Role of CD61+ low-density neutrophils in promoting hepatocellular carcinoma metastasis through CCDC25 upregulation.

BACKGROUND: A subset of neutrophils isolated from the peripheral blood mononuclear cells (PBMC) layer has recently been described in cancer patients. METHODS: Double-gradient centrifugation was used to separate the neutrophil subsets. Western blotting and immunohistochemical assays were performed to assess CCDC25 expression levels. RESULTS: In this study, we found that low-density neutrophils (LDNs) were more highly enriched in metastatic hepatocellular carcinoma (HCC) patients than in non-metastatic HCC patients. We then showed a CD61+ LDNs subset, which displayed distinct functions and gene expression, when compared with high-density neutrophils (HDNs) and CD61- LDNs. Transcriptomic analysis revealed that the CD61+ LDNs were predominantly enhanced in the transcription of glycolysis and angiogenesis associated gene, HMGB1 associated gene and granulation protein gene. These CD61+ LDNs displayed a prominent ability to trigger metastasis, compared with HDNs and CD61- LDNs. Specifically, CD61+ LDN-derived HMGB1 protein increased the invasion of HCC cells by upregulating CCDC25. Mechanistically, the CD61+ LDN-derived HMGB1 protein enhanced the invasiveness of HCC cells and triggered their metastatic potential, which was mediated by TLR9-NF-κB-CCDC25 signaling. Blocking this signaling pathway reversed the invasion of the CD61+ LDN-induced HCC cells. In vivo, we consistently showed that CD61+ LDN-derived HMGB1 enhances HCC metastasis to the lungs. CONCLUSIONS: Overall, our findings showed that a subset of CD61+ LDNs has pro-metastatic effects on HCC, and may be used to target HCC in the clinical setting.

Ligand recognition and G protein coupling of the human itch receptor MRGPRX1.

MRGPRX1, a Mas-related GPCR (MRGPR), is a key receptor for itch perception and targeting MRGPRX1 may have potential to treat both chronic itch and pain. Here we report cryo-EM structures of the MRGPRX1-Gi1 and MRGPRX1-Gq trimers in complex with two peptide ligands, BAM8-22 and CNF-Tx2. These structures reveal a shallow orthosteric pocket and its conformational plasticity for sensing multiple different peptidic itch allergens. Distinct from MRGPRX2, MRGPRX1 contains a unique pocket feature at the extracellular ends of TM3 and TM4 to accommodate the peptide C-terminal "RF/RY" motif, which could serve as key mechanisms for peptidic allergen recognition. Below the ligand binding pocket, the G6.48XP6.50F6.51G6.52X(2)F/W6.55 motif is essential for the inward tilting of the upper end of TM6 to induce receptor activation. Moreover, structural features inside the ligand pocket and on the cytoplasmic side of MRGPRX1 are identified as key elements for both Gi and Gq signaling. Collectively, our studies provide structural insights into understanding itch sensation, MRGPRX1 activation, and downstream G protein signaling.

CD58 acts as a tumor promotor in hepatocellular carcinoma via activating the AKT/GSK-3ß/ß-catenin pathway.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most prevalent malignancies worldwide because of rapid progression and high incidence of metastasis or recurrence. Accumulating evidence shows that CD58-expressing tumor cell is implicated in development of various cancers. The present study aimed to reveal the functional significance of CD58 in HCC progression and the underlying mechanisms. METHODS: Immunohistochemical staining (IHC), and western blotting were used to detect the expression of CD58 in HCC tissues and cells. The levels of sCD58 (a soluble form of CD58) in the cell supernatants and serum were assessed by ELISA. CCK-8, colony formation, and xenograft assays were used to detect the function of CD58 on proliferation in vitro and in vivo. Transwell assay and sphere formation assay were performed to evaluate the effect of CD58 and sCD58 on metastasis and self-renewal ability of HCC cells. Western blotting, immunofluorescence (IF), TOP/FOP Flash reporter assay, and subcellular fractionation assay were conducted to investigate the molecular regulation between CD58/sCD58 and AKT/GSK-3ß/ß-catenin axis in HCC cells. RESULTS: CD58 was significantly upregulated in HCC tissues. Elevation of CD58 expression correlated with more satellite foci and vascular invasion, and poorer tumor-free and overall survival in HCC patients. Higher sCD58 levels were in HCC patients' serum compared to healthy individuals. Functionally, CD58 promotes the proliferation of HCC cells in vitro and in vivo. Meanwhile, CD58 and sCD58 induce metastasis, self-renewal and pluripotency in HCC cells in vitro. Mechanistically, CD58 activates the AKT/GSK-3ß/ß-catenin signaling pathway by increasing phosphorylation of AKT or GSK3ß signaling, promoting expression of Wnt/ß-catenin target proteins and TCF/LEF-mediated transcriptional activity. Furthermore, AKT activator SC-79 or inhibitor LY294002 abolished the inhibitory effect of CD58 silencing on the proliferation, metastasis, and stemness of HCC cells. CONCLUSIONS: Taken together, CD58 promotes HCC progression and metastasis via activating the AKT/GSK-3ß/ß-catenin pathway, suggesting that CD58 is a novel prognostic biomarker and therapeutic target for HCC.

Tumor-associated NK cells facilitate tumor growth via NKp46 in immunocompetent murine hepatocellular carcinoma.

Natural killer(NK) cells comprise one subset of the innate lymphoid cells family. Despite reported anti-tumor activity of NK cells, their tangible contribution to tumor control remains controversial. This is due to the incomplete understanding of NK alterations within tumor microenvironment(TME). Here we showed, using murine hepatocellular carcinoma(HCC) model, that early NK cells deletion markedly attenuated tumor growth in a CD8+T cells dependent manner. This effect was accompanied by an enhanced CD8+T cells effector function in tumor rather than circulating blood. Then, we demonstrated that abundant NKp46+ NK subset, but not NKp46- NK, were recruited towards tumor microenvironment during tumor progression. Frequency of intratumor NKP46+ NK cells were inversely related to CD8+T cells activation, and positively correlated with tumor growth. Intratumor NKp46+ NK cells exhibited dysfunction and increased expression of inhibitory receptors, when compared with NKp46- NK cells. Blockade of NK cells-associated NKp46 effectively attenuated HCC growth. Infusion of tumor-derived NKp46+ NK cells markedly enhanced HCC growth in vivo, in contrast to tumor cells inoculation alone. The further mechanistic investigations unveiled that NK cells boosted tumor growth by NKp46-mediated impairment of CD8+T cells effector function. Overall, this work supported a previously unappreciated regulatory property of tumor-associated NK cells in HCC, and NKp46 as a potential target against HCC in clinical setting.

Lenvatinib inhibited HCC cell migration and invasion through regulating the transcription and ubiquitination of UHRF1 and DNMT1.

Hepatocellular carcinoma (HCC) is one of the most common causes of malignancy-related deaths. Lenvatinib, as a multi-targeted tyrosine kinase inhibitor, has gained increasing attention for its antitumor activity. However, the effect and mechanisms of Lenvatinib on HCC metastasis are virtually unknown. In this study, we revealed that Lenvatinib inhibited HCC cell motility and epithelial mesenchymal transition (EMT), along with cell adhesion and extension. Concomitant high DNMT1 and UHRF1 mRNA levels were in HCC patients and indicated worse prognosis. On the one hand, Lenvatinib modulated the transcription of UHRF1 and DNMT1via negatively regulation of ERK/MAPK pathway. On the other hand, Lenvatinib downregulated DNMT1 and UHRF1 expression by promoting their protein degradation through ubiquitin-proteasome pathway, consequently, resulting in upregulation of E-Cadherin. Moreover, Lenvatinib attenuated Huh7 cell adhesion and metastasis in vivo. Our findings provided insight into the intriguing molecular mechanisms regarding the anti-metastasis effect of Lenvatinib in HCC.

Cross-sectional study of the quality of randomized control trials on problem-based learning in medical education.

Problem-based learning (PBL) is increasingly being used in medical education globally, but its effectiveness in teaching remains controversial. A randomized controlled trial (RCT) is the method of choice for evaluating its effectiveness. The quality of an RCT has a significant effect on this evaluation, but to date we have not seen an assessment of the quality of RCTs for PBL. Two researchers searched MEDLINE and EMBASE for RCTs addressing PBL in medical education. The overall quality of each report was measured on a 28-point overall quality score (OQS) based on the 2010 revised Comprehensive Standards for Reporting and Testing (CONSORT) Statement. Furthermore, to study the key factors affecting OQS more effectively, a linear regression model of those factors was established using SPSS. After literature screening, 30 RCTs were eventually included and analyzed. The median OQS was 15 (range, 7-20), which meant that half of the items in the revised 2010 CONSORT statement were poorly reported in at least 40% of the RCTs analyzed. The regression model showed that the year of publication of RCTs and the impact factors of the journals in which they were published were the main factors affecting OQS. The overall quality of reporting of RCTs on PBL teaching in medical education was not satisfactory. Some RCTs were subjectively selective in reporting certain items, leading to heterogeneity in quality. It is expected that statisticians will develop new standards more suitable for evaluating RCTs related to teaching research and that editors and peer reviewers will be required to review the relevant RCTs more strictly.

Multi-Information Flow CNN and Attribute-Aided Reranking for Person Reidentification.

This paper presents a multi-information flow convolutional neural network (MiF-CNN) model for person reidentification (re-id). It contains several specific multilayer convolutional structures, where the input and output of a convolutional layer are concatenated together on channel dimension. With this idea, layers of model can go deeper and feature maps can be reused by each subsequent layer. Inspired by an image caption, a person attribute recognition network is proposed based on long-short-term memory network and attention mechanism. By fusing identification results of MiF-CNN and attribute recognition, this paper introduces the attribute-aided reranking algorithm to improve the accuracy of person re-id further. Experiments on VIPeR, CUHK01, and Market1501 datasets verify the proposed MiF-CNN can be trained sufficiently with small-scale datasets and obtain outstanding accuracy of person re-id. Contrast experiments also confirm the availability of the attribute-assisted reranking algorithm.