Corneliu Sima

Type 1 diabetes predisposes to enhanced gingival leukocyte margination and macromolecule extravasation in vivo.

BACKGROUND AND OBJECTIVE Diabetes predisposes to periodontal disease. However, the cellular and molecular mechanisms linking the two conditions are not clear. The impact of chronic hyperglycemia on leukocyte margination and macromolecule extravasation was determined in gingival vessels in vivo. MATERIALS AND METHODS Gingival intravital microscopy was employed to measure extravasation of fluorescein isothiocyanate (FITC)-dextran in diabetic Akita and healthy wild-type (WT) mice. Rhodamine 6G and FITC-LY6G were injected for nonspecific and polymorphonuclear-specific leukocyte labeling, respectively. Surface expression of leukocyte adhesion molecules was determined with flow cytometry and western blotting. RESULTS Vascular permeability was significantly increased in Akita gingival vessels compared with WT [permeability index (PI): WT, 0.75 ± 0.05; Akita, 1.1 ± 0.03: p < 0.05). Wild-type gingival vessels reached comparable permeability 2 h after intragingival injection of tumor necrosis factor α (TNFα), used here as positive control (PI, 1.17 ± 0.16). The number of rolling leukocytes was significantly elevated in diabetic gingiva (WT, 25 ± 3.7 cells/min; Akita, 42 ± 8.5 cells/min; p < 0.03). Similar rolling cell counts were obtained in WT after intragingival injection of TNFα (10 ng TNFα, 47 ± 1.3 cells/min; 100 ng TNFα, 57.5 ± 5.85 cells/min). The number of leukocytes firmly attached to the endothelium was similar in WT and Akita mice. Leukocyte cell-surface expression of P-selectin glycoprotein ligand-1 and CD11a was increased in Akita mice, while L-selectin remained unchanged when compared with WT. Moreover, P-selectin expression in Akita gingival tissues was elevated compared with that of WT. CONCLUSION Chronic hyperglycemia induces a proinflammatory state in the gingival microcirculation characterized by increased vascular permeability, and leukocyte and endothelial cell activation. Leukocyte-induced microvascular damage, in turn, may contribute to periodontal tissue damage in diabetes.

Bone Replacement Materials and Techniques Used for Achieving Vertical Alveolar Bone Augmentation

Alveolar bone augmentation in vertical dimension remains the holy grail of periodontal tissue engineering. Successful dental implant placement for restoration of edentulous sites depends on the quality and quantity of alveolar bone available in all spatial dimensions. There are several surgical techniques used alone or in combination with natural or synthetic graft materials to achieve vertical alveolar bone augmentation. While continuously improving surgical techniques combined with the use of auto- or allografts provide the most predictable clinical outcomes, their success often depends on the status of recipient tissues. The morbidity associated with donor sites for auto-grafts makes these techniques less appealing to both patients and clinicians. New developments in material sciences offer a range of synthetic replacements for natural grafts to address the shortcoming of a second surgical site and relatively high resorption rates. This narrative review focuses on existing techniques, natural tissues and synthetic biomaterials commonly used to achieve vertical bone height gain in order to successfully restore edentulous ridges with implant-supported prostheses.

Macrophage subsets and osteoimmunology: tuning of the immunological recognition and effector systems that maintain alveolar bone

Chronic and aggressive periodontal diseases are characterized by the failure to resolve local inflammation against periodontopathogenic bacteria in the subgingival biofilm. Alveolar bone resorption is associated with altered innate and adaptive immune responses to periodontal pathogens. Macrophage-derived cytokines, chemokines and growth factors, present in both destructive and reparative phases of periodontitis, are elevated in numerous animal and human studies. Macrophage polarization to either a predominantly pro-inflammatory or anti-inflammatory phenotype may be a critical target for monitoring disease activity, modulating immune responses to subgingival biofilms in patients at risk and reducing alveolar bone loss.

Untreated type 1 diabetes increases sepsis-induced mortality without inducing a prelethal cytokine response.

Diabetes mellitus is the leading comorbidity in patients with sepsis, but its impact upon survival and immunoinflammatory signaling in sepsis is undetermined. We investigated the effect of untreated diabetes mellitus upon survival and immunoinflammatory responses in the acute phase (days 1-5) of murine polymicrobial sepsis using the AKITA model of type 1 diabetes. Diabetic female C57BL/6-Ins2Akita (AKITA) and C57BL/6 wild-type (WT) mice underwent cecal ligation and puncture (CLP), blood (20 &mgr;L) was sampled for 5 days, and survival was monitored for 28 days. By day 5, all 8 AKITA mice died compared with 10 of 28 deaths in WT mice. Blood glucose declined post-CLP in all groups (most dramatically in AKITAs by 75%). To compare the evolution of inflammatory profiles, mice were retrospectively divided based on outcome into AKITA, WT-Died, and WT-Survived (within days 1-5). Hypoglycemia developed in all groups, which resolved in WT-Survived (97 mg/dL at 96 h) but intensified in WT-Died and AKITAs (∼30 mg/dL). Dramatic increases in both proinflammatory and anti-inflammatory cytokines were observed in WT-Died (i.e., interleukin 6, 38.2 ± 17.8 ng/mL at 24 h), which contrasted with a lack of prelethal cytokine response in AKITA mice (interleukin 6, 4.3 ± 3.4 ng/mL at 24 h). A prelethal composite cytokine score was calculated on values obtained 24 h before death. This score was 3-fold lower for proinflammatory cytokines and 6-fold lower for anti-inflammatory mediators in the AKITA mice compared with the WT-Died mice but identical to the composite score in WT-Survived. These data demonstrate that untreated type I diabetes mellitus severely exacerbates sepsis mortality without inducing a prelethal release of systemic proinflammatory or anti-inflammatory cytokines.

Distinct Oral Neutrophil Subsets Define Health and Periodontal Disease States

Neutrophils exit the vasculature and swarm to sites of inflammation and infection. However, these cells are abundant in the healthy, inflammation-free human oral environment, suggesting a unique immune surveillance role within the periodontium. We hypothesize that neutrophils in the healthy oral cavity occur in an intermediary parainflammatory state that allows them to interact with and contain the oral microflora without eliciting a marked inflammatory response. Based on a high-throughput screen of neutrophil CD (cluster of differentiation) marker expression and a thorough literature review, we developed multicolor flow cytometry panels to determine the surface marker signatures of oral neutrophil subsets in periodontal health and disease. We define here 3 distinct neutrophil subsets: resting/naive circulatory neutrophils, parainflammatory neutrophils found in the healthy oral cavity, and proinflammatory neutrophils found in the oral cavity during chronic periodontal disease. Furthermore, parainflammatory neutrophils manifest as 2 distinct subpopulations—based on size, granularity, and expression of specific CD markers—and exhibit intermediate levels of activation as compared with the proinflammatory oral neutrophils. These intermediately activated parainflammatory populations occur in equal proportions in the healthy oral cavity, with a shift to one highly activated proinflammatory neutrophil population in chronic periodontal disease. This work is the first to identify and characterize oral parainflammatory neutrophils that interact with commensal biofilms without inducing an inflammatory response, thereby demonstrating that not all neutrophils trafficking through periodontal tissues are fully activated. In addition to establishing possible diagnostic and treatment monitoring biomarkers, this oral neutrophil phenotype model builds on existing literature suggesting that the healthy periodontium may be in a parainflammatory state.

Nucleic Acid-Targeting Pathways Promote Inflammation in Obesity-Related Insulin Resistance

Obesity-related inflammation of metabolic tissues, including visceral adipose tissue (VAT) and liver, are key factors in the development of insulin resistance (IR), though many of the contributing mechanisms remain unclear. We show that nucleic-acid-targeting pathways downstream of extracellular trap (ET) formation, unmethylated CpG DNA, or ribonucleic acids drive inflammation in IR. High-fat diet (HFD)-fed mice show increased release of ETs in VAT, decreased systemic clearance of ETs, and increased autoantibodies against conserved nuclear antigens. In HFD-fed mice, this excess of nucleic acids and related protein antigens worsens metabolic parameters through a number of mechanisms, including activation of VAT macrophages and expansion of plasmacytoid dendritic cells (pDCs) in the liver. Consistently, HFD-fed mice lacking critical responders of nucleic acid pathways, Toll-like receptors (TLR)7 and TLR9, show reduced metabolic inflammation and improved glucose homeostasis. Treatment of HFD-fed mice with inhibitors of ET formation or a TLR7/9 antagonist improves metabolic disease. These findings reveal a pathogenic role for nucleic acid targeting as a driver of metabolic inflammation in IR.

Neutrophil Dysfunction and Host Susceptibility to Periodontal Inflammation: Current State of Knowledge

Normal polymorphonuclear neutrophil (PMN) function is critical for the maintenance of host-biofilm equilibrium and periodontal tissue homeostasis. Mounting evidence suggests that PMNs play important roles in the control of commensal periodontal flora and initiation of resolution following inflammation caused by accumulating subgingival plaque. Quantitative and qualitative alterations of PMNs in bone marrow, blood, periodontal tissues, and gingival crevicular fluid contribute to host-microbial dysbiosis and onset of irreversible loss of clinical attachment around teeth. Recent findings of specific PMN phenotypes associated with different disease states bring us closer to understanding disease activity and addressing chronic, non-resolved, periodontal inflammation to better monitor and predict patient-specific treatment outcomes. The present review addresses the current state of knowledge in PMN biology in the pathogenesis of periodontal inflammation and the onset of periodontitis.

Nuclear Factor Erythroid 2-Related Factor 2 Down-Regulation in Oral Neutrophils Is Associated with Periodontal Oxidative Damage and Severe Chronic Periodontitis

The balance between reactive oxygen species and antioxidants plays an important role in periodontal health. We previously demonstrated that high reactive oxygen species production by oral polymorphonuclear neutrophils (oPMNs) in chronic periodontitis (CP) refractory to conventional therapy is associated with severe destruction of periodontium. Herein, we show that inhibition of antioxidant production through down-regulation of nuclear factor erythroid 2-related factor 2 (Nrf2) pathway in oPMN, despite enhanced recruitment in the oral cavity, is associated with severe CP. Twenty-four genes in the Nrf2-mediated oxidative stress response pathway were down-regulated in PMNs of diseased patients. Downstream of Nrf2, levels of oPMN superoxide dismutase 1 and catalase were decreased in severe CP, despite increased recruitment. Nrf2(-/-) mice had more severe loss of periodontium in response to periodontitis-inducing subgingival ligatures compared with wild-types. Levels of 8-hydroxy-deoxyguanosine were increased in periodontal lesions of Nrf2(-/-) mice, indicating high oxidative damage. We report, for the first time, Nrf2 pathway down-regulation in oPMNs of patients with severe CP. PMNs of CP patients may be primed for low antioxidant response in the context of high recruitment in the oral cavity, resulting in increased oxidative tissue damage.

Adseverin plays a role in osteoclast differentiation and periodontal disease-mediated bone loss

Osteoclast differentiation and function are highly dependent on the assembly and turnover of actin filaments, but little is known about the roles of actin binding proteins in these processes. Adseverin (Ads), a member of the gelsolin superfamily of actin capping and severing proteins, regulates actin filament turnover and can regulate the turnover of cortical actin filaments of chromaffin cells during exocytosis. Using a conditional Ads knockout mouse model, we confirmed our previous finding in cultured cells that Ads plays a role in osteoclastogenesis (OCG) and actin cytoskeletal organization in osteoclasts. Here we show that Ads is required for osteoclast formation and that when alveolar bone resorption is experimentally induced in mice, genetic deletion of Ads prevents osteoclast‐mediated bone loss. Further, when Ads‐null osteoclasts are cultured, they exhibit defective OCG, disorganized podosome‐based actin filament superstructures, and decreased bone resorption. Reintroduction of Ads into Ads‐null osteoclast precursor cells restored these osteoclast defects. Collectively, these data demonstrate a unique and osteoclast‐specific role for Ads in OCG and osteoclast function.— Jiang, H., Wang, Y., Viniegra, A., Sima, C., McCulloch, C. A., Glogauer, M. Adseverin plays a role in osteoclast differentiation and periodontal disease‐mediated bone loss. FASEB J. 29, 2281‐2291 (2015). www.fasebj.org

Identification of quantitative trait loci influencing inflammation‐mediated alveolar bone loss: insights into polygenic inheritance of host–biofilm disequilibria in periodontitis

BACKGROUND AND OBJECTIVE The relative contribution of genetic and environmental factors to the onset and progression of periodontitis is inconclusive. Despite the high prevalence, phenotypic heterogeneity and significant local and systemic implications of this disease, early detection and individualized therapy are problematic. Using a murine model of periodontitis in a panel of 17 recombinant inbred mice, the current study addressed the heritability of, and oral dysbiosis associated with, inflammation-mediated alveolar bone loss (iABL), the hallmark of periodontitis. MATERIAL AND METHODS Quantitative trait locus (QTL) genomics and quantitative PCR for over 99% of known murine oral microbiota were used. RESULTS It was found that iABL is a polygenic trait with 32.7% heritability. One suggestive QTL, nicknamed inflammation-mediated alveolar bone loss locus (iABLL), was identified on chromosome 2. Eleven genes involved in innate immune responses and bone metabolism, particularly related to macrophage and osteoblast function, namely Etl4, Pdss1, Cobll1, 9330158F14Rik, Xirp2, Stk39, Mettl5, Metapl1, Itga6, Pdk1 and Sp3, were found in the iABLL using cis expression QTL and nonsynonymous single nucleotide polymorphism analyses. Specific oral microbiome shifts in saliva and tongue mucosa are associated with disease in this model. CONCLUSION Our results indicate that complex host-biofilm interactions generate pathogenic states that extend beyond subgingival biofilms and periodontal tissues. Although no temporal relationship between the onset of iABL and microbiome changes were established, our findings suggest that host factors may be responsible for pathogenic shifts in subgingival biofilms when persistent and undisturbed.