Changes in the alveolar bone morphology among different patterns of incisor inclination during the alignment phase in orthodontic treatment without premolar extraction.

OBJECTIVE: The present study investigated bone remodelling in the upper and lower incisor regions depending on the inclination pattern during the alignment phase of orthodontic treatment (OT). MATERIALS AND METHODS: This prospective clinical study included 71 patients undergoing OT without premolar extraction. Cone beam computed tomography scans were taken before and after the alignment phase and the changes in the inclination, alveolar bone height (ABH) and bone thickness (BT) at levels 2, 3, 4, 6, 8 and 9 mm starting from the cementoenamel junction (CEJ) were determined. RESULTS: Teeth were divided into 'Retroinclination' (lingual crown inclination <0°), 'Proclination-low' (buccal crown inclination between 0° and 5°), or 'Proclination-high' (buccal crown inclination >5°). The alignment phase of OT resulted in ABH loss. The highest ABH loss in the maxilla was observed on the buccal side in the 'Proclination-high' and was 0.71 mm. ABH loss by 1.1 mm was observed in the mandible on the lingual side in the 'Retroinclination' group. The most significant changes in BT by up to 2 mm were observed at levels 6, 8 and 9 mm and these changes exhibited a moderate to strong correlation with the alterations in the inclination of individual incisors. At levels 2, 3 and 4 mm, the highest decrease in BT by up to 0.83 mm was observed on the palatal side of upper incisors in the 'Proclination-high' group. CONCLUSION: The direction and amount of tooth inclination partially determine changes in the bone parameters during the alignment phase.

Effects of customized CAD/CAM abutments on cytokine levels in peri-implant crevicular fluid during early implant healing: a pilot study.

OBJECTIVES: This study aimed to assess levels of biomarkers associated with inflammation and tissue destruction in peri-implant crevicular fluid (PICF) of implants provided with customized or standard healing abutments during early implant healing. MATERIALS AND METHODS: Thirty implants were placed in 22 patients with partial posterior edentulism. Subsequently, test group implants (n=15) received one-piece titanium abutments that were fabricated using computer-aided design/computer-aided manufacturing (CAD/CAM). Control group implants (n=15) were provided with standard abutments. PICF collection and standardized periapical radiographs were carried out at suture removal one week later, following crown delivery after 3 months and at 6 months. Expression of C-reactive protein (CRP), interferon-γ, tumor necrosis factor (TNF)-α, interleukin (IL)-1α, IL-1ß, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12A, IL-17A, macrophage inflammatory protein (MIP)-1α, matrix metalloproteinase (MMP)-13, osteopontin, osteoactivin, Receptor Activator of NF-κB (RANK), and TGF-ß were analyzed using a multiplex ELISA kit. RESULTS: Both groups showed a significant decrease in protein expression of CRP, IL-1ß, IL-6, IL-8, MIP-1α, osteopontin, osteoactivin, and TGF-ß, while MMP-13 levels increased during the observation period. A rise in OPG and RANK levels was detected among customized abutments. Expression of CRP was higher, whereas IL-1ß, IL-1α, and MIP-1α were decreased in control compared to test group implants after 6 months. Marginal bone loss did not depend on abutment modality. CONCLUSIONS: Both abutment types showed distinctive temporal expression of inflammatory biomarkers during 6 months following implant placement. TRIAL REGISTRATION: ISRCTN98477184, registration date 18/05/2022 CLINICAL RELEVANCE: Customized healing abutments exert similar effects on inflammation during early implant healing compared to standard healing abutments.

Effects of enamel matrix derivative in nonsurgical periodontal therapy on pro-inflammatory profiles, microbial environment and clinical outcome: a randomized clinical trial.

OBJECTIVES: This study aimed to evaluate the impact of enamel matrix derivative (EMD) application following subgingival instrumentation of residual pockets in periodontitis patients on inflammatory host response, microbiological composition, and clinical outcome. METHODS: In this double-blinded randomized controlled trial, a total of 22 patients with generalized periodontitis stage III or IV presenting with ≥ 6 mm probing pocket depth (PPD) at re-evaluation after initial periodontal therapy were included. Participants were randomly allocated at a 1:1 ratio to subgingival instrumentation with (EMD +) or without (EMD-) non-surgical EMD application into the pocket. PPD, clinical attachment level (CAL), bleeding on probing (BoP), plaque index (PI), as well as a panel of pro-inflammatory cytokines and periodontal pathogen count in the gingival crevicular fluid (GCF) of the respective sites were evaluated at baseline (T0) and six months afterwards (T1). RESULTS: Both treatment groups showed a significant PPD reduction (EMD + 1.33 ± 1.15 mm, p < 0.001; EMD- 1.32 ± 1.01 mm, p < 0.001) as well as CAL gain (EMD + 1.13 ± 1.58 mm, p < 0.001; EMD- 0.47 ± 1.06 mm, p = 0.005) from T0 to T1. While no intergroup differences for PPD reduction were observed, CAL gain was higher in EMD + sites compared to EMD- (p = 0.009). No essential effects on cytokine expression as well as bacterial count were detected. CONCLUSIONS: Application of EMD as an adjunct to subgingival instrumentation of residual pockets yielded benefits regarding CAL gain; however, effects on PPD reduction, inflammatory cytokines, and bacterial count were negligible. TRIAL REGISTRATION: ClinicalTrials.gov (NCT04449393), registration date 26/06/2020. CLINICAL RELEVANCE: Based on the obtained results, additional non-surgical EMD application compared to subgingival instrumentation alone showed no clinically relevant effects on treatment outcome and underlying biological mechanisms.

Osteogenic Differentiation of Human Periodontal Ligament Stromal Cells Influences Their Immunosuppressive Potential toward Allogenic CD4+ T Cells.

The differentiation ability of human periodontal ligament mesenchymal stromal cells (hPDL-MSCs) in vivo is limited; therefore, some studies considered strategies involving their pre-differentiation in vitro. However, it is not known how the differentiation of hPDL-MSCs influences their immunomodulatory properties. This study investigated how osteogenic differentiation of hPDL-MSCs affects their ability to suppress CD4+ T-lymphocyte proliferation. hPDL-MSCs were cultured for 21 days in osteogenic differentiation or standard culture media. Allogeneic CD4+ T lymphocytes were co-cultured with undifferentiated and differentiated cells in the presence or absence of interferon (IFN)-γ, interleukin (IL)-1ß or tumor necrosis factor (TNF)-α, and their proliferation and apoptosis were measured. Additionally, the effects of these cytokines on the expression of immunomodulatory or pro-inflammatory factors were investigated. Our data show that osteogenic differentiation of hPDL-MSCs reduced their ability to suppress the proliferation of CD4+ T lymphocytes in the presence of IFN-γ and enhanced this ability in the presence of IL-1ß. These changes were accompanied by a slightly decreased proportion of apoptotic CD4+ in the presence of IFN-γ. The osteogenic differentiation was accompanied by decreases and increases in the activity of indoleamine-2,3-dioxygenase in the presence of IFN-γ and IL-1ß, respectively. The basal production of interleukin-8 by hPDL-MSCs was substantially increased upon osteogenic differentiation. In conclusion, this study suggests that pre-differentiation strategies in vitro may impact the immunomodulatory properties of hPDL-MSCs and subsequently affect their therapeutic effectiveness in vivo. These findings provide important insights for the development of MSC-based therapies.

Anti-apoptotic effects of human gingival mesenchymal stromal cells on polymorphonuclear leucocytes.

OBJECTIVES: Polymorphonuclear leucocytes (PMNs) constitute the first line of host defence and are crucial in maintaining periodontal health. Their survival and function are modulated by mesenchymal stromal cells (MSCs) from different origin. Gingival MSCs (GMSCs) play an important role in maintaining oral health and in the initial inflammatory response. The present study aimed to investigate the effects of GMSCs on PMNs apoptosis and reactive oxygen species (ROS) production. METHODS: PMNs were either directly incubated with untreated, interleukin (IL)-1ß- or tumour necrosis factor (TNF)-α-treated GMSCs or stimulated with their conditioned media. Resulting ROS production was evaluated by dichlorofluorescin diacetate staining, whereas PMNs apoptosis was assessed by Annexin V staining, followed by flow cytometry analysis. RESULTS: While conditioned media of untreated and TNF-α-treated GMSCs did not affect apoptosis of PMNs, it was significantly delayed by conditioned media of GMSCs treated with IL-1ß. In direct co-culture, GMSCs exerted anti-apoptotic effects on PMNs independently of the previous stimulation. However, the strongest impact was observed by IL-1ß-treated GMSCs. ROS production of PMNs was not influenced by GMSCs or their conditioned media. CONCLUSION: This study demonstrates for the first time the immunomodulatory properties of GMSCs towards PMNs, revealing that IL-1ß enhances anti-apoptotic effects of GMSCs.

Cyclic tensile strain affects the response of human periodontal ligament stromal cells to tumor necrosis factor-α.

OBJECTIVES: Orthodontic treatment in adult patients predisposed to mild or severe periodontal disease is challenging for orthodontists. Orthodontic malpractice or hyper-occlusal forces may aggravate periodontitis-induced destruction of periodontal tissues, but the specific mechanism remains unknown. In the present study, the combined effect of mechanical stress and tumor necrosis factor (TNF)-α on the inflammatory response in human periodontal ligament stromal cells (hPDLSCs) was investigated. MATERIALS AND METHODS: hPDLSCs from 5 healthy donors were treated with TNF-α and/or subjected to cyclic tensile strain (CTS) of 6% or 12% elongation with 0.1 Hz for 6- and 24 h. The gene expression of interleukin (IL)-6, IL-8 and cell adhesion molecules VCAM and ICAM was analyzed by qPCR. The protein levels of IL-6 and IL-8 in conditioned media was measured by ELISA. The surface expression of VCAM-1 and ICAM-1 was quantified by immunostaining followed by flow cytometry analysis. RESULTS: TNF-α-induced IL-6 gene and protein expression was inhibited by CTS, whereas TNF-α-induced IL-8 expression was decreased at mRNA expression level but enhanced at the protein level in a magnitude-dependent manner. CTS downregulated the gene expression of VCAM-1 and ICAM-1 under TNF-α stimulation, but the downregulation of the surface expression analyzed by flow cytometry was observed chiefly for VCAM-1. CONCLUSIONS: Our findings show that mechanical force differentially regulates TNF-α-induced expression of inflammatory mediators and adhesion molecules at the early stage of force application. The effect of cyclic tensile strain is complex and could be either anti-inflammatory or pro-inflammatory depending on the type of pro-inflammatory mediators and force magnitude. CLINICAL RELEVANCE: Orthodontic forces regulate the inflammatory mediators of periodontitis. The underlying mechanism may have significant implications for future strategies of combined periodontal and orthodontic treatment.

Effect of Titanium and Zirconia Nanoparticles on Human Gingival Mesenchymal Stromal Cells.

Nano- and microparticles are currently being discussed as potential risk factors for peri-implant disease. In the present study, we compared the responses of human gingival mesenchymal stromal cells (hG-MSCs) on titanium and zirconia nanoparticles (<100 nm) in the absence and presence of Porphyromonas gingivalis lipopolysaccharide (LPS). The primary hG-MSCs were treated with titanium and zirconia nanoparticles in concentrations up to 2.000 µg/mL for 24 h, 72 h, and 168 h. Additionally, the cells were treated with different nanoparticles (25−100 µg/mL) in the presence of P. gingivalis LPS for 24 h. The cell proliferation and viability assay and live−dead and focal adhesion stainings were performed, and the expression levels of interleukin (IL)-6, IL-8, and monocyte chemoattractant protein (MCP)-1 were measured. The cell proliferation and viability were inhibited by the titanium (>1000 µg/mL) but not the zirconia nanoparticles, which was accompanied by enhanced apoptosis. Both types of nanoparticles (>25 µg/mL) induced the significant expression of IL-8 in gingival MSCs, and a slightly higher effect was observed for titanium nanoparticles. Both nanoparticles substantially enhanced the P. gingivalis LPS-induced IL-8 production; a higher effect was observed for zirconia nanoparticles. The production of inflammatory mediators by hG-MSCs is affected by the nanoparticles. This effect depends on the nanoparticle material and the presence of inflammatory stimuli.

Effect of vitamin D3 on the osteogenic differentiation of human periodontal ligament stromal cells under inflammatory conditions.

OBJECTIVES: Vitamin D3 is known to activate osteogenic differentiation of human periodontal ligament stromal cells (hPDLSCs). Recently, inflammatory stimuli were shown to inhibit the transcriptional activity of hPDLSCs, but their effect on vitamin D3 -induced osteogenic differentiation is not known. The present study aimed to investigate whether the effects of 1,25-dihydroxvitamin D3 (1,25(OH)2 D3 ) and 25-hydroxvitamin D3 (25(OH)D3 ) on the osteogenic differentiation of hPDLSCs are also altered under inflammatory conditions. Furthermore, the expression of osteogenesis-related factors by hPDLSCs under osteogenic conditions was assessed in the presence of inflammatory stimuli. MATERIALS AND METHODS: Primary hPDLSCs of six donors were cultured in osteogenic induction medium containing either 1,25(OH)2 D3 (0-10 nM) or 25(OH)D3 (0-100 nM) in the presence and absence of Porphyromonas gingivalis lipopolysaccharide (LPS) or Pam3CSK4 for 7, 14 and 21 days. Osteogenic differentiation of hPDLSCs was evaluated by analysis of mineralization as assessed by Alizarin Red S staining and gene expression levels of osteogenesis-related factors osteocalcin, osteopontin and runt-related transcription factor 2 (RUNX2) were analysed with qPCR. RESULTS: Treatment with 1,25(OH)2 D3 significantly enhanced the osteogenic differentiation of hPDLSCs and their expression of osteocalcin and osteopontin. The 1,25(OH)2 D3 -triggered expression of osteogenesis-related factors was significantly lower in the presence of Pam3CSK4, but not P. gingivalis LPS. None of the inflammatory stimuli had significant effects on the 1,25(OH)2 D3 -induced osteogenic differentiation. 25(OH)D3 neither affected gene expression levels nor osteogenic differentiation of hPDLSCs cultured in osteogenic induction medium. CONCLUSION: The results of this study indicate that inflammatory stimuli also diminish the 1,25(OH)2 D3 -induced expression of osteogenesis-related factors in hPDLSCs under osteogenic conditions, while having no effect on the osteogenic differentiation.

Effects of Er:YAG laser irradiation of different titanium surfaces on osteoblast response.

The aim of this in vitro study was to evaluate the effects of erbium-doped yttrium aluminum garnet (Er:YAG) laser irradiation on titanium surface topography and the proliferation and differentiation of osteoblasts using standard clinical treatment settings. Er:YAG laser irradiation at two levels ((1): 160 mJ, pulse at 20 Hz; (2): 80 mJ, pulse at 20 Hz) was applied to moderately rough and smooth titanium disks before MG-63 osteoblast-like cells were cultured on these surfaces. Titanium surface and cell morphology were observed by scanning electron microscopy. Cell proliferation/viability was measured by CCK-8 test. Gene expression of alkaline phosphatase (ALP), osteocalcin (OC), osteoprotegerin (OPG), receptor activator of nuclear factor kappa-B ligand (RANKL), and collagen type 1 was measured by qPCR, and OPG and OC protein production was determined by enzyme-linked immunosorbent assay. Treatment with Er:YAG laser at 160 mJ/20 Hz markedly caused heat-induced fusion of titanium and cell condensation on moderately rough surfaces, but not in smooth surfaces. MG-63 proliferation/viability decreased after 5 days in moderately rough surfaces. The expression of ALP, OC, OPG, and collagen type 1 was unaffected by laser treatment at 160 mJ/20. Laser irradiation at 80 mJ/20 Hz enhanced RANKL gene expression after 5 days in moderately rough surfaces. Study results suggest that Er:YAG laser irradiation at clinically relevant setting has no essential effect on osteogenic gene and protein expression of osteoblasts. However, surface structure, cell attachment, and proliferation are influenced by both treatment protocols, which implies that caution should be taken in the clinical treatment of peri-implant diseases when Er:YAG laser is used.

Salivary MRP-8/14 and the presence of periodontitis-associated bacteria in children with bonded maxillary expansion treatment.

OBJECTIVES: The aim of this study was to investigate changes in saliva concentration of the inflammatory marker MRP-8/14 and the presence of some periodontitis-associated bacteria in patients with mixed dentition treated with a rigid acrylic, bonded maxillary expander. METHODS: Fifteen patients in mixed dentition treated with a bonded palatal expander were enrolled in this longitudinal study. Saliva samples were taken before the therapy, as well as in 2 weeks and 3, 6, 9, and 12 months after the beginning of the therapy. In each sample, the levels of MRP-8/14 were determined by ELISA and the presence of 11 bacteria was detected by PCR followed by DNA-DNA hybridization. RESULTS: Salivary concentration of MRP-8/14 and the amount of Tannerella forsythia, Treponema denticola, and Eikenella corrodens were significantly increased during treatment with bonded maxillary expander. These changes were transient and the maximal levels of MRP-8/14 and periodontitis-associated pathogens were observed 6-9 months after the beginning of the therapy. CONCLUSION: Therapy with bonded maxillary results in higher MRP-8/14 levels and increased prevalence of some periodontitis-associated bacteria, namely T. forsythia, T. denticola, and E. corrodens. The results suggest the detection of salivary MRP-8/14 levels may be a potential tool to reflect the oral health status in children with fixed orthodontic treatment. CLINICAL RELEVANCE: Our data suggest that the treatment with bonded maxillary expander might influence the oral health status and should be accompanied by the careful control of the oral health during the therapy.

Interleukin-1ß Induced Matrix Metalloproteinase Expression in Human Periodontal Ligament-Derived Mesenchymal Stromal Cells under In Vitro Simulated Static Orthodontic Forces.

The periodontal ligament (PDL) responds to applied orthodontic forces by extracellular matrix (ECM) remodeling, in which human periodontal ligament-derived mesenchymal stromal cells (hPDL-MSCs) are largely involved by producing matrix metalloproteinases (MMPs) and their local inhibitors (TIMPs). Apart from orthodontic forces, the synthesis of MMPs and TIMPs is influenced by the aseptic inflammation occurring during orthodontic treatment. Interleukin (IL)-1ß is one of the most abundant inflammatory mediators in this process and crucially affects the expression of MMPs and TIMPs in the presence of cyclic low-magnitude orthodontic tensile forces. In this study we aimed to investigate, for the first time, how IL-1ß induced expression of MMPs, TIMPs and how IL-1ß in hPDL-MSCs was changed after applying in vitro low-magnitude orthodontic tensile strains in a static application mode. Hence, primary hPDL-MSCs were stimulated with IL-1ß in combination with static tensile strains (STS) with 6% elongation. After 6- and 24 h, MMP-1, MMP-2, TIMP-1 and IL-1ß expression levels were measured. STS alone had no influence on the basal expression of investigated target genes, whereas IL-1ß caused increased expression of these genes. In combination, they increased the gene and protein expression of MMP-1 and the gene expression of MMP-2 after 24 h. After 6 h, STS reduced IL-1ß-induced MMP-1 synthesis and MMP-2 gene expression. IL-1ß-induced TIMP-1 gene expression was decreased by STS after 6- and 24-h. At both time points, the IL-1ß-induced gene expression of IL-1ß was increased. Additionally, this study showed that fetal bovine serum (FBS) caused an overall suppression of IL-1ß-induced expression of MMP-1, MMP-2 and TIMP-1. Further, it caused lower or opposite effects of STS on IL-1ß-induced expression. These observations suggest that low-magnitude orthodontic tensile strains may favor a more inflammatory and destructive response of hPDL-MSCs when using a static application form and that this response is highly influenced by the presence of FBS in vitro.

MMPs and TIMPs Expression Levels in the Periodontal Ligament during Orthodontic Tooth Movement: A Systematic Review of In Vitro and In Vivo Studies.

Background: During orthodontic tooth movement (OTM), applied orthodontic forces cause an extensive remodeling of the extracellular matrix (ECM) in the periodontal ligament (PDL). This is mainly orchestrated by different types of matrix metalloproteinases (MMPs) and their tissue inhibitors of matrix metalloproteinases (TIMPs), which are both secreted by periodontal ligament (PDL) fibroblasts. Multiple in vitro and in vivo studies already investigated the influence of applied orthodontic forces on the expression of MMPs and TIMPs. The aim of this systematic review was to explore the expression levels of MMPs and TIMPs during OTM and the influence of specific orthodontic force-related parameters. Methods: Electronic article search was performed on PubMed and Web of Science until 31 January 2021. Screenings of titles, abstracts and full texts were performed according to PRISMA, whereas eligibility criteria were defined for in vitro and in vivo studies, respectively, according to the PICO schema. Risk of bias assessment for in vitro studies was verified by specific methodological and reporting criteria. For in vivo studies, risk of bias assessment was adapted from the Joanna Briggs Institute Critical Appraisal Checklist for analytical cross-sectional study. Results: Electronic article search identified 3266 records, from which 28 in vitro and 12 in vivo studies were included. The studies showed that orthodontic forces mainly caused increased MMPs and TIMPs expression levels, whereas the exact effect may depend on various intervention and sample parameters and subject characteristics. Conclusion: This systematic review revealed that orthodontic forces induce a significant effect on MMPs and TIMPs in the PDL. This connection may contribute to the controlled depletion and formation of the PDLs' ECM at the compression and tension site, respectively, and finally to the highly regulated OTM.

Effect of Enamel Matrix Derivatives on Osteoclast Formation from PBMC of Periodontitis Patients and Healthy Individuals after Interaction with Activated Endothelial Cells.

Background and objectives: Enamel matrix derivative (EMD) is produced from developing porcine tooth buds and represents a complex of low-molecular-weight hydrophobic enamel proteins. EMD is widely applied in periodontal regeneration. Osteoclasts are multinuclear cells, which are responsible for bone resorption. The precursors of osteoclasts, hematopoietic cells, undergo in vivo the process of transendothelial migration before differentiation. EMD is known to affect the process of osteoclastogenesis, but its effect on human osteoclasts precursors after the interaction with activated endothelium was never studied. Materials and Methods: Human umbilical vein endothelial cells (HUVECs)s were seeded in transwell inserts with a pore size of 8 µm and pre-activated by TNF-α and IL-1ß for 18 h. Peripheral blood mononuclear cells (PBMCs), freshly isolated from 16 periodontitis patients and 16 healthy individuals, were added to pre-activated HUVECs. Adherent, non-adherent and transmigrated cells were collected and differentiated to osteoclasts by the standard protocol in the presence or absence of EMD. The number of osteoclasts was determined by tartrate-resistant acid phosphatase staining. Results: PBMCs isolated from periodontitis patients have formed a significantly higher osteoclast number compared to PBMCs isolated from healthy individuals (p < 0.05). EMD induced concentration-dependent inhibition of osteoclast formation from PBMCs. This was true for the different PBMC fractions isolated from both healthy individuals and periodontitis patients. Conclusions: Our data show that EMD inhibits the formation and activity of osteoclasts differentiated from the progenitor cells after the interaction with activated endothelium. This might be associated with bone resorption inhibition and supporting bone regeneration in the frame of periodontal therapy.

Pleiotropic effects of vitamin D3 on CD4+ T lymphocytes mediated by human periodontal ligament cells and inflammatory environment.

AIMS: Both, vitamin D3 and human periodontal ligament cells (hPDLCs) possess immunosuppressive properties, but their combined effect on immune cells has never been investigated. Here, we analysed the impact of vitamin D3 on the immunosuppressive properties of hPDLCs towards CD4+ T lymphocytes. MATERIAL AND METHODS: Allogenic CD4+ T lymphocytes were activated by phytohemagglutinin either in monoculture or co-culture with hPDLCs, in the presence or absence of IFN-γ and 1,25(OH)2 D3 . After 5 days, CD4+ T-lymphocyte proliferation, CD4+ CD25+ FoxP3+ regulatory T lymphocytes (Tregs ) proportion and IL-10, TGF-ß1 and IL-17A production were analysed. RESULTS: In monoculture, 1,25(OH)2 D3 suppressed CD4+ T-lymphocyte proliferation, increased the percentage of CD4+ FoxP3+ CD25+ FoxP3+ Tregs and enhanced IL-10 and TGF-ß1 production. In the presence of IFN-γ treated hPDLCs, 1,25(OH)2 D3 significantly increased CD4+ T-lymphocyte proliferation and decreased the percentage of CD4+ CD25+ FoxP3+ Tregs . IL-10 and IL-17A expression was significantly diminished by 1,25(OH)2 D3 , whereas TGF-ß1 was slightly increased. The effects of 1,25(OH)2 D3 in co-culture were reversed by inhibition of indoleamine-2,3-dioxygenase-1, prostaglandin-endoperoxide synthase and programmed cell death 1 ligand 1. 1,25(OH)2 D3 also suppressed the expression of these proteins in hPDLCs. CONCLUSION: Effects of vitamin D3 on CD4+ T lymphocyte are modified by hPDLCs depending on the microenvironment.

Response of Human Mesenchymal Stromal Cells from Periodontal Tissue to LPS Depends on the Purity but Not on the LPS Source.

Human periodontal ligament stromal cells (hPDLSCs) and gingival mesenchymal stromal cells (hGMSCs) are resident mesenchymal stromal cells (MSCs) of the periodontal tissue. The lipopolysaccharide (LPS) from Porphyromonas gingivalis is structurally distinct from that of other Gram-negative bacteria, and earlier studies linked this structural difference to a distinct virulence activity and the ability to activate toll-like receptor 2 (TLR-2), besides TLR-4 as commonly occurring upon LPS challenge. Later studies, in contrast, argue that TLR-2 activation by P. gingivalis LPS is due to lipoprotein contamination. In the present study, we aimed to define the influence of structure versus purity of P. gingivalis LPS on the immune response of hPDLSCs and hGMSCs. Cells were stimulated with commercially available "standard" P. gingivalis LPS, "ultrapure" P. gingivalis LPS, or "ultrapure" Escherichia coli LPS, and the expression of interleukin- (IL-) 8, IL-6, monocyte chemoattractant protein- (MCP-) 1, TLR-2, and TLR-4 was evaluated. The contribution of TLR-4 to the LPS-induced response was assessed using the specific TLR-4 inhibitor TAK-242. "Standard" P. gingivalis LPS induced significantly higher IL-8, IL-6, and MCP-1 production compared to the "ultrapure" LPS preparations, with no significant difference detectable for "ultrapure" LPS from P. gingivalis and E. coli. By using TAK-242, the response of hPDLSCs and hGMSCs to "ultrapure" LPS preparations was effectively inhibited to the levels comparable to those of nonstimulated controls. In contrast, high levels of response to "standard" LPS were observed, even in the presence of TAK-242. Our data show that the response of MSCs from periodontal tissue to LPS depends more on the purity of the LPS preparation than on the LPS source. Even a small amount of contaminating lipoproteins can drastically enhance the hPDLSCs' and hGMSCs; responsiveness to P. gingivalis LPS, which might also contribute to the progression of periodontal disease.

Effect of bisphosphonate treatment of titanium surfaces on alkaline phosphatase activity in osteoblasts: a systematic review and meta-analysis.

BACKGROUND: Bisphosphonate coating of dental implants is a promising tool for surface modification aiming to improve the osseointegration process and clinical outcome. The biological effects of bisphosphonates are thought to be mainly associated with osteoclasts inhibition, whereas their effects on osteoblast function are unclear. A potential of bisphosphonate coated surfaces to stimulate osteoblast differentiation was investigated by several in vitro studies with contradictory results. The purpose of this systematic review and meta-analysis was to evaluate the effect of bisphosphonate coated implant surfaces on alkaline phosphatase activity in osteoblasts. METHODS: In vitro studies that assessed alkaline phosphatase activity in osteoblasts following cell culture on bisphosphonate coated titanium surfaces were searched in electronic databases PubMed/MEDLINE, Scopus and ISI Web of Science. Animal studies and clinical trials were excluded. The literature search was restricted to articles written in English and published up to August 2019. Publication bias was assessed by the construction of funnel plots. RESULTS: Eleven studies met the inclusion criteria. Meta-analysis showed that coating of titanium surfaces with bisphosphonates increases alkaline phosphatase activity in osteoblasts after 3 days (n = 1), 7 (n = 7), 14 (n = 6) and 21 (n = 3) days. (7 days beta coefficient = 1.363, p-value = 0.001; 14 days beta coefficient = 1.325, p-value < 0.001; 21 days beta coefficient = 1.152, p-value = 0.159). CONCLUSIONS: The meta-analysis suggests that bisphosphonate coatings of titanium implant surfaces may have beneficial effects on osteogenic behaviour of osteoblasts grown on titanium surfaces in vitro. Further studies are required to assess to which extent bisphosphonates coating might improve osseointegration in clinical situations.

Methanandamide diminish the Porphyromonas gingivalis lipopolysaccharide induced response in human periodontal ligament cells.

BACKGROUND: The endocannabinoid system is involved in the regulation of periodontal tissue homeostasis. Synthetic cannabinoid methanandamide (Meth-AEA) has improved stability and affinity to cannabinoid receptors compared to its endogenous analog anandamide. In the present study, we investigated the effect of methanandamide on the production of pro-inflammatory mediators in primary human periodontal ligament cells (hPdLCs). METHODS: hPdLCs were treated with Meth-AEA for 24 h, and the resulting production of interleukin (IL)-6, IL-8, and monocyte chemotactic protein (MCP)-1 was measured in the absence or the presence of Porphyromonas gingivalis lipopolysaccharide (LPS). Additionally, the effect of Meth-AEA on the proliferation/viability of hPdLCs was measured by the MTT method. RESULTS: Methanandamide at a concentration of 10 µM significantly inhibited P. gingivalis LPS induced production of IL-6, IL-8, and MCP-1. Basal production of IL-6 and IL-8 was slightly enhanced by 10 µM Meth-AEA. No effect of Meth-AEA on the basal production of MCP-1 was observed. Meth-AEA in concentrations up to 10 µM did not affect the proliferation/viability of hPdLCs, but significantly inhibited it at a concentration of 30 µM. CONCLUSION: Our study suggests that the inflammatory response in periodontal ligament cells could be influenced by the activation of the cannabinoid system, which might be potentially involved in the progression of periodontal disease.

Soluble CD14 Enhances the Response of Periodontal Ligament Stem Cells to Toll-Like Receptor 2 Agonists.

Human periodontal ligament stem cells (hPDLSCs) do not express membrane-bound CD14, and their responsiveness to bacterial lipopolysaccharide (LPS) is drastically enhanced by soluble CD14 (sCD14), which is due to the facilitation of the interaction between LPS and Toll-like receptor- (TLR-) 4. Several studies also show that sCD14 enhances the responsiveness of different immune cells to TLR-2, but such effect in hPDLSCs has not been studied so far. In the present study, we investigated for the first time the potential effect of sCD14 on the hPDLSC response to two different TLR-2 agonists, in vitro. Primary hPDLSCs were stimulated with synthetic lipopeptide Pam3CSK4 or lipoteichoic acid (LTA) in concentrations 1-1000 ng/ml in the presence/absence of sCD14 (250 ng/ml). Additionally, the effect of different sCD14 concentrations (2.5-250 ng/ml) on the TLR-2 response was determined in Pam3CSK4- or LTA-triggered hPDLSCs. The resulting expression of interleukin- (IL-) 6, chemokine C-X-C motif ligand 8 (CXCL8), and chemokine C-C motif ligand 2 (CCL2) was measured by qPCR and ELISA. The production of IL-6, CXCL8, and CCL2 was gradually increased by both TLR-2 agonists and was significantly enhanced by sCD14. The response of hPDLSCs to low and submaximal concentrations of TLR-2 agonists (1-100 ng/ml) was most effectively enhanced by sCD14. The effect of sCD14 on TLR-2 response in hPDLSCs was concentration-dependent and was already detectable at low sCD14 levels. Our data showed that exogenous sCD14 significantly enhanced the responsiveness of hPDLSCs to TLR-2 agonists and enabled the detection of their small amounts. This effect was already detectable at low sCD14 levels, which are comparable to those in saliva and gingival crevicular fluid. Changes in the local sCD14 level may be considered as a crucial factor influencing the susceptibility of hPDLSCs to different pathogens and thus may contribute to the progression of periodontitis.

Meta-analysis of the prognostic value of the neutrophil-to-lymphocyte ratio in oral squamous cell carcinoma.

BACKGROUND: A number of studies have assessed the prognostic value of the neutrophil-to-lymphocyte ratio (NLR) in oral squamous cell carcinoma (OSCC), but their results regarding the predictive value of NLR in OSCC are inconsistent. We therefore performed a meta-analysis to clarify the association between NLR and clinical outcome in OSCC. METHODS: We searched the MEDLINE and Web of Science to identify potential studies investigating the association between NLR and survival in OSCC. RESULTS: A total of 10 studies, enrolling 2135 patients with OSCC, were included. A higher NLR was a negative predictor for both disease-specific survival (hazard ratio [HR] = 1.93, 95% CI: 1.47-2.54) and overall survival (HR = 1.56, 95% CI: 1.28-1.90). CONCLUSION: This suggests a higher NLR is predictive of a poorer prognosis in OSCC. Because determination of NLR is non-invasive and cost-effective, it could be widely used for predicting prognosis in OSCC.

Serum levels of 25-hydroxyvitamin D are associated with periodontal disease.

OBJECTIVES: Vitamin D plays an essential role in bone metabolism as well as in immunity. Hence, it might affect the development and extent of periodontal disease. The aim of this study was the assessment of 25-hydroxyvitamin D (25(OH)D) status in periodontal disease. MATERIALS AND METHODS: Twenty-nine patients with severe periodontal disease and 29 healthy volunteers were recruited in this case-control-study. Serum 25(OH)D levels, Periodontal Probing Depth (PPD), Clinical Attachment Level (CAL), Bleeding on Probing (BOP), Body Mass Index (BMI), and current smoking status and smoking history (packyears) were assessed in all participants. Serum 25(OH)D levels were compared between controls and cases. Multivariable logistic regression was used to determine the odds ratio (OR) and 95 % confidence interval (CI) for periodontal disease in 25(OH)D deficient probands. RESULTS: Patients with periodontal disease presented a significantly higher proportion of deficient 25(OH)D levels (i.e., <50 nmol/l) compared to healthy controls (48 vs. 14 % respectively). The adjusted OR for periodontal disease with vitamin D deficiency was 1.5 (95 % CI, 1.13-1.98). No correlation between serum 25(OH)D levels and CAL, PPD, and BOP in the group with periodontal disease was found. CONCLUSIONS: In this case-control-study 25(OH)D deficiency is significantly associated with periodontal disease. CLINICAL RELEVANCE: The assessment of vitamin D levels in patients presenting with periodontal disease seems advisable, as vitamin D deficiency might be involved in the onset and progression of periodontal disease.