HDAC1/3-dependent moderate liquid-liquid phase separation of YY1 promotes METTL3 expression and AML cell proliferation.

Methyltransferase-like protein 3 (METTL3) plays critical roles in acute myeloid leukemia (AML) progression, however, the mechanism of abnormal overexpression of METTL3 in AML remain elusive. In the current study, we uncovered that Yin Yang 1 (YY1) binds to the promoter region of METTL3 as a transcription factor and promotes its expression, which in turn enhances the proliferation of AML cells. Mechanistically, YY1 binds to HDAC1/3 and regulates METTL3 expression in a moderate liquid-liquid phase separation (LLPS) manner. After mutation of the HDAC-binding site of YY1 or HDAC inhibitor (HDACi) treatment, YY1 was separated from HDAC1/3, which resulted in an excessive LLPS state, thereby inhibiting the expression of METTL3 and the proliferation of AML cells. In conclusion, our study clarified the regulatory mechanism of the abnormal expression of METTL3 in AML, revealed the precise "Yin-Yang" regulatory mechanism of YY1 from the perspective of LLPS degree, and provided new ideas for the precise diagnosis and treatment of AML.

DT7 peptide-modified lecithin nanoparticles co-loaded with γ-secretase inhibitor and dexamethasone efficiently inhibit T-cell acute lymphoblastic leukemia and reduce gastrointestinal toxicity.

T-cell acute lymphoblastic leukemia (T-ALL) is a serious hematologic malignancy and glucocorticoid resistance is the main recurrent cause for a high relapsed and death rate. Here, we proposed an effective therapeutic regimen of combining gamma-secretase inhibitors (GSIs) with dexamethasone (DEX) to overcome glucocorticoid resistance. Moreover, the bone marrow targeting DT7 peptide-modified lecithin nanoparticles co-loaded with DEX and GSI (TLnp/D&G) were developed to enhance T-ALL cells recognition and endocytosis. In vitro cytotoxicity studies showed that TLnp/D&G significantly inhibited cell survival and promoted apoptosis of T-ALL cells. Mechanically, we found that GSIs promoted DEX-induced cell apoptosis by two main synergetic mechanisms: 1) GSIs significantly upregulated glucocorticoid receptor (GR) expression in T-ALL and restored the glucocorticoid-induced pro-apoptotic response. 2) Both DEX and GSI synergistically inhibited BCL2 and suppressed the survival of T-ALL cells. Furthermore, in vivo studies demonstrated that TLnp/D&G showed high bone marrow accumulation and better antileukemic efficacy both in leukemia bearing models and in systemic Notch1-induced T-ALL models, with excellent biosafety and reduced gastrointestinal toxicity. Overall, our study provides new strategies for the treatment of T-ALL and promising bone marrow targeting systems with high transformation potential.

Shikonin Promotes Apoptosis and Attenuates Migration and Invasion of Human Esophageal Cancer Cells by Inhibiting Tumor Necrosis Factor Receptor-Associated Protein 1 Expression and AKT/mTOR Signaling Pathway.

The aim of this study was to investigate the anticancer effects of shikonin on esophageal cancer (EC) cells and explore the underlying molecular mechanism by identifying dysregulation in shikonin-induced tumor necrosis factor receptor-associated protein 1 (TRAP1) expression. The 3-(4, 5-dimethylthiazol-2-Yl)-2, 5-diphenyltetrazolium bromide assay and EDU assay were performed for cell viability determination. The reactive oxygen species level and mitochondrial membrane potential were evaluated using flow cytometry. The protein expression was detected using Western blot. In addition, cell migration and invasion were estimated. These results demonstrated that shikonin inhibited EC cell growth in a concentration-dependent manner and induced apoptosis through activation of the intracellular apoptotic signaling pathway. Moreover, TRAP1 downregulation promoted shikonin-induced reactive oxygen species release, whereas TRAP1 upregulation blocked it. Meanwhile, shikonin significantly promoted mitochondrial depolarization, accompanied by a large release of cytochrome C. Conversely, shikonin significantly decreased adenosine 5'-triphosphate release, demonstrating a significant intervention in the process of the glucose metabolism. In addition, not only shikonin but also short hairpin RNA (shRNA)-TRAP1 inhibited EC cell migration and invasion. shRNA-TRAP1 enhanced the inhibitory effect of shikonin on matrix metalloproteinase (MMP)2 and MMP9 expression. More interestingly, we demonstrated that shRNA-TRAP1 played a synergistic role in shikonin-mediated regulation of protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling. Collectively, shikonin promoted apoptosis and attenuated migration and invasion of EC cells by inhibiting TRAP1 expression and AKT/mTOR signaling, indicating that shikonin may be a new drug for treating EC.

Natural Potent NAAA Inhibitor Atractylodin Counteracts LPS-Induced Microglial Activation.

N-acylethanolamine-hydrolyzing acid amidase (NAAA) is a lysosomal enzyme that inhibits the degradation of palmitoylethanolamide (PEA), an endogenous lipid that induces analgesic, anti-inflammation, and anti-multiple sclerosis through PPARα activation. Only a few potent NAAA inhibitors have been reported to date, which is mainly due to the restricted substrate-binding site of NAAA. Here, we established a high-throughput fluorescence-based assay for NAAA inhibitor screening. Several new classes of NAAA inhibitors were discovered from a small library of natural products. One of these is atractylodin, a polyethylene alkyne compound from the root of Atractylodes lancea (Thunb) DC., which significantly inhibits NAAA activity and has an IC50 of 2.81 µM. Kinetic analyses and dialysis assays suggested that atractylodin engages in competitive inhibition via reversible reaction to the enzyme. Docking assays revealed that atractylodin occupies the catalytic cavity of NAAA, where the atractylodin furan head group has a hydrophobic-related interaction with the backbone of the Trp181 and Leu152 residues of human NAAA. Further investigation indicated that atractylodin significantly increases PEA and OEA levels and dose-dependently inhibits LPS-induced nitrate, TNF-α, IL-1ß, and IL-6 pro-inflammatory cytokine release in BV-2 microglia. Our results show that atractylodin elevates cellular PEA levels and inhibits microglial activation by inhibiting NAAA activity, which in turn could contribute to NAAA functional research.

Epimedium polysaccharides attenuates hematotoxicity by reducing oxidative stress and enhancing immune function in mice model of benzene-induced bone marrow failure.

Chronic benzene (BZ) exposure is associated with multiple adverse health effects and leads to progressive bone marrow failure (BMF). BZ-induced BMF is an acquired aplastic anemia characterized by severe anemia, neutropenia and thrombocytopenia, which is likely caused by immunotoxicity and oxidative stress. Previous studies showed that Epimedium polysaccharides (EPS), a natural and major herbal compound derived from Epimedium, has immunomodulatory and antioxidant potential. The purpose of this study was to evaluate the potential efficacy of EPS against BZ-induced BMF. BMF mouse model was established by subcutaneous injection of 2 ml/kg BZ in CD1 mice. Mice received daily oral treatment with 100 mg/kg high-dose EPS and 20 mg/kg low-dose EPS for four weeks. Our data showed that EPS treatment alleviated BZ-associated weight loss and increased the number of whole blood cells in peripheral blood and nucleated cells in bone marrow. Furthermore, EPS treatment decreased apoptotic rate and reactive oxygen species production, S-phase arrest in bone marrow cells. Finally, EPS treatment improved T cell-mediated immune suppression by increasing CD3+, CD4 + T-cell counts, and CD4+/CD8+ ratio. and modulated hematopoietic cytokines including EPO, IL-11, and IL-2 in peripheral blood. Our study suggests that EPS is a potential therapeutic target to attenuate hematotoxicity induced by BZ.

Cyclin G2 suppresses estrogen-mediated osteogenesis through inhibition of Wnt/ß-catenin signaling.

Estrogen plays an important role in the maintenance of bone formation, and deficiency in the production of estrogen is directly linked to postmenopausal osteoporosis. To date, the underlying mechanisms of estrogen-mediated osteogenic differentiation are not well understood. In this study, a pluripotent mesenchymal precursor cell line C2C12 was used to induce osteogenic differentiation and subjected to detection of gene expressions or to manipulation of cyclin G2 expressions. C57BL/6 mice were used to generate bilateral ovariectomized and sham-operated mice for analysis of bone mineral density and protein expression. We identified cyclin G2, an unconventional member of cyclin, is involved in osteoblast differentiation regulated by estrogen in vivo and in vitro. In addition, the data showed that ectopic expression of cyclin G2 suppressed expression of osteoblast transcription factor Runx2 and osteogenic differentiation marker genes, as well as ALP activity and in vitro extracellular matrix mineralization. Mechanistically, Wnt/ß-catenin signaling pathway is essential for cyclin G2 to inhibit osteogenic differentiation. To the best of our knowledge, the current study presents the first evidence that cyclin G2 serves as a negative regulator of both osteogenesis and Wnt/ß-catenin signaling. Most importantly, the basal and 17ß-estradiol-induced osteogenic differentiation was restored by overexpression of cyclin G2. These results taken together suggest that cyclin G2 may function as an endogenous suppressor of estrogen-induced osteogenic differentiation through inhibition of Wnt/ß-catenin signaling.