Niemann-Pick C1 (NPC1) is a lysosomal cholesterol storage disorder, that severely affects the brain, and is caused by mutations in the NPC1 gene, which encodes an intracellular membrane transporter ...of non-esterified cholesterol. Therapeutic options for NPC1 are few, and classical enzyme replacement therapy with the recombinant protein is not possible as the NPC1 gene product is an insoluble membrane protein, which increases the need for development of gene therapy for NPC1. While viral based gene therapy is under development, it is important to investigate alternative approaches to brain gene therapy without viral vectors. The present work develops a plasmid DNA approach to gene therapy of NPC1 using Trojan horse liposomes (THLs), wherein the plasmid DNA is encapsulated in 100 nm pegylated liposomes, which are targeted to organs with a monoclonal antibody against the mouse transferrin receptor. THLs were encapsulated with a 8.0 kb plasmid DNA encoding the 3.9 kb human NPC1 open reading frame, under the influence of a 1.5 kb platelet derived growth factor B (PDGFB) promoter. THLs were administered weekly beginning at 6-7 weeks in the NPC1
null mouse, and delivery of the plasmid DNA, and NPC1 mRNA expression in brain, spleen, and liver were confirmed by quantitative PCR. THL treatment reduced tissue inclusion bodies in brain, and peripheral organs, but did not prolong lifespan in these mice. The work suggests that early treatment after birth may be required to reverse this disease model with NPC1 gene replacement therapy.
The MYC oncogene broadly promotes transcription mediated by all nuclear RNA polymerases, thereby acting as a positive modifier of global gene expression. Here, we report that MYC stimulates the ...transcription of DANCR, a long noncoding RNA (lncRNA) that is widely overexpressed in human cancer. We identified DANCR through its overexpression in a transgenic model of MYC-induced lymphoma, but found that it was broadly upregulated in many human cancer cell lines and cancers, including most notably in prostate and ovarian cancers. Mechanistic investigations indicated that DANCR limited the expression of cell-cycle inhibitor p21 (CDKN1A) and that the inhibitory effects of DANCR loss on cell proliferation could be partially rescued by p21 silencing. In a xenograft model of human ovarian cancer, a nanoparticle-mediated siRNA strategy to target DANCR
was sufficient to strongly inhibit tumor growth. Our observations expand knowledge of how MYC drives cancer cell proliferation by identifying DANCR as a critical lncRNA widely overexpressed in human cancers.
These findings expand knowledge of how MYC drives cancer cell proliferation by identifying an oncogenic long noncoding RNA that is widely overexpressed in human cancers.
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TP53, a well-known tumour suppressor gene that encodes p53, is frequently inactivated by mutation or deletion in most human tumours. A tremendous effort has been made to restore p53 activity in ...cancer therapies. However, no effective p53-based therapy has been successfully translated into clinical cancer treatment owing to the complexity of p53 signalling. Here we demonstrate that genomic deletion of TP53 frequently encompasses essential neighbouring genes, rendering cancer cells with hemizygous TP53 deletion vulnerable to further suppression of such genes. POLR2A is identified as such a gene that is almost always co-deleted with TP53 in human cancers. It encodes the largest and catalytic subunit of the RNA polymerase II complex, which is specifically inhibited by α-amanitin. Our analysis of The Cancer Genome Atlas (TCGA) and Cancer Cell Line Encyclopedia (CCLE) databases reveals that POLR2A expression levels are tightly correlated with its gene copy numbers in human colorectal cancer. Suppression of POLR2A with α-amanitin or small interfering RNAs selectively inhibits the proliferation, survival and tumorigenic potential of colorectal cancer cells with hemizygous TP53 loss in a p53-independent manner. Previous clinical applications of α-amanitin have been limited owing to its liver toxicity. However, we found that α-amanitin-based antibody-drug conjugates are highly effective therapeutic agents with reduced toxicity. Here we show that low doses of α-amanitin-conjugated anti-epithelial cell adhesion molecule (EpCAM) antibody lead to complete tumour regression in mouse models of human colorectal cancer with hemizygous deletion of POLR2A. We anticipate that inhibiting POLR2A will be a new therapeutic approach for human cancers containing such common genomic alterations.
Glutamine can play a critical role in cellular growth in multiple cancers. Glutamine‐addicted cancer cells are dependent on glutamine for viability, and their metabolism is reprogrammed for glutamine ...utilization through the tricarboxylic acid (TCA) cycle. Here, we have uncovered a missing link between cancer invasiveness and glutamine dependence. Using isotope tracer and bioenergetic analysis, we found that low‐invasive ovarian cancer (OVCA) cells are glutamine independent, whereas high‐invasive OVCA cells are markedly glutamine dependent. Consistent with our findings, OVCA patients’ microarray data suggest that glutaminolysis correlates with poor survival. Notably, the ratio of gene expression associated with glutamine anabolism versus catabolism has emerged as a novel biomarker for patient prognosis. Significantly, we found that glutamine regulates the activation of STAT3, a mediator of signaling pathways which regulates cancer hallmarks in invasive OVCA cells. Our findings suggest that a combined approach of targeting high‐invasive OVCA cells by blocking glutamine's entry into the TCA cycle, along with targeting low‐invasive OVCA cells by inhibiting glutamine synthesis and STAT3 may lead to potential therapeutic approaches for treating OVCAs.
Synopsis
Glutamine plays an important role in cellular growth in several cancers. In this study, a further link between glutamine dependency and tumor invasiveness is established in ovarian cancer. Glutamine maintains the high‐invasive phenotype by regulating STAT3 signaling.
High‐invasive ovarian cancer (OVCA) cells are glutamine dependent in contrast to low‐invasive cells that are glutamine independent.
Glutamine regulates STAT3 activation in high‐invasive cancer cells.
Glutamine's entry into TCA cycle modulates the invasive potential of high‐invasive cancer cells.
The ratio of glutamine catabolism over glutamine anabolism is associated with worse overall survival in OVCA patients.
Glutamine plays an important role in cellular growth in several cancers. In this study, a further link between glutamine dependency and tumor invasiveness is established in ovarian cancer. Glutamine maintains the high‐invasive phenotype by regulating STAT3 signaling.
The direct feeding value of distillers grains is low due to the presence of higher cellulose, lignin and anti-nutritional factors such as mannan and xylan. In this study, complex enzymes and ...probiotic flora based on “probiotic enzyme synergy” technology were used to produce fermented distillers grains. The optimal substrate ratio, moisture content, fermentation time and temperature were determined. Subsequently, scale-up experiments were conducted to determine the performance of fermented feed. The results showed that multi-probiotic ( Lactobacillus casei , Bacillus subtilis , Saccharomyces cerevisiae , and Aspergillus oryzae ) cooperated with complex enzymes (glucanase, mannanase, xylanase) showed excellent fermentation effect, crude protein, trichloroacetic acid soluble protein and fat increased by 31.25, 36.68, and 49.11% respectively, while crude fiber, acidic fiber and neutral fiber decreased by 34.24, 26.91, and 33.20%, respectively. The anti-nutritional factors mannan and arabinoxylan were reduced by 26.96 and 40.87%, respectively. Lactic acid, acetic acid, and propionic acid in the fermented organic acids increased by 240.93, 76.77, and 89.47%, respectively. Butyric acid increased significantly from scratch, and the mycotoxin degradation effect was not significant. This study provides a potential approach for high-value utilization of distillers grains.
In this paper, carbon nanotubes and graphene are combined with traditional conductive agent (Super-P/KS-15) to prepare a new type of composite conductive agent to study the effect of composite ...conductive agent on the internal resistance and performance of lithium iron phosphate batteries. Through the SEM, internal resistance test and electrochemical performance test, carbon nanotubes and graphene composite traditional conductive agent (Super-P/KS-15) form an effective three-dimensional conductive network of points, lines, and surfaces. When the ratio of graphene to carbon nanotubes is 1:3 to compound the traditional conductive agent, the prepared pole piece has a higher compaction density, and the prepared battery has the smallest battery internal resistance. The increase in internal resistance after the cycle is also minimized, which shows the most excellent processing performance and electrochemical performance.
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•HP reversed body weight gain, dyslipidemia and hepatic steatosis in OVX rats.•HP restored gut microbial composition and metabolite profiles in OVX rats.•HP upregulated CYP7A1 and ...increased fecal TBA, which contributes to lowering cholesterol in OVX rats.•SCFAs may mediate HP’s anti-hypercholesterolemia effect in OVX rats by up-regulating GPR43 and GPR41.
Hypericum perforatum L. (HP), a well-known natural medicine, has a potential effect on menopausal hypercholesterolemia. However, the effect of HP extract on gut microbiota and related metabolites, which play vital roles in metabolic disease occurrence, in the context of estrogen deficiency have not yet been reported. The aims of the present study were to investigate the effects of HP extract on gut microbial composition and related metabolite profiles in ovariectomized (OVX) rats and reveal the relationships between pathological indicators and alterations in both gut microbial composition at the genus level and metabolites. Body weight, serum parameters, liver lipids and histomorphology were determined. Microbial composition was analyzed using 16S rRNA sequencing. Fecal short-chain fatty acids (SCFAs) and serum bile acids were quantitatively measured. Correlations between pathological indicators and alteration in gut microbiota and metabolites were investigated using Spearman's rank correlation test. Gene expression of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, cholesterol 7α-hydroxylase (CYP7A1) and cholesterol 27-hydroxylase (CYP27A1) in the liver and G protein-coupled receptors (GPCRs; GPR43 and GPR41), ZO-1 and occludin in the cecum were determined by PCR. Microbial composition and metabolite profiles were significantly changed in OVX rats compared with sham rats. Twelve bacterial genera, 5 SCFAs and 12 bile acids were identified as differential biomarkers. Differential genera, SCFAs and bile acids were closely associated with weight, total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C). In OVX rats, HP administration can significantly reverse the pathological symptoms of body weight gain, serum lipid disorders and hepatic steatosis, at the meanwhile, reestablish gut microbial composition and metabolite profiles. Moreover, HP administration significantly upregulated the levels of CYP7A1, GPR43 and GPR41. In conclusion, HP can ameliorate estrogen deficiency-induced hypercholesterolemia. The underlying mechanism may be associated with improvements in gut microbiota composition and the profile of related metabolites as well as increases in bile acid secretion.
Circular RNA (circRNA) S-phase cyclin A-associated protein in the endoplasmic reticulum (ER) (circSCAPER, ID: hsa_circ_0104595) has been found to be highly expressed in osteoarthritis (OA) patients ...and has been associated with the severity of OA. Hence, the role and mechanisms underlying circSCAPER in OA were investigated in this study.
In vitro cultured human normal chondrocyte C28/I2 was exposed to interleukin (IL)-1β to mimic the microenvironment of OA. The expression of circSCAPER, microRNA (miR)-140-3p, and enhancer of zeste homolog 2 (EZH2) was detected using quantitative real-time polymerase chain reaction and Western blot assays. The extracellular matrix (ECM) degradation, proliferation, and apoptosis of chondrocytes were determined using Western blot, cell counting kit-8, and flow cytometry assays. Targeted relationships were predicted by bioinformatic analysis and verified using dual-luciferase reporter and RNA immunoprecipitation (RIP) assays. The levels of phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) pathway-related protein were detected using Western blot assays.
CircSCAPER was highly expressed in OA cartilage tissues and IL-1β-induced chondrocytes. Knockdown of circSCAPER reduced IL-1β-evoked ECM degradation, proliferation arrest, and apoptosis enhancement in chondrocytes. Mechanistically, circSCAPER directly bound to miR-140-3p, and miR-140-3p inhibition reversed the effects of circSCAPER knockdown on IL-1β-induced chondrocytes. miR-140-3p was verified to target EZH2, and overexpression of miR-140-3p protected chondrocytes against IL-1β-induced dysfunction via targeting EZH2. Additionally, we confirmed that circSCAPER could regulate EZH2 through sponging miR-140-3p, and the circSCAPER/miR-140-3p/EZH2 axis could activate the PI3K/AKT pathway.
CircSCAPER promoted IL-1β-evoked ECM degradation, proliferation arrest, and apoptosis enhancement in chondrocytes via regulating miR-140-3p/EZH2 axis, which gained a new insight into the pathogenesis of OA. Cite this article:
2022;11(2):61-72.
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Adenylate cyclase (AC) is the key enzyme that catalyzes the formation of cAMP from ATP. In this study, we discovered two novel class III ACs with a halophilic property from ...Thermobifida halotolerans DSM 44931 (ThAC) and Haloactinopolyspora alba DSM 45211 (HaAC), respectively. The recombinant ThAC and HaAC were expressed in Escherichia coli with molecular weights of 36.1 and 36.0 kDa respectively. The presence of 2500 and 2200 mmol·L−1 NaCl significantly enhanced the enzyme activities of ThAC and HaAC, with 22-fold and 7.4-fold higher activities compared to those without NaCl, respectively. Several divalent metal ions were found to activate the recombinant ACs to different extents, and the optimal metal ion was Mg2+ for both ThAC and HaAC with concentrations of 80 mmol·L−1 and 40 mmol·L−1 respectively. Purified ThAC and HaAC had the optimal specific activities ((4.59 ± 0.35) × 104 and (7.76 ± 0.52) × 104 U·mg−1) and catalytic efficiency (4.47 and 5.30 L·mmol−1·s−1) at 45 °C and 40 °C respectively, while the optimum pH of both two recombinant ACs was 10.0. This is the first report of the halophilic Class III ACs, which could make new contributions to explore and study ACs for further associated investigations.
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