A Selection Protocol to Identify Therapeutics to Target NLRP3-Associated Sensory Hearing Loss.

Objective: We propose a selection process to identify a small molecule inhibitor to treat NLRP3-associated sensory hearing loss.

Background: The NLRP3 inflammasome is an innate immune sensor and present in monocytes and macrophages. Once the inflammasome is activated, a cleavage cascade is initiated leading to the release of proinflammatory cytokines IL-1β and IL-18.

Localized application of SAG21k-loaded fibrin hydrogels for targeted modulation of the hedgehog pathway in facial nerve injury.

Given the broad biological effects of the Hedgehog (Hh) pathway, there is potential clinical value in local application of Hh pathway modulators to restrict pathway activation of target tissues and avoid systemic pathway activation. One option to limit Hh pathway activation is using fibrin hydrogels to deliver pathway modulators directly to tissues of interest, bypassing systemic distribution of the drug. In this study, we loaded the potent Hh pathway agonist, SAG21k, into fibrin hydrogels.

Eradication of Bacterial Persister Cells By Leveraging Their Low Metabolic Activity Using Adenosine Triphosphate Coated Gold Nanoclusters.

Bacteria first develop tolerance after antibiotic exposure; later genetic resistance emerges through the population of tolerant bacteria. Bacterial persister cells are the multidrug-tolerant subpopulation within an isogenic bacteria culture that maintains genetic susceptibility to antibiotics. Because of this link between antibiotic tolerance and resistance and the rise of antibiotic resistance, there is a pressing need to develop treatments to eradicate persister cells.

Povidone-Iodine Fails to Eradicate Chronic Suppurative Otitis Media and Demonstrates Ototoxic Risk in Mice.

Hypothesis: Commercially available povidone-iodine solution can eliminate biofilms and persister cells rapidly in in vivo achievable concentrations without inducing ototoxicity.

Background: Chronic suppurative otitis media (CSOM) is a substantial global problem. Current treatment options often induce a temporary remission without leading to a permanent cessation of symptoms secondary to the treatments' inability to eliminate persister cells.

Chronic suppurative otitis media causes macrophage-associated sensorineural hearing loss.

Background: Chronic suppurative otitis media (CSOM) is the most common cause of permanent hearing loss in children in the developing world. A large component of the permanent hearing loss is sensory in nature and our understanding of the mechanism of this has so far been limited to post-mortem human specimens or acute infection models that are not representative of human CSOM. In this report, we assess cochlear injury in a validated Pseudomonas aeruginosa (PA) CSOM mouse model.

Anti-persister and Anti-biofilm Activity of Self-Assembled Antimicrobial Peptoid Ellipsoidal Micelles.

Although persister cells are the root cause of resistance development and relapse of chronic infections, more attention has been focused on developing antimicrobial agents against resistant bacterial strains than on developing anti-persister agents. Frustratingly, the global preclinical antibacterial pipeline does not include any anti-persister drug. Therefore, the central point of this work is to explore antimicrobial peptidomimetics called peptoids (sequence-specific oligo--substituted glycines) as a new class of anti-persister drugs.

Gold nanocluster adjuvant enables the eradication of persister cells by antibiotics and abolishes the emergence of resistance.

Persister cells are responsible for relapses of infections common in cystic fibrosis and chronic suppurative otitis media (CSOM). Yet, there are no Food and Drug Administration (FDA) approved antibiotics to eradicate persister cells. Frustratingly, the global preclinical bacterial pipeline does not contain antibacterial agents targeting persister cells.

Topical Therapy Failure in Chronic Suppurative Otitis Media is Due to Persister Cells in Biofilms.

Objective: Chronic suppurative otitis media (CSOM) is characterized by a chronically draining middle ear. CSOM is typically treated with multiple courses of antibiotics or antiseptics which are successful in achieving quiescence; however, the disease is prone to relapse. Understanding why these treatment failures occur is essential.

Locally administered heparin-binding epidermal growth factor-like growth factor reduces radiation-induced oral mucositis in mice.

Oral mucositis refers to lesions of the oral mucosa observed in patients with cancer being treated with radiation with or without chemotherapy, and can significantly affect quality of life. There is a large unmet medical need to prevent oral mucositis that can occur with radiation either alone or in combination with chemotherapy. We investigated the efficacy of locally administered heparin-binding epidermal growth factor-like growth factor (HB-EGF), a potent epithelial proliferation and migration stimulator of the oral mucosa as a potential therapy to prevent radiation induced oral mucositis.

A novel mouse model of chronic suppurative otitis media and its use in preclinical antibiotic evaluation.

Chronic suppurative otitis media (CSOM) is a neglected pediatric disease affecting 330 million worldwide for which no new drugs have been introduced for over a decade. We developed a mouse model with utility in preclinical drug evaluation and antimicrobial discovery. Our model used immune-competent mice, tympanic membrane perforation and inoculation with luminescent Pseudomonas aeruginosa that enabled bacterial abundance tracking in real-time for 100 days.

Antimicrobial Gold Nanoclusters Eradicate Biofilms and Are Nontoxic by Oral Administration.

biofilms are a major causative agent of many intestinal infections, and there is ongoing research aimed at biofilm eradication. Gold nanoclusters (AuNCs) conjugated with various surface ligands have been extensively investigated for antimicrobial properties and provide a potential solution. There is little known about their safety because current standards of nanosafety research involve incubation of AuNCs with cells to confirm biocompatibility.

Polyelectrolyte stiffness on gold nanorods mediates cell membrane damage.

Charge and surface chemistry of gold nanorods (AuNRs) are often considered the predictive factors for cell membrane damage. Unfortunately, extensive research on AuNR passivated with polyelectrolyte (PE) ligand shell (AuNR-PE) has hitherto left a vital knowledge gap between the mechanical stability of the ligand shell and the cytotoxicity of AuNR-PEs. Here, the agreement between unbiased coarse-grained molecular dynamics (CGMD) simulation and empirical outcomes on hemolysis of red blood cells by AuNR-PEs demonstrates for the first time, a direct impact of the mechanical stability of the PE shell passivating the AuNRs on the lipid membrane rupture.

Quantitative assessment of bacterial growth phase utilizing flow cytometry.

Flow cytometry is currently underutilized for bacterial phenotyping and standard microbiological techniques do not provide phenotypic information about the state of the bacterial disease. Pseudomonas aeruginosa is a human pathogen of increased importance in public health due to both the ability to cause chronic diseases and the prevalence of functionally different subsets that can be difficult to treat and diagnose. In the present study, we used flow cytometry to analyze the growth phase of P.

Quantitative and Label-Free Detection of Protein Kinase A Activity Based on Surface-Enhanced Raman Spectroscopy with Gold Nanostars.

The activity of extracellular protein kinase A (PKA) is known to be a biomarker for cancer. However, conventional PKA assays based on colorimetric, radioactive, and fluorometric techniques suffer from intensive labeling-related preparations, background interference, photobleaching, and safety concerns. While surface-enhanced Raman spectroscopy (SERS)-based assays have been developed for various enzymes to address these limitations, their use in probing PKA activity is limited due to subtle changes in the Raman spectrum with phosphorylation.

Targeted conjugation of breast anticancer drug tamoxifen and its metabolites with synthetic polymers.

Conjugation of antitumor drug tamoxifen and its metabolites, 4-hydroxytamxifen and ednoxifen with synthetic polymers poly(ethylene glycol) (PEG), methoxypoly (ethylene glycol) polyamidoamine (mPEG-PAMAM-G3) and polyamidoamine (PAMAM-G4) dendrimers was studied in aqueous solution at pH 7.4. Multiple spectroscopic methods, transmission electron microscopy (TEM) and molecular modeling were used to characterize the drug binding process to synthetic polymers.

Conjugation of steroids with PAMAM nanoparticles.

We studied the binding process between polyamidoamine PAMAN-G4 dendrimer and testosterone and its aliphatic (alip) and aromatic (arom) dimers in aqueous solution at pH 7.4. Multiple spectroscopic methods, transmission electron microscopy (TEM) and molecular modeling were used to characterize the steroid binding process to PAMAM-G4 nanoparticles.

Transporting testosterone and its dimers by serum proteins.

A substantial part of steroids is bound to serum proteins in vivo. We report the association of testosterone and it aliphatic dimer (alip) and aromatic dimer (arom) with human serum albumin (HSA) and bovine serum albumin (BSA) in aqueous solution at physiological pH. Multiple spectroscopic methods, transmission electron microscopy (TEM) and molecular modeling were used to characterize steroid-protein binding and protein aggregation process.

Structural analysis of doxorubicin-polymer conjugates.

Synthetic polymers poly(ethylene glycol) (PEG), methoxypoly (ethylene glycol) polyamidoamine (mPEG-PAMAM-G3) and polyamidoamine (PAMAM-G4) dendrimers were used for encapsulation of antibiotic drug doxorubicin (Dox) and its analogue N-(trifluoroacetyl) doxorubicin (FDox) in aqueous solution at pH 7.4. Multiple spectroscopic methods, transmission electron microscopy (TEM) and molecular modeling were used to characterize the drug binding process to synthetic polymers.

Structural modeling for DNA binding to antioxidants resveratrol, genistein and curcumin.

Several models are presented here for the bindings of the antioxidant polyphenols resveratrol, genistein and curcumin with DNA in aqueous solution at physiological conditions. Multiple spectroscopic methods and molecular modeling were used to locate the binding sites of these polyphenols with DNA duplex. Structural models showed that intercalation is more stable for resveratrol and genistein than groove bindings, while curcumin interaction is via DNA grooves.

Locating the binding sites of antioxidants resveratrol, genistein and curcumin with tRNA.

We located the binding sites of antioxidants resveratrol, genistein and curcumin on tRNA in aqueous solution at physiological conditions using constant tRNA concentration and various polyphenol contents. FTIR, UV-visible, CD spectroscopic methods and molecular modeling were used to determine polyphenol binding sites, the binding constant and the effects of polyphenol complexation on tRNA conformation and particle formation. Structural analysis showed that polyphenols bind tRNA via G-C and A-U base pairs through hydrophilic, hydrophobic and H-bonding contacts with overall binding constants of K(res-tRNA)=8.

The role of polymer size and hydrophobic end-group in PEG-protein interaction.

We investigated the interaction between polyethylene (glycol) (PEG) and human (HSA) and bovine serum albumin (BSA) in aqueous solution, using multiple spectroscopic methods and molecular modeling. The two important polymer characteristics, size and PEG hydrophobic end-group are studied in order to determine the effect of each one on PEG-protein interaction. The bindings of PEG and mPEG-anthracene with serum albumins occur via hydrophobic and H-bonding contacts with the binding affinity PEG-6000>mPEG-anthracene>PEG-3000 for BSA and EG-6000>PEG-3000>mPEG-anthracene for HSA.

Encapsulation of testosterone and its aliphatic and aromatic dimers by milk beta-lactoglobulin.

The encapsulation of testosterone and it aliphatic dimer (alip) and aromatic dimer (arom) with milk β-lactoglobulin (β-LG) was studied in aqueous solution at pH 7.4. Multiple spectroscopic methods, transmission electron microscopy (TEM) and molecular modeling were used to characterize testosterone-β-LG binding and protein aggregation process.

Effect of polymer molecular weight on chitosan-protein interaction.

We present a comprehensive study of the interactions between chitosan nanoparticles (15, 100 and 200 kDa with the same degree of deacetylation 90%) and two model proteins, i.e., bovine (BSA) and human serum albumins (HSA), with the aim of correlating chitosan molecular weight (Mw) and the binding affinity of these naturally occurring polymers to protein.

Protective action of spermine and spermidine against photoinhibition of photosystem I in isolated thylakoid membranes.

The photo-stability of photosystem I (PSI) is of high importance for the photosynthetic processes. For this reason, we studied the protective action of two biogenic polyamines (PAs) spermine (Spm) and spermidine (Spd) on PSI activity in isolated thylakoid membranes subjected to photoinhibition. Our results show that pre-loading thylakoid membranes with Spm and Spd reduced considerably the inhibition of O2 uptake rates, P700 photooxidation and the accumulation of superoxide anions (O2(-)) induced by light stress.

Association of lipids with milk α- and β-caseins.

Unlabelled: We report the molecular interaction and the binding sites of cholesterol (CHOL), 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), dioctadecyldimethyl-ammoniumbromide (DDAB), and dioleoylphosphatidylethanolamine (DOPE) with milk α- and β-caseins in aquous solution at physiological conditions. Fourier transform infrared (FTIR), fluorescence spectroscopic methods and molecular modeling were used to determine the binding sites of lipid-protein complexes and the effect of lipid interaction on the stability and conformation of α- and β-caseins. Structural analysis showed that lipids bind casein via mainly hydrophobic contact with association constants of KCHOL-α-casein=1.