Unveiling Novel Urease Inhibitors for Helicobacter pylori: A Multi-Methodological Approach from Virtual Screening and ADME to Molecular Dynamics Simulations.

Helicobacter pylori (Hp) infections pose a global health challenge demanding innovative therapeutic strategies by which to eradicate them. Urease, a key Hp virulence factor hydrolyzes urea, facilitating bacterial survival in the acidic gastric environment. In this study, a multi-methodological approach combining pharmacophore- and structure-based virtual screening, molecular dynamics simulations, and MM-GBSA calculations was employed to identify novel inhibitors for Hp urease (HpU). A refined dataset of 8,271,505 small molecules from the ZINC15 database underwent pharmacokinetic and physicochemical filtering, resulting in 16% of compounds for pharmacophore-based virtual screening. Molecular docking simulations were performed in successive stages, utilizing HTVS, SP, and XP algorithms. Subsequent energetic re-scoring with MM-GBSA identified promising candidates interacting with distinct urease variants. Lys219, a residue critical for urea catalysis at the urease binding site, can manifest in two forms, neutral (LYN) or carbamylated (KCX). Notably, the evaluated molecules demonstrated different interaction and energetic patterns in both protein variants. Further evaluation through ADMET predictions highlighted compounds with favorable pharmacological profiles, leading to the identification of 15 candidates. Molecular dynamics simulations revealed comparable structural stability to the control DJM, with candidates 5, 8 and 12 (CA5, CA8, and CA12, respectively) exhibiting the lowest binding free energies. These inhibitors suggest a chelating capacity that is crucial for urease inhibition. The analysis underscores the potential of CA5, CA8, and CA12 as novel HpU inhibitors. Finally, we compare our candidates with the chemical space of urease inhibitors finding physicochemical similarities with potent agents such as thiourea.

Microneedles for oral mucosal delivery - Current trends and perspective on future directions.

INTRODUCTION: Oral cavity drug and vaccine delivery has the potential for local targeting, dose reduction, minimization of systemic side effects, and generation of mucosal immunity. To overcome current limitations of delivery into the oral cavity mucosa, microneedles (MNs) have emerged as a promising technology. AREAS COVERED: We reviewed the literature on MN application in the oral cavity, including in vitro studies, in vivo animal studies, and human clinical trials. EXPERT OPINION: MNs are sufficiently robust to cross the oral cavity epithelium and nearly painless when applied to different parts of the human oral mucosa including the lip, cheek, tongue, and palate. In recent years, MNs have been evaluated for different applications, including vaccination, topical anesthetic delivery, and treatment of local oral pathologies such as oral lesions or carcinomas. MNs are attractive because they have the potential to produce a better treatment outcome with reduced side effects. Over the coming years, we project a significant increase in research related to the development of MNs for use in dentistry and other medical conditions of the mouth.

Cost-Effective Pipeline for a Rational Design and Selection of Capsaicin Analogues Targeting TRPV1 Channels.

Transient receptor potential (TRP) channels constitute a large group of membrane receptors associated with sensory pathways in vertebrates. One of the most studied is TRPV1, a polymodal receptor tuned for detecting heat and pungent compounds. Specific inhibition of the nociceptive transduction at the peripheral nerve represents a convenient approach to pain relief. While acting as a chemoreceptor, TRPV1 shows high sensitivity and selectivity for capsaicin. In contrast to the drugs available on the market that target the inflammatory system, TRPV1 antagonists act as negative modulators of nociceptive transduction. Therefore, the development of compounds modulating TRPV1 activity has expanded dramatically over time. Experimental data suggest that most agonist and antagonist drugs interact at or near capsaicin's binding site. In particular, the properties of capsaicin's head play an essential role in modulating potency and affinity. Here, we explored a cost-efficient pipeline to predict the effects of introducing chemical modifications into capsaicin's head region. An extensive set of molecules was selected by first considering the geometrical properties of capsaicin's binding site and then molecular docking. Finally, the novel ligands were ranked by combining molecular and pharmacokinetic predictions.

Polymicrobial biofilms related to dental implant diseases: unravelling the critical role of extracellular biofilm matrix.

Biofilms are complex tri-dimensional structures that encase microbial cells in an extracellular matrix comprising self-produced polymeric substances. The matrix rich in extracellular polymeric substance (EPS) contributes to the unique features of biofilm lifestyle and structure, enhancing microbial accretion, biofilm virulence, and antimicrobial resistance. The role of the EPS matrix of biofilms growing on biotic surfaces, especially dental surfaces, is largely unravelled. To date, there is a lack of a broad overview of existing literature concerning the relationship between the EPS matrix and the dental implant environment and its role in implant-related infections. Here, we discuss recent advances in the critical role of the EPS matrix on biofilm growth and virulence on the dental implant surface and its effect on the etiopathogenesis and progression of implant-related infections. Similar to other biofilms associated with human diseases/conditions, EPS-enriched biofilms on implant surfaces promote microbial accumulation, microbiological shift, cross-kingdom interaction, antimicrobial resistance, biofilm virulence, and, consequently, peri-implant tissue damage. But intriguingly, the protagonism of EPS role on implant-related infections and the development of matrix-target therapeutic strategies has been neglected. Finally, we highlight the need for more in-depth analyses of polymicrobial interactions within EPS matrix and EPS-targeting technologies' rationale for disrupting the complex biofilm microenvironment with more outstanding translation to implant applications in the near future.

Bacterial-derived extracellular polysaccharides reduce antimicrobial susceptibility on biotic and abiotic surfaces.

OBJECTIVE: Extracellular biofilm matrix plays a role in reducing bacterial susceptibility against antimicrobials. Since the surface where biofilm is growing modulates microbial accumulation and bacterial-derived exopolysaccharides (EPS) synthesis, this study compared the role of EPS to reduce antimicrobial susceptibility on biotic (dental surface) and abiotic (titanium (Ti) material) surfaces and the effect of remaining matrix-enriched biofilms to promote bacterial recolonization. DESIGN: 48 h Streptococcus mutans UA159 strain biofilms were grown on enamel and Ti surfaces. The medium was supplemented with 1% sucrose, substrate for EPS synthesis, or with 0.5% glucose + 0.5% fructose as control. Chlorhexidine (CHX) 0.2% was used for antimicrobial treatment. Biofilms were collected and the following analyses were considered: viable bacterial counts, biofilm pH, EPS content, and biofilm structure by scanning electron microscopy and confocal laser scanning microscopy (CLSM). Substrate surfaces were analyzed by 3D laser scanning confocal microscope. RESULTS: Enamel surface showed a higher amount of EPS content (p < 0.05), which may be explained by the higher bacterial biomass compared to Ti material. EPS content reduced bacterial susceptibility against antimicrobial treatments for both substrates, compared to EPS control (p < 0.05). However, sucrose-treated cells presented the same magnitude of reduction for Ti or enamel. Interestingly, matrix-enriched biofilms favored bacterial recolonization for both substrates. CONCLUSION: The surface where the biofilm is growing modulates the amount of EPS synthesized and matrix content plays a key role in reducing antimicrobial susceptibility and promoting bacterial recolonization.

The Use of Essential Oils and Their Isolated Compounds for the Treatment of Oral Candidiasis: A Literature Review.

In this literature review, we present the main scientific findings on the antifungal activity of essential oils (EOs) applicable for a new drug formulation to treat oral candidiasis. Seven literature databases were systematically searched for eligible in vitro and clinical trials. Selected articles were screened for biological activity, botanical species, phytochemical composition, study design, and methodological quality. A total of 26 articles were included in the review, of which 21 were in vitro studies and 5 clinical trials. The most promising EOs were obtained from Allium tubeorosum, Cinnamomum cassia, Cinnamomum zeylanicum, and Coriandrum sativum L. Among the phytochemicals, citral and thymol were the most active. Clinical trials indicated that the EOs from Pelargonium graveolens and Zataria multiflora are potentially effective to treat oral candidiasis. Further nonclinical and clinical studies with these EO are warranted to determine their potential use and safety for the treatment of oral candidiasis.

Malva sylvestris derivatives as inhibitors of HIV-1 BaL infection.

The emphasis of the present study is to evaluate a natural product and the potential microbicide activity using a dual chamber infection method. Malva sylvestris extracts and fractions were screened for anti-HIV activity by measuring the virus-antibody neutralization. Plant extracts with strong antiviral activity working in nanomolar or picomolar range can be used to enhance the activity of synthetic compounds and work as anti-HIV agents. The aqueous fraction (AF) of M. sylvestris demonstrated antiviral activity in a model with epithelial and blood cell lines. The AF showed an effective antiviral potential on the TZM-bl cells with reduction scores higher than 60% of infectivity. Quantification of p24 in the supernatant of the co-culture model demonstrated a reduction in the number of viral particles after AF treatment (p < 0.05). Cytokines were quantified and all signaling inflammatory markers; IL1-alpha, IL-beta, IL-6, IL-8 and GM-CSF (p < 0.05) were modulated by positive control and AF treatments. In particular, IL-6 had lower levels of expression in Malva groups when compared to the Zidovudine positive control group. Natural occurring derivatives of M. sylvestris demonstrated to work inhibiting reverse transcriptase enzyme action. M. sylvestris contains highly potential anti-HIV-1 BaL components and may be considered a potential source for new formulations in the development of topical microbicides.

Potential Novel Strategies for the Treatment of Dental Pulp-Derived Pain: Pharmacological Approaches and Beyond.

The diagnosis and management of pain is an everyday occurrence in dentistry, and its effective control is essential to ensure the wellbeing of patients. Most tooth-associated pain originates from the dental pulp, a highly vascularized and innervated tissue, which is encased within mineralized dentin. It plays a crucial role in the sensing of stimuli from the local environment, such as infections (i.e. dental caries) and traumatic injury, leading to a local inflammatory response and subsequently to an increase in intra-pulp pressure, activating nerve endings. However, thermal, chemical, and mechanical stimuli also have the ability to generate dental pulp pain, which presents mechanisms highly specific to this tissue and which have to be considered in pain management. Traditionally, the management of dental pulp pain has mostly been pharmacological, using non-steroidal anti-inflammatory drugs (NSAIDs) and opioids, or restorative (i.e. removal of dental caries), or a combination of both. Both research areas continuously present novel and creative approaches. This includes the modulation of thermo-sensitive transient receptor potential cation channels (TRP) by newly designed drugs in pharmacological research, as well as the use of novel biomaterials, stem cells, exosomes and physical stimulation to obtain pulp regeneration in regenerative medicine. Therefore, the aim of this review is to present an up-to-date account of causes underlying dental pain, novel treatments involving the control of pain and inflammation and the induction of pulp regeneration, as well as insights in pain in dentistry from the physiological, pharmacological, regenerative and clinical perspectives.

Assessment of the bioavailability of an antibiotic prophylactic protocol in patients undergoing third molar surgery.

The aim of this prospective study was to determine the antibiotic bioavailability of a prophylactic protocol in patients undergoing third molar surgery. Samples from 25 patients were analysed (average age 21 ± 3.89 years, range 18-33 years; 14 female). The patients received single-dose prophylaxis of 2 g amoxicillin orally 1 hour prior to third molar surgery. Venous blood (1.5 ml) and blood from the third molar socket (1.50 ml) were obtained. The amoxicillin plasma concentration was determined in both samples by high performance liquid chromatography with a diode-array detector (HPLC/DAD). Their associations with demographic variables (age, height, weight, body mass index (BMI), sex) and antibiotic exposure time were analyzed using linear regression models. The mean amoxicillin plasma level detected in the venous blood was 1.21 ± 1.17 µg/ml (range 0.49-6.34 µg/ml) and in the third molar socket was 4.14 ± 2.24 µg/ml (range 0.86-7.46 µg/ml) (P < 0.001). No relationship was observed between the bioavailability of the drug and the patient biometric indices evaluated. The prophylactic administration of 2 g amoxicillin in third molar surgery showed greater bioavailability in the molar socket than the concentrations established as necessary to inhibit the growth of microorganisms that cause oral infections. The results show the need to review the current infection control protocols in oral surgery in light of the overestimated doses observed.

Chalcone derivatives as non-canonical ligands of TRPV1.

Transient receptor potential vanilloid 1 (TRPV1) is a polymodal cation channel activated by heat, voltage, and ligands. Also known as the capsaicin receptor, TRPV1 is expressed in numerous tissues by different cell types, including peripheral sensory fibers where acts as a thermal and chemical detector in nociceptive pathways. TRPV1 channels are able to bind a wide range of ligands, including a number of vanilloid derivatives all modulating channel's activity. When expressed by sensory neurons, activation of TRPV1 channels by heat (>40 °C), capsaicin (sub-micromolar), or acid environment (pH < 6), causes depolarization leading to burning pain sensation in mammals. Naturally occurring chalcones (1,3-diaryl-2-propen-1-ones) have been reported as effective inhibitors of TRPV1. Their relatively simple chemical structure and the possibility for handy chemical modification make them attractive ligands for the treatment of peripheral pain. By taking advantage of the structural information available, here we discuss pharmacological properties of chalcones and their putative mechanism of binding to TRPV1 channels.

Fungal-Host Interaction: Curcumin Modulates Proteolytic Enzyme Activity of Candida albicans and Inflammatory Host Response In Vitro.

Current treatments for Candida albicans infection are limited due to the limited number of antifungal drugs available and the increase in antifungal resistance. Curcumin is used as a spice, food preservative, flavoring, and coloring agent that has been shown to have many pharmacological activities. Thus, this study evaluated the modulatory effects of curcumin on major virulence factors associated with the pathogenicity of C. albicans. The minimum inhibitory concentration (MIC) of curcumin against C. albicans (SC5314) was determined. Biofilm formation was quantified and the proteinase and phospholipase secretion was measured. The cytotoxicity was tested in oral fibroblast cells. A cocultured model was used to analyze the gene expression of proinflammatory cytokines (IL-1ß, IL-1α, and IL-6) from host cells, as well SAP-1 and PLB-1 by RT-PCR. The MIC was between 6.25 and 12.5 µM, and the activity of proteinase enzyme was significantly decreased in biofilms treated with curcumin. However, proteinase gene expression was not downregulated after curcumin treatment. Furthermore, gene expressions of host inflammatory response, IL-1ß and IL-1α, were significantly downregulated after exposure to curcumin. In conclusion, curcumin exhibited antifungal activity against C. albicans and modulated the proteolytic enzyme activities without downregulating the gene expression. In host inflammatory response, curcumin downregulated IL-1ß and IL-1α gene expression.

Single-Dose Bioavailability for Prophylactic Coverage in Patients Undergoing Dental Implant Surgery.

PURPOSE: The use of antibiotic prophylaxis in dental implant surgery is a routine dental practice to prevent implant failure. Current systematic reviews have yet to confirm the effectiveness of antibiotic treatment at reducing postoperative infection rates in implant procedures. The aim of this study was to quantify amoxicillin plasma levels at the peripheral venous blood and at the surgical implant site in patients undergoing dental treatment. MATERIALS AND METHODS: A prospective study was conducted. Patients undergoing dental implant surgery were selected, and preoperative doses of 1 g of amoxicillin, in the form of an oral tablet, were administered 1 hour prior to implant surgery. The blood samples (1.5 mL) were collected from both the venous and implant sites. Data on the antibiotics were measured using high-pressure liquid chromatography. The quantification method was validated for linearity, selectivity, and detection limits. Statistical analysis of plasma antibiotic was performed, comparing both samples and the age, height, weight, and body mass index (BMI) (Wilcoxon test, P < .05; STATA v.10.0). RESULTS: Thirty-two patients (mean age: 56.28 ± 12.2 years; range: 31 to 79 years; 20 men) were selected. Both blood samples were taken at a mean 1.16 ± 0.37 hours after antibiotic intake. The mean amoxicillin concentration at the implant site reached 5 ± 2.63 µg/mL (range: 2.04 to 11.18 µg/mL), while the venous blood level was 4.21 ± 2.12 µg/mL (range: 1.6 to 9.98 µg/mL) (P = .33). No correlations were observed between age, height, weight, and BMI with antibiotic plasma levels. CONCLUSION: These findings demonstrate that plasma concentrations of a prophylactic dose of amoxicillin are higher than the minimum inhibitory concentration needed to prevent the common dental bacteria involved in peri-implantitis and periodontal diseases.

Chemical Characterization and Antioxidant, Antimicrobial, and Anti-Inflammatory Activities of South Brazilian Organic Propolis.

South Brazilian organic propolis (OP), which has never been studied before, was assessed and its chemical composition, scavenging potential of reactive oxygen species, antimicrobial and anti-inflammatory activities are herein presented. Based on the chemical profile obtained using HPLC, OP was grouped into seven variants (OP1-OP7) and all of them exhibited high scavenging activity, mainly against superoxide and hypochlorous acid species. OP1, OP2, and OP3 had the smallest minimal inhibitory concentration (MIC) against Gram-positive bacteria Streptococcus mutans, Streptococcus oralis, and Streptococcus aureus (12.5-100 µg/mL). OP1, OP2, OP3, and OP4 were more effective against Pseudomonas aeruginosa (Gram-negative), with MIC values ranging from 100 to 200 µg/mL. OP6 showed anti-inflammatory activity by decreasing NF-kB activation and TNF-α release in RAW 264.7 macrophages, and expressing the NF-κB-luciferase reporter stable gene. Therefore, south Brazilian OP can be considered an excellent source of bioactive compounds with great potential of application in the pharmaceutical and food industry.

Anti-Inflammatory, Anti-Osteoclastogenic and Antioxidant Effects of Malva sylvestris Extract and Fractions: In Vitro and In Vivo Studies.

Given their medical importance, natural products represent a tremendous source of drug discovery. The aim of this study was to investigate Malva sylvestris L. extract and fractions and their pharmacological activities followed by chemical identification. The aqueous fraction (AF) was identified as the bioactive fraction in the in vitro and in vivo assays. The AF controlled the neutrophil migration to the peritoneal cavity by 66%, inhibited the antiedematogenic activity by 58.8%, and controlled IL-1ß cytokine expression by 54%. The in vitro viability tests showed a concentration-dependent effect, where the MSE and fractions at concentrations under 10 µg/mL were non-toxic to cells. Transcriptional factors of carbonic anhydrase II (CAII), cathepsin K (Ctsk) and tartrate-resistant acid phosphatase (TRAP) were analyzed by qPCR in RAW 264.7 cell lines. The gene expression analysis showed that the AF was the only treatment that could downregulate all the study genes: CAII, Ctsk and TRAP (p<0.05). TRAP staining was used to evaluate osteoclast formation. AF treatments reduced the number of osteoclastogenesis 2.6-fold compared to the vehicle control group. Matrix metalloproteinase 9 (MMP-9) activity decreased 75% with the AF treatment. Moreover, the bioactive fraction had the ability to regulate the oxidation pathway in the ABTS (2,2-Azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) assay with an activity equivalent to 1.30 µmol Trolox/g and DPPH (2,2-diphenyl-1-picrylhydrazyl) radicals 1.01 g/L. Positive ion ESI-mass spectrometry for molecular ions at m/z 611 and 633 confirmed rutin as the major compound in the AF. The AF of M. sylvestris presented anti-inflammatory, controlled osteoclastogenic mechanisms and antioxidant abilities in different in vitro and in vivo methods. In addition, we suggest that given its multi-target activity the bioactive fraction may be a good candidate in the therapy of chronic inflammatory diseases.

In vitro evaluation of antifungal activity of monolaurin against Candida albicans biofilms.

Monolaurin (also known as glycerol monolaurate) is a natural compound found in coconut oil and is known for its protective biological activities as an antimicrobial agent. The nature of oral candidiasis and the increased antifungal resistance demand the search for novel antifungal therapeutic agents. In this study, we examine the antifungal activity of monolaurin against Candida albicans biofilms (strain ATCC:SC5314/MYA2876) in vitro and investigate whether monolaurin can alter gene expression of host inflammatory cytokines, IL-1α and IL-1ß. In a co-culture model, oral fibroblast cells were cultured simultaneously with C. albicans for 24 hrs followed by the exposure to treatments of monolaurin (3.9-2,500 µM), positive control fluconazole (32.2 µM), and vehicle control group (1% ethanol), which was a model used to evaluate the cytotoxicity of monolaurin on fibroblasts as well as to analyze morphological characteristics of biofilms through fluorescence microscopy. In addition, the co-culture model was used for RNA extraction of oral fibroblasts to assess gene expression of host inflammatory cytokines, using quantitative real-time PCR. Our results showed the MIC and MFC of monolaurin were in the range 62.5-125 µM and 125-250 µM, respectively. Biofilm antifungal assay showed significant reduction in Log (CFU/ml) of biofilms treated with 1,250 and 2,500 µM of 1-monolaurin when compared to the control groups . There was also a significant down-regulation of IL-1α and IL-1ß in the co-culture treated with monolaurin. It can be concluded that monolaurin has a potential antifungal activity against C. albicans and can modulate the pro-inflammatory response of the host.

In Vitro and In Vivo Antifungal Activity of Lichochalcone-A against Candida albicans Biofilms.

Oral candidiasis (OC) is an opportunistic fungal infection with high prevalence among immunocompromised patients. Candida albicans is the most common fungal pathogen responsible for OC, often manifested in denture stomatitis and oral thrush. Virulence factors, such as biofilms formation and secretion of proteolytic enzymes, are key components in the pathogenicity of C. albicans. Given the limited number of available antifungal therapies and the increase in antifungal resistance, demand the search for new safe and effective antifungal treatments. Lichochalcone-A is a polyphenol natural compound, known for its broad protective activities, as an antimicrobial agent. In this study, we investigated the antifungal activity of lichochalcone-A against C. albicans biofilms both in vitro and in vivo. Lichochalcone-A (625 µM; equivalent to 10x MIC) significantly reduced C. albicans (MYA 2876) biofilm growth compared to the vehicle control group (1% ethanol), as indicated by the reduction in the colony formation unit (CFU)/ml/g of biofilm dry weight. Furthermore, proteolytic enzymatic activities of proteinases and phospholipases, secreted by C. albicans were significantly decreased in the lichochalcone-A treated biofilms. In vivo model utilized longitudinal imaging of OC fungal load using a bioluminescent-engineered C. albicans (SKCa23-ActgLUC) and coelenterazine substrate. Mice treated with lichochalcone-A topical treatments exhibited a significant reduction in total photon flux over 4 and 5 days post-infection. Similarly, ex vivo analysis of tongue samples, showed a significant decrease in CFU/ml/mg in tongue tissue sample of lichochalcone-A treated group, which suggest the potential of lichochalcone-A as a novel antifungal agent for future clinical use.