Human retinal Müller cells synthesize collagens of the vitreous and vitreoretinal interface in vitro

Purpose: To investigate the capacity of cultured Muller cells to synthesize collagens, since previous studies indicated that Muller cells could be involved in collagen remodeling at the vitreoretinal border in adult human eyes. Methods: Spontaneously immortalized cultured human Muller cells were analyzed for the presence of mRNA of types I-VII, IX, XI, and XVII collagen by RT-PCR. Furthermore, Muller cells were immunocytochemically stained for light microscopic (LM) evaluation of these collagens and their main characteristics. Finally, cell extracts and culture medium were evaluated by western blot (WB) analysis using anticollagen antibodies. Results: Cultured Muller cells contained mRNA for types I-VII, IX, and XI collagen, but not for type XVII collagen. LM and WB confirmed the intracellular expression of all the above-mentioned collagens with the exception of type XVII. Collagen secretion into the medium was established for types I-VII, IX, and XI collagen. Conclusions: Cultured Muller cells can synthesize internal limiting lamina and vitreous collagens. Possible collagen production by Muller cells could explain and expand on previous in vivo morphological findings in the embryonic and postnatal period and in pathologic conditions

Central retinal vein occlusion 36-month outcomes with anti-vascular endothelial growth factors: the Fight Retinal Blindness! registry

PURPOSE To analyze the 3-year outcomes in a broad population of patients starting vascular endothelial growth factor (VEGF) inhibitors for central retinal vein occlusion (CRVO) in routine clinical practice. DESIGN Observational database study PARTICIPANTS: 527 treatment-naïve CRVO eyes that commenced VEGF inhibitors between December 1, 2010-2018 tracked in the Fight Retinal Blindness! registry. METHODS Longitudinal models were used to plot changes in visual acuity (VA) and central subfield thickness (CST). MAIN OUTCOME MEASURES Mean change in VA from baseline to 36 months, injections, visits, completion, switching and suspensions of therapy >180 days at final review. RESULTS Overall (527 eyes) mean VA change (95% CI) was +10 (7, 12) letters, 37% had final VA ≥70 and 30% ≤35 letters, mean CST changed -306μm. Completers (257/527, 49%) had mean 36-month changes in VA and CST of +12 letters and -324μm with a median of 18 injections at 26 visits. The adjusted mean VA change was similar with each VEGF inhibitor (mean, +11.4 letters) despite a greater reduction in CST with aflibercept (-310μm) vs. ranibizumab (-258μm) vs. bevacizumab (-216μm; P 73 letters, 42/527, 8%) lost 7 letters. Switching (160/527, 30%) was most often to aflibercept (79 eyes). Using suspensions and discontinuation reasons we identified similar proportions had ceased therapy (154/527, 29%) as were still receiving it at 36 months (165/527, 31%). Only 62/527 eyes (12%) had resolution of macular edema without treatment for over 6 months. CONCLUSIONS Patients with CRVO that commenced VEGF inhibitors in routine care for whom follow-up was available had VA improvements of around 12 letters at three years, but with more than 50% lost to follow the VA outcome for the entire group is likely worse. The choice of VEGF inhibitor influenced CST but not VA outcomes. We estimate that around half of eyes were still receiving injections after 36 months

Hemiretinal vein occlusion 12-month outcomes are unique with vascular endothelial growth factor inhibitors: data from the Fight Retinal Blindness! Registry

BACKGROUND/AIMS To describe baseline characteristics and 12-month outcomes with vascular endothelial growth factor (VEGF) inhibitors of treatment-naïve hemiretinal vein occlusion (HRVO) compared with branch (BRVO) and central (CRVO) variants in routine clinical care. METHODS A database observational study recruited 79 HRVO eyes, 590 BRVO eyes and 344 CRVO eyes that initiated therapy over 10 years. The primary outcome was mean change in visual acuity (VA-letters read on a logarithm of minimal angle of resolution chart) at 12 months. Secondary outcomes included mean change in central subfield thickness (CST), injections and visits. RESULTS At baseline, mean VA in HRVO (53.8) was similar to CRVO (51.9; p=0.40) but lower than BRVO (59.4; p=0.009). HRVO eyes improved to match BRVO eyes from soon after treatment started through 12 months. Mean change in VA was greater in HRVO (+16.4) than both BRVO (+11.4; p=0.006) and CRVO (+8.5; p<0.001). Mean change in CST in HRVO (-231 µm) was similar to CRVO (-259 µm; p=0.33) but greater than BRVO eyes (-151 µm; p=0.003). The groups had similar median burdens of eight injections and nine visits. CONCLUSIONS HRVO generally experienced the greatest mean change in VA of the three types of RVO when treated with VEGF inhibitors, ending with similar 12-month VA and CST to BRVO despite starting closer to CRVO. Inclusion of HRVO in BRVO or CRVO cohorts of clinical trials would be expected to proportionally inflate and skew the visual and anatomic outcomes

Collagen distribution in the human vitreoretinal interface

PURPOSE. To evaluate the presence of collagen types I to VII, IX, XI, and XVIII at the posterior pole, the equator and the preequatorial area in human donor eyes, since collagens are important macromolecules that contribute to vitreoretinal adhesion at the vitreoretinal interface. METHODS. Freshly isolated human retinectomy samples from the equator were used for reverse transcription-polymerase chain reaction to detect mRNA of the above-mentioned collagens. In addition, human donor eyes and equatorial retinectomy samples were embedded in paraffin, stained with antibodies against the collagens and evaluated by light microscopy (LM). RESULTS. Retinectomy samples expressed mRNA of all tested collagen types. By LM, vitreous cortex was positive for collagen types II, V, IX, and XI. In all three regions within the donor eyes and in the retinectomy samples, the internal limiting membrane (ILM) showed types IV, VI, and XVIII; the retinal vasculature was positive for types I to VI and XVIII in most specimens; and the retinal layers showed condensed spots of type VII. In addition, type VII increased in density and in distribution over the retinal layers toward the posterior pole. CONCLUSIONS. Staining patterns of collagen types I to V, IX, XI, and XVIII confirmed previous observations. Important new findings include the presence of type VI in the ILM and type VII in several layers of the retina. Both collagens can anchor matrix components, and type VI could be involved in vitreoretinal attachment. Furthermore, the presence of collagen mRNA in human retinectomy samples may be an indication of postnatal collagen production by retinal cells

Remodelling of the human vitreous and vitreoretinal interface - A dynamic process

The highly hydrated, almost acellular vitreous body of the human eye consists of only 0.1% macromolecules, of which collagens are the most important for its matrix structure. During embryological development, the human vitreous body is a highly dynamic matrix, in which the primary (vascular) vitreous is gradually replaced by the secondary (avascular) vitreous. With aging, the human vitreous undergoes a slowly progressive remodelling, characterized by the gradual formation of collagenous condensations and liquefied spaces in the gel structure. The former is probably the result of collagen synthesis and the deposition of newly formed collagen into the matrix, while the latter is probably due to collagen breakdown. Therefore, remodelling of the vitreous matrix starts at a very early age and continues into old age, albeit at a slower pace. Older theories and concepts of a strict spatial separation between the primary and secondary vitreous during embryonic development, and morphological changes in the aging vitreous being due to a simple aggregation of collagen fibrils are questioned. This review describes the embryological and postnatal remodelling of the human vitreous matrix and vitreoretinal interface, in addition to the mechanisms and cells that are potentially involved in this process. (C) 2010 Elsevier Ltd. All rights reserved

Human retinal Muller cells synthesize collagens of the vitreous and vitreoretinal interface in vitro

Purpose: To investigate the capacity of cultured Muller cells to synthesize collagens, since previous studies indicated that Muller cells could be involved in collagen remodeling at the vitreoretinal border in adult human eyes. Methods: Spontaneously immortalized cultured human Muller cells were analyzed for the presence of mRNA of types I-VII, IX, XI, and XVII collagen by RT-PCR. Furthermore, Muller cells were immunocytochemically stained for light microscopic (LM) evaluation of these collagens and their main characteristics. Finally, cell extracts and culture medium were evaluated by western blot (WB) analysis using anticollagen antibodies. Results: Cultured Muller cells contained mRNA for types I-VII, IX, and XI collagen, but not for type XVII collagen. LM and WB confirmed the intracellular expression of all the above-mentioned collagens with the exception of type XVII. Collagen secretion into the medium was established for types I-VII, IX, and XI collagen. Conclusions: Cultured Muller cells can synthesize internal limiting lamina and vitreous collagens. Possible collagen production by Muller cells could explain and expand on previous in vivo morphological findings in the embryonic and postnatal period and in pathologic conditions

Mature Enzymatic Collagen Cross-Links, Hydroxylysylpyridinoline and Lysylpyridinoline, in the Aging Human Vitreous

PURPOSE. The vitreous body of the human eye undergoes progressive morphologic changes with aging. Since the enzymatic collagen cross-links hydroxylysylpyridinoline (HP) and lysylpyridinoline (LP) are known to be important for the integrity of the collagen matrix, the presence in the vitreous on aging was studied. METHODS. Vitreous bodies (VBs; n = 143) from 119 donors (age 4-80 years; mean +/- SD, 54.3 +/- 17.0 years) were carefully dissected. After weighing and freeze-drying, all samples were analyzed by high performance liquid chromatography. Left and right eyes of 24 donors were compared and, for age-related phenomena, 119 single eyes were used. RESULTS. Within one donor, no significant differences were found between left and right eyes. On aging, VB wet weight (4.42 +/- 0.84 g) accumulates until 35 years and decreases thereafter. Collagen content (0.30 +/- 0.14 mg), HP per triple helix (TH; 0.55 +/- 0.18), and (HP plus LP)/TH (0.61 +/- 0.19) increase until 50 years followed by a decrease, whereas LP/TH (0.057 +/- 0.018) accumulates until 50 years and remains constant thereafter. The ratio between HP and LP (range, 0.42 - 31.0; median, 10.0) is constant over time. CONCLUSIONS. The accumulation of enzymatic collagen cross-links until 50 years is consistent with collagen maturation and possible collagen synthesis in the human vitreous body. The decline of collagen cross-links after 50 years is consistent with collagen breakdown. (Invest Ophthalmol Vis Sci. 2009; 50: 1041-1046) DOI: 10.1167/iovs.08-171

Immunocytochemical analyses of cultured Müller cells in medium with G5

Types I () and V () collagen show clear extracellular fibrillar threads and less intracellular staining compared to and , respectively. Types II () and XI () collagen show some fine extracellular threads and small granules (arrows) and decreased intracellular staining compared to and , respectively. In the inlays of and , the extracellular collagen is magnified two times. Bars in each panel equal 50 μm.<p><b>Copyright information:</b></p><p>Taken from "Human retinal Müller cells synthesize collagens of the vitreous and vitreoretinal interface in vitro"</p><p></p><p>Molecular Vision 2008;14():652-660.</p><p>Published online 26 Mar 2008</p><p>PMCID:PMC2276182.</p><p></p

Epigenetic Signatures Discriminate Patients With Primary Sclerosing Cholangitis and Ulcerative Colitis From Patients With Ulcerative Colitis

Background: Primary sclerosing cholangitis (PSC) is a chronic inflammatory liver disease affecting the intra- and extrahepatic bile ducts, and is strongly associated with ulcerative colitis (UC). In this study, we explored the peripheral blood DNA methylome and its immune cell composition in patients with PSC-UC, UC, and healthy controls (HC) with the aim to develop a predictive assay in distinguishing patients with PSC-UC from those with UC alone. Methods: The peripheral blood DNA methylome of male patients with PSC and concomitant UC, UC and HCs was profiled using the Illumina HumanMethylation Infinium EPIC BeadChip (850K) array. Differentially methylated CpG position (DMP) and region (DMR) analyses were performed alongside gradient boosting classification analyses to discern PSC-UC from UC patients. As observed differences in the DNA methylome could be the result of differences in cellular populations, we additionally employed mass cytometry (CyTOF) to characterize the immune cell compositions. Results: Genome wide methylation analysis did not reveal large differences between PSC-UC and UC patients nor HCs. Nonetheless, using gradient boosting we were capable of discerning PSC-UC from UC with an area under the receiver operator curve (AUROC) of 0.80. Four CpG sites annotated to the NINJ2 gene were found to strongly contribute to the predictive performance. While CyTOF analyses corroborated the largely similar blood cell composition among patients with PSC-UC, UC and HC, a higher abundance of myeloid cells was observed in UC compared to PSC-UC patients. Conclusion: DNA methylation enables discerning PSC-UC from UC patients, with a potential for biomarker development