Safety and Pharmacokinetics of a Ciprofloxacin and Azithromycin Stent for Chronic Rhinosinusitis.

OBJECTIVES: Previously, we developed a novel double-coated sinus stent containing ciprofloxacin (inner layer) and azithromycin (outer layer) (CASS), but released drug concentrations were found to be insufficient for clinical usage. Our objectives are to improve drug release of CASS and assess safety and pharmacokinetics in rabbits. METHODS: Dip coating was used to create the CASS with 2 mg ciprofloxacin and 5 mg azithromycin. A uniformed double coating was assessed with scanning electron microscopy (SEM), and the release patterns of both drugs and lactate dehydrogenase (LDH) assay were evaluated over 14 days in vitro. Safety, tolerability, and pharmacokinetics of the CASS were tested in rabbits through insertion into the maxillary sinus and evaluated with nasal endoscopy, CT scans, histology, blood counts and chemistries, and in vivo drug release. RESULTS: SEM confirmed the uniformity of the dual coating of ciprofloxacin and azithromycin, and thickness (µm) was found to be 14.7 ± 2.4 and 28.1 ± 4.6, respectively. The inner coated ciprofloxacin showed a sustained release over 14 days (release %) when soaked in saline solution (day 7, 86.2 ± 3.4 vs. day 14,99.2 ± 5.1). In vivo analysis showed that after 12 days, 78.92 ± 7.67% of CP and 84.12 ± 0.45% of AZ were released into the sinus. There were no significant differences in body weight, white blood cell counts, and radiographic changes before and after CASS placement. No significant histological changes were observed compared to the contralateral control side. CONCLUSION: Findings suggest that the CASS is an effective method for delivering therapeutic levels of antibiotics. Further studies are needed to validate efficacy in a preclinical sinusitis model. LEVEL OF EVIDENCE: N/A Laryngoscope, 2024.

Hypoxia-induced cystic fibrosis transmembrane conductance regulator dysfunction is a universal mechanism underlying reduced mucociliary transport in sinusitis.

INTRODUCTION: Hypoxia due to sinus obstruction is a major pathogenic mechanism leading to sinusitis. The objective of the current study is to define the electrophysiologic characteristics of hypoxia in vitro and in vivo. METHODS: Cystic fibrosis bronchoepithelial cells expressing wild-type cystic fibrosis transmembrane conductance regulator (CFTR) and human sinonasal epithelial cells were exposed to 1% or atmospheric O2 for 24 h. Time-dependent production of cytoplasmic free radicals was measured. Cells were subjected to Ussing chamber and patch clamp technique where CFTR currents were recorded in whole-cell and cell-attached mode for single channel studies. Indices of mucociliary transport (MCT) were measured using micro-optical coherence tomography. In a rabbit hypoxic maxillary sinus model, tissue oxygenation, relative mRNA expression of HIF-1α, pH, sinus potential difference (SPD), and MCT were determined. RESULTS: Ussing chamber (p < 0.05), whole-cell (p < 0.001), and single channel patch-clamp (p < 0.0001) showed significant inhibition of Cl- currents in hypoxic cells. Cytoplasmic free radicals showed time-dependent elevation peaking at 4 h (p < 0.0001). Airway surface liquid (p < 0.0001), periciliary liquid (p < 0.001), and MCT (p < 0.01) were diminished. Co-incubation with the free radical scavenger glutathione negated the impact of hypoxia on single channel currents and MCT markers. In sinusitis rabbits, mucosa exhibited low tissue oxygenation (p < 0.0001), increased HIF1α mRNA (p < 0.05), reduced pH (p < 0.01), and decreased MCT (p < 0.001). SPD measurements demonstrated markedly diminished transepithelial Cl- transport (p < 0.0001). CONCLUSION: Hypoxia induces severe CFTR dysfunction via free radical production causing reduced MCT in vitro and in vivo. Improved oxygenation is critical to reducing the impact of persistent mucociliary dysfunction.

Glutathione and bicarbonate nanoparticles improve mucociliary transport in cystic fibrosis epithelia.

INTRODUCTION: Cystic fibrosis (CF) airway disease is characterized by thick mucus and impaired mucociliary transport (MCT). Loss of functional cystic fibrosis transmembrane receptor (CFTR) leads to acidification and oxidation of airway surface mucus. Replacing bicarbonate (HCO3 - ) topically fails due to rapid reabsorption and neutralization, while the scavenging antioxidant, glutathione sulfhydryl (GSH), is also rapidly degraded. The objective of this study is to investigate GSH/NaHCO3 nanoparticles as novel strategy for CF airway disease. METHODS: GSH/NaHCO3 poly (lactic-co-glycolic acid) nanoparticles were tested on primary CF (F508del/F508del) epithelial cultures to evaluate dose-release curves, surface pH, toxicity, and MCT indices using micro-optical coherence tomography. In vivo tests were performed in three rabbits to assess safety and toxicity. After 1 week of daily injections, histopathology, computed tomography (CT), and blood chemistries were performed and compared to three controls. Fluorescent nanoparticles were injected into a rabbit with maxillary sinusitis and explants visualized with confocal microscopy. RESULTS: Sustained release of GSH and HCO3 - with no cellular toxicity was observed over 2 weeks. Apical surface pH gradually increased from 6.54 ± 0.13 (baseline) to 7.07 ± 0.10 (24 h) (p < 0.001) and 6.87 ± 0.05 at 14 days (p < 0.001). MCT, ciliary beat frequency, and periciliary liquid were significantly increased. When injected into the maxillary sinuses of rabbits, there were no changes to histology, CT, or blood chemistries. Nanoparticles penetrated rabbit sinusitis mucus on confocal microscopy. CONCLUSION: Findings suggest that GSH/NaHCO3 - nanoparticles are a promising treatment option for viscous mucus in CF and other respiratory diseases of mucus obstruction such as chronic rhinosinusitis.

Red ginseng aqueous extract improves mucociliary transport dysfunction and histopathology in CF rat airways.

BACKGROUND: We previously discovered that Korean red ginseng aqueous extract (RGAE) potentiates the TMEM16A channel, improved mucociliary transport (MCT) parameters in CF nasal epithelia in vitro, and thus could serve as a therapeutic strategy to rescue the MCT defect in cystic fibrosis (CF) airways. The hypothesis of this study is that RGAE can improve epithelial Cl- secretion, MCT, and histopathology in an in-vivo CF rat model. METHODS: Seventeen 4-month old CFTR-/- rats were randomly assigned to receive daily oral control (saline, n = 9) or RGAE (Ginsenosides 0.4mg/kg/daily, n = 8) for 4 weeks. Outcomes included nasal Cl- secretion measured with the nasal potential difference (NPD), functional microanatomy of the trachea using micro-optical coherence tomography, histopathology, and immunohistochemical staining for TMEM16a. RESULTS: RGAE-treated CF rats had greater mean NPD polarization with UTP (control = -5.48 +/- 2.87 mV, RGAE = -9.49 +/- 2.99 mV, p < 0.05), indicating, at least in part, potentiation of UTP-mediated Cl- secretion through TMEM16A. All measured tracheal MCT parameters (airway surface liquid, periciliary liquid, ciliary beat frequency, MCT) were significantly increased in RGAE-treated CF rats with MCT exhibiting a 3-fold increase (control, 0.45+/-0.31 vs. RGAE, 1.45+/-0.66 mm/min, p < 0.01). Maxillary mucosa histopathology was markedly improved in RGAE-treated cohort (reduced intracellular mucus, goblet cells with no distention, and shorter epithelial height). TMEM16A expression was similar between groups. CONCLUSION: RGAE improves TMEM16A-mediated transepithelial Cl- secretion, functional microanatomy, and histopathology in CF rats. Therapeutic strategies utilizing TMEM16A potentiators to treat CF airway disease are appropriate and provide a new avenue for mutation-independent therapies.

3-O-Ethyl-L-Ascorbic Acid Doped Enteric-Coated Gelatin Capsules towards the Advanced Oral Curcumin Delivery for Cancers.

Among plant-derived polyphenols, curcumin has been recognized as a therapeutically potent nutrient presenting pleiotropic pharmacological effects on various cancers. However, the poor absorption and bioavailability of curcumin limit the use of this excellent naturally occurring polyphenol. 3-O-ethyl-L-ascorbic acid (EA) doped enteric-coated gelatin capsules were studied in the search for advanced oral curcumin delivery. The EA doped enteric-coated gelatin capsules were successfully created based on a developed inner dual enteric coating technique. When placed in four buffer solutions with different pHs (pH 2.0, 5.0, 6.0, and 7.3), the coated gelatin capsules showed delayed-release profiles of curcumin below pH 6.0. In contrast, both pristine and fabricated gelatin capsules showed similar curcumin release profiles at pH 7.3, which is a common pH observed in the lower gastrointestinal tract, especially intestinal regions. In conclusion, these results demonstrated the potential of the EA doped enteric-coated gelatin capsules in developing advanced oral delivery of curcumin targeting intestinal-specific regions.

Bone Mineralization in Electrospun-Based Bone Tissue Engineering.

Increasing the demand for bone substitutes in the management of bone fractures, including osteoporotic fractures, makes bone tissue engineering (BTE) an ideal strategy for solving the constant shortage of bone grafts. Electrospun-based scaffolds have gained popularity in BTE because of their unique features, such as high porosity, a large surface-area-to-volume ratio, and their structural similarity to the native bone extracellular matrix (ECM). To imitate native bone mineralization through which bone minerals are deposited onto the bone matrix, a simple but robust post-treatment using a simulated body fluid (SBF) has been employed, thereby improving the osteogenic potential of these synthetic bone grafts. This study highlights recent electrospinning technologies that are helpful in creating more bone-like scaffolds, and addresses the progress of SBF development. Biomineralized electrospun bone scaffolds are also reviewed, based on the importance of bone mineralization in bone regeneration. This review summarizes the potential of SBF treatments for conferring the biphasic features of native bone ECM architectures onto electrospun-based bone scaffolds.

Cross-Linking Agents for Electrospinning-Based Bone Tissue Engineering.

Electrospun nanofibers are promising bone tissue scaffolds that support bone healing due to the body's structural similarity to the extracellular matrix (ECM). However, the insufficient mechanical properties often limit their potential in bone tissue regeneration. Cross-linking agents that chemically interconnect as-spun electrospun nanofibers are a simple but effective strategy for improving electrospun nanofibers' mechanical, biological, and degradation properties. To improve the mechanical characteristic of the nanofibrous bone scaffolds, two of the most common types of cross-linking agents are used to chemically crosslink electrospun nanofibers: synthetic and natural. Glutaraldehyde (GTA) is a typical synthetic agent for electrospun nanofibers, while genipin (GP) is a natural cross-linking agent isolated from gardenia fruit extracts. GP has gradually gained attention since GP has superior biocompatibility to synthetic ones. In recent studies, much more progress has been made in utilizing crosslinking strategies, including citric acid (CA), a natural cross-linking agent. This review summarizes both cross-linking agents commonly used to improve electrospun-based scaffolds in bone tissue engineering, explains recent progress, and attempts to expand the potential of this straightforward method for electrospinning-based bone tissue engineering.

Microneedles in Action: Microneedling and Microneedles-Assisted Transdermal Delivery.

Human skin is a multilayered physiochemical barrier protecting the human body. The stratum corneum (SC) is the outermost keratinized layer of skin through which only molecules with less or equal to 500 Da (Dalton) in size can freely move through the skin. Unfortunately, the conventional use of a hypothermic needle for large therapeutic agents is susceptible to needle phobia and the risk of acquiring infectious diseases. As a new approach, a microneedle (MN) can deliver therapeutically significant molecules without apparent limitations associated with its molecular size. Microneedles can create microchannels through the skin's SC without stimulating the proprioceptive pain nerves. With recent technological advancements in both fabrication and drug loading, MN has become a versatile platform that improves the efficacy of transdermally applied therapeutic agents (TAs) and associated treatments for various indications. This review summarizes advanced fabrication techniques for MN and addresses numerous TA coating and TA elution strategies from MN, offering a comprehensive perspective on the current microneedle technology. Lastly, we discuss how microneedling and microneedle technologies can improve the clinical efficacy of a variety of skin diseases.

In vitro evaluation of a novel oxygen-generating biomaterial for chronic rhinosinusitis therapy.

BACKGROUND: Hypoxia due to closure at the ostiomeatal complex is widely considered one of the major pathogenic mechanisms leading to chronic inflammation in chronic rhinosinusitis (CRS). The objective of this study was to develop and characterize an oxygen-generating biomaterial (OGB) as an innovative treatment strategy for CRS. METHODS: An OGB was fabricated by coating hydrophobic beeswax (BW, 15mg or 30mg) on the surface of calcium peroxide - catalase complex (CPO-CA, 30mg) and characterized using scanning electron microscopy (SEM). In vitro releases of both oxygen and hydrogen peroxide (H2 O2 ) were spectrophotometrically quantified, and cytotoxicity in human sinonasal epithelial cells (HSNECs) was evaluated. The influence of OGB on transepithelial Cl- secretion was also determined by pharmacologically manipulating HSNECs, cultured under hypoxic conditions, in Ussing chambers. RESULTS: Three groups of OGBs: (1) CPO only; (2) CPO coated with CA and BW (1:1 ratio, CPO-CA(1)-BW(1)); and (3) CPO coated with CA and BW (1:0.5 ratio, CPO-CA(1)-BW(0.5) were analyzed for accumulated oxygen release over 7 days: highest release (mmol/mg) was observed in CPO-CA(1)-BW(1) = 0.11 ± 0.003, followed by CPO-CA(1)-BW(0.5) = 0.08 ± 0.010, and CPO = 0.05 ± 0.004 (p < 0.0001). H2 O2 production (mM) was significantly higher in CPO (1.87 ± 0.50) compared to CPO-CA (1)-BW(1) (0.00 ± 0.00) (p < 0.001) after 24 h. CPO-CA(1)-BW(1) showed significantly reduced cytotoxicity and increased Cl- transport compared to the CPO group. CONCLUSION: A novel OGB (CPO-CA-BW complex) exhibited sustained oxygen release over 7 days without significant cytotoxicity after 24 h in vitro. Preclinical studies evaluating the efficacy of OGB in CRS are warranted, especially for potential therapy in an obstruction-based CRS model.

Ivacaftor restores delayed mucociliary transport caused by Pseudomonas aeruginosa-induced acquired cystic fibrosis transmembrane conductance regulator dysfunction in rabbit nasal epithelia.

BACKGROUND: Abnormal chloride (Cl- ) transport dehydrates airway surface liquid (ASL) in sinonasal epithelium leading to mucus stasis and chronic rhinosinusitis. As an experimental epithelium, rabbit tissue provides an excellent representation of human sinus disease, and the rabbit sinusitis model is both established and well suited for therapeutic interventions in vivo. Our objective in this study was to evaluate whether ivacaftor reverses the consequences of Pseudomonas aeruginosa-induced acquired cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction. METHODS: Rabbit nasal cavities were assessed for responsiveness to ivacaftor in vivo (by nasal potential difference [NPD] assay). Rabbit nasal epithelial (RNE) cultures were incubated with an ultrafiltrate of P aeruginosa (PAO1 strain) for 4 hours and tested for acquired CFTR dysfunction. Markers of mucociliary function, including airway surface liquid depth (ASL), periciliary liquid depth (PCL), ciliary beat frequency (CBF), and mucociliary transport (MCT), were measured by micro-optical coherence tomography (µOCT) after PAO1 and/or ivacaftor incubation. RESULTS: Ivacaftor resulted in a significant mean NPD polarization of 21.8 ± 2.1 mV, which was significantly greater than that seen in the low Cl- control (12.9 ± 1.3; p = 0.01). PAO1 exposure induced a state of acquired CFTR dysfunction in rabbit nasal epithelium as measured by forskolin-stimulated short-circuit current (ISC ) (control, 37.0 ± 1.1 µA/cm2 ; PAO1, 24.4 ± 1.1 µA/cm2 ; p < 0.001). RNE cultures exposed to PAO1 had inhibited mucociliary function, whereas coincubation with ivacaftor restored mucociliary clearance, as measured by µOCT. CONCLUSION: In rabbit nasal epithelium, ivacaftor robustly stimulates CFTR-mediated Cl- secretion and normalizes ASL and CBF in PAO1-induced acquired CFTR dysfunction. Preclinical testing of CFTR potentiators as therapy for P aeruginosa rabbit sinusitis is planned.

Oxygen-Releasing Composites: A Promising Approach in the Management of Diabetic Foot Ulcers.

In diabetes, lower extremity amputation (LEA) is an irreversible diabetic-related complication that easily occurs in patients with diabetic foot ulcers (DFUs). Because DFUs are a clinical outcome of different causes including peripheral hypoxia and diabetic foot infection (DFI), conventional wound dressing materials are often insufficient for supporting the normal wound healing potential in the ulcers. Advanced wound dressing development has recently focused on natural or biocompatible scaffolds or incorporating bioactive molecules. This review directs attention to the potential of oxygenation of diabetic wounds and highlights current fabrication techniques for oxygen-releasing composites and their medical applications. Based on different oxygen-releasable compounds such as liquid peroxides and solid peroxides, for example, a variety of oxygen-releasing composites have been fabricated and evaluated for medical applications. This review provides the challenges and limitations of utilizing current oxygen releasable compounds and provides perspectives on advancing oxygen releasing composites for diabetic-related wounds associated with DFUs.

Methylene Blue-Based Nano and Microparticles: Fabrication and Applications in Photodynamic Therapy.

Methylene blue (MB) has been used in the textile industry since it was first extracted by the German chemist Heinrich Caro. Its pharmacological properties have also been applied toward the treatment of certain diseases such as methemoglobinemia, ifosfamide-induced encephalopathy, and thyroid conditions requiring surgery. Recently, the utilization of MB as a safe photosensitizer in photodynamic therapy (PDT) has received attention. Recent findings demonstrate that photoactivated MB exhibits not only anticancer activity but also antibacterial activity both in vitro and in vivo. However, due to the hydrophilic nature of MB, it is difficult to create MB-embedded nano- or microparticles capable of increasing the clinical efficacy of the PDT. This review aims to summarize fabrication techniques for MB-embedded nano and microparticles and to provide both in vitro and in vivo examples of MB-mediated PDT, thereby offering a future perspective on improving this promising clinical treatment modality. We also address examples of MB-mediated PDT in both cancer and infection treatments. Both in-vitro and in-vivo studies are summarized here to document recent trends in utilizing MB as an effective photosensitizer in PDT. Lastly, we discuss how developing efficient MB-carrying nano- and microparticle platforms would be able to increase the benefits of PDT.

Korean Red Ginseng aqueous extract improves markers of mucociliary clearance by stimulating chloride secretion.

BACKGROUND: Abnormal chloride (Cl-) transport has a detrimental impact on mucociliary clearance in both cystic fibrosis (CF) and non-CF chronic rhinosinusitis. Ginseng is a medicinal plant noted to have anti-inflammatory and antimicrobial properties. The present study aims to assess the capability of red ginseng aqueous extract (RGAE) to promote transepithelial Cl- secretion in nasal epithelium. METHODS: Primary murine nasal septal epithelial (MNSE) [wild-type (WT) and transgenic CFTR-/-], fisher-rat-thyroid (FRT) cells expressing human WT CFTR, and TMEM16A-expressing human embryonic kidney cultures were utilized for the present experiments. Ciliary beat frequency (CBF) and airway surface liquid (ASL) depth measurements were performed using micro-optical coherence tomography (µOCT). Mechanisms underlying transepithelial Cl- transport were determined using pharmacologic manipulation in Ussing chambers and whole-cell patch clamp analysis. RESULTS: RGAE (at 30µg/mL of ginsenosides) significantly increased Cl- transport [measured as change in short-circuit current (ΔISC = µA/cm2)] when compared with control in WT and CFTR-/- MNSE (WT vs control = 49.8±2.6 vs 0.1+/-0.2, CFTR-/- = 33.5±1.5 vs 0.2±0.3, p < 0.0001). In FRT cells, the CFTR-mediated ΔISC attributed to RGAE was small (6.8 ± 2.5 vs control, 0.03 ± 0.01, p < 0.05). In patch clamp, TMEM16A-mediated currents were markedly improved with co-administration of RGAE and uridine 5-triphosphate (8406.3 +/- 807.7 pA) over uridine 5-triphosphate (3524.1 +/- 292.4 pA) or RGAE alone (465.2 +/- 90.7 pA) (p < 0.0001). ASL and CBF were significantly greater with RGAE (6.2+/-0.3 µm vs control, 3.9+/-0.09 µm; 10.4+/-0.3 Hz vs control, 7.3 ± 0.2 Hz; p < 0.0001) in MNSE. CONCLUSION: RGAE augments ASL depth and CBF by stimulating Cl- secretion through CaCC, which suggests therapeutic potential in both CF and non-CF chronic rhinosinusitis.

Azithromycin and ciprofloxacin inhibit interleukin-8 secretion without disrupting human sinonasal epithelial integrity in vitro.

BACKGROUND: We recently developed a ciprofloxacin and azithromycin sinus stent (CASS) to target recalcitrant infections in chronic rhinosinusitis (CRS). The objective of this study was to evaluate the anti-inflammatory activity of azithromycin released from the CASS and assess the impact on the integrity and function of primary human sinonasal epithelial cells (HSNECs). METHODS: Pseudomonas aeruginosa lipopolysaccharide (LPS)-stimulated HSNECs were treated with azithromycin and/or ciprofloxacin at concentrations attainable from CASS release. Interleukin-8 (IL-8) secretion was quantified by enzyme-linked immunosorbent assay (ELISA). Epithelial integrity (transepithelial resistance [TEER], paracellular permeability [fluorescein isothiocyanate-labeled dextran], lactate dehydrogenase [LDH] assays) and function (ciliary beat frequency [CBF]) were also evaluated. RESULTS: Azithromycin significantly reduced secreted IL-8 from P. aeruginosa LPS-stimulated HSNECs at all concentrations tested (mean ± standard deviation; control = 5.77 ± 0.39 ng/mL, azithromycin [6 µg/mL] = 4.58 ± 0.40 ng/mL, azithromycin [60 µg/mL] = 4.31 ± 0.06, azithromycin [180 µg/mL] = 4.27 ± 0.26 ng/mL, p < 0.05). Co-incubation with azithromycin (6 µg/mL) and ciprofloxacin (2.4 µg/mL) in LPS-stimulated HSNECs also displayed a significant reduction in secreted IL-8 when compared to P. aeruginosa LPS alone (co-treatment = 4.61 ± 0.29 ng/mL, P. aeruginosa LPS = 7.35 ± 0.89 ng/mL, p < 0.01). The drugs did not negatively impact TEER, paracellular permeability, LDH release, or CBF, indicating retention of cell integrity and function. CONCLUSION: Azithromycin decreased P. aeruginosa LPS IL-8 production in HSNECs at drug concentrations attainable with sustained release of azithromycin from the CASS. In addition to antibacterial activity, anti-inflammatory properties of the CASS should provide further benefit for patients with recalcitrant CRS.

Antibiotic eluting sinus stents.

OBJECTIVES: Chronic rhinosinusitis (CRS) is a multifactorial disease affecting up to 16% of the United States population and disproportionately affecting the cystic fibrosis (CF) patient population. Despite treating the underlying infection, the use of systemic antibiotics has shown little efficacy in alleviation of symptom burden. This review seeks to discuss recent research on novel antibiotic eluting stent therapy in vitro and within animal models as well as the factors that contribute to its efficacy. DATA SOURCES: PubMed literature review. REVIEW METHODS: A review of all published literature related to antibiotic eluting sinus stents was conducted to integrate and summarize this innovative approach to chronic sinus infections. RESULTS: Placement of the ciprofloxacin sinus stent (CSS) and ciprofloxacin-ivacaftor sinus stent (CISS) exhibited improvement in endoscopic and radiographic findings in rabbit CRS models. While the CSS showed an overall trend toward improvement in microscopic findings and a reduction in biofilm mass, there remained a significant quantity of planktonic bacteria due to antibiotic depletion from an initial burst release in the first 48 hours of stent placement. The CISS and ciprofloxacin-azithromycin sinus stents (CASSs) exhibited controlled antibiotic release over the study period leading to greatly reduced planktonic bacterial load and biofilm mass. In vitro studies indicate that CASS may be just as efficacious at reducing biofilm mass. CONCLUSION: Antibiotic eluting sinus stents show significant promise as a novel therapeutic strategy for CRS. The CISS may have particular promise for the CF patient population by addressing both the infectious and genetic components of disease. Animal studies demonstrate significant promise for translation into human studies. Human clinical trials are warranted to determine the efficacy of antibiotic sinus stents in human patients. LEVEL OF EVIDENCE: NA.

Contribution of Short Chain Fatty Acids to the Growth of Pseudomonas aeruginosa in Rhinosinusitis.

Background: Chronic rhinosinusitis (CRS) is characterized by complex bacterial infections with persistent inflammation. Based on our rabbit model of sinusitis, blockage of sinus ostia generated a shift in microbiota to a predominance of mucin degrading microbes (MDM) with acute inflammation at 2 weeks. This was followed by conversion to chronic sinus inflammation at 3 months with a robust increase in pathogenic bacteria (e.g., Pseudomonas). MDMs are known to produce acid metabolites [short chain fatty acids (SCFA)] that have the potential to stimulate pathogen growth by offering a carbon source to non-fermenting sinus pathogens (e.g., Pseudomonas). The objective of this study is to evaluate the concentrations of SCFA within the mucus and its contribution to the growth of P. aeruginosa. Methods: Healthy and sinusitis mucus from the rabbit model were collected and co-cultured with the PAO1 strain of P. aeruginosa for 72 h and colony forming units (CFUs) were determined with the targeted quantification of three SCFAs (acetate, propionate, butyrate). Quantification of SCFAs in healthy and sinusitis mucus from patients with P. aeruginosa was also performed via high performance liquid chromatography. Results: To provide evidence of fermentative activity, SCFAs were quantified within the mucus samples from rabbits with and without sinusitis. Acetate concentrations were significantly greater in sinusitis mucus compared to controls (4.13 ± 0.53 vs. 1.94 ± 0.44 mM, p < 0.01). After 72 h of co-culturing mucus samples with PAO1 in the presence of mucin medium, the blue-green pigment characteristic of Pseudomonas was observed throughout tubes containing sinusitis mucus. CFUs were higher in cultures containing mucus samples from sinusitis (8.4 × 109 ± 4.8 × 107) compared to control (1.4 × 109 ± 2.0 × 107) or no mucus (1.5 × 109 ± 2.1 × 107) (p < 0.0001). To provide evidence of fermentative activity in human CRS with P. aeruginosa, the presence of SCFAs in human mucus was analyzed and all SCFAs were significantly higher in CRS with P. aeruginosa compared to controls (p < 0.05). Conclusion: Given that SCFAs are solely derived from bacterial fermentation, our evidence suggests a critical role for mucin-degrading bacteria in generating carbon-source nutrients for pathogens. MDM may contribute to the development of recalcitrant CRS by degrading mucins, thus providing nutrients for potential pathogens like P. aeruginosa.

The impact of Lactococcus lactis (probiotic nasal rinse) co-culture on growth of patient-derived strains of Pseudomonas aeruginosa.

BACKGROUND: The Lactococcus strain of bacteria has been introduced as a probiotic nasal rinse for alleged salubrious effects on the sinonasal bacterial microbiome. However, data regarding interactions with pathogenic bacteria within the sinuses are lacking. The purpose of this study is to assess the interaction between L. lactis and patient-derived Pseudomonas aeruginosa, an opportunistic pathogen in recalcitrant chronic rhinosinusitis (CRS). METHODS: Commercially available probiotic suspension containing L. lactis W136 was grown in an anaerobic chamber and colonies were isolated. Colonies were co-cultured with patient-derived P. aeruginosa strains in the presence of porcine gastric mucin (mimicking human mucus) for 72 hours. P. aeruginosa cultures without L. lactis served as controls. Colony forming units (CFUs) were compared. RESULTS: Six P. aeruginosa isolates collected from 5 CRS patients (3 isolates from cystic fibrosis [CF], 1 mucoid strain) and laboratory strain PAO1 were co-cultured with L. lactis. There was no statistical difference in CFUs of 5 P. aeruginosa isolates grown with L. lactis compared to CFUs without presence of L. lactis. CFU counts were much higher when the mucoid strain was co-cultured with L. lactis (CFU+L.lactis = 1.9 × 108 ± 1.44 × 107, CFU-L.lactis = 1.3 × 108 ± 8.9 × 106, p = 0.01, n = 7). L. lactis suppressed the growth of 1 P. aeruginosa strain (CFU+L.lactis = 2.15 × 108 ± 2.9 × 107, CFU-L.lactis = 3.95 × 108 ± 4.8 × 106, p = 0.03, n = 7). CONCLUSION: L. lactis suppressed the growth of 1 patient P. aeruginosa isolate and induced growth of another (a mucoid strain) in in vitro co-culture setting in the presence of mucin. Further experiments are required to assess the underlying interactions between L. lactis and P. aeruginosa.

Controlled delivery of ciprofloxacin and ivacaftor via sinus stent in a preclinical model of Pseudomonas sinusitis.

BACKGROUND: Pseudomonas aeruginosa is common in chronic rhinosinusitus (CRS) and frequently resistant to antibiotic treatment. We recently described the ciprofloxacin and ivacaftor-releasing biodegradable sinus stent (CISS)-a drug-delivery system that administers ciprofloxacin and the mucociliary activator (ivacaftor) at high local concentrations with prolonged mucosal contact time and sustained delivery. The objective of this study is to evaluate the efficacy of the CISS in a rabbit model of P aeruginosa (PAO1 strain) sinusitis. METHODS: Ciprofloxacin/ivacaftor (double layer) was coated on biodegradable poly-D/L-lactic acid (PLLA). A total of 10 sinus stents (5 bare PLLA stent controls, 5 CISSs) were placed unilaterally in rabbit maxillary sinuses via dorsal sinusotomy after inducing infection for 1 week with PAO1. Animals were assessed 3 weeks after stent insertion with sinus culture, nasal endoscopy, computed tomography scan, histopathology, and in-vivo sinus potential difference (SPD) assay. RESULTS: Rabbits treated with CISS had significant reductions in computed tomography (∆ Kerschner scale: control, 0.55 ± 0.92; CISS, -5.92 ± 1.69; p = 0.024) and endoscopy (control, 4.0 ± 0.0; CISS, 1.875 ± 0.74; p = 0.003) scores. A 2-log reduction of PAO1 was observed (control, -2.14 ± 0.77; CISS, 1.84 ± 1.52; p = 0.047). SPD revealed significantly increased Cl- transport in the CISS group compared with the control group (Cl- -free + forskolin ΔPD: control, -4.23 ± 1.04 mV; CISS, -18.36 ± 6.31 mV; p = 0.026). Finally, marked improvements were noted in the histology of the mucosa and submucosa in treated animals. CONCLUSION: The CISS had robust clinical efficacy in treating P aeruginosa rabbit sinusitis. The innovative design of double-layered drug coating on the surface of the biodegradable stent may provide therapeutic advantages over current treatment strategies for P aeruginosa sinusitis.

In-vitro evaluation of a ciprofloxacin and azithromycin sinus stent for Pseudomonas aeruginosa biofilms.

BACKGROUND: Chronic rhinosinusitis (CRS) is a chronic inflammatory disease characterized by persistent inflammation and bacterial infection. Ciprofloxacin and azithromycin are commonly prescribed antibiotics for CRS, but the ability to provide targeted release in the sinuses could mitigate side effects and improve drug concentrations at the infected site. This study was aimed to evaluate the efficacy of the novel ciprofloxacin-azithromycin sinus stent (CASS) in vitro. METHODS: The CASS was created by coating ciprofloxacin (hydrophilic, inner layer) and azithromycin (hydrophobic, outer layer) onto a biodegradable poly-l-lactic acid (PLLA) stent. In-vitro evaluation included: (1) assessment of drug-coating stability within the stent using scanning electron microscopy (SEM); (2) determination of ciprofloxacin and azithromycin release kinetics; and (3) assessment of anti-biofilm activities against Pseudomonas aeruginosa. RESULTS: The ciprofloxacin nanoparticle suspension in the inner layer was confirmed by zeta potential. Both ciprofloxacin (60 µg) and azithromycin (3 mg) were uniformly coated on the surface of the PLLA stents. The CASS showed ciprofloxacin/azithromycin sustained release patterns, with 80.55 ± 11.61% of ciprofloxacin and 93.85 ± 6.9% of azithromycin released by 28 days. The CASS also significantly reduced P aeruginosa biofilm mass compared with bare stents and controls (relative optical density units at 590-nm optical density: CASS, 0.037 ± 0.006; bare stent, 0.911 ± 0.015; control, 1.000 ± 0.000; p < 0.001; n = 3). CONCLUSION: The CASS maintains a uniform coating and sustained delivery of ciprofloxacin and azithromycin, providing anti-biofilm activities against P aeruginosa. Further studies evaluating the efficacy of CASS in a preclinical model are planned.