An inflection point in high-throughput proteomics with Orbitrap Astral: analysis of biofluids, cells, and tissues.

This technical note presents a comprehensive proteomics workflow for the new combination of Orbitrap and Astral mass analyzers across biofluids, cells, and tissues. Central to our workflow is the integration of Adaptive Focused Acoustics (AFA) technology for cells and tissue lysis, to ensure robust and reproducible sample preparation in a high-throughput manner. Furthermore, we automated the detergent-compatible single-pot, solid-phase-enhanced sample Preparation (SP3) method for protein digestion, a technique that streamlines the process by combining purification and digestion steps, thereby reducing sample loss and improving efficiency. The synergy of these advanced methodologies facilitates a robust and high-throughput approach for cells and tissue analysis, an important consideration in translational research. This work disseminates our platform workflow, analyzes the effectiveness, demonstrates reproducibility of the results, and highlights the potential of these technologies in biomarker discovery and disease pathology. For cells and tissues (heart, liver, lung, and intestine) proteomics analysis by data-independent acquisition mode, identifications exceeding 10,000 proteins can be achieved with a 24-minute active gradient. In 200ng injections of HeLa digest across multiple gradients, an average of more than 80% of proteins have a CV less than 20%, and a 45-minute run covers ~90% of the expressed proteome. In plasma samples including naive, depleted, perchloric acid precipitated, and Seer nanoparticle captured, all with a 24-minute gradient length, we identified 87, 108, 96 and 137 out of 216 FDA approved circulating protein biomarkers, respectively. This complete workflow allows for large swaths of the proteome to be identified and is compatible across diverse sample types.

Using Policy Briefs to Communicate Dental Research Findings to Policymakers.

OBJECTIVES: New scientific knowledge is not always available to decision makers. Policy briefs are a way that dental researchers can communicate research findings to policymakers. This study compares usefulness of 2 types of policy briefs about sugar-sweetened beverage (SSB) intake and tooth decay. METHODS: We developed 2 policy brief types (data focused and narrative focused) and emailed a randomly assigned policy brief to 825 policymakers and staff from 3 levels of government (city, county, and state) in Washington State. Participants completed a 22-item online questionnaire. There were 4 study outcomes: whether the brief was understandable, whether the brief was credible, likelihood of use, and likelihood to be shared (each measured on a 5-point Likert-like scale). The t test was used to evaluate whether outcomes differed by policy brief type and government level (α = 0.05). RESULTS: There were 108 respondents (adjusted response rate 14.6%). About 41.6% of participants were in city government, 26.9% were in county government, and 29.6% were in state government. Participants reported that both data- and narrative-focused briefs were understandable (mean rating [MR] and standard deviation [SD]: 4.15 ± 0.68 and 4.09 ± 0.81, respectively; P = 0.65) and credible (MR and SD: 4.13 ± 0.70 and 4.09 ± 0.70, respectively; P = 0.74), but they were not likely to use (MR and SD: 2.71 ± 1.15 and 2.55 ± 1.28, respectively; P = 0.51) or share it (MR and SD: 2.62 ± 1.04 and 2.66 ± 1.30, respectively; P = 0.87). The likelihood of sharing briefs differed significantly by level of government (P = 0.017). Participants at the state level were more likely to share information from the briefs (mean rating and SD: 3.10 ± 0.80) than city- and county-level participants (MR and SD: 2.62 ± 1.27, and 2.24 ± 1.21, respectively). CONCLUSION: Both data- and narrative-focused policy briefs may be a useful way to communicate dental research findings to policymakers, but additional steps are needed to ensure that briefs are used and shared. KNOWLEDGE TRANSFER STATEMENT: Researchers should disseminate their research findings to maximize scientific impact. Our study findings indicate that policy briefs may be a useful way to communicate dental research findings to policymakers, but additional research is needed on the best ways to disseminate findings.

A Qualitative Study on Dentists' Communication Approaches in Managing Fluoride-Hesitant Caregivers.

OBJECTIVES: Guidelines on effective provider-led communication are available but may be underused in dentistry, even if such guidelines could help dentists manage complex clinical scenarios like topical fluoride hesitancy. The purpose of this study was to investigate current chairside communication approaches used by dentists with fluoride-hesitant caregivers. METHODS: A 27-item semistructured interview script was developed and pretested with 3 dentists, revised, and finalized. One-on-one interviews were conducted with a purposive sample of pediatric dentists and general dentists from April to June 2020. Interviews were digitally recorded, transcribed, and analyzed to identify dentists' communication approaches used during clinical interactions with fluoride-hesitant caregivers. Thematic analyses identified themes and subthemes, and exemplary quotes were provided to illustrate each theme. RESULTS: Twenty-seven dentists participated (21 pediatric dentists and 6 general dentists). The mean age of participants was 43.0 ± 8.2 y (range, 30-73). Most participants were women (88.9%), white (51.9%), and non-Hispanic (85.2%). Participants had been practicing dentistry for a mean of 13.2 ± 10.5 y (range, 2-40). There were 4 themes: leaving topical fluoride decisions completely up to the caregiver, educating the caregiver about fluoride, insisting that the caregiver accept fluoride, and engaging the caregiver and child. CONCLUSION: Most communications approaches used by interviewed dentists to manage fluoride hesitancy in clinical settings are not evidence based. Future dental education efforts should ensure that trainees are exposed to and can demonstrate competency in appropriate, evidence-based patient-provider communication strategies. KNOWLEDGE TRANSFER STATEMENT: The study highlights the need for dentists to apply evidence-based communication strategies when managing difficult clinical scenarios like fluoride hesitancy, which is important in optimizing dentist-patient trust.

Periostin as a blood biomarker of muscle cell fibrosis, cardiomyopathy and disease severity in myotonic dystrophy type 1.

BACKGROUND AND PURPOSE: Myotonic dystrophy type 1 (DM1) is the most common form of adult-onset muscular dystrophy and is caused by an repeat expansion [r(CUG)exp] located in the 3' untranslated region of the DMPK gene. Symptoms include skeletal and cardiac muscle dysfunction and fibrosis. In DM1, there is a lack of established biomarkers in routine clinical practice. Thus, we aimed to identify a blood biomarker with relevance for DM1-pathophysiology and clinical presentation. METHODS: We collected fibroblasts from 11, skeletal muscles from 27, and blood samples from 158 DM1 patients. Moreover, serum, cardiac, and skeletal muscle samples from DMSXL mice were included. We employed proteomics, immunostaining, qPCR and ELISA. Periostin level were correlated with CMRI-data available for some patients. RESULTS: Our studies identified Periostin, a modulator of fibrosis, as a novel biomarker candidate for DM1: proteomic profiling of human fibroblasts and murine skeletal muscles showed significant dysregulation of Periostin. Immunostaining on skeletal and cardiac muscles from DM1 patients and DMSXL mice showed an extracellular increase of Periostin, indicating fibrosis. qPCR studies indicated increased POSTN expression in fibroblasts and muscle. Quantification of Periostin in blood samples from DMSXL mice and two large validation cohorts of DM1 patients showed decreased levels in animals and diseased individuals correlating with repeat expansion and disease severity and presence of cardiac symptoms identified by MRI. Analyses of longitudinal blood samples revealed no correlation with disease progression. CONCLUSIONS: Periostin might serve as a novel stratification biomarker for DM1 correlating with disease severity, presence of cardiac malfunction and fibrosis.

Impact of Fluoride on Associations between Free Sugars Intake and Dental Caries in US Children.

OBJECTIVE: Dental caries is the most prevalent chronic disease in US children, with the highest burden among Black and Hispanic youth. Sugars are a primary risk factor, but few studies have specifically measured intakes of free sugars and related this to dental caries or explored the extent to which water fluoride mitigates the cariogenicity of free sugars. Furthermore, the cariogenicity of certain free sugars sources, such as extruded fruit and vegetable products, is unclear. METHODS: Using cross-sectional data on 4,906 children aged 2 to 19 y in the US National Health and Nutrition Examination Survey 2013-2016, we examined associations of free sugars intake with counts of decayed or filled primary tooth surfaces (dfs) and decayed, missing, or filled permanent surfaces (DMFS) in negative binomial regressions. Stratified models examined these associations in children with home water fluoride above or below the Centers for Disease Control and Prevention (CDC)-recommended level of 0.7 ppm. RESULTS: Free sugars accounted for 16.4% of energy, primarily contributed by added sugars. In adjusted models, a doubling in the percentage of energy from free sugars was associated with 22% (95% confidence interval [CI], 1%-47%) greater dfs among children aged 2 to 8. A doubling in energy from added sugars was associated with 20% (95% CI, 1%-42%) greater dfs and 10% (95% CI, 2%-20%) greater DMFS in children aged 6 to 19 y. Beverages were the most important source of added sugars associated with increased caries. Other free sugars were not associated with dfs or DMFS. Associations between free sugars and caries were diminished among children with home water fluoride of 0.7 ppm or greater. CONCLUSIONS: Free sugars intake, especially in the form of added sugars and specifically in sweetened beverages, was associated with higher dental caries. Water fluoride exposures modify these associations, reducing caries risk in the primary dentition of children whose home water meets recommended fluoride levels. KNOWLEDGE TRANSFER STATEMENT: Intake of free sugars, especially in the form of added sugars and specifically in beverages, was associated with higher dental caries in US children in this study. Water fluoride exposure at CDC-recommended levels protected against caries, especially in the primary dentition. These findings suggest that household water fluoridation at CDC-recommended levels protects against the cariogenic potential of free and added sugars during childhood.

Intracellular Lipid Accumulation and Mitochondrial Dysfunction Accompanies Endoplasmic Reticulum Stress Caused by Loss of the Co-chaperone DNAJC3.

Recessive mutations in DNAJC3, an endoplasmic reticulum (ER)-resident BiP co-chaperone, have been identified in patients with multisystemic neurodegeneration and diabetes mellitus. To further unravel these pathomechanisms, we employed a non-biased proteomic approach and identified dysregulation of several key cellular pathways, suggesting a pathophysiological interplay of perturbed lipid metabolism, mitochondrial bioenergetics, ER-Golgi function, and amyloid-beta processing. Further functional investigations in fibroblasts of patients with DNAJC3 mutations detected cellular accumulation of lipids and an increased sensitivity to cholesterol stress, which led to activation of the unfolded protein response (UPR), alterations of the ER-Golgi machinery, and a defect of amyloid precursor protein. In line with the results of previous studies, we describe here alterations in mitochondrial morphology and function, as a major contributor to the DNAJC3 pathophysiology. Hence, we propose that the loss of DNAJC3 affects lipid/cholesterol homeostasis, leading to UPR activation, ß-amyloid accumulation, and impairment of mitochondrial oxidative phosphorylation.

Supply of care by dental therapists and emergency dental consultations in Alaska native communities in the Yukon-Kuskokwim delta: a mixed methods evaluation.

OBJECTIVES: Examine the relationship between supply of care provided by dental therapists and emergency dental consultations in Alaska Native communities. METHODS: Explanatory sequential mixed-methods study using Alaska Medicaid and electronic health record (EHR) data from the Yukon-Kuskokwim Health Corporation (YKHC), and interview data from six Alaska Native communities. From the Medicaid data, we estimated community-level dental therapy treatment days and from the EHR data we identified emergency dental consultations. We calculated Spearman partial correlation coefficients and ran confounder-adjusted models for children and adults. Interview data collected from YKHC providers (N=16) and community members (N=125) were content analysed. The quantitative and qualitative data were integrated through connecting. Results were visualized with a joint display. RESULTS: There were significant negative correlations between dental therapy treatment days and emergency dental consultations for children (partial rank correlation = -0.48; p⟨0.001) and for adults (partial rank correlation = -0.18; p=0.03). Six pediatric themes emerged: child-focused health priorities; school-based dental programs; oral health education and preventive behaviors; dental care availability; healthier teeth; and satisfaction with care. There were four adult themes: satisfaction with care; adults as a lower priority; difficulties getting appointments; and limited scope of practice of dental therapy. CONCLUSIONS: Alaska Native children, and to a lesser extent adults, in communities served more intensively by dental therapists have benefitted. There are high levels of unmet dental need as evidenced by high emergency dental consultation rates. Future research should identify ways to address unmet dental needs, especially for adults.

A multi-omics analysis reveals the unfolded protein response regulon and stress-induced resistance to folate-based antimetabolites.

Stress response pathways are critical for cellular homeostasis, promoting survival through adaptive changes in gene expression and metabolism. They play key roles in numerous diseases and are implicated in cancer progression and chemoresistance. However, the underlying mechanisms are only poorly understood. We have employed a multi-omics approach to monitor changes to gene expression after induction of a stress response pathway, the unfolded protein response (UPR), probing in parallel the transcriptome, the proteome, and changes to translation. Stringent filtering reveals the induction of 267 genes, many of which have not previously been implicated in stress response pathways. We experimentally demonstrate that UPR-mediated translational control induces the expression of enzymes involved in a pathway that diverts intermediate metabolites from glycolysis to fuel mitochondrial one-carbon metabolism. Concomitantly, the cells become resistant to the folate-based antimetabolites Methotrexate and Pemetrexed, establishing a direct link between UPR-driven changes to gene expression and resistance to pharmacological treatment.

PERK-mediated expression of peptidylglycine α-amidating monooxygenase supports angiogenesis in glioblastoma.

PKR-like kinase (PERK) plays a significant role in inducing angiogenesis in various cancer types including glioblastoma. By proteomics analysis of the conditioned medium from a glioblastoma cell line treated with a PERK inhibitor, we showed that peptidylglycine α-amidating monooxygenase (PAM) expression is regulated by PERK under hypoxic conditions. Moreover, PERK activation via CCT020312 (a PERK selective activator) increased the cleavage and thus the generation of PAM cleaved cytosolic domain (PAM sfCD) that acts as a signaling molecule from the cytoplasm to the nuclei. PERK was also found to interact with PAM, suggesting a possible involvement in the generation of PAM sfCD. Knockdown of PERK or PAM reduced the formation of tubes by HUVECs in vitro. Furthermore, in vivo data highlighted the importance of PAM in the growth of glioblastoma with reduction of PAM expression in engrafted tumor significantly increasing the survival in mice. In summary, our data revealed PAM as a potential target for antiangiogenic therapy in glioblastoma.

A sensitive and simple targeted proteomics approach to quantify transcription factor and membrane proteins of the unfolded protein response pathway in glioblastoma cells.

Many cellular events are driven by changes in protein expression, measurable by mass spectrometry or antibody-based assays. However, using conventional technology, the analysis of transcription factor or membrane receptor expression is often limited by an insufficient sensitivity and specificity. To overcome this limitation, we have developed a high-resolution targeted proteomics strategy, which allows quantification down to the lower attomol range in a straightforward way without any prior enrichment or fractionation approaches. The method applies isotope-labeled peptide standards for quantification of the protein of interest. As proof of principle, we applied the improved workflow to proteins of the unfolded protein response (UPR), a signaling pathway of great clinical importance, and could for the first time detect and quantify all major UPR receptors, transducers and effectors that are not readily detectable via antibody-based-, SRM- or conventional PRM assays. As transcription and translation is central to the regulation of UPR, quantification and determination of protein copy numbers in the cell is important for our understanding of the signaling process as well as how pharmacologic modulation of these pathways impacts on the signaling. These questions can be answered using our newly established workflow as exemplified in an experiment using UPR perturbation in a glioblastoma cell lines.

Recruiting Adolescents from Medicaid Enrollment Files into a Neighborhood Oral Health Study.

INTRODUCTION: Strategies are needed to improve recruitment of low-income adolescents into oral health studies. OBJECTIVES: In this study, we assessed the feasibility of recruiting Medicaid-enrolled adolescents into a neighborhood-level oral health study using Medicaid enrollment files and to evaluate the degree of bias in the final recruited study population. METHODS: We obtained Medicaid enrollment files from the Oregon Health Authority for 15,440 Medicaid enrollees aged 12 to 17 y from Multnomah, Hood River, and Tillamook counties. We attempted to contact the primary caregiver of each adolescent by telephone, and we tracked contact, recruitment, enrollment, and study completion rates. We further assessed if these rates were different across county-level rurality, neighborhood-level income, and caregiver-level language preference (Spanish vs. English). The Pearson chi-square test was used to compare rates (α = 0.05). We contacted 6,202 caregivers (40.2%), recruited 738 adolescents (11.9%), enrolled 335 (45.4%), and had complete data for 284 (84.8%). The overall enrollment yield from contacted caregivers was 5.4%. Contact rates did not differ significantly by rurality (P = 0.897), but they were significantly lower in the lowest-income neighborhoods (P = 0.023). Recruitment rates were significantly higher for adolescents from rural counties (P = 0.001), but they did not differ by income or language preference. Enrollment rates were significantly higher among adolescents from rural counties (P < 0.001) and were significantly associated with income (P = 0.041), but they were not different by language preference (P = 0.083). Among participants with complete data, there were no differences by rurality or income, but a significantly larger proportion of adolescents with complete data had caregivers with a language preference for Spanish (P = 0.043). RESULTS AND CONCLUSIONS: It is feasible to recruit Medicaid-enrolled adolescents into a neighborhood oral health study through the use of Medicaid files. County-, neighborhood-, and caregiver-level factors may influence characteristics of the final study population. Additional research is needed to improve recruitment of Medicaid enrollees into neighborhood oral health studies. KNOWLEDGE TRANSFER STATEMENT: Researchers can use the results of this study to plan neighborhood-level oral health studies involving recruitment of low-income adolescents. Findings further underscore the importance of assessing factors related to recruitment to evaluate participant bias and the generalizability of study findings.

Surveying Dentists' Perceptions of Caregiver Refusal of Topical Fluoride.

The objective of this study was to assess dentists' perceptions of caregiver topical fluoride refusal behaviors. We administered an 8-item survey in 2015 and 2016 ( N = 582) and asked dentists about the extent to which fluoride refusal is a problem, refusal trends, comfort talking to caregivers who refuse, and perceived reasons why caregivers refuse. To examine geographic variation, we ran χ2 tests between dentists' location (US West vs. non-West) and the first 3 perception measures (α = 0.05). Nearly 80% of dentists believed fluoride refusal was a problem, and 42.3% believed it was a growing problem. A significantly larger proportion of dentists who saw fluoride refusal as a problem also believed refusal was a growing problem compared to those who thought refusal was not a problem (89.6% and 41.2%, respectively; P < 0.0001). Caregiver characteristics perceived to be associated with fluoride refusal included immunization refusal (41.3%), White race (37.6%), and high income (33.7%). Thirty-seven percent of surveyed dentists were uncomfortable talking to caregivers who refused. There were no geographic differences in perceptions of fluoride refusal as a problem ( P = 0.52). A significantly larger proportion of non-West dentists believed fluoride refusal has gotten worse (non-West: 65.5%, West: 41.2%; P < 0.0001), but more dentists from the West were uncomfortable talking to caregivers who refused (West: 86%, non-West: 67.4%; P < 0.0001). Caregiver refusal of topical fluoride may be a growing problem, and many dentists are uncomfortable talking to caregivers who refuse. Additional interdisciplinary research is needed to identify the reasons why caregivers refuse fluoride, which is an important next step in developing chairside interventions that address fluoride refusal behaviors. Knowledge Transfer Statement: The results of this study can be used by researchers to develop chairside strategies to help dentists identify and manage fluoride refusal behaviors in clinical settings. This could help preserve topical fluoride as an evidence-based preventive therapy and address a growing public health problem.

The Effects of Medical Well Baby Visits in Promoting Earlier First Dental Visits for Children.

The objective of the study was to assess the effects of medical well baby visits in promoting earlier first dental visits. We analyzed Iowa Medicaid claims data (2000-2013). The sample included 4 cohorts of children born in 2000, 2003, 2007, or 2010 and enrolled in Medicaid from birth (N = 38,211). Children were followed for 3 y. The independent variables were cohort year and medical well baby visit frequency during 3 time periods (birth to age 10 mo, ages 11-19 mo, ages 20-36 mo). We used survival analyses to estimate first dental visit rates. First dental visit rates improved significantly from 2000 to 2013, with children in latter cohorts having significantly earlier first dental visits. Children with more medical well baby visits before age 11 mo had significantly delayed first dental visit rates than children with fewer medical well baby visits. The opposite was observed for children with more medical well baby visits between ages 11 to 19 mo and ages 20 to 36 mo. First dental visit rates for Medicaid-enrolled children have improved, but there continues to be a need for early interventions to improve age 1 dental visits and other preventive oral health behaviors. Knowledge Transfer Statement: The results of this study can be used by policy makers when developing strategies to improve access to dental care for young children in Medicaid. With consideration to promoting earlier preventive dental visits for publicly insured children, this study could lead to early interventions and improved health outcomes.

Improving children's oral health: an interdisciplinary research framework.

Despite the concerted efforts of research and professional and advocacy stakeholders, recent evidence suggests that improvements in the oral health of young children in the United States has not followed the prevailing trend of oral health improvement in other age groups. In fact, oral health disparities in the youngest children may be widening, yet efforts to translate advances in science and technology into meaningful improvements in populations' health have had limited success. Nevertheless, the great strides in genomics, biological, behavioral, social, and health services research in the past decade have strengthened the evidence base available to support initiatives and translational efforts. Concerted actions to accelerate this translation and implementation process are warranted; at the same time, policies that can help tackle the upstream determinants of oral health disparities are imperative. This article summarizes the proceedings from the symposium on the interdisciplinary continuum of pediatric oral health that was held during the 43rd annual meeting of the American Association for Dental Research, Charlotte, North Carolina, USA. This report showcases the latest contributions across the interdisciplinary continuum of pediatric oral health research and provides insights into future research priorities and necessary intersectoral synergies. Issues are discussed as related to the overwhelming dominance of social determinants on oral disease and the difficulty of translating science into action.

Economic modeling of sealing primary molars using a "value of information" approach.

The objective was to evaluate 2 primary molar sealant strategies for publicly insured children using an "expected value of perfect information" (EVPI) approach. We converted a 10,000-observation tooth-level cost-effectiveness simulation model comparing 2 primary molar sealant strategies - always seal (AS) and standard care (SC) - with a 1,250-observation child-level model. Costs per child per restoration or extraction averted were estimated. Opportunity losses under the AS strategy were determined for children for whom SC was the optimal choice. We determined the EVPI by multiplying mean opportunity losses by the projected incident population of publicly insured 3-year-olds in the US over 10 years with costs discounted at 2%. All analyses were conducted under assumptions of high and low intrachild correlations between at-risk teeth. The AS strategy cost $43.68 over SC (95% CI: -$5.50, $92.86) per child per restoration or extraction averted under the high intrachild correlation assumption and $15.54 (95% CI $7.86, $23.20) under the low intrachild correlation. Under high intrachild correlation, mean opportunity losses were $80.28 (95% CI: $76.39, $84.17) per child, and AS was the optimal strategy in 31% of children. Under low correlation, mean opportunity losses were $14.61 (95% CI: $12.20, $17.68) and AS was the optimal strategy in 87% of children. The EVPI was calculated at $530,813,740 and $96,578,389 (for high and low intrachild correlation, respectively), for a projected total incident population of 8,059,712 children. On average, always sealing primary molars is more effective than standard care, but widespread implementation of this preventive approach among publicly insured children would result in large opportunity losses. Additional research is needed to identify the subgroups of publicly insured children who would benefit the most from this effective and potentially cost-saving public health intervention.

A serial cross-sectional study of pediatric inpatient hospitalizations for non-traumatic dental conditions.

Investigators have examined children's dental utilization in various settings (e.g., dental offices, emergency departments, operating rooms), but no studies have examined inpatient hospitalizations for non-traumatic dental conditions (NTDCs). The authors examined NTDC-related hospitalization trends in the United States and identified the relationship between complex chronic condition (CCCs) and NTDC-related inpatient hospitalizations. We analyzed data from the U.S. Nationwide Inpatient Sample (2000-2010) for children ages 3 to 17 years (N = 3,030,970). The predictor variable was number of CCCs (0/1/2+). The outcome variable was whether the child had a NTDC-related hospitalization (no/yes). Covariate-adjusted multivariable logistic regression models were used to estimate prevalence odds ratios (PORs). From 2000 to 2010, there were 17,993 NTDC-related hospitalizations (0.59%) and a slight increase in NTDC-related hospitalizations (p = .049). This increase was not significant in the final regression model. There was no difference in odds of NTDCs for children with 0 or 1 CCCs (POR = 1.08; 95%CI = 0.99, 1.18), but children with 2+ CCCs had significantly greater odds (POR = 1.61; 95%CI = 1.42, 1.83), as did non-White, publicly insured, and lower income children. NTDC-related hospitalizations for children did not increase from 2000 to 2010. Children with 2+ CCCs had the greatest odds of being hospitalized for NTDCs, which highlights the need to develop preventive interventions targeting children with 2+ CCCs.

Liposomal sphingomyelin influences the cellular lipid profile of human lymphoblastic leukemia cells without effect on P-glycoprotein activity.

Sphingomyelin (SM)/cholesterol liposomes are currently investigated as drug carriers in cancer therapy. However, no data is available on the influence of SM itself on P-glycoprotein (P-gp) mediated multidrug resistance. P-gp is at least partly located in sphingolipid-enriched lipid raft domains of the plasma membrane, and its activity depends on the lipid profile of the membrane, which could be altered by therapeutical SM liposomes. Therefore, the aim of this study was to analyze the effect of liposomal SM on P-gp activity, P-gp distribution in microdomains, SM content of the membrane domains, and sensitivity of human lymphoblastic CEM cells toward cytotoxic drugs in vitro. Assays were conducted in CEM and multidrug resistant CEM/ADR5000 cells. SM-only liposomes were prepared by a newly developed ethanol injection protocol and thoroughly characterized. Inclusion of SM into the membrane was analyzed by fluorescence microscopy and flow cytometry. Influence of SM liposomes on P-gp activity was assessed by rhodamine efflux and calcein assay, and sensitivity toward cytotoxic drugs was analyzed by flow cytometric 7-AAD staining. Influence on P-gp distribution was analyzed by Western blot after density gradient centrifugation. SM 16:0, 18:0, and 24:1 were quantified by liquid chromatography coupled to tandem mass spectrometry. P-gp was mainly located in nonraft fractions, which did not change upon liposome treatment. Liposomes increased SM 16:0 and SM 24:1 content in nonraft domains, but not in raft domains of multidrug resistant cells. SM-only liposomes did not influence P-gp activity and chemosensitivity. In conclusion, SM-only liposomes in therapeutic amounts did not influence P-gp mediated multidrug resistance in CEM cells.

A first look: determinants of dental care for children in foster care.

PURPOSE: This hypothesis-generating study sought to identify potential determinants of dental care use and oral health among children living in foster care. METHOD: Using a grounded theory approach, fourteen key informant interviews were conducted among health and social services professionals experienced with children in foster care and families in western Washington State. RESULTS: The identified potential determinants of oral health and dental use among children living in foster care included: (1) linguistic and cultural barriers; (2) lack of dentists willing to accept children's Medicaid dental insurance; (3) lack of resources available to case workers (i.e., large caseload burden) (4) lack of federal funding for specialized dental care; (5) lack of systematic health record-keeping; (6) child transience, leading to the lack of a dental home; (8) foster parents' competing needs; (7) child behavior problems; and (9) lack of dental "buy in" from adolescents. CONCLUSION: Additional studies are needed to determine whether children living in foster care achieve oral health, and the extent of their unmet dental need.

Clinical evidence for polyol efficacy.

Xylitol is a safe dental caries preventive when incorporated into chewing gum or confections used habitually. The goal of this paper is to identify and assess the work on xylitol and other polyols and dental caries since 2008. Xylitol is effective when used by the mother prenatally or after delivery to prevent mutans transmission and subsequent dental caries in the offspring. One new completed trial confirmed that children of mothers who used xylitol lozenges after delivery had less dental caries than a comparison group. A similar study confirmed that the use of xylitol gum by the mother either prevented or postponed MS transmission to the offspring. Xylitol use among schoolchildren delivered via a gummy bear confection reduced S. mutans levels, but a once per day use of xylitol-containing toothpaste did not. Randomized trials, with caries outcomes, assessing xylitol-containing lozenges in adults and xylitol-containing gummy bears in children will release results in the coming year. Other studies are ongoing but are not systematic and will fail to answer important questions about how xylitol, or other polyols, can address the global dental caries problem.