Coagulation Factor VII Fine-tunes Hepatic Steatosis by Blocking AKT-CD36-Mediated Fatty Acid Uptake.

Nonalcoholic fatty liver disease (NAFLD) is considered a risk factor for cardiovascular and cerebrovascular disease owing to its close association with coagulant disturbances. However, the precise biological functions and mechanisms that connect coagulation factors to NAFLD pathology remain inadequately understood. Herein, with unbiased bioinformatics analyses followed by functional testing, we demonstrate that hepatic expression of coagulation factor VII (FVII) decreases in patients and mice with NAFLD/nonalcoholic steatohepatitis (NASH). By using adenovirus-mediated F7-knockdown and hepatocyte-specific F7-knockout mouse models, our mechanistic investigations unveil a noncoagulant function of hepatic FVII in mitigating lipid accumulation and lipotoxicity. This protective effect is achieved through the suppression of fatty acid uptake, orchestrated via the AKT-CD36 pathway. Interestingly, intracellular FVII directly interacts with AKT and PP2A, thereby promoting their association and triggering the dephosphorylation of AKT. Therapeutic intervention through adenovirus-mediated liver-specific overexpression of F7 results in noteworthy improvements in liver steatosis, inflammation, injury, and fibrosis in severely afflicted NAFLD mice. In conclusion, our findings highlight coagulation factor FVII as a critical regulator of hepatic steatosis and a potential target for the treatment of NAFLD and NASH.

Induced Self-Assembly of Vitamin E-Spermine/siRNA Nanocomplexes via Spermine/Helix Groove-Specific Interaction for Efficient siRNA Delivery and Antitumor Therapy.

Gene therapy has been one of potential strategies for the treatment of different diseases, where efficient and safe gene delivery systems are also extremely in need. Current lipid nanoparticles (LNP) technology highly depends on the packing and condensation of nucleic acids with amine moieties. Here, an attempt to covalently link two natural compounds, spermine and vitamin E, is made to develop self-assembled nucleic acid delivery systems. Among them, the spermine moieties specifically interact with the major groove of siRNA helix through salt bridge interaction, while vitamin E moieties are located around siRNA duplex. Such amphiphilic vitamin E-spermine/siRNA complexes can further self-assemble into nanocomplexes like multiblade wheels. Further studies indicate that these siRNA nanocomplexes with the neutrally charged surface of vitamin E can enter cells via caveolin/lipid raft mediated endocytosis pathway and bypass lysosome trapping. With these self-assembled delivery systems, efficient siRNA delivery is successfully achieved for Eg5 and Survivin gene silencing as well as DNA plasmid delivery. Further in vivo study indicates that VE-Su-Sper/DSPE-PEG2000/siSurvivin self-assembled nanocomplexes can accumulate in cancer cells and gradually release siRNA in tumor tissues and show significant antitumor effect in vivo. The self-assembled delivery system provides a novel strategy for highly efficient siRNA delivery.

Comparative physiological and transcriptome analysis provide insights into the inhibitory effect of 6-pentyl-2H-pyran-2-one on Clarireedia jacksonii.

Clarireedia spp. is a destructive phytopathogenic fungus that causes turf dollar spot of bent-grass, leading to widespread lawn death. In this study, we explored the antifungal capability of 6-pentyl-2H-pyran-2-one (6PP), a natural metabolite volatilized by microorganisms, which plays an important role in the biological control of turfgrass dollar spot. However, the mechanisms by which 6PP inhibits Clarireedia jacksonii remain unknown. In the present study, C. jacksonii mycelial growth was inhibited by the 6PP treatment and the 6PP treatment damaged cell membrane integrity, causing an increase in relative conduc-tivity. Furthermore, physiological and biochemistry assay showed that 6PP treatment can enhance reactive oxygen species (ROS) levels, malondialdehyde (MDA) content obviously increased with 6PP exposure, increased alchohol dehydrogenase (ADH) and depleted acetalde-hyde dehydrogenase (ALDH), and activated the activities of many antioxidant enzymes in C. jacksonii. Gen Ontology and Kyoto Encyclopedia of Genes and Genomes analysis revealed that some genes in C. jacksonii after 6PP treatment related to integrity of the cell wall and membrane, and oxidative stress were significantly downregulated. It is worth mentioning that the fatty acid degradation pathway is significantly upregulated, with an increase in ATP content and ATP synthase activity, which may promote fungal cell apoptosis. Moreover, we found that the expression of ABC transporters, and glutathione metabolism encoding genes were increased to respond to external stimuli. Taken together, these findings revealed the potential antifungal mechanism of 6PP against Clarireedia spp., which also provides a theoretical basis for the commercial utilization of 6PP as a green pesticide in the future.

Betaine-homocysteine methyltransferase promotes adipocyte commitment and insulin resistance via p38 MAPK/Smad signaling.

OBJECTIVE: Betaine-homocysteine methyltransferase (Bhmt) belongs to the family of methyltransferases and is involved in the one-carbon metabolic cycle, which is associated with the risk of diabetes and adiposity. This study aimed to explore whether Bhmt participated in the development of obesity or its associated diabetes, as well as the mechanism involved. METHODS: The expression levels of Bhmt were examined in stromal vascular fraction cells and mature adipocytes in obesity and nonobesity. Knockdown and overexpression of Bhmt in C3H10T1/2 cells were used to investigate Bhmt's function in adipogenesis. Bhmt's role in vivo was analyzed using an adenovirus-expressing system and a high-fat diet-induced obesity mouse model. RESULTS: Bhmt was highly expressed in stromal vascular fraction cells rather than mature adipocytes of adipose tissue and was upregulated in adipose tissue in obesity and C3H10T1/2-commited preadipocytes. Overexpression of Bhmt promoted adipocyte commitment and differentiation in vitro and exacerbated adipose tissue expansion in vivo, with a concomitant increase in insulin resistance, whereas Bhmt silencing exhibited opposite effects. Mechanistically, Bhmt-induced adipose expansion was mediated by stimulating the p38 MAPK/Smad pathway. CONCLUSIONS: The findings of this study highlight the obesogenic and diabetogenic role of adipocytic Bhmt and propose Bhmt as a promising therapeutic target for obesity and obesity-related diabetes.

Boosting expression level of plectasin in recombinant Pichia pastoris via 2A self-processing peptide assembly.

Plectasin is a promising and potent antimicrobial peptide isolated from the fungus Pseudoplectania nigrella which has been heterologously expressed in various hosts. In this study, a four-copy cassette of plectasin was constructed via 2A peptide assembly to further increase its expression level in recombinant Pichia pastoris. The yeast transformant 4Ple-61 harboring four-copy cassette of plectasin could secrete 183.2 mg/L total protein containing 60.8% of plectasin at the flask level within 120 h, which was 2.3 times higher than that of the yeast transformant Ple-6 carrying one-copy cassette of plectasin. Western blot confirmed the significant peptide expression level in the transformant 4Ple-61. Furthermore, it yielded as high as 426.3 mg/L total protein within 120 h during a 5-L fermentation. The purified plectasin shows superior stability and good antimicrobial activity against conventional Staphylococcus aureus ATCC 26,001 and some food-borne antibiotic-resistant S. aureus strains with the MICs ranging from 8 to 32 µg/mL. Therefore, the strategy based on 2A peptide assembly can enhance the expression of plectasin and further expand its application prospect. KEY POINTS: • A yeast transformant 4Ple-61 with four-copy cassette of plectasin was constructed. • The plectasin level yield by the transformant 4Ple-61 was boosted by 2.3 times. • The plectasin showed good activity against food-borne antibiotic-resistant S. aureus.

Enhanced photodynamic therapy of TiO2/N-succinyl-chitosan composite for killing cancer cells

Abstract The aim of this study was to investigate the effect of TiO2/N-succinyl-chitosan composite (TiO2/ NSCS) photodynamic therapy (PDT), while considering the effects of various light sources on the activation of photosensitizer. The methyl thiazolyl tetrazolium assay was used to examine the cell survival rate of the cells. The results showed that glioma cell strain (U251) was the most sensitive cancer cell strain to TiO2/NSCS. When the concentration of TiO2/NSCS was between 0 and 800 μg·mL-1, there was no obvious cytotoxicity to normal liver cells (HL-7702) and U251 cells. During the PDT process, the photokilling effect of TiO2/NSCS on U251 cells under ultraviolet-A (UVA) light irradiation was stronger than that of pure TiO2, and its killing effects were positively correlated with concentration and irradiation time. In addition, both UVA and visible light could excite TiO2/ NSCS, which had significant killing effect on U251 cells. The results of acridine orange/ethidium bromide fluorescent double staining and Annexin V/propidium iodide double staining indicated that TiO2/NSCS under UVA and visible light irradiation could kill U251 cells by inducing apoptosis, and the apoptosis rate of TiO2/NSCS treatment groups was higher than that of TiO2 treatment groups. Therefore, TiO2/NSCS might be used as a potential photosensitizer in PDT.