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@#By regulating gene expression, nucleic acid drugs functioning in the cytoplasm or nucleus are of great significance in the treatment of acquired or inherited diseases and vaccine development.A variety of nucleic acid delivery vectors currently developed are suffering from low transfection efficiency due to endosome/lysosome entrapment.This paper introduces and summarizes the nucleic acid delivery strategies that bypass the endosomal/lysosomal pathway, including membrane translocation, membrane fusion, receptor/transporter-mediated non-endocytic uptake and caveolae-mediated endocytosis, and discusses the problems and challenges facing such strategies, aiming to facilitate the development of intracellular delivery of nucleic acid drugs bypassing lysosomal pathway.
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The high altitude/hypoxic environment induced skeletal muscle atrophy is considered to be the interaction of multi-system and multi-organ, but the direct mechanism of hypoxia on muscle cells in this process is not clear. This study intended to investigate the effects of hypoxia exposure on proteins in ubiquitin and autophagy pathways, and explored the possible mechanism of hypoxia induced change of myotube diameter. The expression of myosin, hypoxia inducible factor-1 α (HIF-1α), forkhead box protein O1 (FoxO1), and ubiquitin protease pathway (MuRF1 and Atrogin1) and autophagy lysosomal pathway (p62, Beclin1, LC3) related proteins were detected by Western blot; The integrated optical density (IOD) of Myosin and LC3 was detected by IF. The results showed that the diameters of myotube at 6 h and 12 h were significantly reduced, and the expression of myosin was significantly reduced at 6 h after hypoxia exposure (P<0. 05); the protein levels of HIF-1α and FoxO1 were significantly increased at 6 h (P<0. 05); The expression of MuRF1 in each time points of hypoxia was significantly higher than 0 h (P<0. 05), but no difference of Atrogin1 expression was detected; Compared with 0 h, the expression of p62 was reduced significantly in response to hypoxia. The protein expression of Beclin1 and the IOD of LC3 was increased significantly at 6 h, and the LC3Ⅱ/Ⅰ ratio was significantly higher at 6 h, but significantly lower at 12 h and 24 h (P<0. 05).The results above indicated that the reduction of the myotube diameter of L6 skeletal muscle cells was induced by hypoxia exposure (1% O
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Objective: To investigate the changes of expressions of tubulin and endosome-lysosome in the neurons in hippocampus tissue of the mice after status epilepticus (SE), and to elucidate the change rule of microtubule and endosome-lysosome system in the process of delayed neuronal death. Methods: A total of 40 male ICR mice were divided into control group ( n= 7, given normal saline) and experiment group (w=33» give pilocarpine); the mice in experiment group met the SE standand were divided intoSE 1 d» SE 2 d, SE 3 d and SE 7 d groups according to the time after SE ( n=5). Nissl and Fluoro-Jade B (F-JB) staining methods were used to detect the damage of neurons in hippocampus tissue of the mice in various groups. The expression intensities of (3-tubulin? endosom protein Rab5 and lysosome constitutive protein LAMP1 and the percentages of (3-tubulin∗ Rab5 and LAMP1 positive areas in neurons in hippocampus tissue of the mice in various groups were detected by immunofluorescence method. The relationships between the expression of
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OBJECTIVE@#To screen potential traditional Chinese medicine and their active monomer ingredients for treatment of diabetic nephropathy (DN) through the mechanism of caspase-1-mediated pyroptosis.@*METHODS@#Using the Chinese Medicine System Pharmacology Analysis Platform (TCMSP), we screened traditional Chinese drugs and their active monomer components targeting caspase-1, and searched for the potential gene targets of the monomer components using GeneCards database. Cytoscape was used to construct the monomer compound-gene target network. Gene ontology (GO) functional enrichment analysis and Kyoto Gene and Gene Encyclopedia (KEGG) pathway enrichment analysis were used to predict the molecular mechanism of the screened traditional Chinese medicine and monomers. In SD rat models of diabetic mellitus (DM), we tested the therapeutic effect of ginsenoside Rh2 (daily dose of 20 mg/kg for 12 weeks) by examining renal pathology with HE staining and detecting the expressions of pyroptosis marker proteins caspase-1, GSDMD, IL-1β and IL-18 in the renal tissues using Western blotting, the serum levels of IL-1β and IL-18 and activities of cathepsin B and cathepsin L.@*RESULTS@#Ginsenoside Rh2 could effectively dock with caspase-1 molecule. Fourteen targets were identified in ginsenoside Rh2 target network. GO function enrichment analysis revealed 27 GO terms associated with molecular function (4 terms), cell component (10 terms) and biological process (13 terms). KEGG pathyway enrichment analysis identified 4 signaling pathways involving lysosomes, glycosaminoglycan degradation, galactose metabolism, and sphingolipid metabolism pathways. In the animal experiment, treatment with ginsenoside Rh2 significantly alleviated renal pathologies and down-regulated the expressions of pyroptosis marker proteins (cleaved caspase-1, GSDMD-N, IL-1β and IL-18) ( < 0.05 or 0.01), lowered serum levels of IL-1β and IL-18 ( < 0.01), and enhanced the activities of cathepsin B and cathepsin L in the serum of the diabetic rats.@*CONCLUSIONS@#Ginsenoside Rh2 may inhibit caspase-1-mediated pyroptosis through the lysosome pathway to improve kidney damages in rat models of DN.
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OBJECTIVE: To explore the effects of human A30P mutant α-synuclein (α-syn) on autophagy lysosome pathway and ubiquitin proteasome system. METHODS: Constructing stable cell lines expressing human A30P mutant α-syn in PC12 cells, and detecting the levels of autophagy related proteins LC3-II and p62 as well as ubiquitinated proteins. Furthermore, the influence of A30P mutant α-syn on cell viability upon normal and stress conditions was conducted by MTT assay. RESULTS: Expressing A30P mutant α-syn could significantly reduce autophagy related protein LC3-II level and increase level of autophagy substrate p62, as well as promote aggregation of ubiquitinated proteins. Otherwise, expressing A30P mutant α-syn did not reduce cell viability under normal conditions. However, cell viability was significantly reduced under treatment with neurotoxin rotenone or serum free condition in A30P mutant α-syn groups compared with empty vector group. CONCLUSION: Over expression of human A30P mutant α-syn could impair the protein degradation system and increase the sensitivity of cells to toxic insults.