Evaluation of immune and pyroptosis status in a model of sepsis-induced secondary pneumonia.

In recent years, researchers have focused on studying the mechanism of sepsis-induced immunosuppression, but there is still a lack of suitable animal models that accurately reflect the process of sepsis-induced immunosuppression. The aim of this study was to evaluate the immune status at various stages in a model of sepsis-induced secondary pneumonia and to demonstrate whether pyroptosis is one of the modes of immune cell death in sepsis. Firstly, we established a sepsis model in C57BL/6J mice using cecal ligation and puncture (CLP). The surviving mice were treated with a 40 µL suspension of P.aeruginosa (Pa) under anesthesia on day 4 post-CLP to establish a sepsis-induced secondary pneumonia model. Secondly, routine blood tests, serum ALT and PCT levels, gross lung specimens, and H&E staining of the lung and liver tissues were used to assess the successful establishment of this model. Serum levels of TNF-α and IL-6, the CD4+/CD8+ratio in blood, H&E staining of the spleen, and immunohistochemistry of CD4 and CD8 in the spleen were detected to evaluate the immune status of the model mice. Finally, the expression levels of pyroptosis-related proteins in the spleen were detected by Western blot. The expression of GSDMD was assessed using immunohistochemistry, and pyroptosis was directly observed through transmission electron microscopy. The experimental results above confirmed the successful construction of the model for sepsis-induced secondary pneumonia, demonstrating its ability to reflect sepsis-induced immunosuppression. Moreover, the expression of pyroptosis-related proteins, immunohistochemical GSDMD, and transmission electron microscopy of the spleen showed that pyroptosis was one of the modes of immune cell death in sepsis.

Integrative analysis reveals that SLC38A1 promotes hepatocellular carcinoma development via PI3K/AKT/mTOR signaling via glutamine mediated energy metabolism.

Although hepatocellular carcinoma (HCC) is rather frequent, little is known about the molecular pathways underlying its development, progression, and prognosis. In the current study, we comprehensively analyzed the deferentially expressed metabolism-related genes (MRGs) in HCC based on TCGA datasets attempting to discover the potentially prognostic genes in HCC. The up-regulated MRGs were further subjected to analyze their prognostic values and protein expressions. Twenty-seven genes were identified because their high expressions were significant in OS, PFS, DFS, DSS, and HCC tumor samples. They were then used for GO, KEGG, methylation, genetics changes, immune infiltration analyses. Moreover, we established a prognostic model in HCC using univariate assays and LASSO regression based on these MRGs. Additionally, we also found that SLC38A1, an amino acid metabolism closely related transporter, was a potential prognostic gene in HCC, and its function in HCC was further studied using experiments. We found that the knockdown of SLC38A1 notably suppressed the growth and migration of HCC cells. Further studies revealed that SLC38A1 modulated the development of HCC cells by regulating PI3K/AKT/mTOR signaling via glutamine mediated energy metabolism. In conclusion, this study identified the potentially prognostic MRGs in HCC and uncovered that SLC38A1 regulated HCC development and progression by regulating PI3K/AKT/mTOR signaling via glutamine mediated energy metabolism, which might provide a novel marker and potential therapeutic target in HCC.

Altered Signaling Pathways Revealed by Comprehensive Genomic Profiling in Patients With Unknown Primary Tumors.

Purpose: Carcinoma of unknown primary (CUP) is a clinically aggressive disorder with early tumor dissemination. Identifying molecular traits of CUP can be not only beneficial for a better therapeutic approach but also potentially valuable for patients with general metastatic dissemination. Patients and Methods: We retrospectively investigated a total of 35 unique CUP cases. Tumor tissue samples were available in 26 patients, and plasma samples were available in 22 patients. Targeted sequencing was performed with a panel of 416 pan cancer-related genes. Results: A genomic landscape of the CUP cohort showed that TP53 mutation was the most frequently observed mutation while MYC amplification was the most common CNV. Aberrant TP53, RTK-RAS, and PI3K signaling pathways were also prevalent, identified in more than half of the cases with tumor tissue. Around 58% of the CUP cases harbored homologous recombinant repair (HRR) pathway gene alterations. The tumor mutational load of CUP patients with altered HRR pathway displayed a significant increase than that of patients with intact HRR. Clinically actionable mutations were identified in eight patients, which may benefit from targeted therapies. Eight patients were treated with platinum-based chemotherapy, showing different responses, HRR, and LOH status. Conclusion: Collectively, our data have provided much-need insights into the treatment options for patients diagnosed with CUP in the era of precision medicine.

The Role of HMGB1 in Invasive Candida albicans Infection.

INTRODUCTION: High mobility group box 1 (HMGB1) is an important "late" inflammatory mediator in bacterial sepsis. Ethyl pyruvate (EP), an inhibitor of HMGB1, can prevent bacterial sepsis by decreasing HMGB1 levels. However, the role of HMGB1 in fungal sepsis is still unclear. Therefore, we investigated the role of HMGB1 and EP in invasive C. albicans infection. METHODS: We measured serum HMGB1 levels in patients with sepsis with C. albicans infection and without fungal infection, and control subjects. We collected clinical indices to estimate correlations between HMGB1 levels and disease severity. Furthermore, we experimentally stimulated mice with C. albicans and C. albicans + EP. Then, we examined HMGB1 levels from serum and tissue, investigated serum levels of tumor necrosis factor α (TNF-α) and interleukin 6 (IL-6), determined pathological changes in tissues, and assessed mortality. RESULTS: Serum HMGB1 levels in patients with severe sepsis with C. albicans infection were elevated. Increased HMGB1 levels were correlated with procalcitonin (PCT), C-reactive protein (CRP), 1,3-ß-D-Glucan (BDG) and C. albicans sepsis severity. HMGB1 levels in serum and tissues were significantly increased within 7 days after mice were infected with C. albicans. The administration of EP inhibited HMGB1 levels, decreased tissue damage, increased survival rates and inhibited the release of TNF-α and IL-6. CONCLUSIONS: HMGB1 levels were significantly increased in invasive C. albicans infections. EP prevented C. albicans lethality by decreasing HMGB1 expression and release. HMGB1 may provide an effective diagnostic and therapeutic target for invasive C. albicans infections.

Efficacy and safety of raltitrexed-based transcatheter arterial chemoembolization for intermediate and advanced hepatocellular carcinoma: A multicenter real-world study.

AIM: This study aimed to evaluate efficacy and safety of raltitrexed-based transcatheter arterial chemoembolization (TACE) for intermediate and advanced hepatocellular carcinoma (HCC) using real-world evidence. METHODS: All eligible HCC cases were collected from multiple centers in Chongqing, China, from January 2013 to December 2018 and divided into the raltitrexed group (raltitrexed + lobaplatin + pirarubicin) and control group (lobaplatin + pirarubicin). Propensity score matching (PSM) with a 1:1 ratio was used to eliminate the imbalance of potential confounding factors between groups. The primary end-point was overall survival (OS) and the secondary end-points were progression-free survival (PFS) and disease control rate. RESULTS: The median follow-up period for patients in the raltitrexed and control groups was 8.7 and 5.9 months, respectively. After PSM, median OS was 10.0 months in the raltitrexed group and 7.0 months in the control group (p = 0.002). The 6-month, 1-year, and 2-year OS rates of the raltitrexed group were significantly higher than those of the control group (78.2% vs. 60.9%, p = 0.010; 43.5% vs. 22.8%, p = 0.030; and 17.4% vs. 2.2% p = 0.001, respectively). Multivariate analysis of these propensity score-matched HCC patients revealed treatment, age, tumor size, lipiodol accumulation, and the number of TACE cycles as independent predictors of OS (all p < 0.05). The disease control rate of the raltitrexed and control groups was 87.4% and 65.8%, respectively (p < 0.001). CONCLUSIONS: Raltitrexed-based TACE can prolong the OS of patients with intermediate and advanced HCC in a real-world clinical setting, and is safe and tolerable.

Comparative analysis of first-line treatment regimens for advanced EGFR-mutant non-small cell lung cancer patients with stable brain metastases.

BACKGROUND: To compare the survival outcomes of first-line treatment regimens for advanced epidermal growth factor receptor (EGFR)-mutant non-small cell lung cancer (NSCLC) patients with stable brain metastases. METHODS: We conducted a systematic review of available data from randomized controlled trials (RCTs) of first-line treatment regimens of NSCLC patients with stable brain metastases. Progression free survival (PFS) and overall survival (OS) were extracted and analysed from the RCT subgroups. A network meta-analysis was constructed using the Bayesian statistical model to synthesize the survival outcomes of all the treatments. RESULTS: The analysis included 6 eligible RCT subgroups with 417 patients and 7 treatment regimens osimertinib, afatinib, first-generation EGFR-TKI (gefitinib or erlotinib), erlotinib + bevacizumab, gefitinib + pemetrexed + carboplatin, gemcitabine + cisplatin, and pemetrexed + cisplatin. Of these seven treatment regimens, gefitinib + pemetrexed + carboplatin had the highest potential for favorable PFS and OS, followed by osimertinib, in the treatment of advanced EGFR-mutant NSCLC patients with stable brain metastases. None of the results met the predetermined statistical significance of P<0.05. CONCLUSIONS: The regimens of "Gefitinib + pemetrexed + carboplatin" and "Osimertinib" were associated with the most favorable PFS and OS compared to the other therapies in advanced EGFR-mutant NSCLC patients with stable brain metastases, although the difference between these regimens and the others was not statistically significantly different.

Effect of Autologous Bone Marrow Stem Cell Therapy in Patients with Liver Cirrhosis: A Meta-analysis.

Background and Aims: Although autologous bone marrow stem cell (BMSC) transplantation is an effective treatment for liver cirrhosis, there are few reports describing the optimal delivery route and number of injected BMSCs. Methods: A literature search was conducted using PubMed, ISI Web of Science, Cochrane Central Register of Controlled Trials, and EBSCO. A meta-analysis was performed to assess the effect of BMSCs on liver and coagulation function indices. Subgroup analysis was performed based on number of injected BMSCs, delivery route, and length of follow-up. Results: A total of 15 studies were selected from among 1903 potential studies for analysis. Autologous BMSC transplantation significantly improved aspartate aminotransferase, total bilirubin, albumin, prothrombin time, prothrombin activity, prothrombin concentration, Child-Pugh score, and model for end-stage liver disease. In the subgroup analysis of cell numbers, all four of the indices were significantly improved when the number of BMSCs was >4 × 108. The subgroup analysis referring to the delivery route showed that arterial infusion increased the therapeutic effect over venous infusion. Finally, in the subgroup analysis of follow-up length, the results showed that BMSC therapy significantly improved liver function at 2 weeks after transplantation. In addition, this therapy improved coagulation 4 weeks after the transplant, with a maintenance of efficacy for up to 24 weeks. Conclusions: Autologous BMSC therapy is beneficial for liver improvement and coagulation in patients with liver cirrhosis. The therapeutic effect was generated at 2-4 weeks after transplantation. The effect lasted for 24 weeks but no more than 48 weeks. The greatest benefit to patients was observed with a 4 × 108 autologous BMSC transplant via the hepatic artery.

Large-dynamic-range time pre-compensation scheme for fiber optic time transfer.

In this paper we experimentally demonstrate a transmission delay compensation scheme for precise fiber-optic time transfer. The scheme is based on a clock counter and an electronic variable delay line, which theoretically can provide unlimited compensation range. We perform successive tests in three optical fiber links of different lengths in which both continuous drifts and abrupt hop of the transmission delay are effectively compensated. The total transmission delay variation induced in the experiments is much larger than most of the reported cases. This large-dynamic compensation scheme is quite suitable for time transfer links whose transmission delay varies a lot.

Microrna Expression Profiling of Macrophage Line Raw264.7 Infected by Candida Albicans.

INTRODUCTION: The aim of this study was to clarify the microRNA (miRNA) expression profiles of RAW264.7 macrophages infected by Candida albicans to elucidate the roles of differentially expressed miRNAs and to further explore the mechanisms underlying the immune response to C. albicans infection. METHODS: High-throughput miRNA microarray analysis was performed to detect differentially expressed miRNAs in control and C. albicans-infected RAW264.7 cells. Quantitative real-time PCR analysis was used to verify the microarray results. Target genes of differentially expressed miRNAs were predicted with bioinformatics software. The cell biological processes and signaling pathways of these miRNA-targeted genes involved in C. albicans infection were predicted by gene ontology (GO) enrichment and pathway analyses. RESULTS: Significant upregulation of eight miRNAs (mmu-miR-140-5p, mmu-miR-96-5p, mmu-miR-8109, mmu-miR-466i-3p, mmu-miR-222-5p, mmu-miR-301b-3p, mmu-miR-466g, and mmu-miR-7235-5p) and downregulation of eight miRNAs (mmu-miR-3154, mmu-miR-223-3p, mmu-miR-494-3p, mmu-miR-6908-5p, mmu-miR-188-5p, mmu-miR-6769b-5p, mmu-miR-7002-5p, and mmu-miR-1224-5p) were observed, as compared with the control (fold change ≥2.0 and P < 0.05). GO analysis revealed that both mmu-miR-140-5p and mmu-miR-223-3p participated in immune responses, inflammatory reactions, and cell apoptosis in C. albicans infection. Also, the MAPK signaling pathway was found to play an important role in the immune response against C. albicans infection. CONCLUSIONS: This study revealed comprehensive expression and functional profiles of differentially expressed miRNAs in macrophage RAW264.7 cells infected by C. albicans. These findings should help to further elucidate the mechanisms underlying the immune response to C. albicans infection.

WDM-based radio frequency dissemination in a tree-topology fiber optic network.

In this paper, we propose and demonstrate a scheme to achieve point-to-multipoint dissemination of radio frequency (RF) signals in a local area fiber optic network with tree topology based on wavelength-division multiplexing (WDM) technique. The phase changes caused by the fluctuations of the transfer links are passively canceled at remote end instead of at local end, which makes it feasible to flexibly build a tree-topology local area dissemination network with great cost-effectiveness. For the first time, we study the limit of long-term performance which is caused by temperature-induced variation of group velocity dispersion (TIVGVD) in dissemination networks using WDM techniques. In the proof-of-concept experiments, 38.5 km and 50 km fiber links are established to disseminate a 1 GHz frequency signal with fractional instability of 10(-17) order after 10(4) s averaging time. Then 17.4 nm wavelength spacing is introduced between local carrier and user carrier to verify the theoretical analysis. Under a controlled fiber temperature variation of about 21 °C, the obtained overlapping Allan deviation (ADEV) agrees well with the simulation results after 10(4) s time scales, which proves the validity of our theory. The theory has practical values in predicting and optimizing the capacity and performance of a WDM-based local area RF dissemination network.

Role of high-mobility group box 1 in patients with acute obstructive suppurative cholangitis-induced sepsis.

BACKGROUND: High-mobility group box 1 (HMGB1) is a proinflammatory cytokine that plays an active role during the pathogenesis of inflammatory processes. The primary aim of this study was to detect whether HMGB1 is involved in the pathogenesis of acute obstructive suppurative cholangitis (AOSC). METHODS: We collected peripheral blood samples from 23 patients with AOSC and 23 healthy volunteers who served as normal controls. All participants were tested for HMGB1 mRNA level, HMGB1 protein, tumor necrosis factor alpha (TNF-alpha), and interleukin 10 (IL-10). HMGB1 mRNA levels were tested using real-time polymerase chain reaction. HMGB1 protein expression was measured using Western blot. TNF-alpha and IL-10 were tested using enzyme-linked immunosorbent assay. RESULTS: The expression of HMGB1 mRNA and HMGB1 protein was higher in the AOSC group than in the normal controls (P<0.01), and the levels gradually decreased to normal after treatment of the disease (P<0.01). The content of TNF-alpha and IL-10 in peripheral blood of patients with AOSC was significantly higher than that of normal controls (P<0.01) but decreased to normal levels after the necessary treatment (P<0.01). CONCLUSION: The levels of HMGB1 mRNA and HMGB1 protein were elevated in patients with AOSC, which may play an important role in the inflammation of the bile duct and appears to be associated with the development of sepsis. This suggests the importance of monitoring HMGB1 levels in the management of AOSC-induced sepsis.

Inhibition effect of glycyrrhizin in lipopolysaccharide-induced high-mobility group box 1 releasing and expression from RAW264.7 cells.

INTRODUCTION: High-mobility group box 1 (HMGB1) is a therapeutic target for sepsis. Glycyrrhizin (GL) is the aglycone of glycyrrhizin derived from licorice. We clarified the anti-inflammatory effects of GL. We explored the anti-HMGB1 effect of GL and elucidated its molecular mechanism, which will be of benefit for sepsis treatment. METHODS: We stimulated murine macrophage-like RAW 264.7 cells with lipopolysaccharide (LPS) and LPS + GL, then measured the expression and release of HMGB1. The expression of related signal transduction factors was detected. RESULTS: High-mobility group box 1 was distributed mainly in the nucleus with lower cytoplasmic levels in RAW 264.7 cells before LPS stimulation. After stimulation, cytoplasmic HMGB1 levels increased gradually, whereas in nuclear fluctuation a trend of HMGB1 expression was observed. Significant upregulation of HMGB1 mRNA occurred 12 h after LPS stimulation. Glycyrrhizin prevented the transfer of HMGB1 from the nucleus to the cytoplasm and inhibited upregulation of HMGB1 mRNA induced by LPS. Phospho-p38 mitogen-activated protein kinase and activated activating protein 1 increased significantly 8 h after LPS stimulation. Tumor necrosis factor α and interleukin 6 increased 4 h after LPS stimulation and peaked at 48 h, and HMGB1 increased at 8 h. The Toll-like receptor 4/MD2/nuclear factor κB signaling pathway was activated 4 h after LPS stimulation. Glycyrrhizin inhibited this pathway. CONCLUSIONS: Glycyrrhizin inhibited the expression and release of HMGB1 through blocking the p38 mitogen-activated protein kinase/activating protein 1 signaling pathway then inhibited the massive release of tumor necrosis factor α and interleukin 6.

Stable radio frequency dissemination by simple hybrid frequency modulation scheme.

In this Letter, we propose a fiber-based stable radio frequency transfer system by a hybrid frequency modulation scheme. Creatively, two radio frequency signals are combined and simultaneously transferred by only one laser diode. One frequency component is used to detect the phase fluctuation, and the other one is the derivative compensated signal providing a stable frequency for the remote end. A proper ratio of the frequencies of the components is well maintained by parameter m to avoid interference between them. Experimentally, a stable 200 MHz signal is transferred over 100 km optical fiber with the help of a 1 GHz detecting signal, and fractional instability of 2×10(-17) at 10(5) s is achieved.

The role of high mobility group box chromosomal protein 1 expression in the differential diagnosis of hepatic actinomycosis: a case report.

INTRODUCTION: Primary hepatic actinomycosis is a rare disease, but is important in the differential diagnosis of hepatoma in endemic areas. As high mobility group box chromosomal protein 1 plays an important role in the pathogenesis of both acute and chronic inflammatory conditions, we postulate that high mobility group box chromosomal protein 1 may have a possible pathogenic role in hepatic actinomycosis. To the best of our knowledge, our report is the first to detect an association between highly elevated high mobility group box chromosomal protein 1 expression and hepatic actinomycosis. CASE PRESENTATION: A 67-year-old Chinese man was admitted to our hospital with a three-month history of epigastric pain, anorexia, and subjective weight loss. Ultrasonography and computed tomography of the patient's abdomen confirmed a hypodense mass measuring seven cm in diameter in the left lateral segment of his liver. A hepatic tumor was suspected and surgical resection was scheduled. Histopathologic examination revealed that the overall features of the hepatic tissues were consistent with hepatic actinomycosis. Whole blood and hepatic tissue samples of the patient, of patients who had hepatocellular carcinoma and of healthy donors were collected. Serum high mobility group box chromosomal protein 1 concentration in actinomycosis was 8.5ng/mL, which was higher than the hepatocellular carcinoma level of 5.2ng/mL and the normal level of

Evaluation of high mobility group box 1 protein as a presurgical diagnostic marker reflecting the severity of acute appendicitis.

OBJECTIVES: To validate the role of high mobility group box-1(HMGB1) in diagnosis of acute appendicitis (AA) with different pathological severity. METHODS: According to the pathologically diagnosis, 150 patients underwent appendectomies between Jan. 2007 and Dec, 2010 were divided into acute simple, acute suppurative and acute gangrenous appendicitis as group 1, 2 and 3, respectively. Each patient group contains 50 sex and age matched cases to make comparison with 50 healthy volunteers. The mRNA and protein expression levels of serum HMGB1 were determined by real-time quantitative PCR and enzyme linked immunosorbent assay (ELISA). Serum High-sensitivity C-reactive protein (hs-CRP) levels were determined by rate nephelometric immunoassay. RESULTS: In comparison with health volunteers, relative HMGB1 mRNA levels in group 1, 2 and 3 were significantly increased 3.05 ± 0.51,8.33 ± 0.75 and 13.74 ± 1.09 folds, reflecting a tendency of augmented severity. In accordance, serum protein levels of HMGB1 were 10.97 ± 1.64, 14.42 ± 1.56 and 18.08 ± 2.41 ng/ml in 3 patient groups, which are significantly higher than that of healthy volunteers' 5.47 ± 0.73 ng/ml. hs-CRP levels were 12.85 ± 3.41, 21.04 ± 1.98 and 31.07 ± 5.46 ng/ml in 3 patients groups compared with 2.06 ± 0.77 ng/ml in controls. The concentrations of HMGB1 and hs-CRP were both positively correlated with disease severity. CONCLUSION: Serum HMGB1 constitutes as a valuable marker in diagnosis of AA. Positively correlated with hs-CRP level, mRNA and protein expression of HMGB1 to a certain extent reflected the severity of AA.

Utility of clot formation and lysis assay to monitor global coagulation state of patients with severe acute pancreatitis.

BACKGROUND: Systemic inflammation affects hemostasis during severe acute pancreatitis (SAP). A hypercoagulable state occurs more frequently in SAP, which is not fully detected by traditional coagulation testing. AIMS: The aim of this study was to evaluate the contribution of clot formation and lysis (CloFAL) assay to improve monitoring of global coagulation in patients with SAP. METHODS: Twenty-five patients with SAP who were treated from December 2009 to April 2011 were studied. Plasma was collected at the time of admission, and CloFAL was measured using the CloFAL analyzer. The parameters evaluated include coagulation time (CT), fibrinolysis time (FT), and maximum amplitude (MA), from which the accelerating coagulation extent (ACE, MA/CT), accelerating fibrinolytic extent (AFE, MA/FT), and balance level exponent (BLE, ACE/AFE) were calculated. In addition, laboratory values for the traditional coagulation testing were measured. Values were compared to a control group of 20 healthy subjects. RESULTS: The MA, FT, ACE, and BLE values of the CloFAL assay were significantly increased in the SAP group compared to the control group (p\0.05 for all measurements). For the traditional coagulation testing, fibrinogen, plasminogen, and D-dimer levels were higher in patients in the SAP group compared to the control group (p\0.05). CONCLUSIONS: Our findings using the CloFAL analyzer indicate that the hypercoagulable state was due to increased fibrin generation and invariable fibrinolysis in patients with SAP. CloFAL assay is a simple and useful global coagulation assay to monitor hypercoagulable states during SAP.

LPS induces HMGB1 relocation and release by activating the NF-κB-CBP signal transduction pathway in the murine macrophage-like cell line RAW264.7.

BACKGROUND: High mobility group protein B1 (HMGB1) is an important late inflammatory mediator in sepsis. Understanding the mechanisms that regulate HMGB1 release from cells and their downstream signal transduction pathways may lead to the ability to develop anti-HMGB1 therapies to treat inflammation. MATERIALS AND METHODS: We stimulated murine macrophage-like RAW 264.7 cells with lipopolysaccharide (LPS) and LPS+ ethylpyruvate (EP) and examined the resulting HMGB1 expression and release. We also studied the expression of related signal transduction factors (NF-κB, p38 MAPK, and CBP). RESULTS AND CONCLUSION: Gene expression of HMGB1 mRNA in RAW264.7 cell showed no significant change at 0-18 h after stimulation with LPS, but increased significantly at 24, 36, and 48 h. HMGB1 mRNA expression in the LPS+EP group was significantly lower than in LPS alone. HMGB1 was distributed mainly in the nucleus; the cytoplasmic level was low before LPS stimulation. After stimulation with LPS, cytoplasmic HMGB1 increased gradually and plateaued at a high level at 12-48 h. Nuclear HMGB1 decreased gradually at 12-24 h, then increased, maintaining a comparatively high level at 36-48 h. EP prevented this pattern significantly. LPS induced p38 MAPK activation and NF-κB signal pathways first, followed by CBP activation. Activated CBP acetylated HMGB1 was stored in a crino-lysosome and secreted activated NF-κB resulted in increased transcription and synthesis of HMGB1, but the expression of up-regulated HMGB1 mRNA was delayed. Extracellular HMGB1 originated from early synthetic reserves present in the nucleus. New HMGB1 protein was synthesized in the nucleus and transferred into the cytoplasm, causing an increase in HMGB1 in the nucleus and cytoplasm. EP inhibits HMGB1 mRNA up-regulation and release from LPS- stimulated macrophages. The molecular function of EP is to attenuate the activation p38 MAPK, NF-κB, and CBP signaling pathways.