Pharmacological Targeting of BMP6-SMAD Mediated Hepcidin Expression Does Not Improve the Outcome of Systemic Infections With Intra-Or Extracellular Gram-Negative Bacteria in Mice

IntroductionHepcidin is the systemic master regulator of iron metabolism as it degrades the cellular iron exporter ferroportin. In bacterial infections, hepcidin is upregulated to limit circulating iron for pathogens, thereby increasing iron retention in macrophages. This mechanism withholds iron from extracellular bacteria but could be of disadvantage in infections with intracellular bacteria. We aimed to understand the role of hepcidin in infections with intra- or extracellular bacteria using different hepcidin inhibitors.MethodsFor the experiments LDN-193189 and oversulfated heparins were used, which interact with the BMP6-SMAD pathway thereby inhibiting hepcidin expression. We infected male C57BL/6N mice with either the intracellular bacterium Salmonella Typhimurium or the extracellular bacterium Escherichia coli and treated these mice with the different hepcidin inhibitors.ResultsBoth inhibitors effectively reduced hepcidin levels in vitro under steady state conditions and upon stimulation with the inflammatory signals interleukin-6 or lipopolysaccharide. The inhibitors also reduced hepcidin levels and increased circulating iron concentration in uninfected mice. However, both compounds failed to decrease liver- and circulating hepcidin levels in infected mice and did not affect ferroportin expression in the spleen or impact on serum iron levels. Accordingly, both BMP-SMAD signaling inhibitors did not influence bacterial numbers in different organs in the course of E.coli or S.Tm sepsis.ConclusionThese data indicate that targeting the BMP receptor or the BMP-SMAD pathway is not sufficient to suppress hepcidin expression in the course of infection with both intra- or extracellular bacteria. This suggests that upon pharmacological inhibition of the central SMAD-BMP pathways during infection, other signaling cascades are compensatorily induced to ensure sufficient hepcidin formation and iron restriction to circulating microbes.

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ESSENTIAL OILS AND METHYLGLYOXAL: A POSSIBLE ALTERNATIVE TREATMENT FOR ANTIBIOTIC RESISTANT BACTERIAL INFECTIONS

International Journal of Pharmacy and Pharmaceutical Sciences ◽

10.22159/ijpps.2016.v8i9.12242 ◽

2016 ◽

Vol 8 (9) ◽

pp. 107

Author(s):

Erin Cieslak ◽

James P. Mack ◽

Albert Rojtman

Keyword(s):

Essential Oil ◽

Essential Oils ◽

Alternative Treatment ◽

Bacterial Infections ◽

Cost Effective ◽

Diffusion Method ◽

Antibiotic Resistant ◽

Medicinal Properties

Objective: Essential oils are of significant interest in today’s world of healthcare because these compounds have a variety of medicinal properties. In this study, we evaluated the in vitro antibiotic role of essential oils as a possible alternative treatment in combatting Methicillin-resistant Staphylococcus aureus (MRSA).Methods: In conjunction with carrier oils, three essential oils (cassia, cinnamon bark, and thyme), as well as methylglyoxal were tested on MRSA using the Kirby-Bauer disc diffusion method.Results: The minimum inhibitory concentration of each tested essential oil and methylglyoxal in carrier oil was determined to be 25% essential oil and 75% carrier oil mixture. This concentration worked much more effectively than the standard antibiotic, vancomycin, which is currently used to treat MRSA infections.Conclusion: Antibacterial emollients made from naturally occurring products like essential oils can be cost-effective alternatives to antibiotics. The results of this research show that these emollients are more effective against MRSA than standard antibiotics in cell culture.

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Src-dependent NM2A tyrosine-phosphorylation regulates actomyosin dynamics

10.1101/2020.08.12.246090 ◽

2020 ◽

Author(s):

Cláudia Brito ◽

Francisco S. Mesquita ◽

Daniel S. Osório ◽

Joana Pereira ◽

Neil Billington ◽

...

Keyword(s):

Tyrosine Phosphorylation ◽

Cell Biology ◽

Bacterial Infections ◽

Focal Adhesions ◽

The Status ◽

Fine Control ◽

Actomyosin Cytoskeleton

AbstractNon-muscle myosin 2A (NM2A) is a key cytoskeletal enzyme that along with actin assembles into actomyosin filaments inside cells. NM2A is fundamental in cellular processes requiring force generation such as cell adhesion, motility and cell division, and plays important functions in different stages of development and during the progression of viral and bacterial infections. We previously identified at the motor domain of the NM2A, a novel Src-dependent tyrosine phosphorylation on residue 158 (pTyr158), which is promoted by Listeria monocytogenes infection. Despite the central role of NM2A in several cell biology processes, the pTyr at this specific residue had never been reported. Here we showed that LLO, a toxin secreted by Listeria, is sufficient to trigger NM2A pTyr158 by activating Src, which coordinates actomyosin remodeling. We further addressed the role of NM2A pTyr158 on the organization and dynamics of the actomyosin cytoskeleton and found that by controlling the activation of the NM2A, the status of the pTyr158 alters cytoskeletal organization, dynamics of focal adhesions and cell motility, without affecting NM2A enzymatic activity in vitro. Ultimately, by using Caenorhabditis elegans as a model to assess the role of this pTyr158in vivo, we found that the status of the pTyr158 has implications in gonad function and is required for organism survival under stress conditions. We conclude that the fine control of the NM2A pTyr158 is required for cell cytoskeletal remodeling and dynamics, and we propose Src-dependent NM2A pTyr158 as a novel layer of regulation of the actomyosin cytoskeleton.

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In vitro and in vivo antibacterial activities of CS-940, a new 6-fluoro-8-difluoromethoxy quinolone.

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.40.5.1201 ◽

1996 ◽

Vol 40 (5) ◽

pp. 1201-1207 ◽

Cited By ~ 7

Author(s):

N Masuda ◽

Y Takahashi ◽

M Otsuki ◽

E Ibuki ◽

H Miyoshi ◽

...

Keyword(s):

Bacterial Infections ◽

Oral Treatment ◽

Horse Serum ◽

Antibacterial Activities ◽

Inoculum Size ◽

The Other ◽

Gram Positive Bacteria ◽

Systemic Infections

The in vitro and in vivo activities of CS-940, a new 6-fluoro-8-difluoromethoxy quinolone, were compared with those of ciprofloxacin, tosufloxacin, sparfloxacin, and levofloxacin. The in vitro activity of CS-940 against gram-positive bacteria was nearly equal to or greater than those of the other quinolones tested. In particular, CS-940 was two to eight times more active against methicillin-resistant Staphylococcus aureus than the other quinolones, at the MIC at which 90% of the clinical isolates are inhibited. Against gram-negative bacteria, the activity of CS-940 was comparable to or greater than those of tosufloxacin, sparfloxacin, and levofloxacin, while it was lower than that of ciprofloxacin. The activity of CS-940 was largely unaffected by medium, inoculum size, or the addition of horse serum, but it was decreased under acidic conditions, as was also seen with the other quinolones tested. CS-940 showed potent bactericidal activity against S. aureus, Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa. In oral treatment of mouse systemic infections caused by S. aureus, Streptococcus pneumoniae, Streptococcus pyogenes, E. coli, K. pneumoniae, Serratia marcescens, and P. aeruginosa, CS-940 was more effective than ciprofloxacin, sparfloxacin, and levofloxacin against all strains tested. Against experimental pneumonia with K. pneumoniae in mice, CS-940 was the most effective of all the quinolones tested. These results suggest that CS-940 may be effective in the therapy of various bacterial infections.

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Hepcidin Regulation by the BMP Pathway.

Blood ◽

10.1182/blood.v114.22.sci-25.sci-25 ◽

2009 ◽

Vol 114 (22) ◽

pp. SCI-25-SCI-25

Author(s):

Jodie L. Babitt

Keyword(s):

Signaling Pathway ◽

Serum Iron ◽

Iron Homeostasis ◽

Iron Absorption ◽

Bmp Signaling ◽

Smad Pathway ◽

Hepcidin Expression ◽

Iron Administration ◽

Smad Proteins

Abstract Abstract SCI-25 Systemic iron balance is regulated by the key iron regulatory hormone hepcidin. Secreted by the liver, hepcidin inhibits iron absorption from the diet and iron mobilization from body stores by decreasing cell surface expression of the iron export protein ferroportin. Iron administration increases hepcidin expression, thereby providing a feedback mechanism to limit further iron absorption, while anemia and hypoxia inhibit hepcidin expression, thereby increasing iron availability for erythropoiesis. Hepcidin excess is thought to have a role in the anemia of inflammation, while hepcidin deficiency is thought to be the common pathogenic mechanism underlying the iron overload disorder hereditary hemochromatosis, due to mutations in the genes encoding hepcidin itself (HAMP), HFE, transferrin receptor 2 (TFR2), or hemojuvelin (HFE2). Notably the precise molecular mechanisms by which iron levels are “sensed” and how this iron “signal” is transduced to modulate hepcidin expression have remained elusive. We have recently discovered that hemojuvelin is a co-receptor for the bone morphogenetic protein (BMP) signaling pathway, and that hemojuvelin-mediated BMP signals increase hepcidin expression at the transcriptional level. In addition to patients with HFE2 mutations and Hfe2 knockout mice, other genetic mouse models associated with impaired hepatic BMP signaling through a global knockout of the ligand Bmp6, or selective hepatic knockout of an intracellular mediator of BMP signaling, Smad4, also cause inappropriately low hepcidin expression and iron overload. Exogenous BMP6 administration in mice increases hepatic hepcidin expression and reduces serum iron, while BMP6 antagonists inhibit hepatic hepcidin expression, mobilize reticuloendothelial cell iron stores and increase serum iron. Not only does the BMP6-hemojuvelin-SMAD pathway regulate hepcidin expression and thereby systemic iron homeostasis, but also the BMP6-SMAD pathway itself is regulated by iron. Acute iron administration in mice increases phosphorylation of Smad proteins in the liver, and chronic changes in dietary iron modulate hepatic Bmp6 mRNA expression and phosphorylation of Smad proteins concordantly with Hamp mRNA expression. Together, these data support the paramount role of the BMP6-hemojuvelin-SMAD signaling pathway in the iron-mediated regulation of hepcidin expression and systemic iron homeostasis, and suggest that modulators of this pathway may be an alternative therapeutic strategy for treating disorders of iron homeostasis. Recent work elucidating the role of the BMP signaling pathway in hepcidin regulation and systemic iron homeostasis will be presented. Disclosures Babitt: Ferrumax Pharmaceuticals, Inc.: Equity Ownership.

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Gasdermin D restricts Burkholderia cenocepacia infection in vitro and in vivo

Scientific Reports ◽

10.1038/s41598-020-79201-5 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Shady Estfanous ◽

Kathrin Krause ◽

Midhun N. K. Anne ◽

Mostafa Eltobgy ◽

Kyle Caution ◽

...

Keyword(s):

Bacterial Infections ◽

Critical Role ◽

In Vitro Study ◽

Burkholderia Cenocepacia ◽

Inflammasome Activation ◽

Bacterial Survival ◽

Life Threatening ◽

Systemic Infections

AbstractBurkholderia cenocepacia (B. cenocepacia) is an opportunistic bacterium; causing severe life threatening systemic infections in immunocompromised individuals including cystic fibrosis patients. The lack of gasdermin D (GSDMD) protects mice against endotoxin lipopolysaccharide (LPS) shock. On the other hand, GSDMD promotes mice survival in response to certain bacterial infections. However, the role of GSDMD during B. cenocepacia infection is not yet determined. Our in vitro study shows that GSDMD restricts B. cenocepacia replication within macrophages independent of its role in cell death through promoting mitochondrial reactive oxygen species (mROS) production. mROS is known to stimulate autophagy, hence, the inhibition of mROS or the absence of GSDMD during B. cenocepacia infections reduces autophagy which plays a critical role in the restriction of the pathogen. GSDMD promotes inflammation in response to B. cenocepacia through mediating the release of inflammasome dependent cytokine (IL-1β) and an independent one (CXCL1) (KC). Additionally, different B. cenocepacia secretory systems (T3SS, T4SS, and T6SS) contribute to inflammasome activation together with bacterial survival within macrophages. In vivo study confirmed the in vitro findings and showed that GSDMD restricts B. cenocepacia infection and dissemination and stimulates autophagy in response to B. cenocepacia. Nevertheless, GSDMD promotes lung inflammation and necrosis in response to B. cenocepacia without altering mice survival. This study describes the double-edged functions of GSDMD in response to B. cenocepacia infection and shows the importance of GSDMD-mediated mROS in restriction of B. cenocepacia.

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Duodenal cytochrome b: a novel ferrireductase in airway epithelial cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00342.2005 ◽

2006 ◽

Vol 291 (2) ◽

pp. L272-L280 ◽

Cited By ~ 17

Author(s):

Jennifer L. Turi ◽

Xinchao Wang ◽

Andrew T. McKie ◽

Eva Nozik-Grayck ◽

Lisa B. Mamo ◽

...

Keyword(s):

Epithelial Cells ◽

Cytochrome B ◽

Iron Transport ◽

Iron Concentration ◽

Airway Epithelial Cells ◽

Airway Epithelial ◽

Cellular Iron ◽

Duodenal Cytochrome B

Catalytically active iron in the lung causes oxidative stress and promotes microbial growth that can be limited by intracellular sequestration of iron within ferritin. Because cellular iron uptake requires membrane ferrireductase activity that in the gut can be provided by duodenal cytochrome b (Dcytb), we sought Dcytb in the lung to test the hypothesis that it contributes to epithelial iron regulation by reducing Fe3+ for cellular iron transport. Dcytb expression was found in respiratory epithelium in vitro and in vivo and was responsive to iron concentration. Iron transport was measured in human bronchial epithelial (HBE) cells using inductively coupled plasma atomic emission spectroscopy and was demonstrated to be partially inhibited in the presence of Dcytb-blocking antibody, suggesting that Dcytb reduces Fe3+ for cellular iron transport. A definite source of reducing equivalents for Dcytb was sought but not identified. We found no evidence that ascorbate was involved but did demonstrate that O2−· production decreased when Dcytb function was blocked. The presence of Dcytb in airway epithelial cells and its regulation by iron therefore may contribute to pulmonary cytoprotection.

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Synthetic Porcine Hepcidin Exhibits Different Roles in Escherichia coli and Salmonella Infections

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02638-16 ◽

2017 ◽

Vol 61 (10) ◽

Cited By ~ 8

Author(s):

Dan Liu ◽

Zhen-Shun Gan ◽

Wan Ma ◽

Hai-Tao Xiong ◽

Yun-Qing Li ◽

...

Keyword(s):

Escherichia Coli ◽

Bacterial Infections ◽

Iron Status ◽

Iron Chelator ◽

Intracellular Iron ◽

Content Type ◽

Defense Strategies ◽

Salmonella Infections

ABSTRACT Hepcidin, an antimicrobial peptide, was discovered to integrate diverse signals from iron status and an infection threat and orchestrate a series of host-protective responses. Several studies have investigated the antimicrobial role of hepcidin, but the results have been controversial. Here, we aimed to examine the role of hepcidin in bacterial adherence and invasion in vitro. We found that porcine hepcidin could decrease the amount of the extracellular pathogen enterotoxigenic Escherichia coli (ETEC) K88 that adhered to cells because it caused the aggregation of the bacteria. However, addition of hepcidin to macrophages infected with the intracellular pathogen Salmonella enterica serovar Typhimurium enhanced the intracellular growth of the pathogen through the degradation of ferroportin, an iron export protein, and then the sequestration of intracellular iron. Intracellular iron was unavailable by use of the iron chelator deferiprone (DFO), which reduced intracellular bacterial growth. These results demonstrate that hepcidin exhibits different functions in extracellular and intracellular bacterial infections, which suggests that different defense strategies should be taken to prevent bacterial infection.

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The Effect of Bacterial Infections, Probiotics and Zonulin on Intestinal Barrier Integrity

International Journal of Molecular Sciences ◽

10.3390/ijms222111359 ◽

2021 ◽

Vol 22 (21) ◽

pp. 11359

Author(s):

Paweł Serek ◽

Monika Oleksy-Wawrzyniak

Keyword(s):

Bacterial Infections ◽

Intestinal Barrier ◽

The Body ◽

Entire Body ◽

Gut Barrier ◽

Selective Permeability ◽

Current State ◽

Viable Approach ◽

Systemic Infections

The intestinal barrier plays an extremely important role in maintaining the immune homeostasis of the gut and the entire body. It is made up of an intricate system of cells, mucus and intestinal microbiota. A complex system of proteins allows the selective permeability of elements that are safe and necessary for the proper nutrition of the body. Disturbances in the tightness of this barrier result in the penetration of toxins and other harmful antigens into the system. Such events lead to various digestive tract dysfunctions, systemic infections, food intolerances and autoimmune diseases. Pathogenic and probiotic bacteria, and the compounds they secrete, undoubtedly affect the properties of the intestinal barrier. The discovery of zonulin, a protein with tight junction regulatory activity in the epithelia, sheds new light on the understanding of the role of the gut barrier in promoting health, as well as the formation of diseases. Coincidentally, there is an increasing number of reports on treatment methods that target gut microbiota, which suggests that the prevention of gut-barrier defects may be a viable approach for improving the condition of COVID-19 patients. Various bacteria–intestinal barrier interactions are the subject of this review, aiming to show the current state of knowledge on this topic and its potential therapeutic applications.

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CovR and VicRKX Regulate Transcription of the Collagen Binding Protein Cnm ofStreptococcus mutans

Journal of Bacteriology ◽

10.1128/jb.00141-18 ◽

2018 ◽

Vol 200 (23) ◽

Cited By ~ 2

Author(s):

Lívia Araújo Alves ◽

Tridib Ganguly ◽

Renata O. Mattos-Graner ◽

Jessica Kajfasz ◽

Erika N. Harth-Chu ◽

...

Keyword(s):

Streptococcus Mutans ◽

Bacterial Infections ◽

Regulatory Region ◽

Core Gene ◽

Negative Regulator ◽

Mobility Shift ◽

Content Type ◽

Intracellular Invasion ◽

Systemic Infections

ABSTRACTCnm is a surface-associated protein present in a subset ofStreptococcus mutansstrains that mediates binding to extracellular matrices, intracellular invasion, and virulence. Here, we showed thatcnmtranscription is controlled by the global regulators CovR and VicRKX.In silicoanalysis identified multiple putative CovR- and VicR-binding motifs in the regulatory region ofcnmas well as in the downstream genepgfS, which is associated with the posttranslational modification of Cnm. Electrophoretic mobility shift assays revealed that CovR and VicR specifically and independently bind to thecnmandpgfSpromoter regions. Quantitative real-time PCR and Western blot analyses of ΔcovRand ΔvicKstrains as well as of a strain overexpressingvicRKXrevealed that CovR functions as a positive regulator ofcnm, whereas VicRKX acts as a negative regulator. In agreement with the role of VicRKX as a repressor, the ΔvicKstrain showed enhanced binding to collagen and laminin and higher intracellular invasion rates. Overexpression ofvicRKXwas associated with decreased rates of intracellular invasion but did not affect collagen or lamin binding activities, suggesting that this system controls additional genes involved in binding to these extracellular matrix proteins. As expected, based on the role of CovR incnmregulation, the ΔcovRstrain showed decreased intracellular invasion rates, but, unexpectedly collagen and laminin binding activities were increased in this mutant strain. Collectively, the results presented here expand the repertoire of virulence-related genes regulated by CovR and VicRKX to include the core genepgfSand the noncore genecnm.IMPORTANCEStreptococcus mutansis a major pathogen associated with dental caries and also implicated in systemic infections, in particular, infective endocarditis. The Cnm adhesin ofS. mutansis an important virulence factor associated with systemic infections and caries severity. Despite its role in virulence, the regulatory mechanisms governingcnmexpression are poorly understood. Here, we describe the identification of two independent regulatory systems controlling the transcription ofcnmand the downstreampgfS-pgfM1-pgfE-pgfM2operon. A better understanding of the mechanisms controlling expression of virulence factors like Cnm can facilitate the development of new strategies to treat bacterial infections.

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In Vitro and In Vivo Antibacterial Activities of CS-023 (RO4908463), a Novel Parenteral Carbapenem

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.49.8.3239-3250.2005 ◽

2005 ◽

Vol 49 (8) ◽

pp. 3239-3250 ◽

Cited By ~ 42

Author(s):

Tetsufumi Koga ◽

Tomomi Abe ◽

Harumi Inoue ◽

Takashi Takenouchi ◽

Akiko Kitayama ◽

...

Keyword(s):

Bacterial Infections ◽

Systemic Infection ◽

Antibacterial Activities ◽

Infection Model ◽

Gram Positive ◽

Methicillin Resistant ◽

Lung Infections ◽

Systemic Infections

ABSTRACT CS-023 (RO4908463, formerly R-115685) is a novel 1β-methylcarbapenem with 5-substituted pyrrolidin-3-ylthio groups, including an amidine moiety at the C-2 position. Its antibacterial activity was tested against 1,214 clinical isolates of 32 species and was compared with those of imipenem, meropenem, ceftazidime, ceftriaxone, ampicillin, amikacin, and levofloxacin. CS-023 exhibited a broad spectrum of activity against gram-positive and -negative aerobes and anaerobes, including methicillin-resistant Staphylococcus aureus (MRSA), methicillin-resistant Staphylococcus epidermidis, penicillin-resistant Streptococcus pneumoniae (PRSP), β-lactamase-negative ampicillin-resistant Haemophilus influenzae, and Pseudomonas aeruginosa. CS-023 showed the most potent activity among the compounds tested against P. aeruginosa and MRSA, with MICs at which 90% of isolates tested were inhibited of 4 μg/ml and 8 μg/ml, respectively. CS-023 was stable against hydrolysis by the β-lactamases from Enterobacter cloacae and Proteus vulgaris. CS-023 also showed potent activity against extended-spectrum β-lactamase-producing Escherichia coli. The in vivo efficacy of CS-023 was evaluated with a murine systemic infection model induced by 13 strains of gram-positive and -negative pathogens and a lung infection model induced by 2 strains of PRSP (serotypes 6 and 19). Against the systemic infections with PRSP, MRSA, and P. aeruginosa and the lung infections, the efficacy of CS-023 was comparable to those of imipenem/cilastatin and vancomycin (tested against lung infections only) and superior to those of meropenem, ceftriaxone, and ceftazidime (tested against P. aeruginosa infections only). These results suggest that CS-023 has potential for the treatment of nosocomial bacterial infections by gram-positive and -negative pathogens, including MRSA and P. aeruginosa.

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