Comprehensive analysis of miRNAs-lncRNAs-mRNAs modules and ceRNA network in acute liver failure: Hsa-miR3175 and C-reactive protein determination.

Acute liver failure (ALF), also known as fulminant hepatitis, coagulation disorders, and worsening mental status. It has a poor prognosis and high mortality rate. Among these, the top 10 upregulated genes included GKA-DPA1, IGLL5, PLA2G7, CCL5, IGLJ, GUSBP11, RHOBT1, IGLL3P, CCL18, and ADRBK2. On the other hand, the top 10 downregulated genes were SLC6A1, PID1, AVPR1A, PP1R1A, ST3GAL6, TPST, ERO1LB, SLCO4C1, and KLF15. Furthermore, the DEGs were found to be enriched in processes related to LIAO metastasis and creighton endocrine therapy resistance. To explore the interactions among the DEGs, we constructed a PPI network. This network revealed 16 hub genes that play crucial roles in ALF pathogenesis. Within this network, hsa-mir-375 and hsa-mir-650 were identified as central nodes, indicating their potential importance in ALF. By identifying and analyzing the transcriptional-level ceRNA network, we have provided valuable insights into the etiology of ALF.

Modified Respiratory Rate Oxygenation Index: An Early Warning Index for the Need of Intubation in COVID-19 Patients with High-Flow Nasal Cannula Therapy.

BACKGROUND: High-flow nasal cannula oxygen therapy (HFNC) is recommended for patients with COVID-19. However, the increasing use of HFNC brings a risk of delayed intubation. The optimal timing of switching from HFNC to invasive mechanical ventilation (IMV) remains unclear. An effective predictor is needed to assist in deciding on the timing of intubation. Respiratory rate and oxygenation (ROX) index, defined as (SpO2/FiO2) / respiratory rate, has already shown good diagnostic accuracy. Modified ROX (mROX) index, defined as (PaO2 /FiO2) / respiratory rate, might be better than the ROX index in predicting HFNC failure. OBJECTIVE: The aim was to evaluate the predictive value of mROX for HFNC failure in patients with COVID-19. METHODS: Severe or critical patients with COVID-19 treated with HFNC were enrolled in two clinical centers. Laboratory indicators, respiratory parameters, and mROX index at 0 h and 2 h after initial HFNC were collected. Based on the need for IMV after HFNC initiation, the patients were divided into an HFNC failure group and an HFNC success group. The predictive value of mROX index for IMV was evaluated by the area under the receiver operating characteristic curve (AUROC) and logistic regression analysis. We performed Kaplan-Meier survival analysis using the log-rank test. RESULTS: Sixty patients with COVID-19 (mean ± SD age, 62.8 ± 14.1 years; 42 patients were male) receiving HFNC were evaluated, including 18 critical and 42 severe cases. A total of 33 patients had hypertension; 14 had diabetes; 17 had chronic cardiac disease; 11 had chronic lung disease; 13 had chronic kidney disease; and 17 had a history of stroke. The AUROC of mROX index at 2 h was superior to that of other respiratory parameters to predict the need for IMV (0.959; p < 0.001). At the mROX index cutoff point of 4.45, predicting HFNC failure reached the optimal threshold, with specificity of 94% and sensitivity of 92%. Logistic regression analysis showed that 2-h mROX index < 4.45 was a protective factor for IMV (odd radio 0.18; 95% CI 0.05-0.64; p = 0.008). In the HFNC failure group, the median time from HFNC to IMV was 22.5 h. The 28-day mortality of the late intubation patients (≥ 22.5 h) was higher than that of the early intubation patients (< 22.5 h) (53.8% vs. 8.3%; p = 0.023). CONCLUSIONS: mROX at 2 h is a good early warning index of the need for IMV in patients with COVID-19 after HFNC initiation. Early intubation may lead to better survival in patients with 2-h mROX index < 4.45.

Creation and Reaction of Solvated Electrons at and near the Surface of Water.

Solvated electrons (es-) are among nature's most powerful reactants, with over 2600 reactions investigated in bulk water. These electrons can also be created at and near the surface of water by exposing an aqueous microjet in vacuum to gas-phase sodium atoms, which ionize into es- and Na+ within the top few layers. When a reactive surfactant is added to the jet, the surfactant and es- become coreactants localized in the interfacial region. We report the reaction of es- with the surfactant benzyltrimethylammonium in a 6.7 M LiBr/water microjet at 235 K and pH = 2. The reaction intermediates trimethylamine (TMA) and benzyl radical are identified by mass spectrometry after they evaporate from solution into the gas phase. Their detection demonstrates that TMA can escape before it is protonated and benzyl before it combines with itself or a H atom. Diffusion-reaction calculations indicate that es- reacts on average within 20 Å of the surface and perhaps within the surfactant monolayer itself, while unprotonated TMA evaporates from the top 40 Å. The escape depth exceeds 1300 Å for the more slowly reacting benzyl radical. These proof-of-principle experiments establish an approach for exploring the near-interfacial analogues of aqueous bulk-phase radical chemistry through the evaporation of reaction intermediates into the gas phase.

Using Postoperative Coagulation Parameters Within 24 h After Liver Transplantation to Predict Incidence of Stage 3 Acute Kidney Injury (AKI).

BACKGROUND This study aimed to investigate the relationship between coagulation function and the incidence of acute kidney injury (AKI) stage 3 within 24 h after liver transplantation (LT) and explore the predictive value of coagulation parameters for post-LT stage 3 AKI. MATERIAL AND METHODS A retrospective study was conducted on 241 patients who underwent LT at the Renji Hospital affiliated with Shanghai Jiao Tong University School of Medicine between February 2021 and February 2022. The coagulation parameters within 24 h after LT and the incidence of post-LT AKI within 7 days were recorded. The correlation between post-LT coagulation function and post-LT stage 3 AKI was determined using binary logistic regression analysis. RESULTS Post-LT AKI occurred in 99 cases (41.1%), 28 (28.3%) of which developed AKI stage 3. In univariate logistic regression analysis, multiple coagulation indexes of the AKI stage 3 group were worse than in the AKI stage 0-2 group. In multivariate logistic regression analysis, lower post-LT ADP-induced PLT aggregation rate (cut-off: 15.75%), higher D-dimer level (cut-off: 3.52 ug/ml), and prolonged R-value (cut-off: 7.5 min) within 24 h were independent risk factors for post-LT AKI stage 3. The AUROC value for predicting the incidence of post-LT AKI stage 3 combining the 3 indices was 0.835 (sensitivity: 83.3%, specificity: 76.9%). The decision curve showed that combining D-dimer, R-value, and ADP-induced PLT aggregation rate yielded the highest net benefit for predicting the incidence of stage 3 AKI. CONCLUSIONS Post-LT coagulation function within 24 h correlated with the incidence of post-LT AKI stage 3. Lower ADP-induced PLT aggregation rate, higher D-dimer level, and prolonged R-value from the TEG were independent risk factors for the incidence of post-LT AKI stage 3.

d-Amino Acid-Based Metabolic Labeling Enables a Fast Antibiotic Susceptibility Test of Both Isolated Bacteria and Bronchoalveolar Lavage Fluid.

The threat of multidrug-resistant bacteria has escalated rapidly, increasing the demand for accurate antibiotic susceptibility tests (ASTs). Traditional bacterial growth yield-based ASTs often take overnight to report, delaying the timely guidance of antibiotic use. Here, a fluorescent d-amino acid (FDAA) labeling-based AST (FaAST) is reported, which can quickly provide accurate minimum inhibitory concentrations (MICs). The FDAA-labeling signals that reflect the bacterial metabolic status underlie the flow cytometry-based strategy for MIC determination. Resistant bacteria show a reluctant decline in FDAA-labeling (inhibited metabolism) after treatment with the corresponding antibiotics, whereas susceptible bacteria demonstrate quick responses to low doses of drugs. The MICs are determined based on the changing trends in labeling. After testing 23 clinical isolates and laboratory strains of the most critical drug-resistant bacteria against a panel of representative antibiotics, FaAST shows a high susceptibility category with an accuracy of 98.13%. Moreover, FaAST can also make quick and accurate diagnosis against bronchoalveolar lavage fluids collected from hospital-acquired pneumonia patients, saving 2-4 days in guiding antibiotic use for this life-threatening infection. Thus, the speed, accuracy, and broad applicability of FaAST will be valuable in informing antibiotic decisions when treating critical infections caused by drug-resistant bacteria.

Global trends and hotspots in research of carbapenem-resistant Enterobacteriaceae (CRE): a bibliometric analysis from 2010 to 2020.

BACKGROUND: Infectious disease caused by carbapenem-resistant Enterobacteriaceae (CRE) has become one of the most serious challenges due to its high morbidity and mortality and research on it has aroused great concern worldwide in the last decade. Thus, a bibliometric analysis of relevant publications is needed to identify the situation of current investigations and prioritize the future research areas. METHODS: The current study retrieved articles related to CRE published between 2010 and 2020 from the Web of Science core collection database. The search strategy syntax included "carbapenem-resistant Enterobacteriaceae", "carbapenem-resistant Klebsiella pneumoniae", "carbapenemase producing Enterobacteriaceae" and "carbapenemase producing Klebsiella pneumoniae" which were searched in both Medical Subject Headings (MeSH) and titles. A bibliometric analysis was conducted using VOSviewer, Bibliographic Item Co-Occurrence Matrix Builder, gCLUTO and other machine learning tools. Key words, subject terms, contributions as well as collaborations were assessed. Moreover, hot off the press and future research trends were demonstrated. RESULTS: A total of 1,671 publications on CRE were finally included in the bibliometric analysis and 5 related theme clusters were identified which mainly focused on epidemiology, resistance mechanisms, antibiotics treatment and infection control. A total of 142 keywords occurred more than 5 times and the most frequent keyword was "carbapenem-resistant Enterobacteriaceae" with 247 occurrences and a total link strength of 559. The output on CRE has gradually increased during the last decade, and the USA has made the greatest contribution due to the 533 research papers. Agents that act against CRE, especially ceftazidime-avibactam (occurrences, 85; average publication year, 2018.26), and the early detection of CRE by genome sequencing techniques (occurrences, 97; average publication year, 2017.94) were emerging hot topics which would probably attract future research interest. CONCLUSIONS: The bibliometric analysis revealed that development of antibacterial agents, early etiological detection and genome sequencing techniques were the hotspots and would probably direct the future research directions which would also facilitate a better understanding of the epidemiology of drug-resistant bacteria and implementing the antibiotic stewardship program.

Competing Segregation of Br- and Cl- to a Surface Coated with a Cationic Surfactant: Direct Measurements of Ion and Solvent Depth Profiles.

Ion-surface scattering experiments can be used to measure elemental depth profiles on the angstrom scale in complex liquid mixtures. We employ NICISS (neutral impact collision ion scattering spectroscopy) to measure depth profiles of dissolved ions and solvent in liquid glycerol containing the cationic surfactant tetrahexylammonium bromide (THA+/Br-) at 0.013 M and mixtures of NaBr + NaCl at 0.4 M total concentration. The experiments reveal that Br- outcompetes Cl- in its attraction to surface THA+, and that THA+ segregates more extensively when more Br- ions are present. Intriguingly, the depths spanned by THA+, Br-, and Cl- ions generally increase with Br- bulk concentration, expanding from ∼10 to ∼25 Šfor both Br- and Cl- depth profiles. This broadening likely occurs because of an increasing pileup of THA+ ions in a multilayer region that spreads the halide ions over a wider depth. The experiments indicate that cationic surfactants enhance Br- and Cl- concentrations in the surface region far beyond their bulk-phase values, making solutions coated with these surfactants potentially more reactive toward gases that can oxidize the halide ions.

Management of Bivalirudin Anticoagulation Therapy for Extracorporeal Membrane Oxygenation in Heparin-Induced Thrombocytopenia: A Case Report and a Systematic Review.

Extracorporeal membrane oxygenation (ECMO) can provide respiratory and cardiac support to patients in reversible devastated conditions. Heparin is the mainstay for anticoagulation during ECMO. Bivalirudin, a direct thrombin blocker, may represent an effective alternative for patients suffering from heparin-induced thrombocytopenia (HIT). We present the first case of a Chinese patient who experienced HIT and received bivalirudin anticoagulation during ECMO. In addition, we present a systematic review for this topic. We searched PubMed, EMBASE, and Cochrane Library (up to April 20, 2020) for studies that included patients undergoing ECMO, presenting with HIT, requiring bivalirudin treatment, and reporting relevant outcomes. The literature review yielded 15 studies involving 123 patients, amongst whom 58 patients were confirmed or suspected HIT patients, and 76 patients received bivalirudin as an anticoagulant for ECMO. Twelve studies were included for quantitative synthesis, and 46 patients were retrieved. The mean age of these patients was 46 years, and 30 patients were males. The average maintenance rate of bivalirudin was 0.27 ± 0.37 mg/kg/h, in order to maintain a target of activated clotting time (ACT) of 160-220 s. Additionally, bivalirudin doses in patients with continuous renal replacement therapies (CRRT) and patients without CRRT were 0.15 ± 0.06 mg/kg/h vs 0.28 ± 0.36 mg/kg/h, respectively (p=0.15). Most of the patients with confirmed HIT improved platelet counts in 3.3 ± 2.8 days after switching to bivalirudin anticoagulation. The patient-level data showed that 29 cases survived, 1 reported major bleeding, and 4 reported thrombotic events. Bivalirudin might be a promising optimal choice for ECMO anticoagulation in patients with HIT. A tailored protocol for management of bivalirudin treatment during ECMO should be developed with caution. Further prospective studies are necessary to standardise the use of bivalirudin. SYSTEMATIC REVIEW REGISTRATION: PROSPERO, identifier CRD42020160907.

Experimental Depth Profiles of Surfactants, Ions, and Solvent at the Angstrom Scale: Studies of Cationic and Anionic Surfactants and Their Salting Out.

Neutral impact ion scattering spectroscopy (NICISS) is used to measure the depth profiles of ionic surfactants, counterions, and solvent molecules on the angstrom scale. The chosen surfactants are 0.010 m tetrahexylammonium bromide (THA+/Br-) and 0.0050 m sodium dodecyl sulfate (Na+/DS-) in the absence and presence of 0.30 m NaBr in liquid glycerol. NICISS determines the depth profiles of the elements C, O, Na, S, and Br through the loss in energy of 5 keV He atoms that travel into and out of the liquid, which is then converted into depth. In the absence of NaBr, we find that THA+ and its Br- counterion segregate together because of charge attraction, forming a narrow double layer that is 10 Šwide and 150 times more concentrated than in the bulk. With the addition of NaBr, THA+ is "salted out" to the surface, increasing the interfacial Br- concentration by 3-fold and spreading the anions over a ∼30 Šdepth. Added NaBr similarly increases the interfacial concentration of DS- ions and broadens their positions. Conversely, the dissolved Br- ions are significantly depleted over a depth of 0-40 Šfrom the surface because of charge repulsion from DS- ions within the interfacial region. These different interfacial Br- propensities correlate with previously measured gas-liquid reactivities: gaseous Cl2 readily reacts with Br- ions in the presence of THA+ but drops 70-fold in the presence of DS-, demonstrating that surfactant headgroup charge controls the reactivity of Br- through changes in its depth profile.

Treatment with Recombinant Interleukin-15 (IL-15) Increases the Number of T Cells and Natural Killer (NK) Cells and Levels of Interferon-γ (IFN-γ) in a Rat Model of Sepsis.

BACKGROUND This study aimed to investigate the effects of treatment with recombinant interleukin-15 (IL-15) on T cells, natural killer (NK) cells, and interferon-γ (IFN-γ) on the immune response in a rat cecal ligation and perforation model of sepsis. MATERIAL AND METHODS Sprague-Dawley rats (n=120) were divided into four groups (n=30). A rat model of clinical sepsis was created using cecal ligation and perforation, and 109 rats successfully developed sepsis. Rats were then injected intraperitoneally with 0.5, 1.0, and 1.5 µg of recombinant rat IL-15 or saline. Survival was determined, and the numbers of T cells and NK cells, and the expression levels of IL-15 and IFN-γ were detected in the peripheral blood of rats in each group at 24 h and 48 h. RESULTS The levels of IL-15 and IFN-γ, as well as the numbers of T cells and NK cells, were significantly increased in the IL-15-treated groups compared with the control group at both 24 h and 48 h (P<0.05). Levels of IL-15 and IFN-γ were significantly increased in the IL-15-treated groups at 48 h compared with 24 h in the control group. Levels of IL-15, the numbers of T cells and NK cells, and the levels of IFN-γ in peripheral blood were significantly lower at 48 h when compared with 24 h (P<0.05). CONCLUSIONS In a rat model of sepsis, treatment with recombinant IL-15 significantly increased T cell and NK cell numbers, and levels of IFN-γ, and prolonged the survival of rats with sepsis.

The efficacy of extracorporeal membrane oxygenation in liver transplantation from non-heart-beating donors: A systemic review and meta-analysis.

BACKGROUND: A systematic review and meta-analysis was made to see whether extracorporeal membrane oxygenation (ECMO) in liver transplantation could improve non-heart-beating donors (NHBDs) recipients' outcomes compared with donors after brain death (DBDs) recipients. METHODS: We searched MEDLINE, EMBASE, and Cochrane Central Register of Controlled Trials for eligible studies. The study eligible criteria are cohort or case-control studies using ECMO in all NHBDs; studies involved a comparison group of DBDs; and studies evaluated 1-year graft and patient survival rate in NHBDs and DBDs groups. RESULTS: Four studies with 704 patients fulfilled the inclusion criteria. The pooled odds ratio (OR) of 1-year patient survival rate in NHBDs recipients compared with DBDs recipients was 0.8 (95% confidence interval [CI], 0.41-1.55). The pooled OR of 1-year graft survival rate in NHBDs recipients compared with DBDs recipients was 0.46 (95% CI, 0.26-0.81). NHBDs recipients were at greater risks to the occurrence of primary nonfunction (PNF) (OR = 7.12, 95% CI, 1.84-27.52) and ischemic cholangiopathy (IC) (OR = 9.46, 95% CI, 2.76-32.4) than DBDs recipients. CONCLUSIONS: ECMO makes 1-year patient survival acceptable in NHBDs recipients. One-year graft survival rate was lower in NHBDs recipients than in DBDs recipients. Compared with DBDs recipients, the risks to develop PNF and IC were increased among NHBDs recipients.

P27 Protects Cardiomyocytes from Sepsis via Activation of Autophagy and Inhibition of Apoptosis.

BACKGROUND It has been reported that p27Kip1 plays an important role not only in the inhibition of cyclin-dependent kinases but also in the regulation of autophagy under various metabolically related stress conditions, including glucose deprivation and endoplasmic reticulum stress. However, its effect on lipopolysaccharide (LPS)-induced cardiomyocyte stress in vitro remains unclear. Here, we measured the increased expression of LC3-II and visualized autophagosomes in vitro by immunofluorescent assays after treatment with a p27 fusion protein. MATERIAL AND METHODS Cardiomyocyte contractile properties were assessed by measuring cell shortening and re-lengthening. Apoptosis was evaluated by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. Western blot, colorectal ligation puncture (CLP) surgery, silencing of Atg5 expression by small interfering RNA (siRNA), and immunofluorescent assays were also performed in this study. RESULTS After exogenous delivery of the p27 fusion protein and overexpression of p27 in LPS-induced cardiomyocytes, we found lower expressions of caspase-3 and caspase-8 and reduced positive TUNEL staining. Improved cardiomyocyte mechanical functions and reduced apoptosis were diminished after treatment with various autophagy inhibitors. Intravenous injections of p27-expressing adeno-associated virus serotype 9 (AAV9) vectors resulted in cardiac specific overexpression of p27, and echocardiography was used to assess cardiac function and structure in sepsis rat models. We observed improved cardiac function and reversed adverse ventricular remolding after the introduction of AAV9 vectors. Meanwhile, apoptosis was reduced, and expression of LC3-II was elevated in septic rat models treated with AAV9 vectors compared to controls. CONCLUSIONS The study data demonstrated that the overexpression of p27 protects cardiomyocytes from sepsis-induced cardiac depression via the activation of autophagy and inhibition of apoptosis.

Control of Interfacial Cl2 and N2O5 Reactivity by a Zwitterionic Phospholipid in Comparison with Ionic and Uncharged Surfactants.

Gas-liquid scattering experiments reveal that charge-separated but neutral (zwitterionic) surfactants catalyze the oxidation of dissolved Br- to Br2 by gaseous Cl2 at the surface of a 0.3 M NaBr/glycerol solution. Solutions of NaBr dissolved in glycerol with no surfactant were compared with solutions coated with zwitterionic, cationic, and anionic surfactants at dilute surface concentrations of 1.1 to 1.5 × 1014 cm-2 (less than 65% of maximum chain packing). The zwitterionic phospholipid enhances Cl2 conversion of Br- to Br2 by a factor of 1.61 ± 0.15, in comparison with a 14-fold enhancement by a cationic surfactant (tetrahexylammonium) and a five-fold suppression by an anionic surfactant (dodecyl sulfate). Further studies indicate that even an uncharged surfactant, monododecanoylglycerol, enhances Cl2 → Br2 production. Similar behavior is observed for the oxidation of Br- to Br2 by N2O5; it is just slightly suppressed by the phospholipid and strongly enhanced by the cationic surfactant. Collectively, these results suggest that attractions and repulsions between the negative Br- ions and the positive and negative charges of the surfactant headgroups draw Br- ions to the surface or repel them away. At low coverages, ion-induced dipole and dispersion interactions between the CH2 groups and Br- or Cl2 may also enhance reactivity. These results demonstrate that the hydrocarbon chains of loosely packed surfactants do not necessarily block gas-liquid reactions but that positively charged, and even uncharged, groups can instead facilitate reactions by bringing gas-phase and solution-phase reagents together in the interfacial region.

Evaluation of the efficacy and safety of ceftazidime/avibactam in the treatment of Gram-negative bacterial infections: a systematic review and meta-analysis.

Data on the efficacy and safety of ceftazidime/avibactam (CAZ-AVI) are limited. A systematic review and meta-analysis was conducted to clarify the role of CAZ-AVI for patients with serious Gram-negative bacterial infections. The PubMed, EMBASE and Cochrane Library databases were searched for randomised controlled trials (RCTs) and cohort studies involving CAZ-AVI. Summary risk ratios (RRs) and 95% confidence intervals (CIs) were calculated using a fixed- or random-effects model. Twelve articles (4951 patients) were included, consisting of nine RCTs and three observational studies comparing CAZ-AVI with other regimens, e.g. carbapenems or colistin. CAZ-AVI showed a comparable clinical response (RR = 0.99, 95% CI 0.96-1.02; I2 = 0%) and non-inferior bacterial eradication (RR = 1.04, 95% CI 0.93-1.17; I2 = 79.1%) to carbapenems. No significant difference was detected between groups regarding mortality and adverse events. Moreover, subgroup analyses demonstrated that CAZ-AVI improved the clinical response (RR = 1.61, 95% CI 1.13-2.29) with reduced mortality (RR = 0.29, 95% CI 0.13-0.63) in patients infected by carbapenem-resistant Enterobacteriaceae versus comparators. Likewise, CAZ-AVI improved the clinical cure rate of bloodstream infections (RR = 2.11, 95% CI 1.54-2.88). An improved ability of CAZ-AVI in microbiological eradication was also detected in patients with complicated urinary tract infections (RR = 1.13, 95% CI 1.05-1.21). CAZ-AVI exhibited comparable efficacy and safety with carbapenems. Therefore, this agent might be a potential powerful agent for patients with serious Gram-negative bacterial infections.

When CMRP met alkyl vinyl ketone: visible light induced living radical polymerization (LRP) of ethyl vinyl ketone (EVK).

Photo responsive polyethyl vinyl ketone (PEVK) and various block copolymers were prepared through the visible light induced cobalt mediated radical polymerization (CMRP) process. The first order kinetics and linear increase in molecular weight with conversion demonstrated a well-behaved system. Chain extension experiments showed the retention of the living end of PEVK.

HDAC is essential for epigenetic regulation of Thy-1 gene expression during LPS/TLR4-mediated proliferation of lung fibroblasts.

Lipopolysaccharide (LPS)-induced proliferation of lung fibroblasts is closely correlated with loss of gene expression of thymocyte differentiation antigen-1 (Thy-1), accompanied with deacetylation of histones H3 and H4 at the Thy-1 gene promoter region; however, the mechanism remains enigmatic. We report here that LPS downregulates Thy-1 gene expression by activating histone deacetylases (HDACs) via Toll-like receptor 4 (TLR4) signaling. Treatment of primary cultured mouse lung fibroblasts with LPS resulted in significant upregulation of TLR4 and enhanced cell proliferation that was abolished by silencing TLR4 with lentivirus-delivered TLR4 shRNA. Interestingly, LPS increased the mRNA and protein levels of HDAC-4, -5, and -7, an effect that was abrogated by HDAC inhibitor trichostatin A (TSA) or TLR4-shRNA-lentivirus. Consistent with these findings, Ace-H3 and Ace-H4 were decreased by LPS that was prevented by TSA. Most importantly, chromosome immunoprecipitation (ChIP) analysis demonstrated that LPS decreased the association of Ace-H4 at the Thy-1 promoter region that was efficiently restored by pretreatment with TSA. Accordingly, LPS decreased the mRNA and protein levels of Thy-1 that was inhibited by TSA. Furthermore, silencing the Thy-1 gene by lentivirus-delivered Thy-1 shRNA could promote lung fibroblast proliferation, even in the absence of LPS. Conversely, overexpressing Thy-1 gene could inhibit lung fibroblast proliferation and reduce LPS-induced lung fibroblast proliferation. Our data suggest that LPS upregulates and activates HDACs through TLR4, resulting in deacetylation of histones H3 and H4 at the Thy-1 gene promoter that may contribute to Thy-1 gene silencing and lung fibroblast proliferation.

High-mobility group box 1 accelerates lipopolysaccharide-induced lung fibroblast proliferation in vitro: involvement of the NF-κB signaling pathway.

The mechanism underlying lipopolysaccharide (LPS)-induced aberrant proliferation of lung fibroblasts in Gram-negative bacilli-associated pulmonary fibrosis is unknown. High-mobility group box 1 (HMGB1) is a ubiquitous nuclear protein that is released from the nuclei of lung fibroblasts after LPS stimulation. It can exasperate LPS-induced inflammation and hasten cell proliferation. Thus, this study investigated the effects of LPS- and/or HMGB1-stimulating murine lung fibroblasts on gene expression using various assays in vitro. Thiazolyl-diphenyl-tetrazolium bromide (MTT) assay data showed that either LPS or HMGB1 could induce lung fibroblast proliferation. Endogenous HMGB1 secreted from lung fibroblasts was detected by enzyme-linked immunosorbent assay (ELISA) 48 h after LPS stimulation. Pretreatment with an anti-HMGB1 antibody inhibited the proliferative effects of LPS on lung fibroblasts. DNA microarray data showed that the NF-κB signaling genes were upregulated in cells after stimulated with LPS, HMGB1, or both. Secretion of matrix metalloproteinase (MMP)-2 and MMP-9, and tissue inhibitor of metalloproteinase 2 (TIMP-2) was significantly upregulated after treatment with LPS, HMGB1, or their combination. However, an NF-κB inhibitor was able to downregulate levels of these proteins. In addition, levels of Toll-like receptor 4 (TLR4), Toll-like receptor 2 (TLR2), and receptors for advanced glycation end products (RAGE) mRNA and proteins were also upregulated in these cells after LPS treatment and further upregulated by LPS plus HMGB1. In conclusion, the data from the current study demonstrate that LPS-induced lung fibroblast secretion of endogenous HMGB1 can augment the proproliferative effects of LPS and, therefore, may play a key role in exacerbation of pulmonary fibrosis. The underlying molecular mechanisms are related to the activation of the TLR4/NF-κB signaling pathway and its downstream targets.

Overexpression of PTEN suppresses lipopolysaccharide-induced lung fibroblast proliferation, differentiation and collagen secretion through inhibition of the PI3-K-Akt-GSK3beta pathway.

BACKGROUND: Abnormal and uncontrolled proliferation of lung fibroblasts may contribute to pulmonary fibrosis. Lipopolysaccharide (LPS) can induce fibroblast proliferation and differentiation through activation of phosphoinositide3-Kinase (PI3-K) pathway. However, the detail mechanism by which LPS contributes to the development of lung fibrosis is not clearly understood. To investigate the role of phosphatase and tensin homolog (PTEN), a PI3-K pathway suppressor, on LPS-induced lung fibroblast proliferation, differentiation, collagen secretion and activation of PI3-K, we transfected PTEN overexpression lentivirus into cultured mouse lung fibroblasts with or without LPS treatment to evaluate proliferation by MTT and Flow cytometry assays. Expression of PTEN, alpha-smooth muscle actin (alpha-SMA), glycogen synthase kinase 3 beta (GSK3beta) and phosphorylation of Akt were determined by Western-blot or real-time RT-PCR assays. The PTEN phosphorylation activity was measured by a malachite green-based assay. The content of C-terminal propeptide of type I procollagen (PICP) in cell culture supernatants was examined by ELISA. RESULTS: We found that overexpression of PTEN effectively increased expression and phosphatase activity of PTEN, and concomitantly inhibited LPS-induced fibroblast proliferation, differentiation and collagen secretion. Phosphorylation of Akt and GSK3beta protein expression levels in the LPS-induced PTEN overexpression transfected cells were significantly lower than those in the LPS-induced non-transfected cells, which can be reversed by the PTEN inhibitor, bpV(phen). CONCLUSIONS: Collectively, our results show that overexpression and induced phosphatase activity of PTEN inhibits LPS-induced lung fibroblast proliferation, differentiation and collagen secretion through inactivation of PI3-K-Akt-GSK3beta signaling pathways, which can be abrogated by a selective PTEN inhibitor. Thus, expression and phosphatase activity of PTEN could be a potential therapeutic target for LPS-induced pulmonary fibrosis. Compared with PTEN expression level, phosphatase activity of PTEN is more crucial in affecting lung fibroblast proliferation, differentiation and collagen secretion.

Multistep DNA-templated synthesis using a universal template.

We report a DNA-templated synthesis method that allows construction of the entire DNA-encoded library with a single DNA template. Taking advantage of deoxyinosine's indiscriminate base-pairing property, we designed a "universal template" that is capable of directing chemical reactions with multiple reactant DNAs with different sequences. In combination with other design features including photocleavable linkers and direct encoding by the reactant DNA, we demonstrated the capabilities of the universal template in library synthesis, target selection, and hit decoding. Our method can be generally and straightforwardly applied to prepare a variety of chemically diverse DNA-encoded libraries.

Epigenetic regulation of Thy-1 gene expression by histone modification is involved in lipopolysaccharide-induced lung fibroblast proliferation.

Lipopolysaccharide (LPS)-induced pulmonary fibrosis is characterized by aberrant proliferation and activation of lung fibroblasts. Epigenetic regulation of thymocyte differentiation antigen 1 (Thy-1) is associated with lung fibroblast phenotype transformation that results in aberrant cell proliferation. However, it is not clear whether the epigenetic regulation of Thy-1 expression is required for LPS-induced lung fibroblast proliferation. To address this issue and better understand the relative underlying mechanisms, we used mouse lung fibroblasts as model to observe the changes of Thy-1 expression and histone deacetylation after LPS challenge. The results showed that cellular DNA synthesis, measured by BrdU incorporation, was impacted less in the early stage (24 hrs) after the challenge of LPS, but significantly increased at 48 or 72 hrs after the challenge of LPS. Meanwhile, Thy-1 expression, which was detected by real-time PCR and Western blot, in lung fibroblasts decreased with increased time after LPS challenge and diminished at 72 hrs. We also found that the acetylation of either histone H3 or H4 decreased in the LPS-challenged lung fibroblasts. ChIP assay revealed that the acetylation of histone H4 (Ace-H4) decreased in the Thy-1 promoter region in response to LPS. In addition, all the above changes could be attenuated by depletion of TLR4 gene. Our studies indicate that epigenetic regulation of Thy-1 gene expression by histone modification is involved in LPS-induced lung fibroblast proliferation.