Doxycycline-doped collagen membranes accelerate in vitro osteoblast proliferation and differentiation.

OBJECTIVE: The aim of the study was to evaluate the effect of doxycycline- and dexamethasone-doped collagen membranes on the proliferation and differentiation of osteoblasts. BACKGROUND: Collagen barrier membranes are frequently used to promote bone regeneration and to boost this biological activity their functionalization with antibacterial and immunomodulatory substances has been suggested. METHODS: The design included commercially available collagen membranes doped with doxycycline (Dox-Col-M) or dexamethasone (Dex-Col-M), as well as undoped membranes (Col-M) as controls, which were placed in contact with cultured MG63 osteoblast-like cells (ATCC). Cell proliferation was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) assay and differentiation by measuring the alkaline phosphatase (ALP) activity using spectrophotometry. Real-time quantitative polymerase chain reaction was used to study the expression of the genes: Runx-2, OSX, ALP, OSC, OPG, RANKL, Col-I, BMP-2, BMP-7, TGF-ß1, VEGF, TGF-ßR1, TGF-ßR2, and TGF-ßR3. Scanning electron microscopy was used to study osteoblast morphology. Data were assessed using one-way analysis of variance or Kruskal-Wallis tests, once their distribution normality was assessed by Kolmogorov-Smirnov tests (p > .05). Bonferroni for multiple comparisons were carried out (p < .05). RESULTS: Osteoblast proliferation was significantly enhanced in the functionalized membranes as follows: (Col-M < Dex-Col-M < Dox-Col-M). ALP activity was significantly higher on cultured osteoblasts on Dox-Col-M. Runx-2, OSX, ALP, OSC, BMP-2, BMP-7, TGF-ß1, VEGF, TGF-ßR1, TGF-ßR2, and TGF-ßR3 were overexpressed, and RANKL was down-regulated in osteoblasts cultured on Dox-Col-M. The osteoblasts cultured in contact with the functionalized membranes demonstrated an elongated spindle-shaped morphology. CONCLUSION: The functionalization of collagen membranes with Dox promoted an increase in the proliferation and differentiation of osteoblasts.

Role of Salivary MicroRNA and Cytokines in the Diagnosis and Prognosis of Oral Squamous Cell Carcinoma.

Oral squamous cell carcinoma (OSCC) is the most prevalent oral malignant tumor worldwide. An early diagnosis can have a major positive impact on its prognosis. Human saliva contains cytokines, DNA and RNA molecules, circulating cells, and derivatives of tissues and extracellular vesicles, among other factors that can serve as biomarkers. Hence, the analysis of saliva may provide useful information for the early diagnosis of OSCC for its prognosis. The objective of this review was to determine the potential usefulness of salivary biomarkers (cytokines and microRNA) to diagnose OSCC and improve its prognosis. A combination of salivary miRNA and proteomic data could allow a definitive and early diagnosis to be obtained. However, there remains a need to optimize and standardize the protocols used to quantify miRNAs.

Preoperative oral pregabalin for anxiety control: a systematic review.

OBJECTIVE: The objective of this systematic review was to determine the effectiveness of preoperative oral pregabalin for anxiety control, the most effective dosage regimen, its impact on postoperative pain, and its adverse effects. MATERIALS AND METHODS: A search was conducted of PubMed/Medline and clinicaltrials.gov (National Library of Medicine, Washington, DC), Scopus, Web of Science, and Cochrane databases for studies published between January 2009 and November 2018, with no language restriction. Based on PRISMA guidelines, the specific question was: is preoperative oral pregabalin effective and safe for anxiety control in patients undergoing surgery? The critical reading of retrieved studies followed questions prepared by the CASPe Network, and their methodological quality was evaluated using the Jadad Scale. RESULTS: Twelve randomized controlled trials were selected for review. All twelve studies were trials of high quality. A dose of 75 mg preoperative oral pregabalin has been found to reduce anxiety and stabilize intraoperative hemodynamics, although a more significant improvement appears to be achieved with a single dose of 150 mg pregabalin at least 1 h before the surgery. It is not associated with any severe adverse effects. CONCLUSION: Preoperative administration of oral pregabalin in a single dose of 150 mg appears to be effective to significantly reduce the anxiety of patients, intraoperative hemodynamic changes, and postoperative pain. CLINICAL RELEVANCE: These findings suggest that pregabalin is useful and safe for preoperative and intraoperative anxiety control in patients undergoing surgery.

Salivary Biomarkers and Their Application in the Diagnosis and Monitoring of the Most Common Oral Pathologies.

Saliva is a highly versatile biological fluid that is easy to gather in a non-invasive manner-and the results of its analysis complement clinical and histopathological findings in the diagnosis of multiple diseases. The objective of this review was to offer an update on the contribution of salivary biomarkers to the diagnosis and prognosis of diseases of the oral cavity, including oral lichen planus, periodontitis, Sjögren's syndrome, oral leukoplakia, peri-implantitis, and medication-related osteonecrosis of the jaw. Salivary biomarkers such as interleukins, growth factors, enzymes, and other biomolecules have proven useful in the diagnosis and follow-up of these diseases, facilitating the early evaluation of malignization risk and the monitoring of disease progression and response to treatment. However, further studies are required to identify new biomarkers and verify their reported role in the diagnosis and/or prognosis of oral diseases.

Impact of bisphosphonates on the proliferation and gene expression of human fibroblasts.

The aim of this study was to elucidate the role of fibroblasts in bisphosphonate-related osteonecrosis of the jaw (BRONJ), evaluating the effect of zoledronate, alendronate, and ibandronate on the proliferation of fibroblasts and on their expression of genes essential for fibroblast physiology. Human CCD-1064Sk epithelial fibroblast cells were incubated in culture medium with 10-5, 10-7, or 10-9 M zoledronate, alendronate, or ibandronate. The proliferative capacity of fibroblasts was determined by spectrophotometry (MTT) at 24 of culture. Real-time polymerase chain reaction (RT-PCR) was used to study the effects of BPs at a dose of 10-9 M on the expression of FGF, CTGF, TGF-ß1, TGFßR1, TGFßR2, TGFßR3, DDR2, α-actin, fibronectin, decorin, and elastin. Fibroblasts proliferation was significantly increased at the lowest dose (10-9M) of each BP but was not affected at the higher doses (10-5 and 10-7M). The proliferation increase may be related to the rise in TGF-ß1 and TGFßR1 expression detected after the treatment of cells with 10-9M of zoledronate, alendronate, or ibandronate. However, the expression of CTGF, DDR2, α-actin, fibronectin, and decorin decreased versus controls. The results of this in vitro study indicate that a very low BP dose (10-9 M) can significantly affect the physiology of fibroblasts, increasing their proliferative capacity and modulating the expression of multiple genes involved in their growth and differentiation.

Influence of pH on osteoclasts treated with zoledronate and alendronate.

OBJECTIVES: The objectives of this study were to analyze the effect of pH on the growth and activity of osteoclasts treated with different doses of two nitrogen-containing BPs, zoledronate and alendronate. MATERIALS AND METHODS: Murine osteoclasts cultured on dentine disks were treated with zoledronate (50 or 500 nM) or alendronate (500 or 5 µM) at two different pH values (7.4 or 7.0). Osteoclasts were counted with transmitted light microscopy, apoptosis/necrosis was studied with flow cytometry and confocal microscopy, and resorption pit number and depth were calculated using reflected light and scanning electron microscopy. RESULTS: The osteoclast count on dentine disks was significantly (p < 0.001) reduced by zoledronate or alendronate treatment at pH 7.0 in comparison to treatment with the same doses at pH 7.4 and untreated disks (controls). The percentage of apoptotic cells was significantly increased by treatment with 500 nM zoledronate or 5 µM alendronate at pH 7.0 in comparison to the same doses at pH 7.4. The number and depth of resorption pits were significantly lower in disks treated at each BP dose studied than in untreated controls at pH 7.0. CONCLUSIONS: Zoledronate and alendronate at therapeutic doses have an adverse effect on the viability and resorptive activity of osteoclasts when the local medium pH is reduced. CLINICAL RELEVANCE: These findings suggest that periodontal or peri-implant oral cavity infection may be a key trigger of the cascade of events that lead to BRONJ.

Effect of olive oil phenolic compounds on osteoblast differentiation.

BACKGROUND: Osteoporosis is a skeletal disorder characterized by compromised bone strength that predisposes individuals to an increased risk of fracture. Previous in vivo and in vitro studies have reported that phenolic compounds present in extra virgin olive oil have a beneficial effect on osteoblasts in terms of increase cell proliferation. The aim of this study was to determine whether phenolic compounds present in olive oil could modify the expression of cell differentiation markers on osteoblasts. STUDY DESIGN: An in vitro experimental design was performed using MG-63 osteoblasts cell line. METHODS: MG63 cells were exposed to different doses of luteolin, apigenin, or p-coumaric, caffeic or ferulic acid. Alkaline phosphatase (ALP) was evaluated by spectrophotometry and antigen expression (cluster of differentiation [CD] 54, CD80, CD86 and HLA-DR) by flow cytometry. RESULTS: At 24 hour, treated groups showed an increased ALP and modulated antigen profile, with respect to the nontreated group. CONCLUSION: These results demonstrate that the phenolic compounds studied induce cell maturation in vitro, increasing ALP synthesis and reducing the expression of antigens involved in immune functions of the osteoblast which would improve bone density.

Bisphosphonate Modulation of the Gene Expression of Different Markers Involved in Osteoblast Physiology: Possible Implications in Bisphosphonate-Related Osteonecrosis of the Jaw.

The aim of the present study was to elucidate the role of osteoblasts in bisphosphonates-related osteonecrosis of the jaw (BRONJ). The specific objective was to evaluate the effect on osteoblasts of two nitrogen-containing BPs (zoledronate and alendronate) and one non-nitrogen-containing BP (clodronate) by analyzing modulations in their expression of genes essential for osteoblast physiology. Real-time polymerase chain reaction (RT-PCR) was used to study the effects of zoledronate, alendronate, and clodronate at doses of 10-5, 10-7, or 10-9 M on the expression of Runx-2, OSX, ALP, OSC, OPG, RANKL, Col-I, BMP-2, BMP-7, TGF-ß1, VEGF, TGF-ßR1, TGF-ßR2, and TGF-ßR3 by primary human osteoblasts (HOBs) and MG-63 osteosarcoma cells. Expression of these markers was found to be dose-dependent, with no substantive differences between these cell lines. In general, results demonstrated a significant increase in TFG-ß1, TGF-ßR1, TGF-ßR2, TGF-ßR3, and VEGF expressions and a significant reduction in RUNX-2, Col-1, OSX, OSC, BMP-2, BMP-7, ALP, and RANKL expressions, while OPG expression varied according to the dose and cell line. The results of this in vitro study of HOBS and MG-63 cell lines indicate that low BP doses can significantly affect the expression of genes essential for osteoblast growth and differentiation and of genes involved in regulating osteoblast-osteoclast interaction, possibly by increasing TGF-ß1 production. These findings suggest that osteoblasts may play an important role in BRONJ development, without ruling out other factors.

Oral pregabalin for acute pain relief after cervicofacial surgery: a systematic review.

OBJECTIVE: The objectives of this systematic review were to unify criteria on the effectiveness of oral pregabalin to treat acute post-operative pain after cervicofacial surgery, to establish the most effective dose regimens, and to determine its effect on rescue medicine consumption and its association with adverse effects. MATERIALS AND METHODS: PubMed/Medline (National Library of Medicine, Washington, DC), Scopus, Web of Science, and Cochrane databases were searched for studies in any language published between January 2000 and September 2016. The following question was posed, in accordance with PRISMA guidelines: Is oral pregabalin effective and safe for the relief of acute pain after cervicofacial surgery? The critical reading of the literature utilized a list of questions prepared by the CASPe Network, applying the Jadad scale for evaluation of the methodological quality of trials. RESULTS: Eleven randomized controlled clinical trials were selected. The 11 trials obtained a score ≥ 3, considered as Ib evidence level and high quality. A single oral dose of 75-mg pregabalin before or after cervicofacial surgery alleviates pain and lessens the need for rescue analgesia consumption, while the statistical significance of these effects is higher with a single dose of 150-mg pregabalin, either before or after the surgery. CONCLUSION: Oral pregabalin appears to significantly alleviate post-operative pain and reduce rescue analgesia consumption, with no severe adverse effects. However, the ideal dose and most effective administration regimen remain controversial issues that need to be addressed in further high-quality clinical trials. CLINICAL RELEVANCE: These findings suggest that pregabalin may be useful for acute pain relief after cervicofacial surgery.

Double-blind, randomized controlled clinical trial on analgesic efficacy of local anesthetics articaine and bupivacaine after impacted third molar extraction.

OBJECTIVE: The objective of this randomized controlled clinical trial (RCT) was to compare the effect of bupivacaine and articaine at habitual doses on pain intensity and the need for analgesics after lower third molar extraction. MATERIALS AND METHODS: The final study sample comprised 50 Caucasian volunteers (26 males and 24 females; age range, 18-30 years) undergoing scheduled surgical extraction of impacted lower third molar. A computer-generated random sequence was used to allocate participants to the articaine (4%) or bupivacaine (0.5%) group. Surgeons and patients were blinded by labeling the articaine and bupivacaine carpules with numbers (1 and 2, respectively). Postoperative pain intensity (primary outcome) was evaluated with a visual analogue scale (VAS), while the requirement for and timing of rescue medication and the quality of intraoperative anesthesia were also measured (secondary outcomes). RESULTS: VAS-measured pain intensity was significantly higher (p < 0.05) in the articaine group than in the bupivacaine group at all time points except for 8 h post-surgery (p = 0.052). Rescue medication was required by 13 (52%) patients in the articaine group and 8 (32%) patients in the bupivacaine group, although the difference did not reach statistical significance (p = 0.252). The groups did not significantly differ (p = 0.391) in the quality of the intraoperative anesthesia. CONCLUSIONS: Bupivacaine is a valid alternative to articaine in third molar surgery and may offer residual anesthesia as a means of reducing postoperative pain. However, further well-designed RCTs are required in larger study populations to verify the effectiveness of bupivacaine to achieve residual analgesia after oral surgery. CLINICAL RELEVANCE: These findings suggest that bupivacaine may be useful as a coadjuvant to control acute postoperative pain. TRIAL REGISTRATION: ACTRN12617001138370.

Effect of Clodronate on Antigenic Profile, Growth, and Differentiation of Osteoblast-Like Cells.

PURPOSE: To evaluate the role of osteoblasts in bisphosphonate-related osteonecrosis of the jaw (BRONJ) by studying the effects of different concentrations of clodronate, a non-nitrogen-containing bisphosphonate, on osteoblast growth, differentiation, and antigenic profile. MATERIALS AND METHODS: Osteoblast-like cells (MG63) were incubated in culture medium with different doses of clodronate. Their proliferative capacity was determined with a spectrophotometric technique (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium assay). Flow cytometry was used to study the antigenic profile. Cell differentiation was evaluated by nodule formation and alkaline phosphatase (ALP) activity was measured by spectrophotometric assay. RESULTS: Clodronate had a significant stimulatory effect on osteoblast-like cell (MG63) proliferation (P < .05). A significant decrease in the expression of CD54, CD80, CD86, and HLA-DR membrane antigens versus controls was observed after 24 hours of treatment with the different clodronate doses assayed (P < .05). A significant decrease (P = .004) in ALP activity was found after 24 hours of treatment with the lowest dose (10(-9) mol/L), and a significant decrease in calcium deposition was found after 15 and 21 days of treatment (P < .05). CONCLUSION: Clodronate increases the proliferation of MG63 osteoblast-like cells and decreases their differentiation capacity, generally at low doses, and modulates the expression of costimulatory molecules associated with immune function. Clodronate exerts its effect on osteoblasts by altering their physiology and impairing their repair capacity, which could be related to the development of BRONJ. However, further research is warranted to elucidate fully the mechanisms by which bisphosphonates can produce this disease.

Nitrogen-containing bisphosphonates modulate the antigenic profile and inhibit the maturation and biomineralization potential of osteoblast-like cells.

OBJECTIVES: The aim was to evaluate the effect of three nitrogen-containing bisphosphonates at different concentrations on osteoblast growth, differentiation, and antigenic profile, using the MG-63 cell line as osteoblast model, in order to determine the role of osteoblasts in bisphosphonate-related osteonecrosis of the jaw (BRONJ). MATERIALS AND METHODS: Osteoblasts were incubated in culture medium with 10(-5), 10(-7), or 10(-9) M of pamidronate, alendronate, or ibandronate. Proliferative capacity of the osteoblasts was determined by spectrophotometry (MTT) at 24 and 48 h of culture. Flow cytometry was used to study antigenic profile (CD54, CD80, CD86, HLA-DR) and phagocytic activity. Cell differentiation was evaluated at 7, 15, and 21 days by the study of nodule formation and alkaline phosphatase activity (ALP) at 24 h by spectrophotometric assay. RESULTS: Pamidronate, alendronate, and ibandronate each exerted a significant stimulatory effect on MG63 proliferation that depended on the dose and treatment duration (p < 0.05). In general, a significantly decreased expression of CD54, CD80, and HLA-DR membrane antigens was observed after 24 h of treatment with each nitrogen-containing bisphosphonate (p < 0.05), but there was no significant difference in phagocytic activity versus controls. A decrease in ALP activity was observed after 24 h of treatment and a decrease in calcium deposition after 15 and 21 days (p < 0.05). CONCLUSION: Nitrogen-containing bisphosphonates can increase the proliferation of MG-63 osteoblast-like cells, modulate their expression of co-stimulatory molecules associated with immune function, and decrease their differentiation capacity, generally at low doses. CLINICAL RELEVANCE: These findings suggest that low doses of nitrogen-containing bisphosphonates exert their effect on osteoblasts by altering their physiology, which would explain the disruption of their repair capacity and may be directly related to the development of BRONJ.

The effects of low-level diode laser irradiation on differentiation, antigenic profile, and phagocytic capacity of osteoblast-like cells (MG-63).

Previous in vivo and in vitro studies have reported that low-level diode laser therapy induces a biostimulatory effect, such as cell proliferation. The aim of the present study was to evaluate whether the laser irradiation of osteoblast-like cells (MG-63) can modify alkaline phosphatase activity (ALP), antigenic profile, and phagocytic capacity. The MG-63 cell line was exposed to diode laser (ezLase) of 940 nm at 1-1.5 W/cm(2) and 3-4 J. ALP was evaluated by a spectrophotometric technique and antigenic expression analysis (CD 54, CD80, CD86, HLA-DR), and phagocytic activity was analyzed by flow cytometry. At 24 h, the treated groups showed an increased ALP, and the highest increase versus controls (P = 0.002) was at the dose of 1 W/cm(2) and 3 J; this modulation of the antigenic profile translated into a reduced expression of CD54, CD86, and HLA-DR and a slightly decreased phagocytic capacity with respect to the nonirradiated control group at the different intensities and fluencies assayed. These results demonstrate that laser therapy can exert a biostimulatory effect on osteoblastic cells at different levels, which may be clinically useful in the regeneration of bone tissue.

NP-12 peptide functionalized nanoparticles counteract the effect of bacterial lipopolysaccharide on cultured osteoblasts.

OBJECTIVE: To evaluate whether nanoparticles (NPs) functionalized with Tideglusib (TDg, NP-12), and deposited on titanium surfaces, would counteract the effect of bacterial lipopolysaccharide (LPS) on osteoblasts. METHODS: Experimental groups were: (a) Titanium discs (TiD), (b) TiD covered with undoped NPs (Un-NPs) and (c) TiD covered with TDg-doped NPs (TDg-NPs). Human primary osteoblasts were cultured onto these discs, in the presence or absence of bacterial LPS. Cell proliferation was assessed by MTT-assay and differentiation by measuring the alkaline phosphatase activity. Mineral nodule formation was assessed by the alizarin red test. Real-time quantitative polymerase chain reaction was used to study the expression of Runx-2, OSX, ALP, OSC, OPG, RANKL, Col-I, BMP-2, BMP-7, TGF-ß1, VEGF, TGF-ßR1, TGF-ßR2, and TGF-ßR3 genes. Osteoblasts morphology was studied by Scanning Electron Microscopy. One-way ANOVA or Kruskal-Wallis and Bonferroni multiple comparisons tests were carried out (p < 0.05). RESULTS: TDg-NPs enhanced osteoblasts proliferation. Similarly, this group increased ALP production and mineral nodules formation. TDg-NPs on titanium discs resulted in overexpression of the proliferative genes, OSC and OSX, regardless of LPS activity. In the absence of LPS, TDg-NPs up-regulated Runx2, COL-I, ALP, BMP2 and BMP7 genes. OPG/RANKL gene ratios were increased about 2500 and 4,000-fold by TDg-NPs, when LPS was added or not, respectively. In contact with the TDg-NPs osteoblasts demonstrated an elongated spindle-shaped morphology with extracellular matrix production. SIGNIFICANCE: TDg-NPs on titanium discs counteracted the detrimental effect of LPS by preventing the decrease on osteoblasts proliferation and mineralization, and produced an overexpression of proliferative and bone-promoting genes on human primary osteoblasts.

Effect of amoxicillin and clindamycin on the gene expression of markers involved in osteoblast physiology.

Background/purpose: Amoxicillin and clindamycin are the most effective decontaminants for intraoral bone grafts before their application in bone regeneration without cytotoxic effects on osteoblasts, but their effects on the gene expression of markers involved in osteoblast growth and differentiation remain unclear. The study objective was to determine the effects of amoxicillin and clindamycin on the gene expression of markers involved in osteoblast growth and differentiation. Materials and methods: Real-time polymerase chain reaction (RT-PCR) was performed to explore the effect of 150 µg/mL clindamycin or 400 µg/mL amoxicillin on the gene expression by primary human osteoblasts (HOBs) of runt-related transcription factor 2 (Runx-2), osterix (OSX), alkaline phosphatase (ALP), osteocalcin (OSC), osteoprotegerin (OPG), receptor activator for nuclear factor κ B ligand (RANKL), type I collagen (Col-I), bone morphogenetic proteins 2 and 7 (BMP-2 and BMP-7), TGF-ß1 and TGF-ß receptors (TGF-ßR1, TGF-ßR2, and TGF-ßR3), and vascular endothelial growth factor (VEGF). Results: Treatment with 150 µg/mL clindamycin significantly increased the gene expression of TFG-ß1, TGF-ßR1, TGF-ßR2, TGF-ßR3, RUNX-2, Col-1, OSX, OSC, BMP-2, BMP-7, ALP, VEGF, and RANKL by HOBs. Treatment with 400 µg/mL amoxicillin significantly increased the gene expression of TGF-ß R1, Col-I, OSC, RANKL, and OPG alone. Conclusion: These findings suggest that 150 µg/mL clindamycin is the decontaminant of choice to treat intraoral bone grafts before their application in bone regeneration. The osteogenic and antibacterial properties of clindamycin can favor and accelerate the integration of bone grafts in the oral cavity.

Proliferation and osteogenic differentiation of osteoblast-like cells obtained from two techniques for harvesting intraoral bone grafts.

OBJECTIVES: The aims of our study were to verify the presence of viable osteoblasts in samples of bone tissue obtained by drilling or from cortico-cancellous bone blocks and to assess their growth and differentiation capacities. MATERIALS AND METHODS: Bone tissue samples were processed independently and cultured in Dulbecco's modified Eagle medium, in a CO2 incubator at 37 °C. The proliferative capacity of osteoblasts was determined by spectrophotometry (MTT) at 24 and 48 h of culture. Cell cycle was analysed by flow cytometry. Cell differentiation was studied by red alizarin staining of nodules formed in mineralisation medium and by analysis of alkaline phosphatase activity. RESULTS: In comparison to bone block-derived osteoblasts, the proliferative capacity was greater at 24 and 48 h of culture (P < 0.001) in the drilling-derived osteoblasts, which showed significantly increased G2/M (P = 0.014) and S (P < 0.001) phases in the cell cycle study. The number of mineralised nodules was proportional to the incubation time, with no differences between the two types of sample, which also did not significantly differ in alkaline phosphatase activity. CONCLUSION: Superior autograft material is obtained by harvesting particulate bone from low-speed drilling fragments than from a cortico-cancellous bone block. CLINICAL RELEVANCE: These results suggest that bone obtained from low-speed drilling is a simple and effective alternative to the classic procedure for obtaining bone tissue.

Regeneration Membranes Loaded with Non-Antibiotic Anti-2 Microbials: A Review.

Both guided bone and guided tissue regeneration are techniques that require the use of barrier membranes. Contamination and infection of the surgical area is one of the most feared complications. Some current lines of research focus on functionalizing these membranes with different antimicrobial agents. The objective of this study was to carry out a review of the use and antibacterial properties of regeneration membranes doped with antimicrobials such as zinc, silver, chlorhexidine, and lauric acid. The protocol was based on PRISMA recommendations, addressing the PICO question: "Do membranes doped with non-antibiotic antimicrobials have antibacterial activity that can reduce or improve infection compared to membranes not impregnated with said antimicrobial?" Methodological quality was evaluated using the RoBDEMAT tool. A total of 329 articles were found, of which 25 met the eligibility criteria and were included in this review. Most studies agree that zinc inhibits bacterial growth as it decreases colony-forming units, depending on the concentration used and the bacterial species studied. Silver compounds also decreased the secretion of proinflammatory cytokines and presented less bacterial adhesion to the membrane. Some concentrations of chlorhexidine that possess antimicrobial activity have shown high toxicity. Finally, lauric acid shows inhibition of bacterial growth measured by the disk diffusion test, the inhibition zone being larger with higher concentrations. Antimicrobial agents such as zinc, silver, chlorhexidine, and lauric acid have effective antibacterial activity and can be used to dope regenerative membranes in order to reduce the risk of bacterial colonization.

Dexamethasone and doxycycline functionalized nanoparticles enhance osteogenic properties of titanium surfaces.

OBJECTIVES: To evaluate the effect of doxycycline and dexamethasone doped nanoparticles covering titanium surfaces, on osteoblasts proliferation and differentiation. METHODS: Doxycycline and dexamethasone doped polymeric nanoparticles were applied on titanium discs (Ti-DoxNPs and Ti-DexNPs). Undoped NPs and uncovered Ti discs were used as control. Human MG-63 osteoblast-like cells were cultured. Osteoblasts proliferation was tested by MTT assay. Alkaline phosphatase activity was analyzed. Differentiation gene expression was assessed by real-time quantitative polymerase chain reaction. Scanning Electron Microscopy was performed to assess osteoblasts morphology. Mean comparisons were conducted by ANOVA and Wilcoxon or Tukey tests (p < 0.05). RESULTS: No differences in osteoblasts proliferation were found. Osteoblasts grown on Ti-DoxNPs significantly increased alkaline phosphatase activity. Doxycycline and dexamethasone nanoparticles produced an over-expression of the main osteogenic proliferative genes (TGF-ß1, TGF-ßR1 and TGF-ßR2). The expression of Runx-2 was up-regulated. The osteogenic proteins (AP, OSX and OPG) were also overexpressed on osteoblasts cultured on Ti-DoxNPs and Ti-DexNPs. The OPG/RANKL ratio was the highest when DoxNPs were present (75-fold increase with respect to the control group). DexNPs also produced a significantly higher OPG/RANKL ratio with respect to the control (20 times higher). Osteoblasts grown on titanium discs were mainly flat and polygonal in shape, with inter-cellular connections. In contrast, osteoblasts cultured on Ti-DoxNPs or Ti-DexNPs were found to be spindle-shaped and had abundant secretions on their surfaces. SIGNIFICANCE: DoxNPs and DexNPs were able to stimulate osteoblasts differentiation when applied on titanium surfaces, being considered potential inducers of osteogenic environment when performing regenerative procedures around titanium dental implants.

Biomimetic Collagen Membranes as Drug Carriers of Geranylgeraniol to Counteract the Effect of Zoledronate.

To counteract the effect of zoledronate and decrease the risk of osteonecrosis of the jaw (BRONJ) development in patients undergoing guided bone regeneration surgery, the use of geranylgeraniol (GGOH) has been proposed. Collagen membranes may act as biomimetical drug carriers. The objective of this study was to determine the capacity of collagen-based membranes doped with GGOH to revert the negative impact of zoledronate on the growth and differentiation of human osteoblasts. MG-63 cells were cultured on collagen membranes. Two groups were established: (1) undoped membranes and (2) membranes doped with geranylgeraniol. Osteoblasts were cultured with or without zoledronate (50 µM). Cell proliferation was evaluated at 48 h using the MTT colorimetric method. Differentiation was tested by staining mineralization nodules with alizarin red and by gene expression analysis of bone morphogenetic proteins 2 and 7, alkaline phosphatase (ALP), bone morphogenetic proteins 2 and 7 (BMP-2 and BMP-7), type I collagen (Col-I), osterix (OSX), osteocalcin (OSC), osteoprotegerin (OPG), receptor for RANK (RANKL), runt-related transcription factor 2 (Runx-2), TGF-ß1 and TGF-ß receptors (TGF-ßR1, TGF-ßR2, and TGF-ßR3), and vascular endothelial growth factor (VEGF) with real-time PCR. One-way ANOVA or Kruskal-Wallis and post hoc Bonferroni tests were applied (p < 0.05). Scanning electron microscopy (SEM) observations were also performed. Treatment of osteoblasts with 50 µM zoledronate produced a significant decrease in cell proliferation, mineralization capacity, and gene expression of several differentiation markers if compared to the control (p < 0.001). When osteoblasts were treated with zoledronate and cultured on GGOH-doped membranes, these variables were, in general, similar to the control group (p > 0.05). GGOH applied on collagen membranes is able to reverse the negative impact of zoledronate on the proliferation, differentiation, and gene expression of different osteoblasts' markers.

Antibiotic-Loaded Polymeric Barrier Membranes for Guided Bone/Tissue Regeneration: A Mini-Review.

Polymeric membranes are frequently used for bone regeneration in oral and periodontal surgery. Polymers provide adequate mechanical properties (i.e., Young's modulus) to support oral function and also pose some porosity with interconnectivity to permit for cell proliferation and migration. Bacterial contamination of the membrane is an event that may lead to infection at the bone site, hindering the clinical outcomes of the regeneration procedure. Therefore, polymeric membranes have been proposed as carriers for local antibiotic therapy. A literature search was performed for papers, including peer-reviewed publications. Among the different membranes, collagen is the most employed biomaterial. Collagen membranes and expanded polytetrafluoroethylene loaded with tetracyclines, and polycaprolactone with metronidazole are the combinations that have been assayed the most. Antibiotic liberation is produced in two phases. A first burst release is sometimes followed by a sustained liberation lasting from 7 to 28 days. All tested combinations of membranes and antibiotics provoke an antibacterial effect, but most of the time, they were measured against single bacteria cultures and usually non-specific pathogenic bacteria were employed, limiting the clinical relevance of the attained results. The majority of the studies on animal models state a beneficial effect of these antibiotic functionalized membranes, but human clinical assays are scarce and controversial.