Candela Reyes-Botella

Antigenic Phenotype of Cultured Human Osteoblast-Like Cells

Background/Aims: Osteoblasts are classically considered to play an important role during bone tissue development, and to be involved in the formation of mineralized bone matrix. Recent reports have suggested that they can also exert some activities directly associated with the immune system (cytokine synthesis and antigen presentation). Moreover, some authors have found antigens on osteoblast-like cells normally expressed by other cells with a common origin in bone marrow.Methods: We isolated and cultured human osteoblast-like lines and studied their antigenic phenotype with flow cytometry using monoclonal antibodies against antigens associated with hematopoietic cells.Results: Cultured cells expressed CD34, but were negative for CD45. B cell antigens CD20 and CD23 and myelomonocytic antigens CD11b, CD13, and CD16 were detected. Expression of CD3, CD14, CD15 and CD68 was negative, whereas CD25 expression was positive. CD56, an antigen expressed on NK cells, was positive. These cells were CD10, CD44, CD54, CD80, CD86 and HLA-DR positive, as previously described. An antigen specific to follicular dendritic cells was also observed on cultured osteoblast-like cells.Conclusions: The antigenic phenotypes of human osteoblast-like cells and FDC are similar. These data suggest that osteoblasts may be functionally related to certain dendritic cells and may play an additional role in bone tissue to that classically assigned.

Expression of cytokines IL-4, IL-12, IL-15, IL-18, and IFNγ and modulation by different growth factors in cultured human osteoblast-like cells

The antigenic phenotype of cultured human osteoblast-like cells, their ability to phagocytose particles of different nature and size, and their capacity to stimulate allogeneic T cells suggest that they are related to other cell populations with which they may also have immunological functions in common. The objective of this study was to investigate the intracytoplasmatic presence of cytokines and their modulation by different biomolecules. Immunocytochemistry and flow cytometry were used to study the expression of IL-4, IL-12, IL-15, IL-18, and IFNγ cytokines. To investigate whether FGF, TGF, PDGF, IL-1, and IFNγ modulate expression of these cytokines in cultured human osteoblast-like cells we used flow cytometry. IL-4, IL-12, IL-15, IL-18, and IFNγ cytokines were expressed by all the cultured human osteoblast-like cells studied. Treatment with FGF and TGFβ1 reduced the percentage expression and fluorescence intensity of the cytokines. PDGF treatment enhanced their fluorescence intensity but did not modify their expression. IL-1 treatment produced a small reduction in expression and fluorescence intensity of IL-12 and IL-15, but did not produce major changes in the expression of IL-4, IL-18, or IFNγ. IFNγ markedly increased the fluorescence intensity of the cytokines. The results indicate that human osteoblast-like cells may perform immunological functions (e.g., synthesizing cytokines with immune regulator function) that can be modulated by different biomolecules related to bone tissue and/or immune response.

Phagocytosis and Allogeneic T Cell Stimulation by Cultured Human Osteoblast-Like Cells

Background/Aims: The antigen phenotype of human osteoblast-like cells suggests that they are related to other cellular populations and may also have immunologic functions in common. Methods: Flow cytometry and transmission electron microscopy were used to show the phagocytotic activity of osteoblast-like cells in culture. The allogeneic stimulation of T cells by human osteoblast-like cells was determined by the measurement of T cell proliferation. Results: We demonstrated in vitro that human osteoblast-like cells isolated from normal bone specimens obtained during mandibular osteotomy can phagocytose particles of different nature and size and can stimulate allogeneic T cells. Phagocytosis of microorganisms (E.coli, Klebsiella or C. albicans) was observed, although at a very low rate of activity in comparison with the phagocytosis of latex particles. Conclusion: Our results suggest that human osteoblast-like cells may perform immunologic functions and act as antigen presentation cells.p>

Effect of Platelet-Rich Plasma on Growth and Antigenic Profile of Human Osteoblasts and Its Clinical Impact

PURPOSE In recent years, there has been widespread clinical use of platelet-rich plasma (PRP) to facilitate the regeneration of different tissues. However, few data are available on the effect of PRP on parameters other than cell growth. The aim of the present study was to evaluate the effect of PRP on the cell cycle, antigenic profile, and proliferation of primary cultured human osteoblasts. MATERIALS AND METHODS The cells in the present study were derived from human bone sections obtained from healthy volunteers during third molar surgery. PRP was prepared from human venous blood and used to culture the cell line obtained from the same patient. Flow cytometry was used to study the cell cycle, antigenic profile, and proliferation. RESULTS The treatment of osteoblasts with PRP modified the expression of CD54, CD80, CD86, and HLA-DR antigens. PRP treatment increased cell proliferation in the short term, but the cell proliferation capacity diminished in the long term, perhaps owing to cell exhaustion. No change in the cell cycle profile was observed in the PRP-cultured cells. CONCLUSIONS These results suggest that PRP treatment accelerates bone neoformation with no cell cycle changes that might carry a risk of malignant transformation.

Effect of Different Growth Factors on Human Cultured Osteoblast-Like Cells

Background/Aims: Proliferation and differentiation of osteoblast-like cells are regulated by complex interactions among systemic hormones, cytokines, and local growth factors. The success of oral rehabilitation using biomaterial implants depends on the growth of osteoblasts and their adhesivity to the surface of the implant. The study aimed to investigate the effect of different growth factors on the proliferation and adhesivity of human osteoblast-like cells in vitro.Methods: The effects of transforming growth factor β1 (TGFβ1), fibroblast growth factor (FGF), and interleukin-2 (IL-2) on the growth and adhesivity of human osteoblast-like cells in monolayer culture were studied. Their growth was measured by 3H-thymidine incorporation and their adhesivity by flow cytometry.Results: Incorporation of 3H-thymidine was increased by all growth factors. The effect did not appear to be dose dependent. No synergic effect was found between TGFβ1 and FGF. Treatment with TGFβ1 or FGF increased cell adhesivity by shortening the time needed for cell adhesion to the culture dish surface and hydroxyapatite-coated implants. IL-2 did not modify cell adhesivity.Conclusions: Growth factors can modulate cell proliferation and adhesivity, which may help to increase the success of implantation therapy.

Factors Affecting Dental Implant Stability Measured Using the Ostell Mentor Device: A Systematic Review.

Objectives:The aim of this study was to review the literature on factors that may affect dental implant stability as measured with the Ostell mentor device. Materials and Methods:A systematic search of the literature was performed in Pubmed, Scopus, and Cochrane databases using dental implants, stability, and resonance frequency analysis as key words. Results:The most relevant randomized controlled trials and clinical trials (n = 39) were selected from among 264 articles. Conclusions:Many factors can affect dental implant stability as measured with the Ostell mentor device. This may be a useful instrument for deciding the timing of implant loading, but additional research is required to establish the reliability and predictability of resonance frequency analysis for the future osseointegration of dental implants, which remains controversial.

Long-Term Survival of Dental Implants with Different Prosthetic Loading Times in Healthy Patients: A 5-Year Retrospective Clinical Study

PURPOSE To compare survival rates among dental implants restored with immediate, early, and conventional loading protocols, also comparing between maxillary and mandibular implants, and to evaluate the influence of implant length and diameter and the type of prosthesis on treatment outcomes. MATERIALS AND METHODS This retrospective cohort study initially included all 52 patients receiving dental implants between July 2006 and February 2008 at a private oral surgery clinic in Granada (Southern Spain). Clinical and radiographic examinations were performed, including periapical or panoramic radiographs, and incidences during completion of the restoration were recorded at 1 week, 3 months, 6 months, and at 1, 2, 3, 4, and 5 years. After a 5-year follow-up, 1 patient had died, 3 were lost to follow-up, and 6 required grafting before implant placement; therefore, the final study sample comprised 42 patients with 164 implants. RESULTS Variables associated with the survival/failure of the restoration were: number of implants (higher failure rate with fewer implants), bone type (higher failure rate in type III or IV bone), and type of prosthesis (higher failure rate with single crowns). No significant association was found in univariate or multivariate analyses between survival rate and the loading protocol, implant length or diameter, or maxillary/mandibular location. CONCLUSIONS Immediate occlusal loading, immediate provisionalization without occlusal loading, and early loading are viable treatment options with similar survival rates to those obtained with conventional loading. Bone quality and number of implants per patient were the most influential factors.

Bacterial contamination levels of autogenous bone particles collected by 3 different techniques for harvesting intraoral bone grafts.

PURPOSE The aim of this study was to compare levels of bacterial contamination of autogenous bone collected when using low-speed drilling, a back-action chisel, and a bone filter. MATERIALS AND METHODS Bone tissue samples were taken from 31 patients who underwent surgical extraction of their third lower molars. Before surgical removal of the molar, bone particles were collected by a low-speed drill or a back-action chisel. Then, a stringent aspiration protocol was applied during the ostectomy to collect particulate bone by a bone filter. Processing of samples commenced immediately by incubation in an anaerobic or a CO2-rich atmosphere. The number of colony-forming units (CFUs) was determined at 48 hours of culture. RESULTS No significant difference in the number of CFUs per milliliter was observed between the low-speed drilling group and the back-action chisel group in the anaerobic or CO2-rich condition (P = .34). However, significantly more micro-organisms were found in the bone filter group than in the low-speed drilling group or the back-action chisel group in the anaerobic and CO2-rich conditions (P < .001). CONCLUSIONS Particulate bone harvested with low-speed drilling or a back-action chisel is safer for use as an autograft than are bone particles collected with a bone filter. These results suggest that bone obtained from low-speed drilling is safe and straightforward to harvest and could be the method of choice for collecting particulate bone. Further research is needed to lower the bacterial contamination levels of autogenous bone particles used as graft material.

Aminopeptidase Activity Profile in Cultured Human Osteoblasts

Aminopeptidases (APs) are enzymes involved in a wide variety of biological processes and present in a variety of different cell populations. The authors studied these enzymes in primary cultured human osteoblasts in order to establish an activity profile and thereby contribute to knowledge of bone tissue. The authors used 13 different substrates (N-terminal amino acids) and a fluorimetric assay to examine AP activity associated with the membranes of cultured osteoblasts. The authors demonstrated activity > 10 pmol/min/104 cells when glycine, alanine, leucine, arginine, phenylalanine, methionine, and lysine were used as substrates. The activity was markedly lower (<1.6 pmol/min/104 cells) when the other N-terminal amino acids were used. Puromycin and bestatin inhibited AP activity, though not completely, when we used AlaNA or LeuNA as substrates. Further studies are warranted to determine the role of these enzymes in bone tissue physiology.

Modulation of antigenic phenotype by IL-1β, IFNγ and TGFβ1 on cultured human decidual stromal cells

Decidual stromal cells (DSC) constitute the most abundant population in normal human decidua together with leukocytes. Both populations may be involved in the immunological role of the decidua by favoring gestational functions, participating in physiological mechanisms to eliminate the fetus, or providing local defense against infection. Using flow cytometry, we investigated whether different cytokines modulate the expression on cultured DSC of antigen-presenting molecules. The treatment with IFNγ or IL-1β enhanced the expression of CD54. The percentage of expression of HLA-DR was enhanced by IL-1β treatment but was not modified by IFNγ. The expression of CD80 and CD86 was enhanced by IFNγ treatment but was not modified by IL-1β; the expression of CD86 and HLA-DR was reduced by TGFβ1 treatment. The response of DSC and dendritic cells to these cytokines appears to be similar, suggesting a phenotypic and functional relationship between these cell types.