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Haemophilus influenzae serotype b has been the main cause of invasive infections in children, during the prevaccination period. More than 20 years after the introduction of the conjugate vaccine against Hib, HiNT has emerged as the cause of localized infections in children and adults. The main objective of this work is to evaluate the susceptibility and resistance mechanisms of H. influenzae strains from carriers and describe the molecular epidemiology and their clonal relationships by multilocus sequence typing (MLST). Sixty-nine strains from clinical cases and asymptomatic carriers from 2009 to 2019 were analyzed, confirmed as H. influenzae, and serotyped by polymerase chain reaction. The susceptibility to antibiotics was evaluated by E-test strips. Genotyping was performed by MLST. HiNT was the most frequent in all age groups. Resistance to ampicillin, sulfamethoxazole+trimethoprim, and amoxicillin+clavulanic acid was detected, with the production of β-lactamase being the main resistance mechanism. Among 21 HiNT strains with complete allelic MLST profiles, 19 new sequence types were described, reinforcing the already reported heterogeneity of nontypeable strains, and only one clonal complex (cc-1355) was observed. Our results show a high percentage of colonization regardless of age, increased antimicrobial resistance, and high genetic diversity, along with an increased number of cases caused by HiNT strains. These findings reinforce the need for continuous surveillance for HiNT strains as it has been reported worldwide after the introduction of the Hib conjugate vaccine.

Ethnopharmacological Relevance: Studies have shown interest in nutraceuticals for the prevention of liver diseases. Methoxyeugenol, is a molecule found in foods, such as nutmeg (Myristica fragrans Houtt.) and Brazilian red propolis. These two sources of methoxyeugenol, propolis and nutmeg, are used in folk medicine for the treatment of hepatic and gastrointestinal disorders, although little is known about their effects on the prevention of liver fibrosis. Natural PPAR (Peroxisome proliferator-activated receptor) agonists would represent unique molecules for therapy, considering the lack of therapeutics to treat liver fibrosis in chronic liver disease. Thus, investigation on new alternatives are necessary, including the search for natural compounds from renewable and sustainable sources. Liver fibrosis is a pathological process characterized by an exacerbated cicatricial response in the hepatic tissue, which compromises liver function. Therefore, inhibition of HSC (hepatic stellate cell) activation and hepatocyte damage are considered major strategies for the development of new anti-fibrotic treatments., Aim of the Study: This study aimed to investigate the effects of methoxyeugenol treatment on HSC phenotype modulation in human and murine cells, hepatocyte damage prevention, and protective effects in vivo, in order to evaluate its therapeutic potential for liver fibrosis prevention., Methods: We investigated the effects of methoxyeugenol in (i) in vitro models using human and murine HSC and hepatocytes, and (ii) in vivo models of CCl 4 (carbon tetrachloride) -induced liver fibrosis in mice., Results: We herein report that methoxyeugenol decreases HSC activation through the activation of PPAR-ɣ, ultimately inducing a quiescent phenotype highlighted by an increase in lipid droplets, loss of contraction ability, and a decrease in the proliferative rate and mRNA expression of fibroblast markers. In addition, methoxyeugenol prevented hepatocytes from oxidative stress damage. Moreover, in mice submitted to chronic liver disease through CCl 4 administration, methoxyeugenol decreased the inflammatory profile, liver fibrosis, mRNA expression of fibrotic genes, and the inflammatory pathway signaled by NF-kB (Nuclear factor kappa B)., Conclusion: We propose methoxyeugenol as a novel and potential therapeutic approach to treat chronic liver disease and fibrosis., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Potassium 5-cyano-4-methyl-6-oxo-1,6-dihydropyridine-2-olate (CPBMF65) is a potent inhibitor of the uridine phosphorylase 1 (UPP1) enzyme. Its non-ionized analog has already demonstrated biological properties by reducing adverse effects caused by the chemotherapeutic 5-fluorouracil (5-FU). In addition, it has been demonstrated that uridine inhibits inflammation and fibrosis in bleomycin lung injury, decreasing collagen production. The purpose of this study was to investigate the in vitro and in vivo effects of CPBMF65 on activated hepatic stellate cells (HSC) and on carbon tetrachloride-induced liver fibrosis in mice. After incubation with CPBMF65, decreased cell proliferation and phenotype reversion were observed in vitro. In addition, CPBMF65 promoted a protective effect on tetrachloride-induced liver fibrosis in mice, demonstrated by its antifibrotic and anti-inflammatory actions. The results of the present study indicate that the UPP1 inhibitor (CPBMF65) may have potential as a novel therapeutic agent for the treatment of liver fibrosis., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Octyl gallate (OG) is an antioxidant commonly used in food, although there is no definition of its acceptable daily intake. There are reports in vitro and in vivo showing that food additives and drugs can alter lipid metabolism. Lipid droplet accumulation in hepatic cells is one of the main findings in the unregulated lipid metabolism and is strongly related to the development of nonalcoholic fatty liver disease (NAFLD). In this study, we investigated the effects of OG on lipid metabolism in the hepatocellular carcinoma cell line (HepG2). The results have shown, for the first time, that treatment with OG increased the overall amount of lipids, the triglyceride concentration, the lipid droplet area, and SREBP-1c and PPAR-γ gene expression. Taken together, the findings indicate that OG induces lipid droplet accumulation in HepG2 cells through the regulation of SREBP-1c and PPAR-γ gene expression without involving mTOR/S6K1 and may contribute to NAFLD when used as a food additive., (Copyright © 2020 Lima et al.)

Background & Aims: Cirrhosis and its clinical consequences can be aggravated by bacterial infections, ultimately leading to the development of acute on chronic liver failure (ACLF), characterized by acute decompensation, organ failure, and high mortality within 28 days. Little is known about cellular and molecular mechanisms of ACLF in patients with cirrhosis, so no therapeutic options are available. We developed a sepsis-associated preclinical model of ACLF to facilitate studies of pathogenesis and evaluate the protective effects of simvastatin., Methods: Male Wistar rats inhaled CCl 4 until they developed cirrhosis (at 10 weeks) or cirrhosis with ascites (at 15-16 weeks). Male Sprague-Dawley rats received bile-duct ligation for 28 days or intraperitoneal thioacetamide for 10 weeks to induce cirrhosis. After induction of cirrhosis, some rats received a single injection of lipopolysaccharide (LPS) to induce ACLF; some were given simvastatin or vehicle (control) 4 hours or 24 hours before induction of ACLF. We collected data on changes in hepatic and systemic hemodynamics, hepatic microvascular phenotype and function, and survival times. Liver tissues and plasma were collected and analyzed by immunoblots, quantitative polymerase chain reaction, immuno(fluoro)histochemistry and immunoassays., Results: Administration of LPS aggravated portal hypertension in rats with cirrhosis by increasing the severity of intrahepatic microvascular dysfunction, exacerbating hepatic inflammation, increasing oxidative stress, and recruiting hepatic stellate cells and neutrophils. Rats with cirrhosis given LPS had significantly shorter survival times than rats with cirrhosis given the control. Simvastatin prevented most of ACLF-derived complications and increased survival times. Simvastatin appeared to increase hepatic sinusoidal function and reduce portal hypertension and markers of inflammation and oxidation. The drug significantly reduced levels of transaminases, total bilirubin, and ammonia, as well as LPS-mediated activation of hepatic stellate cells in liver tissues of rats with cirrhosis., Conclusions: In studies of rats with cirrhosis, we found administration of LPS to promote development of ACLF, aggravating the complications of chronic liver disease and decreasing survival times. Simvastatin reduced LPS-induced inflammation and liver damage in rats with ACLF, supporting its use in treatment of patients with advanced chronic liver disease., (Copyright © 2018 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Cells respond to different kind of stress through the coordinated activation of signaling pathways such as MAPK or p53. To find which molecular mechanisms are involved, we need to understand their cell adaptation. The ribosomal protein, S6 kinase 1 (S6K1), is a common downstream target of signaling by hormonal or nutritional stress. Here, we investigated the initial contribution of S6K1/MAPK signaling pathways in the cell response to oxidative stress produced by hydrogen peroxide (H2O2). To analyze S6K1 activation, we used the commercial anti-phospho-Thr389-S6K1 antibody most frequently mentioned in the bibliography. We found that this antibody detected an 80-90 kDa protein that was rapidly phosphorylated in response to H2O2 in several human cells. Unexpectedly, this phosphorylation was insensitive to both mTOR and PI3K inhibitors, and knock-down experiments showed that this protein was not S6K1. RSK and MSK proteins were candidate targets of this phosphorylation. We demonstrated that H2O2 stimulated phosphorylation of RSK and MSK kinases at residues that are homologous to Thr389 in S6K1. This phosphorylation required the activity of either p38 or ERK MAP kinases. Kinase assays showed activation of RSK and MSK by H2O2. Experiments with mouse embryonic fibroblasts from p38 animals' knockout confirmed these observations. Altogether, these findings show that the S6K1 signaling pathway is not activated under these conditions, clarify previous observations probably misinterpreted by non-specific detection of proteins RSK and MSK by the anti-phospho-Thr389-S6K1 antibody, and demonstrate the specific activation of MAPK signaling pathways through ERK/p38/RSK/MSK by H2O2.

The phenolic compounds present in cocoa seeds have been studied regarding health benefits, such as antioxidant and anti-inflammatory activities. Fibrosis is a wound healing response that occurs in almost all patients with chronic liver injury. A large number of cytokines and soluble intercellular mediators are related to changes in the behavior and phenotype of the hepatic stellate cell (HSC) that develop a fibrogenic and contractile phenotype leading to the development of fibrosis. The objective of this study was to assess the catechin effect in GRX liver cells in activities such as cell growth and inflammation. The GRX cells treatment with catechin induced a significant decrease in cell growth. This mechanism does not occur by apoptosis or even by autophagy because there were no alterations in expression of caspase 3 and PARP (apoptosis), and LC3 (autophagy). The expression of p27 and p53 proteins, regulators of the cell cycle, showed increased expression, while COX-2 and IL-6 mRNA showed a significant decrease in expression. This study shows that catechin decreases cell growth in GRX cells and, probably, this decrease does not occur by apoptosis or autophagy but through an anti-inflammatory effect and cell cycle arrest. Catechin also significantly decreased the production of TGF-β by GRX cells, showing a significant antifibrotic effect.

Background: The aim of this study was to examine the effects of the N-methyl-D-aspartate receptor (NMDAR) channel blocker dizocilpine (MK-801) on lung injury in rats submitted to experimental sepsis induced by cecal ligation and perforation (CLP)., Methods: Adult male Wistar rats submitted to CLP were given a single systemic injection of MK-801 (subcutaneously at 0.3 mg/kg) administered 4 or 7 h after CLP induction. Twelve hours after CLP BAL was performed to determine total cell count, protein content, and inflammatory parameters. In addition, lung was excised for histopathologic analyses and determination of NMDAR subunits content. In a separate cohort of animals mortality was recorded for 5 days., Results: Animals submitted to sepsis induced by CLP showed an increase in the content of NMDAR subunits NR1 and NR2A in the lung. Administration of MK-801 4 h after CLP induction resulted in a decrease in BAL fluid cellular content and decreased levels of proinflammatory cytokines. In addition, MK-801 decreased lung oxidative stress markers and histopathologic alterations and improved survival., Conclusions: These findings indicate that NMDAR blockade might represent a promising novel therapeutic strategy for the treatment of sepsis and inflammatory disorders.

Sepsis remains an important and life-threatening problem, and is the most common cause of death in the intensive care unit. One promising therapeutic candidate for protection against injury in sepsis is fructose-1,6-bisphosphate (FBP), a high-energy glycolytic pathway intermediate. The objective of the study was to establish a role for FBP on the immune system, especially in lymphocyte proliferation. Peripheral blood mononuclear cells (PBMCs) were isolated from the blood of healthy humans by gradient centrifugation. T-lymphocytes were stimulated for 96 h with phytohemagglutinin (PHA) and varying concentration of FBP. Fructose-1,6-bisphosphate at concentrations between 1.2 and 10 mM decreased proliferation of T-lymphocytes and reduced the viability only at concentrations 5.0 and 10 mM. The levels of soluble IL-2 receptor were reduced at FBP concentrations between 1.2 and 10 mM. In conclusion, this study demonstrates that FBP has important effect on immunomodulatory and this result can be correlated with the protection against injury in sepsis., (Copyright 2002 Elsevier Science B.V.)

Galactosamine (GalN) is a hepatotoxic agent, which under determined situations provokes metabolic and energetic depletion as well as alterations in permeability, leading to cellular death. At the same time, it is known that fructose-1,6-bisphosphate (FBP) helps maintain cell energy levels and protects the cell against this lesive agent. We submitted two groups of male Wistar rats to the harmful intraperitoneal doses of GalN (400 mg/kg), one of which simultaneously received FBP (2 g/kg). Techniques were used in the analysis of the cellular components, adenosine triphosphate (ATP) and hepatic calcium and a close relationship between the types of cellular death unchained by these agents was established. The liver of the rats treated with GalN showed energy depletion and concomitant increase calcium in the hepatic tissue, which provoked higher levels of necrosis leading to reduce cellular viability. On the other hand, the group which had received GalN+FBP maintained calcium levels close to the control values and the energy rate did not decrease as much as in the GalN only group, but recovered the control values, within a period of 48 h. At the same time, the degree of apoptosis was greater than in the GalN group. This fact suggests that the FBP maintains cellular levels of ATP, thus protecting the cell from the toxic action of GalN, reducing the percentage of dead cells and causing an alteration in the pattern of the cell death, whereby there is an increase in the rate of apoptosis and a decrease in that of necrosis.

Objective: The aim of this research project was to test the ability of fructose 1,6-bisphosphate (FBP), which has anti-inflammatory effects and maintains cellular energy levels, to inhibit the septic process in an experimental model in rats., Design: Prospective, controlled animal trial., Setting: Research laboratory., Subjects: Fed male Wistar rats., Interventions: Three experimental groups were formed for the test: control group, untreated septic group, and septic group treated with FBP (500 mg/kg)., Measurements and Main Results: In the control group, there were no deaths; in the untreated septic group, the mortality rate was 100% within 15 hrs; in the septic group treated with FBP, the mortality rate reached 20% within 15 hrs. The blood cell tests revealed that concentrations of hematocrit, leukocytes, monocytes, and immature cells increased significantly in the untreated septic group compared with both the FBP-treated septic group and the control group. The histologic lesions verified in the heart, lungs, liver, and kidneys of septic animals were smaller and even absent in those treated with FBP., Conclusion: FBP reduced the mortality rate provoked by experimental sepsis and ameliorated hematologic and histologic alterations.

Administration of carrageenan (0.5 mg) to the plantar tissue of rats resulted in reversible inflammatory injury. This damage was monitored as changes in foot volume, using a plethysmometer. Administration of fructose 1,6-bisphosphate at different doses, orally or intraperitoneally, prevented the inflammatory action induced by the simultaneous injection of carrageenan in the rat paw. The effect was dose and time dependent. In contrast, fructose or fructose 6-phosphate afforded no significant protection. In order to extend the average half-life of the drug, we prepared liposomes of fructose 1,6-bisphosphate which, administered orally or intraperitoneally, showed a greater and more prolonged antiinflammatory action. The significance of these findings with respect to the mechanism of the antiinflammatory action of fructose 1,6-bisphosphate is discussed.