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Background 20(S)-protopanaxadiol (20(S)-PPD), one of the main active metabolites of ginseng, performs a broad spectrum of anti-tumor effects. Our aims are to search out new strategies to enhance anti-tumor effects of natural products, including 20(S)-PPD. In recent years, fasting has been shown to be multi-functional on tumor progression. Here, the effects of fasting combined with 20(S)-PPD on hepatocellular carcinoma growth, apoptosis, migration, invasion and cell cycle were explored. Methods CCK-8 assay, trypan blue dye exclusion test, imagings photographed by HoloMonitorTM M4, transwell assay and flow cytometry assay were performed for functional analyses on cell proliferation, morphology, migration, invasion, apoptosis, necrosis and cell cycle. The expressions of genes on protein levels were tested by western blot. Tumor-bearing mice were used to evaluate the effects of intermittent fasting combined with 20(S)-PPD. Results We firstly confirmed that fasting-mimicking increased the anti-proliferation effect of 20(S)-PPD in human HepG2 cells in vitro. In fasting-mimicking culturing medium, the apoptosis and necrosis induced by 20(S)-PPD increased and more cells were arrested at G0-G1 phase. Meanwhile, invasion and migration of cells were decreased by down-regulating the expressions of matrix metalloproteinase (MMP)-2 and MMP-9 in fasting-mimicking medium. Furthermore, the in vivo study confirmed that intermittent fasting enhanced the tumor growth inhibition of 20(S)-PPD in H22 tumor-bearing mice without obvious side effects. Conclusion Fasting significantly sensitized HCC cells to 20(S)-PPD in vivo and in vitro. These data indicated that dietary restriction can be one of the potential strategies of chinese medicine or its active metabolites against hepatocellular carcinoma., Graphical abstract Image 1

Irisin, as a novel and versatile hormone secreted by skeletal myocytes and fat tissues, is reported to be involved in inflammation-related diseases; one of the main characteristics of severe acute pancreatitis (SAP) is inflammatory responses. This study aims to describe the characteristics of irisin in SAP.Here, we enrolled 50 controls, 20 with no acute pancreatitis (AP), 20 AP, and 12 SAP patients, determined white blood cell, blood glucose, C-reactive protein, urine amylase, blood lipase, and serum irisin using an analyzer and enzyme-linked immunosorbent assay at the indicated time-points, analyzed the correlations of irisin with blood glucose, sex, and age, and then predicted the morality and complications of organ failure and/or exacerbations of comorbidities in SAP by irisin.The results showed no significant difference in all groups in the clinical parameters (P0.05), except that white blood cell was significantly higher in no AP, AP, and SAP than the controls (P0.05). In addition, irisin levels were significantly lower and maintained a steadily low trend in the process of SAP than others (P0.05), whereas C-reactive protein, urine amylase, and blood lipase in the SAP and AP groups were higher than others and kept decreasing tendency (P0.01). Moreover, the irisin level in female SAP patients was significantly higher than that in male patients, but no differences were found in the other groups (P0.05). In addition, the correlation between irisin levels and blood glucose was better in the SAP group than that between irisin levels and age in SAP patients and controls, although a relatively better correlation was found in SAP patients than the controls. Finally, the prognostic significance of mortality and complications of SAP according to irisin levels represented significantly, especially for complications of organ failure and/or exacerbations of comorbidities in female SAP.Therefore, serum irisin level has unique characteristics and may be an independent factor and useful to predict the mortality, and complications in SAP patients, especially in female SAP patients.

Background Infection in patients with acute pancreatitis, especially severe acute pancreatitis patients, is a common and important phenomenon, and the distributions and drug resistance profiles of bacteria causing biliary infection and related risk factors are dynamic. We conducted this study to explore the characteristics of and risk factors for bacterial infection in the biliary tract to understand antimicrobial susceptibility, promote the rational use of antibiotics, control multidrug-resistant bacterial infections and provide guidance for the treatment of acute pancreatitis caused by drug-resistant bacteria. Methods The distribution of 132 strains of biliary pathogenic bacteria in patients with acute pancreatitis from January 2016 to December 2020 were analyzed. We assessed drug resistance in the dominant Gram-negative bacteria and studied the drug resistance profiles of multidrug-resistant bacteria by classifying Enterobacteriaceae and nonfermentative bacteria. We then retrospectively analyzed the clinical data and risk factors associated with 72 strains of Gram-negative bacilli, which were divided into multidrug-resistant bacteria (50 cases) and non-multidrug-resistant bacteria (22 cases). Results The main bacteria were Escherichia coli, Acinetobacter baumannii, Klebsiella pneumoniae and Pseudomonas aeruginosa. Extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli had a 66.67% detection rate. Acinetobacter baumannii had more than 50.00% drug resistance to carbapenems, ESBL-producing Klebsiella pneumoniae had 100.00% drug resistance, and Pseudomonas aeruginosa had 66.67% resistance to carbapenems. Multivariate logistic regression analysis suggested that the administration of third- or fourth-generation cephalosporins was an independent risk factor for Gram-negative multidrug-resistant biliary bacterial infection in acute pancreatitis patients. Conclusion Drug resistance among biliary pathogens in acute pancreatitis patients remains high; therefore, rational antimicrobial drug use and control measures should be carried out considering associated risk factors to improve diagnosis and treatment quality in acute pancreatitis patients.

Huganpian (HGP), a traditional chinese medicine composed of 6 herbs, possesses excellent therapeutic effects in clinical application. In this study, we aimed to elucidate the anti-tumor activity and the underlying mechanisms of HGP in liver cancer. The results of this study indicated that HGP effectively inhibited liver cancer growth in vitro and in vivo in a dose-dependent manner. Mechanistically, HGP exerted its anti-tumor effects by triggering autophagy with increased LC3Ⅱ and beclin1 levels and arrested the cell cycle on G0-G1 phase by downregulating the expressions of cyclin-dependent kinase 2 (CDK2), cyclin-dependent kinase 4 (CDK4) and cyclinE1 in vitro and in vivo. Meanwhile, HGP did not induce apoptosis significantly. Importantly, we also confirmed that there were fewer side effects of HGP on immune system. Taken together, our findings suggest for the first time that HGP may become a promising drug or adjuvant drug with a lower toxicity for liver cancer treatment in the future.

Macrophages exhibit phenotypic heterogeneity under both physiological and pathological conditions. Applications targeting M2-like tumor-associated macrophages (TAMs) improve outcome in solid tumors. Considerable differences are detected between leukemia-associated macrophages (LAMs) and TAMs. However, application to induce M1 characteristics in heterogeneous LAMs has not been established. Here we analyzed clinical relevance of macrophage phenotypes in human acute myeloid leukemia (AML), studied phenotypic evolution of bone marrow (BM) and spleen (SP) LAMs in mouse AML and T cell acute lymphoblastic leukemia (T-ALL) models, explored mechanism leading to different LAM phenotypes and tried to eliminate pro-leukemic effects by inducing M1 characteristics. The results showed that more M2-like LAMs but not total LAMs correlated with worse prognosis in AML patients. Heterogeneity of LAM activation in tissue-specific leukemic microenvironments was observed in both AML and ALL models, i.e. SP LAMs evolved with more M2 characteristics while BM LAMs with more M1 characteristics. Furthermore, IRF7 contributed to M1 characteristics through the activation of SAPK/JNK pathway. Moreover, targeting IRF7-SAPK/JNK pathway to induce M1 characteristics in LAMs contributed to prolonged survival in leukemia mice. Our study provides the potential target for macrophage based immuno-therapy strategy against leukemia.

Macrophages, which have remarkable plasticity, are indispensable cellular components and play essential roles in both innate and adaptive immune responses. Peritoneal macrophages show unique gene expression profile and peritoneal cavity is also involved in leukemia. However, the characteristics of peritoneal leukemia-associated macrophages (Per LAMs) have not been established. Here we studied the phenotype of Per LAMs, their subpopulations in Notch1-induced acute lymphoblastic leukemia mice and compared with LAMs from BM or spleen in the same model. Peritoneal macrophages and Per LAMs simultaneously expressed high level iNOS and Arg1, which was not commonly observed in macrophages from different origins. Furthermore, LAMs from peritoneal, BM and spleen expressed lower level CSF-1, TGF-β1 and VEGF-A than tumor-associated macrophages (TAMs). Moreover, diverse responses in the expression of some phenotype-associated genes to leukemia microenvironments were detected among those LAMs. In addition, Per LAMs can be sub-divided into CD206+ and CD206- sub-populations, which expressed both M1- and M2-associated genes. These results revealed the unique phenotype of Per macrophages and Per LAMs and contributed to better understanding of macrophage plasticity and their pathological roles in leukemia.

Tumor-associated macrophages are widely studied in solid tumors. The distribution of macrophages in lymph node samples was found to be associated with the prognosis of lymphoma patients. However, the role of macrophages in leukemia and their functional and phenotypic characteristics in hematopoietic malignancies have not been defined. In this study, we examined the distribution and functional and phenotypic characteristics of macrophages in a Notch1-induced mouse model of T cell acute lymphoblastic leukemia (T-ALL). The distribution of macrophages in bone marrow (BM) and spleen, which are proposed as BM and spleen leukemia-associated macrophages (LAMs), were different during the development of leukemia. LAMs stimulated the proliferation of T-ALL cells and had higher migration activity. RNA-sequencing analysis revealed that gene expression profiles of BM and spleen LAMs showed considerable differences. RT-PCR analysis showed that LAMs expressed both M1- and M2-associated phenotypic genes, but they expressed much lower levels of TGF-β1, VEGF-A, and CSF-1 than did tumor-associated macrophages from B16 melanoma. Furthermore, spleen LAMs more potently stimulated the proliferation of T-ALL cells compared with BM LAMs. Moreover, LAMs could be subdivided into M1-like (CD206−) and M2-like (CD206+) groups. Both CD206+ and CD206− LAMs stimulated the proliferation of T-ALL cells, although CD206+ LAMs expressed higher levels of most M1- and M2-associated genes. These results suggested the functional and phenotypic characteristics of LAMs, which were modified by organ specific microenvironments. Our results broaden our knowledge about macrophages in malignant microenvironments from solid tumors to leukemia.

Nucleotides are important players in intercellular signaling communication network. P2X family receptors (P2XRs) are ATP-gated plasma membrane ion channels with diverse biological functions. Macrophages are important components in the microenvironment of hematopoiesis participating in both physiological and pathological processes. However, the role of P2XRs in macrophages in leukemia has not been established. Here we investigated expression pattern and functions of P2XRs in macrophages from bone marrow (BM) and spleen of Notch1-induced T-ALL mice. Real-time PCR showed that P2XRs except P2X5R were expressed in BM and spleen macrophages. Furthermore, with the development of leukemia, the expression of P2X7R increased in both BM and spleen macrophages whereas expression of P2X1R increased in spleen macrophages. Live cell imaging recoding the Ca(2+) response demonstrated that P2X7R expressed in macrophages was functional. TUNEL and electron microscopy analysis found that apoptotic macrophages were frequently observed in BM and spleen at late stage of leukemia, which was partly contributed by the activation of overexpressed P2X7R. Our results suggested that the intercellular communication mediated by nucleotides might orchestrate in the pathological process of leukemia and could be a potential target for the treatment of leukemia.

Fbxw11, as a member of F-box proteins family, is a constituent of the SCF (Skp1-Cul1-F box) ubiquitin ligase complex. This ligase ubiquitinates specifically phosphorylated substrates and controls the degradation and half-life of key cellular regulators. So, Fbxw11 play a pivotal role in many aspects of hematopoiesis and tumorigenesis through regulating various signal transduction pathways. We found two transcript variants (Fbxw11c and Fbxw11d) in mouse bone marrow. However the role of Fbxw11 variants in the development of leukemia and the underlying mechanisms remain largely unknown. In this study, we cloned three transcript variants (Fbxw11a, Fbxw11c and Fbxw11d) to study the biological function of Fbxw11 in leukemia. In order to investigate the role of Fbxw11 variants in leukemia, we established L1210 cell lines with over-expression of Fbxw11a, Fbxw11c and Fbxw11d respectively using the lentivirus system. The effect of Fbxw11 variants on proliferation of leukemia cells in vitro was first detected. Growth curve of leukemia cells with Fbxw11a, Fbxw11c or Fbxw11d over-expression was established by cell counting. The results suggested that over-expression of Fbxw11 variants stimulated the growth of leukemia cells. Then MTT experiment was carried out to study the effect of Fbxw11 variants on leukemia cell proliferation and the results showed that Fbxw11 variants increased the proliferation of L1210 cells in vitro. To further confirm the effects of Fbxw11 variants on proliferation of leukemia cells in vivo, tumor xenografts model with over-expression of Fbxw11a, Fbxw11c and Fbxw11d in DBA/2 mice was established. Leukemia cells L1210 with over-expression of Fbxw11a, Fbxw11c and Fbxw11d respectively were transplanted into DBA/2 mice by hypodermic injection. The tumor growth curves showed that tumor growth was increased in Fbxw11 variants over-expression group compared to the control group. Mice were sacrificed on day 28 after transplantation, greater volume of the xenograft tumors were obtained from Fbxw11 variants over-expression group than control group. Therefore, Over-expression of Fbxw11 variants could increase growth of tumor in vivo. To further investigate the molecular mechanism under the effect of Fbxw11 variants on proliferation of leukemia cells, we tested the apoptosis and cell cycle of leukemia cells with Fbxw11 variants over-expression. Over-expression of Fbxw11 variants did not affect the cell apoptosis but accelerated the process of cell cycle. These results revealed that the increased cell proliferation was not due to decrease in cell apoptosis but due to increase in cell cycle. In addition, we tested the effect of Fbxw11 variants on the signal transduction by dual-luciferase reporter gene system. The results showed that over-expression of Fbxw11 variants caused the activation of NF-κB signaling pathway. In conclusion, our findings suggest that Fbxw11 variants have promoting effect on cell proliferation of leukemia cells. The effect of Fbxw11 variants on cell proliferation are due to accelerated the process of cell cycle but not decreasing in cell apoptosis. Further study demonstrated that Fbxw11 variants promote cell proliferation through activating the NF-κB signaling pathway. The important role of Fbxw11 in regulating the development of leukemia suggests that a potent rationale for developing Fbxw11 as a potential therapeutic target against leukemia. Disclosures No relevant conflicts of interest to declare.

F-box proteins serve as the substrate recognition subunit for the SCF E3 ubiquitin ligases. These ligases ubiquitinate specifically phosphorylated substrates and play a pivotal role in the regulation of various signal transduction pathways, which, in turn, are essential for many aspects of tumorigenesis. However the role of Fbxw11 in the development of leukemia and the underlying mechanisms remain largely unknown. Here, we found two transcript variants (Fbxw11c and Fbxw11d) expressed in mouse bone marrow. More importantly, the expression of Fbxw11 in normal hematopoietic stem cells (HSCs) from Notch1-induced leukemia mice was higher than that from control mice. In order to investigate the role of Fbxw11 in leukemia, we established L1210 cell lines over-expressing Fbxw11c or Fbxw11d respectively using the lentivirus system. Over-expression of both Fbxw11 variants stimulated the proliferation of L1210 cells in vitro by cell counting and MTT assay. The tumor xenografts model with over-expression of Fbxw11c or Fbxw11d in DBA/2 mice was further established. Over-expression of both variants resulted in the increased tumor growth in vivo. Investigation of the molecular mechanism revealed that the increased cell proliferation was not due to decrease in cell apoptosis but due to increase in cell cycle. Further studies showed that overexpression of both Fbxw11c and Fbxw11d caused the activation of NF-κB signaling pathway. These findings suggest the important role of Fbxw11 in regulating the development of leukemia and indicate a potent rationale for developing Fbxw11 as a potential therapeutic target against leukemia. Citation Format: Lina Wang, Jinfeng Liao, Xiao Yang, Wenli Feng, Shayan Chen, Guoguang Zheng. Fbxw11 promotes leukemia development by activating the NF-κB signaling pathway. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4433. doi:10.1158/1538-7445.AM2014-4433