To improve the quality control of drugs, we predicted the absorption, distribution, metabolism, excretion, and toxicity (ADMET) of ceftazidime (CAZ) and its impurities via in silico methods. We used three types of quantitative structure-activity relationship and docking software for precise prediction: Discovery Studio 4.0, OECD QSAR Toolbox 4.1, Toxtree, and the pkCSM approach. The pharmacokinetics and toxicity of ceftazidime and impurity A (Δ-2-CAZ) are similar. The biological properties of impurity B (CAZ E-isomer) are different from CAZ. Therefore, we focused on drug stability to analyze impurity B. Impurities D and I have strong lipophilicity, good intestinal absorption, and poor excretion in the body. Impurity D is particularly neurotoxic and genotoxic. It is important to control the content of impurity D. The toxicity of impurity F is low, but the toxicity is enhanced when it becomes the C-3 side chain of CAZ and forms a quaternary amine group. We conclude that the beta-lactam ring of nucleus, the quaternary amine group at the C-3 side chain, and the acetates at the C-7 side chain of CAZ are the main toxic functional groups. Impurities B and D may be the genetic impurity in CAZ and may also have neurotoxicity. This in silico approach can predict the toxicity of other cephalosporins and impurities.

Parkinson's disease (PD) is one of the most epidemic neurodegenerative diseases, and is characterized by movement disorders arising from loss of midbrain dopaminergic (DA) neurons. Recently, the relationship between PD and autophagy has received considerable attention, but information about the mechanisms involved is lacking. Here, we report that autophagy-related gene 5 (ATG5) is potentially important in protecting dopaminergic neurons in a 1-methyl-4phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD model in zebrafish. Using analyses of zebarfish swimming behavior, in situ hybridizatiton, immunofluorescence and expressions of genes and proteins related to PD and autophagy, we found that ATG5 expression level was decreased and autophagy flux was blocked in this model. The ATG5 down-regulation led to the upgrade of PDassociated proteins, such as β-synuclein, Parkin, and PINK1, aggravation of MPTP-induced PDmimicking pathological locomotor behavior, DA neuron loss labelled by tyrosine hydroxylase (TH) or dopamine transporter (DAT), and blocked autophagy flux in the zebrafish model. ATG5 overexpression alleviated or reversed these PD pathological features, rescued DA neuron cells as indicated by elevated TH /DAT levels, and restored autophagy flux. The role of ATG5 in protecting DA neurons was confirmed by expression of the human atg5 gene in the zebrafish model. Our findings reveal that ATG5 has a role in neuroprotection, and upregulation of ATG5 may serve as a goal in the development of drugs for PD prevention and management.

Dietary composition has important impact on nonalcoholic fatty liver disease (NAFLD). The purpose of this study was to explore the relationship between NAFLD and major dietary components using zebrafish larvae fed different diets. Zebrafish larvae fed with high cholesterol (HC), high fructose (HF) and extra feeding (EF) diets for 10 days displayed varying degrees steatosis. The incidence and degree of steatosis were the most severe in the EF group. A HC diet severely promoted lipid deposits in the caudal vein. The triglyceride and glucose contents of zebrafish significantly increased in the HF and EF groups compared with the control group. Moreover, the mRNA expression of oxidative stress gene gpx1a, endoplasmic reticulum stress genes ddit3 and grp78, inflammatory genes tnfa, glucose metabolism genes irs2, glut1 and glut2, and lipid metabolism genes cidec, chrebp, ppara and cpt1a were significantly increased in the HF group. The HC diet was associated with upregulation of grp78, and downregulation of irs2, glut1 and glut2. The mRNA expression of lipogenesis and glucose metabolism associated genes were decreased in the EF group. In addition, the autophagy associated genes atg3, atg5, atg7, and atg12, and protein expression of ATG3 and LC3BII were reduced and P62 were elevated in the HC group. We also performed comparative transcriptome analysis of the four groups. A total of 2,492 differentially expressed genes were identified, and 24 statistically significant pathways were enriched in the diet treatment groups. This study extends our understanding of the relationships between diet ingredients and host factors that contribute to the pathogenesis of NAFLD, which may provide new ideas for the treatment of NAFLD.

Type 2 diabetes mellitus is characterized by insulin resistance. However, the complete molecular mechanism remains unclear. In this study, zebrafish were fed a long-term high-fat diet to induce type 2 diabetes, which resulted in a higher body weight, body mass index, more lipid vacuoles in liver, increased insulin transcription level in liver, brain and muscle, and high fasting blood glucose in the high-fat diet zebrafish. Oppositely, the transcription levels of insulin substrate-2 and glucose transporter 2 were significantly decreased, indicating insulin signaling pathway and glucose transport impaired in the insulin-targeting tissues. Transcription of the autophagy-related genes, ATG3, ATG4B, ATG5, ATG7, ATG12, and FOXO3, were decreased but autophagy inhibitor gene m-TOR increased, and autophagy-flux was inhibited in liver of the high-fat diet zebrafish. Main of these changes were confirmed in palmitic acid-treated HepG2 cells. Further, in co-immunoprecipitation and subcellular co-localization experiments, the conjunction of preproinsulin with cargo-recognition protein p62 increased, but conjuncts of autophagosome with p62-cargo, lysosomes with p62-cargo, and autolysosomes decreased apparently. Interestingly, lysosomes, autolysosomes and conjuncts of p62-insulin localized at the periphery of palmitic acid-treated cells, the margination of lysosomes may mediate deactivation of proteases activity. These findings suggest that intracellular high-lipid may trigger defective autophagy, defective downstream signaling of insulin and accumulated intracellular preproinsulin, leading to dysregulation of cell homeostasis mechanism, which may be one of reasons involved in insulin-resistance in type 2 diabetes.