Effect of Ducrosia anethifolia methanol extract against methicillin resistant Staphylococcus aureus and Pseudomonas aeruginosa biofilms on excision wound in diabetic mice.

Background: Ducrosia anethifolia is an aromatic desert plant used in Saudi folk medicine to treat skin infections. It is widely found in Middle Eastern countries. Methods: A methanolic extract of the plant was prepared, and its phytoconstituents were determined using LC-MS. In-vitro and in-vivo antibacterial and antibiofilm activities of the methanolic extract were evaluated against multidrug-resistant bacteria. The cytotoxic effect was assessed using HaCaT cell lines in-vitro. Diabetic mice were used to study the in-vivo antibiofilm and wound healing activity using the excision wound method. Results: More than 50 phytoconstituents were found in the extract after LC-MS analysis. The extract exhibited antibacterial activity against both the tested pathogens. The extract was free of irritant effects on mice skin, and no cytotoxicity was observed on HaCaT cells with an IC50 value of 1381 µg/ml. The ointment formulation of the extract increased the healing of diabetic wounds. The microbial load of both pathogens in the wounded tissue was also reduced after the treatment. The extract was more effective against methicillin-resistant Staphylococcus aureus (MRSA) than MDR-P. aeruginosa in both in vitro and in vivo experiments. Further, skin regeneration was also observed in histological studies. Conclusions: The results showed that D. anethifolia methanol extract supports wound healing in infected wounds in diabetic mice through antibacterial, antibiofilm, and wound healing activities.

Assessment of the anti-cancer potential of Ephedra foeminea leaf extract on MDA-MB-231, MCF-7, 4 T1, and MCF-10 breast cancer cell lines: Cytotoxic, apoptotic and oxidative assays.

Ephedra foeminea is traditionally used to treat breast cancer in several Arab countries. Scientific studies have reported different effects of this plant on some cancer cell lines. The current study determined the anti-cancer potential of the methanolic extract of Ephedra foeminea against four different types of breast cancer cell lines in-vitro. The extract was prepared by maceration and phytoconstituents were identified by LC-MS analysis. The IC50 value was determined against MDA-MB-231, MCF-7, 4 T1, and MCF-10 cell lines using the MTT assay. Further investigations were carried out using IC50 concentration of the extract (40.09 µg/ml) to determine live/dead cells by acridine orange/ethidium bromide staining. The effect on the expression of reactive oxygen species (ROS) was evaluated by flow cytometry. The results were analyzed using one-way ANOVA followed by Tukey's test. The LC-MS analysis revealed the presence of 34 and 30 phytoconstituents in positive and negative modes respectively. The Ephedra foeminea extract was most effective against 4 T1 cells in a dose-dependent manner (P < 0.001) with an IC50 value of 40.09 µg/ml and showed negligible effect against MCF-10 cells. It increased apoptosis in 77.84 % of 4 T1 cells, as determined by acridine orange/ethidium bromide staining. The extract also increased the ROS expression in the 39.57 % of 4 T1 cells. The study results showed that Ephedra foeminea extract possesses an anti-cancer effect against 4 T1 cells by increasing the expression of ROS and inducing apoptosis in the 4 T1 cells. The result suggests Ephedra foemenia methanolic extract possesses a reasonable anti-cancer effect due to its effect on apoptosis and oxidative pathways. The results confirm the traditional belief that Ephedra is effective against breast cancerز.

Effect of Achillea fragrantissima Extract on Excision Wound Biofilms of MRSA and Pseudomonas aeruginosa in Diabetic Mice.

Achillea fragrantissima, a desert plant commonly known as yarrow, is traditionally used as an antimicrobial agent in folklore medicine in Saudi Arabia. The current study was undertaken to determine its antibiofilm activity against methicillin-resistant Staphylococcus aureus (MRSA) and multi-drug-resistant Pseudomonas aeruginosa (MDR-P. aeruginosa) using in vitro and in vivo studies. A biofilm model induced through an excision wound in diabetic mice was used to evaluate its effect in vivo. The skin irritation and cytotoxic effects of the extract were determined using mice and HaCaT cell lines, respectively. The Achillea fragrantissima methanolic extract was analyzed with LC-MS to detect different phytoconstituents, which revealed the presence of 47 different phytoconstituents. The extract inhibited the growth of both tested pathogens in vitro. It also increased the healing of biofilm-formed excision wounds, demonstrating its antibiofilm, antimicrobial, and wound-healing action in vivo. The effect of the extract was concentration-dependent, and its activity was stronger against MRSA than MDR-P. aeruginosa. The extract formulation was devoid of a skin irritation effect in vivo and cytotoxic effect on HaCaT cell lines in vitro.

Investigating the Effect of Cigarette Smoking on Serum Uric Acid Levels in Multiple Sclerosis Patients: A Cross Sectional Study.

OBJECTIVES: The present study is aimed at determining the effect of cigarette smoking (CS) on serum uric acid (UA) levels quantitatively before and after smoking cessation among people with MS (pwMS). Additionally, a possible correlation between UA levels and both disability progression and disease severity was also investigated. A retrospective cross-sectional study was conducted using the Nottingham University Hospitals MS Clinics database. It involves 127 people with definite MS recorded when reporting the latest smoking status and the clinical diagnosis. All necessary demographics and clinical characteristics were collected. We found that smoker pwMS had significantly lower serum UA levels than non-smoker pwMS (p-value = 0.0475), and this reduction was recovered after smoking cessation (p-value = 0.0216). However, the levels of disability or disease severity were not correlated with the levels of serum UA in current smoker pwMS, measured by the expanded disability status scale (EDSS; r = -0.24; p-value = 0.38), multiple sclerosis impact scale 29 (MSIS-29; r = 0.01; p-value = 0.97) and MS severity score (MSSS; r = -0.16; p-value = 0.58), respectively. Our result suggests that the reduction in UA levels is more likely a consequence of oxidative stress triggered by many risk factors, including CS, and could be considered a potential indicator of smoking cessation. In addition, the absence of a correlation between UA levels and disease severity and disability suggests that UA is not an optimal biomarker for disease severity and disability prediction among current smoker, ex-smoker or non-smoker pwMS.

A Simple In-Vivo Method for Evaluation of Antibiofilm and Wound Healing Activity Using Excision Wound Model in Diabetic Swiss Albino Mice.

The study developed a simple and inexpensive method to induce biofilm formation in-vivo for the evaluation of the antibiofilm activity of pharmacological agents using Swiss albino mice. Animals were made diabetic using streptozocin and nicotinamide. A cover slip containing preformed biofilm along with MRSA culture was introduced into the excision wound in these animals. The method was effective in developing biofilm on the coverslip after 24 h incubation in MRSA broth which was confirmed by microscopic examination and a crystal violet assay. Application of preformed biofilm along with microbial culture induced a profound infection with biofilm formation on excision wounds in 72 h. This was confirmed by macroscopic, histological, and bacterial load determination. Mupirocin, a known antibacterial agent effective against MRSA was used to demonstrate antibiofilm activity. Mupirocin was able to completely heal the excised wounds in 19 to 21 days while in the base-treated group, healing took place between 30 and 35 days. The method described is robust and can be reproduced easily without the use of transgenic animals and sophisticated methods such as confocal microscopy.

Anti-tubercular activity evaluation of natural compounds by targeting Mycobacterium tuberculosis resuscitation promoting factor B inhibition: An in silico study.

Tuberculosis (TB), an infectious disease caused by the Mycobacterium tuberculosis (Mtb), has been responsible for the deaths of millions of individuals around the globe. A vital protein in viral pathogenesis known as resuscitation promoting factor (RpfB) has been identified as a potential therapeutic target of anti-tuberculosis drugs. This study offered an in silico process to examine possible RpfB inhibitors employing a computational drug design pipeline. In this study, a total of 1228 phytomolecules were virtually tested against the RpfB of Mtb. These phytomolecules were sourced from the NP-lib database of the MTi-OpenScreen server, and five top hits (ZINC000044404209, ZINC000059779788, ZINC000001562130, ZINC000014766825, and ZINC000043552589) were prioritized for compute intensive docking with dock score ≤ - 8.5 kcal/mole. Later, molecular dynamics (MD) simulation and principal component analysis (PCA) were used to validate these top five hits. In the list of these top five hits, the ligands ZINC000044404209, ZINC000059779788, and ZINC000043552589 showed hydrogen bond formation with the functional residue Glu292 of the RpfB protein suggesting biological significance of the binding. The RMSD study showed stable protein-ligand complexes and higher conformational consistency for the ligands ZINC000014766825, and ZINC000043552589 with RMSD 3-4 Å during 100 ns MD simulation. The overall analysis performed in the study suggested promising binding of these compounds with the RpfB protein of the Mtb at its functional site, further experimental investigation is needed to validate the computational finding.

Fabrication, assessment, and optimization of alendronate sodium nanoemulsion-based injectable in-situ gel formulation for management of osteoporosis.

Low bone mass, degeneration of bone tissue, and disruption of bone microarchitecture are all symptoms of the disease osteoporosis, which can decrease bone strength and increase the risk of fractures. The main objective of the current study was to use a phospholipid-based phase separation in-situ gel (PPSG) in combination with an alendronate sodium nanoemulsion (ALS-NE) to help prevent bone resorption in rats. The effect of factors such as concentrations of the ALS aqueous solution, surfactant Plurol Oleique CC 497, and Maisine CC oil on nanoemulsion characteristics such as stability index and globular size was investigated using an l-optimal coordinate exchange statistical design. Injectable PPSG with the best nanoemulsion formulation was tested for viscosity, gel strength, water absorption, and in-vitro ALS release. ALS retention in the rats' muscles was measured after 30 days. The droplet size and stability index of the optimal nanoemulsion were 90 ± 2.0 nm and 85 ± 1.9%, respectively. When mixed with water, the optimal ALS-NE-loaded PPSG became viscous and achieved 36 seconds of gel strength, which was adequate for an injectable in-situ formulation. In comparison with the ALS solution-loaded in-situ gel, the newly created optimal ALS-NE-loaded PPSG produced the sustained and regulated release of ALS; hence, a higher percentage of ALS remained in rats' muscles after 30 days. PPSG that has been loaded with an ALS-NE may therefore be a more auspicious, productive, and effective platform for osteoporosis treatment than conventional oral forms.

Antibacterial Activity of Syzygium aromaticum (Clove) Bud Oil and Its Interaction with Imipenem in Controlling Wound Infections in Rats Caused by Methicillin-Resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the leading causes of infection worldwide. Clove oil's ability to inhibit the growth of MRSA was studied through in vitro and in vivo studies. The phytochemical components of clove oil were determined through gas chromatography-mass spectrometry (GC-MS) analysis. The antibacterial effects of clove oil and its interaction with imipenem were determined by studying MIC, MBC, and FIC indices in vitro. The in vivo wound-healing effect of the clove oil and infection control were determined using excision wound model rats. The GC-MS analysis of clove oil revealed the presence of 16 volatile compounds. Clove oil showed a good antibacterial effect in vitro but no interaction was observed with imipenem. Clove bud oil alone or in combination with imipenem healed wounds faster and reduced the microbial load in wounds. The findings of this study confirmed the antibacterial activity of clove oil in vitro and in vivo and demonstrated its interaction with imipenem.

In-Vitro and In-Vivo Antibacterial Effects of Frankincense Oil and Its Interaction with Some Antibiotics against Multidrug-Resistant Pathogens.

Frankincense (Boswellia sacra oleo gum resin) is reported to possess antimicrobial activity against several pathogens in-vitro. The antimicrobial effects of frankincense oil and its interaction with imipenem and gentamicin against methicillin-resistant Staphylococcus aureus (MRSA) and multidrug-resistant P. aeruginosa were determined through in-vitro methods and an in-vivo study using a rat pneumonia model. Frankincense oil was subjected to GC-MS analysis to determine the different volatile components. Antibacterial effects against MRSA and MDR-P. aeruginosa was evaluated and its MIC and MBC were determined. For the rat pneumonia model (in-vivo), oil was administered at a dose of 500 mg/kg and 1000 mg/kg followed by determination of CFU in lung tissue and histological studies. Frankincense oil did not show a very potent inhibitory effect against MRSA or MDR-P. aeruginosa; the oil did not affect the zone of inhibition or FIC when combined with imipenem or gentamicin indicating a lack of interaction between the oil and the antibiotics. Furthermore, there was no interaction between the antibiotics and the frankincense oil in the in-vivo model. The result of the study revealed that frankincense oil has a weak inhibitory effect against MRSA and MDR-P. aeruginosa, and it did not show any interaction with imipenem or gentamicin.

Carbohydrates from Pseudomonas aeruginosa biofilms interact with immune C-type lectins and interfere with their receptor function.

Bacterial biofilms represent a challenge to the healthcare system because of their resilience against antimicrobials and immune attack. Biofilms consist of bacterial aggregates embedded in an extracellular polymeric substance (EPS) composed of polysaccharides, nucleic acids and proteins. We hypothesised that carbohydrates could contribute to immune recognition of Pseudomonas aeruginosa biofilms by engaging C-type lectins. Here we show binding of Dendritic Cell-Specific Intercellular adhesion molecule-3-Grabbing Non-integrin (DC-SIGN, CD209), mannose receptor (MR, CD206) and Dectin-2 to P. aeruginosa biofilms. We also demonstrate that DC-SIGN, unlike MR and Dectin-2, recognises planktonic P. aeruginosa cultures and this interaction depends on the presence of the common polysaccharide antigen. Within biofilms DC-SIGN, Dectin-2 and MR ligands appear as discrete clusters with dispersed DC-SIGN ligands also found among bacterial aggregates. DC-SIGN, MR and Dectin-2 bind to carbohydrates purified from P. aeruginosa biofilms, particularly the high molecular weight fraction (HMW; >132,000 Da), with KDs in the nM range. These HMW carbohydrates contain 74.9-80.9% mannose, display α-mannan segments, interfere with the endocytic activity of cell-associated DC-SIGN and MR and inhibit Dectin-2-mediated cellular activation. In addition, biofilm carbohydrates reduce the association of the DC-SIGN ligand Lewisx, but not fucose, to human monocyte-derived dendritic cells (moDCs), and alter moDC morphology without affecting early cytokine production in response to lipopolysaccharide or P. aeruginosa cultures. This work identifies the presence of ligands for three important C-type lectins within P. aeruginosa biofilm structures and purified biofilm carbohydrates and highlights the potential for these receptors to impact immunity to P. aeruginosa infection.

Gamma Interferon and Interleukin-17A Differentially Influence the Response of Human Macrophages and Neutrophils to Pseudomonas aeruginosa Infection.

Macrophages are important orchestrators of inflammation during bacterial infection, acting as both effector cells and regulators of neutrophil recruitment and life span. Differently activated macrophage populations with distinct inflammatory and microbicidal potentials have been described. Our previous work unveiled a positive and a negative correlation between levels of gamma interferon (IFN-γ) and interleukin-17A (IL-17A), respectively, and lung function in cystic fibrosis, particularly in patients chronically infected with Pseudomonas aeruginosa This study sought to define key parameters in human antibacterial immunity under Th1- and Th17-dominated inflammatory conditions; the final aim was to identify unique characteristics that could be fine-tuned therapeutically to minimize tissue damage while maximizing bacterial clearance. Toward this aim, neutrophils were incorporated into cultures of macrophages treated with IFN-γ or IL-17A and infected with P. aeruginosa The intent of this design was to model (i) initiation of inflammation by infected macrophages and (ii) delayed arrival of neutrophils and their exposure to macrophage-derived cytokines. Under these conditions, IFN-γ decreased bacterial killing and promoted the production of monocyte chemoattractant protein 1 (MCP-1). In contrast, IL-17A promoted bacterial killing but did not affect MCP-1 production. The level of secretion of the pyrogen IL-1ß was significantly lower in the presence of IFN-γ than in the presence of IL-17A and correlated with levels of the IL1B transcript in infected macrophages. These findings support the validity of this model to investigate human antibacterial immunity. Based on these observations, the protective and damaging roles of IFN-γ and IL-17A, respectively, during P. aeruginosa infection could be caused by their contrasting effects on IL-1ß and MCP-1 production.