PGRN inhibits CD8+T cell recruitment and promotes breast cancer progression by up-regulating ICAM-1 on TAM.

BACKGROUND: Tumor microenvironment actually reduces antitumor effect against the immune attack by exclusion of CD8+T cells. Progranulin (PGRN) is a multifunctional growth factor with significant pathological effects in multiple tumors; however, its role in immunity evasion of breast cancer (BCa) is not completely understood. METHODS: We depleted GRN (PGRN gene) genetically in mice or specifically in PY8119 murine BCa cell line, and mouse models of orthotopic or subcutaneous transplantation were used. Chimeric mice-deficient of PGRN (Grn-/-) in bone marrow (BM) compartment was also generated. Association of PGRN expression with chemokine production or BCa development was investigated by histological and immunological assays. RESULTS: We found PGRN was involved in exhaustion of cytotoxic CD8+T cell in BCa with the increasing expressions of M2 markers and intercellular cell adhesion molecule-1 (ICAM-1) on macrophages. Specifically, ablation of PGRN in PY8119 cells reduced tumor burden, accompanied by the infiltrating of cytotoxic CD8+T cells into tumor nests. Moreover, our result revealed that blockade of PD-1 in PGRN-depleted tumors exhibited better antitumor effect in vivo and significantly decreased tumor burden. CONCLUSION: These findings suggest that inhibition of PGRN may act as a potential immune-therapeutic strategy by recovering infiltration of CD8+T cell in BCa tissue and thereby enhancing the response to anti-PD-1 therapy.

Possible Origin of Low-Frequency Magnetic Flux Noise in Superconducting Devices.

The magnetic flux noise caused by surface spin fluctuations in superconducting quantum interference devices (SQUIDs) limits their development. In this work, we report that different adsorbents such as H, O2, NO, and NO2 that adsorb on the surfaces of Mg-based and Pb-based SQUIDs, respectively, producing large local magnetic moments ranging from 0.7-1.6 µB, with energy barriers for thermal spin fluctuation as low as 10-30 mK. Moreover, we observe that the presence of H atoms on the surface of MgO can cause the coadsorption of other molecules, which generates additional spin sources. Monte Carlo simulations of the weakly coupled spin on a two-dimensional square lattice produce a low-frequency flux noise spectrum. We suggest eliminating the surface magnetism by coating the surface with monolayer indium phosphide or protecting the surface from other molecules by nonmagnetic preoccupants with a larger adsorption energy. The work provides important physical insights and feasible strategies for reducing magnetic noise sources in superconducting circuits.

[Cerebrospinal Fluid ß2-Microglobulin for Prognostic Evaluation of Patients with Central Nervous System Infiltration in Acute Lymphoblastic Leukemia].

OBJECTIVE: To investigate the prognostic value of serum and cerebrospinal fluid ß2-microglobulin (ß2-MG) in acute lymphoblastic leukemia (ALL) with central nervous system invasion after chemotherapy. METHODS: 40 patients with leukemia who had been confirmed to have central nervous system infiltration were selected for treatment at the Second Affiliated Hospital of Chongqing Medical University from January 2015 to May 2017, and the serum levels of ß2-MG and CSF-ß2MG were dynamically monitored and performed statistical analysis. RESULTS: After chemotherapy, the changes in serum ß2-MG were not statistically significant (P>0.05); the absolute level of CSF-ß2MG and the percentage of relative baseline changes were statistically different in different clinical outcome groups(P<0.05), and the decreasing CSF-ß2MG levels suggest a better prognosis, with cut-off values of 1.505 and -25%, respectively. CONCLUSION: The best cut-off point may be a predictor of complete remission; the reduction of the absolute and relative levels of CSF-ß2MG can suggest a good prognosis for patients.

Self-assembled tetrahedral framework nucleic acid mediates tumor-associated macrophage reprogramming and restores antitumor immunity.

There is increasing interest in depleting or repolarizing tumor-associated macrophages (TAMs) to generate a proinflammatory effect. However, TAMs usually display an immunosuppressive M2-like phenotype in the tumor microenvironment. Apparently, developing a macrophage-targeting delivery system with immunomodulatory agents is urgent. In this study, an efficient siRNA and CpG ODNs delivery system (CpG-siRNA-tFNA) was prepared with nucleic acid stepwise self-assembled. The tFNA composed of CpG ODNs and siRNA showed a higher stability and an enhanced cellular uptake efficiency. Moreover, the CpG-siRNA-tFNA effectively reprogrammed TAMs toward M1 phenotype polarization with increased proinflammatory cytokine secretion and NF-κB signal pathway activation, which triggers dramatic antitumor immune responses. Additionally, the CpG-siRNA-tFNA exhibited superior antitumor efficacy in a breast cancer xenograft mouse model without obvious systemic side effects. Taken together, CpG-siRNA-tFNA displayed greatly antitumor effect by facilitating TAM polarization toward M1 phenotypes in favor of immunotherapy. Hence, we have developed an efficient therapeutic strategy with immunomodulatory agents for clinical applications.

Organic-Inorganic Hybrid Cathode with Dual Energy-Storage Mechanism for Ultrahigh-Rate and Ultralong-Life Aqueous Zinc-Ion Batteries.

The exploitation of cathode materials with high capacity as well as high operating voltage is extremely important for the development of aqueous zinc-ion batteries (ZIBs). Yet, the classical high-capacity materials (e.g., vanadium-based materials) provide a low discharge voltage, while organic cathodes with high operating voltage generally suffer from a low capacity. In this work, organic (ethylenediamine)-inorganic (vanadium oxide) hybrid cathodes, that is, EDA-VO, with a dual energy-storage mechanism, are designed for ultrahigh-rate and ultralong-life ZIBs. The embedded ethylenediamine (EDA) can not only increase the layer spacing of the vanadium oxide, with improved mobility of Zn ions in the V-O layered structure, but also act as a bidentate chelating ligand participating in the storage of Zn ions. This hybrid provides a high specific capacity (382.6 mA h g-1 at 0.5 A g-1 ), elevated voltage (0.82 V) and excellent long-term cycle stability (over 10 000 cycles at 5 A g-1 ). Assistant density functional theory (DFT) calculations indicate the cathode has remarkable electronic conductivity, with an ultralow diffusion barrier of 0.78 eV for an optimal Zn-ion diffusion path in the EDA-VO. This interesting idea of building organic-inorganic hybrid cathode materials with a dual energy-storage mechanism opens a new research direction toward high-energy secondary batteries.

Characterization of Host Cell Potential Proteins Interacting with OsHV-1 Membrane Proteins.

The interaction between viral membrane associate proteins and host cellular surface molecules should facilitate the attachment and entry of OsHV-1 into host cells. Thus, blocking the putative membrane proteins ORF25 and ORF72 of OsHV-1 with antibodies that have previously been reported to subdue OsHV-1 replication in host cells, especially ORF25. In this study, prey proteins in host hemocytes were screened by pull-down assay with recombinant baits ORF25 and ORF72, respectively. Gene Ontology (GO) analysis of these prey proteins revealed that most of them were mainly associated with binding, structural molecule activity and transport activity in the molecular function category. The protein-protein interaction (PPI) network of the prey proteins was constructed by STRING and clustered via K-means. For both ORF25 and ORF72, three clusters of these prey proteins were distinguished that were mainly associated with cytoskeleton assembly, energy metabolism and nucleic acid processing. ORF25 tended to function in synergy with actins, while ORF72 functioned mainly with tubulins. The above results suggest that these two putative membrane proteins, ORF25 and ORF72, might serve a role in the transport of viral particles with the aid of a cytoskeleton inside cells.

Qualitative Histopathological Classification of Primary Bone Tumors Using Deep Learning: A Pilot Study.

BACKGROUND: Histopathological diagnosis of bone tumors is challenging for pathologists. We aim to classify bone tumors histopathologically in terms of aggressiveness using deep learning (DL) and compare performance with pathologists. METHODS: A total of 427 pathological slides of bone tumors were produced and scanned as whole slide imaging (WSI). Tumor area of WSI was annotated by pathologists and cropped into 716,838 image patches of 256 × 256 pixels for training. After six DL models were trained and validated in patch level, performance was evaluated on testing dataset for binary classification (benign vs. non-benign) and ternary classification (benign vs. intermediate vs. malignant) in patch-level and slide-level prediction. The performance of four pathologists with different experiences was compared to the best-performing models. The gradient-weighted class activation mapping was used to visualize patch's important area. RESULTS: VGG-16 and Inception V3 performed better than other models in patch-level binary and ternary classification. For slide-level prediction, VGG-16 and Inception V3 had area under curve of 0.962 and 0.971 for binary classification and Cohen's kappa score (CKS) of 0.731 and 0.802 for ternary classification. The senior pathologist had CKS of 0.685 comparable to both models (p = 0.688 and p = 0.287) while attending and junior pathologists showed lower CKS than the best model (each p < 0.05). Visualization showed that the DL model depended on pathological features to make predictions. CONCLUSION: DL can effectively classify bone tumors histopathologically in terms of aggressiveness with performance similar to senior pathologists. Our results are promising and would help expedite the future application of DL-assisted histopathological diagnosis for bone tumors.

Tailoring the Structural and Electronic Properties of Graphene through Ion Implantation.

Ion implantation is a superior post-synthesis doping technique to tailor the structural properties of materials. Via density functional theory (DFT) calculation and ab-initio molecular dynamics simulations (AIMD) based on stochastic boundary conditions, we systematically investigate the implantation of low energy elements Ga/Ge/As into graphene as well as the electronic, optoelectronic and transport properties. It is found that a single incident Ga, Ge or As atom can substitute a carbon atom of graphene lattice due to the head-on collision as their initial kinetic energies lie in the ranges of 25-26 eV/atom, 22-33 eV/atom and 19-42 eV/atom, respectively. Owing to the different chemical interactions between incident atom and graphene lattice, Ge and As atoms have a wide kinetic energy window for implantation, while Ga is not. Moreover, implantation of Ga/Ge/As into graphene opens up a concentration-dependent bandgap from ~0.1 to ~0.6 eV, enhancing the green and blue light adsorption through optical analysis. Furthermore, the carrier mobility of ion-implanted graphene is lower than pristine graphene; however, it is still almost one order of magnitude higher than silicon semiconductors. These results provide useful guidance for the fabrication of electronic and optoelectronic devices of single-atom-thick two-dimensional materials through the ion implantation technique.

[Progranulin (PGRN) promotes invasion and migration of mouse breast cancer 4T1 cells by promoting epithelial-mesenchymal transition of cancer cells and activating ERK1/2 pathway].

Objective To investigate the effect of progranulin (PGRN) on the invasion and migration of mouse breast cancer 4T1 cells and its mechanism. Methods After treated with PGRN (1 µg/mL) for 24 hours, the invasion ability of breast cancer 4T1 cells was detected by TranswellTM invasion assay, the migration ability was detected by scratch test, and the epithelial cadherin (E-cadherin), vimentin mRNA expression was detected by real-time fluorescent quantitative PCR. Western blot assay was used to detect the expression of E-cadherin, vimentin, extracellular signal-regulated kinase 1/2 (ERK1/2) and phosphorylated ERK1/2 (p-ERK1/2). After treated with 1 µg/mL PGRN and ERK1/2 signaling pathway inhibitor U0126 (10 µmol/L) simultaneously, the migration and invasion ability of 4T1 cells and the changes in the expression of E-cadherin, vimentin and p-ERK proteins were detected again. Results After treated with PGRN, the migration and invasion capabilities of breast cancer 4T1 cells were significantly enhanced; E-cadherin expression decreased; vimentin and p-ERK1/2 expression increased. After treated with ERK1/2 signaling pathway inhibitor, the ability of PGRN to promote breast cancer 4T1 cell migration, invasion and epithelial-mesenchymal transition (EMT) was significantly inhibited. Conclusion PGRN can promote the migration and invasion of breast cancer 4T1 cells by promoting EMT and activating the ERK1/2 pathway.

Progranulin induces immune escape in breast cancer via up-regulating PD-L1 expression on tumor-associated macrophages (TAMs) and promoting CD8+ T cell exclusion.

BACKGROUND: Progranulin (PGRN), as a multifunctional growth factor, is overexpressed in multiple tumors, but the role of PGRN on tumor immunity is still unclear. Here, we studied the effect of PGRN on breast cancer tumor immunity and its possible molecular mechanism. METHODS: The changes of macrophage phenotypes after PGRN treatment were detected by western blot, quantitative polymerase chain reaction (PCR) and flow cytometry. Western blot was used to study the signal molecular mechanism of PGRN regulating this process. The number and localization of immune cells in Wild-type (WT) and PGRN-/- breast cancer tissues were analyzed by immunohistochemical staining and immunofluorescence techniques. The activation and proliferation of CD8+ T cells were measured by flow cytometry. RESULTS: After being treated with PGRN, the expressions of M2 markers and programmed death ligand 1 (PD-L1) on macrophages increased significantly. Signal transducer and activator of transcription 3 (STAT3) signaling pathway inhibitor Stattic significantly inhibited the expression of PD-L1 and M2 related markers induced by PGRN. In WT group, CD8 were co-localized with macrophages and PD-L1, but not tumor cells. The number of immune cells in PGRN-/- breast cancer tissue increased, and their infiltration into tumor parenchyma was also enhanced. Moreover, in the co-culture system, WT peritoneal macrophages not only reduced the ratio of activated CD8+ T cells but also reduced the proportion of proliferating CD8+ T cells. The addition of programmed death receptor 1 (PD-1) and PD-L1 neutralizing antibodies effectively reversed this effect and restored the immune function of CD8+ T cells. CONCLUSION: These results demonstrate that PGRN promotes M2 polarization and PD-L1 expression by activating the STAT3 signaling pathway. Furthermore, through PD-1/PD-L1 interaction, PGRN can promote the breast tumor immune escape. Our research may provide new ideas and targets for clinical breast cancer immunotherapy.

PGRN-/- TAMs-derived exosomes inhibit breast cancer cell invasion and migration and its mechanism exploration.

Breast cancer is one of the most malignant diseases world-wide and ranks the first among female cancers. Progranulin (PGRN) plays a carcinogenic role in breast cancer, but its mechanisms are not clear. In addition, there are few reports on the relationship between PGRN and tumor-associated macrophages (TAMs). AIMS: To investigate the effects of exosomes derived from PGRN-/- TAMs on invasion and migration of breast cancer cells. MAIN METHODS: Mouse breast cancer xenograft model was constructed to explore the effect of PGRN-/- tumor environment (TME) on breast cancer. Flow cytometry was used to compare TAMs of wild type (WT) and PGRN-/- tumor tissue. Transwell assay, wound healing assay and western blot were used to explore the effect of WT and PGRN-/- TAMs and their exosomes on invasion, migration and epithelial-mesenchymal transition (EMT) of breast cancer cells. MicroRNA (miRNA) assay was used to find out the differentially expressed miRNA of negative control (NC) and siPGRN-TAMs exosomes. Quantitative PCR and luciferase report assay were used to explore the target gene. KEY FINDINGS: The lung metastasis of breast cancer of PGRN-/- mice was inhibited. PGRN-/- TAMs inhibited invasion, migration and EMT of breast cancer cells through their exosomes. MiR-5100 of PGRN-/- TAMs-derived exosomes was up-regulated, which might regulate expression of CXCL12, thereby inhibiting the CXCL12/CXCR4 axis, and ultimately inhibiting the invasion, migration and EMT of breast cancer cells. SIGNIFICANCE: Our study elucidates a new molecular mechanism of lung metastasis of breast cancer, so it may contribute to efficient prevention and therapeutic strategies.

Nucleus-located PDK1 regulates growth, invasion and migration of breast cancer cells.

AIMS: It is well known that pyruvate dehydrogenase kinase 1 (PDK1) is highly expressed in breast cancer (BC) tissues and promotes tumor growth, but the underlying mechanisms of this process are unclear. Here, we investigated the effects of nuclear PDK1 on growth, migration and invasion in human BC cells. MAIN METHODS: The sub-cellular localization of PDK1 in BC cells was performed with subcellular fractionation followed by Western blot and immunofluorescence. The localization of PDK1 in breast normal tissue and breast duct carcinoma was detected by Immunohistochemistry. Then the protein-protein interaction between PDK1 and Importin ß was verified by co-immunoprecipitation assay. Finally, the effects of nuclear PDK1 on cell proliferation, apoptosis, migration and invasion of BC cells were assessed. KEY FINDINGS: In addition to its well-known sub-cellular localization, PDK1 was present in the nucleus of BC cells, and EGF treatment increased nucleus distribution of PDK1. Moreover, the level of nuclear PDK1 accumulation facilitated the growth of BC cells. We also found that the entry of PDK1 into nucleus mainly relied on the nuclear localization signal (NLS), and NLS mutation inhibited the entry of PDK1 into nucleus; as a result, the migration and invasion abilities of BC cells were impaired, and the number of apoptotic cells was significantly increased. SIGNIFICANCE: Our findings provided a new supplement to the sub-cellular localization of PDK1 in BC cells and uncovered the function of nuclear PDK1 in facilitating BC cells growth, migration and invasion.

Honokiol inhibits the growth of SKBR3 cells.

BACKGROUND: Breast cancer is one of the most malignant tumors in the reproductive system and has a poor prognosis. Finding drugs with high efficiency, low side-effects, and low cost has become a research hotspot. METHODS: In the present study, we treated SK-BR-3 cells with different doses of honokiol. Crystal violet staining method was used to detect changes in the total number of living cells; 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was used to detect the effect of honokiol on SK-BR-3 cell proliferation. Cell migration ability change was determined by wound healing assay. Cell invasion ability change was determined by Transwell migration assay. Flow cytometry was used to detect the apoptotic rate of SK-BR-3 cells, and Western blot was used to detect the expression levels of proliferation-associated protein (PCNA); migration- and invasion-related protein matrix metalloproteinase-2 (MMP-2); vimentin; apoptosis-related proteins Bcl-xl, caspase 3, and cleaved caspase 3 (CC3); and ß-catenin and its downstream target molecule c-Myc. RESULTS: Compared with the control group, different doses of honokiol have different degrees of inhibitory effects on cells, including proliferation and invasion and migration (P<0.01). After treatment with 50 or 60 µmol·L-1 honokiol, the apoptotic rate of SK-BR-3 cells increased (both P<0.01); PCNA expression was significantly downregulated (P<0.01). Intracellular accumulation of apoptosis-related proteins Bcl-xl and caspase-3 decreased but C-C3 increased. We also found downregulation of MMP-2 expression, a protein related to invasion and migration (P<0.01), and a decrease in the expression levels of the Wnt/ß-catenin signaling pathway-related proteins ß-catenin and c-Myc (P<0.01). CONCLUSIONS: Honokiol can promote the apoptosis of SK-BR-3 cells and can inhibit the proliferation, migration, and invasion of human breast cancer SK-BR-3 cells. The underlying mechanism may be through inhibiting the activation of the Wnt signaling pathway.

[Molecular mechanism of macrophages derived from PGRN gene knockout mice inhibit invasion and migration of breast cancer cells].

Objective To explore the functions and mechanisms of macrophages derived from PGRN gene knockout (PGRN-/- ) C57BL/6 mice in the invasion and migration of breast cancer cells. Methods Breast cancer cells were cultured in conditioned medium of macrophages derived from WT and PGRN-/- mice. TranswellTM assay and scratch assay were used to detect the invasion and migration ability of cancer cells. Western blot analysis was used to detect the expression of E-cadherin and N-cadherin in cancer cells. Cytokine array, real-time quantitative PCR and ELISA were performed to investigate the differences of cytokines secreted by macrophages derived from WT and PGRN-/- mice. Breast cancer cells were treated by the differentially expressed cytokine interleukin-6 (IL-6), and then the above methods were used to investigate its effect on cancer cells. Western blot analysis was used to verify the roles of NF-κB and JAK/STAT3 signaling pathways. Results The macrophages derived from PGRN-/- mice blocked NF-κB signaling pathway, reduced IL-6 secretion, and inhibited the invasion and migration of breast cancer cells. IL-6 activated JAK/STAT3 signaling pathway to promote the invasion and migration of breast cancer cells. Conclusion The macrophages derived from PGRN-/- mice can block the NF-κB and JAK/STAT3 signaling pathways, down-regulate IL-6 expression, and inhibit the invasion and migration of breast cancer cells.