Publications by authors named "Elizabeth Ferreira Martinez"

57 Publications

Physical characterization of biphasic bioceramic materials with different granulation sizes and their influence on bone repair and inflammation in rat calvaria.

Sci Rep 2021 Feb 24;11(1):4484. Epub 2021 Feb 24.

Division of Cell Biology and Oral Pathology, São Leopoldo Mandic Research Institute, Campinas, SP, 13045-755, Brazil.

Biphasic calcium phosphate bioceramics (BCP) consist of a mixture of hydroxyapatite (HA) and beta-tricalcium phosphate (β-TCP) within the same particle. Due to their osteoconductive properties, biocompatibility and resemblance to natural bone, these materials have become a promising and suitable alternative to autologous bone grafting. First, the topography characteristics, specific surface area, and total pore volume of BCP were evaluated using scanning electron microscopy and the BET and BJH methods. Next, this study aimed to evaluate the intensity of the inflammatory process and the bone neoformation capacity of various particle sizes of BCP in the repair of critical defects in the calvaria of rats. A xenogeneic biomaterial was used in the control group. After 30, 60, and 90 days, the animals were euthanized, followed by the processing of the samples to measure the intensity of inflammatory infiltrates and the areas of bone neoformation. Our results indicate that no considerable differences were observed in the inflammatory scores in sites treated with distinct BCP grain sizes. A greater area of bone neoformation was measured in the xenogeneic group at all analysis times, with no substantial differences in bone formation between the BCP particle size in the range of 250-500 µm and 500-1000 µm.
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http://dx.doi.org/10.1038/s41598-021-84033-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7904940PMC
February 2021

Effect of Hyaluronic Acid and Poly-L-Lactic Acid Dermal Fillers on Collagen Synthesis: An in vitro and in vivo Study.

Clin Cosmet Investig Dermatol 2020 29;13:701-710. Epub 2020 Sep 29.

Division of Cell Biology and Oral Pathology, Faculdade São Leopoldo Mandic, Campinas, São Paulo, Brazil.

Purpose: Skin ageing is marked by structural and functional changes in epidermis and dermis, which result clinically in wrinkles, loss of elasticity, and rough-textured appearance. In this context, different dermal fillers have been used to overcome these negative effects associated with skin ageing, such as hyaluronic acid (HA) and poly-L-lactic acid (PLLA). Despite their low immunogenicity, these materials can cause an inflammatory reaction after application.

Materials And Methods: Considering high demand of HA and PLLA as filler material, this study aimed to evaluate their in vitro and in vivo effects. For the in vitro study, human dermal fibroblast cell cultures were supplemented with HA or PLLA for 24, 48, and 72 h. The following parameters were assayed: 1) cell proliferation, 2) cell viability, and 3) quantification of type I collagen by ELISA. For the in vivo study, HA or PLLA was injected in the dermis of Wistar rats and the tissues were collected after 15, 30, and 60 days for histologic evaluation and for quantification of type I collagen by Western blotting. The quantitative data were statistically analyzed using an ANOVA two-way. The significance level was set at 5%.

Results: At 72 h, high cell proliferation was observed for HA compared to control (p<0.05). Cultures exposed to PLLA exhibited a reduction in both cell proliferation and viability compared to control in all time points (p<0.05). Type I collagen expression was greater in cultures exposed to HA or PLLA compared to control (p<0.05). Histologic analysis showed the presence of multinucleated cells only in the PLLA group in all experimental time points. Western blotting analysis revealed high content of type I collagen in HA compared to PLLA (p<0.05).

Conclusion: The present study addresses a potentially unfavorable effect of dermal PLLA filler on the fibroblast phenotype, with possible clinical complications, unlike HA.
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http://dx.doi.org/10.2147/CCID.S266015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7533910PMC
September 2020

Secreted Osteoclastogenic Factor of Activated T Cells (SOFAT) Is Associated With Rheumatoid Arthritis and Joint Pain: Initial Evidences of a New Pathway.

Front Immunol 2020 28;11:1442. Epub 2020 Jul 28.

Laboratoy of Neuroimmune Interface of Pain Research, Faculdade São Leopoldo Mandic, Instituto de Pesquisas São Leopoldo Mandic, Campinas, Brazil.

Rheumatoid arthritis (RA) has an inflammatory milieu in the synovial compartment, which is regulated by a complex cytokine and chemokine network that induces continuously degenerative and inflammatory reactions. The secreted osteoclastogenic factor of activated T cells (SOFAT) is a unique cytokine and represents an alternative pathway for osteoclast activation. In this study, we examined whether SOFAT is able to induce joint pain and investigated the presence of SOFAT in a Collagen-induced Arthritis (CIA) model and in human subjects. Here, we found that an intra-articular stimulation with SOFAT (1, 10, 100, or 1,000 ng/10 μl) in the knee joint significantly decreases the mechanical threshold in the hind paw of mice ( < 0.05). Moreover, after a second injection of SOFAT, the mechanical threshold decrease was sustained for up to 8 days ( < 0.05). In the CIA model, the immunohistochemical assay of knee joint showed positivity stained for SOFAT, and the mRNA and protein expression of SOFAT were significantly higher in the affected-group ( < 0.05). Besides, the mRNA of RANKL, IL-1β, IL-6, and IL-15 were significantly higher in the affected-group ( < 0.05). Finally, SOFAT was detected in the synovial fluid of RA patients, but not in OA patients ( < 0.05). In conclusion, SOFAT is up regulated in inflammatory milieu such as RA but not in non-inflammatory OA. SOFAT may be a novel molecule in the complex inflammatory phenotype of RA.
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http://dx.doi.org/10.3389/fimmu.2020.01442DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7399082PMC
July 2020

Inflammatory cell profile using different autologous fibrin protocols.

Tissue Cell 2020 Dec 10;67:101407. Epub 2020 Jul 10.

Faculdade São Leopoldo Mandic, Instituto de Pesquisas São Leopoldo Mandic, Neuroimmune Interface of Pain Research Lab, Campinas, SP, Brazil. Electronic address:

Autologous fibrin has been widely used in surgical procedures for both soft and hard tissue repair. There are different protocols and devices to obtain this matrix, with varying centrifugal time, gravity force, speed, angle of the sample tube and spinning radius. The aim of this study was to compare three methods of obtaining autologous fibrin: L-PRF using the Intra-Spin L-PRF centrifuge (Dohan protocol), the advanced PRF (A-PRF) using the Intra-Spin L-PRF centrifuge and autologous leukocyte fibrin (ALF), using the Kasvi centrifuge. Venous blood was collected from 7 healthy volunteers, which were submitted to the 3 different methods of centrifugation. The membranes were tissue-processed and evaluated by immunohistochemistry for CD3, CD20, CD68 and CD138. For CD68+, a lower number of cells was immunolabelled in the L-PRF group when compared to the other groups (A-PRF and ALF). For CD3+, a lower number of immunolabellated cells was observed in the ALF group when compared to the remaining groups (p < 0.05). In the A-PRF group, the CD20+ cell count was lower than in the remaining groups. No difference was observed in CD138+ cell counts between the groups. The 3 protocols tested are suitable for obtaining autologous fibrin membranes.
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http://dx.doi.org/10.1016/j.tice.2020.101407DOI Listing
December 2020

Topographic characterization and biofilm adhesion to titanium and polypropylene membranes used for alveolar preservation.

J Indian Soc Periodontol 2020 Jul-Aug;24(4):316-321. Epub 2020 Jul 1.

Division of Oral Pathology and Cell Biology, São Leopoldo Mandic Research Institute, Campinas, São Paulo, Brazil.

Background: Nonresorbable membranes have been widely used in guided bone regeneration (GBR) procedures in posttooth extraction alveoli. In this context, one of the properties suggested by the GBR technique is that these barriers, when exposed to the oral environment, control or prevent the infiltration of connective and epithelial tissue cells, favoring the proliferation of bone cells inside the alveolus, without the growth of biofilm.

Materials And Methods: This study evaluated the topographic characteristics and biofilm adhesion on membranes used for alveolar preservation, bone Heal™ and Titanium Seal™. Fragments of these membranes (5 mm × 5 mm) were used for all experiments. The topographical morphology and chemical characterization of the membranes were analyzed by scanning electron microscope and dispersive energy X-ray spectroscopy, respectively. For the biofilm adhesion assay, samples were immersed in (American Type Culture Collection [ATCC] 10231) and (ATCC 25923) mixed biofilm for 7 and 14 days. Biofilm formation was measured by quantitative analysis with crystal violet aqueous solution, in a spectrophotometer, with a wavelength of 590 nm.

Results: The ultrastructural images showed a rough surface for the titanium membrane, without homogeneity in the surface structure, while the polypropylene membrane presented a smoother surface without depressions. The chemical composition of the membranes by Ehlers-Danlos syndrome has identified the presence of copolymer and traces of zinc for the polypropylene membrane; in contrast, the titanium membrane revealed the unique presence of titanium. In addition, there was a decrease in biofilm formation on the surface of the titanium membrane compared to polypropylene ( < 0.05), at both evaluated times.

Conclusions: It can be concluded that despite the greater heterogeneity of the titanium membrane surface, the results showed less biofilm formation on this membrane ( < 0.05), which may be indicated in cases of oral cavity exposure.
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http://dx.doi.org/10.4103/jisp.jisp_602_19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7418540PMC
July 2020

DC-STAMP and TACE Levels are Higher in Patients with Periodontitis.

Braz Dent J 2020 Mar-Apr;31(2):122-126

Laboratory of Neuroimmune Interface of Pain Research Instituto de Pesquisas São Leopoldo Mandic, Faculdade São Leopoldo Mandic, Campinas, SP, Brazil.

Although periodontitis is one of the commonest infectious inflammatory diseases in humans, the mechanisms involved with its immunopathology remain ill understood. Numerous molecules may induce inflammation and lead to bone resorption, secondary to activation of monocytes into osteoclasts. TACE (TNF-α converting enzyme) and DC-STAMP (dendritic cell-specific transmembrane protein) appear to play a role on bone resorption since TACE induces the release of sRANKL (soluble receptor activator of nuclear factor kappa-β ligand) whereas DC-STAMP is a key factor in osteoclast induction. The present study evaluated the levels of TACE and DC-STAMP in patients with and without periodontitis. Twenty individuals were selected: 10 periodontally healthy participants undergoing gingivectomy for esthetic reasons and 10 diagnosed with periodontitis. Protein levels of such molecules in gingival tissue were established using Western blotting. Protein levels of both TACE and DC-STAMP were higher in the periodontitis group than in the control group (p<0.05; Student t-test). In conclusion, TACE and DC-STAMP protein levels are elevated in patients with periodontitis, favoring progression of bone resorption.
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http://dx.doi.org/10.1590/0103-6440202002939DOI Listing
June 2020

Microvesicles derived from squamous cell carcinoma induce cell death, autophagy, and invasion of benign myoepithelial cells.

J Oral Pathol Med 2020 Sep 11;49(8):761-770. Epub 2020 Jun 11.

Cell Biology and Oral Pathology Division, Faculdade São Leopoldo Mandic, Campinas, SP, Brazil.

Background: There has been great interest recently in the mechanisms of cell-to-cell communication through microvesicles (MV). These structures are produced by many different cell types and can modulate cellular activity by induction of epigenetic alterations. These vesicles may promote tumor mass increase either by stimulating cell proliferation via growth factors or by inhibiting apoptosis, which reinforces the role of such vesicles as important modulators of tumor progression.

Methods: The present in vitro study aimed to characterize MV derived from malignant neoplastic epithelial cell cultures (EP) and their effect on the expression of apoptosis/autophagy and invasion related genes of benign myoepithelial (Myo) cell cultures.

Results: The results revealed round structures with a mean size of 153.6 (±0.2) nm, with typical MV morphology. CD63 quantification indicated that EP cell culture at 70%-80% confluence secreted 3.088 × 10 MV/mL. Overall, Myo exposed to MVs derived from EP showed both up- and downregulation of tumorigenesis promoting genes. MVs from EP cells promoted cell death of Myo cells and positively modulate BAX, SURVIVIN, LC3B, MMP-2, and MMP-9 expression. Furthermore, an increasing of MMP-2 and MMP-9 secretion by Myo was observed after MV exposure.

Conclusions: These findings suggest that MVs from EP modulate autophagy of Myo cells, which may, in part, explain the disappearance of these cells in in situ areas of invasive carcinoma ex-pleomorphic adenoma. Additionally, the overexpression of MMPs contributes to the development of an invasive phenotype of Myo cells, which could favor the dissolution of the basement membrane during tumorigenesis process.
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http://dx.doi.org/10.1111/jop.13037DOI Listing
September 2020

Long-term Evaluation of Dentin Matrix Stability and Gelatinolytic Activity after Dentin Pretreatment with Caffeic Acid Phenethyl Ester.

J Adhes Dent 2020 ;22(3):285-296

Purpose: To investigate the long-term effect of 0.05% or 0.1% caffeic acid phenethyl ester (CAPE) on dentin matrix stability and hybrid layer stability, using an etch-and-rinse (Adper Scotchbond Multipurpose/ASB) or a self-etch adhesive (Clearfil SE Bond/CSE).

Materials And Methods: Dentin matrix specimens were assigned to five groups: 0.05% or 0.1% CAPE, green tea (GT), and the controls distilled water (DW) and dimethyl sulfoxide (DMSO). Following immersion of specimens for 1 h, modulus of elasticity (ME) and dentin mass change (MG) were determined at 3 post-treatment time points: immediately afterwards and at 3 and 6 months. Collagen solubilization (CS) was estimated by hydroxyproline (HYP) quantification. Resin-dentin interfaces with both adhesives were assessed with in situ zymography tests to evaluate gelatinolytic activity (GA). The dentin pretreatments were actively applied for 60 s. The sealing ability of aged resin-bonded slices was assessed by nanoleakage tests.

Results: GT increased immediate ME, which decreased significantly after 3 months (p < 0.0001). The CAPE groups did not differ from the control groups. GT provided a significant increase in dentin matrix mass after treatment (p < 0.0001). No significant differences regarding MG were observed for CAPE 0.1%, CAPE 0.05%, DW, and DMSO groups after 3 and 6 months. Cumulative HYP release revealed that CAPE groups and GT were statistically similar to DW and DMSO; the GT group exhibited statistically significantly less HYP release than did CAPE groups (p = 0.0073). Treatment with 0.05% or 0.1% CAPE presented lower GA when applied to ASB before acid conditioning (p < 0.05), but no differences were detected when the CAPE groups were applied to CSE. CAPE at 0.1% significantly reduced nanoleakage for CSE, and 0.05% CAPE with CSE presented levels of nanoleakage similar to those of the CSE control group.

Conclusion: CAPE at 0.05% or 0.01% did not influence ME, MG, or CS, but reduced GA when applied to ASB before acid conditioning. CAPE at 0.1% with CSE promoted adhesive layer integrity.
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http://dx.doi.org/10.3290/j.jad.a44552DOI Listing
May 2020

Preliminary findings on the possible role of B-lymphocyte stimulator (BLyS) on diabetes-related periodontitis.

Braz Oral Res 2020 30;34:e038. Epub 2020 Apr 30.

Faculdade São Leopoldo Mandic , Instituto de Pesquisas São Leopoldo Mandic , Campinas , SP , Brazil .

The possible role of B-cell growth and differentiation-related cytokines on the pathogenesis of diabetes-related periodontitis has not been addressed so far. The aim of this study was to evaluate the effects of diabetes mellitus (DM) on the gene expression of proliferation-inducing ligand (APRIL) and B-lymphocyte stimulator (BLyS), two major cytokines associated to survival, differentiation and maturation of B cells in biopsies from gingival tissue with periodontitis. Gingival biopsies were obtained from subjects with periodontitis (n = 17), with periodontitis and DM (n = 19) as well as from periodontally and systemically healthy controls (n = 10). Gene expressions for APRIL, BLyS, RANKL, OPG, TRAP and DC-STAMP were evaluated using qPCR. The expressions APRIL, BLyS, RANKL, OPG, TRAP and DC-STAMP were all higher in both periodontitis groups when compared to the control group (p < 0.05). Furthermore, the expressions of BLyS, TRAP and RANKL were significantly higher in the subjects with periodontitis and DM when compared to those with periodontitis alone (p < 0.05). The mRNA levels of BLyS correlated positively with RANKL in the subjects with periodontitis and DM (p < 0.05). BLyS is overexpressed in periodontitis tissues of subjects with type 2 DM, suggesting a possible role of this cytokine on the pathogenesis DM-related periodontitis.
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http://dx.doi.org/10.1590/1807-3107bor-2020.vol34.0038DOI Listing
May 2020

Effect of sodium ascorbyl phosphate on osteoblast viability and differentiation.

J Periodontal Res 2020 Oct 22;55(5):660-666. Epub 2020 Apr 22.

Department of Pathology, São Leopoldo Mandic Institute and Research Center, Campinas, Brazil.

Objectives And Background: Sodium ascorbyl phosphate (SAP) is a hydrophilic and stable L-ascorbic acid derivative, being converted by the cell phosphatases into free ascorbic acid (AA), which allows its sustained release in the medium. AA participates in the maintenance and healing of the periodontium. It presents a regulatory role of the osteoblastic activity, stimulating the deposition of collagen extracellular matrix followed by the induction of genes associated with the osteoblastic phenotype. It also acts in the elimination of reactive oxygen species, abundantly produced by defense cells in periodontal disease. The aim of this study was to evaluate the effect of SAP on osteoblast viability and differentiation.

Methods: Mouse preosteoblastic cells of the MC3T3-E1 strain were used. Cell viability was assessed by the trypan blue dye exclusion assay and the expression of genes related to osteoblast differentiation by quantitative PCR. Collagen I secretion was evaluated by ELISA, and mineralized matrix formation was assayed by Alizarin red S staining.

Results: The results showed that SAP at concentrations from 50 to 500 µmol/L does not influence preosteoblast cell viability, but stimulates their differentiation, observed by the induction of RUNX2, COL1A1, and BGLAP2; by the higher secreted levels of collagen I; and also by the increase in the mineralization of the extracellular matrix in cells exposed to this agent at 200 or 400 µmol/L, compared with those not exposed.

Conclusion: By its stability and capacity to induce preosteoblastic cell differentiation, our results indicate that the incorporation of SAP into local release devices, membranes/scaffolds or biomaterials, could favor bone tissue formation and therefore periodontal healing.
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http://dx.doi.org/10.1111/jre.12752DOI Listing
October 2020

Lithium chloride improves bone filling around implants placed in estrogen-deficient rats.

Arch Oral Biol 2020 Mar 24;111:104644. Epub 2019 Dec 24.

Faculdade São Leopoldo Mandic, Instituto São Leopoldo Mandic, Laboratory of Neuroimmune Interface of Pain Research, Campinas, SP, Brazil. Electronic address:

Objective: This study evaluated the ability of lithium chloride (LiCl) to increase bone filling (BF) around threaded titanium implants inserted in estrogen-deficient rats and, thein-vitro effects of this drug on osteoblast-like cell viability, proliferation, mineralization and expression of bone-related markers.

Design: In vivo: Rats received sham surgery plus water (Estrogen-sufficient group), ovariectomy plus water (Estrogen-deficient group) or ovariectomy plus LiCl (150 mg/kg/every other day) (LiCl/estrogen-deficient group). On the 21 day after ovariectomy/sham surgeries, a threaded titanium implant was inserted in the rat tibia. BF and the number of TRAP + cells were assessed at 10, 20 and 30 days after implant placement. In vitro: Osteosarcoma SAOS-2 cells were exposed to 0, 0.01, 0.05, and 0.1 mM of LiCl; cell proliferation, viability, mineralization (alizarin red staining) and gene expressions of RUNX-2, OCN, OPN, BSP and ALP (Real Time PCR) were estimated in the cultures.

Results: In vivo: The estrogen-sufficient and LiCl/estrogen-deficient groups demonstrated higher percentages of BF, within the limits of implant threads, than the estrogen-deficient group at 20 and 30 days (p < 0.05). The number of TRAP + cells was lower in LiCl/estrogen-deficient than in the estrogen-deficient group at all experimental times (p < 0.05). In vitro: Cell cultures exposed to LiCl (0.01 or 0.05 mM) exhibited larger areas of mineralized matrix than the non-exposed cultures (p < 0.05) and demonstrated the highest expressions of the genes investigated.

Conclusion: LiCl treatment improved BF around threaded titanium implants inserted in estrogen-deficient rats and stimulated matrix mineralization and overexpression of bone-formation markers in osteoblastic cells in culture.
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http://dx.doi.org/10.1016/j.archoralbio.2019.104644DOI Listing
March 2020

Evaluation of toxicity and response to oxidative stress generated by orthodontic bands in human gingival fibroblasts.

Angle Orthod 2020 03 5;90(2):285-290. Epub 2019 Dec 5.

Objective: To evaluate the cytotoxicity of stainless-steel orthodontic bands and their influence on the expression of the antioxidant genes in human gingival fibroblasts.

Materials And Methods: Ten bands of each brand (Dentsply-Sirona, Dentaurum, TP Orthodontics, and Morelli) were conditioned in 0.2 g/mL culture medium at 37°C for 14 days, and the corresponding conditioned media were applied over the fibroblasts. Cell viability was assessed after 24, 48, and 72 hours of exposure to the conditioned media by trypan blue exclusion assay. Expression of the antioxidant defense genes peroxiredoxin 1 (), superoxide dismutase 1 (), and glutathione peroxidase 1 () were evaluated by quantitative polymerase chain reaction after 24 hours of exposure. These parameters were compared to those of the cells not exposed to the conditioned media of the bands (control).

Results: All bands promoted a reduction in the number of viable cells in the periods of 48 and 72 hours ( < .01). Analysis of gene expression showed a significant increase in the levels of transcripts caused by the conditioned media of the Dentsply-Sirona, TP Orthodontics, and Morelli bands ( < .01) as well as induction of by the conditioned media of the Dentaurum and Morelli ( < .01). Expression of was not influenced by the conditioned media.

Conclusions: The orthodontic bands showed toxicity to fibroblasts and increased the expression of and antioxidant genes, indicating induction of oxidative stress in the cells.
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http://dx.doi.org/10.2319/110717-761.1DOI Listing
March 2020

Viability and collagen secretion by fibroblasts on titanium surfaces with different acid-etching protocols.

Int J Implant Dent 2019 Nov 21;5(1):41. Epub 2019 Nov 21.

Division of Oral Biology and Cell Biology, Faculdade São Leopoldo Mandic, Rua José Rocha Junqueira, 13, Campinas, SP, 13045-755, Brazil.

Background: From the consolidation of surface treatments of dental implants and knowledge on the cellular mechanisms of osseointegration, studies have highlighted the importance of a connective tissue seal against the implant to prevent contamination from the oral environment and consequent biofilm formation.

Objective: This in vitro study aimed to evaluate whether different titanium surface treatments using acid solutions promoted an increase in collagen secretion, proliferation, and viability of fibroblasts.

Material And Methods: Commercially pure grade-4 titanium disks (6 × 2 mm) were treated with different acid solutions (hydrochloric, nitric, and sulfuric) for 20 and 60 min, respectively, obtaining mean surface roughness of 0.1 to 0.15 μm and 0.5 to 0.7 μm. Human fibroblasts were seeded onto different surfaces and assessed after 24 h, 48 h, and 72 h for cell proliferation and viability using Trypan blue staining and MTT, respectively, as well as the secretion of type I collagen on to such surfaces using ELISA. Machined titanium surfaces were used as controls. Data were statistically analyzed using one-way ANOVA and Fisher's LSD test for multiple comparisons, adopting a significance level of 5%.

Results: No significant difference was observed in cell proliferation for the different surfaces analyzed. Cell viability was significantly lower on the machined surface, after 48 h, when compared to the groups treated with acid for 20 or 60 min, which did not differ from each other. The expression of type I collagen was lowest on the acid-treated surfaces.

Conclusion: The results showed that the acid treatment proposed did not promote fibroblast proliferation and viability nor favor type I collagen synthesis.
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http://dx.doi.org/10.1186/s40729-019-0192-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6868076PMC
November 2019

Mechanical and histological evaluation of a titanium device for orthodontic anchorage, placed with or without cyanoacrylate adhesive.

Dental Press J Orthod 2019 Aug 1;24(3):71-78. Epub 2019 Aug 1.

Instituto e Centro de Pesquisas São Leopoldo Mandic (Campinas/SP, Brazil).

Objective: The objective of the present study was to perform a histological evaluation of a titanium mini-implant for orthodontic anchorage. Shear strength and fracture patterns that occurred immediately, 30 and 60 days after insertion with or without N-2-butyl-cyanoacrylate adhesive were evaluated.

Methods: Ninety-six mini-implants (Arrow, Peclab, Brazil) were placed in the tibia of 9 male rabbits, with or without an adhesive (Vetbond™, 3M, USA). Histological evaluation was done by optical light microscope. Shear strength testing was performed, followed by fracture analysis with visual inspection.

Results: Close contact between the newly formed bone and the device was evidenced in the group without adhesive, whereas gaps in the group with adhesive were found. Tukey test showed similar values in both groups at the immediate time point (20.70 N without adhesive and 24.69 N with adhesive), and higher values for the non-adhesive group, after 30 and 60 days (43.98 N and 78.55 N, respectively). The values for the adhesive group were similar for the immediate time point (24.69 N), 30 days (18.23 N) and 60 days (31.98 N). The fractures were adhesive for both groups at the immediate time point. The fractures were cohesive in bone for the non-adhesive group after 30 and 60 days.

Conclusions: The mini-implants showed close bone contact and required higher shear strength for removal at 30 and 60 days for the non-adhesive group. Further studies are needed to assess the proper way to remove the orthodontic anchorage without cohesive fractures in bone.
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http://dx.doi.org/10.1590/2177-6709.24.3.071-078.oarDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6677335PMC
August 2019

TiF4 Incorporated into a Self-etching Primer in Different Concentrations: Antimicrobial Properties and Effects on Demineralisation Inhibition Around the Restoration/Enamel-Dentin Interface.

Oral Health Prev Dent 2019;17(1):57-67

Purpose: To apply titanium tetrafluoride (TiF4) in an aqueous solution or incorporated into the primer of a self-etching adhesive (Clearfil SE Bond) as dentin pre-treatment and evaluate its antimicrobial effect, determine the minimum bactericidal concentraion (MBC) against Streptococcus mutans and Lactobacillus casei and analyse its potential to inhibit the development of carious lesions at the restoration interface.

Materials And Methods: For MBC, an aqueous solution or primer with different concentrations (in %) of TiF4 were used (from 0.0 to 4.0). Also, 50 cavities were prepared at the enamel/dentin junction of third molars and received the following dentin pre-treatments (n = 10): Clearfil SE Bond (CL); aqueous solution of 2.5% TiF4 + CL (T2.5%); aqueous solution of 4% TiF4 + CL (T4%); 2.5% TiF4 incorporated into the primer (P2.5%); 4% TiF4 incorporated into the primer (P4%). Cavities were restored and submitted to pH cycling to create artificial caries lesions. Microhardness tests were performed after sectioning the restorations to assess the demineralisation at margins.

Results: ANOVA and Tukey's tests showed that TiF4 in aqueous solution presented MBC against S. mutans and L. casei of over 2.0%. TiF4 in the primer of a self-etching adhesive presented MBC of over 1% for L. casei. For enamel, CL showed no significant differences in microhardness between the depths.

Conclusions: The aqueous solution had an antimicrobial effect against Streptococcus mutans and Lactobacillus casei of over 2.0%. Pretreatment with the aqueous solution or primer did not inhibit demineralisation at enamel or dentin restoration interfaces.
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http://dx.doi.org/10.3290/j.ohpd.a41978DOI Listing
November 2019

Comparison of p63/p40 Expression With Myoepithelial Markers in Minor Salivary Gland Tumors.

Int J Surg Pathol 2019 Jun 27;27(4):360-371. Epub 2018 Nov 27.

1 São Leopoldo Mandic Institute and Research Center, Campinas, São Paulo, Brazil.

The present study aimed to compare the expression of p63/p40 with smooth muscle actin (SMA) and vimentin (VIM) by myoepithelial cells in minor salivary gland tumors. Fifty-two formalin-fixed paraffin-embedded samples of minor salivary gland tumors derived from intercalated duct (pleomorphic adenoma [PA], adenoid cystic carcinoma [ACC], epithelial-myoepithelial carcinoma [EMC], polymorphous adenocarcinoma [PAC], and secretory carcinoma [SC]) and 3 samples of minor salivary gland tumors derived from excretory duct (mucoepidermoid carcinoma [MEC]) were evaluated by means of immunohistochemistry. The data were analyzed qualitatively. The results indicated that p63 and p40 expression were detected in myoepithelial cells present in PA, ACC, and EMC. However, both proteins were also observed in squamous areas of PA and all cases of MEC. SMA were noticed in some myoepithelial cells of PA, ACC, and EMC. Expression of SMA was negative in the other salivary gland tumors evaluated. VIM was constantly expressed by myoepithelial cells in PA, ACC, and EMC. VIM was also observed in cells of PAC and SC, but not in squamous areas of PA and MEC. In conclusion, p63 expression is almost comparable with VIM in detecting myoepithelial cells, an immunolabeling pattern not followed by p40, and consequently, caution has to be taken during the interpretation of salivary gland tumor exhibiting an p63/p40 phenotype in order to avoid a misdiagnosis.
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http://dx.doi.org/10.1177/1066896918813678DOI Listing
June 2019

Histologic Evaluation of Leucocyte- and Platelet-Rich Fibrin in the Inflammatory Process and Repair of Noncritical Bone Defects in the Calvaria of Rats.

Int J Oral Maxillofac Implants 2018 Nov/Dec;33(6):1206-1212

Purpose: This study aimed to evaluate the effects of leucocyte- and platelet-rich fibrin (L-PRF) on the inflammatory process, tissue repair, and expression of vascular endothelial growth factor (VEGF) on bone defects in the calvaria of rats.

Materials And Methods: L-PRF was obtained from three animals submitted to cardiac puncture to prepare the membranes. Two noncritical defects with a diameter of 2 mm were created in the calvaria of 15 Wistar rats. The defects on the right side were filled with a blood clot (CTRL) and the left side with L-PRF. After 5, 15, and 30 days, the animals were euthanized and the specimens processed for histologic, histomorphometric, and immunohistochemical analyses. In order to measure the intensity of the inflammatory infiltrate and VEGF expression, scores were assigned from 0 to 3, with 0 being no expression, 1 discrete (up to 25%), 2 moderate (between 25% and 50%), and 3 intense (> 50%) expression. The area of bone neoformation at the edges of the defects was also quantified.

Results: A less intense inflammatory infiltrate was observed in the defects filled with L-PRF compared with CTRL at all times analyzed (P < .05). At 5 days, no bone neoformation was observed in any of the groups evaluated. After 15 and 30 days, greater bone neoformation was observed in the group treated with L-PRF compared with the CTRL group (P < .05). At 15 days, 3,871.8 (1,070.15) μm were recorded for the CTRL and 49,978.5 (14,360.7) μm in the L-PRF. At 30 days, 62,284.5 (3,579.5) μm were observed in the CTRL and 154,076.6 (31,464.9) μm in the L-PRF. At all evaluated times, a lower inflammatory infiltrate was observed in the group treated with L-PRF compared with the CTRL. VEGF expression was observed in the initial phase and throughout the tissue repair process in both groups. At 5 days, there was no difference in VEGF expression between the groups. VEGF was present at the initial phase and throughout the tissue repair process in both groups. In the L-PRF group, a decrease in VEGF expression was observed at 15 and 30 days compared with the CTRL group.

Conclusion: L-PRF had a positive effect on the regenerative process of bony defects, with a reduced inflammatory response and greater bone neoformation.
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http://dx.doi.org/10.11607/jomi.6604DOI Listing
December 2018

Sustained Release from Ionic-Gradient Liposomes Significantly Decreases ETIDOCAINE Cytotoxicity.

Pharm Res 2018 Oct 10;35(12):229. Epub 2018 Oct 10.

Department of Biochemistry and Tissue Biology, Institute of Biology, University of Campinas - Unicamp, P.O. Box - 6109, CEP, Campinas, SP, 13083-862, Brazil.

Purpose: Etidocaine (EDC) is a long lasting local anesthetic, which alleged toxicity has restricted its clinical use. Liposomes can prolong the analgesia time and reduce the toxicity of local anesthetics. Ionic gradient liposomes (IGL) have been proposed to increase the upload and prolong the drug release, from liposomes.

Methods: First, a HPLC method for EDC quantification was validated. Then, large unilamellar vesicles composed of hydrogenated soy phosphatidylcholine:cholesterol with 250 mM (NH)SO - inside gradient - were prepared for the encapsulation of 0.5% EDC. Dynamic light scattering, nanotracking analysis, transmission electron microscopy and electron paramagnetic resonance were used to characterize: nanoparticles size, polydispersity, zeta potential, concentration, morphology and membrane fluidity. Release kinetics and in vitro cytotoxicity tests were also performed.

Results: IGL showed average diameters of 172.3 ± 2.6 nm, low PDI (0.12 ± 0.01), mean particle concentration of 6.3 ± 0.5 × 10/mL and negative zeta values (-10.2 ± 0.4 mV); parameters that remain stable during storage at 4°C. The formulation, with 40% encapsulation efficiency, induced the sustained release of EDC (ca. 24 h), while reducing its toxicity to human fibroblasts.

Conclusion: A novel formulation is proposed for etidocaine that promotes sustained release and reduces its cytotoxicity. IGL can come to be a tool to reintroduce etidocaine in clinical use.
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http://dx.doi.org/10.1007/s11095-018-2512-4DOI Listing
October 2018

Influence of Transmucosal Height on Loss of Prosthetic Abutment Torque After Mechanical Cycling.

J Oral Implantol 2018 Dec 16;44(6):423-426. Epub 2018 Jul 16.

2   Department of Cell Biology, São Leopoldo Mandic Institute and Research Center, Campinas, São Paulo, Brazil.

The aim of this study was to measure and record the universal transmucosal abutment height, and then evaluate whether it influenced loosening of the abutment screw by analyzing the torque and detorque values after mechanical cycling. Thirty-six implants, model CM Unitite, with internal conical connections (3.5 × 10 mm) and respective universal prosthetic abutments (n = 36, 3.25 × 6 mm), were divided into three groups (n = 12 each) with respective transmucosal heights of 0.8, 3.5, and 5.5 mm. Insertion torque of 20 Ncm was used in accordance with the manufacturer's specifications. Afterward, the samples were submitted to fatigue tests consisting of 500,000 cycles at a frequency of 2Hz, a dynamic compressive load of 120N, and an angle of 30°. The detorque values were measured with a digital torque meter and tabulated to perform statistical analyses; a level of significance of 5% was adopted. The mean detorque values (SD) obtained were 22.83 (6.30), 22.5 (5.45), and 19.41 (4.69) Ncm for transmucosal abutments with heights of 0.8, 3.5, and 5.5 mm, respectively, and showed no statistically significant difference ( P = .262). The authors of this study concluded that the transmucosal height of prosthetic abutments submitted to mechanical fatigue did not influence the detorque values.
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http://dx.doi.org/10.1563/aaid-joi-D-17-00250DOI Listing
December 2018

Effects of caffeic acid phenethyl ester application on dentin MMP-2, stability of bond strength and failure mode of total-etch and self-etch adhesive systems.

Arch Oral Biol 2018 Oct 11;94:16-26. Epub 2018 Jun 11.

São Leopoldo Mandic Institute and Dental Research Center, Rua José Rocha Junqueira 13, Bairro Swift, Campinas, CEP: 13045-755, São Paulo, Brazil. Electronic address:

Objective: Investigate the long-term effect of dentin pretreatment with 0.05 or 0.1% caffeic acid phenethyl ester (CAPE) on (1) bond strength of resin composite to dentin by a three-step etch-and-rinse (Adper Scotchbond Multipurpose/ ASB) or a two-step self-etch adhesive system (Clearfil SE Bond/ CSE), (2) their fracture mode, (3) the micromorphological features of the hybrid layer formed; and (4) the level of MMP-2 in dentin (after application, using a correlative immunoexpression/quantification approach).

Design: Composite resin blocks were fabricated on 48 third molars (n = 6), according to the type of adhesive and treatment (control, CAPE 0.05% and CAPE 0.1%). Slices were obtained for scanning electron microscopy (SEM) evaluation, and sticks were fabricated for microtensile tests (24 h and 1 year). Aliquots of dentin powder were distributed (n = 12) according to the treatment and the MMP-2 concentration was determined by ELISA.

Results: Tukey test showed that ASB groups presented higher BS in 24 h than CSE groups. ASB presented a reduction in BS values after 1-year. ASB and CSE presented no significant differences in BS after 1-year. CAPE had no effect on BS for both adhesive systems. The predominant failure mode for the ASB groups were adhesive; when 0.1% CAPE was applied there was a predominance of mixed fractures. Regarding the CSE group, 0.05% CAPE led to more adhesive failures, and the 0.1% concentration resulted in a higher number of cohesive failures in dentin. Higher MMP-2 concentrations were detected for the groups that did not undergo demineralization treatment, and the lowest values for the ASB groups treated with CAPE. SEM analysis showed no influence of pretreatment with CAPE.

Conclusions: CAPE did not influence the BS of the adhesives tested, or the micromorphology of the hybrid layer, irrespective of concentration or storage time. CAPE affected the fracture pattern at 24 h, depending on the concentration and the adhesive system used. Immunoassay analysis showed that CAPE 0.1% reduced the MMP-2 concentration in the ASB adhesive without affecting bond strength to dentin.
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http://dx.doi.org/10.1016/j.archoralbio.2018.06.012DOI Listing
October 2018

Caries experience and salivary aspects in individuals with fragile X syndrome.

Braz Oral Res 2017 Sep 28;31:e79. Epub 2017 Sep 28.

São Leopoldo Mandic Institute and Research Center, Departament of Oral Pathology, Campinas, SP, Brazil.

Fragile X syndrome (FXS) is the most common cause of hereditary mental retardation, but studies on the oral health condition of these patients are rare. The aim of this study was to determine the experience of dental caries in individuals with FXS, by examining the saliva profile, oral hygiene, socioeconomic characteristics and use of controlled drugs in these patients. Dental health was estimated using the decayed, missing and filled teeth index (DMF-T) and sialometry, and the pH value and buffering capacity of the saliva, colony forming units of S. mutans (CFU/mL), visible biofilm index, and socioeconomic status were all examined. The sample, comprising 23 individuals, had an average age of 17.3 ± 5.6 years, a DMF-T index of 5.5, a diminished salivary flow (78.3%), and a low (73.9%) saliva buffering capacity. Most (52.2%) individuals presented with a high abundance (CFU/mL) of S. mutans. The experience of caries was correlated with salivary parameters, poor oral hygiene, lower socioeconomic status and an increased count of S. mutans in saliva.
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http://dx.doi.org/10.1590/1807-3107BOR-2017.vol31.0079DOI Listing
September 2017

Evaluation of a Titanium Surface Treated with Hydroxyapatite Nanocrystals on Osteoblastic Cell Behavior: An In Vitro Study.

Int J Oral Maxillofac Implants 2018 May/June;33(3):597–602. Epub 2017 Sep 22.

Purpose: In the context of macrostructural and microstructural modifications to the design of dental implants, surface topography changes with different treatments have the purpose of accelerating bone formation. The aim of this study was to evaluate in vitro the influence of aggregated hydroxyapatite nanocrystals to surfaces treated with double acid etching (Nano) on osteoblastic cell behavior compared with a conventional double acid-etched surface (DE).

Materials And Methods: Commercially pure Grade 4 titanium discs (6 × 2 mm) were selected, and both cell proliferation and viability were assessed at 24, 48, and 72 hours using Trypan blue vital dye and MTT, respectively. The expression of type I collagen and osteopontin on such surfaces was evaluated using ELISA. Immunostaining for fibronectin was also performed. Quantitative data were analyzed statistically using two-way analysis of variance (ANOVA) followed by Bonferroni post-test with a 5% significance level.

Results: The results showed that in all evaluated time periods, cells expressed fibronectin on both surfaces. The cells presented greater morphologic spreading on the Nano surface when compared with the conventional DE surface in all assessed times. Increased cell proliferation and viability were detected in the Nano surface (P < .05) when compared with the conventional DE surface, especially after 72 hours. Osteopontin expression was higher after 24 hours in the Nano surface when compared with the conventional DE surface (P < .05). For type I collagen, a higher expression was observed with the Nano surface than with the DE surface, again after 72 hours (P < .05).

Conclusion: This in vitro study showed that the treated Nano surface tested promoted increased cell proliferation and viability when compared with the control surface. Additionally, increased cell spreading as well as type I collagen and osteopontin secretion were observed, favoring the early events of osseointegration.
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http://dx.doi.org/10.11607/jomi.5887DOI Listing
September 2018

Effect of epithelial growth factor on matrix metalloproteinase-2 and E-cadherin/β-catenin expression in an model of tumorigenesis.

Oncol Lett 2017 Sep 30;14(3):3136-3140. Epub 2017 Jun 30.

Department of Oral Pathology, São Leopoldo Mandic Institute and Research Center, Campinas, São Paulo 13045-755, Brazil.

The aim of the present study was to analyze the effect of various doses of epidermal growth factor (EGF; 5 and 10 ng/ml) on matrix metalloproteinase-2 (MMP-2) secretion and E-cadherin/β-catenin expression by co-cultured cells that mimic an carcinoma ex-pleomorphic adenoma, where benign myoepithelial cells from a pleomorphic adenoma surround malignant epithelial cells. EGF was supplemented in various doses and the effects were evaluated following four days of cell culture. ELISA was performed to determine MMP-2 secretion levels. Gene expression for E-cadherin and β-catenin was analyzed using quantitative polymerase chain reaction. The results revealed that E-cadherin expression decreased when the cells were supplemented with 5 ng/ml EGF. ELISA results indicated that MMP-2 secretion increased when EGF was supplemented at concentrations of 5 and 10 ng/ml. The present findings demonstrated that EGF may be involved in the epithelial-mesenchymal transition process via altering the E-cadherin/β-catenin complex and increasing MMP-2 secretion, which may then favor the dissolution of the basement membrane to the benefit of malignant cell clusters, contributing to the development of an invasive phenotype in this model of tumorigenesis.
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http://dx.doi.org/10.3892/ol.2017.6513DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5588072PMC
September 2017

Establishment of a primary culture of polymorphous low grade adenocarcinoma cells.

Arch Oral Biol 2017 Oct 15;82:188-193. Epub 2017 Jun 15.

Department of Oral Pathology, São Leopoldo Mandic Institute and Research Center, Rua José Rocha Junqueira 13, Swift, 13045-755 Campinas, SP, Brazil.

Objective: The aim of the present study was to establish a primary cell culture derived from polymorphous low grade adenocarcinoma (PLGA).

Design: The neoplastic cells were derived from a 57-year-old female patient diagnosed with PLGA. A fragment of the tumor was collected and submitted to enzymatic digestion followed by centrifugation on a Percoll gradient. The cell population was characterized by means of immunofluorescence and detection of PRKD1 gene mutations.

Results: Epifluorescence analysis of the primary culture revealed that the malignant epithelial cells were predominantly polygonal in shape and positive for cytokeratin 7, vimentin, and S100. The doubling time of the cell culture was 86.73h. The restriction digestion assay showed that the neoplastic cells possess PRKD1 gene mutations.

Conclusion: The establishment of primary cell culture derived from PLGA should be considered a useful tool for molecular analysis of this salivary gland tumor.
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http://dx.doi.org/10.1016/j.archoralbio.2017.06.015DOI Listing
October 2017

Simvastatin modulates gingival cytokine and MMP production in a rat model of ligature-induced periodontitis.

Clin Cosmet Investig Dent 2017 15;9:33-38. Epub 2017 May 15.

Laboratory of Cell and Molecular Biology, São Leopoldo Mandic Institute and Research Center, Campinas.

Purpose: The aim of this study was to evaluate the effect of simvastatin on the synthesis of cytokines TNF-α and IL-10 and metalloproteinase (MMPs) 2 and 9 in a rat model of ligature-induced periodontitis.

Materials And Methods: Twenty Wistar rats were used, and a cotton ligature was place in a subgingival position encircling the entire cervix of the first molar of the left (ipsilateral) side of the mandible. The right (contralateral) side of the mandible had no ligature placed and was used as control. After the ligature placement, animals were randomly assigned to two experimental groups (n=10): 1) rats with ligature + vehicle (saline; 10 mL/kg; orally) and 2) rats with ligature + simvastatin (25 mg/kg; orally). After 14 days of treatment, the animals were euthanized by anesthetic overdose and the gingival tissue was removed and homogenized in appropriate buffer. MMP-2 and -9 release as well as the IL-10 and TNF-α levels were detected by enzyme-linked immunosorbent assay. Statistical comparison was performed by unpaired Student's -test, with <0.05 representing significance.

Results: No differences were observed for TNF-α production between the groups (>0.05). However, IL-10 was upregulated in simvastatin-treated animals (1.8-fold increase) in comparison with the vehicle-treated group (<0.05). Simvastatin reduced the gingival levels of MMP-9 (64.3%) in comparison with vehicle-treated samples (<0.05).

Conclusion: Oral treatment with simvastatin increased the release of IL-10 and reduced the MMP-9 in ligature-induced periodontitis model in rats.
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http://dx.doi.org/10.2147/CCIDE.S134125DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5439939PMC
May 2017

Presence of Cells in Fresh-Frozen Allogeneic Bone Grafts from Different Tissue Banks.

Braz Dent J 2017 Mar-Apr;28(2):152-157

São Leopoldo Mandic Institute and Research Center, Campinas, SP, Brazi.

Bone replacement materials have been widely used to reconstruct atrophic jawbones. Based on previous reports demonstrating the presence of viable cells in bone blocks even after processing by musculoskeletal tissue banks for orthopedic use, the aim of this study was to evaluate the presence of cells in bone blocks from three Brazilian tissue banks for maxillary reconstructions. All samples were processed by the respective tissue banks, according to the guidelines of the Brazilian National Sanitary Surveillance Agency. Three samples were removed from each block for subsequent histological processing and stained using hematoxylin & eosin. Further evaluation included section staining by the Feulgen method and ultrastructural analysis using scanning electron microscopy (SEM). Light microscopy images from all bone samples showed presence of osteocyte-like cells in all groups and intense Feulgen staining, demonstrating presence of DNA in bone even after tissue processing. The ultrastructural analysis showed red blood cells in lacunae within the bone tissue. In conclusion, despite bone tissue processing by the musculoskeletal tissue banks, cells may be found within the bone used for allogeneic grafts.
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http://dx.doi.org/10.1590/0103-6440201701206DOI Listing
March 2018

Behavior of MC3T3-E1 Osteoblastic Cells Cultured on Titanium and Zirconia Surfaces: An In Vitro Study.

Implant Dent 2017 Jun;26(3):373-377

*Resident, Department of Oral Biology Implantology, São Leopoldo Mandic School of Dentistry, Campinas, Brazil. †Professor, Department of Cellular and Molecular Biology, São Leopoldo Mandic School of Dentistry, Campinas, Brazil.

Purpose: The goal from this in vitro study was to evaluate osteoblast protein expression of collagen type 1 (col 1) and osteopontin (OPN) on titanium surface treated with double etching compared with a zirconia implant surface.

Materials And Methods: Discs were selected on both surfaces with respective treatments. Mouse MC3T3-E1 osteoblastic cells were tested for cell viability using the MTT assay. Subsequently, the quantification of col 1 and OPN secreted by osteoblastic cells plated on different surfaces was evaluated by enzyme linked immunosorbent assay (ELISA).

Results: Regarding the cell proliferation, statistical analysis showed no significant effect of surface-time interaction and viable cell count on the surfaces of titanium versus zirconia. No statistical differences between the surfaces of titanium and zirconia on cell viability was found. The zirconia surface expression of OPN was significantly higher, which occurred in all times. Furthermore, zirconia demonstrated significantly greater in col 1 expression in 48 and 96 hours compared with titanium.

Conclusion: In this study, the zirconia surface demonstrated OPN and col 1 expression significantly higher as compared with titanium.
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http://dx.doi.org/10.1097/ID.0000000000000543DOI Listing
June 2017

Microbiological Sealing Analysis of a Tapered Connection and External Hexagon System.

Int J Dent 2017 28;2017:3849085. Epub 2017 Feb 28.

São Leopoldo Mandic Institute and Research Center, Campinas, SP, Brazil.

Considering the variety of implant connection systems available in the market and the contrasting literature regarding tapered connection systems in terms of bacterial leakage, the aim of this in vitro study was to compare the effectiveness of the bacterial seal at the implant/abutment interface between an external hexagon and a tapered connection system. Twelve sets of indexed tapered connection components and twelve sets of external hexagon connection components were used for microbiological analysis. In addition, for each model, an implant with its respective prosthetic abutment was used as a negative control and another as a positive control of microbial contamination. Failure of the abutment/implant interface seal was observed via turbidity or presence of deposits in the culture. Descriptive analysis of the data and relative frequency (percentage) as well as Fisher's exact test were used at a significance level of 5%. Two of ten (20%) external hexagon specimens showed contamination against 0/10 (0%) tapered connection implants. In conclusion, both implant/abutment connections were able to prevent bacterial leakage in vitro.
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http://dx.doi.org/10.1155/2017/3849085DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5350418PMC
February 2017