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During a search for natural inflammatory inhibitors, 1-O-acetylbritannilactone (ABL), a sesquiterpene lactone, was isolated from the flowers of Inula britannica. ABL significantly inhibited human neutrophil elastase (HNE) with a half-maximal inhibitory concentration (IC50) of 3.2 ± 0.3 µM, thus did so more effectively than the positive control material (epigallocatechin gallate) (IC50 7.2 ± 0.5 µM). An enzyme kinetic study was performed. ABL noncompetitively inhibited HNE with an inhibition constant Ki of 2.4 µM. ABL inhibited lipopolysaccharide-induced nitric oxide and prostaglandin E2 production by RAW 264.7 cells in a dose-dependent manner, as well as the protein-level expression of inducible nitric oxide synthase and cyclooxygenase-2. The anti-inflammatory effect of ABL was confirmed using a transgenic Tg(mpx:EGFP) zebrafish larval model. The exposure of the larvae to ABL inhibited neutrophil recruitment to the site of injury after tail fin amputation. [ABSTRACT FROM AUTHOR]

The evaluation of retinal vascular structures is important for analyzing various ophthalmic diseases. Conventional trypsin digestion was used for separating retinal vasculatures in mouse, rat, and other animal models; however, the trypsin method alone is technically difficult to perform and has not been reported in zebrafish to date. In this study, we introduced a rapid and convenient method that allows the investigation of fine vessel structures at a cellular level in the relatively intact retinal vasculature of adult zebrafish. Using an anti-ZO-1 antibody, tight junction structures in retinal vessels were examined in detail and several different cell types constituting blood vessels in arterial and capillary areas were identified. In addition, using cell type-specific antibodies, we identified smooth muscle cells, blood cells, and endothelial cells in the retinal vasculature. Finally, using the hyperglycemic model, we observed the dilation of retinal vessels, the downregulation of tight junction proteins, and the reduction in smooth muscle cells. Based on these results, we provide a rapid and convenient method for the study of retinal vasculature disease in the zebrafish animal model. [ABSTRACT FROM AUTHOR]

Vanishing white matter (VWM) disease is a genetic leukodystrophy leading to severe neurological disease and early death. VWM is caused by bi-allelic mutations in any of the five genes encoding the subunits of the eukaryotic translation factor 2B (EIF2B). Previous studies have attempted to investigate the molecular mechanism of VWN by constructing models for each subunit of EIF2B that causes VWM disease. The underlying molecular mechanisms of the way in which mutations in EIF2B3 result in VWM are largely unknown. Based on our recent results, we generated an eif2b3 knockout (eif2b3−/−) zebrafish model and performed quantitative proteomic analysis between the wild-type (WT) and eif2b3−/− zebrafish, and identified 25 differentially expressed proteins. Four proteins were significantly upregulated, and 21 proteins were significantly downregulated in eif2b3−/− zebrafish compared to WT. Lon protease and the neutral amino acid transporter SLC1A4 were significantly increased in eif2b3−/− zebrafish, and crystallin proteins were significantly decreased. The differential expression of proteins was confirmed by the evaluation of mRNA levels in eif2b3−/− zebrafish, using whole-mount in situ hybridization analysis. This study identified proteins which candidates as key regulators of the progression of VWN disease, using quantitative proteomic analysis in the first EIF2B3 animal model of VWN disease. [ABSTRACT FROM AUTHOR]

Ocular pathologic angiogenesis is an important causative risk factor of blindness in retinopathy of prematurity, proliferative diabetic retinopathy, and neovascular macular degeneration. Guibi-tang (GBT) is a frequently used oriental herbal formula in East Asian countries, and is also called Qui-pi-tang in Chinese and Kihi-To in Japanese. In the present study, we investigated the preventive effect of GBT on retinal pathogenic neovascularization in a mouse model of oxygen-induced retinopathy (OIR). C57BL/6 mice were exposed to 75% hyperoxia for five days on postnatal day 7 (P7). The mice were then exposed to room air from P12 to P17 to induce ischemic proliferative retinopathy. GBT (50 or 100 mg/kg/day) was intraperitoneally administered daily for five days (from P12 to P16). On P17, Retinal neovascularization was measured on P17, and the expression levels of 55 angiogenesis-related factors were analyzed using protein arrays. GBT significantly decreased retinal pathogenic angiogenesis in OIR mice, and protein arrays revealed that GBT decreased PAI-1 protein expression levels. Quantitative real-time PCR revealed that GBT reduced vascular endothelial growth factor (VEGF), fibroblast growth factor 2 (FGF2), and plasminogen activator inhibitor 1 (PAI-1) mRNA levels in OIR mice. GBT promotes potent inhibitory activity for retinal neovascularization by decreasing VEGF, FGF2, and PAI-1 levels.

Choroidal neovascularization (CNV) is a common pathology in age-related macular degeneration. In this study, we evaluated in a rat model the effect of an extract of Cinidium officinale Makino and its bioactive compound, butylidenephthalide, on laser-induced CNV. Experimental CNV was induced in Long-Evans rats by laser photocoagulation. C. officinale extract (COE) and butylidenephthalide was intraperitoneally injected once per day for ten days after laser photocoagulation. Choroidal flat mounts were prepared to measure CNV areas and macrophage infiltration. We used a protein array to evaluate the expression levels of angiogenic factors. The CNV area and macrophage infiltration in COE-treated rats were significantly lower than in vehicle-treated rats. COE decreased the expression levels of IGFBP-1, MCP-1, PAI-1, and VEGF. Additionally, butylidenephthalide also inhibited the laser-induced CNV formation and macrophage infiltration and down-regulated the expression of IGFBP-1, MCP-1 and VEGF. These results suggest that COE exerts anti-angiogenic effects on laser-induced CNV by inhibiting the expression of IGFBP-1, MCP-1, and VEGF, indicating that anti-angiogenic activities of COE may be in part due to its bioactive compound, butylidenephthalide.