Human Leukemia Cells Research Articles

A novel type IIb L-asparaginase from Latilactobacillus sakei LK-145: characterization and application.

We succeeded in homogeneously expressing and purifying L-asparaginase from Latilactobacillus sakei LK-145 (Ls-Asn1) and its mutated enzymes C196S, C264S, C290S, C196S/C264S, C196S/C290S, C264S/C290S, and C196S/C264S/C290S-Ls-Asn1. Enzymological studies using purified enzymes revealed that all cysteine residues of Ls-Asn1 were found to affect the catalytic activity of Ls-Asn1 to varying degrees. The mutation of Cys196 did not affect the specific activity, but the mutation of Cys264, even a single mutation, significantly decreased the specific activity. Furthermore, C264S/C290S- and C196S/C264S/C290S-Ls-Asn1 almost completely lost their activity, suggesting that C290 cooperates with C264 to influence the catalytic activity of Ls-Asn1. The detailed enzymatic properties of three single-mutated enzymes (C196S, C264S, and C290S-Ls-Asn1) were investigated for comparison with Ls-Asn1. We found that only C196S-Ls-Asn1 has almost the same enzymatic properties as that of Ls-Asn1 except for its increased stability for thermal, pH, and the metals NaCl, KCl, CaCl2, and FeCl2. We measured the growth inhibitory effect of Ls-Asn1 and C196S-Ls-Asn1 on Jurkat cells, a human T-cell acute lymphoblastic leukemia cell line, using L-asparaginase from Escherichia coli K-12 as a reference. Only C196S-Ls-Asn1 effectively and selectively inhibited the growth of Jurkat T-cell leukemia, which suggested that it exhibited antileukemic activity. Furthermore, based on alignment, phylogenetic tree analysis, and structural modeling, we also proposed that Ls-Asn1 is a so-called "Type IIb" novel type of asparaginase that is distinct from previously reported type I or type II asparaginases. Based on the above results, Ls-Asn1 is expected to be useful as a new leukemia therapeutic agent.

Minnelide exhibits antileukemic activity by targeting the Ars2/miR-190a-3p axis

BackgroundThe identification of a novel and effective strategy for the clinical treatment of acute leukemia (AL) is a long-term goal. Minnelide, a water-soluble prodrug of triptolide, has recently been evaluated in phase I and II clinical trials in patients with multiple cancers and has shown promise as an antileukemic agent. However, the molecular mechanism underlying minnelide's antileukemic activity remains unclear. PurposeTo explore the molecular mechanisms by which minnelide exhibits antileukemic activity. MethodsAL cells, primary human leukemia cells, and a xenograft mouse model were treated with triptolide and minnelide. The molecular mechanism was elucidated using western blotting, immunoprecipitation, flow cytometry, GSEA and liquid chromatography-mass spectrometry analysis. ResultsMinnelide was highly effective in inhibiting leukemogenesis and improving survival in two complementary AL mouse models. Triptolide, an active form of minnelide, causes cell cycle arrest in G1 phase and induces apoptosis in both human AL cell lines and primary AL cells. Mechanistically, we identified Ars2 as a new chemotherapeutic target of minnelide for AL treatment. We found that triptolide directly targeted Ars2, resulting in the downregulation of miR-190a-3p, which led to the disturbance of PTEN/Akt signaling and culminated in G1 cell cycle arrest and apoptosis. ConclusionsOur findings demonstrate that targeting Ars2/miR-190a-3p signaling using minnelide could represent a novel chemotherapeutic strategy for AL treatment and support the evaluation of minnelide for the treatment of AL in clinical trials.

Study on the inhibitory and pro-apoptotic effects of different concentrations of total tanshinone alone and in combination with tyrosine kinase inhibitors on human myeloid leukemia cell lines

Objective: To explore the effect and investigate the molecular mechanism of different concentrations of total tanshinones alone and in combination with tyrosine kinase inhibitors (TKIs) on the proliferation inhibition and apoptosis of human myeloid leukemia cell lines. Methods: K562 and Kasumi-1 cell lines were purchased from the Shanghai Cell Bank of the Chinese Academy of Sciences, and the TKIs-resistant strain K562/T315I cell line was constructed in Molecular Medicine Research Center, Beijing Lu Daopei Institute of Hematology. Logarithmic growth phase cells were taken and divided into intervention groups with total tanshinone of 0, 2.19, 4.38, 8.75, 17.50 and 35.00 μg/ml intervention groups, which were inoculated in 96-well plates at a density of 1×104 cells/well and exposed to the drug for 24 h, and a control group treated with dimethyl sulfoxide was also set up simultaneously. All experiments were repeated independently 3-5 times. The proliferative activity of the cells was assessed using the CCK-8 assay, the apoptotic rates were measured by flow cytometry, and the expression levels of apoptosis-regulating proteins Bcl-2 and Bax were analyzed by Western blotting. The cell lines treated and untreated with total tanshinone were subjected to transcriptome sequencing and gene set enrichment analysis to identify differentially expressed genes. Results: The half-inhibitory concentration (IC50) values of 8.75 μg/ml total tanshinone at 24 h for K562, K562/T315I and Kasumi-1 cells were (4.11±0.02), (4.95±0.04) and (3.98±0.01) μg/ml, respectively. When combined with 0.25 μmol/L imatinib, 8.75 μg/ml total tanshinone could enhance the induction of apoptosis effects on K562 and K562/T315I cell lines. After being treated with 4.38, 8.75, and 17.50 μg/ml of total tanshinone for 24 h, compared with the control group, total tanshinone upregulated the expression level of Bax protein, downregulated the expression level of Bcl-2 protein, and decreased the Bcl-2/Bax ratio (all P<0.05). Total tanshinone inhibited the proliferation-related signaling pathway and DNA damage repair pathway of myeloid leukemia cell lines, and activated the signaling pathway that induces apoptosis in leukemia cells. Conclusion: Different concentrations of total tanshinoneinhibites proliferation and promote apoptosis in K562, Kasumi-1 and TKIs-resistant K562/T315I cell lines, and further enhance the anti-leukemic effect when combined with TKIs.

Sulfated polysaccharides from the viscera of Mustelus shark: Characterization and antioxidant, anticoagulant and anti-proliferative activities

In the present study, sulfated polysaccharides (SPs) from smooth hound shark were extracted using different precipitation agents, the cetylpyridinium chloride or ethanol, named SHSP-I and SHSP-II, respectively. The UV–visible scan demonstrated the absence of nucleic acids or proteins in the extracted SHSPs. The antioxidant, anticoagulant and anti-proliferative activities were also investigated. Results showed that SHSPs displayed reducing and scavenging capacities, as shown by the oxygen radical absorbance capacity, the total antioxidant capacity and the hydroxyl radical-scavenging activities. Moreover, extracted SPs were able to prolong the activated partial thromboplastin time (aPTT), the prothrombin time (PT) and the thrombin time (TT) and they reduced the fibrinogen level (FL), without inducing erythrocytes hemolysis, which indicated that they inhibited the intrinsic and extrinsic coagulation pathways and thrombin-mediated fibrin formation. The SHSP-I was more effective on prolonging the blood clotting time by more than 120 s at 1 mg/ml in the aPTT and about 70 s and 60 s at 2.5 mg/ml in the PT and TT, respectively, compared to the SHSP-II. The anti-proliferative activity of SHSPs was tested against the growth of K562 human myelogenous leukemia cells and Caco-2 colon cancer cells. Data showed that both polysaccharides have a dose-dependent inhibitory effect, where SHSP-I exhibited the strongest activity against the tested cells and the K562 was more sensitive than Caco-2 line cells to the action of SHSPs. The overall data suggested that SPs from shark viscera could be useful to be used as nutraceutical agents.

Variable characteristics overlooked in human K-562 leukemia cell lines with a common signature

K-562 is a well-known in vitro cellular model that represents human leukemia cell lines. Although the K-562 cells have been extensively characterized, there are inconsistencies in the data across publications, showing the presence of multiple K-562 cell lines. This suggests that analyzing a single K-562 cell line is insufficient to provide reliable reference data. In this study, we compared three K-562 cell lines with different IDs (RCB0027, RCB1635, and RCB1897) to investigate the fundamental characteristics of K-562 cells. Amplifications of the BCR-ABL1 fusion gene and at 13q31 were detected in all three cell lines, whereas each genome exhibited distinctive features of sequence variants and loss of heterozygosity. This implies that each K-562 cell line can be characterized by common and unique features through a comparison of multiple K-562 cell lines. Variations in transcriptome profiles and hemoglobin synthesis were also observed among the three cell lines, indicating that they should be considered sublines that have diverged from the common ancestral K-562 despite no changes from the original cell name. This leads to unintentional differences in genotypes and/or phenotypes among cell lines that share the same name. These data show that characterizing a single K-562 cell line does not necessarily provide data that are applicable to other K-562 cells. In this context, it is essential to modify cell names in accordance with changes in characteristics during cell culture. Furthermore, our data could serve as a reference for evaluating other K-562 sublines, facilitating the discovery of new K-562 sublines with distinct characteristics. This approach results in the accumulation of K-562 sublines with diverged characteristics and expands the options available, which may help in selecting the most suitable K-562 subline for each experiment.

TurboID-Based IRE1 Interactome Reveals Participants of the Endoplasmic Reticulum-Associated Protein Degradation Machinery in the Human Mast Cell Leukemia Cell Line HMC-1.2.

The unfolded protein response is an intricate system of sensor proteins in the endoplasmic reticulum (ER) that recognizes misfolded proteins and transmits information via transcription factors to either regain proteostasis or, depending on the severity, to induce apoptosis. The main transmembrane sensor is IRE1α, which contains cytoplasmic kinase and RNase domains relevant for its activation and the mRNA splicing of the transcription factor XBP1. Mast cell leukemia (MCL) is a severe form of systemic mastocytosis. The inhibition of IRE1α in the MCL cell line HMC-1.2 has anti-proliferative and pro-apoptotic effects, motivating us to elucidate the IRE1α interactors/regulators in HMC-1.2 cells. Therefore, the TurboID proximity labeling technique combined with MS analysis was applied. Gene Ontology and pathway enrichment analyses revealed that the majority of the enriched proteins are involved in vesicle-mediated transport, protein stabilization, and ubiquitin-dependent ER-associated protein degradation pathways. In particular, the AAA ATPase VCP and the oncoprotein MTDH as IRE1α-interacting proteins caught our interest for further analyses. The pharmacological inhibition of VCP activity resulted in the increased stability of IRE1α and MTDH as well as the activation of IRE1α. The interaction of VCP with both IRE1α and MTDH was dependent on ubiquitination. Moreover, MTDH stability was reduced in IRE1α-knockout cells. Hence, pharmacological manipulation of IRE1α-MTDH-VCP complex(es) might enable the treatment of MCL.

Discovery of a Novel Orally Bioavailable FLT3-PROTAC Degrader for Efficient Treatment of Acute Myeloid Leukemia and Overcoming Resistance of FLT3 Inhibitors.

Fms-like tyrosine receptor kinase 3 (FLT3) proteolysis-targeting chimeras (PROTACs) represent a promising approach to eliminate the resistance of FLT3 inhibitors. However, due to the poor druggability of PROTACs, the development of orally bioavailable FLT3-PROTACs faces great challenges. Herein, a novel orally bioavailable FLT3-ITD degrader A20 with excellent pharmacokinetic properties was discovered through reasonable design. A20 selectively inhibited the proliferation of FLT3-ITD mutant acute myeloid leukemia (AML) cells and potently induced FLT3-ITD degradation through the ubiquitin-proteasome system. Notably, oral administration of A20 resulted in complete tumor regression on subcutaneous AML xenograft models. Furthermore, on systemic AML xenograft models, A20 could completely eliminate the CD45+CD33+ human leukemic cells in murine and significantly prolonged the survival time of mice. Most importantly, A20 exerted significantly improved antiproliferative activity against drug-resistant AML cells compared to existing FLT3 inhibitors. These findings suggested that A20 could serve as a promising drug candidate for relapsed or refractory AML.

Association of poly(rC)-binding protein-2 with sideroflexin-3 through TOM20 as an iron entry pathway to mitochondria

Iron is essential for all the lives and mitochondria integrate iron into heme and Fe-S clusters for diverse use as cofactors. Here, we screened mitochondrial proteins in KU812 human chronic myelogenous leukemia cells by glutathione S-transferase pulldown assay with PCBP2 to identify mitochondrial receptors for PCBP2, a major cytosolic Fe(II) chaperone. LC–MS analyses identified TOM20, sideroflexin-3 (SFXN3), SFXN1 and TOM70 in the affinity-score sequence. Stimulated emission depletion microscopy and proteinase-K digestion of mitochondria in HeLa cells revealed that TOM20 is located in the outer membrane of mitochondria whereas SFXN3 is located in the inner membrane. Although direct association was not observed between PCBP2 and SFXN3 with co-immunoprecipitation, proximity ligation assay demonstrated proximal localization of PCBP2 with TOM20 and there was a direct binding between TOM20 and SFXN3. Single knockdown either of PCBP2 and SFXN3 in K562 leukemia cells significantly decreased mitochondrial catalytic Fe(II) and mitochondrial maximal respiration. SFXN3 but not MFRN1 knockout (KO) in mouse embryonic fibroblasts decreased FBXL5 and heme oxygenase-1 (HO-1) but increased transferrin uptake and induced ferritin, indicating that mitochondrial iron entry through SFXN3 is distinct. MFRN1 KO revealed more intense mitochondrial Fe(II) deficiency than SFXN3 KO. Insufficient mitochondrial heme synthesis was evident under iron overload both with SFXN3 and MFRN KO, which was partially reversed by HO-1 inhibitor. Conversely, SFXN3 overexpression caused cytosolic iron deficiency with mitochondrial excess Fe(II), which further sensitized HeLa cells to RSL3-induced ferroptosis. In conclusion, we discovered a novel pathway of iron entry into mitochondria from cytosol through PCBP2-TOM20-SFXN3 axis.

Near-infrared inducible supports in bio-catalysts design: A useful and versatile tool in enhancement of enzyme activity

Immobilized enzymes have encountered two main challenges: Reduced enzyme activity compared to free enzymes and exhausted immobilized enzymes due to reusability. Herein, we suggested a promising activity enhancement strategy to overcome these challenges. The emission from upconversion nanoparticles (UCNPs) under near-infrared (NIR) excitation can increase the activity of PEG-l-ASNase due to Förster Resonance Energy Transfer. For this purpose, UCNPs were initially synthesized using the hydrothermal method. Subsequently, these UCNPs were functionalized with a polycationic polymer, branched polyethyleneimine (PEI), and the immobilization of PEG-l-ASNase was achieved through adsorption. We preliminarily explored the parameters such as enzyme concentration, incubation time, pH, temperature, reusability, storage stability, and kinetic study, etc. Further, the in vitro biocompatibility, hemolytic behavior, and anticancer activity of the produced UCNPs were also analyzed as crucial parameters. The results showed the pH durance, thermal and storage stability of the immobilized PEG-l-ASNases were enhanced. The immobilized PEG-l-ASNases maintained their activity to ≥55 % after 20 cycles. Enzyme immobilization led to a decrease in Km and Vmax compared to PEG-l-ASNase. In vitro assays revealed that immobilized enzyme further reduced the proliferation of human leukemia cell line (HL-60) upon NIR irradiation exposure but did not cause toxicity. This research may provide a new strategy to promote the catalytic activity of l-ASNase and demonstrates its potential application on human leukemia cells. Finally, these outcomes are valuable for the use of NIR induction in enzymatic reactions.

Elesclomol-Cu Induces Cuproptosis in Human Acute Myeloid Leukemia Cells

To investigate the effects of elesclomol-Cu (ES-Cu) on the proliferation and cuproptosis of human acute myeloid leukemia (AML) cells. The effects of ES-Cu on the proliferation of AML cells and the AML cells pre-treated with ammonium tetrathiomolybdate (TTM) were examined by CCK-8 assay. The Calcein/PI kit was used to detected the changes in activity and cytotoxicity of AML cells induced by ES-Cu. Flow cytometry and Cytation3 fully automated cell imaging multifunctional detection system were used to analyze DCFH-DA fluorescence intensity, so as to determine the level of reactive oxygen species (ROS). The GSH and GSSG detection kits were used to measure the intracellular GSH content. Western blot was used to detected the expression of cuproptosis-related proteins ATP7B, FDX1, DLAT and DPYD. ES-Cu inhibited the proliferation of Kasumi-1 and HL-60 cells in a concentration-dependent manner (r Kasumi-1=-0.99， r HL-60=-0.98). As the concentration of ES-Cu increased, the level of intracellular ROS also increased (P <0.01-0.001). TTM could significantly reverse the inhibitory effect of ES-Cu on cell proliferation and its promoting effect on ROS. With the increase of ES-Cu concentration, the content of GSH was decreased (r =-0.98), and Western blot showed that the protein expressions of ATP7B, FDX1, DLAT and DPYD were significantly reduced (P <0.05). ES-Cu can induce cuproptosis in AML cells, which provides a new idea for the treatment of AML.

Effect of Shikonin on Autophagy and Apoptosis of Human Promyelocytic Leukemia Cells

To explore the effect of shikonin on autophagy and apoptosis of human promyelocytic leukemia cells and its possible mechanism. Human promyelocytic leukemia cells NB4 in the logarithmic growth phase were divided into control group (untreated NB4 cells), shikonin group (0.3 µmol/L shikonin treatment), 740Y-P group (15 µmol/L PI3K/Akt/mTOR pathway activator 740Y-P treatment), shikonin+740Y-P group (0.3 µmol/L shikonin and 15 µmol/L 740Y-P co-treatment), after 24 hours of treatment, the cells were used for subsequent experiments. CCK-8 method was used to detect cell viability, monodansylcadaverine (MDC) staining to detect the aggregation of autophagic vesicles, flow cytometry to detect cell apoptosis, and Western blot to detect the expression of Beclin1, LC3, p62, Bax, cleaved caspase-3, Bcl-2 and PI3K/Akt/mTOR pathway related proteins. Compared with the control group, the purple punctate fluorescence intensity, apoptosis rate, Beclin1, LC3-Ⅱ/LC3-Ⅰ, cleaved caspase-3, and Bax protein expression in NB4 cells were increased in the shikonin group, while OD450 value (24, 48 h) and the expressions of Bcl-2 and p62 proteins were decreased (all P < 0.05). Compared with the control group, the purple punctate fluorescence intensity, apoptosis rate, Beclin1, LC3-Ⅱ/LC3-Ⅰ, cleaved caspase-3, and Bax protein expression in NB4 cells were decreased, while OD450 value (24, 48 h) and the expressions of Bcl-2 and p62 proteins were increased in the 740Y-P group (all P < 0.05). Compared with the shikonin group, the purple punctate fluorescence intensity, apoptosis rate, Beclin1, LC3-Ⅱ/LC3-Ⅰ, cleaved caspase-3, and Bax protein expression in NB4 cells were decreased, while OD450 value (24, 48 h) and the expressions of Bcl-2 and p62 proteins were increased in the shikonin+740Y-P group (all P < 0.05). Compared with the control group, the expression of PI3K/Akt/mTOR pathway related proteins p-PI3K, p-Akt, and p-mTOR in NB4 cells were significantly decreased in the shikonin group, while those in the 740Y-P group were increased (all P < 0.05). Compared with the shikonin group, the expressions of p-PI3K, p-Akt, and p-mTOR proteins in NB4 cells were significantly increased in the shikonin+740Y-P group (all P < 0.05). Shikonin may promote autophagy and apoptosis of NB4 cells by inhibiting PI3K/Akt/mTOR pathway.

Critical view on antimicrobial, antibiofilm and cytotoxic activities of quinazolin-4(3H)-one derived schiff bases and their Cu(II) complexes

A series of nine 2,3-disubstituted-quinazolin-4(3H)-one derived Schiff bases and their three Cu(II) complexes was prepared and tested for their antimicrobial activities against reference strains Staphylococcus aureus ATCC 29213 and Enterococcus faecalis ATCC 29212 and resistant clinical isolates of methicillin-resistant S. aureus (MRSA) and vancomycin-resistant E. faecalis (VRE). All the substances were tested in vitro against Mycobacterium tuberculosis H37Ra ATCC 25177, M. kansasii DSM 44162 and M. smegmatis ATCC 700084. While anti-enterococcal and antimycobacterial activities were insignificant, 3-[(E)-(2-hydroxy-5-nitrobenzylidene)amino]-2-(2-hydroxy-5-nitrophenyl)-2,3-dihydroquinazolin-4(1H)-one (SB3) and its Cu(II) complex (SB3–Cu) demonstrated bacteriostatic antistaphylococcal activity. In addition, both compounds, as well as the other two prepared complexes, showed antibiofilm activity, which resulted in a reduction of biofilm formation and eradication of mature S. aureus biofilm by 80% even at concentrations lower than the values of their minimum inhibitory concentrations. In addition, the compounds were tested for their cytotoxic effect on the human monocytic leukemia cell line THP-1. The antileukemic efficiency was improved by the preparation of Cu(II) complexes from the corresponding non-chelated Schiff base ligands.

LINC01686 affects LPS-induced cytokine expression via the miR-18a-5p/A20/STAT1 axis in THP-1 cells.

Long noncoding RNAs (lncRNAs) are crucial in regulating various physiological and pathological processes, including immune responses. LINC01686 is a lncRNA with previously uncharacterized functions in immune regulation. This study aims to investigate the function of LINC01686 in lipopolysaccharide (LPS)-induced inflammatory responses in the human monocytic leukemia cell line THP-1 and its potential regulatory mechanisms involving miR-18a-5p and the anti-inflammatory protein A20. THP-1 cells were stimulated with LPS to induce inflammatory responses, followed by analysis of LINC01686 expression levels. The role of LINC01686 in regulating the expression of interleukin (IL)-6, IL-8, A20, and signal transducer and activator of transcription 1 (STAT1) was examined using small interfering RNA-mediated knockdown. Additionally, the involvement of miR-18a-5p in LINC01686-mediated regulatory pathways was assessed by transfection with decoy RNAs mimicking the miR-18a-5p binding sites of LINC01686 or A20 messenger RNA. LINC01686 expression was upregulated in THP-1 cells following LPS stimulation. Suppression of LINC01686 enhanced LPS-induced expression of IL-6 and IL-8, mediated through increased production of reactive oxygen species. Moreover, LINC01686 knockdown upregulated the expression and activation of IκB-ζ, STAT1, and downregulated A20 expression. Transfection with decoy RNAs reversed the effects of LINC01686 suppression on A20, STAT1, IL-6, and IL-8 expression, highlighting the role of LINC01686 in sponging miR-18a-5p and regulating A20 expression. This study provides the first evidence that LINC01686 plays a critical role in modulating LPS-induced inflammatory responses in THP-1 cells by sponging miR-18a-5p, thereby regulating the expression and activation of A20 and STAT1. These findings shed light on the complex regulatory mechanisms involving lncRNAs in immune responses and offer potential therapeutic targets for inflammatory diseases.

The S1 spike protein of SARS-CoV-2 upregulates the ERK/MAPK signaling pathway in DC-SIGN-expressing THP-1 cells.

Dendritic cells, macrophages, neutrophils, and other antigen-presenting cells express various C-type lectin receptors that function to recognize the glycans associated with pathogens. The dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) binds various pathogens such as HIV glycoprotein 120, the Ebola glycoprotein, hemagglutinin, and the dengue virus glycoprotein in addition to the SARS-CoV-2 spike protein, and also triggers antigen-presenting cell endocytosis and immune escape from systemic infections. Many studies on the binding of SARS-CoV-2 spike protein with glycans have been published, but the underlying mechanism by which intracellular signaling occurs remains unclear. In this study, we report that the S1 spike protein of SARS-CoV-2 induces the phosphorylation of extracellular signal-regulated kinases (ERKs) in THP-1 cells, a DC-SIGN-expressing human monocytic leukemic cell line. On the other hand, the phosphorylation level of NF-κB remained unchanged under the same conditions. These data suggest that the major cell signaling pathway regulated by the S1 spike protein is the ERK pathway, which is superior to the NF-κB pathway in these DC-SIGN-expressing THP-1 cells and may contribute to immune hyperactivation in SARS-CoV-2 infections. Additionally, several glycans such as mannans, mannosylatedbovine serum albumin, the serum amyloid beta protein, andintracellular adhesion molecule 3 suppressed ERKphosphorylation,suggesting that these molecules are target molecules for SARS-CoV-2 infection by suppressing immune hyperactivation that occurs in the ERK signaling pathway.

Reversal Effect of Arctigenin on the Drug Resistance in Leukemia K562/A02 Cells and Its Mechanism

To study the effect of arctigenin(ARG) on adriamycin(ADM) resistance of leukemia cell line K562/A02 and the underlying mechanism. Human leukemia cell line K562 and ADM-resistant cell line K562/A02 were cultured and treated with 2.5-50 µmol/L ADM. Cell proliferation was measured using CCK-8 method, and half maximal inhibitory concentration (IC50) was calculated. K562/A02 cells were treated with different concentrations of ARG (1, 2, 4, 8, 16 mmol/L) to detect the effect of ARG on K562/A02 cells, and a suitable concentration (2 mmol/L) was selected for subsequent experiments. K562/A02 cells were treated with 2 mmol/L ARG and 5 µmol/L ADM, and cell apoptosis was detected by flow cytometry, the expression of P-gp, MRP, cleaved caspase-3, Bax, Bcl-2 proteins and the TLR4/NF-κB signaling pathway-related proteins were measured by Western blot. TLR4 overexpression plasmid was transfected into K562/A02 cells which were co-treated with ARG and ADM, then drug sensitivity and cell apoptosis were measured. The IC50 value of ADM on K562/A02 cells was 36.57 µmol/L, which was significantly higher than that on K562 cells (1.30 µmol/L). ARG with a concentration of ≤2 mmol/L did not have a significant effect on K562/A02 cells. 2 mmol/L ARG significantly reduced the IC50 of ADM on K562/A02 cells. In 5 µmol/L ADM-treated K562/A02 cells, compared with the control group, the apoptosis rate of K562/A02 cells in the ARG group was significantly increased, the expressions of cleaved caspase-3, Bax proteins were significantly upregulated, the expressions of P-gp, MRP, Bcl-2, TLR4, MyD88, and p-NF-κB proteins were significantly downregulated, and the differences were statistically significant (P < 0.05). After transfection with TLR4 overexpression plasmid, the sensitivity of ARG-treated K562/A02 cells to ADM was reduced (P < 0.05), the cell apoptosis was decreased, and the expressions of P-gp, MRP, Bcl-2 and TLR4/NF-κB signaling pathway-related proteins were significantly elevated, while the expressions of cleaved caspase-3 and Bax proteins were significantly decreased (all P < 0.05). ARG may reverse the resistance of human leukemia cell line K562/A02 to ADM by inhibiting TLR4/NF-κB signaling pathway.

Ethnobotanical survey and biological activities of plants used for cancer treatment in traditional Senegalese medicine

Female breast cancer is known to be one of the leading causes of death in Senegal. In Senegal, the absence of a national cancer control program, the lack of specialized infrastructure and qualified human resources and the exorbitant cost of care have contributed to the extensive use of traditional medecine, particularly in rural areas. This study aims to inventory the medicinal plants used by these healers and to assess the cytotoxic and antioxidant activities of the most widely used one. Data on healers and their use practices and information on plants were collected through the administration of a structured questionnaire. Based on their citation frequencies during the survey, Antiaris Africana Engler, Hymenocardia Acida Tul. and Halouf Halal (local name) were selected for chemical and biological studies. Their hydroalcoholic extracts were analyzed in terms of antioxidant capacity and cytotoxic effects, again, in the human cancer cell line. The study revealed a total of 65 medicinal plants belonging to 35 different families. The plant parts used by traditional healers are leaves (63.89%), roots (11.11%), bark (15.28%), fruits (2.78%), and others (6.94%). Generally, herbal medicine is prepared as a powder and mixed with water by maceration (55.38%) and administered orally. A. Africana ranked first with a citation frequency of 5.7% and its hydroalcoholic extract had the highest antioxidant activity in TEAC (6533.64±7 μmol ET/g dry plant) and in ORAC (3745.17±4.8 μmol ET/g dry plant) followed by H. Acida in TEAC (3115.6±145 μmol ET/g dry plant) and in ORAC (4105.29±872 μmol ET/g dry plant). The hydroalcoholic extract of A. Africana exhibited the highest cytotoxic activity in MCF-7 (Human mammary) and THP-1 (Human acute monocytic leukemia cell line) but had low activity against HTC-116 (Human carcinoma colorectal) and A-375 (Human skin malignant melanoma). The percentages of proapoptotic cells were, respectively, 68.85±6.22, 58.1±1.90 and 48.58±1.4%. These results provide scientific support for the traditional use of medicinal plants in cancer treatment and constitute a database for biological screening to isolate cytotoxic plant-based molecules.

A complex interplay of intra- and extracellular factors regulates the outcome of fetal- and adult-derived MLL-rearranged leukemia

Infant and adult MLL1/KMT2A-rearranged (MLLr) leukemia represents a disease with a dismal prognosis. Here, we present a functional and proteomic characterization of in utero-initiated and adult-onset MLLr leukemia. We reveal that fetal MLL::ENL-expressing lymphomyeloid multipotent progenitors (LMPPs) are intrinsically programmed towards a lymphoid fate but give rise to myeloid leukemia in vivo, highlighting a complex interplay of intra- and extracellular factors in determining disease subtype. We characterize early proteomic events of MLL::ENL-mediated transformation in fetal and adult blood progenitors and reveal that whereas adult pre-leukemic cells are mainly characterized by retained myeloid features and downregulation of ribosomal and metabolic proteins, expression of MLL::ENL in fetal LMPPs leads to enrichment of translation-associated and histone deacetylases signaling proteins, and decreased expression of inflammation and myeloid differentiation proteins. Integrating the proteome of pre-leukemic cells with their secretome and the proteomic composition of the extracellular environment of normal progenitors highlights differential regulation of Igf2 bioavailability, as well as of VLA-4 dimer and its ligandome, upon initiation of fetal- and adult-origin leukemia, with implications for human MLLr leukemia cells’ ability to communicate with their environment through granule proteins. Our study has uncovered opportunities for targeting ontogeny-specific proteomic vulnerabilities in in utero-initiated and adult-onset MLLr leukemia.

Injury of Macrophages Induced by Clostridium perfringens Type C Exotoxins.

Clostridium perfringens is a kind of anaerobic Gram-positive bacterium that widely exists in the intestinal tissue of humans and animals. And the main virulence factor in Clostridium perfringens is its exotoxins. Clostridium perfringens type C is the main strain of livestock disease, its exotoxins can induce necrotizing enteritis and enterotoxemia, which lead to the reduction in feed conversion, and a serious impact on breeding production performance. Our study found that treatment with exotoxins reduced cell viability and triggered intracellular reactive oxygen species (ROS) in human mononuclear leukemia cells (THP-1) cells. Through transcriptome sequencing analysis, we found that the levels of related proteins such as heme oxygenase 1 (HO-1) and ferroptosis signaling pathway increased significantly after treatment with exotoxins. To investigate whether ferroptosis occurred after exotoxin treatment in macrophages, we confirmed that the protein expression levels of antioxidant factors glutathione peroxidase 4/ferroptosis-suppressor-protein 1/the cystine/glutamate antiporter solute carrier family 7 member 11 (GPX4/FSP1/xCT), ferroptosis-related protein nuclear receptor coactivator 4/transferrin/transferrin receptor (NCOA4/TF/TFR)/ferritin and the level of lipid peroxidation were significantly changed. Based on the above results, our study suggested that Clostridium perfringens type C exotoxins can induce macrophage injury through oxidative stress and ferroptosis.

Modification of Polyethylene Glycol-Hydroxypropyl Methacrylate Polymeric Micelles Loaded with Curcumin for Cellular Internalization and Cytotoxicity to Wilms Tumor 1-Expressing Myeloblastic Leukemia K562 Cells.

Curcumin loaded in micelles of block copolymers of ω-methoxypoly(ethylene glycol) and N-(2-hydroxypropyl) methacrylamide modified with aliphatic dilactate (CD) or aromatic benzoyl group (CN) were previously reported to inhibit human ovarian carcinoma (OVCAR-3), human colorectal adenocarcinoma (Caco-2), and human lymphoblastic leukemia (Molt-4) cells. Myeloblastic leukemia cells (K562) are prone to drug resistance and differ in both cancer genotype and phenotype from the three mentioned cancer cells. In the present study, CD and CN micelles were prepared and their effects on K562 and normal cells were explored. The obtained CD and CN showed a narrow size distribution with diameters of 63 ± 3 and 50 ± 1 nm, respectively. The curcumin entrapment efficiency of CD and CN was similarly high, above 80% (84 ± 8% and 91 ± 3%). Both CD and CN showed suppression on WT1-expressing K562 and high cell-cycle arrest at the G2/M phase. However, CD showed significantly higher cytotoxicity to K562, with faster cellular uptake and internalization than CN. In addition, CD showed better compatibility with normal red blood cells and peripheral blood mononuclear cells than CN. The promising CD will be further investigated in rodents and possibly in clinical studies for leukemia treatment.

Different Influence Pattern of Conventional and Alternative Sources of Smoking on Adhesion Molecules and Cytokine Secretion in THP-1 Monocytes.

Tobacco is a carcinogen that is closely associated with the occurrence of lung cancer and head and neck squamous cell carcinoma (HNSCC). The consumption of tobacco is also leading to alterations in different immune cell subtypes. However, the impact of different conventional and alternative smoking sources on human monocytes remains elusive. In this study, we investigated the influence of aqueous extracts of different sources of smoking (cigarettes; heated tobacco product IQOS; e-cigarettes with and without nicotine; nicotine pouches) on different monocytic adhesion molecules, chemokine receptors and checkpoint molecule PD-L1 by flow cytometry. Cytokine expression patterns were evaluated using human cytokine arrays and the human monocyte leukemia cell line THP-1 as a model. Data revealed differential effects of the analyzed conventional and alternative smoking devices on monocyte adhesion molecules and cytokine secretion. The examined smoking devices can be assigned to two differential monocyte activation patterns. Monocytes stimulated with aqueous extracts of cigarettes, e-cigarette without nicotine, and heat not burn product IQOS revealed distinct alterations of surface markers and cytokines compared to the monocyte activation pattern in response to aqueous extracts of nicotine, nicotine pouches, and e-cigarette with nicotine. Our data indicate differential immunological consequences of different conventional and alternative smoking sources with and without nicotine. Further comprehensive analysis as well as in vivo investigations on peripheral blood monocyte subsets from smoking individuals using different smoking sources are required to better understand the impact on monocyte characteristics, especially with regard to the development of cancer.