Osteoarthritis of the Temporomandibular Joint: Clinical and Imagenological Diagnosis, Pathogenic Role of the Immuno- Inflammatory Response, and Immunotherapeutic Strategies Based on T Regulatory Lymphocytes

Osteoarthritis is a degenerative disease affecting the TMJ. It is the most common TMJ disorder and shows a higher prevalence in women and older people. TMJ osteoarthritis (TMJ-OA) is characterized by variable degrees of inflammation, destruction of the articular cartilage, and sub-chondral bone resorption. In this context, diverse pro-inflammatory cytokines, chemokines, enzymes, and bone-resorptive associated factors have been considered as possible markers of active TMJ-OA. The molecular balance is determinant not only for initiation and progression, but also for the clinical expression of the disease. Recent advances in the biochemical analysis of synovial fluid from affected patients have provided new insights into the patho-physiology of the TMJ-OA; however, its molecular pathogenesis still remains unclear. Recently, a Th1 and Th17-dominated immune response has been associated with the inflammatory and destructive events characteristic of TMJ-OA and, in particular, the Th17 lymphocyte pathway has a pivotal role in the increased production of RANKL, which is involved in osteoclast activation and subsequent sub-chondral bone resorption. Understanding the TMJ physiology and pathogenesis of the TMJ-OA, together with the key molecular determinants of the TMJ tissue destruction, will enable the development of new chair-side point of care diagnostics and more conservative treatment modalities with minimal complications

Determination of Cr(VI) and Cr(III) species in parenteral solutions using a nanostructured material packed-microcolumn and electrothermal atomic absorption spectrometry

A sequential on-line preconcentration and separation system for Cr(VI) and Cr(III) species determination was developed in this work. For this purpose, a microcolumn filled with nanostructured α-alumina was used for on-line retention of Cr species in a flow-injection system. The method involves the selective elution of Cr(VI) with concentrated ammonia and Cr(III) with 1 mol L-1 nitric acid for sequential injection into an electrothermal atomic absorption spectrometer (ETAAS). Analytical parameters including pH, eluent type, flow rates of sample and eluent, interfering effects, etc., were optimized. The preconcentration factors for Cr(VI) and Cr(III) were 41 and 18, respectively. The limit of detection (LOD) was 1.9 ng L-1 for Cr(VI) and 6.1 ng L-1 for Cr(III). The calibration graph was linear with a correlation coefficient of 0.999. The relative standard deviation (RSD) was 8.6% for Cr(VI) and 6.1% for Cr(III) (c=10 μg L-1, n=10, sample volume=25 mL). Verification of the accuracy was carried out by analysis of a standard reference material (NIST SRM 1643e "Trace elements in natural water") with a reported Cr content of 20.40±0.24 μg L-1. Using the proposed methodology the total Cr content, computed as sum of Cr(III) and Cr(VI), in this SRM was 20.26±0.96 μg L-1. The method was successfully applied to the determination of Cr(VI) and Cr(III) species in parenteral solutions. Concentration of Cr(III) species was found to be in the range of 0.29-3.62 μg L-1, while Cr(VI) species was not detected in the samples under study.Fil: Monasterio, Romina Paula. Universidad Nacional de La Pampa; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Regional de Investigaciones Cientifícas y Tecnológicas; ArgentinaFil: Lascalea, Gustavo Enrique. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza. Instituto Argentino de Nivología, Glaciología y Ciencias Ambientales. Provincia de Mendoza. Instituto Argentino de Nivología, Glaciología y Ciencias Ambientales. Universidad Nacional de Cuyo. Instituto Argentino de Nivología, Glaciología y Ciencias Ambientales; ArgentinaFil: Martinez, Luis Dante. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - San Luis. Instituto de Química de San Luis. Universidad Nacional de San Luis. Facultad de Química, Bioquímica y Farmacia. Instituto de Química de San Luis; ArgentinaFil: Wuilloud, Rodolfo German. Universidad Nacional de Cuyo. Facultad de Ciencias Exactas y Naturales; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza; Argentin

The therapeutic potential of regulatory T lymphocytes in periodontitis: A systematic review.

This systematic review aimed to: (a) generate a descriptive synthesis of preclinical studies assessing the therapeutic potential of regulatory T lymphocytes (Tregs) to arrest periodontitis, (b) evaluate the methodological heterogeneity of the reviewed animal studies and (c) assess the risk of bias (RoB) of the included studies. The electronic search for animal studies included the MEDLINE, EMBASE, Web of Science and LILACS databases. In addition, a manual search assessed the high-ranked scientific journals in "periodontics/immunology" and the references listed in the included studies. There were no language, year or publication status restrictions. Two independent reviewers selected and extracted the data, and Cohen's Kappa coefficient was calculated to determine the inter-examiner agreement. The Systematic Review Center for Laboratory Animal Experimentation's (SYRCLE) tool was used to assess the RoB. A total of 21 of the 425 studies obtained from the database search were included. Treg function was mainly described in Porphyromonas gingivalis-induced periodontitis (57.1%) in mice (76.2%), where Treg suppression was strongly related to disease progression and Treg induction was strongly related to immuno-inflammatory response reduction. Of those 21 studies, eight included eight animal experiments using three distinct therapeutic approaches, including: P. gingivalis-driven immunization (n = 3), retinoic acid inoculation (n = 2) and anti-inflammatory molecules in polymeric carriers (n = 3), which could modulate the Treg activity through cytokine production (interleukin-10 and transforming growth factor-β1), CC-chemokine- and CC-chemokine receptor-mediated chemoattraction (CCL22 and CCR4) or Th17-associated receptor activator of nuclear factor κB ligand (RANKL) downregulation. However, the studies with animal experiments did not specify the randomization sequences and housing conditions that were used, and therefore, 42.11% of the entries were rated as unclear RoB. Distinct therapeutic strategies involving Tregs could potentially suppress the immuno-inflammatory response and restore alveolar bone homeostasis during periodontitis. Nevertheless, important methodological variability, poor reporting of treatment effect estimates and unclear RoB suggest using caution when assessing the results of these studies

Regulatory T lymphocytes in periodontitis: A translational view

Periodontitis is a chronic immuno-inflammatory disease in which the disruption of the balance between host and microbiota interactions is key to the onset and progression of the disease. The immune homeostasis associated with periodontal health requires a regulated immuno-inflammatory response, during which the presence of regulatory T cells (Tregs) is essential to ensure a controlled response that minimizes collateral tissue damage. Since Tregs modulate both innate and adaptive immunity, pathological conditions that may resolve by the acquisition of immuno-tolerance, such as periodontitis, may benefit by the use of Treg immunotherapy. In recent years, many strategies have been proposed to take advantage of the immuno-suppressive capabilities of Tregs as immunotherapy, including the ex vivo and in vivo manipulation of the Treg compartment. Ongoing research in both basic and translational studies let us gain a better understanding of the diversity of Treg subsets, their phenotypic plasticity, and suppressive functions, which can be used as a substrate for new immunotherapies. Certainly, as our knowledge of Treg biology increases, we will be capable to develop new therapies designed to enhance the stability and function of Tregs during periodontitis.Comision Nacional de Investigacion Cientifica y Tecnologica (CONICYT) from the Chilean Government FONDECYT 1140904 FONDECYT 118178

Immunostimulatory activity of low-molecular-weight hyaluronan on dendritic cells stimulated with Aggregatibacter actinomycetemcomitans or Porphyromonas gingivalis

Objectives Periodontitis is a chronic inflammatory disease characterized by tooth-supporting tissue destruction, which is elicited by the host’s immune response triggered against periodonto-pathogen bacteria. During periodontal tissue destruction, extracellular matrix components are metabolized and fragmented. Some extracellular matrix component-derived fragments, such as lowmolecular-weight hyaluronan (LMW-HA), have potent immunogenic potential, playing a role as damage-associated molecular patterns (DAMPs) during activation of immune cells. Dendritic cells (DCs) play a central role in the host’s immune response displayed during periodontitis; thus, this study aimed to analyze whether LMW-HA has an immunostimulatory activity on DCs when stimulated with periodonto-pathogen bacteria. Materials and methods LMW-HA-treated and non-treated DCs were stimulated with Aggregatibacter actinomycetemcomitans or Porphyromonas gingivalis and the mRNA expression for cytokines tumor necrosis factor-α (TNF-alpha), interleukin-1β (IL1B), interleukin-6 (IL-6), and interleukin-23 (IL-23A) was quantified by RT-qPCR. In addition, transcription factors interferon regulatory factor 4 (IRF4), interferon regulatory factor 8 (IRF8), neurogenic locus notch homolog protein 2 (NOTCH2), and basic leucine zipper ATF-like transcription factor 3 (BATF3), involved in DC activation, were analyzed. Results Higher expression levels of TNF-alpha, IL-1B, IL-6, and IL-23A were detected in LMW-HA-treated DCs after bacterial infection, as compared with non-treated DCs. When LMW-HA-treated DCs were infected with A. actinomycetemcomitans, higher levels of IRF4, NOTCH2, and BATF3 were detected compared with non-treated cells; whereas against P. gingivalis infection, increased levels of IRF4 and NOTCH2 were detected. Conclusion LMW-HA plays an immunostimulatory role on the immune response triggered by DCs during infection with A. actinomycetemcomitans or P. gingivalis. Clinical relevance Detection of extracellular matrix component-derived fragments produced during periodontal tissue destruction, such as LMW-HA, could explain at least partly unsuccessful periodontal treatment and the chronicity of the disease

Serotype a of Aggregatibacter actinomycetemcomitans down-regulates the increased serotype b-induced cytokine and chemokine production in dendritic cells

Objective: In Aggregatibacter actinomycetemcomitans, different serotypes have been described based on LPS antigenicity. Mixed infection with the different A. actinomycetemcomitans serotypes is frequent in periodontitis patients; accordingly, the role of this bacterial species in the pathogenesis of periodontitis may differ depending whether patients or periodontal lesions harbour one or more of the A. actinomycetemcomitans serotypes. We hypothesized that different combinations of these serotypes could be associated with distinct host responses and hence different inflammatory patterns. This investigation was aimed to assess whether the increased immuno-stimulatory potential attributed to the serotype b of A. actinomycetemcomitans on immune cells is able to be modified during co-infection with other A. actinomycetemcomitans serotypes. Methods: Dendritic cells (DCs) were obtained from healthy subjects and stimulated with the different A. actinomycetemcomitans serotypes or their purified LPS using the following stimulatory conditions: serotype a, b, or c, and the combinations a+b, a+c, b+c, or a+b+c. The cytokine, CCR, and CCL levels were quantified by qPCR and ELISA. Results: Higher levels of cytokines, CCRs, and CCLs were induced when DCs were stimulated with the serotype b of A. actinomycetemcomitans compared with the same cells stimulated with the other serotypes. When DCs were co-infected, these levels decreased in comparison with the serotype b-stimulation alone, in particular when the serotype a was present in the mixed infection. Conclusions: The increased immuno-stimulatory potential attributed to the serotype b was modified when DCs were co-infected with other A. actinomycetemcomitans serotypes, in particular, when the serotype a was present, the DC response diminished

Differential human Th22-lymphocyte response triggered by Aggregatibacter actinomycetemcomitans serotypes

Objective: In Aggregatibacter actinomycetemcomitans, different serotypes have been described based on lipopolysaccharide (LPS) antigenicity. When T lymphocytes were stimulated with these serotypes, different patterns of T-helper (Th)1 and Th17-type of immune responses were reported. Recently, two new Th phenotypes have been described and named Th9 and Th22 lymphocytes; however, their role in the pathogenesis of periodontitis remains unclear. This study aimed to investigate the potential Th9 and/ or Th22 lymphocyte responses when stimulated with autologous dendritic cells infected with different A. actinomycetemcomitans serotypes. Methods: Monocyte-derived dendritic cells and naïve CD4+ T lymphocytes were obtained from healthy donors and stimulated with different serotypes of A. actinomycetemcomitans at a multiplicity of infection MOI = 102 or their purified LPS (10–50 ng/ml). The levels for the Th9 and Th22-associated cytokines, as well as the transcription factor master-switch genes implied in their differentiation Spi-B and AhR, were quantified by qPCR and ELISA. Results: When stimulated with the serotype b of A. actinomycetemcomitans, higher levels of interleukin (IL)-6 and tumor necrosis factor (TNF)-a were detected in dendritic cells, as well as higher levels of IL-22 and AhR were detected in T lymphocytes, when compared with stimulation with the other serotypes. Conclusions: The serotype b of A. actinomycetemcomitans has a higher capacity of trigger Th22-type of immune response in both dendritic cells and T lymphocytes. These data allow us to suggest that, when the serotype b of A. actinomycetemcomitans is a significant part of the subgingival biofilm, the Th22 polarization might be triggered within the periodontal lesion

Regulatory T cell phenotype and anti osteoclastogenic function in experimental periodontitis

The alveolar bone resorption is a distinctive feature of periodontitis progression and determinant for tooth loss. Regulatory T lymphocytes (Tregs) display immuno-suppressive mechanisms and tissue repairing functions, which are critical to support periodontal health. Tregs may become unstable and dysfunctional under inflammatory conditions, which can even accelerate tissue destruction. In this study, experimental periodontitis was associated with the progressive and increased presence of Th17 and Treg-related mediators in the gingiva (IL-6, IL-17A, IL-17F, RANKL, IL-10, TGF-beta and GITR; P 25% MFI loss) and increased IL-17A expression (> 15%), compared with Tregs from spleen and healthy controls. Tregs gene expression analysis showed a differential signature between health and disease, with increased expression of Th17-associated factors in periodontitis-derived Tregs. The ex vivo suppression capacity of Tregs on osteoclastic differentiation was significantly lower in Tregs obtained from periodontally diseased animals compared to controls (P < 0.05), as identified by the increased number of TRAP(+) osteoclasts (P < 0.01) in the Tregs/pre-osteoclast co-cultures. Taken together, these results demonstrate that Tregs become phenotypically unstable and lose anti-osteoclastogenic properties during experimental periodontitis; thus, further promoting the Th17-driven bone loss.United States Department of Health & Human Services National Institutes of Health (NIH) - USA NIH National Institute on Aging (NIA) R01AG062496 Agencia Nacional de Investigacion y Desarrollo, Chilean Government FONDECYT 1181780 Millennium Science Initiative from Ministry for the Economy, Development and Tourism, Chile P09/016-F Chilean Government CONICYT 2116125

Alveolar bone resorption and TH1/TH17-associated immune response triggered during Aggregatibacter actinomycetemcomitans-induced experimental periodontitis are serotype-dependent

BACKGROUND: Aggregatibacter actinomycetemcomitans expresses several virulence factors that may contribute to the pathogenesis of periodontitis. Based on the antigenicity of the O-polysaccharide component of the lipopolysaccharide (LPS), different A. actinomycetemcomitans serotypes have been described. Among them, serotype b has demonstrated a stronger capacity to trigger Th1 and Th17-associated cytokine, CC-chemokine, and CC-chemokine receptor production on immune cells in vitro. With a murine model of experimental periodontitis, this investigation aimed to analyze the alveolar bone resorption and the pattern of immune response triggered by the different A. actinomycetemcomitans serotypes within periodontal lesions. METHODS: For periodontal lesion induction, mice were orally infected with the different A. actinomycetemcomitans serotypes or their purified LPS. Alveolar bone resorption was analyzed using microcomputed tomography and scanning electron microscopy. Bacterial infection, receptor activator of nuclear factor-kappa B ligand (RANKL) and Th1 and Th17-associated cytokine, CC-chemokine, and CC-chemokine receptor levels were quantified by quantitative polymerase chain reaction (qPCR). T lymphocytes isolated from periodontal lesions were analyzed by flow cytometry. RESULTS: In periodontal lesions, serotype b of A. actinomycetemcomitans induced higher alveolar bone resorption and expression of RANKL compared with the other serotypes. In addition, serotype b induced greater levels of Th1- and Th17-related cytokines, CC-chemokines, and CC-chemokine receptors than the others. Similarly, higher numbers of infiltrating Th1 and Th17 lymphocytes were detected in serotype b-induced periodontal lesions. CONCLUSIONS: These results demonstrate that periodontal lesions induced with different A. actinomycetemcomitans serotypes elicited distinct alveolar bone resorption and immune response. In particular, serotype b was more pathogenic than the others and induced stronger Th1 and Th17 patterns of immune responses during experimental periodontitis