Publications by authors named "Dimitri T Azar"

121 Publications

Corneal Lymphangiogenesis as a Potential Target in Dry Eye Disease - A Systematic Review.

Surv Ophthalmol 2021 Mar 31. Epub 2021 Mar 31.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, Illinois.

Dry eye disease (DED) is a common ocular surface condition causing symptoms of significant discomfort, visual disturbance, and pain. With recent advancements, DED has become recognized as a chronic self-perpetuating inflammatory condition triggered by various internal and environmental factors. DED has been shown to arise from the activation of both the innate and adaptive immune systems, leading to significant corneal epithelium and lacrimal gland dysfunction. While the cornea is normally avascular and thus imbued with angiogenic and lymphangiogenic privilege, various DED models have revealed activated corneal antigen-presenting cells in regional lymph nodes, suggesting the formation of new corneal lymphatic vessels in DED. The recent availability of reliable lymphatic cell surface markers such as LYVE-1 has made it possible to study lymphangiogenesis. Accordingly, numerous studies have been published within the last decade discussing the role of lymphangiogenesis in DED pathology. We systematically review the literature to identify and evaluate studies presenting data on corneal lymphangiogenesis in DED. There is significant evidence supporting corneal lymphangiogenesis as a central mediator of DED pathogenesis. These findings suggest that anti-lymphangiogenic therapeutic strategies may be a viable option for the treatment of DED, a conclusion supported by the limited number of reported clinical trials examining anti-lymphangiogenic modalities in DED.
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http://dx.doi.org/10.1016/j.survophthal.2021.03.007DOI Listing
March 2021

The AI Revolution and How to Prepare for It.

Transl Vis Sci Technol 2020 03 18;9(2):16. Epub 2020 Mar 18.

Department of Ophthalmology and Visual Sciences, University of Illinois at Chicago, Chicago, IL, USA.

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http://dx.doi.org/10.1167/tvst.9.2.16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7395668PMC
March 2020

Artificial intelligence in ophthalmology during COVID-19 and in the post COVID-19 era.

Curr Opin Ophthalmol 2020 Sep;31(5):447-453

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, Illinois.

Purpose Of Review: To highlight artificial intelligence applications in ophthalmology during the COVID-19 pandemic that can be used to: describe ocular findings and changes correlated with COVID-19; extract information from scholarly articles on SARS-CoV-2 and COVID-19 specific to ophthalmology; and implement efficient patient triage and telemedicine care.

Recent Findings: Ophthalmology has been leading in artificial intelligence and technology applications. With medical imaging analysis, pixel-annotated distinguishable features on COVID-19 patients may help with noninvasive diagnosis and severity outcome predictions. Using natural language processing (NLP) and data integration methods, topic modeling on more than 200 ophthalmology-related articles on COVID-19 can summarize ocular manifestations, viral transmission, treatment strategies, and patient care and practice management. Artificial intelligence for telemedicine applications can address the high demand, prioritize and triage patients, as well as improve at home-monitoring devices and secure data transfers.

Summary: COVID-19 is significantly impacting the way we are delivering healthcare. Given the already successful implementation of artificial intelligence applications and telemedicine in ophthalmology, we expect that these systems will be embraced more as tools for research, education, and patient care.
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http://dx.doi.org/10.1097/ICU.0000000000000685DOI Listing
September 2020

Simultaneous fluorescence imaging of distinct nerve and blood vessel patterns in dual Thy1-YFP and Flt1-DsRed transgenic mice.

Angiogenesis 2020 08 5;23(3):459-477. Epub 2020 May 5.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, IL, USA.

Blood vessels and nerve tissues are critical to the development and functionality of many vital organs. However, little is currently known about their interdependency during development and after injury. In this study, dual fluorescence transgenic reporter mice were utilized to observe blood vessels and nervous tissues in organs postnatally. Thy1-YFP and Flt1-DsRed (TYFD) mice were interbred to achieve dual fluorescence in the offspring, with Thy1-YFP yellow fluorescence expressed primarily in nerves, and Flt1-DsRed fluorescence expressed selectively in blood vessels. Using this dual fluorescent mouse strain, we were able to visualize the networks of nervous and vascular tissue simultaneously in various organ systems both in the physiological state and after injury. Using ex vivo high-resolution imaging in this dual fluorescent strain, we characterized the organizational patterns of both nervous and vascular systems in a diverse set of organs and tissues. In the cornea, we also observed the dynamic patterns of nerve and blood vessel networks following epithelial debridement injury. These findings highlight the versatility of this dual fluorescent strain for characterizing the relationship between nerve and blood vessel growth and organization.
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http://dx.doi.org/10.1007/s10456-020-09724-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7316607PMC
August 2020

Proteomics-Based Characterization of the Effects of MMP14 on the Protein Content of Exosomes from Corneal Fibroblasts.

Protein Pept Lett 2020 ;27(10):979-988

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, IL 60612, United States.

Background: Exosomes secreted by corneal fibroblasts contain matrix metalloproteinase (MMP) 14, which is known to influence pro-MMP2 accumulation on exosomes. Accordingly, we hypothesized that the enzymatic activity of MMP14 may alter the protein content of corneal fibroblast- secreted exosomes.

Objective: The aim of this study was to investigate the effects of MMP14 on the composition and biological activity of corneal fibroblast-derived exosomes.

Methods: Knock out of the catalytic domain (ΔExon4) of MMP14 in corneal fibroblasts was used to determine the effect of MMP14 expression on the characteristics of fibroblast-secreted exosomes. The amount of secreted proteins and their size distribution were measured using Nano Tracking Analysis. Proteins within exosomes from wild-type (WT) and ΔExon4-deficient fibroblasts were identified by liquid chromatography-tandem mass spectrometry (MS/MS) proteomics analysis. The proteolytic effects of MMP14 were evaluated in vitro via MS identification of eliminated proteins. The biological functions of MMP14-carrying exosomes were investigated via fusion to endothelial cells and flow cytometric assays.

Results: Exosomes isolated from WT and ΔExon4-deficient fibroblasts exhibited similar size distributions and morphologies, although WT fibroblasts secreted a greater amount of exosomes. The protein content, however, was higher in ΔExon4-deficient fibroblast-derived exosomes than in WT fibroblast-derived exosomes. Proteomics analysis revealed that WT-derived exosomes included proteins that regulated cell migration, and ΔExon4 fibroblast-derived exosomes contained additional proteins that were cleaved by MMP14.

Conclusion: Our findings suggest that MMP14 expression influences the protein composition of exosomes secreted by corneal fibroblasts, and through those biological components, MMP14 in corneal fibroblasts derived-exosomes may regulate corneal angiogenesis.
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http://dx.doi.org/10.2174/0929866527666200408142827DOI Listing
January 2021

Transgenic models for investigating the nervous system: Currently available neurofluorescent reporters and potential neuronal markers.

Biochim Biophys Acta Gen Subj 2020 07 12;1864(7):129595. Epub 2020 Mar 12.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, IL, United States of America. Electronic address:

Recombinant DNA technologies have enabled the development of transgenic animal models for use in studying a myriad of diseases and biological states. By placing fluorescent reporters under the direct regulation of the promoter region of specific marker proteins, these models can localize and characterize very specific cell types. One important application of transgenic species is the study of the cytoarchitecture of the nervous system. Neurofluorescent reporters can be used to study the structural patterns of nerves in the central or peripheral nervous system in vivo, as well as phenomena involving embryologic or adult neurogenesis, injury, degeneration, and recovery. Furthermore, crucial molecular factors can also be screened via the transgenic approach, which may eventually play a major role in the development of therapeutic strategies against diseases like Alzheimer's or Parkinson's. This review describes currently available reporters and their uses in the literature as well as potential neural markers that can be leveraged to create additional, robust transgenic models for future studies.
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http://dx.doi.org/10.1016/j.bbagen.2020.129595DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7196036PMC
July 2020

Quantification of Angiogenesis and Lymphangiogenesis in the Dual ex vivo Aortic and Thoracic Duct Assay.

Protein Pept Lett 2020 ;27(1):30-40

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, IL, United States.

Background: Lymphatic vessel formation (lymphangiogenesis) plays important roles in cancer metastasis, organ rejection, and lymphedema, but the underlying molecular events remain unclear. Furthermore, despite significant overlap in the molecular families involved in angiogenesis and lymphangiogenesis, little is known about the crosstalk between these processes. The ex vivo aortic ring assay and lymphatic ring assay have enabled detailed studies of vessel sprouting, but harvesting and imaging clear thoracic duct samples remain challenging. Here we present a modified ex vivo dual aortic ring and thoracic duct assay using tissues from dual fluorescence reporter Prox1- GFP/Flt1-DsRed (PGFD) mice, which permit simultaneous visualization of blood and lymphatic endothelial cells.

Objective: To characterize the concurrent sprouting of intrinsically fluorescent blood and lymphatic vessels from harvested aorta and thoracic duct samples.

Methods: Dual aorta and thoracic duct specimens were harvested from PGFD mice, grown in six types of endothelial cell growth media (one control, five that each lack a specific growth factor), and visualized by confocal fluorescence microscopy. Linear mixed models were used to compare the extent of vessel growth and sprouting over a 28-day period.

Results: Angiogenesis occurred prior to lymphangiogenesis in our assay. The control medium generally induced superior growth of both vessel types compared with the different modified media formulations. The greatest decrease in lymphangiogenesis was observed in vascular endothelial growth factor-C (VEGF-C)-devoid medium, suggesting the importance of VEGF-C in lymphangiogenesis.

Conclusion: The modified ex vivo dual aortic ring and thoracic duct assay represents a powerful tool for studying angiogenesis and lymphangiogenesis in concert.
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http://dx.doi.org/10.2174/0929866526666190925145842DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6978644PMC
March 2020

Application of corneal injury models in dual fluorescent reporter transgenic mice to understand the roles of the cornea and limbus in angiogenic and lymphangiogenic privilege.

Sci Rep 2019 08 23;9(1):12331. Epub 2019 Aug 23.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, Illinois, USA.

The role of the corneal epithelium and limbus in corneal avascularity and pathological neovascularization (NV) is not well understood. To investigate the contributions of the corneal and limbal epithelia in angiogenic and lymphangiogenic privilege, we designed five injury models involving debridement of different portions of the cornea and limbus and applied them to the dual-fluorescence reporter Prox1-GFP/Flt1-DsRed mouse, which permits in vivo imaging of blood and lymphatic vessels via fluorescence microscopy. Debridement of the whole cornea resulted in significant hemangiogenesis (HA) and lymphangiogenesis (LA), while that of the whole limbus yielded minimal corneal HA or LA. Following hemilimbal plus whole corneal debridement, corneal NV occurred only through the non-injured aspect of the limbus. Overall, these results suggest that the integrity of the corneal epithelium is important for (lymph)angiogenic privilege, whereas the limbus does not act as a physical or physiologic barrier to invading vessels. In CDh5-CreERT2VEGFR2lox/PGFD mice, conditional deletion of vascular endothelial growth factor receptor 2 in vascular endothelial cells abolished injury-induced HA and LA, demonstrating the utility of this transgenic mouse line for identifying important factors in the process of neovascularization.
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http://dx.doi.org/10.1038/s41598-019-48811-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6707148PMC
August 2019

MMP14-Containing Exosomes Cleave VEGFR1 and Promote VEGFA-Induced Migration and Proliferation of Vascular Endothelial Cells.

Invest Ophthalmol Vis Sci 2019 05;60(6):2321-2329

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois-Chicago, Chicago, Illinois, United States.

Purpose: Investigate the impact matrix metalloproteinase 14 (MMP14) delivered via exosomes produced by corneal fibroblasts on vascular endothelial growth factor receptor 1 (VEGFR1) cleavage on endothelial cells, and other key processes of angiogenesis.

Methods: Proteolysis of VEGFR1 and R2 by the catalytic domain of MMP14 was investigated via immunocytochemistry with anti-VEGFR1, anti-VEGFR2, and anti-MMP14 antibodies. Exosomes were isolated via precipitation and serial ultracentrifugation from wild-type (WT) and MMP14 exon4-deficient corneal fibroblasts. Transmission electron microscopy and nanotracking analysis were used to characterize the isolated exosomes. The presence of MMP14 in exosomes from WT fibroblasts was confirmed by Western blotting. VEGFR1 cleavage upon treatment with WT-derived exosomes, Δexon4-derived exosomes, or the pan-MMP inhibitor GM60001 was examined via in vitro proteolysis analysis using recombinant mouse (rm) VEGFR1/R2. Endothelial cell migration and proliferation were investigated using a Boyden chamber assay and BrdU incorporation, respectively.

Results: WT-derived exosomes specifically cleaved rmVEGFR1 in vitro, whereas Δexon4-derived exosomes did not. Treatment with the pan-MMP inhibitor GM6001 effectively inhibited VEGFR1 cleavage by WT-derived exosomes, confirming the role of MMP14 in this cleavage. WT-derived exosomes induced greater endothelial cell migration (P < 0.01) and proliferation (P < 0.5) compared to Δexon4-derived exosomes.

Conclusions: MMP14-containing exosomes may be involved in the regulation of corneal neovascularization through degradation of VEGFR1 and VEGFA-induced endothelial cell proliferation and migration.
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http://dx.doi.org/10.1167/iovs.18-26277DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6532701PMC
May 2019

Direct and Indirect Flap Measurements in Femtosecond Laser-Assisted In Situ Keratomileusis.

Cornea 2019 Mar;38(3):297-303

Department of Ophthalmology and Visual Sciences, Illinois Eye & Ear Infirmary, University of Illinois at Chicago, Chicago, IL.

Purpose: To compare direct and indirect LASIK flap thickness measurements using ultrasound and Scheimpflug technology.

Methods: Eighty-two eyes treated with laser-assisted in situ keratomileusis refractive surgery using a femtosecond laser (IntraLase FS150) were prospectively included in the study. Flap thickness was set to 115 μm. Corneal flap thickness was measured using the direct method-ie, ultrasound pachymetry immediately after flap construction in the presence of cavitation bubbles-and indirect methods, with subtraction of intraoperative post-lift corneal thickness measured using ultrasound pachymetry (intrastroma) from preoperative central corneal thickness using ultrasound (Indirect-US) or Scheimpflug thinnest pachymetry (Indirect-Scheimpflug).

Results: Mean flap thickness was overestimated using the indirect methods, Indirect-US and Indirect-Scheimpflug (122.6 ± 24.5 μm and 128.1 ± 26.1 μm, respectively; P < 0.0060 and P < 0.0001, respectively). There were no significant correlations between the direct and indirect methods. Indirect-Scheimpflug was significantly higher (P = 0.0122) than Indirect-US. The closest average flap thickness compared with the set parameter of 115 μm was that of the direct method (115.6 ± 8.6 μm; 95% confidence interval: -1.3 to 2.5; P = 0.5163). The direct method provided the lowest SD of all groups (SD: 8.64).

Conclusions: The direct method of flap thickness measurement was the most comparable to the set parameter compared with the indirect subtraction methods. Additional studies are needed to determine which method allows for the most accurate measurement of flap thickness.
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http://dx.doi.org/10.1097/ICO.0000000000001836DOI Listing
March 2019

Fluorescent reporter transgenic mice for in vivo live imaging of angiogenesis and lymphangiogenesis.

Angiogenesis 2018 11 3;21(4):677-698. Epub 2018 Jul 3.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, IL, USA.

The study of lymphangiogenesis is an emerging science that has revealed the lymphatic system as a central player in many pathological conditions including cancer metastasis, lymphedema, and organ graft rejection. A thorough understanding of the mechanisms of lymphatic growth will play a key role in the development of therapeutic strategies against these conditions. Despite the known potential of this field, the study of lymphatics has historically lagged behind that of hemangiogenesis. Until recently, significant strides in lymphatic studies were impeded by a lack of lymphatic-specific markers and suitable experimental models compared to those of the more immediately visible blood vasculature. Lymphangiogenesis has also been shown to be a key phenomenon in developmental biological processes, such as cell proliferation, guided migration, differentiation, and cell-to-cell communication, making lymphatic-specific visualization techniques highly desirable and desperately needed. Imaging modalities including immunohistochemistry and in situ hybridization are limited by the need to sacrifice animal models for tissue harvesting at every experimental time point. Moreover, the processes of mounting and staining harvested tissues may introduce artifacts that can confound results. These traditional methods for investigating lymphatic and blood vasculature are associated with several problems including animal variability (e.g., between mice) when replicating lymphatic growth environments and the cost concerns of prolonged, labor-intensive studies, all of which complicate the study of dynamic lymphatic processes. With the discovery of lymphatic-specific markers, researchers have been able to develop several lymphatic and blood vessel-specific, promoter-driven, fluorescent-reporter transgenic mice for visualization of lymphatics in vivo and in vitro. For instance, GFP, mOrange, tdTomato, and other fluorescent proteins can be expressed under control of a lymphatic-specific marker like Prospero-related homeobox 1 (Prox1), which is a highly conserved transcription factor for determining embryonic organogenesis in vertebrates that is implicated in lymphangiogenesis as well as several human cancers. Importantly, Prox1-null mouse embryos develop without lymphatic vessels. In human adults, Prox1 maintains lymphatic endothelial cells and upregulates proteins associated with lymphangiogenesis (e.g., VEGFR-3) and downregulates angiogenesis-associated gene expression (e.g., STAT6). To visualize lymphatic development in the context of angiogenesis, dual fluorescent-transgenic reporters, like Prox1-GFP/Flt1-DsRed mice, have been bred to characterize lymphatic and blood vessels simultaneously in vivo. In this review, we discuss the trends in lymphatic visualization and the potential usage of transgenic breeds in hemangiogenesis and lymphangiogenesis research to understand spatial and temporal correlations between vascular development and pathological progression.
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http://dx.doi.org/10.1007/s10456-018-9629-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6472480PMC
November 2018

Potential lymphangiogenesis therapies: Learning from current antiangiogenesis therapies-A review.

Med Res Rev 2018 09 12;38(6):1769-1798. Epub 2018 Mar 12.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, IL.

In recent years, lymphangiogenesis, the process of lymphatic vessel formation from existing lymph vessels, has been demonstrated to have a significant role in diverse pathologies, including cancer metastasis, organ graft rejection, and lymphedema. Our understanding of the mechanisms of lymphangiogenesis has advanced on the heels of studies demonstrating vascular endothelial growth factor C as a central pro-lymphangiogenic regulator and others identifying multiple lymphatic endothelial biomarkers. Despite these breakthroughs and a growing appreciation of the signaling events that govern the lymphangiogenic process, there are no FDA-approved drugs that target lymphangiogenesis. In this review, we reflect on the lessons available from the development of antiangiogenic therapies (26 FDA-approved drugs to date), review current lymphangiogenesis research including nanotechnology in therapeutic drug delivery and imaging, and discuss molecules in the lymphangiogenic pathway that are promising therapeutic targets.
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http://dx.doi.org/10.1002/med.21496DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6135718PMC
September 2018

Angiogenesis and lymphangiogenesis in corneal transplantation-A review.

Surv Ophthalmol 2018 Jul - Aug;63(4):453-479. Epub 2017 Dec 27.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, Illinois, USA. Electronic address:

Corneal transplantation has been proven effective for returning the gift of sight to those affected by corneal disorders such as opacity, injury, and infections that are a leading cause of blindness. Immune privilege plays an important role in the success of corneal transplantation procedures; however, immune rejection reactions do occur, and they, in conjunction with a shortage of corneal donor tissue, continue to pose major challenges. Corneal immune privilege is important to the success of corneal transplantation and closely related to the avascular nature of the cornea. Corneal avascularity may be disrupted by the processes of angiogenesis and lymphangiogenesis, and for this reason, these phenomena have been a focus of research in recent years. Through this research, therapies addressing certain rejection reactions related to angiogenesis have been developed and implemented. Corneal donor tissue shortages also have been addressed by the development of new materials to replace the human donor cornea. These advancements, along with other improvements in the corneal transplantation procedure, have contributed to an improved success rate for corneal transplantation. We summarize recent developments and improvements in corneal transplantation, including the current understanding of angiogenesis mechanisms, the anti-angiogenic and anti-lymphangiogenic factors identified to date, and the new materials being used. Additionally, we discuss future directions for research in corneal transplantation.
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http://dx.doi.org/10.1016/j.survophthal.2017.12.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6003844PMC
September 2018

TFOS DEWS II Report Executive Summary.

Ocul Surf 2017 10 8;15(4):802-812. Epub 2017 Aug 8.

Schepens Eye Research Institute, Massachusetts Eye and Ear, and Department of Ophthalmology, Harvard Medical School, Boston, MA, USA. Electronic address:

This article presents an Executive Summary of the conclusions and recommendations of the 10-chapter TFOS DEWS II report. The entire TFOS DEWS II report was published in the July 2017 issue of The Ocular Surface. A downloadable version of the document and additional material, including videos of diagnostic and management techniques, are available on the TFOS website: www.TearFilm.org.
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http://dx.doi.org/10.1016/j.jtos.2017.08.003DOI Listing
October 2017

Prox1-GFP/Flt1-DsRed transgenic mice: an animal model for simultaneous live imaging of angiogenesis and lymphangiogenesis.

Angiogenesis 2017 Nov 9;20(4):581-598. Epub 2017 Aug 9.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, IL, 60612, USA.

The roles of angiogenesis in development, health, and disease have been studied extensively; however, the studies related to lymphatic system are limited due to the difficulty in observing colorless lymphatic vessels. But recently, with the improved technique, the relative importance of the lymphatic system is just being revealed. We bred transgenic mice in which lymphatic endothelial cells express GFP (Prox1-GFP) with mice in which vascular endothelial cells express DsRed (Flt1-DsRed) to generate Prox1-GFP/Flt1-DsRed (PGFD) mice. The inherent fluorescence of blood and lymphatic vessels allows for direct visualization of blood and lymphatic vessels in various organs via confocal and two-photon microscopy and the formation, branching, and regression of both vessel types in the same live mouse cornea throughout an experimental time course. PGFD mice were bred with CDh5CreERT2 and VEGFR2lox knockout mice to examine specific knockouts. These studies showed a novel role for vascular endothelial cell VEGFR2 in regulating VEGFC-induced corneal lymphangiogenesis. Conditional deletion of vascular endothelial VEGFR2 abolished VEGFA- and VEGFC-induced corneal lymphangiogenesis. These results demonstrate the potential use of the PGFD mouse as a powerful animal model for studying angiogenesis and lymphangiogenesis.
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http://dx.doi.org/10.1007/s10456-017-9572-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5782821PMC
November 2017

TFOS DEWS II iatrogenic report.

Ocul Surf 2017 07 20;15(3):511-538. Epub 2017 Jul 20.

Aston University, Birmingham, UK.

Dry eye can be caused by a variety of iatrogenic interventions. The increasing number of patients looking for eye care or cosmetic procedures involving the eyes, together with a better understanding of the pathophysiological mechanisms of dry eye disease (DED), have led to the need for a specific report about iatrogenic dry eye within the TFOS DEWS II. Topical medications can cause DED due to their allergic, toxic and immuno-inflammatory effects on the ocular surface. Preservatives, such as benzalkonium chloride, may further aggravate DED. A variety of systemic drugs can also induce DED secondary to multiple mechanisms. Moreover, the use of contact lens induces or is associated with DED. However, one of the most emblematic situations is DED caused by surgical procedures such as corneal refractive surgery as in laser-assisted in situ keratomileusis (LASIK) and keratoplasty due to mechanisms intrinsic to the procedure (i.e. corneal nerve cutting) or even by the use of postoperative topical drugs. Cataract surgery, lid surgeries, botulinum toxin application and cosmetic procedures are also considered risk factors to iatrogenic DED, which can cause patient dissatisfaction, visual disturbance and poor surgical outcomes. This report also presents future directions to address iatrogenic DED, including the need for more in-depth epidemiological studies about the risk factors, development of less toxic medications and preservatives, as well as new techniques for less invasive eye surgeries. Novel research into detection of early dry eye prior to surgeries, efforts to establish appropriate therapeutics and a greater attempt to regulate and oversee medications, preservatives and procedures should be considered.
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http://dx.doi.org/10.1016/j.jtos.2017.05.004DOI Listing
July 2017

Risk profiles of ectasia after keratorefractive surgery.

Curr Opin Ophthalmol 2017 Jul;28(4):337-342

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, Illinois, USA.

Purpose Of Review: To identify and evaluate the risk factors of iatrogenic ectasia after refractive surgery.

Recent Findings: We reviewed recently published papers that identified various risk factors associated with ectasia after LASIK, photorefractive keratectomy, small incision lenticule extraction, and other refractive surgical procedures. We also attempted to evaluate the relative contributions of these factors to the development of ectasia following refractive surgery. Forme fruste keratoconus, genetic predisposition to keratoconus, low residual stromal bed thickness (through high myopia, thin preoperative cornea, or thick LASIK flap), and irregular corneal topography have been identified as risk factors for keratectasia development after refractive surgical procedures. A newly proposed metric, percentage tissue altered, has been reported to be a robust indicator for post LASIK ectasia risk calculation. Several cases of keratectasia have also been reported 6 to 12 months following minimally invasive small incision lenticule extraction procedure. Other risk factors associated with iatrogenic ectasia include eye rubbing, young age, and pregnancy.

Summary: Ectasia after refractive surgery is a relatively rare complication which can lead to sight-threatening complications if not detected and treated in time. It is important to continue our quest to improve our methods of identifying absolute and relative risk factors of ectasia and their cut-off values following various keratorefractive surgical procedures.
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http://dx.doi.org/10.1097/ICU.0000000000000383DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6444927PMC
July 2017

Potential role of corneal epithelial cell-derived exosomes in corneal wound healing and neovascularization.

Sci Rep 2017 02 6;7:40548. Epub 2017 Feb 6.

Schepens Eye Research Institute/MEE, Boston, Massachusetts, United States.

Specific factors from the corneal epithelium underlying the stimulation of stromal fibrosis and myofibroblast formation in corneal wound healing have not been fully elucidated. Given that exosomes are known to transfer bioactive molecules among cells and play crucial roles in wound healing, angiogenesis, and cancer, we hypothesized that corneal epithelial cell-derived exosomes may gain access to the underlying stromal fibroblasts upon disruption of the epithelial basement membrane and that they induce signaling events essential for corneal wound healing. In the present study, exosome-like vesicles were observed between corneal epithelial cells and the stroma during wound healing after corneal epithelial debridement. These vesicles were also found in the stroma following anterior stromal keratectomy, in which surgical removal of the epithelium, basement membrane, and anterior stroma was performed. Exosomes secreted by mouse corneal epithelial cells were found to fuse to keratocytes in vitro and to induce myofibroblast transformation. In addition, epithelial cell-derived exosomes induced endothelial cell proliferation and ex vivo aortic ring sprouting. Our results indicate that epithelial cell-derived exosomes mediate communication between corneal epithelial cells and corneal keratocytes as well as vascular endothelial cells. These findings demonstrate that epithelial-derived exosomes may be involved in corneal wound healing and neovascularization, and thus, may serve as targets for potential therapeutic interventions.
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http://dx.doi.org/10.1038/srep40548DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5292698PMC
February 2017

MMP14 Regulates VEGFR3 Expression on Corneal Epithelial Cells.

Protein Pept Lett 2016 ;23(12):1095-1102

Department of Ophthalmology and Visual Sciences, College of Medicine, University of Illinois at Chicago, Chicago, IL 60612.

Vascular endothelial growth factor receptor 3 (VEGFR3) regulates the growth and differentiation of blood and lymphatic vessels. To determine whether matrix metalloproteinase 14 (MMP14) modulates VEGFR3 expression in the corneal epithelium to influence the avascularity of the cornea, VEGFR3 expression was compared between wild-type and MMP14-deficient (MMP14 Δexon4) corneal epithelial cells. Western blot analysis showed that VEGFR3 protein expression was higher on MMP14 Δexon4 corneal epithelial cells than on wild-type cells, and quantitative RT-PCR analysis showed that VEGFR3 gene expression was highly induced in MMP14 Δexon4 corneal epithelial cells but not in wild-type corneal epithelial cells or wild-type and MMP14 Δexon4 corneal keratocytes. Unlike in epithelial cells, MMP14 Δexon4 keratocytes did not express relatively higher levels of VEGFR3 than wild-type keratocytes. Interestingly, in vitro proteolysis experiments showed that MMP14 does not cleave VEGFR3 in vitro as it does VEGFR1, indicating that other genes may be involved in the modulation of VEGFR3 expression by MMP14. Using proteomic analysis to identify candidate factors, we found that 39 nuclear proteins were differentially expressed between wildtype and MMP14 Δexon4 corneal epithelial cells. These findings suggest that MMP14 may regulate VEGFR3 expression at the transcriptional level on corneal epithelial cells but not on corneal keratocytes.
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http://dx.doi.org/10.2174/0929866523666161024142824DOI Listing
March 2017

Proangiogenic Interactions of Vascular Endothelial MMP14 With VEGF Receptor 1 in VEGFA-Mediated Corneal Angiogenesis.

Invest Ophthalmol Vis Sci 2016 06;57(7):3313-22

Department of Ophthalmology and Visual Sciences Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, Illinois, United States.

Purpose: Matrix metalloproteinase 14 (MMP14) has been shown to be required for corneal angiogenesis. We hypothesized that the proangiogenic activity of MMP14 may be based on its selective binding to, and cleaving of, vascular endothelial growth factor receptor 1 (VEGFR1), but not VEGFR2 or VEGFR3.

Methods: Recombinant human (rh)VEGFR1, R2, and R3 were incubated with human MMP14, and the reaction mixtures were analyzed by SDS-PAGE and Coomassie blue staining. Surface plasmon resonance was used to determine the equilibrium constants (KD) for binding between MMP14 and VEGFA versus rhVEGFR1, R2, and R3. Extracellular signal-regulated kinase (ERK) phosphorylation was assayed in vascular endothelial cells after incubation with VEGF and various concentrations of MMP14. Ex vivo aortic ring tube formation assays and VEGFA micropocket corneal neovascularization assays were performed using Flk1Cre/Flk1mCherry/MMP14lox and Flk1mCherry/MMP14lox control mice.

Results: Maxtrix metalloproteinase 14 increased VEGFA-induced ERK phosphorylation in a time- and concentration-dependent manner in vascular endothelial cells. Aortic ring assays showed diminished vessel sprouting in vitro in response to VEGFA, but not to basic fibroblast growth factor, in mice with conditional deletion of vascular MMP14 (Flk1creMMP14lox) compared with that in MMP14lox control mice. In addition, diminished VEGFA-induced corneal angiogenesis was seen in flk1creMMP14lox mice compared with MMP14lox mice in vivo.

Conclusions: Our findings indicate that VEGFR1 interaction with MMP14 and the enzymatic activity of MMP14 are necessary for VEGFA-induced angiogenesis. Additionally, selective cleavage of VEGFR1 by MMP14 may play an important role in VEGFA-induced corneal angiogenesis.
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http://dx.doi.org/10.1167/iovs.16-19420DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5993529PMC
June 2016

Limited versus total epithelial debridement ocular surface injury: Live fluorescence imaging of hemangiogenesis and lymphangiogenesis in Prox1-GFP/Flk1::Myr-mCherry mice.

Biochim Biophys Acta 2016 10 24;1860(10):2148-56. Epub 2016 May 24.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, College of Medicine, University of Illinois at Chicago, Chicago, IL, USA. Electronic address:

Background: Immunohistochemical staining experiments have shown that both hemangiogenesis and lymphangiogenesis occur following severe corneal and conjunctival injury and that the neovascularization of the cornea often has severe visual consequences. To better understand how hemangiogenesis and lymphangiogenesis are induced by different degrees of ocular injury, we investigated patterns of injury-induced corneal neovascularization in live Prox1-GFP/Flk1::myr-mCherry mice, in which blood and lymphatic vessels can be imaged simultaneously in vivo.

Methods: The eyes of Prox1-GFP/Flk1::myr-mCherry mice were injured according to four models based on epithelial debridement of the: A) central cornea (a 1.5-mm-diameter circle of tissue over the corneal apex), B) total cornea, C) bulbar conjunctiva, and D) cornea+bulbar conjunctiva. Corneal blood and lymphatic vessels were imaged on days 0, 3, 7, and 10 post-injury, and the percentages of the cornea containing blood and lymphatic vessels were calculated.

Results: Neither central corneal nor bulbar conjunctival debridement resulted in significant vessel growth in the mouse cornea, whereas total corneal and corneal+bulbar conjunctival debridement did. On day 10 in the central cornea, total cornea, bulbar conjunctiva, and corneal+bulbar conjunctival epithelial debridement models, the percentage of the corneal surface that was occupied by blood vessels (hemangiogenesis) was 1.9±0.8%, 7.14±2.4%, 2.29±1%, and 15.05±2.14%, respectively, and the percentage of the corneal surface that was occupied by lymphatic vessels (lymphangiogenesis) was 2.45±1.51%, 4.85±0.95%, 2.95±1.27%, and 4.15±3.85%, respectively.

Conclusions: Substantial corneal debridement was required to induce corneal neovascularization in the mouse cornea, and the corneal epithelium may therefore be partially responsible for maintaining corneal avascularity.

General Significance: Our study demonstrates that GFP/Flk1::myr-mCherry mice are a useful model for studying coordinated hemangiogenic and lymphangiogenic responses.
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http://dx.doi.org/10.1016/j.bbagen.2016.05.027DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4961530PMC
October 2016

Understanding lymphangiogenesis in knockout models, the cornea, and ocular diseases for the development of therapeutic interventions.

Surv Ophthalmol 2016 May-Jun;61(3):272-96. Epub 2015 Dec 17.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, University of Illinois at Chicago, Chicago, Illinois, USA. Electronic address:

A major focus of cancer research for several decades has been understand the ability of tumors to induce new blood vessel formation, a process known as angiogenesis. Unfortunately, only limited success has been achieved in the clinical application of angiogenesis inhibitors. We now know that lymphangiogenesis, the growth of lymphatic vessels, likely also plays a major role in tumor progression. Thus, therapeutic strategies targeting lymphangiogenesis or both lymphangiogenesis and angiogenesis may represent promising approaches for treating cancer and other diseases. Importantly, research progress toward understanding lymphangiogenesis is significantly behind that related to angiogenesis. A PubMed search of "angiogenesis" returns nearly 80,000 articles, whereas a search of "lymphangiogenesis" returns 2,635 articles. This stark contrast can be explained by the lack of molecular markers for identifying the invisible lymphatic vasculature that persisted until less than 2 decades ago, combined with the intensity of research interest in angiogenesis during the past half century. Still, significant strides have been made in developing strategies to modulate lymphangiogenesis, largely using ocular disease models. Here we review the current knowledge of lymphangiogenesis in the context of knockout models, ocular diseases, the biology of activators and inhibitors, and the potential for therapeutic interventions targeting this process.
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http://dx.doi.org/10.1016/j.survophthal.2015.12.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4835265PMC
September 2016

Matrix metalloproteinase 14 modulates signal transduction and angiogenesis in the cornea.

Surv Ophthalmol 2016 Jul-Aug;61(4):478-97. Epub 2015 Dec 2.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, University of Illinois at Chicago, Chicago, Illinois, USA. Electronic address:

The cornea is transparent and avascular, and retention of these characteristics is critical to maintaining vision clarity. Under normal conditions, wound healing in response to corneal injury occurs without the formation of new blood vessels; however, neovascularization may be induced during corneal wound healing when the balance between proangiogenic and antiangiogenic mediators is disrupted to favor angiogenesis. Matrix metalloproteinases (MMPs), which are key factors in extracellular matrix remodeling and angiogenesis, contribute to the maintenance of this balance, and in pathologic instances, can contribute to its disruption. Here, we elaborate on the facilitative role of MMPs, specifically MMP-14, in corneal neovascularization. MMP-14 is a transmembrane MMP that is critically involved in extracellular matrix proteolysis, exosome transport, and cellular migration and invasion, processes that are critical for angiogenesis. To aid in developing efficacious therapies that promote healing without neovascularization, it is important to understand and further investigate the complex pathways related to MMP-14 signaling, which can also involve vascular endothelial growth factor, basic fibroblast growth factor, Wnt/β-catenin, transforming growth factor, platelet-derived growth factor, hepatocyte growth factor or chemokines, epidermal growth factor, prostaglandin E2, thrombin, integrins, Notch, Toll-like receptors, PI3k/Akt, Src, RhoA/RhoA kinase, and extracellular signal-related kinase. The involvement and potential contribution of these signaling molecules or proteins in neovascularization are the focus of the present review.
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http://dx.doi.org/10.1016/j.survophthal.2015.11.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6417502PMC
May 2017

Endostatin's emerging roles in angiogenesis, lymphangiogenesis, disease, and clinical applications.

Biochim Biophys Acta 2015 Dec 12;1850(12):2422-38. Epub 2015 Sep 12.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, University of Illinois at Chicago, Chicago, IL, USA.

Background: Angiogenesis is the process of neovascularization from pre-existing vasculature and is involved in various physiological and pathological processes. Inhibitors of angiogenesis, administered either as individual drugs or in combination with other chemotherapy, have been shown to benefit patients with various cancers. Endostatin, a 20-kDa C-terminal fragment of type XVIII collagen, is one of the most potent inhibitors of angiogenesis.

Scope Of Review: We discuss the biology behind endostatin in the context of its endogenous production, the various receptors to which it binds, and the mechanisms by which it acts. We focus on its inhibitory role in angiogenesis, lymphangiogenesis, and cancer metastasis. We also present emerging clinical applications for endostatin and its potential as a therapeutic agent in the form a short peptide.

Major Conclusions: The delicate balance between pro- and anti-angiogenic factors can be modulated to result in physiological wound healing or pathological tumor metastasis. Research in the last decade has emphasized an emerging clinical potential for endostatin as a biomarker and as a therapeutic short peptide. Moreover, elevated or depressed endostatin levels in diseased states may help explain the pathophysiological mechanisms of the particular disease.

General Significance: Endostatin was once sought after as the 'be all and end all' for cancer treatment; however, research throughout the last decade has made it apparent that endostatin's effects are complex and involve multiple mechanisms. A better understanding of newly discovered mechanisms and clinical applications still has the potential to lead to future advances in the use of endostatin in the clinic.
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http://dx.doi.org/10.1016/j.bbagen.2015.09.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4624607PMC
December 2015

Selective Binding of Endostatin Peptide 4 to Recombinant VEGF Receptor 3 In Vitro.

Protein Pept Lett 2015 ;22(11):1025-30

Department of Ophthalmology and Visual Sciences, University of Illinois at Chicago, 1855 W. Taylor Street, Chicago, IL 60612, USA.

We previously reported that neostatin, a proteolytic fragment of collagen XVIII that includes endostatin, inhibits basic fibroblast growth factor-induced corneal angiogenesis and lymphangiogenesis. In experiments to determine which fragments in neostatin are responsible for binding to VEGF receptors (VEGFRs), we previously showed that a 28- mer sequence at the C-terminal of endostatin, known as endostatin peptide 9, preferentially binds VEGFR3-Fc over VEGFR1-Fc and VEGFR2-Fc. In the present study, we show that a different endostatin fragment, endostatin peptide 4 (26 mers long), also selectively binds VEGFR3-Fc and not VEGFR1-Fc or VEGFR2-Fc. From surface plasmon resonance data, the KD and Chi² (RU²) values for endostatin peptide 4 binding to VEGFR3-Fc are 5.72 × 10⁻⁸ M and 0.354, respectively. In conclusion, endostatin peptides 4 and 9 may be responsible for endostatin binding to VEGFR3-Fc, and this improved understanding of endostatin peptide binding to VEGFR3-Fc may support the development of therapeutics targeting lymphangiogenic processes.
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http://dx.doi.org/10.2174/0929866522666150907111953DOI Listing
July 2016

MMP14 Cleavage of VEGFR1 in the Cornea Leads to a VEGF-Trap Antiangiogenic Effect.

Invest Ophthalmol Vis Sci 2015 Aug;56(9):5450-6

Department of Ophthalmology and Visual Sciences Illinois Eye and Ear Infirmary, University of Illinois at Chicago, Chicago, Illinois, United States.

Purpose: To determine the possible antiangiogenic effect of metalloproteinase (MMP) 14 cleavage of vascular endothelial growth factor receptor 1 (VEGFR1) in the cornea.

Methods: Recombinant mouse (rm) VEGFR1 was incubated with various concentrations of recombinant MMP14 to examine proteolysis in vitro. The reaction mixture was analyzed by SDS-PAGE and stained with Coomassie blue. The fragments resulting from rmVEGFR1 cleavage by MMP14 were subjected to Edman degradation, and the amino acid sequences were aligned with rmVEGFR1 sequences. Surface plasmon resonance was used to determine the equilibrium dissociation constant (KD) between MMP14 and rmVEGFR1. The KD value of rmVEGFR1 and the 59.8-kDa cleavage product binding to VEGF-A₁₆₅ was also determined. Cell proliferation assays were performed in the presence of VEGF-A₁₆₅ plus the 59.8-kDa VEGFR1 fragment or VEGF-A₁₆₅ alone.

Results: Matrix metalloproteinase 14 binds and cleaves rmVEGFR1 to produce 59.8-kDa (N-terminal fragment, Ig domains 1-5), 35-kDa (C-terminal fragment containing IgG and His-tag), and 21-kDa (Ig domains 6-7) fragments. The 59.8-kDa fragment showed binding to VEGF-A₁₆₅ and inhibited VEGF-induced endothelial cell mitogenesis.

Conclusions: Our findings suggest that VEGFR1 cleavage by MMP14 in the cornea leads to a VEGF-trap effect, reducing the proangiogenic effect of VEGF-A₁₆₅, thereby reducing corneal angiogenesis.
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http://dx.doi.org/10.1167/iovs.14-16248DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4544186PMC
August 2015

Simultaneous in vivo imaging of blood and lymphatic vessel growth in Prox1-GFP/Flk1::myr-mCherry mice.

FEBS J 2015 Apr 6;282(8):1458-1467. Epub 2015 Mar 6.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, University of Illinois at Chicago, Chicago, IL, USA.

The ability to visually observe angiogenesis and lymphangiogenesis simultaneously and repeatedly in living animals would greatly enhance our understanding of the inter-dependence of these processes. To generate a mouse model that allows such visualization via in vivo fluorescence imaging, we crossed Prox1-GFP mice with Flk1::myr-mCherry mice to generate Prox1-GFP/Flk1::myr-mCherry mice, in which lymphatic vessels emit green fluorescence and blood vessels emit red fluorescence. Corneal neovascularization was induced in these mice using three injury models: implantation of a vascular endothelial growth factor (VEGF) pellet, implantation of a basic fibroblast growth factor (bFGF) pellet, and alkali burn injury. Vessel growth was observed in vivo by stereomicroscopy on days 0, 3, 7 and 10 after pellet implantation or alkali injury as well as in flat-mounted corneas via confocal microscopy after the final in vivo imaging time point. We observed blood and lymphatic vessel growth in all three models, with the most significant growth occurring from days 0-7. Upon VEGF stimulation, the growth kinetics of blood and lymphatic vessels were similar. Blood vessels exhibited similar growth patterns in VEGF- and bFGF-stimulated corneas. Alkali burn injury induced robust angiogenesis and lymphangiogenesis. The intrinsic fluorescence of blood and lymphatic endothelial cells in Prox1-GFP/Flk1::myr-mCherry mice permitted simultaneous in vivo imaging of angiogenesis and lymphangiogenesis. This allowed us to differentiate the processes as well as observe their inter-dependence, and will be valuable in development of therapies targeting angiogenesis and/or lymphangiogenesis.
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http://dx.doi.org/10.1111/febs.13234DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4400230PMC
April 2015

Effect of anterior corneal surface asphericity modification on fourth-order zernike spherical aberrations.

J Refract Surg 2014 Oct;30(10):708-15

Purpose: To evaluate the theoretical influence of the change in corneal asphericity (ΔQ) on the change in fourth-order Zernike spherical aberration coefficient (ΔC(4)0) with customized aspheric refractive correction of myopia and hyperopia.

Methods: The initial anterior corneal surface profile was modeled as a conic section of apical radius of curvature R0 and asphericity Q₀. The postoperative corneal profile was modeled as a conic section of apical curvature R1 and asphericity Q1, where R1 was computed from defocus D, and Q₁ selected for controlling the postoperative asphericity. The corresponding change in fourth-order spherical aberration (ΔC(0)4) was computed within a 6-mm optical zone using inner products applied to the incurred optical path changes. These calculations were repeated for different values of D, R₀, Q₀, and various intended ΔC(4)0 values.

Results: Increasing negative spherical aberration (ΔC(4)(0) < 0) requires a change toward more negative values of asphericity (increased prolateness; ΔQ < 0) for hyperopic and low myopic corrections, but more positive values (ΔQ < 0) for high myopic correction. The larger the intended change in corneal spherical aberration (ΔC(4)(0)), the more myopic the threshold value for which the required change in asphericity, ΔQ, becomes positive. The influence of the magnitude of paraxial defocus correction is less pronounced when larger changes in C(4)(0) are intended.

Conclusions: These results provide a basis for controlling the direction (sign) and the magnitude of spherical aberration changes when using customized aspheric profiles of ablation.
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http://dx.doi.org/10.3928/1081597X-20140903-10DOI Listing
October 2014

Involvement of lysosomal degradation in VEGF-C-induced down-regulation of VEGFR-3.

FEBS Lett 2014 Nov 2;588(23):4357-63. Epub 2014 Oct 2.

Department of Ophthalmology and Visual Sciences, Illinois Eye and Ear Infirmary, University of Illinois at Chicago, Chicago, IL, United States.

The vascular endothelial growth factor (VEGF)-C-induced down-regulation of VEGF receptor (VEGFR)-3 is important in lymphangiogenesis. Here, we demonstrate that VEGF-C, -D, and -C156S, but not VEGF-A, down-regulate VEGFR-3. VEGF-C stimulates VEGFR-3 tyrosyl phosphorylation and transient phosphorylation of extracellular signal-regulated kinase (ERK), p38, and c-Jun N-terminal kinases in lymphatic endothelial cells. VEGF-C-induced down-regulation of VEGFR-3 was blocked by a VEGF-C trap, tyrosine kinase inhibitor, and leupeptin, pepstatin, and E64 (LPE), but was unaffected by Notch 1 activator and γ-secretase inhibitors. Our findings indicate that VEGF-C down-regulates VEGFR-3 in lymphatic endothelial cells through VEGFR-3 kinase activation and, in part, via lysosomal degradation.
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http://dx.doi.org/10.1016/j.febslet.2014.09.034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4254303PMC
November 2014