Silversol® (a Colloidal Nanosilver Formulation) Inhibits Growth of Antibiotic-Resistant Staphylococcus aureus by Disrupting Its Physiology in Multiple Ways.

Antibiotic-resistant strains of Staphylococcus aureus are being viewed as a serious threat by various public health agencies. Identifying novel targets in this important pathogen is crucial to the development of new effective antibacterial formulations. We investigated the antibacterial effect of a colloidal nanosilver formulation, Silversol®, against an antibiotic-resistant strain of S. aureus using appropriate in vitro assays. Moreover, we deciphered the molecular mechanisms underlying this formulation's anti-S. aureus activity using whole transcriptome analysis. Lower concentrations of the test formulation exerted a bacteriostatic effect against this pathogen, and higher concentrations exerted a bactericidal effect. Silversol® at sub-lethal concentration was found to disturb multiple physiological traits of S. aureus such as growth, antibiotic susceptibility, membrane permeability, efflux, protein synthesis and export, biofilm and exopolysaccharide production, etc. Transcriptome data revealed that the genes coding for transcriptional regulators, efflux machinery, transferases, ß-lactam resistance, oxidoreductases, metal homeostasis, virulence factors, and arginine biosynthesis are expressed differently under the influence of the test formulation. Genes (argG and argH) involved in arginine biosynthesis emerged among the major targets of Silversol®'s antibacterial activity against S. aureus.

Current insights into transcriptional role(s) for the nutraceutical Withania somnifera in inflammation and aging.

The health-beneficial effects of nutraceuticals in various diseases have received enhanced attention in recent years. Aging is a continuous process wherein physiological activity of an individual declines over time and is characterized by various indefinite hallmarks which contribute toward aging-related comorbidities in an individual which include many neurodegenerative diseases, cardiac problems, diabetes, bone-degeneration, and cancer. Cellular senescence is a homeostatic biological process that has an important function in driving aging. Currently, a growing body of evidence substantiates the connection between epigenetic modifications and the aging process, along with aging-related diseases. These modifications are now being recognized as promising targets for emerging therapeutic interventions. Considering that almost all the biological processes are modulated by RNAs, numerous RNA-binding proteins have been found to be linked to aging and age-related complexities. Currently, studies have shed light on the ability of the nutraceutical Withania somnifera (Ashwagandha) to influence RNA expression, stability, and processing, offering insights into its mechanisms of action. By targeting RNA-related pathways, Withania somnifera may exhibit promising effects in ameliorating age-associated molecular changes, which include modifications in gene expression and signaling networks. This review summarizes the potential role of Withania somnifera as a nutraceutical in modulating RNA-level changes associated with aging, encompassing both in vitro and in vivo studies. Taken together, the putative role(s) of Withania in modulation of key RNAs will provide insights into understanding the aging process and facilitate the development of various preventive and therapeutic strategies employing nutraceuticals for healthy aging.

Emerging Vistas for the Nutraceutical Withania somnifera in Inflammaging.

Inflammaging, a coexistence of inflammation and aging, is a persistent, systemic, low-grade inflammation seen in the geriatric population. Various natural compounds have been greatly explored for their potential role in preventing and treating inflammaging. Withania somnifera has been used for thousands of years in traditional medicine as a nutraceutical for its numerous health benefits including regenerative and adaptogenic effects. Recent preclinical and clinical studies on the role of Withania somnifera and its active compounds in treating aging, inflammation, and oxidative stress have shown promise for its use in healthy aging. We discuss the chemistry of Withania somnifera, the etiology of inflammaging and the protective role(s) of Withania somnifera in inflammaging in key organ systems including brain, lung, kidney, and liver as well as the mechanistic underpinning of these effects. Furthermore, we elucidate the beneficial effects of Withania somnifera in oxidative stress/DNA damage, immunomodulation, COVID-19, and the microbiome. We also delineate a putative protein-protein interaction network of key biomarkers modulated by Withania somnifera in inflammaging. In addition, we review the safety/potential toxicity of Withania somnifera as well as global clinical trials on Withania somnifera. Taken together, this is a synthetic review on the beneficial effects of Withania somnifera in inflammaging and highlights the potential of Withania somnifera in improving the health-related quality of life (HRQoL) in the aging population worldwide.

Sub-lethal concentration of a colloidal nanosilver formulation (Silversol®) triggers dysregulation of iron homeostasis and nitrogen metabolism in multidrug resistant Pseudomonas aeruginosa.

BACKGROUND: Pseudomonas aeruginosa is a notorious pathogen. Its multidrug resistant strains are listed among priority pathogens against whom discovery of novel antibacterial agents and, elucidation of new anti-pathogenicity mechanisms are urgently warranted. This study describes multiple antibacterial effects of a colloidal nano-silver formulation- Silversol® against a multi-drug resistant strain of P. aeruginosa. RESULTS: Minimum inhibitory concentration (MIC) of Silversol® against P. aeruginosa was found to be 1.5 ppm; and at sub-MIC of 1 ppm, it was able to alter quorum-sensing regulated pigmentation (pyocanin 82%↓; pyoverdine 48%↑), exopolysaccharide synthesis (76%↑) and biofilm formation, susceptibility to antibiotics (streptomycin and augmentin), protein synthesis and export (65%↑), nitrogen metabolism (37%↑ nitrite accumulation), and siderophore production in this pathogen. Network analysis of the differentially expressed genes in the transcriptome of the silversol-treated bacterium identified ten genes as the potential molecular targets: norB, norD, nirS, nirF, nirM, nirQ, nosZ, nosY, narK1, and norE (all associated with nitrogen metabolism or denitrification). Three of them (norB, narK1, and norE) were also validated through RT-PCR. CONCLUSIONS: Generation of nitrosative stress and disturbance of iron homeostasis were found to be the major mechanisms associated with anti-Pseudomonas activity of Silversol®.

Potent anthelmintic activity of a colloidal nano-silver formulation (Silversol®) against the model worm Caenorhabditis elegans.

OBJECTIVE: In the background of a very small number of effective anthelmintics available today with a narrow activity spectrum, and a rise in resistance against them among parasitic helminths, there is an urgent need for discovery of novel broad-spectrum anthelmintics displaying no or minimal toxicity towards the host. Silver being used since centuries for therapeutic purposes and considered safe for human consumption, we investigated anthelmintic activity of a colloidal nanosilver formulation Silversol®. Anthelmintic efficacy of the test formulation was assayed employing the nematode Caenorhabditis elegans as a model worm through a live-dead count. RESULTS: Silversol® exerted anthelmintic action superior to one of the positive controls (Benzimidazole), and almost at par to another positive control (Ivermectin). At concentrations ≥ 2 ppm, it could kill all the worms present in the experimental well. Lower concentrations of silver were found to have a cuticle-damaging action on worms. Further investigation is warranted to assess whether Silversol® can exert similar potent activity against different species of parasitic helminths, and elucidate the underlying molecular mechanisms of action.

The in vitro efficacy of betadine antiseptic solution and colloidal silver gel combination in inhibiting the growth of bacterial biofilms.

INTRODUCTION: Betadine (Povidone-Iodine) solution is a topically applied antiseptic, which has been used routinely used in wound care and general surgery to prevent skin and wound infections. However, several studies have documented the ineffectiveness of betadine. Other topical antimicrobial dressings, including those that contain silver, have been used in the management of infected wounds. The present study was undertaken to determine if the combination of 5% betadine solution and silver colloidal gel (Ag-gel) is more effective than either substance alone in inhibiting the growth gram-negative and gram-positive bacteria. METHODS: The effectiveness of 5% betadine solution and Ag-gel as anti-microbial agents were assessed using both colony forming unit (CFU) assay and confocal laser scanning microscopy (CLSM). RESULTS: Ag-gel showed complete inhibition on all the bacteria species examined except the Klebsiella pneumoniae clinical isolate (CL) strain while 5% betadine concentrations did not completely kill any of the tested bacteria. In contrast, K. pneumoniae was completely eliminated in the presence of both 5% betadine solution and Ag-gel together. The CLSM showed similar findings to the CFU results examining the 5% betadine solution and Ag-gel combination. CONCLUSIONS: This study demonstrated that while the individual treatments using either 5% betadine solution and Ag-gel alone were infective antimicrobial agents, the combination of 5% betadine solution and Ag-gel was superior at eliminating all tested bacteria, including K. pneumoniae CL.

Current Insights into miRNA and lncRNA Dysregulation in Diabetes: Signal Transduction, Clinical Trials and Biomarker Discovery.

Diabetes is one of the most frequently occurring metabolic disorders, affecting almost one tenth of the global population. Despite advances in antihyperglycemic therapeutics, the management of diabetes is limited due to its complexity and associated comorbidities, including diabetic neuropathy, diabetic nephropathy and diabetic retinopathy. Noncoding RNAs (ncRNAs), including microRNAs (miRNAs) and long noncoding RNAs (lncRNAs), are involved in the regulation of gene expression as well as various disease pathways in humans. Several ncRNAs are dysregulated in diabetes and are responsible for modulating the expression of various genes that contribute to the 'symptom complex' in diabetes. We review various miRNAs and lncRNAs implicated in diabetes and delineate ncRNA biological networks as well as key ncRNA targets in diabetes. Further, we discuss the spatial regulation of ncRNAs and their role(s) as prognostic markers in diabetes. We also shed light on the molecular mechanisms of signal transduction with diabetes-associated ncRNAs and ncRNA-mediated epigenetic events. Lastly, we summarize clinical trials on diabetes-associated ncRNAs and discuss the functional relevance of the dysregulated ncRNA interactome in diabetes. This knowledge will facilitate the identification of putative biomarkers for the therapeutic management of diabetes and its comorbidities. Taken together, the elucidation of the architecture of signature ncRNA regulatory networks in diabetes may enable the identification of novel biomarkers in the discovery pipeline for diabetes, which may lead to better management of this metabolic disorder.

Molecular Pathways and Roles for Vitamin K2-7 as a Health-Beneficial Nutraceutical: Challenges and Opportunities.

Vitamin K2-7, also known as menaquinone-7 (MK-7) is a form of vitamin K that has health-beneficial effects in osteoporosis, cardiovascular disease, inflammation, cancer, Alzheimer's disease, diabetes and peripheral neuropathy. Compared to vitamin K1 (phylloquinone), K2-7 is absorbed more readily and is more bioavailable. Clinical studies have unequivocally demonstrated the utility of vitamin K2-7 supplementation in ameliorating peripheral neuropathy, reducing bone fracture risk and improving cardiovascular health. We examine how undercarboxylated osteocalcin (ucOC) and matrix Gla protein (ucMGP) are converted to carboxylated forms (cOC and cMGP respectively) by K2-7 acting as a cofactor, thus facilitating the deposition of calcium in bones and preventing vascular calcification. K2-7 is beneficial in managing bone loss because it upregulates osteoprotegerin which is a decoy receptor for RANK ligand (RANKL) thus inhibiting bone resorption. We also review the evidence for the health-beneficial outcomes of K2-7 in diabetes, peripheral neuropathy and Alzheimer's disease. In addition, we discuss the K2-7-mediated suppression of growth in cancer cells via cell-cycle arrest, autophagy and apoptosis. The mechanistic basis for the disease-modulating effects of K2-7 is mediated through various signal transduction pathways such as PI3K/AKT, MAP Kinase, JAK/STAT, NF-κB, etc. Interestingly, K2-7 is also responsible for suppression of proinflammatory mediators such as IL-1α, IL-1ß and TNF-α. We elucidate various genes modulated by K2-7 as well as the clinical pharmacometrics of vitamin K2-7 including K2-7-mediated pharmacokinetics/pharmacodynamics (PK/PD). Further, we discuss the current status of clinical trials on K2-7 that shed light on dosing strategies for maximum health benefits. Taken together, this is a synthetic review that delineates the health-beneficial effects of K2-7 in a clinical setting, highlights the molecular basis for these effects, elucidates the clinical pharmacokinetics of K2-7, and underscores the need for K2-7 supplementation in the global diet.

Vitamin K Insufficiency in the Indian Population: Pilot Observational Epidemiology Study.

BACKGROUND: The fat-soluble K vitamins K1 and K2 play an essential role in the blood coagulation cascade and are made available predominantly through selective dietary intakes. They are less known for their nonessential roles in a family of vitamin K-dependent proteins that promote various functions of organs and systems in the body. A lack of vitamin K can characterize vitamin and nutritional element insufficiency, which is different from a clinically apparent vitamin deficiency. OBJECTIVE: This epidemiological study evaluated the nutritional status of vitamin K in a sample of the Indian population and vitamin K content in staple Indian foods. METHODS: Serum levels of vitamin K1 and vitamin K2 in the form of menaquinone-7 (MK-7) were assessed via high-performance liquid chromatography coupled with fluorescence detection in 209 patients with type 2 diabetes, 50 healthy volunteers, and common staple foods in India. RESULTS: After comparing populations with high and low serum vitamin K levels from various geographical regions, our results indicated that the sample of healthy Indian individuals and the sample of Indian patients with type 2 diabetes had low (insufficient) levels of vitamin K2 (MK-7; range 0.3-0.4 ng/mL). No significant differences existed in vitamin K1-related and MK-7-related values between healthy male and female subjects, between male and female subjects with diabetes, and between the healthy sample and the sample of patients with diabetes. The staple, commonly consumed Indian foods that were tested in this study had undetectable levels of vitamin K2, while levels of vitamin K1 varied widely (range 0-37 µg/100 g). CONCLUSIONS: Based on our sample's low serum levels of vitamin K2 (MK-7) as well as the low levels of vitamin K2 in their typical diet, we propose that the general Indian population could benefit from the consumption of vitamin K2 in the form of MK-7 supplements. TRIAL REGISTRATION: Clinical Trials Registry - India CTRI/2019/05/014246; http://ctri.nic.in/Clinicaltrials/showallp.php?mid1=21660&EncHid=&userName=014246; Clinical Trials Registry - India CTRI/2019/03/018278; http://ctri.nic.in/Clinicaltrials/showallp.php?mid1=32349&EncHid=&userName=018278.

New Vistas in microRNA Regulatory Interactome in Neuropathic Pain.

Neuropathic pain is a chronic pain condition seen in patients with diabetic neuropathy, cancer chemotherapy-induced neuropathy, idiopathic neuropathy as well as other diseases affecting the nervous system. Only a small percentage of people with neuropathic pain benefit from current medications. The complexity of the disease, poor identification/lack of diagnostic and prognostic markers limit current strategies for the management of neuropathic pain. Multiple genes and pathways involved in human diseases can be regulated by microRNA (miRNA) which are small non-coding RNA. Several miRNAs are found to be dysregulated in neuropathic pain. These miRNAs regulate expression of various genes associated with neuroinflammation and pain, thus, regulating neuropathic pain. Some of these key players include adenylate cyclase (Ac9), toll-like receptor 8 (Tlr8), suppressor of cytokine signaling 3 (Socs3), signal transducer and activator of transcription 3 (Stat3) and RAS p21 protein activator 1 (Rasa1). With advancements in high-throughput technology and better computational power available for research in present-day pharmacology, biomarker discovery has entered a very exciting phase. We dissect the architecture of miRNA biological networks encompassing both human and rodent microRNAs involved in the development of neuropathic pain. We delineate various microRNAs, and their targets, that may likely serve as potential biomarkers for diagnosis, prognosis, and therapeutic intervention in neuropathic pain. miRNAs mediate their effects in neuropathic pain by signal transduction through IRAK/TRAF6, TLR4/NF-κB, TXIP/NLRP3 inflammasome, MAP Kinase, TGFß and TLR5 signaling pathways. Taken together, the elucidation of the landscape of signature miRNA regulatory networks in neuropathic pain will facilitate the discovery of novel miRNA/target biomarkers for more effective management of neuropathic pain.

Bacillus subtilis HU58 and Bacillus coagulans SC208 Probiotics Reduced the Effects of Antibiotic-Induced Gut Microbiome Dysbiosis in An M-SHIME® Model.

Benefits associated with probiotic use have been reported; however, the mechanisms behind these benefits are poorly understood. The effects of a probiotic formulation (MegaDuo™) containing Bacillus coagulans SC208 and Bacillus subtilis HU58 on intestinal permeability and immune markers was assessed using a combination of the in vitro gut model, the mucosal simulator of the human intestinal microbial ecosystem (M-SHIME®), and an in vitro inflammatory bowel disease-like Caco-2/THP1 co-culture model in both healthy and antibiotic-induced dysbiosis conditions. Established M-SHIME® proximal colon vessels were treated with/without clindamycin (1 week) and then with/without daily MegaDuo™ treatment (2 weeks). The mucosal and luminal microbial communities were sampled weekly. Suspensions were removed from the proximal colon vessels after 1 and 2 weeks of MegaDuo™ treatment and added to the co-culture system. Transepithelial resistance (membrane barrier function), cytokine/chemokine release, and NFκB activity were then measured. Under conditions of antibiotic-induced dysbiosis, suspensions from MegaDuo™ treated vessels showed reduced gut membrane barrier damage and decreased levels of TNFα and IL-6 compared with suspensions from untreated vessels; no appreciable differences were observed under healthy conditions. MegaDuo™ treatment had no effect on NFκB activity of THP1-Blue™ cells. The potential benefits of MegaDuo™ treatment appeared most evident after 2 weeks of treatment.

Efficacy of a silver colloidal gel against selected oral bacteria in vitro.

Background: It is necessary to develop new strategies to protect against bacteria such as S treptococcus mutans, S treptococcus sanguis, and Streptococcus salivarius, which contribute to tooth decay and plaque formation. Our current study investigated the efficacy of a colloidal silver gel in inhibiting biofilm formation by these principal oral bacteria , in vitro. The aim of this study was to assess the efficacy of a colloidal silver gel formulation for inhibiting bacterial biofilm formation (Ag-gel) by the principal bacteria that cause plaque formation and tooth decay. Methods: The effect of Ag-gel on viability of S. mutans, S. sanguis, and S. salivarius was assessed by quantifying their colony forming units (CFU) in presence or absence of the test gel. The effect of this formulation on biofilm-forming ability of these bacteria was studied through scanning electron microscopy. Results: Using the CFU assays, over 6 logs of inhibition (100%) were found for S. mutans, S. sanguis, and S. salivarius for the Ag-gel-treated bacteria when compared with the control gel. In addition, the Ag-gel also inhibited biofilm formation by these three bacteria mixed together. These results were confirmed by scanning electron microscopy. Conclusions: The Ag-gel was effective in preventing biofilm formation by S. mutans, S. sanguis, and S. salivarius. This Ag-gel should be tested for the ability to block plaque formation in the mouth, through its use as a tooth paste.

The ability of quaternary ammonium groups attached to a urethane bandage to inhibit bacterial attachment and biofilm formation in a mouse wound model.

For proper wound healing, control of bacteria or bacterial infections is of major importance. While caring for a wound, dressing material plays a key role as bacteria can live in the bandage and keep re-infecting the wound. They do this by forming biofilms in the bandage, which slough off planktonic bacteria and overwhelm the host defense. It is thus necessary to develop a wound dressing that will inhibit bacterial growth. This study examines the effectiveness of a polyurethane foam wound dressing bound with polydiallyl-dimethylammonium chloride (pDADMAC) to inhibit the growth of bacteria in a wound on the back of a mouse. This technology does not allow pDADMAC to leach away from the dressing into the wound, thereby preventing cytotoxic effects. Staphylococcus aureus, Pseudomonas aeruginosa and Acinetobacter baumannii were chosen for the study to infect the wounds. S. aureus and P. aeruginosa are important pathogens in wound infections, while A. baumannii was selected because of its ability to acquire or upregulate antibiotic drug resistance determinants. In addition, two different isolates of methicillin-resistant S. aureus (MRSA) were tested. All the bacteria were measured in the wound dressing and in the wound tissue under the dressing. Using colony-forming unit (CFU) assays, over six logs of inhibition (100%) were found for all the bacterial strains using pDADMAC-treated wound dressing when compared with control-untreated dressing. The CFU assay results obtained with the tissues were significant as there were 4-5 logs of reduction (100%) of the test organism in the tissue of the pDADMAC-covered wound versus that of the control dressing-covered wound. As the pDADMAC cannot leave the dressing (like other antimicrobials), this would imply that the dressing acts as a reservoir for free bacteria from a biofilm and plays a significant role in the development of a wound infection.

Inhibition of TNF-α, IL-1α, and IL-1ß by Pretreatment of Human Monocyte-Derived Macrophages with Menaquinone-7 and Cell Activation with TLR Agonists In Vitro.

Circulatory markers of low-grade inflammation such as tumor necrosis factor-alpha (TNF-α), interleukin-1 alpha (IL-1α), and interleukin-1 beta (IL-1ß) positively correlate with endothelial damage, atheroma formation, cardiovascular disease, and aging. The natural vitamin K2-menaquinone-7 (MK-7) added to the cell culture of human monocyte-derived macrophages (hMDMs) at the same time as toll-like receptor (TLR) agonists did not influence the production of TNF-α. When the cells were pretreated up to 6 h with MK-7 before treatment with TLR agonists, MK-7 did not inhibit significantly the production of TNF-α after the TLR activation. However, 30 h pretreatment of hMDMs with at least 10 µM of MK-7 effectively and dose dependently inhibited the proinflammatory function of hMDMs. Pretreatment of hMDMs with 10 µM of MK-7 for 30 h resulted in 20% inhibition of TNF-α production after lipopolysaccharide (LPS) activation (P < .05) and 43% inhibition after macrophage-activating lipopeptide (MALP) activation (P < .001). Pathogen-associated molecular pattern (PMPP) activation was inhibited by 20% with MK-7 pretreatment; however, this inhibition was not statistically significant. The 30 h pretreatment of a THP-1-differentiated monocyte cell line with MK-7 resulted in a dose-dependent downregulation of TNFα, IL-1α, and IL-1ß gene expression as evaluated by RNA semiquantitative reverse transcription polymerase chain reaction (RT-PCR). MK-7 is able to modulate immune and inflammatory reactions in the dose-response inhibition of TNF-α, IL-1α, and IL-1ß gene expression and protein production by the healthy hMDMs in vitro.

Safety assessment of menaquinone-7 for use in human nutrition.

Vitamin K occurs widely in foods and has been shown to have a beneficial effect on the cardiovascular system, as well as anticancer, anti-inflammatory, and antiosteoporosis properties. A previous study indicates that long-chain menaquinone-7 may be more bioavailable than vitamin K and short-chain menaquinones. In the present study, acute, subacute toxicity and genotoxicity assays were carried out to evaluate the safety of oral menaquinone-7 in albino Wistar rats. Oral administration of menaquinone-7, at a concentration of 2000 mg/kg, did not cause toxic symptoms in either male or female rats. A subacute toxicity study also proved the safety and tolerance of prolonged treatment (for 90 days) with menaquinone-7 in rats, as evidenced by biochemical, hematological, and urine parameters as well as by histopathological analysis. Genotoxicity and mutagenicity studies were performed by comet, micronucleus, and Ames tests on Salmonella typhimurium strains, which showed cellular safety and nonmutagenicity of menaquinone-7. The results indicate the safety of menaquinone-7 for human consumption.

A study on the ability of quaternary ammonium groups attached to a polyurethane foam wound dressing to inhibit bacterial attachment and biofilm formation.

Bacterial infection of acute and chronic wounds impedes wound healing significantly. Part of this impediment is the ability of bacterial pathogens to grow in wound dressings. In this study, we examined the effectiveness of a polyurethane (PU) foam wound dressings coated with poly diallyl-dimethylammonium chloride (pDADMAC-PU) to inhibit the growth and biofilm development by three main wound pathogens, Staphylococcus aureus, Pseudomonas aeruginosa, and Acinetobacter baumannii, within the wound dressing. pDADMAC-PU inhibited the growth of all three pathogens. Time-kill curves were conducted both with and without serum to determine the killing kinetic of pDADMAC-PU. pDADMAC-PU killed S. aureus, A. baumannii, and P. aeruginosa. The effect of pDADMAC-PU on biofilm development was analyzed quantitatively and qualitatively. Quantitative analysis, colony-forming unit assay, revealed that pDADMAC-PU dressing produced more than eight log reduction in biofilm formation by each pathogen. Visualization of the biofilms by either confocal laser scanning microscopy or scanning electron microscopy confirmed these findings. In addition, it was found that the pDADMAC-PU-treated foam totally inhibited migration of bacteria through the foam for all three bacterial strains. These results suggest that pDADMAC-PU is an effective wound dressing that inhibits the growth of wound pathogens both within the wound and in the wound dressing.