Increased Choroidal Neovascularization following Laser Induction in Mice Lacking Lysyl Oxidase-like 1

PURPOSE. Age-related degradation of the elastic lamina in Bruch's membrane may have a permissive effect on the growth of choroidal neovascularization (CNV). This study investigated the influence of defective elastic fiber maintenance in the development of laser-induced CNV. METHODS. A mouse lacking lysyl oxidase-like (LOXL)-1, an enzyme essential for elastin polymerization, was studied. The morphologic characteristics of the elastic lamina within Bruch's membrane were examined in mutant and wild-type (WT) eyes. Laser-induced CNV was evaluated by fluorescein angiography and choroidal flat mounts. Immunohistochemistry for elastin was performed on the CNV lesions, and vascular endothelial growth factor (VEGF) levels were determined by ELISA. Soluble elastin and matrix metalloproteinase (MMPs) levels were also analyzed by immunoblotting. RESULTS. The elastic lamina of Bruch's membrane in the LOXL1-deficient mice was fragmented and less continuous than in the WT controls. The mutant mice showed increased levels of soluble elastin peptides and reduced elastin polymer deposition in neovascular membranes. Significantly larger CNV with greater leakage on fluorescein angiography developed in mutant mice. VEGF levels in the RPE/choroid were higher in the knockout mice on days 7 and 14 after laser (P Ͻ 0.05). MT1-MMP (MMP14) was also elevated after laser in the LOXL1 mutant eyes compared to the WT controls. CONCLUSIONS. These results show that a systemic defect in elastic fiber deposition affects Bruch's membrane integrity and leads to more aggressive CNV growth. The latter may be partially mediated by abnormal signaling from the accumulation of soluble elastin peptides. (Invest Ophthalmol Vis Sci. 2008;49:2599 -2605) DOI:10.1167/iovs.07-1508 C horoidal neovascularization (CNV) is the predominant cause of severe visual loss in age-related macular degeneration (AMD) and other macular diseases. Bruch's membrane integrity is known to be compromised in many conditions characterized by choroidal neovascularization, such as traumatic choroidal rupture, angioid streaks, and myopia. Additionally, laser rupture of Bruch's membrane is a well-established means of inducing experimental choroidal neovascularization. Such evidence indicates that Bruch's membrane serves as an important barrier to CNV formation, but the structural characteristics important in this barrier function are still under investigation. Bruch's membrane lies between the neural retina and the choriocapillaris. There are five layers to this structure: the basement membrane of the retinal pigment epithelium, an inner collagenous layer, the central elastic lamina composed of elastic fibers, the outer collagenous layer, and the basement membrane of the choriocapillaris endothelium. Numerous morphologic changes in Bruch's membrane have been described with aging, particularly the development of drusen and basal laminar deposits, but also calcification, changes in thickness, and decreases in hydraulic conductivity. 1 Loss of elastic fibers is a hallmark of connective tissue aging, and the central elastic lamina of Bruch's membrane has long been postulated to have a barrier function against CNV. 2,3 Recently, Chong et al. 5 Elastic fibers are amorphous polymers composed of the protein elastin, known as tropoelastin in its monomeric form. Oxidative deamination of lysine residues, an initial step of elastin polymerization, requires lysyl oxidases. 6 Lysyl oxidaselike (LOXL) protein 1 has been shown to guide the spatially defined deposition of elastin and is essential for the maintenance of elastic fibers. 7 LOXL1-deficient mice develop multiple systemic defects including pelvic organ prolapse, emphysematous changes in the lungs, and vascular abnormalities attributable to a failure in elastic fiber maintenance

Sex difference and intra-operative tidal volume: Insights from the LAS VEGAS study

BACKGROUND: One key element of lung-protective ventilation is the use of a low tidal volume (VT). A sex difference in use of low tidal volume ventilation (LTVV) has been described in critically ill ICU patients.OBJECTIVES: The aim of this study was to determine whether a sex difference in use of LTVV also exists in operating room patients, and if present what factors drive this difference.DESIGN, PATIENTS AND SETTING: This is a posthoc analysis of LAS VEGAS, a 1-week worldwide observational study in adults requiring intra-operative ventilation during general anaesthesia for surgery in 146 hospitals in 29 countries.MAIN OUTCOME MEASURES: Women and men were compared with respect to use of LTVV, defined as VT of 8 ml kg-1 or less predicted bodyweight (PBW). A VT was deemed 'default' if the set VT was a round number. A mediation analysis assessed which factors may explain the sex difference in use of LTVV during intra-operative ventilation.RESULTS: This analysis includes 9864 patients, of whom 5425 (55%) were women. A default VT was often set, both in women and men; mode VT was 500 ml. Median [IQR] VT was higher in women than in men (8.6 [7.7 to 9.6] vs. 7.6 [6.8 to 8.4] ml kg-1 PBW, P < 0.001). Compared with men, women were twice as likely not to receive LTVV [68.8 vs. 36.0%; relative risk ratio 2.1 (95% CI 1.9 to 2.1), P < 0.001]. In the mediation analysis, patients' height and actual body weight (ABW) explained 81 and 18% of the sex difference in use of LTVV, respectively; it was not explained by the use of a default VT.CONCLUSION: In this worldwide cohort of patients receiving intra-operative ventilation during general anaesthesia for surgery, women received a higher VT than men during intra-operative ventilation. The risk for a female not to receive LTVV during surgery was double that of males. Height and ABW were the two mediators of the sex difference in use of LTVV.TRIAL REGISTRATION: The study was registered at Clinicaltrials.gov, NCT01601223

Generating Recombinant Antibodies to Membrane Proteins through Phage Display

One of the most important classes of proteins in terms of drug targets is cell surface membrane proteins, and yet it is a challenging set of proteins for generating high-quality affinity reagents. In this review, we focus on the use of phage libraries, which display antibody fragments, for generating recombinant antibodies to membrane proteins. Such affinity reagents generally have high specificity and affinity for their targets. They have been used for cell staining, for promoting protein crystallization to solve three-dimensional structures, for diagnostics, and for treating diseases as therapeutics. We cover publications on this topic from the past 10 years, with a focus on the various formats of membrane proteins for affinity selection and the diverse affinity selection strategies used. Lastly, we discuss the challenges faced in this field and provide possible directions for future efforts

Generating Recombinant Antibodies to Membrane Proteins through Phage Display

One of the most important classes of proteins in terms of drug targets is cell surface membrane proteins, and yet it is a challenging set of proteins for generating high-quality affinity reagents. In this review, we focus on the use of phage libraries, which display antibody fragments, for generating recombinant antibodies to membrane proteins. Such affinity reagents generally have high specificity and affinity for their targets. They have been used for cell staining, for promoting protein crystallization to solve three-dimensional structures, for diagnostics, and for treating diseases as therapeutics. We cover publications on this topic from the past 10 years, with a focus on the various formats of membrane proteins for affinity selection and the diverse affinity selection strategies used. Lastly, we discuss the challenges faced in this field and provide possible directions for future efforts

Increased choroidal neovascularization following laser induction in mice lacking lysyl oxidase-like 1

PURPOSE: Age-related degradation of the elastic lamina in Bruch's membrane may have a permissive effect on the growth of choroidal neovascularization (CNV). This study investigated the influence of defective elastic fiber maintenance in the development of laser-induced CNV. METHODS: A mouse lacking lysyl oxidase-like (LOXL)-1, an enzyme essential for elastin polymerization, was studied. The morphologic characteristics of the elastic lamina within Bruch's membrane were examined in mutant and wild-type (WT) eyes. Laser-induced CNV was evaluated by fluorescein angiography and choroidal flat mounts. Immunohistochemistry for elastin was performed on the CNV lesions, and vascular endothelial growth factor (VEGF) levels were determined by ELISA. Soluble elastin and matrix metalloproteinase (MMPs) levels were also analyzed by immunoblotting. RESULTS: The elastic lamina of Bruch's membrane in the LOXL1-deficient mice was fragmented and less continuous than in the WT controls. The mutant mice showed increased levels of soluble elastin peptides and reduced elastin polymer deposition in neovascular membranes. Significantly larger CNV with greater leakage on fluorescein angiography developed in mutant mice. VEGF levels in the RPE/choroid were higher in the knockout mice on days 7 and 14 after laser (P < 0.05). MT1-MMP (MMP14) was also elevated after laser in the LOXL1 mutant eyes compared to the WT controls. CONCLUSIONS: These results show that a systemic defect in elastic fiber deposition affects Bruch's membrane integrity and leads to more aggressive CNV growth. The latter may be partially mediated by abnormal signaling from the accumulation of soluble elastin peptides

Inferring model parameters in network-based disease simulation

Stochastic simulation, Network dynamics, Concurrent partnerships, Sexually transmitted diseases,