Geroscience: Aging and Oral Health Research.

Research in aging has significantly advanced; scientists are now able to identify interventions that slow the biologic aging processes (i.e., the "hallmarks of aging"), thus delaying the onset and progression of multiple diseases, including oral conditions. Presentations given during the 3-part session "Geroscience: Aging and Oral Health Research," held during the 2023 American Association for Dental, Oral, and Craniofacial Research meeting, are summarized in this publication. Speakers' topics spanned the translational research spectrum. Session 1 provided an overview of the geroscience and health span (disease-free and functional health throughout life) concepts. The common molecular mechanisms between oral cancer and aging were discussed, and research was presented that showed periodontal microflora as a potential factor in Alzheimer's disease progression. Session 2 focused on behavioral and social science aspects of aging and their oral health significance. The keynote provided evidence that loneliness and isolation can have major health effects. These social conditions, along with poor oral health, tooth loss, and cognitive decline, could potentially affect healthy eating ability and systemic health in older adults. Research could help elucidate the directions and pathways connecting these seemingly disparate conditions. Session 3 focused on the delivery of oral care in different settings and the many barriers to access care faced by older adults. Research is needed to identify and implement effective technology and strategies to improve access to dental care, including new delivery and financing mechanisms, workforce models, interprofessional provider education and practice, and use of big data from medical-dental integration of electronic health records. Research to improve the "oral health span," reduce oral health disparities, and increase health equity must be tackled at all levels from biologic pathways to social determinants of health and health policies.

Sex-based differential regulation of bacterial-induced bone resorption.

BACKGROUND AND OBJECTIVE: Periodontal disease pathogenesis is comprised of the complex inflammatory immune response to oral bacterial dysbiosis. Like other inflammatory diseases, there is sexual dimorphism evident in periodontal diseases. During periodontitis, inflammatory chemokines direct neutrophils to migrate to the site of infection to neutralize the pathogen. Interestingly, these same chemokines are also involved in regulating pathogen-induced osteoclast formation. Previous reports show differences in bone turnover and lymphocyte recruitment between sexes. We hypothesize that chemokine expression is differentially regulated by sex and thus results in differential osteoclast formation. MATERIAL AND METHODS: Male and female mice were utilized to isolate neutrophils based on expression of Ly6G-specific, as well as defined osteoclast progenitors. Cells were stimulated with lipopolysaccharide (LPS; 100 ng/mL) then analyzed for neutrophil infiltration and gene expression. Defined osteoclast progenitors were primed: macrophage-colony stimulating factor (25 ng/mL), receptor activator of NF-κB ligand (50 ng/mL), then stimulated with LPS. Osteoclasts were enumerated via TRAP stain and mRNA isolated for gene expression analysis via quantitative polymerase chain reaction. RESULTS: In response to LPS, male neutrophils in vitro respond with increased chemokine expression and significantly more osteoclast formed in response to LPS compared to females. CONCLUSIONS: Findings support observations in humans regarding a sexual dimorphism in oral bacterial infections of alveolar bone loss. Males have a strong inflammatory response to bacterial infection, resulting in increased inflammatory microenvironment, reduced pathogenic bacteria clearance and increased osteoclast-driven bone loss in response to differential expression of key chemokines.

CD36 is upregulated in mice with periodontitis and metabolic syndrome and involved in macrophage gene upregulation by palmitate.

BACKGROUND: We reported that high-fat diet (HFD)-induced metabolic syndrome (MetS) exacerbates lipopolysaccharide (LPS)-stimulated periodontitis and palmitate, the major saturated fatty acid in the HFD, amplified LPS-stimulated gene expression in vitro. As CD36 is a major receptor for fatty acids, we investigated periodontal CD36 expression in mice with periodontitis and MetS, and the role of CD36 in inflammatory gene expression in macrophages stimulated by palmitate. METHODS: MetS and periodontitis were induced in mice by HFD and periodontal injection of LPS, respectively. The periodontal CD36 expression and its relationship with alveolar bone loss were studied using immunohistochemistry, real-time PCR, and correlation analysis. The role of CD36 in upregulation of inflammatory mediators by LPS and palmitate in macrophages was assessed using pharmacological inhibitor and small interfering RNA. RESULTS: Periodontal CD36 expression was higher in mice with both MetS and periodontitis than that in mice with periodontitis or MetS alone and was correlated with osteoclastogenesis and alveolar bone loss. In vitro studies showed that CD36 expression in macrophages was upregulated by LPS and palmitate, and targeting CD36 attenuated palmitate-enhanced gene expression. CONCLUSION: CD36 expression is upregulated in mice with periodontitis and MetS and involved in gene expression in macrophages stimulated by palmitate and LPS.

Simvastatin inhibits lipopolysaccharide-induced osteoclastogenesis and reduces alveolar bone loss in experimental periodontal disease.

BACKGROUND AND OBJECTIVE: Statins are inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase and have anti-inflammatory effects independent of cholesterol lowering. Recent clinical studies have indicated that statin intake has a beneficial effect on periodontal disease. However, the underlying mechanisms have not been well understood. In the current study, we employed a rat model with lipopolysaccharide (LPS)-induced periodontal disease and determined the effect of simvastatin, a commonly prescribed statin, on osteoclastogenesis, gingival inflammation and alveolar bone loss. MATERIAL AND METHODS: Sprague-Dawley rats were injected with Aggregatibacter actinomycetemcomitans LPS in periodontal tissue three times per week for 8 wk and part of the rats with LPS injection were also given simvastatin via gavage. After the treatments, the rat maxillae were scanned by microcomputed tomography and the images were analyzed to determine alveolar bone loss. To explore the underlying mechanisms, the effect of simvastatin on osteoclastogenesis and gingival expression of proinflammatory cytokines were also determined by tartrate-resistant acid phosphatase staining and real-time polymerase chain reaction assays, respectively. RESULTS: Results showed that LPS treatment markedly increased bone loss, but administration of simvastatin significantly alleviated the bone loss. Results also showed that LPS treatment stimulated osteoclastogenesis and the expression of inflammatory cytokines, but simvastatin significantly modulates the stimulatory effect of LPS on osteoclastogenesis and cytokine expression. CONCLUSION: This study demonstrated that simvastatin treatment inhibits LPS-induced osteoclastogenesis and gingival inflammation and reduces alveolar bone loss, indicating that the intake of simvastatin may hinder the progression of periodontal disease.

Alveolar bone loss: mechanisms, potential therapeutic targets, and interventions.

This article reviews recent research into mechanisms underlying bone resorption and highlights avenues of investigation that may generate new therapies to combat alveolar bone loss in periodontitis. Several proteins, signaling pathways, stem cells, and dietary supplements are discussed as they relate to periodontal bone loss and regeneration. RGS12 is a crucial protein that mediates osteoclastogenesis and bone destruction, and a potential therapeutic target. RGS12 likely regulates osteoclast differentiation through regulating calcium influx to control the calcium oscillation-NFATc1 pathway. A working model for RGS10 and RGS12 in the regulation of Ca(2+) oscillations during osteoclast differentiation is proposed. Initiation of inflammation depends on host cell-microbe interactions, including the p38 mitogen-activated protein kinase (MAPK) signaling pathway. Oral p38 inhibitors reduced lipopolysaccharide (LPS)-induced bone destruction in a rat periodontitis model but showed unsatisfactory safety profiles. The p38 substrate MK2 is a more specific therapeutic target with potentially superior tolerability. Furthermore, MKP-1 shows anti-inflammatory activity, reducing inflammatory cytokine biosynthesis and bone resorption. Multipotent skeletal stem cell (SSC) populations exist within the bone marrow and periosteum of long bones. These bone-marrow-derived SSCs and periosteum-derived SSCs have shown therapeutic potential in several applications, including bone and periodontal regeneration. The existence of craniofacial bone-specific SSCs is suggested based on existing studies. The effects of calcium, vitamin D, and soy isoflavone supplementation on alveolar and skeletal bone loss in post-menopausal women were investigated. Supplementation resulted in stabilization of forearm bone mass density and a reduced rate of alveolar bone loss over 1 yr, compared with placebo. Periodontal attachment levels were also well-maintained and alveolar bone loss suppressed during 24 wk of supplementation.

Randomized Trial to Test a Chemo-Mechanical Antiplaque Regimen as Adjunct to Periodontal Therapy.

BACKGROUND: The control of dental biofilm regrowth after nonsurgical periodontal therapy is associated with better clinical outcomes. However, many patients have difficulty achieving optimal plaque control. Subjects with diabetes, in which immune and wound-healing responses are typically impaired, may benefit from intensive antiplaque control regimens after scaling and root planing (SRP). OBJECTIVES: This study aimed to evaluate the effects of an intensive, at-home, chemical, and mechanical antiplaque regimen as an adjunct to SRP for the treatment of moderate to severe periodontitis. A secondary objective was to compare responses in subjects with type 2 diabetes and nondiabetics. METHODS: This was a 6-mo, single-center, parallel-group, randomized trial. The test group received SRP and oral hygiene instructions, and subjects were instructed to use a 0.12% chlorhexidine gluconate mouthrinse twice a day for 3 mo and utilize rubber interproximal bristle cleaners twice a day for 6 mo. The control group received SRP and oral hygiene instructions. The main outcome was change in mean probing depth (PD) from baseline to 6 mo. Secondary outcomes included change in sites with deep PDs, mean clinical attachment level, bleeding on probing, plaque index, hemoglobin A1C, fasting blood glucose, C-reactive protein, and taste assessment. This study was registered at ClinicalTrials.gov as NCT04830969. RESULTS: In total, 114 subjects were randomized to either treatment. Eighty-six subjects completed the trial with no missing visits. Neither an intention-to-treat nor a per-protocol analysis showed statistically significant differences between treatment groups in mean PD at 6 mo. In a subgroup analysis, subjects with diabetes in the test group showed a statistically significant greater reduction in mean PD at 6 mo when compared to subjects with diabetes receiving the control treatment (Δ = 0.15, P = 0.04), while there were no differences within nondiabetics (Δ = 0.02, P = 0.75). CONCLUSION: Outcomes in subjects with diabetes may be improved by chemo-mechanical antiplaque measures after nonsurgical periodontal therapy. KNOWLEDGE TRANSFER STATEMENT: This study suggests diabetic subjects may benefit from an intensive, at-home, chemical, and mechanical antiplaque regimen to improve nonsurgical periodontal therapy outcomes.

Topical Vitamin D Prevents Bone Loss and Inflammation in a Mouse Model.

There is a strong association between vitamin D levels and periodontal disease based on numerous epidemiological studies. We have previously shown that experimental deficiency of serum vitamin D in mice leads to gingival inflammation and alveolar bone loss. Treatment of cultured oral epithelial cells with the active form of vitamin D, 1,25(OH)2 vitamin D3 (1,25(OH)2D3), inhibits the extracellular growth and intracellular invasion of bacteria associated with periodontal disease. Maintenance of periodontal health may be due in part to the anti-inflammatory activities of vitamin D. Furthermore, this hormone can induce the expression of an antimicrobial peptide in cultured oral epithelial cells. We have shown that oral epithelial cells are capable of converting inactive vitamin D to the active form, suggesting that topical treatment of the oral epithelium with inactive vitamin D could prevent the development of periodontitis. We subjected mice to ligature-induced periodontitis (LIP), followed by daily treatment with inactive vitamin D or 1,25(OH)2D3. Treatment with both forms led to a reduction in ligature-induced bone loss and inflammation. Gingival tissues obtained from vitamin D-treated LIP showed production of specialized proresolving mediators (SPM) of inflammation. To examine the mechanism, we demonstrated that apical treatment of 3-dimensional cultures of primary gingival epithelial cells with vitamin D prevented lipopolysaccharide-induced secretion of proinflammatory cytokines and led to a similar production of SPM. Analysis of the oral microbiome of the mice treated with vitamin D showed significant changes in resident bacteria, which reflects a shift toward health-associated species. Together, our results show that topical treatment of oral tissues with inactive vitamin D can lead to the maintenance of periodontal health through the regulation of a healthy microbiome and the stimulation of resolution of inflammation. This strongly supports the development of a safe and effective vitamin D-based topical treatment or preventive agent for periodontal inflammation and disease.

Novel Preosteoclast Populations in Obesity-Associated Periodontal Disease.

Although there is a clear relationship between the degree of obesity and periodontal disease incidence, the mechanisms that underpin the links between these conditions are not completely understood. Understanding that myeloid-derived suppressor cells (MDSCs) are expanded during obesity and operate in a context-defined manner, we addressed the potential role of MDSCs to contribute toward obesity-associated periodontal disease. Flow cytometry revealed that in the spleen of mice fed a high-fat diet (HFD), expansion in monocytic MDSCs (M-MDSCs) significantly increased when compared with mice fed a low-fat diet (LFD). In the osteoclast differentiation assay, M-MDSCs isolated from the bone marrow of HFD-fed mice showed a larger number and area of osteoclasts with a greater number of nuclei. In the M-MDSCs of HFD-fed mice, several osteoclast-related genes were significantly elevated when compared with LFD-fed mice according to a focused transcriptomic platform. In experimental periodontitis, the number and percentage of M-MDSCs were greater, with a significantly larger increase in HFD-fed mice versus LFD-fed mice. In the spleen, the percentage of M-MDSCs was significantly higher in HFD-fed periodontitis-induced (PI) mice than in LFD-PI mice. Alveolar bone volume fraction was significantly reduced in experimental periodontitis and was further decreased in HFD-PI mice as compared with LFD-PI mice. The inflammation score was significantly higher in HFD-PI mice versus LFD-PI mice, with a concomitant increase in TRAP staining for osteoclast number and area in HFD-PI mice over LFD-PI mice. These data support the concept that M-MDSC expansion during obesity to become osteoclasts during periodontitis is related to increased alveolar bone destruction, providing a more detailed mechanistic appreciation of the interconnection between obesity and periodontitis.

Anti-inflammatory effect of MAPK phosphatase-1 local gene transfer in inflammatory bone loss.

Alveolar bone loss associated with periodontal diseases is the result of osteoclastogenesis induced by bacterial pathogens. The mitogen-activated protein kinase (MAPK) phosphatase 1 (MKP-1) is a critical negative regulator of immune response as a key phosphatase capable of dephosphorylating activated MAPKs. In this study, rat macrophages transduced with recombinant adenovirus (Ad.)MKP-1 specifically dephosphorylated activated MAPKs induced by lipopolysaccharide (LPS) compared with control cells. Bone marrow macrophages from MKP-1 knockout (KO) mice exhibited higher interleukin (IL)-6, IL-10, tumor necrosis factor (TNF)-α, and select chemokine compared with wild-type (WT) mice when stimulated by LPS. In addition, bone marrow cultures from MKP-1 KO mice exhibited significantly more osteoclastogenesis induced by LPS than when compared with WT mice. Importantly, MKP-1 gene transfer in bone marrow cells of MKP-1 KO mice significantly decreased IL-6, IL-10, TNF-α and chemokine levels, and formed fewer osteoclasts induced by LPS than compared with control group of cells. Furthermore, MKP-1 gene transfer in an experimental periodontal disease model attenuated bone resorption induced by LPS. Histological analysis confirmed that periodontal tissues transduced with Ad. MKP-1 exhibited less infiltrated inflammatory cells, less osteoclasts and less IL-6 than compared with rats of control groups. These studies indicate that MKP-1 is a key therapeutic target to control of inflammation-induced bone loss.

Potent anti-inflammatory effects of systemically administered curcumin modulate periodontal disease in vivo.

BACKGROUND AND OBJECTIVE: Curcumin is a plant-derived dietary spice with various biological activities, including anticarcinogenic and anti-inflammatory effects. Its therapeutic applications have been studied in a variety of conditions, including rheumatoid arthritis, colon cancer and depression, but no studies have evaluated the effects of curcumin on periodontal disease in vivo. MATERIAL AND METHODS: Experimental periodontal disease was induced in rats by placing cotton ligatures around both lower first molars. Curcumin was given to the rats by the intragastric route daily at two dosages (30 and 100 mg/kg) for 15 d. Control animals received ligatures but only the corn oil vehicle by gavage, and no treatment-negative control animals were included. Bone resorption was assessed by micro-computed tomography, and the inflammatory status was evaluated by stereometric analysis. Both RT-qPCR and ELISA were used to determine the expression of interleukin-6, tumor necrosis factor-α and prostaglandin E(2) synthase in the gingival tissues. Modulation of p38 MAPK and nuclear factor-κB activation were assessed by western blotting. RESULTS: Bone resorption was effectively induced in the experimental period, but it was not affected by either dose of curcumin. Curcumin effectively inhibited cytokine gene expression at both the mRNA and the protein level and produced a dose-dependent inhibition of the activation of nuclear factor-κB in the gingival tissues. Activation of p38 MAPK was not inhibited by curcumin. Curcumin-treated animals also presented a marked reduction of the inflammatory cell infiltrate and increased collagen content and fibroblastic cell numbers. CONCLUSION: Curcumin did not prevent alveolar bone resorption, but its potent anti-inflammatory effect suggests that it may have a therapeutic potential in periodontal diseases.

A dominant function of p38 mitogen-activated protein kinase signaling in receptor activator of nuclear factor-kappaB ligand expression and osteoclastogenesis induction by Aggregatibacter actinomycetemcomitans and Escherichia coli lipopolysaccharide.

BACKGROUND AND OBJECTIVE: Lipopolysaccharide from gram-negative bacteria is one of the microbial-associated molecular patterns that initiate the immune/inflammatory response, leading to the tissue destruction observed in periodontitis. The aim of this study was to evaluate the role of the p38 mitogen-activated protein kinase (MAPK) signaling pathway in lipopolysaccharide-induced receptor activator of nuclear factor-kappaB ligand (RANKL) expression by murine periodontal ligament cells. MATERIAL AND METHODS: Expression of RANKL and osteoprotegerin mRNA was studied by reverse transcription-polymerase chain reaction upon stimulation with lipopolysaccharide from Escherichia coli and Aggregatibacter actinomycetemcomitans. The biochemical inhibitor SB203580 was used to evaluate the contribution of the p38 MAPK signaling pathway to lipopolysaccharide-induced RANKL and osteoprotegerin expression. Stable cell lines expressing dominant-negative forms of MAPK kinase (MKK)-3 and MKK6 were generated to confirm the role of the p38 MAPK pathway. An osteoclastogenesis assay using a coculture model of the murine monocytic cell line RAW 264.7 was used to determine if osteoclast differentiation induced by lipopolysaccharide-stimulated periodontal ligament was correlated with RANKL expression. RESULTS: Inhibiting p38 MAPK prior to lipopolysaccharide stimulation resulted in a significant decrease of RANKL mRNA expression. Osteoprotegerin mRNA expression was not affected by lipopolysaccharide or p38 MAPK. Lipopolysaccharide-stimulated periodontal ligament cells increased osteoclast differentiation, an effect that was completely blocked by osteoprotegerin and significantly decreased by inhibition of MKK3 and MKK6, upstream activators of p38 MAPK. Conditioned medium from murine periodontal ligament cultures did not increase osteoclast differentiation, indicating that periodontal ligament cells produced membrane-bound RANKL. CONCLUSION: Lipopolysaccharide resulted in a significant increase of RANKL in periodontal ligament cells. The p38 MAPK pathway is required for lipopolysaccharide-induced membrane-bound RANKL expression in these cells.

Sexual Dimorphism in Immunity to Oral Bacterial Diseases: Intersection of Neutrophil and Osteoclast Pathobiology.

Sex is a biological variable that affects immune responses to bacterial and other types of infectious agents. Males and females are known to have differential oral bacterial disease burden in periodontal and endodontic disease. Understanding that there is a contribution from both sex and gender to these oral diseases, we discuss in this review recent sex-based findings that provide a pathobiological basis for differences observed between males and females. Sexual dimorphism of immune responses with respect to neutrophil trafficking and osteoclast differentiation and formation is presented as a plausible mechanism to explain the sexual differences. We also emphasize that sex, as a biological variable, should be considered in these types of oral immunologic studies.

Tristetraprolin Is Required for Alveolar Bone Homeostasis.

Tristetraprolin (TTP) is an RNA-binding protein that targets numerous immunomodulatory mRNA transcripts for degradation. Many TTP targets are key players in the pathogenesis of periodontal bone loss, including tumor necrosis factor-α. To better understand the extent that host immune factors play during periodontal bone loss, we assessed alveolar bone levels, inflammation and osteoclast activity in periodontal tissues, and immune response in draining cervical lymph nodes in TTP-deficient and wild-type (WT) mice in an aging study. WT and TTP-deficient (knockout [KO]) mice were used for all studies under specific pathogen-free conditions. Data were collected on mice aged 3, 6, and 9 mo. Microcomputed tomography (µCT) was performed on maxillae where 3-dimensional images were generated and bone loss was assessed. Decalcified sections of specimens were scored for inflammation and stained with tartrate-resistant acid phosphate (TRAP) to visualize osteoclasts. Immunophenotyping was performed on single-cell suspensions isolated from primary and peripheral lymphoid tissues using flow cytometry. Results presented indicate that TTP KO mice had significantly more alveolar bone loss over time compared with WT controls. Bone loss was associated with significant increases in inflammatory cell infiltration and an increased percentage of alveolar bone surfaces apposed with TRAP+ cells. Furthermore, it was found that the draining cervical lymph nodes were significantly enlarged in TTP-deficient animals and contained a distinct pathological immune profile compared with WT controls. Finally, the oral microbiome in the TTP KO mice was significantly different with age from WT cohoused mice. The severe bone loss, inflammation, and increased osteoclast activity observed in these mice support the concept that TTP plays a critical role in the maintenance of alveolar bone homeostasis in the presence of oral commensal flora. This study suggests that TTP is required to inhibit excessive inflammatory host responses that contribute to periodontal bone loss, even in the absence of specific periodontal pathogens.

p38 MAPK signaling in oral-related diseases.

Multiple dental diseases are characterized by chronic inflammation, due to the production of cytokines, chemokines, and prostanoids by immune and non-immune cells. Membrane-bound receptors provide a link between the extracellular environment and the initiation of intracellular signaling events that activate common signaling components, including p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and nuclear factor (NF)-kappaB. Although ERK pathways regulate cell survival and are responsive to extracellular mitogens, p38 MAPK, JNK, and NF-kappaB are involved in environmental stress responses, including inflammatory stimuli. Over the past decade, significant advances have been made relative to our understanding of the fundamental intracellular signaling mechanisms that govern inflammatory cytokine expression. The p38 MAPK pathway has been shown to play a pivotal role in inflammatory cytokine and chemokine gene regulation at both the transcriptional and the post-transcriptional levels. In this review, we present evidence for the significance of p38 MAPK signaling in diverse dental diseases, including chronic pain, desquamative disorders, and periodontal diseases. Additional information is presented on the molecular mechanisms whereby p38 signaling controls post-transcriptional gene expression in inflammatory states.

Autoimmunity to deltaNp63alpha in chronic ulcerative stomatitis.

Chronic ulcerative stomatitis (CUS) is a recently described mucocutaneous condition in which patients experience chronic, painful, ulcerative lesions of the oral mucosa. CUS is diagnosed by immunofluorescence studies that demonstrate antinuclear antibodies. These autoantibodies are specific for a protein, deltaNp63alpha, which is normally expressed in basal cell nuclei of stratified squamous epithelia. The purpose of this study was to characterize the autoimmune response in CUS. Protein antigens were produced by in vitro transcription/translation of polymerase chain-reaction (PCR)-amplified cDNAs. We used immunoblotting and immunoprecipitation experiments with serum from CUS patients to examine the (1) antibody isotype, (2) immunogenic functional domains of the deltaNp63alpha antigen, and (3) cross-reactivity with homologous p53, p73, and p63 proteins. Results demonstrate CUS patient antibodies to deltaNp63alpha, and 52% of cases have circulating IgA isotype antibodies. The N-terminal and DNA-binding domains are the immunodominant regions, and antibody cross-reactivity with p53, p63, and p73 isoforms is limited.

Recent Trends in Oral Cavity Cancer Research Support in the United States.

The objectives were to characterize oral cavity cancer (OCC) funding from the National Institutes of Health (NIH) with a secondary aim of comparing NIH support provided to OCC and other malignancies. NIH awards supporting OCC inquiry from 2000 to 2014 were accessed from the NIH RePORTER database. These data were used to evaluate temporal trends and the role of human papilloma virus and to determine the academic training and professional profiles of the principal investigators. Comparison of 2014 funding levels with other malignancies was also performed, controlling for incidence. Overall funding totals decreased considerably after 2009. Funding administered through the National Institute of Dental and Craniofacial Research (NIDCR) was 6.5 times greater than dollars awarded by the National Cancer Institute in 2000. During the period evaluated, NIDCR support decreased in most years, while National Cancer Institute support increased and approached NIDCR funding levels. Funding for human papilloma virus-related projects gradually rose, from 3.4% of dollars in 2000 to 2004 to 6.2% from 2010 to 2014 ( P < 0.05). A majority of principal investigators had a PhD omnia solus (57%), and 13% possessed dual PhD/clinical degrees. Among clinicians with specialty training, otolaryngologists and oral/maxillofacial pathologists garnered the most funding. OCC had a 2014 funding:incidence ratio of $785, much lower than for other malignancies. There has been increased volatility in funding support in recent years possibly due to budget cuts and sequestration. The National Cancer Institute has played an increasingly important role in supporting OCC research, concomitant with decreasing NIDCR support. Our findings suggest that OCC is underfunded relative to other non-oral cavity malignancies, indicating a need to increase the focus on rectifying the disparity.

Inhibition of human breast cancer cell proliferation in tissue culture by the neuroleptic agents pimozide and thioridazine.

Permanent cell culture lines derived from human breast cancer tissue are important experimental models in the study of human breast cancer cell proliferation. In the present work, pimozide, thioridazine, W-13, and W-12 were shown to inhibit MCF-7 human breast cancer cell growth. The 50% inhibition concentration values determined in two proliferation assays, [3H]thymidine incorporation and cell number, were in close agreement for each compound tested. The order of potency for growth inhibition in the presence of 2% stripped calf serum was pimozide (Ki 2 microM) greater than thioridazine (Ki 5 microM) greater than W-13 (Ki 15 microM) greater than W-12 (Ki 39 microM). Similar concentrations of these compounds blocked estradiol-induced growth of MCF-7 cells, but estrogen receptor (ER) interactions do not seem to be involved. Pimozide and thioridazine had no effect on the estradiol binding properties of the MCF-7 ER, nor did pimozide interfere with the induction of progesterone receptors by estradiol. Furthermore, pimozide also inhibited incorporation of [3H]thymidine into MCF-7 cells stimulated by polypeptide hormones in serum-free medium. The Ki for pimozide in serum-free medium alone, 0.46 microM, was similar to that determined in the presence of insulin (0.42 microM), insulin-like growth factor I (0.54 microM), and epidermal growth factor (0.43 microM). The effects of pimozide on breast cancer cell growth were not limited to the MCF-7 cell line. Pimozide also blocked cell growth and [3H]thymidine incorporation into the ER-positive T47D and ZR75-1B human breast cancer cell lines and the ER-negative human breast cancer cell line, MDA-MB-231. Although numerous mechanisms of action of pimozide and thioridazine have been identified, both drugs are calmodulin antagonists at drug concentrations that inhibit breast cancer cell growth in vitro. Inhibition of MCF-7 cell growth by the selective calmodulin antagonists W-13 and W-12 is consistent with a role for calmodulin antagonism in the broad growth-inhibitory properties of pimozide. We conclude that pimozide and thioridazine may be useful in the control of estradiol- and polypeptide hormone-induced growth of ER-positive and ER-negative human breast tumors.

Aggregatibacter actinomycetemcomitans, a potent immunoregulator of the periodontal host defense system and alveolar bone homeostasis.

Aggregatibacter actinomycetemcomitans is a perio-pathogenic bacteria that has long been associated with localized aggressive periodontitis. The mechanisms of its pathogenicity have been studied in humans and preclinical experimental models. Although different serotypes of A. actinomycetemcomitans have differential virulence factor expression, A. actinomycetemcomitans cytolethal distending toxin (CDT), leukotoxin, and lipopolysaccharide (LPS) have been most extensively studied in the context of modulating the host immune response. Following colonization and attachment in the oral cavity, A. actinomycetemcomitans employs CDT, leukotoxin, and LPS to evade host innate defense mechanisms and drive a pathophysiologic inflammatory response. This supra-physiologic immune response state perturbs normal periodontal tissue remodeling/turnover and ultimately has catabolic effects on periodontal tissue homeostasis. In this review, we have divided the host response into two systems: non-hematopoietic and hematopoietic. Non-hematopoietic barriers include epithelium and fibroblasts that initiate the innate immune host response. The hematopoietic system contains lymphoid and myeloid-derived cell lineages that are responsible for expanding the immune response and driving the pathophysiologic inflammatory state in the local periodontal microenvironment. Effector systems and signaling transduction pathways activated and utilized in response to A. actinomycetemcomitans will be discussed to further delineate immune cell mechanisms during A. actinomycetemcomitans infection. Finally, we will discuss the osteo-immunomodulatory effects induced by A. actinomycetemcomitans and dissect the catabolic disruption of balanced osteoclast-osteoblast-mediated bone remodeling, which subsequently leads to net alveolar bone loss.

Metabolic syndrome exacerbates inflammation and bone loss in periodontitis.

Clinical studies have shown that metabolic syndrome (MetS) is associated with increased risk of developing periodontitis. However, the underlying mechanisms remain largely unknown. Since it is known that lipopolysaccharide (LPS)-activated toll-like receptor 4 signaling pathways play a crucial role in periodontitis, we hypothesized that MetS enhances LPS-induced periodontal inflammation and alveolar bone loss. In this study, we induced MetS in C57BL/6 mice by feeding them high-fat diet (HFD), and we induced periodontitis by periodontal injection of Aggregatibacter actinomycetemcomitans LPS. We found that mice fed a HFD had significantly increased body weight, plasma lipids, insulin, and insulin resistance when compared with mice fed regular chow, indicating that the mice developed MetS. We also found that a HFD markedly increased LPS-induced alveolar bone loss, osteoclastogenesis, and inflammatory infiltration. Analysis of gene expression in periodontal tissue revealed that HFD and LPS injection cooperatively stimulated expression of cytokines that are known to be involved in periodontal tissue inflammation and osteoclastogenesis-such as interleukin 6, monocyte-chemotactic protein 1, receptor activator of nuclear factor kappa-B ligand, and macrophage colony-stimulating factor. To further understand the potential mechanisms involved in MetS-boosted tissue inflammation, our in vitro studies showed that palmitic acid-the most abundant saturated fatty acid (SFA) and the major SFA in the HFD used in our animal study-potently enhanced LPS-induced proinflammatory gene expression in macrophages. In sum, this study demonstrated that MetS was associated with increased periodontal inflammation and alveolar bone loss in an LPS-induced periodontitis animal model. This study also suggests that SFA palmitic acid may play an important role in MetS-associated periodontitis by enhancing LPS-induced expression of inflammatory cytokines in macrophages.

Inositol trisphosphate receptor gene expression and hormonal regulation in osteoblast-like cell lines and primary osteoblastic cell cultures.

The inositol trisphosphate receptor (IP3R) is an intracellular calcium channel that mediates the cellular actions of a wide variety of hormones, growth factors, and cytokines. In osteoblastic cell cultures, many bone resorbing hormones increase phosphoinositide turnover, inositol trisphosphate production, mobilization of intracellular calcium, and the secretion of osteoclast recruitment and activating factors. In this study, the effects of 17 beta-estradiol, 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), phrobol ester, and serum on IP3R mRNA levels were evaluated in osteogenic-osteosarcoma cells and in primary osteoblastic cultures derived from neonatal rat calvaria. Type-specific reverse transcription polymerase chain reaction (RT-PCR) indicated that all cell types evaluated (G-292, U-2 OS, Saos-2, MC3T3-E1, UMR-106, and calvarial osteoblastic cells) express IP3R mRNA type I; G-292, U-2 OS, MC3T3-E1, and calvarial osteoblastic cells also express type II IP3R mRNA; and UMR-106 and the calvarial osteoblastic cells express type III IP3R mRNA. Northern blot and RT-PCR analyses of human G-292 osteosarcoma cells and rat calvarial osteoblastic cells showed that phorbol ester and serum increase IP3R mRNA levels, whereas 17 beta-estradiol and 1,25(OH)2D3 decrease these levels. In G-292 cells, the effect of 17 beta-estradiol was not due to accelerated IP3R mRNA degradation and required continued protein synthesis. The results show that multiple IP3R types are expressed in osteoblasts and osteoblastic osteosarcoma cells and that this expression is regulated by 17 beta-estradiol and other osteoporotic and antiosteoporotic hormones. These findings indicate that hormonal control of IP3R expression may be relevant in the chronic regulation of osteoblast secretory activity.