Matrix metalloproteinase-8 regulates dendritic cell tolerance in late polymicrobial sepsis via the nuclear factor kappa-B p65/ß-catenin pathway.

Background: Tolerogenic dendritic cells (DCs) are associated with poor prognosis of sepsis. Matrix metalloproteinases (MMPs) have been shown to have immunomodulatory effects. However, whether MMPs are involved in the functional reprogramming of DCs is unknown. The study aims to investigate the role of MMPs in sepsis-induced DCs tolerance and the potential mechanisms. Methods: A murine model of late sepsis was induced by cecal ligation and puncture (CLP). The expression levels of members of the MMP family were detected in sepsis-induced tolerogenic DCs by using microarray assessment. The potential roles and mechanisms underlying MMP8 in the differentiation, maturation and functional reprogramming of DCs during late sepsis were assessed both in vitro and in vivo. Results: DCs from late septic mice expressed higher levels of MMP8, MMP9, MMP14, MMP19, MMP25 and MMP27, and MMP8 levels were the highest. MMP8 deficiency significantly alleviated sepsis-induced immune tolerance of DCs both in vivo and in vitro. Adoptive transfer of MMP8 knockdown post-septic bone marrow-derived DCs protected mice against sepsis-associated lethality and organ dysfunction, inhibited regulatory T-cell expansion and enhanced Th1 response. Furthermore, the effect of MMP8 on DC tolerance was found to be associated with the nuclear factor kappa-B p65/ß-catenin pathway. Conclusions: Increased MMP8 levels in septic DCs might serve as a negative feedback loop, thereby suppressing the proinflammatory response and inducing DC tolerance.

Risk model for predicting mortality in patients with necrotizing soft tissue infections in the intensive care unit.

BACKGROUND: The goal of this study is to look into the factors that lead to death in patients with necrotizing soft tissue infections（NSTIs） in the intensive care unit and create a mortality risk model. METHODS: The clinical data of 106 patients with necrotizing soft tissue infections admitted to intensive care unit(ICU) of the First Affiliated Hospital of Wenzhou Medical University between January 2008 and December 2021 were retrospectively analyzed. Univariate analysis and multivariate analysis were performed to evaluate the risk factors impacting patient mortality. The regression coefficient in binary logistic regression analysis was converted into the item score in the model, and then the model score of each patient was calculated. Finally, an ROC curve was constructed to evaluate the efficiency of the model for predicting mortality. Thirteen patients with NSTIs admitted to ICU between January 2022 and November 2022 were used to validate the model. RESULTS: The death group had 44 patients, while the survival group had 62 patients. The overall mortality was 41.5%. Binary logistic regression analysis showed that risk factors for mortality were age≥ 60 years(OR:4.419; 95%CI:1.093-17.862; P = 0.037), creatinine ≥ 132µmol/L(OR:11.166; 95%CI:2.234-55.816; P = 0.003), creatine kinase ≥ 1104 U/L(OR:4.019; 95%CI:1.134-14.250; P = 0.031), prothrombin time ≥ 24.4 s(OR:11.589; 95%CI:2.510-53.506; P = 0.002), and invasive mechanical ventilation (OR:17.404; 95%CI:4.586-66.052; P＜0.000). The AUC of the model for predicting mortality was 0.940 (95% CI:0.894-0.986). When the cut-off value for the model was 4 points, the sensitivity was 95.5% and the specificity was 83.9%. CONCLUSION: The death risk model in this study for NSTIs patients in the intensive care unit shows high sensitivity and specificity. Patients with a score of ≥ 4 points have a higher risk of mortality.

SUPPLEMENTATION WITH NICOTINAMIDE RIBOSIDE ATTENUATES T CELL EXHAUSTION AND IMPROVES SURVIVAL IN SEPSIS.

ABSTRACT: T cell exhaustion is the main cause of sepsis-induced immunosuppression and is associated with the poor prognosis. Nicotinamide adenine dinucleotide (NAD + ) is well known for its anti-aging effect, but its role in sepsis-induced T cell exhaustion remains to be elucidated. In the present study, using a classic septic animal model, we found that the levels of NAD + and its downstream molecule, which is sirtuins 1 (SIRT1), in T cells in sepsis were decreased. Supplementation with nicotinamide ribose (NR), the precursor of NAD + , right after cecal ligation and puncture significantly increased the levels of NAD + and SIRT1. Supplementation with NR alleviated the depletion of mononuclear cells and T lymphocytes in spleen in sepsis and increased the levels of CD3 + CD4 + and CD3 + CD8 + T cells. Interestingly, both Th1 and Th2 cells were expanded after NR treatment, but the balance of Th1/Th2 was partly restored. Nicotinamide ribose also inhibited the regulatory T cells expansion and programmed cell death 1 expression in CD4 + T cells in sepsis. In addition, the bacteria load, organ damage (lung, heart, liver, and kidney), and the mortality of septic mice were reduced after NR supplementation. In summary, these results demonstrate the beneficial effect of NR on sepsis and T cell exhaustion, which is associated with NAD + /SIRT1 pathway.

Mutual promotion of mitochondrial fission and oxidative stress contributes to mitochondrial-DNA-mediated inflammation and epithelial-mesenchymal transition in paraquat-induced pulmonary fibrosis.

BACKGROUND: Pulmonary fibrosis (PF) is one of the main causes of death in patients with paraquat (PQ) poisoning. This study aimed to evaluate the relationship between mitochondrial fission and oxidative stress in PQ-induced epithelial-mesenchymal transition (EMT) and PF. METHODS: C57BL/6 mice and MLE-12 cells were exposed to PQ to construct a PF model in vivo and in vitro. Histological changes in the lungs were examined by hematoxylin and eosin (H&E) staining. Mitochondrial morphology was detected by MitoTracker® Deep Red FM or transmission electron microscopy (TEM). Western blotting and immunofluorescence were used to determine the expression of protein. The migration ability of the cells was detected by the cell scratch test. Mitochondrial DNA (mtDNA) levels were assessed by real-time polymerase chain reaction (PCR). Enzyme-linked immunosorbent assay (ELISA) was applied to detect cytokine levels. Superoxide dismutase (SOD) activity and the levels of glutathione (GSH) and malondialdehyde (MDA) were detected by chemichromatometry. RESULTS: PQ exposure caused EMT and PF in vivo and in vitro. PQ destroyed mitochondrial structure and enhanced the expression of dynamin-related protein 1 (Drp1), which were accompanied by oxidative stress. Inhibiting mitochondrial fission using mitochondrial division inhibitor-1 (Mdivi-1), a selective inhibitor of Drp1, attenuated PQ-induced EMT and oxidative damage. Treatment with N-acetyl-L-cysteine (NAC), an antioxidant, reduced Drp1 expression, attenuated mitochondrial structure damage and inhibited PQ-induced EMT and PF. Both Mdivi-1 and NAC treatment markedly suppressed mtDNA release, the expression of Toll-like receptor 9 (TLR9) and phosphorylation (P)-NF-κB p65 as well as cytokines (interleukin 6 [IL-6], interleukin-1ß [IL-1ß], and tumor necrosis factor-α [TNF-α]) production. CONCLUSION: Mutual promotion of mitochondrial fission and oxidative stress contributes to EMT in PQ-induced PF, which is associated with the mtDNA/TLR9/NF-κB pathway.

Vesicle transport related protein Synaptotagmin-1 mediates paraquat transport to antagonize paraquat toxicity.

In this study, A549/PQ cells with moderate resistance to paraquat (PQ) were obtained by treating A549 cells with PQ, their growth rate was slowed down, the accumulation concentration of PQ and the levels of growth inhibition, injury and early apoptosis induced by PQ were significantly lower than those of parental A549 cells. Microarray screening and RT-qPCR detection found that Synaptotagmin-1 (SYT1) expression in drug-resistant cells was significantly increased, and PQ further enhanced its expression. After inhibiting SYT1 expression in A549/PQ cells, cell viability, intracellular PQ concentration and the expression of Bcl-2, SNAP25 and RAB26 were significantly reduced, while the mortality, early apoptosis rate and Bax expression were significantly increased. In vivo experiments also further showed that PQ promoted the expression of SYT1, SNAP25 and RAB26 in PQ-poisoned mice; when inhibiting SYT1 expression, PQ concentration in lung tissues was significantly increased, and the levels of lung injury and apoptosis were also significantly enhanced, while the expression of SNAP25 and RAB26 was significantly reduced. This indicates that PQ poisoning leads to compensatory up-regulation of vesicle transport related proteins such as SYT1 in vivo, thereby promoting PQ transmembrane transport, and then reducing the pulmonary accumulation of PQ and PQ-caused lung injury.

CD71+ Erythroid Cell Expansion in Adult Sepsis: Potential Causes and Role in Prognosis and Nosocomial Infection Prediction.

BACKGROUND: Immune suppression contributes to nosocomial infections (NIs) and poor prognosis in sepsis. Recent studies revealed that CD71+ erythroid cells had unappreciated immunosuppressive functions. This study aimed to investigate the values of CD71+ erythroid cells (CECs) in predicting NIs and prognosis among adult septic patients. The potential factors associated with the expansion of CECs were also explored. METHODS: In total, 112 septic patients and 32 critically ill controls were enrolled. The frequencies of CD71+ cells, CD71+CD235a+ cells, and CD45+ CECs were measured by flow cytometry. The associations between CECs and NIs and 30-day mortality were assessed by ROC curve analysis and Cox and competing-risk regression models. Factors associated with the frequency of CECs were identified by linear regression analysis. RESULTS: The percentage of CD71+ cells, CECs, and CD45+ CECs were higher in septic patients than critically ill controls. In septic patients, the percentages of CD71+ cells, CECs, and CD45+ CECs were associated with NI development, while CD71+ cells and CECs were independently associated with 30-day mortality. Linear regression analysis showed that the levels of interleukin (IL)-6 and interferon (IFN)-γ were positively associated with the frequencies of CD71+ cells, CECs, and CD45+ CECs, while IL-10 was negatively associated with them. Additionally, the levels of red blood cells (RBCs) were negatively associated with the percentage of CD45+ CECs. CONCLUSIONS: CECs were expanded in sepsis and can serve as independent predictors of the development of NI and 30-day mortality. Low levels of RBCs and high levels of IL-6 and IFN-γ may contribute to the expansion of CECs in sepsis. TRIAL REGISTRATION: ChiCTR, ChiCTR1900024887. Registered 2 August 2019, http://www.chictr.org.cn/showproj.aspx?proj=38645.

Arterial oxygen pressure targets in critically ill patients: Analysis of a large ICU database.

BACKGROUND: Providing supplemental oxygen is common in the management of critically ill patients, yet the optimal oxygen regimen remains unclear. OBJECTIVES: To explore the optimal range of PaO2 in critically ill patients. METHODS: This is a retrospective study conducted in the Medical Information Mart for Intensive Care III (MIMIC-III) database. The patients with a least 48 h of oxygen therapy were included. Nonlinear regression was used to analyze the association between PaO2 and mortality. We derived an optimal range of PaO2 and evaluated the association between the proportion of PaO2 measurements within this range and mortality. RESULTS: In total, 8401 patients were included in the study. A J-shaped relationship was observed between median PaO2 and hospital mortality. Compared with the reference group of 100-120 mmHg, patients with values of 80-100 mmHg and 120-140 mmHg had higher hospital mortality (adjusted odds ratio [aOR], 1.23; 95% CI, 1.05-1.43 and 1.29; 95%CI, 1.08-1.54, respectively). Similarly, mortality rates were significantly higher for PaO2 <80 mmHg and ≥140 mmHg (aOR, 1.97; 95%CI, 1.58-2.45 and 1.42; 95%CI, 1.19-1.69, respectively). Patients spent a greater proportion of time within 100-120 mmHg tended to have a lower mortality rate. CONCLUSION: Among critically ill patients, the relationship between median PaO2 and hospital mortality was J-shaped. The lowest rates of mortality was observed in those with PaO2 levels within 100 to 120 mmHg.

Partial Depletion of Regulatory T Cells Enhances Host Inflammatory Response Against Acute Pseudomonas aeruginosa Infection After Sepsis.

Immune dysfunction contributes to secondary infection and worse outcomes in sepsis. Regulatory T cells (Tregs) have been implicated in sepsis-induced immunosuppression. Nevertheless, the role of Tregs in secondary infection after sepsis remains to be determined. In the present study, a two-hit model which mimics clinical conditions was used and the potential role of Tregs in secondary Pseudomonas aeruginosa infection post-sepsis was investigated. Results showed that mice were susceptible to secondary P. aeruginosa infection 3 days, but not 7 days, post-cecal ligation and puncture (CLP). The levels of IL-17A, IL-1ß, and IL-6 remained low in CLP mice after P. aeruginosa infection, while the levels of IL-10 increased significantly. Additionally, increased number of Tregs in both lung and spleen was observed in "two-hit" mice. Injection with PC61 (anti-CD25) mAb reduced the number of Tregs by 50% in spleen and 60% in lung of septic mice. This partial depletion of Tregs elevated IL-17A, IL-1ß, and IL-6 production and decreased IL-10 levels in septic mice with P. aeruginosa infection, leading to lower bacterial load, attenuation of lung injury, and improvement of survival. The present findings demonstrate that Tregs play a crucial role in secondary P. aeruginosa infection after sepsis by modulating the inflammatory response.

Ethyl pyruvate reverses development of Pseudomonas aeruginosa pneumonia during sepsis-induced immunosuppression.

Sepsis is characterized by an innate immune response and the following immune dysfunction which can increase the emergence of secondary infections. Ethyl pyruvate (EP) has multiple immunoregulation functions in several serious illnesses, such as burn injury, severe sepsis and acute respiratory syndrome. However, little data was shown the effect of EP administration on immunosuppression post-CLP and the following secondary infection. The cecal ligation and puncture (CLP) followed by the induction of Pseudomonas aeruginosa (PA) was used as a clinically relevant two-hit model of sepsis. We assessed the survival rate, lung damage and lung bacterial clearance in vehicle or EP treatment group to demonstrate the lung response to Pseudomonas aeruginosa of septic mice. Then cytokines including lung IL-6, IL-1ß, IL-10 and plasma HMGB1, apoptosis of splenic immune cells and Foxp3 level on regulatory T cells (Tregs) were studied to demonstrate the mechanisms of EP administration on two-hit mice. We found that the susceptibility of septic mice to Secondary Pseudomonas aeruginosa pneumonia could be down-regulated by ethyl pyruvate treatment and the protective effects of EP may via decreasing lung IL-10 and plasma HMGB1 expression, inhibiting the function of Tregs and relieving the apoptosis of splenic immune cells. The "immune paralysis" post-sepsis still remains a rigorous challenge for curing sepsis, our study may aid in the development of new therapeutic strategies to this problem.

Growth Arrest-Specific 6 Enhances the Suppressive Function of CD4+CD25+ Regulatory T Cells Mainly through Axl Receptor.

Background. Growth arrest-specific (Gas) 6 is one of the endogenous ligands of TAM receptors (Tyro3, Axl, and Mertk), and its role as an immune modulator has been recently emphasized. Naturally occurring CD4+CD25+ regulatory T cells (Tregs) are essential for the active suppression of autoimmunity. The present study was designed to investigate whether Tregs express TAM receptors and the potential role of Gas6-TAM signal in regulating the suppressive function of Tregs. Methods. The protein and mRNA levels of TAM receptors were determined by using Western blot, immunofluorescence, flow cytometry, and RT-PCR. Then, TAM receptors were silenced using targeted siRNA or blocked with specific antibody. The suppressive function of Tregs was assessed by using a CFSE-based T cell proliferation assay. Flow cytometry was used to determine the expression of Foxp3 and CTLA4 whereas cytokines secretion levels were measured by ELISA assay. Results. Tregs express both Axl and Mertk receptors. Gas6 increases the suppressive function of Tregs in vitro and in mice. Both Foxp3 and CTLA-4 expression on Tregs are enhanced after Gas6 stimulation. Gas6 enhances the suppressive activity of Tregs mainly through Axl receptor. Conclusion. Gas6 has a direct effect on the functions of CD4+CD25+Tregs mainly through its interaction with Axl receptor.

Design, synthesis and biological evaluation of novel benzo-α-pyrone containing piperazine derivatives as potential BRAFV600E inhibitors.

The increasingly acquired resistance to vemurafenib and side effects of known inhibitors motivate the search for new and more effective anti-melanoma drugs. In this Letter, virtual screening and scaffold growth were combined together to achieve new molecules as BRAFV600E inhibitors. Along with docking simulation, a primary screen in vitro was performed to filter the modifications for the lead compound, which was then substituted, synthesized and evaluated for their inhibitory activity against BRAFV600E and several melanoma cell lines. Out of the obtained compounds, derivative 3l was identified as a potent BRAFV600E inhibitor and exerted an anticancer effect through BRAFV600E inhibition. The following biological evaluation assays confirmed that 3l could induce cell apoptosis and marked DNA fragmentation. Furthermore, 3l could arrest the cell cycle at the G0/G1 phase in melanoma cells. The docking simulation displayed that 3l could tightly bind with the crystal structure of BRAFV600E at the active site. Overall, the biological profile of 3l suggests that this compound may be developed as a potential anticancer agent.

Metronidazole containing pyrazole derivatives potently inhibit tyrosyl-tRNA synthetase: design, synthesis, and biological evaluation.

As an important enzyme in bacterial protein biosynthesis, tyrosyl-tRNA synthetase (TyrRS) has been an absorbing therapeutic target for exploring novel antibacterial agents. A series of metronidazole-based antibacterial agents has been synthesized and identified as TyrRS inhibitors with low cytotoxicity and significant antibacterial activity, especially against Gram-negative organisms. Of the compounds obtained, 4f is the most potent agent which inhibited the growth of Pseudomonas aeruginosa ATCC 13525 (MIC = 0.98 µg/mL) and exhibited TryRS inhibitory activity (IC50  = 0.92 µm). Docking simulation was performed to further understand its potency. Membrane-mediated apoptosis in P. aeruginosa was verified by flow cytometry.

Telomerase inhibitors: a patent review (2010-2015).

INTRODUCTION: Telomerase is a ribonucleoprotein that catalyses the addition of telomeric repeat sequences (having the sequence 5'-TTAGGG-3' in humans) to the ends of chromosomes. Telomerase activity is detected in most types of human tumours, but it is almost undetectable in normal somatic cells. Therefore, telomerase is a promising therapeutic target. To date, the known inhibitors of telomerase include nucleoside analogues, oligonucleotides and G-quadruplex stabilizers. This review highlights recent advances in our understanding of telomerase inhibitors, the relationships between telomerase inhibitors, cancer, and fields such as inflammation. AREAS COVERED: This review summarizes new patents published on telomerase inhibitors from 2010 to 2015. EXPERT OPINION: The review provides a brief account of the background, development, and on-going issues involving telomerase inhibitors. In particular, this review emphasizes imetelstat (GRN163L) and some typical G-quadruplex stabilizers that participate in telomerase inhibition. Overall, the research scope of antineoplastic is becoming broader and telomerase inhibitors have been shown to be a promising therapeutic target. Therefore, novel antineoplastic agents with greater activity and higher specificity must be developed.

Protective Effects of Growth Arrest-Specific Protein 6 (Gas6) on Sepsis-Induced Acute Kidney Injury.

Acute kidney injury (AKI) is a serious complication of sepsis, which has a high mortality rate. Growth arrest-specific protein 6 (Gas6), the protein product of the growth arrest specific gene 6, has been shown to have an anti-apoptotic effect as well as pro-survival capability. Here, we investigated the effects of Gas6 on sepsis-associated AKI in mice subjected to cecal ligation and puncture (CLP). We found that the administration of rmGas6 significantly reduced serum urea nitrogen and creatinine and improved the survival of septic mice. Furthermore, the renal pathological damage induced by CLP was attenuated by rmGas6 treatment. Finally, rmGas6 reduced the renal tissue apoptotic index and the expression of Bax, while it upregulated the expression of Bcl-2. The data suggest that rmGas6 might be used as a potential therapeutic agent for sepsis-induced AKI.

Design, synthesis and biological evaluation of pyrazolyl-nitroimidazole derivatives as potential EGFR/HER-2 kinase inhibitors.

A series of novel pyrazole-nitroimidazole derivatives had been arranged and evaluated for their EGFR/HER-2 tyrosine kinase inhibitory activity as well as their antiproliferative properties on four kinds of cancer cell lines (MCF-7, Hela, HepG2, B16-F10). The bioassay results showed most of the designed compounds exhibited potential antiproliferation activity, with the IC50 values ranging from 0.13µM to 128.06µM in four tumor cell lines. Among them, compound 5c exhibited remarkable inhibitory activity against EGFR/HER-2 tyrosine kinase with IC50 value of 0.26µM/0.51µM, respectively, comparable to the positive control erlotinib (IC50=0.41µM for HER-2 and IC50=0.20µM for EGFR) and lapatinib (IC50=0.54µM for HER-2 and IC50=0.28µM for EGFR). Molecular modeling simulation studies were performed in order to predict the biological activity of the proposed compounds and activity relationship (SAR) of these pyrazole-nitroimidazole derivatives.

Sulfonamide derivatives containing dihydropyrazole moieties selectively and potently inhibit MMP-2/MMP-9: Design, synthesis, inhibitory activity and 3D-QSAR analysis.

New series of sulfonamide derivatives containing a dihydropyrazole moieties inhibitors of MMP-2/MMP-9 were discovered using structure-based drug design. Synthesis, antitumor activity, structure-activity relationship and optimization of physicochemical properties were described. In vitro the bioassay results revealed that most target compounds showed potent inhibitory activity in the enzymatic and cellular assays. Among the compounds, compound 3i exhibited the most potent inhibitory activity with IC50 values of 0.21 µM inhibiting MMP-2 and 1.87 µM inhibiting MMP-9, comparable to the control positive compound CMT-1 (1.26 µM, 2.52 µM). Docking simulation was performed to position compound 3i into the MMP-2 active site to determine the probable binding pose. Docking simulation was further performed to position compound 3i into the MMP-2 active site to determine the probable binding model the 3D-QSAR models were built for reasonable design of MMP-2/MMP-9 inhibitors at present and in future.

Design, synthesis, biological evaluation and molecular modeling of dihydropyrazole sulfonamide derivatives as potential COX-1/COX-2 inhibitors.

Novel dihydropyrazole sulfonamide derivatives (30-56) were designed, synthesized, and evaluated for their biological activities as COX-1 and COX-2 inhibitors. In vitro biological evaluation against three human tumor cell lines revealed that most target compounds showed antiproliferative activities. Among the compounds, compound 48 exhibited the most potent and selective COX-2 inhibitor (COX-2 IC50=0.33 µM; COX-1 IC50=68.49 µM) relative to the reference drugs celecoxib (IC50=0.052 µM). Docking simulation was performed to position compound 48 into the COX-2 active site and the result showed that compound 48 could bind well at the COX-2 active site and it indicated that compound 48 could be a potent and selective COX-2 inhibitor.

[Application of the wavelet transform in terahertz time-domain spectroscopy].

With the development of ultrafast laser technique and deeper understanding of the terahertz (THz)electromagnetic wave, THz time domain spectroscopy (THz-TDS)has become a novel and rapidly developed technique. It has been applied in many fields such as biomedical diagnostics, semiconductor wafer diagnostics, and inspection of artwork However, the achieved spectrum will exhibit strong oscillations when it is measured in air environment due to the absorption of water vapor. In the present paper, spectra in the range of 0.2-1.9 THz for seven kinds of samples were measured by THz-TDs in both nitrogen and air environment. The wavelet transform was used to process the data measured in the air environment to eliminate the influence of water vapor. The feasibility of this method was demonstrated by the experimental results.