Changes in oral health during aging in a novel non-human primate model.

Oral health plays a significant role in the quality of life and overall well-being of the aging population. However, age-related changes in oral health are not well understood due to challenges with current animal models. In this study, we analyzed the oral health and microbiota of a short-lived non-human primate (i.e., marmoset), as a step towards establishing a surrogate for studying the changes that occur in oral health during human aging. We investigated the oral health of marmosets using cadaveric tissues in three different cohorts: young (aged ≤6 years), middle-aged, and older (>10 years) and assessed the gingival bacterial community using analyses of the V3-V4 variable region of 16S rRNA gene. The oldest cohort had a significantly higher number of dental caries, increased dental attrition/erosion, and deeper periodontal pocket depth scores. Oral microbiome analyses showed that older marmosets had a significantly greater abundance of Escherichia-Shigella and Propionibacterium, and a lower abundance of Agrobacterium/Rhizobium at the genus level. Alpha diversity of the microbiome between the three groups showed no significant differences; however, principal coordinate analysis and non-metric multidimensional scaling analysis revealed that samples from middle-aged and older marmosets were more closely clustered than the youngest cohort. In addition, linear discriminant analysis effect size (LEFSe) identified a higher abundance of Esherichia-Shigella as a potential pathogenic biomarker in older animals. Our findings confirm that changes in the oral microbiome are associated with a decline in oral health in aging marmosets. The current study suggests that the marmoset model recapitulates some of the changes in oral health associated with human aging and may provide opportunities for developing new preventive strategies or interventions which target these disease conditions.

Extracellular matrix turnover in salivary gland disorders and regenerative therapies: Obstacles and opportunities.

Salivary gland (SG) extracellular matrix (ECM) has a major influence on tissue development, homeostasis, and tissue regeneration after injury. During aging, disease, and physical insult, normal remodeling of the SG microenvironment (i.e. ECM) becomes dysregulated, leading to alterations in matrix composition which disrupt tissue architecture/structure, alter cell activity, and negatively impact gland function. Matrix metalloproteinases (MMPs) are a large and diverse family of metalloendopeptidases which play a major role in matrix degradation and are intimately involved in regulating development and cell function; dysregulation of these enzymes leads to the production of a fibrotic matrix. In the SG this altered fibrotic ECM (or cell microenvironment) negatively impacts normal cell function and the effectiveness of gene and stem cell therapies which serve as a foundation for many SG regenerative therapies. For this reason, prospective regenerative strategies should prioritize the maintenance and/or restoration of a healthy SG ECM. Mesenchymal stem cells (MSCs) have great potential for mitigating damage to the SG microenvironment by ameliorating inflammation, reducing fibrosis, and repairing the damaged milieu of extracellular regulatory cues, including the matrix. This review addresses our current understanding of the impact of aging and disease on the SG microenvironment and suggests critical deficiencies and opportunities in ECM-targeted therapeutic interventions.

Autologous mesenchymal stem cells offer a new paradigm for salivary gland regeneration.

Salivary gland (SG) dysfunction, due to radiotherapy, disease, or aging, is a clinical manifestation that has the potential to cause severe oral and/or systemic diseases and compromise quality of life. Currently, the standard-of-care for this condition remains palliative. A variety of approaches have been employed to restore saliva production, but they have largely failed due to damage to both secretory cells and the extracellular matrix (niche). Transplantation of allogeneic cells from healthy donors has been suggested as a potential solution, but no definitive population of SG stem cells, capable of regenerating the gland, has been identified. Alternatively, mesenchymal stem cells (MSCs) are abundant, well characterized, and during SG development/homeostasis engage in signaling crosstalk with the SG epithelium. Further, the trans-differentiation potential of these cells and their ability to regenerate SG tissues have been demonstrated. However, recent findings suggest that the "immuno-privileged" status of allogeneic adult MSCs may not reflect their status post-transplantation. In contrast, autologous MSCs can be recovered from healthy tissues and do not present a challenge to the recipient's immune system. With recent advances in our ability to expand MSCs in vitro on tissue-specific matrices, autologous MSCs may offer a new therapeutic paradigm for restoration of SG function.

New horizons in understanding oral health and swallowing function within the context of frailty.

Frailty is a complex and multidimensional condition wherein declines in physiologic reserve and function place individuals in a state of heightened vulnerability and decreased resiliency. There has been growing interest in both research and clinical settings to understand how to best define, assess and characterise frailty in older adults. To this end, various models and clinical assessment tools have been used to define and measure frailty. While differences exist among these models and tools, a common unifying theme is a focus on physical function and activity. Notably absent across many available conceptual models and clinical tools are items directly related to oral and swallowing function. This is an important oversight as widespread changes to both oral and swallowing function are evident in older adults. Indeed, emerging evidence suggests many of the functional domains affected in frail older adults, such as nutrition and sarcopenia, have cyclical relationships with impairments in oral (oral hypofunction) and swallowing function (dysphagia) as well. The increasing appreciation for the interrelationships among oral hypofunction, dysphagia and frailty provides an opportunity for refinement of frailty assessment and characterisation in older adults to incorporate metrics specific to oral and swallowing function.

Periodontopathic bacterial adhesion to different restorative materials used to elevate proximal subgingival margins.

This study compared the periodontopathic bacterial adhesion to four restorative materials used for deep margin elevation at 2, 24, and 48-h after incubation. Discs were produced from four restorative materials: resin modified glass ionomer, glass hybrid, flowable bulk fill resin composite, and bioactive ionic resin. Root dentin was used as control. Specimens were coated with saliva and used to culture a biofilm comprised of three strains of periodontopathic bacteria; Porphyromonas gingivalis, Prevotella intermedia, and Aggregatibacter actinomycetemcomitans. Bacterial adherence was assessed by colony count assay, crystal violet staining, and visualized using confocal laser scanning microscopy. Data were analyzed by two-way ANOVA followed by Tukey's post hoc tests. The adhesion values for the control specimens were significantly higher than for other materials, while those for the flowable bulk fill were significantly lower than for any other material within all evaluation assays. The 2-h incubation period showed the lowest adhesion values regardless of the group. The 48-h adhesion values were higher than the 24-h results in all groups except the flowable bulk fill. Microscopic imaging partially supported the findings of the measurements. In terms of periodontopathic bacterial adhesion, the tested flowable bulk fill may be preferable for subgingival use over other tested materials.

Follow-up study of veterans with white and red oral mucosal lesions at Veterans Affairs Dental Clinics.

OBJECTIVES: This analysis examined the clinical and histopathological characteristics of white and red oral mucosal lesions and patient lifestyle behaviors to understand how the lesions changed over 19-23 years, including among patients who developed oral and pharyngeal cancer. MATERIALS AND METHODS: Seventy-five individuals with red and/or white oral mucosal lesions with clinical diagnoses of smokeless tobacco lesions, leukoplakia, erythroplakia, lichen planus, ulcer, and virus-associated lesions were identified in six Veterans Affairs Medical Center Dental Clinics (VAMC) from 1996 to 2001. Biopsy results and patients' sociodemographic, medical, and tobacco/alcohol use characteristics were obtained. Study dentists used standardized forms to capture information about the lesions. Study participants were re-examined at intervals through January 2002. In 2020, a retrospective review of VAMC and public records ascertained whether participants developed oral cancer or died. RESULTS: The most common red or white oral mucosal lesions among the 75 study participants were leukoplakia (36.0%), smokeless tobacco lesions (26.7%), virus-associated lesions (18.7%), and lichen planus (16.0%). Lesions in 11% of participants with leukoplakia and one-third of participants with lichen planus persisted for 5 years or more. Dysplasia was present in four participants with leukoplakia. Seventeen percent of participants developed a new white or red oral mucosal lesion. Five patients (6.1%) developed oral or pharyngeal cancer, four among participants with leukoplakia (one with prior dysplasia) and one among participants with lichen planus. Four of the cancers developed 6-20 years after enrollment, and only one was at the original lesion site. CONCLUSIONS: The occurrence of oral and pharyngeal cancers in some study participants with white and red oral mucosal lesions many years after enrollment reinforces the need for patients, dentists, and health care systems to have better methods to identify and assess the malignant potential of oral lesions, monitor patients over time, and intercept high-risk oral lesions before they become cancerous.

Organ-specific extracellular matrix directs trans-differentiation of mesenchymal stem cells and formation of salivary gland-like organoids in vivo.

BACKGROUND: Current treatments for salivary gland (SG) hypofunction are palliative and do not address the underlying cause or progression of the disease. SG-derived stem cells have the potential to treat SG hypofunction, but their isolation is challenging, especially when the tissue has been damaged by disease or irradiation for head and neck cancer. In the current study, we test the hypothesis that multipotent bone marrow-derived mesenchymal stem cells (BM-MSCs) in a rat model are capable of trans-differentiating to the SG epithelial cell lineage when induced by a native SG-specific extracellular matrix (SG-ECM) and thus may be a viable substitute for repairing damaged SGs. METHODS: Rat BM-MSCs were treated with homogenates of decellularized rat SG-ECM for one hour in cell suspension and then cultured in tissue culture plates for 7 days in growth media. By day 7, the cultures contained cell aggregates and a cell monolayer. The cell aggregates were hand-selected under a dissecting microscope, transferred to a new tissue culture dish, and cultured for an additional 7 days in epithelial cell differentiation media. Cell aggregates and cells isolated from the monolayer were evaluated for expression of SG progenitor and epithelial cell specific markers, cell morphology and ultrastructure, and ability to form SG-like organoids in vivo. RESULTS: The results showed that this approach was very effective and guided the trans-differentiation of a subpopulation of CD133-positive BM-MSCs to the SG epithelial cell lineage. These cells expressed amylase, tight junction proteins (Cldn 3 and 10), and markers for SG acinar (Aqp5 and Mist 1) and ductal (Krt 14) cells at both the transcript and protein levels, produced intracellular secretory granules which were morphologically identical to those found in submandibular gland, and formed SG-like organoids when implanted in the renal capsule in vivo. CONCLUSIONS: The results of this study suggest the feasibility of using autologous BM-MSCs as an abundant source of stem cells for treating SG hypofunction and restoring the production of saliva in these patients.

Matrix-bound Cyr61/CCN1 is required to retain the properties of the bone marrow mesenchymal stem cell niche but is depleted with aging.

Previously, we showed that extracellular matrices (ECMs), produced ex vivo by various types of stromal cells, direct bone marrow mesenchymal stem cells (BM-MSCs) in a tissue-specific manner and recapitulate physiologic changes characteristic of the aging microenvironment. In particular, BM-MSCs obtained from elderly donors and cultured on ECM produced by young BM stromal cells showed improved quantity, quality and osteogenic differentiation. In the present study, we searched for matrix components that are required for a functional BM-MSC niche by comparing ECMs produced by BM stromal cells from "young" (≤25 y/o) versus "elderly" (≥60 y/o) donors. With increasing donor age, ECM fibrillar organization and mechanical integrity deteriorated, along with the ability to promote BM-MSC proliferation and responsiveness to growth factors. Proteomic analyses revealed that the matricellular protein, Cyr61/CCN1, was present in young, but undetectable in elderly, BM-ECM. To assess the role of Cyr61 in the BM-MSC niche, we used genetic methods to down-regulate the incorporation of Cyr61 during production of young ECM and up-regulate its incorporation in elderly ECM. The results showed that Cyr61-depleted young ECM lost the ability to promote BM-MSC proliferation and growth factor responsiveness. However, up-regulating the incorporation of Cyr61 during synthesis of elderly ECM restored its ability to support BM-MSC responsiveness to osteogenic factors such as BMP-2 and IGF-1. We next examined aging bone and compared bone mineral density and Cyr61 content of L4-L5 vertebral bodies in "young" (9-11 m/o) and "elderly" (21-33 m/o) mice. Our analyses showed that low bone mineral density was associated with decreased amounts of Cyr61 in osseous tissue of elderly versus young mice. Our results strongly demonstrate a novel role for ECM-bound Cyr61 in the BM-MSC niche, where it is responsible for retention of BM-MSC proliferation and growth factor responsiveness, while depletion of Cyr61 from the BM niche contributes to an aging-related dysregulation of BM-MSCs. Our results also suggest new potential therapeutic targets for treating age-related bone loss by restoring specific ECM components to the stem cell niche.

In vitro effect of an oral spray and mouthrinses on dual species cariogenic bacteria biofilm.

PURPOSE: To determine the efficacy of an oral spray and oral rinses to inhibit oral cariogenic dual species biofilm formation on hydroxyapatite (HA) discs. METHODS: The Streptococcus mutans (NCTC 10449, ATCC), Lactobacilli casei (NCIB 8820, ATCC) dual species biofilm formation and inhibition on HA disc was tested using five antimicrobial products, i.e., oral spray (Oral Shield), Mouthrinse (Listerine Ultra Clean, Listerine Cool Mint, Crest Pro-Health, ACT Restoring). An untreated group served as control. The established biofilm on the surface of each disc was treated or untreated with oral spray and mouthrinse for 2 minutes after 24 or 48 hours. The dual species biofilm formation and inhibition on HA discs was determined using the spread plate method and colonies were counted and expressed as colony forming units (CFU/mL). Further, the HA disc was subjected to confocal laser scanning microscope (CLSM) examination to determine the viability of cells using live-dead staining and a scanning electron microscope (SEM) to examine the effect on bacteria biofilm and morphology. The cytotoxic effect of test spray and mouthrinse was tested on OKF6/TERT-2 cells using the MTT method. RESULTS: At each time point, 24- or 48-hours, S. mutans and L. casei mixed biofilm on HA discs had a significantly (P> 0.001) fewer number of bacteria in the treated groups than the untreated one. The oral spray and mouthrinses had a detrimental effect on bacteria biofilm, morphology and cell wall, whereas no significant changes were observed in the untreated group. Cytotoxic assay revealed that the oral spray was safe for human oral keratinocyte cells. CLINICAL SIGNIFICANCE: The tested oral spray could offer potential to inhibit the cariogenic bacteria and protect the tooth enamel from cariogenic bacterial biofilm.

Stress-Busting Program for Family Caregivers: Validation of the Spanish version using biomarkers and quality-of-life measures.

Hispanic family caregivers of people with dementia experience higher levels of stress compared to non-Hispanic white caregivers. Long-term stress causes depression, caregiver burden, cellular aging, and dysregulation of the immune, nervous, and endocrine systems. The purpose of this study was to determine the validity of the Spanish version of the English Stress-Busting Program (SBP) for Family Caregivers by determining changes in quality-of-life measures and biomarkers. Thirty-six caregivers completed the SBP in the language of their choice (14 Spanish-speaking Hispanics [HS], 8 English-speaking Hispanics [HE], and 14 non-Hispanic English [NHE] speakers). Quality-of-life measures included the Perceived Stress Scale, the Screen for Caregiver Burden, and the Center for Epidemiologic Studies Depression Scale. Assessment of oral health and immunity included salivary flow rate, pH, buffer capacity, total protein, and secretory immunoglobulin A (sIgA). Indicators of stress (salivary cortisol), inflammation (C-reactive protein), and cellular aging (leukocyte telomere length) were assessed. Following completion of the SBP, the Spanish-speaking group had less depression and caregiver burden along with improved oral health and reduced cellular aging. When comparing baseline values to post-intervention, all three groups showed significant improvement in subjective caregiver burden. When the data from all three groups were combined, biomarkers that showed improvement after nine weeks of SBP included the stress hormone cortisol, salivary pH, and leukocyte telomere length. The results indicate that the Spanish SBP reduces caregiver stress as assessed by quality-of-life indicators and biomarkers. The Spanish SBP can help to mitigate health disparities in Hispanic Spanish-speaking caregivers.

ß2-Adrenergic receptor agonist induced hepatic steatosis in mice: modeling nonalcoholic fatty liver disease in hyperadrenergic states.

Nonalcoholic fatty liver disease (NAFLD) is a spectrum of disorders ranging from hepatic steatosis [excessive accumulation of triglycerides (TG)] to nonalcoholic steatohepatitis, which can progress to cirrhosis and hepatocellular carcinoma. The molecular pathogenesis of steatosis and progression to more severe NAFLD remains unclear. Obesity and aging, two principal risk factors for NAFLD, are associated with a hyperadrenergic state. ß-Adrenergic responsiveness in liver increases in animal models of obesity and aging, and in both is linked to increased hepatic expression of ß2-adrenergic receptors (ß2-ARs). We previously showed that in aging rodents intracellular signaling from elevated hepatic levels of ß2-ARs may contribute to liver steatosis. In this study we demonstrate that injection of formoterol, a highly selective ß2-AR agonist, to mice acutely results in hepatic TG accumulation. Further, we have sought to define the intrahepatic mechanisms underlying ß2-AR mediated steatosis by investigating changes in hepatic expression and cellular localization of enzymes, transcription factors, and coactivators involved in processes of lipid accrual and disposition-and also functional aspects thereof-in livers of formoterol-treated animals. Our results suggest that ß2-AR activation by formoterol leads to increased hepatic TG synthesis and de novo lipogenesis, increased but incomplete ß-oxidation of fatty acids with accumulation of potentially toxic long-chain acylcarnitine intermediates, and reduced TG secretion-all previously invoked as contributors to fatty liver disease. Experiments are ongoing to determine whether sustained activation of hepatic ß2-AR signaling by formoterol might be utilized to model fatty liver changes occurring in hyperadrenergic states of obesity and aging, and thereby identify novel molecular targets for the prevention or treatment of NAFLD.NEW & NOTEWORTHY Results of our study suggest that ß2-adrenergic receptor (ß2-AR) activation by agonist formoterol leads to increased hepatic TG synthesis and de novo lipogenesis, incomplete ß-oxidation of fatty acids with accumulation of long-chain acylcarnitine intermediates, and reduced TG secretion. These findings may, for the first time, implicate a role for ß2-AR responsive dysregulation of hepatic lipid metabolism in the pathogenetic processes underlying NAFLD in hyperadrenergic states such as obesity and aging.

Culture on a native bone marrow-derived extracellular matrix restores the pancreatic islet basement membrane, preserves islet function, and attenuates islet immunogenicity.

Islet transplantation in man is limited by multiple factors including islet availability, islet cell damage caused by collagenase during isolation, maintenance of islet function between isolation and transplantation, and allograft rejection. In this study, we describe a new approach for preparing islets that enhances islet function in vitro and reduces immunogenicity. The approach involves culture on native decellularized 3D bone marrow-derived extracellular matrix (3D-ECM), which contains many of the matrix components present in pancreas, prior to islet transplantation. Compared to islets cultured on tissue culture plastic (TCP), islets cultured on 3D-ECM exhibited greater attachment, higher survival rate, increased insulin content, and enhanced glucose-stimulated insulin secretion. In addition, culture of islets on 3D-ECM promoted recovery of vascular endothelial cells within the islets and restored basement membrane-related proteins (eg, fibronectin and collagen type VI). More interestingly, culture on 3D-ECM also selectively decontaminated islets of "passenger" cells (co-isolated with the islets) and restored basement membrane-associated type VI collagen, which were associated with an attenuation in islet immunogenicity. These results demonstrate that this novel approach has promise for overcoming two major issues in human islet transplantation: (a) poor yield of islets from donated pancreas tissue and (b) the need for life-long immunosuppression.

Oral and Craniofacial Stem Cells: An Untapped Source for Neural Tissue Regeneration.

Nerve tissue regeneration continues to represent an intractable obstacle to realizing the promise of tissue engineering. Although neurobiology works to shed light on the mechanisms governing neuronal growth and repair, considerable technical gaps remain that hinder progress. Chief among these is the absence of an appropriate culture environment to faithfully reproduce the neuronal niche ex vivo. We propose that the various multipotent cells found in the oral cavity may represent an important yet underutilized resource for preparing such neurogenic microenvironments. Similar to those of nerve tissue, these cell populations are of ectodermal origin and have clinically demonstrated neurogenic potential. Although there is a lack of consensus on whether putative types of oral and craniofacial stem cells constitute distinct populations, their contribution to neural tissue engineering may be twofold: as a cellular feedstock for neoneurogenesis and for the production of specialized in vitro environments for neurogenic differentiation, phenotype maintenance, and use in therapeutic applications. Impact statement We propose that addressing gaps in understanding the neurogenic role of dental stem cells and their microenvironment may yield efficient and reliable strategies for long-term neuronal cell culture and open new avenues for neural regeneration in both dental, nerve, and other tissues.

Native extracellular matrix, synthesized ex vivo by bone marrow or adipose stromal cells, faithfully directs mesenchymal stem cell differentiation.

Mesenchymal stem cells (MSCs) are highly responsive to cues in the microenvironment (niche) that must be recapitulated ex vivo to study their authentic behavior. In this study, we hypothesized that native bone marrow (BM)- and adipose (AD)-derived extracellular matrices (ECM) were unique in their ability to control MSC behavior. To test this, we compared proliferation and differentiation of bone marrow (BM)-derived MSCs when maintained on native decellularized ECM produced by BM versus AD stromal cells (i.e. BM- versus AD-ECM). We found that both ECMs contained similar types of collagens but differed in the relative abundance of each. Type VI collagen was the most abundant (≈60% of the total collagen present), while type I was the next most abundant at ≈30%. These two types of collagen were found in nearly equal proportions in both ECMs. In contrast, type XII collagen was almost exclusively found in AD-ECM, while types IV and V were only found in BM-ECM. Physically and mechanically, BM-ECM was rougher and stiffer, but less adhesive, than AD-ECM. During 14 days in culture, both ECMs supported BM-MSC proliferation better than tissue culture plastic (TCP), although MSC-related surface marker expression remained relatively high on all three culture surfaces. BM-MSCs cultured in osteogenic (OS) differentiation media on BM-ECM displayed a significant increase in calcium deposition in the matrix, indicative of osteogenesis, while BM-MSCs cultured on AD-ECM in the presence of adipogenic (AP) differentiation media showed a significant increase in Oil Red O staining, indicative of adipogenesis. Further, culture on BM-ECM significantly increased BM-MSC-responsiveness to rhBMP-2 (an osteogenic inducer), while culture on AD-ECM enhanced responsiveness to rosiglitazone (an adipogenic inducer). These findings support our hypothesis and indicate that BM- and AD-ECMs retain unique elements, characteristic of their tissue-specific microenvironment (niche), which promote retention of MSC differentiation state (i.e. "stemness") during expansion and direct cell response to lineage-specific inducers. This study provides a new paradigm for precisely controlling MSC fate to a desired cell lineage for tissue-specific cell-based therapies.

Can oral health and oral-derived biospecimens predict progression of dementia?

Growing evidence indicates that oral health and brain health are interconnected. Declining cognition and dementia coincide with lack of self-preservation, including oral hygiene. The oral microbiota plays an important role in maintaining oral health. Emerging evidence suggests a link between oral dysbiosis and cognitive decline in patients with Alzheimer's disease. This review showcases the recent advances connecting oral health and cognitive function during aging and the potential utility of oral-derived biospecimens to inform on brain health. Collectively, experimental findings indicate that the connection between oral health and cognition cannot be underestimated; moreover, oral biospecimens are abundant and readily obtainable without invasive procedures, which may help inform on cognitive health.

Implementation of a modified quality assurance protocol for CBCT machines within UT health San Antonio School of Dentistry.

OBJECTIVES: To implement the quality control assurance protocol (including the re-establishment of baseline data from 2016) to monitor the stability of image quality of CBCT machines located within the UT Health San Antonio School of Dentistry. METHODS AND MATERIALS: Five CBCT machines ProMax 3D Mid® (Planmeca Oy, Helsinki, Finland), 3D Accuitomo XYZ Slice View Tomograph® (Model MCT-1, Type EX-1F8; Fushimi-ku, Kyoto: J. Morita Mfg. Corp), Veraviewepocs 3D (Model R100; Fushimi-ku, Kyoto: J. Morita Mfg. Corp), PreXion3D Excelsior® (PreXion, San Mateo, CA), and i-CAT FLX Series® (Imaging Sciences International, Hatfield, PA) were tested for Artifact, Contrast-to-Noise Ratio, Noise, Spatial Resolution, and Contrast Resolution using a custom insert configuration in the SEDENTEXCT IQ phantom. RESULTS: Four-scan benchmark mean values for Artifact, Contrast-to-Noise, Noise, Spatial Resolution, and Contrast Resolution were determined for the five machines tested with associated alert and action level thresholds calculated. CONCLUSION: This newly developed QA protocol established image quality baseline values. Recommended tests, frequency, and actions levels have been updated and control charts established for future trend analysis to enable proper implementation of a QA protocol monitoring CBCT machines at UT health San Antonio.

Wheat germ agglutinin liposomes with surface grafted cyclodextrins as bioadhesive dual-drug delivery nanocarriers to treat oral cells.

Topical management of oral infection requires combined use of multiple classes of drugs and frequent dosing due to low drug retention rates. The sustained, co-delivery of drugs with different solubilities to cells using nanoparticle drug delivery systems remains a challenge. Here, we developed wheat germ agglutinin (WGA) conjugated liposomes with surface grafted cyclodextrin (WGA-liposome-CD) as bioadhesive dual-drug nanocarriers. We effectively encapsulated two physiochemically different drugs (ciprofloxacin and betamethasone) and demonstrated sustained co-drug release in saliva over a 24 h period in vitro. As proof of therapeutic utility in oral cells, we infected oral keratinocytes with Aggregatibacter actinomycetemcomitans, a bacterial pathogen responsible for chronic periodontal disease. Drug release, resulting from nanocarrier cell binding, produced a significant increase in oral cell survival and synergistically reduced inflammation. These results suggest that WGA-liposome-CD nanocarriers are novel cyto-adhesive candidates for delivering multiple drugs with sustained therapeutic activity for localized drug delivery to oral cells.

Amphiphilic quaternary ammonium chitosan/sodium alginate multilayer coatings kill fungal cells and inhibit fungal biofilm on dental biomaterials.

Formation of fungal biofilms on health care-related materials causes serious clinical consequences. This study reports a novel fungal repelling strategy to control fungal biofilm formation on denture biomaterials through layer-by-layer self-assembly (LBL). Amphiphilic quaternary ammonium chitosans (CS612) were synthesized and used as the antimicrobial positive layer, and sodium alginate (SA) was chosen as the negative layer to construct LBL multilayers on poly (methyl methacrylate) (PMMA)-based denture materials. The presence of LBL multilayers on denture disc was confirmed and characterized by surface zeta potential, water contact angle, AFM, and FT-IR analyses. The multilayer coatings, especially CS612 as the outmost layer, effectively prevented the fungal initial adhesion and biofilm formation. The Candida cells avoided the multilayer coatings and suspended in broth solution instead of forming biofilms, suggesting that the LBL multilayers had fungal repelling effects. The LBL multilayers were biocompatible toward mammalian cells. In stability tests, after immersion in PBS for 4 weeks under constant shaking and repeated brushing with a denture brush for up to 3000 times, the biofilm-controlling effects of the LBL multilayers were not affected, pointing to a novel long-term strategy in controlling fungal biofilms on denture and other related biomaterials.

Salivary polypeptide/hyaluronic acid multilayer coatings act as "fungal repellents" and prevent biofilm formation on biomaterials.

Candida-associated denture stomatitis (CADS) is a common, recurring clinical complication in denture wearers that can lead to serious oral and systemic health problems. Current management strategies are not satisfactory due to their short-acting and ineffective therapeutic effects. Here, we describe a new fungal biofilm controlling strategy using the polyelectrolyte layer-by-layer (LBL) self-assembly technology on denture materials. Conventional poly(methyl methacrylate) (PMMA) denture material discs were functionalized with negatively charged poly(methacrylic acid) (PMAA) via plasma-initiated surface grafting, followed by repetitive alternating coating with the salivary antimicrobial polypeptide histatin 5 (H-5; cationic polymer) and hyaluronic acid (HA; anionic polymer). On the other hand, the H-5/HA LBL coatings (i.e., the outermost layer was H-5) inhibited fungal attachment/adhesion, significantly reduced fungal biofilm formation, and showed synergistic effects with the antifungal drug miconazole. LBL surface hydrophilicity was not the key mechanism in controlling Candida biofilm formation. The current approach demonstrates the utility of a new design principle for fabricating anticandidal denture materials, as well as potentially other related medical devices, for controlling fungal biofilm formation and combating insidious infections.

Lectin-Conjugated Liposomes as Biocompatible, Bioadhesive Drug Carriers for the Management of Oral Ulcerative Lesions.

Oral ulcerative lesions are painful and debilitating, particularly for immunosuppressed patients undergoing chemotherapeutic/irradiation treatment. Their clinical management requires multiple drugs to be administered simultaneously. Current formulations available to patients require frequent dosing, leading to poor compliance and suboptimal clinical outcomes. In this study, we prepared wheat germ agglutinin (WGA)-conjugated liposomes (WGA liposomes) to serve as bioadhesive drug carriers that can encapsulate various classes of drugs, rapidly bind to oral epithelial cells within minutes, and stay on the cells to provide sustained, localized drug release for days. Fluorescence binding studies found a significant increase (p < 0.05) in the binding of WGA liposomes to oral cells in as short an incubation time as 1 min compared to that for nonconjugated liposomes. WGA liposomes encapsulating model drug amoxicillin showed sustained in vitro drug release, and the released drugs provided potent antimicrobial activity against Streptococcus mutans in an oral epithelial-bacterial coculture system. Exocytosis studies confirmed that the WGA liposomes stayed within the oral cells for 48 h, after which the cells completely removed the liposomes. Moreover, cell viability studies showed that there was a significant reduction in oral cell damage when the bacterially infected cells were treated with amoxicillin-loaded WGA liposomes compared to the untreated controls. These results point to the great potential of the lectin-conjugated liposomes as cell-binding drug-delivery systems in achieving localized, sustained drug release for the management of oral ulcerative lesions and other related complications.