patients were divided into four groups:RRD with macular involvement treated with gas tamponade,RRD without macular involvement treated with gas tamponade,RRD with macular involvement treated with silicone oil tamponade and RRD without macular involvement treated with silicone oil tamponade.A one-way ANOVA test combined with post hoc Bonferroni corrections compared FAZ sizes and PVD in all four groups.RESULTS:The FAZ size was statistically significantly larger in eyes with RRD involving the macula than in those not involving it(P=0.005).There was no statistically significant difference in the FAZ sizes of the eyes treated with silicone oil tamponade compared to those treated with gas tamponade(P=0.54).There was no statistically significant difference in the PVD comparing all four groups.CONCLUSION:Despite the known risks associated with silicone oil,our findings suggest that the type of tamponade used during PPV to treat an RRD has no significant effect on the future integrity of the PVD or the size of the FAZ in the superficial capillary plexus as measured by OCTA.
文摘AIM： To evaluate the anterior and posterior corneal surfaces using scanning-slit topography and to determine the diagnostic ability of the measured corneal parameters in keratoconus.METHODS： Orbscan II measurements were taken in 39 keratoconic corneas previously diagnosed by corneal topography and in 39 healthy eyes. The central minimum, maximum, and astigmatic simulated keratometry（K） and anterior axial power values were determined. Spherical and cylindrical mean power diopters were obtained at the central and at the steepest point of the cornea both on anterior and on posterior mean power maps. Pachymetry evaluations were taken at the center and paracentrally in the 3 mm zone from the center at a location of every 45 degrees. Receiver operating characteristic（ROC） analysis was used to determine the best cut-off values and to evaluate the utility of the measured parameters in identifying patients with keratoconus.RESULTS： The minimum, maximum and astigmatic simulated K readings were 44.80±3.06 D, 47.17±3.67 D and 2.42±1.84 D respectively in keratoconus patients and these values differed significantly（P〈0.0001 for all comparisons） from healthy subjects. For all pachymetry measurements and for anterior and posterior mean power values significant differences were found between the two groups. Moreover, anterior central cylindrical power had the best discrimination ability（area under the ROC curve=0.948）.CONCLUSION： The results suggest that scanning-slit topography and pachymetry are accurate methods both for keratoconus screening and for confirmation of the diagnosis.
文摘High-quality DNA extraction is a crucial step in metagenomic studies.Bias by different isolation kits impairs the comparison across datasets.A trending topic is,however,the analysis of multiple metagenomes from the same patients to draw a holistic picture of microbiota associated with diseases.We thus collected bile,stool,saliva,plaque,sputum,and conjunctival swab samples and performed DNA extraction with three commercial kits.For each combination of the specimen type and DNA extraction kit,20-gigabase(Gb)metagenomic data were generated using short-read sequencing.While profiles of the specimen types showed close proximity to each other,we observed notable differences in the alpha diversity and composition of the microbiota depending on the DNA extraction kits.No kit outperformed all selected kits on every specimen.We reached consistently good results using the Qiagen QiAamp DNA Microbiome Kit.Depending on the specimen,our data indicate that over 10 Gb of sequencing data are required to achieve sufficient resolution,but DNA-based identification is superior to identification by mass spectrometry.Finally,longread nanopore sequencing confirmed the results(correlation coefficient>0.98).Our results thus suggest using a strategy with only one kit for studies aiming for a direct comparison of multiple microbiotas from the same patients.
