Nasopharyngeal carcinoma (NPC) is an endemic malignant disease of the head and neck region with unique features including striking ethnic and geographic variations as well as multifactorial etiology. ...Previous studies have demonstrated the anticancer properties of genistein, the major soy isoflavonoid, in several human cancer cells such as breast, prostate, colon, gastric, lung, and hepatoma. However, the action of genistein in NPC cells has not been determined. In this study, we investigated the inhibitory effects of genistein on NPC cells and its possible underlying mechanisms. We found that genistein dose-dependently inhibited the proliferation of human NPC cell line CNE2 cells. DNA flow cytometric analysis revealed that 30 to 120 μM genistein induced dramatic G2/M phase arrest in NPC cells. The mRNA expression levels, as shown by gene expression array and quantitative real-time polymerase chain reaction, and the protein expression levels of the cell cycle regulators p2(Cip1) and ATR (Ataxia telangiectasia and Rad3 related) were elevated following genistein treatment. Interestingly, we also observed concomitant induction of p15(Ink4b) in genistein induced inhibitory effects in NPC cells. Moreover, selective estrogen receptor modulators did not affect genistein induced growth inhibition. These findings provide new insights into the potential intervention of NPC with genistein.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, OILJ, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK, VSZLJ
We previously found that r-hu-IFNgamma exerts a potent anti-tumor effect on human nasopharyngeal carcinoma xenografts in vivo. Considering the fact that the clinical use of recombinant IFNgamma is ...limited by its short half-life and systemic side effects, we developed a recombinant adenovirus, Ad-IFNgamma.
Dynamic distribution of the adenovirus vector and expression of IFNgamma were evaluated by Q-PCR and ELISA after intratumoral administration of Ad-IFNgamma into CNE-2 xenografts.
Ad-IFNgamma DNA was mainly enriched in tumors where the Ad-IFNgamma DNA was injected (P < 0.05, compared to blood or parenchymal organs), as well as in livers (P < 0.05). Concentrations of Ad-IFNgamma DNA in other organs and blood were very low. Intratumoral Ad-IFNgamma DNA decreased sharply at high concentrations (9 x 10(5) copies/microg tissue DNA), and slowly at lower concentrations (1.7-2.9 x 10(5) copies/microg tissue DNA). IFNgamma was detected in the tumors and parenchymal organs. The concentration of IFNgamma was highest in the tumor (P < 0.05), followed by the liver and kidney (P < 0.05). High-level intratumoral expression of IFNgamma was maintained for at least 7 days, rapidly peaking on day 3 after injection of Ad-IFNgamma DNA.
An IFNgamma gene delivered by an adenoviral vector achieved high and consistent intratumoral expression. Disseminated Ad-IFNgamma DNA and the transgene product were mainly enriched in the liver.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
e19019
Background: Nucleophosmin (NPM1) is predominant nucleolar localization. NPM1-mutant acute myeloid leukemia (AML) account for 25-35% of adult AML patients, has been defined as a distinct AML ...entity in the 2022 WHO classification. The majority of NPM1 mutations reported affect exon 12 (classical type A mutation), account for 75-80% of adult NPM1-mutated AML cases, which generate an extra nuclear export signal (NES) in the C-terminal leading to aberrant cytoplasmic localization. Similar aberrant cytoplasmic translocation has also been reported in NPM1 mutants that identified in exons 9 and 11. Identification of novel non-type A mutants is paramount for diagnosis, risk stratification, and prognosis for targeted population. Methods: Somatic mutation analysis is performed on 566 AML patients using next-generation sequencing (NGS). Immunofluorescence and immunohistochemistry were used to identify the cytological localization of mutant NPM1. HEK-293T cells transiently transfected with plasmids for ectopic expression of the GFP-mutated NPM1 fusion proteins were treated with the specific Crm1/XPO1 inhibitor leptomycin B and to evaluate the NES dependence of their subcellular localization. Results: We identified one AML patient with NPM1 mutant located on exon 5. This patient was a 59-year-old female with de novo AML, with an 18-nucletide in-frame insertion at position 405 (c.405_406insGCCCTGGAACTGGGGAAC, named MutSong) in the middle of exon 5 variant allele fraction (VAF), 11.9%. It generated a new NPM1 mutant protein (p.135insALELGN), and containing a leucine-rich NES. Notably, different with all NPM1 mutant in other exons reported so far, our exon 5 mutant retained the functional c-terminal but insert one leucine-rich NES, similar finding was recently reported in NPM1 mutants that identified in exon 5 but on different position. This NPM1 mutant in exon 5 resulted in cytoplasmic localization both by confocal and immunohistochemistry in AML patient sample. HEK-293T overexpressing the new GFP-NPM1 exon 5 fusion protein indicated the aberrant localization in the cytoplasm and partially in nucleoli. Moreover, NES-dependent cytoplasmic localization was inhibited by exportin-1 inhibitor leptomycin B, indicated that the novel NES generated by exon 5 mutant is responsible for cytoplasmic localization. This patient carried WT1, IKZF1, JAK2, and NUP98 mutations, and was intravenously treated with daunorubicin plus cytarabine for induction followed by 3 cycles of intermediate-dose cytarabine as consolidation. Then the patient received allogeneic hematopoietic stem cell transplantation (allo-HSCT). Unfortunately, the patient died of severe pulmonary infection and viral encephalitis 14 months after the initial remission, 7 months after allo-HSCT. Conclusions: Our finding of the novel NPM1 mutation in exon 5 supported that beside exon 12, exon 5 mutant is another NPM1 “born to be exported” mutant critical for leukemogenesis.
The serine protease inhibitor clade E member 1 (SERPINE1) is a major inhibitor of tissue plasminogen activator and urokinase, and has been implicated in the development and progression of a variety ...of tumors. In this study, mRNA microarray and TCGA database were used to comprehensively analyze the upregulation of SERPINE1 in gastric cancer (GC) tissues compared with the normal stomach tissues. Kaplan-Meier results confirmed that patients with high SERPINE1 expression exhibited worse overall survival and disease-free survival. In addition, cell proliferation, cell scratches, transwell migration and invasion assay showed that SERPINE1 knockdown inhibited the proliferation, migration and invasion of GC ells. Western blot showed that the expression of VEGF and IL-6 was significantly upregulated after overexpression of SERPINE1. Meanwhile, SERPINE1 was positively correlated with the level of immune infiltration using the online analysis tools TISIDB and TIMER. And SERPINE1 expression increased with the increase of malignancy of GC which were detected by Immunohistochemistry. Finally, tumorigenesis experiments in nude mice further demonstrated that SERPINE1 could promote the occurrence and development of GC, while deletion of SERPINE1 inhibited the progression of GC. In summary, SERPINE1 was highly expressed in GC tissues, and SERPINE1 was helpful for differential diagnosis of pathological grade of gastric mucosal lesions. SERPINE1 might regulate the expression of VEGF and IL-6 through the VEGF signaling pathway and JAK-STAT3 inflammatory signaling pathway, thus ultimately affecting the invasion and migration of GC cells.
•A 3D micromechanical model for self-healing cementitious composites is established.•The evolutionary domains of microcrack growth (DMG) are obtained.•The evolutionary compliances and constitutive ...relations are investigated.•The healing effect is demonstrated on the mesoscale under tensile loading.•The effects of various model parameters on self-healing effect are studied.
For the past several years, research in the field of self-healing construction materials becomes a hotspot with broad application prospects. This paper mainly focuses on the self-healing cementitious composites with microcapsules. To quantitatively interpret the self-healing effect of micro-encapsulated healing agents on microcrack-induced damage, a three-dimensional evolutionary micromechanical model is established to predict the mechanical response of the cementitious composites with the microcapsules subjected to tensile loading during the damage-healing process. The evolutionary domains of microcrack growth (DMG) and the corresponding compliances at the initial, activated and repaired stages are obtained. On the basis of the proposed 3D micromechanical model of the self-healing cementitious composites with microcapsules, elaborate studies of constitutive relations and compliance are conducted to investigate the effects of various system parameters involving the healing efficiency, fracture toughness and preloading-induced damage degrees on the compliances and stress-strain relations. The results indicate that relatively significant healing efficiency, preloading-induced damage degree and the fracture toughness of polymerized healing agent with the matrix will lead to higher tensile strength and stiffness. However, based on the two different failure modes of self-healing concrete, the excessive values of healing efficiency, preloading-induced damage degree and the fracture toughness of polymerized healing agent with the matrix will not affect the tensile strength of the cementitious composites. For the sake of the desired optimal healing effect, the specific parameters of both the matrix and the microcapsules should be selected carefully.
Characterizing changes in rock properties is essential for the hydraulic fracture and re-fracture parameter optimization of shale formations. This paper proposed a hydraulic fracturing model to ...investigate the changes in rock properties during hydraulic fracturing using SPH, and the changes in the stress field and rock properties were quantitatively characterized. The simulation results indicated that the minimum horizontal principal stress increased by 10 MPa~15 MPa during fracture propagation, which is the main reason for the uneven propagation in multi-fracture propagation. Affected by the stress disturbance, the stimulated area was divided into four parts based on the changes in Young’s modulus and permeability; the more seriously the stress disturbance was affected, the higher the permeability of the stimulated zone was, and the smaller the stimulated zone was. Meanwhile, a zone with reduced permeability appeared due to the compression effect caused by the high injection pressure, and this increased with the increase in stress disturbance. The main reason for this was that strain formed because of the compression effect from the high injection pressure. The higher the stress disturbance, the higher the accumulated strain. This new model provides a new method for fracture parameter optimization, which also provides a foundation for the re-fracture parameter optimization of shale formations.
Lycium barbarum has been used as a traditional Chinese medicine to nourish liver, kidneys and the eyes. However, the underlying mechanisms of its hepatic-protective properties remain uncertain. In ...this study, we aimed to investigate whether thioredoxin-interacting protein (TXNIP) and NOD-like receptor 3 (NLRP3) inflammasome mediated the attenuation of ethanol-induced hepatic injury by Lycium barbarum polysaccharide (LBP). Rat normal hepatocyte line BRL-3A was pre-treated with LBP prior to ethanol incubation. Hepatic damages, including apoptosis, inflammation, and oxidative stress, were measured. Then the inhibition of endogenous TXNIP expression was achieved by using its specific siRNA to test its possible involvement in the injury attenuation. We found that 50μg/ml LBP pre-treatment significantly alleviated 24-h ethanol exposure-induced overexpression of TXNIP, increased cellular apoptosis, secretion of inflammatory cytokines, activation of NLRP3 inflammasome, production of ROS, and reduced antioxidant enzyme expression. Silence of TXNIP suppressed the activated NLRP3 inflammasome, increased oxidative stress and worsened apoptosis in the cells. Further addition of LBP did not influence the effects of TXNIP inhibition on the cells. In conclusion, inhibition of hepatic TXNIP by LBP contributes to the reduction of cellular apoptosis, oxidative stress and NLRP3 inflammasome-mediated inflammation.
The presence of FMS-like receptor tyrosine kinase-3 internal tandem duplication (FLT3-ITD) mutations in patients with acute myeloid leukemia (AML) is associated with poor clinical outcome. FLT3 ...tyrosine kinase inhibitors (TKIs), although effective in kinase ablation, do not eliminate primitive FLT3-ITD+ leukemia cells, which are potential sources of relapse. Thus, understanding the mechanisms underlying FLT3-ITD+ AML cell persistence is essential to devise future AML therapies. Here, we show that expression of protein arginine methyltransferase 1 (PRMT1), the primary type I arginine methyltransferase, is increased significantly in AML cells relative to normal hematopoietic cells. Genome-wide analysis, coimmunoprecipitation assay, and PRMT1-knockout mouse studies indicate that PRMT1 preferentially cooperates with FLT3-ITD, contributing to AML maintenance. Genetic or pharmacological inhibition of PRMT1 markedly blocked FLT3-ITD+ AML cell maintenance. Mechanistically, PRMT1 catalyzed FLT3-ITD protein methylation at arginine 972/973, and PRMT1 promoted leukemia cell growth in an FLT3 methylation–dependent manner. Moreover, the effects of FLT3-ITD methylation in AML cells were partially due to cross talk with FLT3-ITD phosphorylation at tyrosine 969. Importantly, FLT3 methylation persisted in FLT3-ITD+ AML cells following kinase inhibition, indicating that methylation occurs independently of kinase activity. Finally, in patient-derived xenograft and murine AML models, combined administration of AC220 with a type I PRMT inhibitor (MS023) enhanced elimination of FLT3-ITD+ AML cells relative to AC220 treatment alone. Our study demonstrates that PRMT1-mediated FLT3 methylation promotes AML maintenance and suggests that combining PRMT1 inhibition with FLT3 TKI treatment could be a promising approach to eliminate FLT3-ITD+ AML cells.
•PRMT1 promotes survival and growth of FLT3-ITD+ AML cells through methylating FLT3 protein at arginine residues 972 and 973.•PRMT1 inhibition enhances elimination of FLT3-ITD+ AML cells by FLT3 TKI treatment.
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Background: Long non-coding RNA (lncRNA) plays an essential regulatory role in the occurrence and development of hepatocellular carcinoma (HCC). This paper aims to establish an immune-related lncRNA ...(irlncRNA) pairs model independent of expression level for risk assessment and prognosis prediction of HCC. Methods: Transcriptome data and corresponding clinical data were downloaded from TCGA. HCC patients were randomly divided into training group and test group. Univariate Cox regression analysis, LASSO regression analysis, and stepwise multiple Cox regression analysis were used to establish a prognostic model. The prediction ability of the model was verified by ROC curves. Next, the patients were divided into low-risk and high-risk groups. We compared the differences between the two groups in survival rate, clinicopathological characteristics, tumor immune cell infiltration status, chemotherapeutic drug sensitivity and immunosuppressive molecules. Results: A prognosis prediction model was established based on 7 irlncRNA pairs, namely irlncRNA pairs (IRLP). ROC curves of the training group and test group showed that the IRLP model had high sensitivity and specificity for survival prediction. Kaplan-Meier analysis showed that the survival rate of the high-risk group was significantly lower than that of the low-risk group. Immune cell infiltration analysis showed that the high-risk group was significantly correlated with various immune cell infiltration. Finally, there were statistically significant differences in chemosensitivity and molecular marker expression between the two groups. Conclusion: The prognosis prediction model established by irlncRNA pairs has a certain guiding significance for the prognosis prediction of HCC. It may provide valuable clinical applications in antitumor immunotherapy. Keywords: hepatocellular carcinoma, long non-coding RNA, immune cell infiltration, prognostic model, immunotherapy
Inversion of chromosome 16, inv(16) inv(16)(p13q22) or t(16;16)(p13.1;q22), is a recurrent chromosomal translocation observed in 5-8% of acute myeloid leukemia (AML) cases. Inv(16) creates a fusion ...gene CBFb-MYH11 (CM) which impairs hematopoietic differentiation and creates abnormal progenitor populations prone to leukemic transformation. We reported that CM interacts with HDAC8 and enhances its activity, and high HDAC8 activity impaired DNA damage response (DDR) in CM knock-in (KI) hematopoietic stem and progenitor cells (HSPCs) (Zhang, L et al, ASH meeting 2022, https://doi.org/10.1182/blood-2022-160280). The recruitment of BRCA1-mRNA splicing machinery is critical for efficient DDR and the U2 small nuclear RNA auxiliary factor 1 (U2AF1) is required for accurate 3'-splice site selection. Here, we show how U2AF1 is post-translationally regulated following DNA damage and its impact on alternative splicing in CM KI HSPCs upon DNA damage. First, we identified TIP60 as the histone acetyltransferase (HAT) responsible for the increased U2AF1 acetylation induced by ionizing radiation (IR). We detected enhanced interaction between U2AF1 and TIP60 but not with other HATs (GCN5, PCAF, CBP) along with the increased U2AF1 acetylation in response to IR. In addition, TIP60 knockdown (KD) reduced U2AF1 acetylation, while TIP60 expression increased U2AF1 acetylation, indicating that TIP60 mediates the acetylation of U2AF1. To pinpoint the specific lysine residue subjected to TIP60 catalysis, we introduced lysine site-specific mutants U2AF1-HA-K15R, K23R, K39R, K175R, or 4 lysine sites mutant (U2AF1-HA-4mut) with TIP60. Co-immunoprecipitation (anti-HA) followed by western blotting (anti-acetyl-lysine) showed undetectable acetylation of U2AF1-HA-K23R and K175R mutants (similar to U2AF1-HA-4mut), suggesting that K23 and K175 residues are essential for TIP60-mediated acetylation. To investigate the impact of K23 or K175 acetylation on the assembly of the BRCA1-mRNA splicing complex and DDR, we subjected U2AF1-K23R or K175R expressing 32D cells to IR (3.5 Gy). Both U2AF1-K23R and K175R mutants exhibited markedly reduced acetylation levels after IR, confirming that K23 and K175 sites are the acetylation sites responding to DNA damage. Furthermore, the interaction of U2AF1-K23R and K175R with BRCA1 and BCLAF1 was impaired. We observed increased apoptosis in U2AF1-K23R and K175R cells compared to wild-type (WT) U2AF1 cells (K23R vs WT, 51.975±7.867% vs 6.055±0.634%, P=0.0011; K175R vs WT,22.645±3.037% vs 6.055±0.634%, P=0.0017) after KD of endogenous U2AF1 and subjected to IR (3.5 Gy). These results indicate that U2AF1 is acetylated by TIP60 upon DNA damage on K23 and K175 which is critical for cell survival and assembly of the BRCA1-mRNA splicing complex in DDR. Our studies revealed that HDAC8 interacts with U2AF1 and modulates U2AF1 acetylation following DNA damage and acetylation on U2AF1-K23 is important for HDAC8 binding. Consistent with the enhanced HDAC8 activity induced by CM, we show that CM also diminished U2AF1 acetylation and impaired the assembly of the BRCA1-mRNA splicing complex after IR. To examine the consequences in alternative splicing events upon IR, we sorted CM KI (n=5) or control (n=3) Lin-Sca1+Kit+ (LSK) cells and exposed them to IR (2.0 Gy) followed by RNA-seq. We identified a total of 829 splicing events in WT-IR vs WT-NIR (NIR: no IR) and 643 splicing events in CM-IR vs CM-NIR. Specifically, we found that skipped exon (SE) and retained intron (RI) events are selectively reduced in CM vs WT (SE events: 496 vs 653; RI events: 2 vs 26). Gene set enrichment analysis revealed striking differential enrichment in 38 BRCA1-related pathways, including regulation of response to DNA damage stimulus (NES: -1.0816 vs 0.8707), regulation of DNA repair (NES: -1.1557 vs 0.7729), cell cycle checkpoint signaling (NES: -0.9719 vs 0.7895), and recombinational repair (NES: -0.9215 vs 0.7299). Altogether, these studies reveal mechanistic insights into DNA damage induced regulation of post-translational acetylation of U2AF1 splicing factor and provide a mechanistic link to the dysregulated alternative splicing and impaired DNA damage response in inv(16) AML.
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