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Aims Neurodegenerative diseases are the leading cause of morbidity and mortality worldwide. Several studies have shown that tetramethylpyrazine (TMP) is an effective therapy for neurodegenerative diseases and that it acts by inhibiting the activation of microglial cells in response to inflammatory stimuli. However, the molecular mechanisms underlying the action of TMP remain unknown. Main methods Proteomic analysis was used to generate novel insights into the mechanism by which TMP inhibits microglial activation, and western blotting was used to validate candidate proteins. Key findings To identify candidate proteins affected by TMP in lipopolysaccharide-activated microglia, we performed proteomic analysis using iTRAQ labelling coupled with LC TRIPLE-TOF, and we identified 5187 unique proteins. Among these, 266 proteins were differentially expressed and considered putative candidate proteins. Protein annotation revealed that the differentially expressed proteins, such as inducible nitric oxide synthase (iNOS) and ERO1-like protein (ERO1L), might be involved in reducing cellular oxidation in response to stress. Ingenuity pathway analysis revealed that the differentially expressed proteins were involved in a variety of signalling pathways, including liver X receptor/retinoid X receptor (LXR/RXR) activation and the production of nitric oxide and reactive oxygen species in macrophages. Furthermore, one of the differentially expressed protein candidates detected by iTRAQ, iNOS, was confirmed by western blotting. Significance Our data suggest that iTRAQ technology is an effective tool to study the mechanism by which TMP inhibits activated microglia. TMP decreased the expression of LXR/RXR-mediated iNOS, which reduced microglial activation in response to inflammatory stimuli. [ABSTRACT FROM AUTHOR]

Two‐photon excited fluorescence is a process in which the excited fluorophore relaxes and emits fluorescence after two‐photon absorption. Combined with appropriate two‐photon fluorescent probes, two‐photon microscopy enables observation deep inside tissue without damage. Former reports showed that the two‐photon absorption cross sections of donor–acceptor (D–A) dipoles, as well as D‐π‐D and D–A–D quadrupoles can be strengthened by using strong D–A groups. In this study, a novel water‐soluble fluorescent probe with D‐π‐A structure containing pyridine‐2,6‐dicarboxylic acid unit was synthesized. The maximum excitation wavelength and emission wavelength of the probe in aqueous solution were 328 nm and 471 nm, respectively. The fluorescence emitted by the probe can be quenched by various metal cations. The pyridyl N and carboxyl O in the probe formed chelates with Cu2+, which resulted in a fluorescence quenching response, and showed a linear relationship between fluorescent intensity and Cu2+ concentration with wide linear range and low limit of detection. CCK‐8 tests showed the probe had no obvious toxicity to 4T1 cells in the range of 0.4–50 μmol L−1, and the cell survival rate was above 86 %. The probe had a large two‐photon absorption cross section ranging from 680 nm to 820 nm. The two‐photon absorption cross section reached 100 GM at 760 nm of excitation wavelength. By means of microscopic imaging of 4T1 cells stained with the probe, one‐photon fluorescence represented blue emission, while two‐photon fluorescence represented bright green emission. The probe was proved to have good cell permeability, and preferred to target nucleus, showing application potential in two‐photon fluorescence imaging technology. [ABSTRACT FROM AUTHOR]

Successful mouse embryo implantation requires a receptive uterus and an activated blastocyst. A large number of genes, cytokines, and other factors are involved in the process. MicroRNAs (miRNAs) regulate the expression of many genes, and previous studies have investigated the relationship between miRNA expression and embryo implantation. In this study, we show that mmu-microRNA-200a (mmu-miR-200a) is expressed in a spatiatemporal manner during implantation in mouse uterus and found that phosphatase and tensin homolog (PTEN), SON, and programmed cell death 4 (Pdcd4) are the target genes of mmu-miR-200a by bioinformatics analysis. In vitro gain and loss of function experiments confirm that PTEN, a critical gene for cell proliferation and apoptosis, is the target gene of mmu-miR-200a. Our experiments also show that injection of the uterine horn with mmu-miR-200a lentivirus leads to a decreased implantation rate. Collectively, our results suggest that mmu-miR-200a affects embryo implantation by regulating PTEN protein expression. Thus, clarifying the physiological functions of uterine miRNAs will help to elucidate the embryo implantation process and may even contribute to curing infertility and inventing new contraceptives. [ABSTRACT FROM PUBLISHER]

Abstract: This study aimed to investigate the protective effect of chitosan oligosaccharides (COS) on human umbilical vein endothelial cell (HUVEC) apoptosis induced by hydrogen peroxide (H2O2). We found that COS not only reversed the decrease of cell viability and proliferation activity, but ameliorated nuclear chromatin damage in H2O2-induced HUVECs. Additionally, COS contributed to the decrease of cytosolic Ca2+ level, increase of mitochondrial membrane potential, up-regulation of Bcl-2 mRNA, down-regulation of Bax mRNA, reduction of cleaved caspase-3 protein, inhibition of phosphorylated p38 mitogen-activated protein kinase (MAPK) and induction of phosphorylated Akt in HUVECs. Further, H2O2-induced caspase-3 activation could be suppressed by p38 MAPK inhibitor (SB203580) and up-regulated by phosphatidylinositol-3-kinase (PI3K) inhibitor (LY294002), indicating the involvement of p38 MAPK and PI3K/Akt signaling pathways. In conclusion, the results show that COS may effectively inhibit the H2O2-induced HUVEC apoptosis. [Copyright &y& Elsevier]

Abstract: Aim of the study: To determine the anti-inflammatory effects of Tetramethylpyrazine (TMP) and to investigate the inhibitory effect of TMP on IL-8 production in human umbilical vein endothelial cells (HUVECs) induced by LPS might be mediated by inhibiting p38, ERK and NF-κB signaling pathways. Materials and methods: HUVECs were treated with or without TMP for 24h before exposure to LPS for 4h. IL-8 gene and protein expressions were determined by RT-PCR and ELISA. Cell viability was determined by methyl thiazoyltetrazolium (MTT) assay. Phosphorylation of ERK1/2 and p38 were examined by western blotting. Results: TMP inhibits LPS-induced IL-8 production in HUVECs at both the protein and mRNA levels, suggesting that TMP has an antiinflammatory effect on endothelial cells. TMP also inhibited U937 monocyte adhesion to HUVECs stimulated by LPS. LPS-induced phosphorylation of ERK1/2 and p38 were inhibited by TMP. The inhibitory effect of TMP on NF-κB (p65) activity was mediated by blocking the consequent translocation of p65 into the nucleus. Conclusions: The inhibitory effect of TMP on the LPS–induced IL-8 production is mediated by the NF-κB–dependent pathway, and TMP also separately affects the ERK and p38 MAPK pathway. TMP may be beneficial in the treatment of cardiovascular disorders such as atherosclerosis. [Copyright &y& Elsevier]

Evidence recently showed that pleiotropic cytokine interferon-gamma (IFN-γ) in the tumor microenvironment (TME) plays a positive role in hepatocellular carcinoma (HCC) progression through the regulation of liver cancer stem cells (LCSCs) in HCC. The present study explored the role and potential mechanism of mitochondrial programmed cell death-ligand 1 (PD-L1) and its regulation of ferroptosis in modulating the cancer stemness of LCSCs. It was shown that mimicking TME IFN-γ exposure increased the LCSCs ratio and cancer stemness phenotypes in HCC cells. IFN-γ exposure inhibited sorafenib (Sora)-induced ferroptosis by enhancing glutathione peroxidase 4 (GPX4) expression as well reactive oxygen species (ROS) and lipid peroxidation (LPO) generation in LCSCs. Furthermore, IFN-γ exposure upregulated PD-L1 expression and its mitochondrial translocation, inducing dynamin-related protein 1 (Drp1)-dependent mitochondrial fission and correlating with glycolytic metabolism reprogramming in LCSCs. The genetic intervention of PD-L1 promoted ferroptosis-dependent anti-tumor effects of Sora, reduced glycolytic metabolism reprogramming, and inhibited cancer stemness of HCC in vitro and in vivo. Our results revealed a novel mechanism that IFN-γ exposure-induced mitochondrial translocation of PD-L1 enhanced glycolytic reprogramming to mediate the GPX4-dependent ferroptosis resistance and cancer stemness in LCSCs. This study provided new insights into the role of mitochondrial PD-L1-Drp1-GPX4 signal axis in regulating IFN-γ exposure-associated cancer stemness in LCSCs and verified that PD-L1-targeted intervention in combination with Sora might achieve promising synergistic anti-HCC effects. Interventing mitochondrial PD-L1 suppressed IFN-γ-induced cancer stemness in hepatocellular carcinoma by sensitizing sorafenib-induced ferroptosis (Created with BioRender.com). [Display omitted] • IFN-γ promoted ferroptosis resistance of LCSCs to sorafenib via mediating cancer stemness. • IFN-γ exposure upregulated PD-L1 and its mitochondrial translocation to reprogram glycolytic metabolism. • Mitochondrial PD-L1-Drp1 signal axis modulated GPX4-dependent ferroptosis in sorafenib-treated LCSCs. • Targeting intervention of PD-L1 promoted anti-HCC effects via enhancing ferroptosis susceptibility to sorafenib. [ABSTRACT FROM AUTHOR]

Embryo implantation is an essential and complex process in mammalian reproduction. However, little evidence has indicated the involvement of autophagy during embryo implantation. To determine the possible role of autophagy in uterine of pregnant mice during the peri-implantation stage, we first examined the expression of autophagy-related markers ATG5 and LC3 on day 4, 5, and 6 of pregnancy (D4, D5, and D6, respectively). Compared with expression on D4, downregulation of the autophagy-related markers was observed on D5 and D6, the days after the embryo attached to the receptivity endometrium. Further examination showed that autophagy-related markers ATG5, ATG12, LC3, cathepsin B, and P62 at the implantation site were significantly decreased when comparing with the inter-implantation site. Fewer number of autophagosomes at the implantation site were also observed by transmission electron microscopy. To confirm the functional role of autophagy during embryo implantation in mice, we administered the autophagy inhibitor 3-methyladenine and chloroquine to mice. After treated with 3-methyladenine, the expression of decidual markers HOXA10 and progesterone receptor were significantly reduced. Furthermore, a reduction in implantation sites and increase in the HOXA10 and PR protein levels were observed in response to chloroquine treatment. In addition, impaired uterine decidualization and dysregulation of the PR and HOXA10 protein levels was observed after autophagy inhibited by 3-methyladenine and chloroquine in in vivo artificial decidualization mouse model. In the last, LC3 and P62 were also observed in normal human proliferative, secretory, and decidua tissues. In conclusion, endometrial autophagy may be essential for embryo implantation, and it may be associated with endometrial decidualization during early pregnancy. Key message: • Autophagy-related markers were significantly decreased at implantation site. • Autophagy inhibition results in abnormal decidualization. • Autophagy is essential for embryo implantation. [ABSTRACT FROM AUTHOR]

Three aryl-tetralin-type lignans, including 2 previously undescribed compounds, were isolated from the root of Sanguisorba officinalis. The structures of the new compounds were elucidated by 1D- and 2D-NMR spectroscopic analyses and mass spectrometry. Experimental and calculated ECD were used to determine the absolute configurations. The isolated compounds were evaluated for cytotoxicity against two cell lines (MV4-11 and MDA-MB-231) and compound 1 exhibited moderate growth inhibition against MDA-MB-231 cell line with IC50 value of 15.76 μM. [ABSTRACT FROM AUTHOR]

Embryo implantation is essential for normal pregnancy, and the process of decidualization is critical for embryo implantation. However, the mechanism of decidualization during early pregnancy is still unknown. Forkhead box O3a (FOXO3a) is the most important functional transcription factor of the forkhead box family and is a highly conserved transcription factor of apoptosis‐related genes. In the mouse uterus, FOXO3a was found to be expressed regularly from Days 1–7 of early pregnancy. Upon further exploration, it was found that FOXO3a was expressed at significantly higher levels at the implantation site than at the interimplantation site on Days 5–7 of pregnancy. Under artificial decidualization, FOXO3a was highly expressed in the first and second decidual zones. After decidualization, the expression of FOXO3a was significantly increased both in vivo and vitro. In primary stromal cells, apoptosis was reduced by decreased expression of FOXO3a after inducing decidualization. Moreover, when FOXO3a‐small interfering RNA was transfected into the uteri of mice, the expression of decidualization‐ and apoptosis‐related factors was impaired. Thus, FOXO3a might play an important role in decidualization during early pregnancy, and cell apoptosis might be one of pathways for FOXO3a‐regulated decidualization. Embryo implantation is essential for normal pregnancy, and the process of decidualization is critical for embryo implantation. The expression of forkhead box O3a (FOXO3a) was significantly increased when decidualization both in vivo and vitro. FOXO3a might play an important role in decidualization during early pregnancy. [ABSTRACT FROM AUTHOR]

Highlights: [•] Benzo(a)pyrene affects the behavior of rats in Morris water maze. [•] Benzo(a)pyrene affects rat hippocampus gene expression on signal transduction. [•] Benzo(a)pyrene alters protein expression involved in cell death. [•] Benzo(a)pyrene down-regulates major vault protein. [ABSTRACT FROM AUTHOR]

Aim of the study: To determine the inhibitory effect of tetramethylpyrazine (TMP) on lipopolysaccharide (LPS)-induced over-production of nitric oxide (NO) and inducible nitric oxide synthase (iNOS) in N9 microglial cells. Materials and methods: N9 cells were pretreated with vehicle or TMP and then exposed to LPS for the time indicated. Cell viability was determined by methylthiazoyltetrazolium (MTT) assay. Nitrite assay was performed by Griess reaction. Expression of iNOS mRNA was examined by RT-PCR. Protein levels of iNOS, p38 mitogen-activated protein kinase (MAPK), ERK1/2, JNK, phosphatidylinositol 3-kinase (PI3K) and Akt were determined by western blot analysis. Formation of reactive oxygen species (ROS) was evaluated by fluorescence image system. Results: TMP inhibited LPS-induced over-production of NO and iNOS in N9 cells. TMP also inhibited the NF-κB translocation from cytoplasm into nucleus of N9 cells. In addition, TMP showed blocking effect on the phosphorylation of p38 MAPK, ERK1/2, JNK and Akt, but not PI3K. Further, TMP suppressed the formation of intracellular ROS in LPS-induced N9 cells. Conclusions: TMP inhibited production of NO and iNOS in LPS-induced N9 cells through blocking MAPK and PI3K/Akt activation and suppressing ROS production. [Copyright &y& Elsevier]

Erlotinib has potential therapeutic effect on acute myeloid leukemia (AML) in patients, but the mechanism is not clear. Effective tumor biomarkers for erlotinib in the treatment of AML remain poorly defined. Here, we demonstrate that erlotinib in vitro significantly inhibits the growth of the FLT3‐ITD mutant AML cell MV4‐11 and Ba/F3‐FLT3‐ITD cell via targeting FLT3, a certified valid target for the effective treatment of AML. In vivo, oral administration of erlotinib at 100 mg/kg/day induced rapid MV4‐11 tumor regression and significantly prolonged the survival time of bone marrow engraftment AML mice via inhibiting the FLT3 signal. Thus, the therapeutic benefits of erlotinib on AML are due to its ability to target FLT3. FLT3‐ITD mutation is an effective biomarker for erlotinib during AML treatment. In addition, we also demonstrate that erlotinib inhibits the activity of AML cell KG‐1 (no FLT3 expression) by targeting Lyn. Recently, single cell analysis demonstrated that intratumoral heterogeneity are one of the contributors in the relapse and FLT3 inhibitor resistance. Erlotinib could effectively inhibit the MV4‐11 cells via targeting FLT3, and inhibit KG‐1 cells via targeting Lyn. Therefore, Erlotinib also has the potential to overcome intratumoral heterogeneity via targeting FLT3 and Lyn. [ABSTRACT FROM AUTHOR]

• Seven compounds including 3 previously undescribed compounds were obtained from Valeriana jatamansi. • Compounds 5 – 7 exhibited selective cytotoxicity against HCT116 cells, with IC 50 values of 9.3, 1.7 and 2.2 μM, respectively. • The cytotoxicity effect of 6 was attributed to Akt/mTOR activation blockade via inhibition of PDK1 phosphorylation. • Compound 6 Induces autophagosome formation involving PDK1/Akt/mTOR axis in human colorectal carcinoma HCT116 cells. Chlorovaltrates U-W (1 – 3), three previously undescribed iridoids, together with four known analogues were isolated from the roots of Valeriana jatamansi. Their structures were elucidated by means of spectroscopic analyses (HRESIMS, NMR). The cytotoxicity of all isolates was evaluated. Compounds 5 – 7 exhibited selective cytotoxicity against HCT116 cells, with IC 50 values of 9.3, 1.7 and 2.2 μM, respectively. The preliminary mechanistic study revealed that, the cytotoxicity effect of 6 was attributed to Akt/mTOR activation blockade via inhibition of PDK1 phosphorylation. Meanwhile, compound 6 could induce autophagosome formation in HCT116 cells via suppressing its downstream Akt/mTOR. These findings show that compound 6 could be of great importance to the development of anti-colon cancer agents. [ABSTRACT FROM AUTHOR]

Cover: The cover image, by Long et al., is based on the Original Research Article, FOXO3a is essential for murine endometrial decidualization through cell apoptosis during early pregnancy, DOI: 10.1002/jcp.27167. [ABSTRACT FROM AUTHOR]

The molecular mechanisms underlying endometrial stromal cell proliferation and differentiation (decidualization) are still not fully understood. This study revealed that increased Slp-2 expression is a significant factor modulating endometrial stromal cell proliferation and decidualization in both mice and humans. Our results showed a significant difference in the mRNA and protein levels between the implantation site and inter-implantation site on day 5 and day 6 of pregnancy in mice (all P < 0.05). Strong Slp-2 immunostaining was mainly localized within the decidual zone of mice through the post-implantation period. Mice with artificially induced deciduoma showed significantly higher expression of Slp-2 compared with uninduced controls ( P < 0.005). Human stromal cells in the middle and late-secretory phases demonstrated significantly (all P < 0.05) upregulated SLP-2, compared with cells in the proliferative phase and early secretory phases. Further analyses of the SLP-2 gene knocked down revealed a significant ( P < 0.005) repression of both the decidualization marker gene's expression (decidual/trophoblast prolactin-related protein in mice, insulin-like growth factor binding protein and prolactin in human) and the cell proliferation in in vitro-induced decidualized primary endometrial stromal cells in mice and humans. [ABSTRACT FROM AUTHOR]

Beta-cypermethrin (β-CYP) is a highly effective broad-spectrum insecticide that can potentially affect female reproduction. However, little is known about the effect of β-CYP on uterine decidualisation, which is a vital process by which the uterus provides a suitable microenvironment for pregnancy maintenance. Therefore, we focused on the effect and mechanism of β-CYP on endometrial decidualisation during early pregnancy in mice. The results indicated that the expression levels of HOXA10, BMP2, and IGFBP1 was significantly downregulated in the decidual tissue and primary endometrial stromal cells of pregnant and pseudopregnant mice following β-CYP treatment. Serum E 2 concentration was significantly increased, whereas P 4 concentration and oestrogen receptor (ERα) and progesterone receptor (PRA) expression were significantly downregulated following β-CYP exposure. The number of polyploid decidual cells was lower in the β-CYP-treated group. Furthermore, β-CYP significantly downregulated the protein expression levels of CDK4 and CDK6, and the mRNA expression levels of cyclin D3 and p21. The number of foetuses per female in the first litter was markedly reduced following exposure to β-CYP. In summary, early pregnancy exposure to β-CYP may result in defective endometrial decidualisation via compromised proliferation of uterine stromal cells and reduced expressions of cyclin D3, CDK4/6, and p21 in mice. [Display omitted] • β-CYP administration inhibited the proliferation of uterine stromal cells in mice. • β-CYP administration lowered the number of polyploid decidual cells. • β-CYP administration downregulated the expressions of cyclin D3, CDK4/6, and p21. • β-CYP administration resulted in defective endometrial decidualisation. • β-CYP administration resulted in abnormal fertility in pregnant mice. [ABSTRACT FROM AUTHOR]

The tri-axial creep tests of frozen sand with different dry densities and grain size distributions were performed at 0.1 MPa confining pressure at different temperatures. The results show that the rheological properties of frozen sand are significantly affected by the temperature, the creep deformation of the frozen sand is obvious even under a small load when the temperature is higher than - 1.5 °C. Firstly, based on the Nishihara model, the elastic modulus of viscoelastic elements is modified to be a function of stress, the coefficient of viscosity in viscoelastic part is modified considering the effect of time and stress, and the viscoplastic element is improved by introducing the damage variable. Then a new non-linear creep constitutive model for frozen sand is proposed, and the new model is verified based on test results. It can be found that the predicted results agree well with the corresponding experimental results from low stress to high stress, and the proposed model can describe the whole creep process of frozen sand with different dry densities and grain size distributions accurately under different shear stress levels and temperatures. Besides, the function relationship between the model parameters and the shear stress under different temperatures is obtained through the regression analysis. Finally, the influences of the temperature and shear stress on the parameters are analyzed as well. This study is important to further study the stability of permafrost subgrade, and can accumulate experimental data for the study of creep theory. • The tri-axial creep tests of frozen sand with different dry densities and grain size distributions were performed at 0.1 MPa confining pressure at different temperatures. • The effect of particle content ranging from 0.075 to 1 mm on creep properties of sand has been discussed in this paper. • A new non-linear creep constitutive model for frozen sand was proposed and verified based on the test results. [ABSTRACT FROM AUTHOR]