Daniel Jönsson

The fit of cobalt–chromium three-unit fixed dental prostheses fabricated with four different techniques: A comparative in vitro study

OBJECTIVES This study sought to evaluate and compare the marginal and internal fit in vitro of three-unit FDPs in Co-Cr made using four fabrication techniques, and to conclude in which area the largest misfit is present. METHODS An epoxy resin master model was produced. The impression was first made with silicone, and master and working models were then produced. A total of 32 three-unit Co-Cr FDPs were fabricated with four different production techniques: conventional lost-wax method (LW), milled wax with lost-wax method (MW), milled Co-Cr (MC), and direct laser metal sintering (DLMS). Each of the four groups consisted of eight FDPs (test groups). The FDPs were cemented on their cast and standardised-sectioned. The cement film thickness of the marginal and internal gaps was measured in a stereomicroscope, digital photos were taken at 12× magnification and then analyzed using measurement software. Statistical analyses were performed with one-way ANOVA and Tukeys test. RESULTS Best fit based on the means (SDs) in μm for all measurement points was in the DLMS group 84 (60) followed by MW 117 (89), LW 133 (89) and MC 166 (135). Significant differences were present between MC and DLMS (p<0.05). The regression analyses presented differences within the parameters: production technique, tooth size, position and measurement point (p < 0.05). SIGNIFICANCE Best fit was found in the DLMS group followed by MW, LW and MC. In all four groups, best fit in both abutments was along the axial walls and in the deepest part of the chamfer preparation. The greatest misfit was present occlusally in all specimens.

The human periodontal ligament cell: a fibroblast-like cell acting as an immune cell

BACKGROUND Periodontal ligament cells are fibroblast-like cells characterized by collagen production but also possessing some osteoblastic features. In the light of numerous studies presented during recent times, which show that human periodontal ligament cells also produce cytokines and chemokines in response to inflammation promoters, it is reasonable to suggest that periodontal ligament cells play a role as promoters of periodontal inflammation through these mechanisms. MATERIAL AND METHODS The periodontal ligament, which harbours the periodontal ligament cells, is a part of the attachment apparatus comprised of periodontal ligament cells, extracellular matrix and fibres, attaching the root cement to the alveolar bone. Periodontal ligament cells are in close proximity to bacteria within the plaque and the pocket, and thus these cells are readily accessible to bacterial endotoxins and other promoters of inflammation. RESULTS Cytokines and chemokines, released by periodontal ligament cells upon stimulation with inflammation promoters, reach the blood vessels easily thanks to rich vascularization of the periodontium stimulating recruitment of white blood cells to the site of inflammation. In addition to classical inflammatory cells, such as leucocytes, macrophages and mast cells, the periodontal ligament cells also contribute to periodontal inflammation via their production and release of cytokines and chemokines. CONCLUSION Therefore, pharmacological treatment of periodontitis should aim to reduce the release of proinflammatory agents not only from classical inflammatory cells but also from periodontal ligament cells.

Impact of monocytic cells on recovery of uncultivable bacteria from atherosclerotic lesions

Abstract.  Rafferty B, Jönsson D, Kalachikov S, Demmer RT, Nowygrod R, Elkind MSV, Bush Jr H, Kozarov E. (Columbia University Medical Center, New York, NY; and Weill Cornell Medical College, New York, NY; USA) Impact of monocytic cells on recovery of uncultivable bacteria from atherosclerotic lesions. J Intern Med 2011; 270: 273–280.

LPS-induced MCP-1 and IL-6 production is not reversed by oestrogen in human periodontal ligament cells

OBJECTIVE Periodontal ligament (PDL) cells express oestrogen receptors but the functional importance of oestrogen in PDL cells exposed to bacterial endotoxins is not known. Here we investigate if the inflammation promoter lipopolysaccharide (LPS) affects PDL cell production of interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1), C-reactive protein (CRP) and/or normal functional PDL cell characteristics such as collagen synthesis and cell proliferation and if oestrogen modulates the effects of LPS. METHODS PDL cells were obtained from periodontal ligament of premolars. PDL cells were treated with Escherichia coli LPS in the absence or presence of oestrogen (17beta-oestradiol, E2). Cellular concentration of IL-6, MCP-1 and CRP was determined by enzyme-linked immunosorbent assay (ELISA). Collagen synthesis was determined by l-[3H]proline incorporation. Cell proliferation was assessed by DNA synthesis measurement using [3H]thymidine incorporation. RESULTS Stimulation with LPS (500 ng/ml to 10 microg/ml) increased IL-6 production in a concentration-dependent manner. Lower concentration (100 ng/ml) of LPS had no effect. LPS-induced stimulation of IL-6 was not reversed by a physiologically high concentration (100 nM) of E2. LPS increased also MCP-1 production which was unaffected by E2. Treatment with E2 alone had no effect on either IL-6 or MCP-1. Stimulation with LPS had no effect on CRP. LPS did not affect collagen synthesis and cell proliferation, reflecting normal physiological properties of PDL cells. CONCLUSIONS LPS stimulates PDL cell IL-6 and MCP-1 production but has no effect on the normal physiological properties of PDL cells. LPS-induced IL-6 and MCP-1 is not reversed by oestrogen suggesting that oestrogen exerts no anti-inflammatory effect via this mechanism.

Immunocytochemical demonstration of estrogen receptor beta in human periodontal ligament cells.

Two transcription associated estrogen receptor (ER) subtypes have been identified and named ERalpha and ERbeta. In the present study we investigate the expression of these ER subtypes in cultured human periodontal ligament (PDL) cells by immunocytochemistry. ERbeta immunoreactivity was observed in the nuclei of about 40% of the PDL cells, while no ERalpha immunoreactivity was detected. In human breast cancer MCF-7 cells, serving as positive controls, both ERalpha and ERbeta immunoreactivities were demonstrated. No immunoreactivity was observed after omission of the primary antibodies. This study suggests that estrogen acts on gene transcription preferentially via ERbeta in human PDL cells.

The antimicrobial peptide LL-37 alters human osteoblast Ca2+ handling and induces Ca2+-independent apoptosis

The human antimicrobial peptide cathelicidin LL-37 has, besides its antimicrobial properties, also been shown to regulate apoptosis in a cell type-specific manner. Mechanisms involved in LL-37-regulated apoptotic signaling are not identified. Here, we show that LL-37 reduces the human osteoblast-like MG63 cell number and cell viability in the micromolar concentration range with an IC₅₀ value of about 5 µ<smlcap>M</smlcap>. Treatment with 4 µ<smlcap>M</smlcap> LL-37 increased the number of annexin V-positive cells and stimulated activation of caspase 3 showing that LL-37 promotes apoptosis. Treatment with 4 µ<smlcap>M</smlcap> LL-37 caused an acute and sustained rise in intracellular Ca²⁺ concentration assessed by laser-scanning confocal microscopy of Fluo-4-AM-loaded MG63 cells. LL-37 increased Ca²⁺ also in the presence of the respective L- and T-type voltage-sensitive Ca²⁺ channel blockers nifedipine and NiCl₂. LL-37 had no effect on Ca²⁺ in cells incubated with Ca²⁺-free solution. LL-37 (4 and 8 µ<smlcap>M</smlcap>) reduced the MG63 cell number both in the presence and absence of Ca²⁺ in the medium. In conclusion, LL-37 reduces the osteoblast cell number by promoting apoptosis, and furthermore, LL-37 stimulates Ca²⁺ inflow via a mechanism independent of voltage-sensitive Ca²⁺ channels. Interestingly, LL-37-induced lowering of the cell number seems to be mediated via a mechanism independent of Ca²⁺.

Differential regulation of chemokine expression by estrogen in human periodontal ligament cells

BACKGROUND AND OBJECTIVE Estrogen modulates inflammatory responses, but the mechanisms involved have not yet been identified. Periodontal ligament (PDL) cells produce chemokines (a group of chemoattractant molecules that recruit leukocytes) and it has been suggested that estrogen modulates periodontal inflammation by regulating the expression of chemokines by PDL cells. Therefore, the objectives of this study were to investigate the regulation of chemokine ligand 2 [CCL2/monocyte chemoattractant protein 1 (MCP-1)], chemokine ligand 3 [CCL3/macrophage inflammatory protein-1α (MIP-1α)] and chemokine ligand 5 (CCL5/RANTES) by estrogen in human PDL cells. MATERIAL AND METHODS PDL cells were obtained from the PDL of premolars, extracted for orthodontic reasons, from two boys and two girls (16 and 17 years of age). PDL cell CCL2, CCL3 and CCL5 mRNA transcripts were determined by quantitative real-time PCR. The concentrations of CCL2, CCL3 and CCL5 proteins were determined by ELISAs. RESULTS Treatment with 0.5 μg/mL of lipopolysaccharide (LPS, from Escherichia coli) + 100 nm 17β-estradiol (E(2) ) for 24 h reduced the expression of CCL3 mRNA by about 40% compared to PDL cells treated with LPS alone. Attenuation of CCL3 mRNA was not associated with a decrease in CCL3 protein within 48 h, suggesting a slow turnover of the CCL3 protein. Interindividual differences in the effects of E(2) on CCL5 mRNA expression were observed. E(2) (100 nm) increased the expression of CCL5 by 40-60% in PDL cells derived from two subjects but reduced the expression of CCL5 by about 30% in cells from another subject. CCL2 mRNA and CCL2 protein were highly expressed, but not regulated by E(2) . Similar data were observed in cells obtained from both boys and girls. CONCLUSION Regulation, by estrogen, of chemokine expression in PDL cells shows a complex pattern involving the down-regulation as well as the up-regulation of chemokines, suggesting that estrogen exerts both anti-inflammatory and proinflammatory effects through these mechanisms.

The antimicrobial peptide LL‐37 is anti‐inflammatory and proapoptotic in human periodontal ligament cells

BACKGROUND AND OBJECTIVE The antimicrobial peptide LL-37 is expressed in periodontal tissue, and variations in LL-37 levels have been associated with periodontal disease. The effects of LL-37 on periodontal ligament cell function have not been described before. Here, we assess anti-inflammatory properties of LL-37 and investigate the effects of LL-37 on cell differentiation, cell proliferation and apoptosis in human periodontal ligament cells. MATERIAL AND METHODS Periodontal ligament cells were obtained from teeth extracted for orthodontic reasons. Cytokine (interleukin-6) and chemokine (monocyte chemoattractant protein-1) expression was determined by quantitative PCR, cell differentiation by alkaline phosphatase activity, cell proliferation by counting cells in a Bürker chamber, DNA synthesis by incorporation of radiolabeled thymidine and apoptosis by cell morphology and activated caspase 3 quantities. RESULTS Treatment with 0.1 and 1 μm of LL-37 totally reversed lipopolysaccharide-induced monocyte chemoattractant protein-1 expression and suppressed lipopolysaccharide-induced interleukin-6 expression by 50-70%. LL-37 had no effect on alkaline phosphatase activity. Incubation with 8 μm LL-37 strongly reduced cell number. DNA synthesis was attenuated by about 90% in response to 8 μm LL-37, confirming its antiproliferative effect. Cell morphology was altered in an apoptosis-like fashion in cells treated with 8 μm LL-37. Furthermore, the quantity of activated caspase 3 was increased in cells treated with 1 and 8 μm of LL-37, suggesting apoptosis. CONCLUSION LL-37 strongly attenuates lipopolysaccharide-induced cytokine and chemokine expression and, in high concentrations, reduces cell proliferation through inhibition of DNA synthesis and by promoting apoptosis in human periodontal ligament cells.

Secretory leukocyte protease inhibitor regulates human periodontal ligament cell production of pro-inflammatory cytokines

ObjectiveRegulation of immune-like cell properties of periodontal ligament (PDL) cells is not understood. We investigate the importance of secretory leukocyte protease inhibitor (SLPI) for production of pro-inflammatory cytokines in human PDL cells.Materials and methodsPDL cells were isolated from teeth extracted for orthodontic reasons. Cellular location of SLPI was investigated by immunocytochemistry. Cytokine transcript and protein expression were assessed by quantitative real-time RT-PCR and Western blotting. SLPI gene activity was knocked-down by siRNA. NF-κB signaling was assessed by measuring IκBα, and phosphorylated p65 and p105 protein expression.ResultsPDL cells showed cytoplasmic expression of SLPI. Cellular expression level of SLPI negatively correlated to LPS-induced stimulation of IL-6 and MCP-1. Both SLPI gene activity and protein were reduced by about 70% in PDL cells treated with SLPI siRNA compared to cells treated with non-coding construct. Treatment with SLPI siRNA was associated with up-regulation of both basal and LPS-stimulated IL-6, MCP-1 and TLRs mRNA expression. The up-regulation of MCP-1 transcript in SLPI siRNA-treated cells was confirmed on protein level. SLPI siRNA-treatment enhanced the phosphorylated NF-κB p105 protein expression.ConclusionsSLPI regulates PDL cell pro-inflammatory cytokine expression and modulates NF-κB signaling, suggesting that SLPI governs the immune cell-like properties of PDL cells.

Antimicrobial Peptides: Roles in Periodontal Health and Disease

Why do cockroaches and rats survive in sewers and other extremely challenging conditions? The answer is that through evolution they have been equipped with antimicrobial peptides (AMPs) that protect them from microbiological insult. In the case of cockroaches and rats, the AMPs are very potent, allowing for these challenging environments [1–3]. AMPs are the most ancient and primitive arm of the human immune system and are expressed in mammals, insects, fungus, trees—virtually every multicellular organism that coexist with bacteria, including bacteria. AMPs cover the outer barriers of our body, such as epithelium and skin, enabling us to live in coexistence with what some consider a complex organ—the microbiome [4]. As we now, through the technological advancements in microbiology, start to comprehend the complexity of the microbiome, we can also appreciate the complexity of the AMP-profile.