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OBJECTIVE: To establish nomograms for linear measurements of the frontal and occipital horns of the lateral ventricle and their relationship, in pregnant patients between 18 and 40 weeks of gestation and having attended 2 units of Maternal Fetal Medicine in Bogotá-Colombia. METHODOLOGY: A descriptive cross-sectional study with an analytical component was carried out on pregnant patients who utilized the ultrasound services at 2 Maternal-Fetal Medicine units in Bogotá, between 18 and 40 weeks of pregnancy who underwent measurement. From the anterior and posterior horns of the lateral ventricles, the fronto-occipital ratio was calculated at each gestational week, and nomograms were created for each of these variables. RESULTS: Nine hundred and seventy-eight patients were included in the study. The distance of the frontal horns ranged between 6.9 and 51.6 mm with a mean of 19.1 ± 5.8 mm; that of the occipital horns had a measurement between 8.7 and 53 mm with a mean of 28, 1 ± 8.9 mm; on the other hand, the fronto-occipital ratio (FOR) yielded a mean of 0.365 ± 0.067 (0.136-0.616) without bearing any relation to gestational age. The trend of normal values for the studied population is displayed, plotted in percentile curves and nomograms for each gestational age. CONCLUSION: The measurement of the frontal and occipital horns, and the calculation of the fronto-occipital relationship is technically possible between 18 and 40 weeks, finding that the anterior and posterior horns have a positive linear relationship with gestational age. Contrarily, the FOR does not correlate with the gestational age, it was possible to establish a table of percentiles that allows determining the normal values for these measurements during pregnancy.
Subject(s)
Fetus , Perinatology , Pregnancy , Female , Humans , Colombia , Reference Values , Cross-Sectional Studies , Fetus/diagnostic imaging , Gestational Age , Ultrasonography, PrenatalABSTRACT
PURPOSE: To evaluate histopathologic features of preretinal tissues removed from eyes with myopic traction maculopathy (MTM). METHODS: We retrospectively studied preretinal tissue specimens from eyes with MTM removed during pars plana vitrectomy. A control group of six idiopathic epiretinal membranes was studied for comparison. RESULTS: Six MTM specimens were studied histopathologically. Outer retinal schisis-like thickening was present in 100% of preoperative optical coherence tomography images; four of the six eyes had subfoveal neurosensory retinal detachment. Postoperative optical coherence tomography images demonstrated complete resolution of the schisis-like appearance in all eyes; a full-thickness macular hole occurred in two of the six eyes. Histopathologic examination disclosed fibrocellular tissue that was strongly positive for glial fibrillary acidic protein, weak to moderately positive for cytokeratin, and weakly positive for smooth muscle actin and CD68. There were no apparent histopathologic or immunohistochemical differences between preretinal tissues from eyes with MTM and idiopathic epiretinal membranes from control eyes. CONCLUSION: The outer retinal schisis-like thickening, commonly associated with subretinal fluid, that characterizes MTM is associated with preretinal tissues that are histopathologically indistinguishable from idiopathic epiretinal membranes. These findings suggest that anteroposterior traction caused by axial elongation rather than a uniquely abnormal cellular process is the etiologic mechanism of MTM.
Subject(s)
Macula Lutea/diagnostic imaging , Macular Degeneration/diagnosis , Myopia, Degenerative/complications , Visual Acuity , Aged , Aged, 80 and over , Female , Humans , Macular Degeneration/etiology , Macular Degeneration/surgery , Middle Aged , Retrospective Studies , Tomography, Optical Coherence/methods , VitrectomyABSTRACT
Rose Bengal Photodynamic Antimicrobial Therapy (RB-PDAT) is a novel potential treatment for progressive infectious keratitis. The principle behind this therapy is using Rose Bengal as a photosensitizer that can be activated by green light and results in the production of oxygen free radicals which in turn eradicate the microorganism. Given RB-PDAT's mechanism of action and the potential cytotoxic effects, concerns regarding the safety of this technique have arisen. The purpose of this study was to evaluate the effect of RB-PDAT on keratocytes, while focusing on the safety profile that the photo-chemical reaction has on the limbal stem cell (LSC) niche and endothelial cell layer of the treated cornea. To perform RB-PDAT, Rose Bengal solution (0.1% RB in BSS) was applied to the right cornea of rabbits for 30â¯min and then irradiated by a custom-made green LED light source (525â¯nm, 6â¯mW/cm2) for 15â¯min (5.4â¯J/cm2). Three rabbits were sacrificed and enucleated after 24â¯h for evaluation. TUNEL assay and immunohistochemistry for endothelium and limbal stem cell viability were performed on whole mounts and frozen sections in treated and control eyes. LSC of both eyes were isolated and cultured to perform MTT viability and proliferation, and scratch wound healing assays under time-lapse microscopy. Interestingly, while Rose Bengal dye penetration was superficial, yet associated cellular apoptosis was evidenced in up to 1/3 of the stromal thickness on frozen sections. TUNEL assay on whole mounts showed no endothelial cell death following treatment. Immunohistochemistry on frozen sections of LSC displayed no structural difference between treated and non-treated eyes. There was no difference in LSC proliferation rates and scratch wound healing assay demonstrated adequate cell migration from treated and non-treated eyes. The current study suggests that even though penetration of the RB dye has been shown to be limited, oxidative stress produced by RB-PDAT can reach deeper into the corneal stroma. Nevertheless, our results show that performing RB-PDAT is safe on the corneal endothelium and has no effect on LSC viability or function.
Subject(s)
Anti-Infective Agents/pharmacology , Corneal Keratocytes/drug effects , Endothelium, Corneal/drug effects , Fluorescent Dyes/pharmacology , Photochemotherapy , Rose Bengal/pharmacology , Stem Cell Niche/drug effects , Animals , Apoptosis , Biomarkers/metabolism , Cell Proliferation , Cell Survival , Cells, Cultured , Corneal Keratocytes/metabolism , Corneal Keratocytes/pathology , Corneal Stroma/drug effects , Corneal Stroma/metabolism , Corneal Stroma/pathology , Endothelium, Corneal/metabolism , Endothelium, Corneal/pathology , Immunohistochemistry , In Situ Nick-End Labeling , Limbus Corneae/drug effects , Limbus Corneae/metabolism , Limbus Corneae/pathology , RabbitsABSTRACT
G protein-coupled receptors (GPCRs) are integral membrane proteins involved in cellular signaling and constitute major drug targets. Despite their importance, the relationship between structure and function of these receptors is not well understood. In this study, the role of extracellular disulfide bonds on the trafficking and ligand-binding activity of the human A2A adenosine receptor was examined. To this end, cysteine-to-alanine mutations were conducted to replace individual and both cysteines in three disulfide bonds present in the first two extracellular loops. Although none of the disulfide bonds were essential for the formation of plasma membrane-localized active GPCR, loss of the disulfide bonds led to changes in the distribution of the receptor within the cell and changes in the ligand-binding affinity. These results indicate that in contrast to many class A GPCRs, the extracellular disulfide bonds of the A2A receptor are not essential, but can modulate the ligand-binding activity, by either changing the conformation of the extracellular loops or perturbing the interactions of the transmembrane domains.
Subject(s)
Alanine/chemistry , Cell Membrane/chemistry , Cysteine/chemistry , Disulfides/chemistry , Receptor, Adenosine A2A/chemistry , Alanine/metabolism , Amino Acid Sequence , Amino Acid Substitution , Cell Membrane/metabolism , Cysteine/metabolism , Gene Expression , HEK293 Cells , Humans , Kinetics , Ligands , Models, Molecular , Molecular Sequence Data , Protein Binding , Protein Structure, Secondary , Protein Transport , Receptor, Adenosine A2A/genetics , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Sequence Alignment , Structure-Activity RelationshipABSTRACT
The adenosine A2A receptor (A2AR) is a much-studied class A G protein-coupled receptor (GPCR). For biophysical studies, A2AR is commonly purified in a detergent mixture of dodecylmaltoside (DDM), 3-(3-cholamidopropyl) dimethylammoniopropane sulfonate (CHAPS), and cholesteryl hemisuccinate (CHS). Here we studied the effects of CHAPS on the ligand binding activity and stability of wild type, full-length human A2AR. We also tested the cholesterol requirement for maintaining the active conformation of the receptor when solubilized in detergent micelles. To this end, the receptor was purified using DDM, DDM/CHAPS, or the short hydrocarbon chain lipid 1,2-dihexanoyl-sn-glycero-3-phosphocholine (DHPC, di-6:0PC). After solubilization in DDM, DDM/CHAPS, or DHPC micelles, although A2AR was found to retain its native-like fold, its binding ability was significantly compromised compared to DDM or DDM/CHAPS with CHS. It therefore appears that although cholesterol is not needed for A2AR to retain a native-like, α-helical conformation, it may be a critical component for high affinity ligand binding. Further, this result suggests that the conformational differences between the active and inactive protein may be so subtle that commonly used spectroscopic methods are unable to differentiate between the two forms, highlighting the need for activity measurements. The studies presented in this paper also underline the importance of the protein's purification history; i.e., detergents that interact with the protein during purification affect the ligand binding properties of the receptor in an irreversible manner.
Subject(s)
Cholesterol Esters/chemistry , Cholic Acids/chemistry , Phospholipid Ethers/chemistry , Receptor, Adenosine A2A/chemistry , Receptor, Adenosine A2A/isolation & purification , Recombinant Proteins/chemistry , Humans , Protein Structure, SecondaryABSTRACT
Corneal crosslinking (CXL) represents a paradigm shift in the management of corneal ectatic disorders. Before CXL was introduced, patients would need specialty contact lenses and possible corneal transplantation. CXL involves a biochemical reaction in which ultraviolet A light is used in conjunction with Riboflavin to form crosslinks in between corneal stromal collagen. This leads to strengthening and stabilizing of the collagen lamellae, resulting in mechanical stiffening of the cornea. Multiple protocols have been proposed including epithelium on versus off and varying light intensity and duration of treatment. All protocols appear to be safe and effective with few reported complications including infection, stromal haze, scarring, and endothelial toxicity. Overall, CXL has demonstrated to halt the progression of the disease clinically and in keratometry readings and improve the quality of life for patients. It is a minimally invasive, cost-effective procedure that can be performed in an outpatient setting with a fast recovery time and long-lasting results.
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Purpose: To report a case of branch retinal artery occlusion (BRAO) followed by branch retinal vein occlusion (BRVO) and paracentral acute middle maculopathy (PAMM) in a patient with confirmed calciphylaxis. Observations: A 52-year-old female with a history of BRAO in the right eye one-year prior presented with decreased vision and a new inferotemporal scotoma. Computed tomography angiography of the head and neck demonstrated vascular calcifications at the origin of both ophthalmic arteries, which were otherwise poorly visualized. Ophthalmic examination demonstrated retinal whitening superiorly with intraretinal hemorrhages inferiorly. Optical coherence tomography (OCT) demonstrated middle retinal hyperreflectivity and a mild epiretinal membrane. Fluorescein angiography (FFA) demonstrated delayed perfusion of superior retinal arcade. On further questioning, patient was found to have a history of IgA nephropathy with end-stage renal disease, secondary hyperparathyroidism and calciphylaxis. Calciphylaxis is a systemic disease, characterized by high levels of calcium and progressive calcification of the vascular medial layer leading to ischemia. Anterior ischemic optic neuropathy (AION) and crystalline retinopathy have been reported as ocular manifestations of calciphylaxis, however, there are very few reports on ophthalmic manifestations of calciphylaxis. Conclusion and importance: Clinical manifestations of calciphylaxis are variable and a detailed clinical history is important to suspect calciphylaxis. Calciphylaxis should be considered in the differential diagnosis of BRAO, BRVO, PAMM or any ophthalmic vascular manifestation in patients with end-stage renal disease.
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PURPOSE: The purpose of this study was to report the indications, ocular and systemic comorbidities, and surgical outcomes of corneal transplantation in patients older than 90 years. METHODS: A retrospective review was conducted to identify individuals 90 years and older who underwent corneal transplantation surgery at the Bascom Palmer Eye Institute between January 2013 and October 2020. Outcomes included best-corrected visual acuity and graft survival over time. Paired t tests were used to compare visual acuity preoperatively versus postoperatively. Graft survival was evaluated with Kaplan-Meier curves. RESULTS: Fifty-eight eyes of 52 consecutive individuals were included. The mean age of individuals was 92 ± 2 years; 26.9% were male; and 48.1% self-identified as non-Hispanic White and 38.5% as Hispanic. Postoperative follow-up was 14.7 ± 12.1 months. Of the 58 eyes, 44.8% (26/58) underwent penetrating keratoplasty, 46.6% (27/58) Descemet stripping automated endothelial keratoplasty, and 6.9% (4/58) keratoprosthesis. All surgeries were performed under monitored local anesthesia, without major complications. Surgical indications included pseudophakic bullous keratopathy (36.2%), glaucoma-associated corneal decompensation (27.6%), Fuchs endothelial dystrophy (25.9%), and perforated corneal ulceration (19.0%). The best-corrected visual acuity improved by 0.32 (95% confidence interval 0.14-0.50; P < 0.01) as early as 1 month postoperatively, and vision gains were sustained for at least 12 months. Graft survival probability at 12 months was 88%. CONCLUSIONS: Corneal transplantation is a safe and successful procedure in restoring the visual acuity for patients older than 90 years after careful preoperative evaluation. Further research is needed to evaluate the impact of corneal transplantation on quality of life in patients in the 10th decade of life.
Subject(s)
Corneal Diseases , Descemet Stripping Endothelial Keratoplasty , Fuchs' Endothelial Dystrophy , Aged, 80 and over , Cornea , Corneal Diseases/surgery , Endothelium, Corneal/transplantation , Female , Fuchs' Endothelial Dystrophy/surgery , Humans , Male , Prostheses and Implants , Quality of Life , Retrospective Studies , Treatment OutcomeABSTRACT
Introduction: Uveal melanoma (UM) is the most common primary intraocular malignancy in adults, and despite treatment of the primary tumor, approximately 15%-50% of patients will develop metastatic disease. Based on gene expression profiling (GEPs), UM can be categorized as Class 1A (low metastatic risk), Class 1B (intermediate metastatic risk), or Class 2 (high metastatic risk). PReferentially expressed Antigen in MElanoma (PRAME) status is an independent prognostic UM biomarker and a potential target for immunotherapy in metastatic UM. PRAME expression status can be detected in tumors using reverse-transcription polymerase chain reaction (RT-PCR). More recently, immunohistochemistry (IHC) has been developed to detect PRAME protein expression. Here, we employed both techniques to evaluate PRAME expression in 18 UM enucleations. Methods: Tumor material from the 18 UM patients who underwent enucleation was collected by fine-needle aspiration before or during enucleation and sent for GEP and PRAME analysis by RT-PCR. Histologic sections from these patients were stained with an anti-PRAME monoclonal antibody. We collected patient demographics and tumor characteristics and included this with our analysis of GEP class, PRAME status by RT-PCR, and PRAME status by IHC. PRAME IHC and RT-PCR results were compared. Results: Twelve males (12/18) and 6 females (6/18) with an average age of 60.6 years underwent enucleation for UM. TNM staging of the UM diagnosed Stage I in 2 patients (2/18), Stage II in 7 patients (7/18), Stage III in 8 patients (8/18), and Stage IV in 1 (1/18). GEP was Class 1A in 6 tumors (6/18), Class 1B in 6 tumors (6/18), and Class 2 in 6 tumors (6/18). PRAME IHC showed diffusely positive labeling of all UM cells in 2/18 enucleations; negative IHC labeling of UM cells in 9/18 enucleations; and IHC labeling of subsets of UM cells in 7/18 enucleations. Eleven of the 17 UMs tested for PRAME by both RT-PCR and IHC had consistent PRAME results. In the remaining 6/17 cases tested by both modalities, PRAME results were discordant between RT-PCR and IHC. Conclusions: We find that PRAME IHC distinguishes PRAME-positive and PRAME-negative UM tumor cells. Interestingly, IHC reveals focal PRAME expression in subsets of tumor cells consistent with tumor heterogeneity. PRAME RT-PCR and IHC provide concordant results in most of our cases. We suggest that discordance in PRAME results could arise from spatial or temporal variation in PRAME expression between tumor cells. Further studies are required to determine the prognostic implications of PRAME IHC in UM.
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Purpose: Conjunctival squamous cell carcinoma (SCC) is a sight-threatening ocular surface malignancy with the primary treatment modality being surgical resection. To evaluate surgical imaging modalities to improve surgical resection, we established a novel murine model for conjunctival SCC to demonstrate the utility of panitumumab-IRDye800, a fluorescently labeled anti-epidermal growth factor receptor (EGFR) antibody. Methods: NOD-scid IL2Rgammanull (NSG) mice received subconjunctival injection of UM-SCC-1 or SCC-9, head and neck SCC cell lines. On tumor growth, mice were injected with Panitumumab-IRDye800CW, and imaged with a small animal imaging system and optical coherence tomography (OCT). Immunohistochemistry for SCC markers were used to confirm tumor origin. Results: Seventy-five percent (N = 4) of the UM-SCC-1 group developed aggressive, rapidly growing tumors that were P40 and EGFR positive within two weeks of inoculation. The SCC-9 tumors failed to demonstrate any growth (N = 4). Ocular tumors demonstrated high fluorescence levels with a tumor to background ratio of 3.8. Conclusions: Subconjunctival injections are an appropriate technique to create in vivo models for assessing treatment modalities and novel therapies in conjunctival SCC. Translational Relevance: This model demonstrates Panitumumab-IRDye800CW's utility in the ophthalmic setting and suggests that clinical trials may be warranted.
Subject(s)
Conjunctival Neoplasms , Head and Neck Neoplasms , Animals , Antibodies, Monoclonal/therapeutic use , Cell Line, Tumor , Conjunctival Neoplasms/drug therapy , Disease Models, Animal , ErbB Receptors/metabolism , Head and Neck Neoplasms/drug therapy , Mice , Mice, Inbred NOD , Panitumumab , Squamous Cell Carcinoma of Head and Neck/drug therapyABSTRACT
γD crystallin is a natively monomeric eye-lens protein that is associated with hereditary juvenile cataract formation. It is an attractive model system as a multidomain Greek-key protein that aggregates through partially folded intermediates. Point mutations M69Q and S130P were used to test (1) whether the protein-design algorithm RosettaDesign would successfully predict mutants that are resistant to aggregation when combined with informatic sequence-based predictors of peptide aggregation propensity and (2) how the mutations affected relative unfolding free energies (ΔΔG(un)) and intrinsic aggregation propensity (IAP). M69Q was predicted to have ΔΔG(un) â« 0, without significantly affecting IAP. S130P was predicted to have ΔΔG(un) â¼ 0 but with reduced IAP. The stability, conformation, and aggregation kinetics in acidic solution were experimentally characterized and compared for the variants and wild-type (WT) protein using circular dichroism and intrinsic fluorescence spectroscopy, calorimetric and chemical unfolding, thioflavin-T binding, chromatography, static laser light scattering, and kinetic modeling. Monomer secondary and tertiary structures of both variants were indistinguishable from WT, while ΔΔG(un) > 0 for M69Q and ΔΔG(un) < 0 for S130P. Surprisingly, despite being the least conformationally stable, S130P was the most resistant to aggregation, indicating a significant decrease of its IAP compared to WT and M69Q.
Subject(s)
Point Mutation , gamma-Crystallins/chemistry , gamma-Crystallins/genetics , Amino Acid Sequence , Circular Dichroism , Computer-Aided Design , Humans , Kinetics , Molecular Sequence Data , Protein Conformation , Protein Folding , Protein Stability , Thermodynamics , gamma-Crystallins/metabolismABSTRACT
Purpose: To report our experience with oral miltefosine (MLT) as an adjunct treatment for progressive Acanthamoeba keratitis (AK).Methods: Retrospective case series of all patients who underwent treatment with oral MLT for AK at Bascom Palmer Eye Institute from 2017 to 2020.Results: Six females from 16 to 55 years old, with a microbiologic diagnosis of Acanthamoeba, were treated with MLT and standard medical treatment. Four of the six cases deteriorated after initiating treatment and three required a therapeutic keratoplasty. Two patients improved after 1 week of MLT and optical penetrating keratoplasty was performed after clinical resolution. Microbiologic culture of corneal buttons was negative in all cases. All corneal grafts remain clear at last follow-up with best-corrected visual acuity of 20/40 or better.Conclusion: Oral MLT may be a viable adjunctive therapy for recalcitrant AK; however, its use may be associated with a severe inflammatory reaction. Further studies are needed to evaluate its efficacy and variable clinical response.
Subject(s)
Acanthamoeba Keratitis/drug therapy , Antiprotozoal Agents/therapeutic use , Phosphorylcholine/analogs & derivatives , Acanthamoeba Keratitis/diagnosis , Acanthamoeba Keratitis/etiology , Administration, Oral , Adolescent , Adult , Contact Lenses/adverse effects , Female , Follow-Up Studies , Humans , Keratoplasty, Penetrating , Middle Aged , Phosphorylcholine/therapeutic use , Retrospective Studies , Risk Factors , Visual Acuity/physiology , Young AdultABSTRACT
PURPOSE: To provide clinicopathologic correlations for retrocorneal membranes associated with Descemet stripping automated endothelial keratoplasty (DSAEK) failure. DESIGN: Retrospective case series. METHODS: The specimens and medical records of the patients diagnosed with clinically significant retrocorneal membranes associated with DSAEK failure at the Bascom Palmer Eye Institute or the University of Miami Veterans Hospital between October 2015 and March 2020 were reviewed for demographics, clinical presentation, comorbidities, and surgeries performed. Histopathologic analysis was performed on hematoxylin-eosin and periodic acid-Schiff sections. Immunohistochemical studies were performed for smooth muscle actin (α-SMA), pancytokeratin, and CK7. Immunofluorescence was performed for vimentin, N-cadherin, ROCK1, RhoA, ZEB1, and Snail. RESULTS: A total of 7 patients (3 male and 4 female) were identified to have a clinically significant retrocorneal membranes at the time of graft failure. The average age at the time of first DSAEK was 70 years (range: 55-85 years). All patients were pseudophakic and had a glaucoma drainage device in place; 1 had a history of failed DSAEK. Ranging from 0 to 47 months after surgery, a variably thick retrocorneal fibrous membrane was observed, eventually leading to graft failure. Four patients underwent subsequent penetrating keratoplasty and 3 underwent repeat DSAEK. On histopathologic evaluation, a pigmented fibrocellular tissue was identified along the posterior margin of the corneas and DSAEK buttons in all cases. Further characterization with immunohistochemistry and immunofluorescence demonstrated membranes to be negative for pancytokeratin and positive for α-SMA, vimentin, CK7, N-cadherin, ZEB1, Snail, ROCK1, and RhoA. CONCLUSIONS: Fibrocellular retrocorneal membrane proliferation may be associated with DSAEK failure in patients with previous glaucoma drainage device surgery. Our results demonstrate myofibroblastic differentiation and a lack of epithelial differentiation. Positivity for markers of an endothelial-to-mesenchymal transition indicates possible endothelial origin and could be the hallmark for future targeted pharmacotherapy.
Subject(s)
Cornea/pathology , Corneal Diseases/surgery , Descemet Stripping Endothelial Keratoplasty/adverse effects , Graft Rejection/diagnosis , Visual Acuity , Aged , Aged, 80 and over , Corneal Diseases/diagnosis , Female , Graft Survival , Humans , Male , Middle Aged , Retrospective StudiesABSTRACT
BACKGROUND: Preeclampsia is a multiorgan disorder associated with maternal and perinatal morbi-mortality. In Peru, incidence is 10% and accounts for 22% of maternal deaths. Genome and genetic epidemiological studies have found an association between preeclampsia and genetic polymorphisms. OBJECTIVE: To determine the association of the vascular endothelial growth factor (VEGF) +936 C/T and +405 G/C, interleukine-6 (IL-6) -174 G/C, IL-1ß-511 C/T, Apo A-1-75 G/A, Apo B-100 2488 C/T (Xbal) polymorphisms with preeclampsia in pregnant Peruvian women. METHODS: Were included preeclamptic and healthy (control) pregnant women. Maternal blood samples were subjected to DNA extraction, and molecular genetic analysis was conducted using the PCR-RFLP technique and following a specific protocol for each gene. Allele and genotypic frequencies in the cases and controls were compared. RESULTS: No association was found between the VEGF+936C/T and VEGF+405 polymorphisms and preeclampsia. The frequencies of the GG genotypes and the G allele of the -174 G/C polymorphism in the IL6 gene in preeclamptic and controls showed significant differences, with higher frequencies in cases. For the -511 C/T polymorphism of the IL-1ß gene, no significant differences were found in the frequencies of TT genotypes compared with CT+CC. The genotypes and alleles of the Apo-A1-75 G/A and Apo-B100 Xbal variants showed no significant differences between cases and controls. CONCLUSION: No association was found between the studied genetic markers and preeclampsia. However, in the -174G/C polymorphism of the IL-6 gene, significant differences were found mainly in the GG genotype and G allele.
Subject(s)
Pre-Eclampsia , Case-Control Studies , Female , Gene Frequency , Genetic Markers , Genetic Predisposition to Disease , Genotype , Humans , Peru/epidemiology , Polymorphism, Single Nucleotide , Pre-Eclampsia/epidemiology , Pre-Eclampsia/genetics , Pregnancy , Vascular Endothelial Growth Factor A/geneticsABSTRACT
PURPOSE: To evaluate the in vivo corneal changes after Rose Bengal photodynamic antimicrobial therapy (RB-PDAT) treatment in New Zealand White rabbits. METHODS: Sixteen rabbits were divided into 5 groups. All groups underwent deepithelialization of an 8 mm diameter area in the central cornea. Group 1: balanced salt solution drops only, group 2: 0.2% RB only, group 3: green light exposure (525 nm, 5.4 J/cm2) only, group 4: 0.1% RB-PDAT, and group 5: 0.1% RB-PDAT. All rabbits were followed clinically. Group 5 rabbits were followed using anterior segment optical coherence tomography (AS-OCT) and clinically. On day 35 after initial treatment, 1 rabbit from group 5 was re-exposed to green light (5.4 J/cm2) to evaluate reactivation of the remaining RB dye, and terminal deoxynucleotyl transferase-mediated UTP-biotin-nick-end labeling assay was performed on corneal cryosections. RESULTS: Complete reepithelization was observed, and corneas remained clear after treatment in all groups. In group 5, AS-OCT revealed a cross-linking demarcation line. AS-OCT showed RB fluorescence and collagen cross-linking in all treated eyes of group 5 animals after 5 weeks of treatment. Photobleached RB retention in the corneal stroma was corroborated by fluorescence confocal microscopy on frozen sections. There was no evidence of a sustained cytotoxic effect through terminal deoxynucleotyl transferase-mediated UTP-biotin-nick-end labeling at 5 weeks. CONCLUSIONS: RB-PDAT with 0.1% RB is a safe procedure. There was no difference clinically and on histopathology compared with control groups. In eyes where RB dye is retained in the corneal stroma after 1 month of treatment, oxidative stress is not evidenced at long term.
Subject(s)
Collagen/therapeutic use , Cross-Linking Reagents/pharmacology , Eye Infections, Bacterial/drug therapy , Keratitis/drug therapy , Photochemotherapy/methods , Photosensitizing Agents/therapeutic use , Rose Bengal/therapeutic use , Animals , Disease Models, Animal , Eye Infections, Bacterial/diagnosis , Female , Fluorescent Dyes/therapeutic use , Keratitis/diagnosis , Microscopy, Confocal , Pilot Projects , Rabbits , Tomography, Optical Coherence/methodsABSTRACT
G protein-coupled receptors (GPCRs) constitute the largest family of integral membrane proteins present in all eukaryotic cells, yet relatively little information about their structure, folding, and stability has been published. In this work, we describe several approaches to characterizing the conformational stability of the human adenosine A(2)a receptor (hA(2)aR). Thermal denaturation and chemical denaturation were not reversible, yet clear differences in the unfolding behavior were observed upon ligand binding via circular dichroism and fluorescence spectrometry. We found that the stability of hA(2)aR was increased upon incubation with the agonist N(6)-cyclohexyladenosine or the antagonist theophylline. When extracellular disulfide bonds were reduced with a chemical reducing agent, the ligand binding activity decreased by ~40%, but reduction of these bonds did not compromise the unfolding transition observed via urea denaturation. Overall, these approaches offer a general strategy for characterizing the effect of surfactant and ligand effects on the stability of GPCRs.
Subject(s)
Disulfides/pharmacology , Receptors, Adenosine A2/chemistry , Adenosine/analogs & derivatives , Adenosine/pharmacology , Humans , Ligands , Protein Denaturation , Protein Stability/drug effects , Receptors, Adenosine A2/metabolism , Receptors, G-Protein-Coupled , Theophylline/pharmacologyABSTRACT
PURPOSE: To describe optical coherence tomography angiography (OCTA) findings in a patient with aniridia and correlate with representative histopathology. OBSERVATIONS: OCTA images of the macula of a pediatric aniridic patient, who has nystagmus and impaired vision bilaterally, demonstrate a complete absence of the foveal avascular zone (FAZ) in both the superficial and deep vascular complexes (SVC and DVC). In addition, larger superficial blood vessels were found to be abnormally diving from the SVC into the DVC. Similarly, immunofluorescence with confocal microscopy imaging of a retinal histopathology specimen from a 2 month old aniridic patient demonstrated larger vessels diving in the same manner. CONCLUSIONS AND IMPORTANCE: This study highlights the clinical, imaging and histopathologic findings of aniridia. Supine OCTA imaging, performed during examination under anesthesia, allowed for visualization of retinal microvasculature in eyes with nystagmus. The histopathology images helped validate OCTA findings that, with further investigation, may lead to new information about the development of abnormal retinal microvasculature.
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A role for the adhesion G-protein coupled receptor ADGRE2 or EMR2 in mechanosensing was revealed by the finding of a missense substitution (p.C492Y) associated with familial vibratory urticaria. In these patients, friction of the skin induces mast cell hyper-degranulation through p.C492Y-ADGRE2, causing localized hives, flushing, and hypotension. We have now characterized the responses and intracellular signals elicited by mechanical activation in human mast cells expressing p.C492Y-ADGRE2 and attached to dermatan sulfate, a ligand for ADGRE2. The presence of p.C492Y-ADGRE2 reduced the threshold to activation and increased the extent of degranulation along with the percentage of mast cells responding. Vibration caused phospholipase C activation, transient increases in cytosolic calcium, and downstream activation of phosphoinositide 3-kinase and extracellular signal-regulated kinases 1 and 2 by Gßγ, Gαq/11, and Gαi/o-independent mechanisms. Degranulation induced by vibration was dependent on phospholipase C pathways, including calcium, protein kinase C, and phosphoinositide 3-kinase but not extracellular signal-regulated kinases 1/2 pathways, along with pertussis toxin-sensitive signals. In addition, mechanoactivation of mast cells stimulated the synthesis and release of prostaglandin D2, to our knowledge a previously unreported mediator in vibratory urticaria, and extracellular signal-regulated kinases 1/2 activation was required for this response together with calcium, protein kinase C, and to some extent, phosphoinositide 3-kinase. Our studies thus identified critical molecular events initiated by mechanical forces and potential therapeutic targets for patients with vibratory urticaria.
Subject(s)
Mast Cells/physiology , Receptors, G-Protein-Coupled/genetics , Urticaria/etiology , Calcium/metabolism , Cell Degranulation , Cells, Cultured , Extracellular Signal-Regulated MAP Kinases/physiology , Humans , Mechanotransduction, Cellular , Mutation, Missense , Phosphatidylinositol 3-Kinases/physiology , Prostaglandin D2/physiology , Protein Kinase C/physiology , Receptors, G-Protein-Coupled/physiology , Signal Transduction/physiology , Tetraspanin 30/physiology , Type C Phospholipases/physiology , Urticaria/genetics , Vibration/adverse effectsABSTRACT
Identification of molecular targets is the first step in developing efficacious therapeutic strategies for tumors. A tumors' biological response to perturbagens yields important information on the molecular determinants for tumor growth. The aim of this study was to characterize the response of adenoid cystic carcinoma of the lacrimal gland (LGACC) to intra-arterial cytoreductive chemotherapy (IACC) in order to identify novel targets to enhance therapy. We performed high-throughput proteomic analysis on paired samples from pre-IACC diagnostic biopsies and post-IACC excised tumor samples from 6 LGACC patients. This proteomic analysis provides a comprehensive landscape of the cellular compartments contained within the excised tumors. Interestingly, we found a strong upregulation across the fibroblast growth factor (FGF) signaling pathway, with FGF receptor 1 (FGFR1) exhibiting a consistent and significant upregulation in all post-IACC samples. We thus evaluated the therapeutic efficacy of a novel FGFR1 selective inhibitor, AZD4547, in combination with cisplatin on LGACC cells in-vitro. The combination index (CI) value (<0.895) demonstrated synergistic effect of AZD4547 and cisplatin in inhibiting cell growth and viability (p<0.02), with a differential response seen in post-IACC cultures when compared to pre-IACC cultures. The combination approach showed synergy of the drugs in the migration assay. Western blot analysis indicated a significant upregulation of cleaved caspase-3 and downregulation the expression of FGFR1 (p<0.05) with the combination treatment as compared to either agent independently. Our findings demonstrate that FGFR1 inhibition potentiates the cytoreductive effects of cisplatin and suggest a potential therapeutic benefit of using AZD4547 in the management of LGACC.
ABSTRACT
Toxic anterior segment syndrome is a surgical complication characterized by a noninfectious anterior chamber inflammatory reaction having multiple etiologies. The clinical signs (prominent limbus-to-limbus corneal edema, anterior chamber inflammation) and symptoms (decreased visual acuity, discomfort) generally occur within the first 12-48 hours after intraocular surgery. Most patients achieve good clinical and visual outcomes when there is a prompt clinical diagnosis and adequate treatment. We review the literature on toxic anterior segment syndrome, emphasizing its etiology, pathophysiology, and clinical and surgical management, as well as prognosis and sequelae. Our goal is to reduce the frequency of toxic anterior segment syndrome by highlighting the importance of prevention, early recognition, and distinguishing toxic anterior segment syndrome from infectious endophthalmitis.