Intracellular Activity of Antibiotics against Staphylococcus aureus in a Mouse Peritonitis Model

ABSTRACT Antibiotic treatment of Staphylococcus aureus infections is often problematic due to the slow response to therapy and the high frequency of infection recurrence. The intracellular persistence of staphylococci has been recognized and could offer a good explanation for these treatment difficulties. Knowledge of the interplay between intracellular antibiotic activity and the overall outcome of infection is therefore important. Several intracellular in vitro models have been developed, but few experimental animal models have been published. The mouse peritonitis/sepsis model was used as the basic in vivo model exploring a quantitative ex vivo extra- and intracellular differentiation assay. The intracellular presence of S. aureus was documented by electron microscopy. Five antibiotics, dicloxacillin, cefuroxime, gentamicin, azithromycin, and rifampin (rifampicin), were tested in the new in vivo model; and the model was able to distinguish between their extra- and intracellular effects. The intracellular effects of the five antibiotics could be ranked as follows as the mean change in the log10 number of CFU/ml (Δlog10 CFU/ml) between treated and untreated mice after 4 h of treatment: dicloxacillin (3.70 Δlog10 CFU/ml) > cefuroxime (3.56 Δlog10 CFU/ml) > rifampin (1.86 Δlog10 CFU/ml) > gentamicin (0.61 Δlog10 CFU/ml) > azithromycin (0.21 Δlog10 CFU/ml). We could also show that the important factors during testing of intracellular activity in vivo are the size, number, and frequency of doses; the time of exposure; and the timing between the start of infection and treatment. A poor correlation between the intracellular accumulation of the antibiotics and the actual intracellular effect was found. This stresses the importance of performing experimental studies, like those with the new in vivo model described here, to measure actual intracellular activity instead of making predictions based on cellular pharmacokinetic and MICs.

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Plectasin Shows Intracellular Activity against Staphylococcus aureus in Human THP-1 Monocytes and in a Mouse Peritonitis Model

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00685-09 ◽

2009 ◽

Vol 53 (11) ◽

pp. 4801-4808 ◽

Cited By ~ 33

Author(s):

Karoline Sidelmann Brinch ◽

Anne Sandberg ◽

Pierre Baudoux ◽

Françoise Van Bambeke ◽

Paul M. Tulkens ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Animal Studies ◽

Kinetic Studies ◽

Response To Therapy ◽

In Vitro Studies ◽

In Vivo Model ◽

Intracellular Activity ◽

Peritonitis Model

ABSTRACT Antimicrobial therapy of infections with Staphylococcus aureus can pose a challenge due to slow response to therapy and recurrence of infection. These treatment difficulties can partly be explained by intracellular survival of staphylococci, which is why the intracellular activity of antistaphylococcal compounds has received increased attention within recent years. The intracellular activity of plectasin, an antimicrobial peptide, against S. aureus was determined both in vitro and in vivo. In vitro studies using THP-1 monocytes showed that some intracellular antibacterial activity of plectasin was maintained (maximal relative efficacy [E max], 1.0- to 1.3-log reduction in CFU) even though efficacy was inferior to that of extracellular killing (E max, >4.5-log CFU reduction). Animal studies included a novel use of the mouse peritonitis model, exploiting extra- and intracellular differentiation assays, and assessment of the correlations between activity and pharmacokinetic (PK) parameters. The intracellular activity of plectasin was in accordance with the in vitro studies, with an E max of a 1.1-log CFU reduction. The parameter most important for activity was fC peak/MIC, where fC peak is the free peak concentration. These findings stress the importance of performing studies of extra- and intracellular activity since these features cannot be predicted from traditional MIC and killing kinetic studies. Application of both the THP-1 and the mouse peritonitis models showed that the in vitro results were similar to findings in the in vivo model with respect to demonstration of intracellular activity. Therefore the in vitro model was a good screening model for intracellular activity. However, animal models should be applied if further information on activity, PK/pharmacodynamic parameters, and optimal dosing regimens is required.

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AS1411 Nucleolin-Specific Binding Aptamers Reduce Pathological Angiogenesis through Inhibition of Nucleolin Phosphorylation

International Journal of Molecular Sciences ◽

10.3390/ijms222313150 ◽

2021 ◽

Vol 22 (23) ◽

pp. 13150

Author(s):

Emilio Iturriaga-Goyon ◽

Oscar Vivanco-Rojas ◽

Fátima Sofía Magaña-Guerrero ◽

Beatriz Buentello-Volante ◽

Ilse Castro-Salas ◽

...

Keyword(s):

Ex Vivo ◽

Specific Binding ◽

Pediatric Population ◽

Tube Formation ◽

In Vitro Models ◽

In Vivo Model ◽

Systemic Complications ◽

Pathological Angiogenesis

Proliferative retinopathies produces an irreversible type of blindness affecting working age and pediatric population of industrialized countries. Despite the good results of anti-VEGF therapy, intraocular and systemic complications are often associated after its intravitreal use, hence novel therapeutic approaches are needed. The aim of the present study is to test the effect of the AS1411, an antiangiogenic nucleolin-binding aptamer, using in vivo, ex vivo and in vitro models of angiogenesis and propose a mechanistic insight. Our results showed that AS1411 significantly inhibited retinal neovascularization in the oxygen induced retinopathy (OIR) in vivo model, as well as inhibited branch formation in the rat aortic ex vivo assay, and, significantly reduced proliferation, cell migration and tube formation in the HUVEC in vitro model. Importantly, phosphorylated NCL protein was significantly abolished in HUVEC in the presence of AS1411 without affecting NFκB phosphorylation and -21 and 221-angiomiRs, suggesting that the antiangiogenic properties of this molecule are partially mediated by a down regulation in NCL phosphorylation. In sum, this new research further supports the NCL role in the molecular etiology of pathological angiogenesis and identifies AS1411 as a novel anti-angiogenic treatment.

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Carbapenem-Resistant Klebsiella pneumoniae Clinical Isolates: In Vivo Virulence Assessment in Galleria mellonella and Potential Therapeutics by Polycationic Oligoethyleneimine

Antibiotics ◽

10.3390/antibiotics10010056 ◽

2021 ◽

Vol 10 (1) ◽

pp. 56

Author(s):

Dalila Mil-Homens ◽

Maria Martins ◽

José Barbosa ◽

Gabriel Serafim ◽

Maria J. Sarmento ◽

...

Keyword(s):

Klebsiella Pneumoniae ◽

Galleria Mellonella ◽

Multidrug Resistant ◽

In Vitro Models ◽

Clinical Isolates ◽

In Vivo Model ◽

Virulence Potential ◽

Hospital Acquired

Klebsiella pneumoniae, one of the most common pathogens found in hospital-acquired infections, is often resistant to multiple antibiotics. In fact, multidrug-resistant (MDR) K. pneumoniae producing KPC or OXA-48-like carbapenemases are recognized as a serious global health threat. In this sense, we evaluated the virulence of K. pneumoniae KPC(+) or OXA-48(+) aiming at potential antimicrobial therapeutics. K. pneumoniae carbapenemase (KPC) and the expanded-spectrum oxacillinase OXA-48 isolates were obtained from patients treated in medical care units in Lisbon, Portugal. The virulence potential of the K. pneumonia clinical isolates was tested using the Galleria mellonella model. For that, G. mellonella larvae were inoculated using patients KPC(+) and OXA-48(+) isolates. Using this in vivo model, the KPC(+) K. pneumoniae isolates showed to be, on average, more virulent than OXA-48(+). Virulence was found attenuated when a low bacterial inoculum (one magnitude lower) was tested. In addition, we also report the use of a synthetic polycationic oligomer (L-OEI-h) as a potential antimicrobial agent to fight infectious diseases caused by MDR bacteria. L-OEI-h has a broad-spectrum antibacterial activity and exerts a significantly bactericidal activity within the first 5-30 min treatment, causing lysis of the cytoplasmic membrane. Importantly, the polycationic oligomer showed low toxicity against in vitro models and no visible cytotoxicity (measured by survival and health index) was noted on the in vivo model (G. mellonella), thus L-OEI-h is foreseen as a promising polymer therapeutic for the treatment of MDR K. pneumoniae infections.

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Lignans from Bursera fagaroides Affect In Vivo Cell Behavior by Disturbing the Tubulin Cytoskeleton in Zebrafish Embryos

Molecules ◽

10.3390/molecules24010008 ◽

2018 ◽

Vol 24 (1) ◽

pp. 8 ◽

Cited By ~ 2

Author(s):

Mayra Antúnez-Mojica ◽

Andrés Rojas-Sepúlveda ◽

Mario Mendieta-Serrano ◽

Leticia Gonzalez-Maya ◽

Silvia Marquina ◽

...

Keyword(s):

Cell Migration ◽

Biological Effects ◽

In Vitro Models ◽

In Vivo Model ◽

Zebrafish Embryos ◽

Microtubule Cytoskeleton ◽

Zebrafish Model ◽

Bioassay Guided Fractionation

By using a zebrafish embryo model to guide the chromatographic fractionation of antimitotic secondary metabolites, seven podophyllotoxin-type lignans were isolated from a hydroalcoholic extract obtained from the steam bark of Bursera fagaroides. The compounds were identified as podophyllotoxin (1), β-peltatin-A-methylether (2), 5′-desmethoxy-β-peltatin-A-methylether (3), desmethoxy-yatein (4), desoxypodophyllotoxin (5), burseranin (6), and acetyl podophyllotoxin (7). The biological effects on mitosis, cell migration, and microtubule cytoskeleton remodeling of lignans 1–7 were further evaluated in zebrafish embryos by whole-mount immunolocalization of the mitotic marker phospho-histone H3 and by a tubulin antibody. We found that lignans 1, 2, 4, and 7 induced mitotic arrest, delayed cell migration, and disrupted the microtubule cytoskeleton in zebrafish embryos. Furthermore, microtubule cytoskeleton destabilization was observed also in PC3 cells, except for 7. Therefore, these results demonstrate that the cytotoxic activity of 1, 2, and 4 is mediated by their microtubule-destabilizing activity. In general, the in vivo and in vitro models here used displayed equivalent mitotic effects, which allows us to conclude that the zebrafish model can be a fast and cheap in vivo model that can be used to identify antimitotic natural products through bioassay-guided fractionation.

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Increased intracellular activity of MP1102 and NZ2114 against Staphylococcus aureus in vitro and in vivo

Scientific Reports ◽

10.1038/s41598-018-22245-5 ◽

2018 ◽

Vol 8 (1) ◽

Cited By ~ 10

Author(s):

Xiao Wang ◽

Xiumin Wang ◽

Da Teng ◽

Ruoyu Mao ◽

Ya Hao ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Intracellular Activity

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Viscum album (L) extracts in cancer treatment: a systematic review of in vitro and in vivo studies

International Journal of High Dilution Research - ISSN 1982-6206 ◽

10.51910/ijhdr.v14i2.787 ◽

2021 ◽

Vol 14 (2) ◽

pp. 52-52

Author(s):

Aloisio Cunha de Carvalho ◽

Leoni Villano Bonamin

Keyword(s):

Systematic Review ◽

Experimental Studies ◽

Pharmaceutical Preparations ◽

Viscum Album ◽

In Vitro Models ◽

In Vivo Studies ◽

Pubmed Database ◽

Anti Cancer

Background: Several reviews about phytotherapy and homeopathy have been published in the last years, including Viscum album (VA.L). VA is a parasite plant whose extract has anti-cancer proprieties and is used alone or in combination with conventional chemotherapy. Methods: We performed a systematic review about the in vivo and in vitro models described in the literature, including veterinary clinical trials. The literature was consulted from Pubmed database. Results: There are several kinds of pharmaceutical preparations about VA and their active principles used in experimental studies, lectin being frequently studied (alone or as an extract compound). More than 50% of available literature about VA is related to the lectin effects. On the other hand, the effects of viscotoxins are less studied. Among the in vivo experimental studies about VA and its compounds, the B16 murine melanoma is the most used model, followed by Ehrlich, Walker and Dalton tumors. The results point to the apoptotic effects, metastasis control and tumor regression. Some veterinary clinical studies about the use of VA in the treatment of sarcoid, fibrosarcoma and neuroblastoma are quoted in literature too, with interesting results. Considering the in vitro models, our review revealed that NALM6 leukemia cells, B16 melanoma and NC1-H460 lung carcinoma were the most studied tumor models, apoptosis signals being the most important findings. Only one study verified immunoglobulin and interleukin production. All consulted papers were related to phytotherapy preparations only. Conclusions: Although the literature about the anti-cancer activity of VA extract and its lectins is enough, there is a marked lack of information about viscotoxin activities and about the effects of homeopathic preparations of this plant on animal tumors and on in vitro cultivated tumor cells.

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Experimental Migraine Models and Their Relevance in Migraine Therapy

Cephalalgia ◽

10.1111/j.1468-2982.2005.01082.x ◽

2006 ◽

Vol 26 (6) ◽

pp. 642-659 ◽

Cited By ~ 31

Author(s):

U Arulmani ◽

S Gupta ◽

A Maassen VanDenBrink ◽

D Centurión ◽

CM Villalón ◽

...

Keyword(s):

Electrical Stimulation ◽

Trigeminal Ganglion ◽

In Vitro Models ◽

In Vivo Model ◽

Trigeminal System ◽

Sagittal Sinus ◽

Migraine Pathophysiology ◽

Stimulation Of

Although the understanding of migraine pathophysiology is incomplete, it is now well accepted that this neurovascular syndrome is mainly due to a cranial vasodilation with activation of the trigeminal system. Several experimental migraine models, based on vascular and neuronal involvement, have been developed. Obviously, the migraine models do not entail all facets of this clinically heterogeneous disorder, but their contribution at several levels (molecular, in vitro, in vivo) has been crucial in the development of novel antimigraine drugs and in the understanding of migraine pathophysiology. One important vascular in vivo model, based on an assumption that migraine headache involves cranial vasodilation, determines porcine arteriovenous anastomotic blood flow. Other models utilize electrical stimulation of the trigeminal ganglion/nerve to study neurogenic dural inflammation, while the superior sagittal sinus stimulation model takes into account the transmission of trigeminal nociceptive input in the brainstem. More recently, the introduction of integrated models, namely electrical stimulation of the trigeminal ganglion or systemic administration of capsaicin, allows studying the activation of the trigeminal system and its effect on the cranial vasculature. Studies using in vitro models have contributed enormously during the preclinical stage to characterizing the receptors in cranial blood vessels and to studying the effects of several putative antimigraine agents. The aforementioned migraine models have advantages as well as some limitations. The present review is devoted to discussing various migraine models and their relevance to antimigraine therapy.

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Impact of Bicarbonate-β-Lactam Exposures on Methicillin-Resistant Staphylococcus aureus (MRSA) Gene Expression in Bicarbonate-β-Lactam-Responsive vs. Non-Responsive Strains

Genes ◽

10.3390/genes12111650 ◽

2021 ◽

Vol 12 (11) ◽

pp. 1650

Author(s):

Selvi C. Ersoy ◽

Blake M. Hanson ◽

Richard A. Proctor ◽

Cesar A. Arias ◽

Truc T. Tran ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Cell Wall ◽

Methicillin Resistant Staphylococcus Aureus ◽

Ex Vivo ◽

In Vivo Models ◽

Methicillin Resistant ◽

Genetic Perturbations ◽

Functional Relevance

Methicillin-resistant Staphylococcus aureus (MRSA) infections represent a difficult clinical treatment issue. Recently, a novel phenotype was discovered amongst selected MRSA which exhibited enhanced β-lactam susceptibility in vitro in the presence of NaHCO3 (termed ‘NaHCO3-responsiveness’). This increased β-lactam susceptibility phenotype has been verified in both ex vivo and in vivo models. Mechanistic studies to-date have implicated NaHCO3-mediated repression of genes involved in the production, as well as maturation, of the alternative penicillin-binding protein (PBP) 2a, a necessary component of MRSA β-lactam resistance. Herein, we utilized RNA-sequencing (RNA-seq) to identify genes that were differentially expressed in NaHCO3-responsive (MRSA 11/11) vs. non-responsive (COL) strains, in the presence vs. absence of NaHCO3-β-lactam co-exposures. These investigations revealed that NaHCO3 selectively repressed the expression of a cadre of genes in strain 11/11 known to be a part of the sigB-sarA-agr regulon, as well as a number of genes involved in the anchoring of cell wall proteins in MRSA. Moreover, several genes related to autolysis, cell division, and cell wall biosynthesis/remodeling, were also selectively impacted by NaHCO3-OXA exposure in the NaHCO3-responsive strain MRSA 11/11. These outcomes provide an important framework for further studies to mechanistically verify the functional relevance of these genetic perturbations to the NaHCO3-responsiveness phenotype in MRSA.

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A Niclosamide-releasing hot-melt extruded catheter prevents Staphylococcus aureus experimental biomaterial-associated infection

10.1101/2022.01.10.475592 ◽

2022 ◽

Author(s):

Jesus Augusto Vazquez-Rodriguez ◽

Bahaa Shaqour ◽

Clara Guarch-Perez ◽

Emilia Choinska ◽

Martijn Riool ◽

...

Keyword(s):

Staphylococcus Aureus ◽

Mouse Model ◽

Bacterial Colonization ◽

Thermoplastic Polyurethane ◽

In Vitro Models ◽

Surrounding Tissue ◽

Preclinical Development ◽

Hot Melt

Biomaterial-associated infections are a major healthcare challenge as they are responsible for high disease burden in critically ill patients. In this study, we have developed drug-eluting antibacterial catheters to prevent catheter-related infections. Niclosamide (NIC), originally a well-studied antiparasitic drug, was incorporated into the polymeric matrix of thermoplastic polyurethane (TPU) via solvent casting, and catheters were fabricated using hot-melt extrusion technology. The mechanical and physicochemical properties of TPU polymers loaded with NIC were studied. NIC was released in a sustained manner from the catheters and exhibited antibacterial activity against Staphylococcus aureus and Staphylococcus epidermidis in different in vitro models. Moreover, the antibacterial efficacy of NIC-loaded catheters was validated in an in vivo biomaterial-associated infection mouse model using a methicillin-susceptible and methicillin-resistant strain of S. aureus. The released NIC from the produced catheters reduced bacterial colonization of the catheter as well as of the surrounding tissue. A sustained in vivo release of NIC from the catheters for at least 14 days was observed. In summary, the NIC-releasing hot-melt extruded catheters prevented implant colonization and reduced the bacterial colonization of peri-catheter tissue by methicillin sensitive as well as resistant S. aureus in a biomaterial-associated infection mouse model and has good prospects for preclinical development.

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Characterizing the Role of Biologically Relevant Fluid Dynamics on Silver Nanoparticle Dependent Oxidative Stress in Adherent and Suspension In Vitro Models

Antioxidants ◽

10.3390/antiox10060832 ◽

2021 ◽

Vol 10 (6) ◽

pp. 832

Author(s):

Katherine E. Burns ◽

Robert F. Uhrig ◽

Maggie E. Jewett ◽

Madison F. Bourbon ◽

Kristen A. Krupa

Keyword(s):

Oxidative Stress ◽

Fluid Dynamics ◽

In Vitro Models ◽

Poor Correlation ◽

Cellular Interactions ◽

Dynamic Flow ◽

In Vivo Models ◽

And Oxidative Stress

Silver nanoparticles (AgNPs) are being employed in numerous consumer goods and applications; however, they are renowned for inducing negative cellular consequences including toxicity, oxidative stress, and an inflammatory response. Nanotoxicological outcomes are dependent on numerous factors, including physicochemical, biological, and environmental influences. Currently, NP safety evaluations are carried out in both cell-based in vitro and animal in vivo models, with poor correlation between these mechanisms. These discrepancies highlight the need for enhanced exposure environments, which retain the advantages of in vitro models but incorporate critical in vivo influences, such as fluid dynamics. This study characterized the effects of dynamic flow on AgNP behavior, cellular interactions, and oxidative stress within both adherent alveolar (A549) and suspension monocyte (U937) models. This study determined that the presence of physiologically relevant flow resulted in substantial modifications to AgNP cellular interactions and subsequent oxidative stress, as assessed via reactive oxygen species (ROS), glutathione levels, p53, NFκB, and secretion of pro-inflammatory cytokines. Within the adherent model, dynamic flow reduced AgNP deposition and oxidative stress markers by roughly 20%. However, due to increased frequency of contact, the suspension U937 cells were associated with higher NP interactions and intracellular stress under fluid flow exposure conditions. For example, the increased AgNP association resulted in a 50% increase in intracellular ROS and p53 levels. This work highlights the potential of modified in vitro systems to improve analysis of AgNP dosimetry and safety evaluations, including oxidative stress assessments.

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