Broad Spectrum Antiangiogenic Treatment for Ocular Neovascular Diseases

Pathological neovascularization is a hallmark of late stage neovascular (wet) age-related macular degeneration (AMD) and the leading cause of blindness in people over the age of 50 in the western world. The treatments focus on suppression of choroidal neovascularization (CNV), while current approved therapies are limited to inhibiting vascular endothelial growth factor (VEGF) exclusively. However, this treatment does not address the underlying cause of AMD, and the loss of VEGF''s neuroprotective can be a potential side effect. Therapy which targets the key processes in AMD, the pathological neovascularization, vessel leakage and inflammation could bring a major shift in the approach to disease treatment and prevention. In this study we have demonstrated the efficacy of such broad spectrum antiangiogenic therapy on mouse model of AMD.

Methods and Findings

Lodamin, a polymeric formulation of TNP-470, is a potent broad-spectrum antiangiogenic drug. Lodamin significantly reduced key processes involved in AMD progression as demonstrated in mice and rats. Its suppressive effects on angiogenesis, vascular leakage and inflammation were studied in a wide array of assays including; a Matrigel, delayed-type hypersensitivity (DTH), Miles assay, laser-induced CNV and corneal micropocket assay. Lodamin significantly suppressed the secretion of various pro-inflammatory cytokines in the CNV lesion including monocyte chemotactic protein-1 (MCP-1/Ccl2). Importantly, Lodamin was found to regress established CNV lesions, unlike soluble fms-like tyrosine kinase-1 (sFlk-1). The drug was found to be safe in mice and have little toxicity as demonstrated by electroretinography (ERG) assessing retinal and by histology.

Conclusions

Lodamin, a polymer formulation of TNP-470, was identified as a first in its class, broad-spectrum antiangiogenic drug that can be administered orally or locally to treat corneal and retinal neovascularization. Several unique properties make Lodamin especially beneficial for ophthalmic use. Our results support the concept that broad spectrum antiangiogenic drugs are promising agents for AMD treatment and prevention.     

Lactobacillus casei Abundance Is Associated with Profound Shifts in the Infant Gut Microbiome

Colonization of the infant gut by microorganisms over the first year of life is crucial for development of a balanced immune response. Early alterations in the gastrointestinal microbiota of neonates has been linked with subsequent development of asthma and atopy in older children. Here we describe high-resolution culture-independent analysis of stool samples from 6-month old infants fed daily supplements of Lactobacillus casei subsp. Rhamnosus (LGG) or placebo in a double-blind, randomized Trial of Infant Probiotic Supplementation (TIPS). Bacterial community composition was examined using a high-density microarray, the 16S rRNA PhyloChip, and the microbial assemblages of infants with either high or low LGG abundance were compared. Communities with high abundance of LGG exhibited promotion of phylogenetically clustered taxa including a number of other known probiotic species, and were significantly more even in their distribution of community members. Ecologically, these aspects are characteristic of communities that are more resistant to perturbation and outgrowth of pathogens. PhyloChip analysis also permitted identification of taxa negatively correlated with LGG abundance that have previously been associated with atopy, as well as those positively correlated that may prove useful alternative targets for investigation as alternative probiotic species. From these findings we hypothesize that a key mechanism for the protective effect of LGG supplementation on subsequent development of allergic disease is through promotion of a stable, even, and functionally redundant infant gastrointestinal community.     

Cardiovascular response to functional electrical stimulation and dynamic tilt table therapy to improve orthostatic tolerance

Orthostatic hypotension is a common condition for individuals with stroke or spinal cord injury. The inability to regulate the central nervous system will result in pooling of blood in the lower extremities leading to orthostatic intolerance. This study compared the use of functional electrical stimulation (FES) and passive leg movements to improve orthostatic tolerance during head-up tilt. Four trial conditions were assessed during head-up tilt: (1) rest, (2) isometric FES of the hamstring, gastrocnemius and quadriceps muscle group, (3) passive mobilization using the Erigo dynamic tilt table; and (4) dynamic FES (combined 2 and 3). Ten healthy male subjects experienced 70 degrees head-up tilt for 15 min under each trial condition. Heart rate, blood pressure and abdominal echograms of the inferior vena cava were recorded for each trial. Passive mobilization and dynamic FES resulted in an increase in intravascular blood volume, while isometric FES only resulted in elevating heart rate. No significant differences in blood pressure were observed under each condition. We conclude that FES combined with passive stepping movements may be an effective modality to increase circulating blood volume and thereby tolerance to postural hypotension in healthy subjects.     

5'HS5 of the human beta-globin locus control region is dispensable for the formation of the beta-globin active chromatin hub

Hypersensitive site 5 (5'HS5) of the beta-globin Locus Control Region functions as a developmental stage-specific border in erythroid cells. Here, we have analyzed the role of 5'HS5 in the three dimensional organization of the beta-gene locus using the Chromatin Conformation Capture (3C) technique. The results show that when 5'HS5 is deleted from the locus, both remote and internal regulatory elements are still able to interact with each other in a three-dimensional configuration termed the Active Chromatin Hub. Thus, the absence of 5'HS5 does not have an appreciable effect on the three dimensional organization of the beta-globin locus. This rules out models in which 5'HS5 nucleates interactions with remote and/or internal regulatory elements. We also determined the binding of CTCF, the only defined insulator protein in mammalian cells, to 5'HS5 by using chromatin immunoprecipitation (ChIP) assays. We detect low levels of CTCF binding to 5'HS5 in primitive erythroid cells, in which it functions as a border element. Surprisingly, we also observe binding levels of CTCF to 5'HS5 in definitive erythroid cells. Thus, binding of CTCF to 5'HS5 per se does not render it a functional border element. This is consistent with the previous data suggesting that CTCF has dual functionality.     

Subfailure damage in ligament: a structural and cellular evaluation.

Subfailure damage in ligaments was evaluated macroscopically from a structural perspective (referring to the entire ligament as a structure) and microscopically from a cellular perspective. Freshly harvested rat medial collateral ligaments (MCLs) were used as a model in ex vivo experiments. Ligaments were preloaded with 0.1 N to establish a consistent point of reference for length (and strain) measurements. Ligament structural damage was characterized by nonrecoverable difference in tissue length after a subfailure stretch. The tissue's mechanical properties (via stress vs. strain curves measured from a preloaded state) after a single subfailure stretch were also evaluated (n = 6 pairs with a different stretch magnitude applied to each stretched ligament). Regions containing necrotic cells were used to characterize cellular damage after a single stretch. It should be noted that the number of damaged cells was not quantified and the difference between cellular area and area of fluorescence is not known. Structural and cellular damage were represented and compared as functions of subfailure MCL strains. Statistical analysis indicated that the onset of structural damage occurs at 5.14% strain (referenced from a preloaded length). Subfailure strains above the damage threshold changed the shape of the MCL stress-strain curve by elongating the toe region (i.e., increasing laxity) as well as decreasing the tangential modulus and ultimate stress. Cellular damage was induced at ligament strains significantly below the structural damage threshold. This cellular damage is likely to be part of the natural healing process in mildly sprained ligaments.     

Asymmetric transformation of enol acetates with esterases from Marchantia polymorpha

Two esterases catalyzing the asymmetric hydrolysis of enol acetates to give optically active -alkylated ketones were isolated from cultured cells of Marchantia polymorpha by a three-step procedure: hydrophobic chromatography, anion exchange chromatography and gel-filtration chromatography. These esterases had opposite stereoselectivities on protonation of the enol intermediate in the hydrolysis and one of them obviously reversed the stereoselectivity when the chain length and the bulkiness of substituents at the β-position to the acetoxyl group were increased. The internal amino acid sequences of peptide fragments obtained by the proteolysis of the esterases with lysyl endopeptidase had no similarity to those of other hydrolytic enzymes.     

Wnt-10b, uniquely among Wnts, promotes epithelial differentiation and shaft growth

Although Wnts are expressed in hair follicles throughout life from embryo to adult, and considered to be critical for their development and maturation, their roles remain largely unknown. In the present study, we investigated the effects of Wnts (Wnt-3a, Wnt-5a, Wnt-10b, and Wnt-11) on epithelial cell differentiation using adult mouse-derived primary skin epithelial cell (MPSEC) cultures and hair growth using hair follicle organ cultures. Only Wnt-10b showed evident promotion of epithelial cell differentiation and hair shaft growth, in contrast to Wnt-3a, 5a, and 11. Our results suggest that Wnt-10b is unique and plays an important role in differentiation of epithelial cells in the hair follicle.     

A viable Bacillus subtilis strain without functional extracytoplasmic function sigma genes

We constructed a Bacillus subtilis Marburg strain that harbors deletion mutations in all seven extracytoplasmic function (ECF) sigma genes. The strain shows wild-type growth at 37 degrees C both in a complex and in a synthetic medium and exhibits wild-type sporulation. ECF sigma genes of B. subtilis are dispensable as long as no stress is imposed, although they seem to be required for quick response to stresses.