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Diabetic retinopathy (DR) is the most prevalently occurring microvascular complication in diabetic patients that triggers severe visual impairments. The anti‐angiogenesis role of FBXW7 has been identified in breast cancer. Therefore, this study intends to decipher the mechanism of FBXW7 in angiogenesis of DR. DR model was induced on mice using high‐glucose (HG) and high‐fat diet, and retinal microvascular endothelial cells (RMECs) isolated from normal mice were induced with HG, followed by evaluation of FBXW7, Ki67, HIF‐1α and VEGF expression by immunofluorescence, immunohistochemistry or Western blot analysis. After gain‐ and loss‐of‐function assays in normal and DR mice, angiogenesis was assessed by CD31 fluorescence staining and Western blot analysis. After ectopic expression and silencing experiments in HG‐induced RMECs, RMEC proliferation, migration and angiogenesis were, respectively, determined by EdU, Transwell and in vitro angiogenesis assays. The impact of FBXW7 on the ubiquitination of c‐Myc was studied by cycloheximide chase assay and proteasome inhibition, and the binding of c‐Myc to HDAC2 promoter by dual‐luciferase reporter gene experiment. DR mice and HG‐induced RMECs possessed down‐regulated FBXW7 and up‐regulated Ki67, HIF‐1α and VEGF. Silencing FBXW7 enhanced angiogenesis in normal mouse retinal tissue, but overexpressing FBXW7 or silencing c‐Myc diminished angiogenesis in DR mouse retinal tissue. Overexpressing FBXW7 or silencing c‐Myc depressed proliferation, migration and angiogenesis in HG‐induced RMECs. FBXW7 induced c‐Myc ubiquitination degradation, and c‐Myc augmented HDAC2 expression by binding to HDAC2 promoter. Conclusively, our data provided a novel sight of anti‐angiogenesis role of FBXW7 in DR by modulating the c‐Myc/HDAC2 axis.

For strain improvement, robust and scalable high-throughput cultivation systems as well as simple and rapid high-throughput detection methods are crucial. However, most of the screening methods for lactic acid bacteria (LAB) strains were conducted in shake flasks and detected by high-performance liquid chromatography (HPLC), making the screening program laborious, time-consuming and costly. In this study, an integrated strategy for high-throughput screening of high L-lactic acid-productivity strains by Bacillus coagulans in deep-well microtiter plates (MTPs) was developed. The good agreement of fermentation results obtained in the MTPs platform with shake flasks confirmed that 24-well U-bottom MTPs could well alternate shake flasks for cell cultivation as a scale-down tool. The high-throughput pH indicator (bromocresol green) and L-lactate oxidase (LOD) assays were subsequently developed to qualitatively and quantitatively analyze L-lactic acid concentration. Together with the color halos method, the pH indicator assay and LOD assay, the newly developed three-step screening strategy has greatly accelerated the screening process for LAB strains with low cost. As a result, two high L-lactic acid-productivity mutants, IH6 and IIIB5, were successfully screened out, which presented, respectively, 42.75 and 46.10 % higher productivities than that of the parent strain in a 5-L bioreactor.

In the production of L-lactic acid, the residual sugar in fermentation broth would lead to increased sugar content in its commercial products, bringing about lactic acid deterioration because of Maillard reactions and thus shortening their shelf life. In this work, the residual sugar components in L-lactic acid industrial fermentation broth with agricultural biomass-derived sugar were determined by means of highperformance anion-exchange chromatography (HPAEC) coupled with pulsed amperometric detection (PAD). A feasible off-line sample clean-up procedure consisting of sodium oxalate extraction, ethanol extraction and ion-exchange column extraction was developed to remove calcium ions, proteins, anion acids and organic acids before analysis, and their removal efficiencies were 100, 98.74, 97.99 and 99.41%, respectively. Under the optimized analytical conditions, the seventeen sugars, including four monosaccharides, eight disaccharides and five oligosaccharides, have been completely separated in one experimental run on a Dionex Carbopac PA 200 column with sodium hydroxide and sodium acetate as eluents. The method was validated, with good linearity (R 2 > 0.99), high precision (RSDs < 5.47%), and sensitive detection limits (0.35-44.61 mg L À1 ); recoveries were >86.95% for short chain carbohydrates (degree of polymerization, DP < 4), while recoveries ranged from 81.87 to 61.41% for DP4 to DP7. The method was successfully applied for analysis of the residual sugar in lactic acid broth with a salt content as high as 120 g L À1 calcium lactate, using different kinds of starch (corn starch, corn flour and cassava starch) hydrolysates as carbon sources. Isomaltose (47.97-60.43%), sucrose (17.50-29.73%) and trehalose (3.05-7.51%) were the main components of residual sugar in these broths.

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