Chronic wounds are characterized by persistent inflammation and microbial colonization, which interfere with the healing process and represent a significant clinical challenge. This study evaluated the anti-inflammatory and reparative potential of ellagic acid and punicalagin, along with their antimicrobial activity. Human dermal fibroblasts were exposed to a simulated inflammatory microenvironment induced by interleukin-1β (IL-1β), Interleukin-6 (IL-6), and Tumor Necrosis Factor-α (TNF) or bacterial lipopolysaccharides (LPS) and subsequently treated with ellagic acid or punicalagin (10−6 M and 10−7 M). Cell proliferation was assessed via MTT assay, and migration was evaluated using the scratch wound assay. IL-1β and IL-6 secretion was quantified by Enzyme-Linked Immunosorbent Assay in LPS-treated fibroblasts. Antimicrobial activity against Candida albicans, Staphylococcus aureus, and Pseudomonas aeruginosa was analyzed using the disk diffusion method. Both compounds significantly enhanced fibroblast viability and migration under inflammation and reduced the secretion of IL-1β and IL-6. However, no antimicrobial activity was observed at the tested concentrations. These findings suggest that ellagic acid and punicalagin may promote wound healing by modulating inflammation and supporting fibroblast function, despite lacking direct antimicrobial effect. Further in vivo studies are needed to validate their therapeutic relevance and explore their potential in the development of novel treatments for chronic wounds.

OBJECTIVE:The objective of this study was to determine the effects of combined treatment with clindamycin and zoledronate on the growth and differentiation of cultured human osteoblasts. DESIGN:Human osteoblasts, obtained by primary culture from mandibular bone fragments, were cultured in the presence of 50 μM zoledronate, 150 μg/mL clindamycin, or the combination of both (zoledronate + clindamycin). The effect on cell proliferation was evaluated at 24 h by the MTT colorimetric method, using a spectrophotometer at 570 nm. The effect on differentiation was examined by measuring alkaline phosphatase (ALP) activity, and mineralization by the osteoblast was studied by staining with alizarin red. Real-time polymerase chain reaction (RT-PCR) was performed for gene expression analysis. Data were expressed as means±standard deviation, and analysis of variance was performed, applying Bonferroni correction when interactions were significant. RESULTS:Treatment of osteoblasts with 50 μM zoledronate significantly reduced cell proliferation and differentiation and the gene expression of certain markers versus controls (p < 0.001). However, treatment with 150 μg/mL clindamycin significantly increased cell proliferation and differentiation and the gene expression of certain markers (p < 0.05). The combination of 150 μg/mL clindamycin and 50 μM zoledronate partially counteracted the loss of osteoblast proliferative and differentiation capacity caused by zoledronate. CONCLUSION:Treatment with low-dose clindamycin can reverse the negative impact of zoledronate on osteoblast proliferation and differentiation. Follow-up animal studies and clinical trials are needed before topical clindamycin can be considered as a possible therapeutic resource for BP-treated patients who require a GBR procedure.

The effects of the phenolic compounds punicalagin and ellagic acid, derived from pomegranate, were analyzed on the gene expression of cultured human fibroblasts. The focus was placed on genes associated with growth factors, differentiation markers, and extracellular matrix (ECM) components, all of which play a crucial role in effective wound healing. The human dermal fibroblast cell line CCD-1064Sk was used, and the bioactive compounds punicalagin and ellagic acid were administered to cultured fibroblasts at concentrations of 10-6 and 10-7 M, respectively. Gene expression was assessed at 24 h using real-time polymerase chain reaction (qRT-PCR). Treatments with punicalagin and ellagic acid dose-dependently increased the expression of platelet-derived growth factor (PDGF), transforming growth factor β (TGF-β), vascular endothelial growth factor (VEGF), connective tissue growth factor (CTGF), collagen 1 (COL1), collagen 3 (COL3), matrix metalloproteinase-2 (MMP2), α-actin, and fibronectin genes. Both compounds modulate the expression of genes related to the synthesis of growth factors, differentiation, and ECM components in human fibroblasts. These findings suggest the potential of these phenolic compounds to develop therapies that improve wound healing. Further studies are needed to better understand the mechanisms of cellular activation.

Extra virgin olive oil phenolic compounds have been identified as possible biostimulant agents against different pathological processes, including alterations in healing processes. However, there is little evidence on the molecular mechanisms involved in this process. The aim was to analyse the effect of hydroxytyrosol, tyrosol, and oleocanthal on fibroblast gene expression. PCR was used to determine the expression of different differentiation markers, extracellular matrix elements, and growth factors in cultured human fibroblasts CCD-1064Sk treated with different doses of hydroxytyrosol (10−5 M and 10−6 M), tyrosol (10−5 M and 10−6 M), and oleocanthal (10−6 M and 10−7 M). After 24 h of hydroxytyrosol treatment, increased expression of connective tissue growth factor, fibroblast growth factor (FGF), platelet-derived growth factor, vascular endothelial growth factor, transforming growth factor β1 (TGF-β1), and their receptors was observed. Tyrosol and olecanthal modulated the expression of FGF and TGFβR1. All phytochemicals tested modified the expression of differentiation markers and extracellular matrix elements, increasing gene expression of actin, fibronectin, decorin, collagen I, and III. Phenolic compounds present in extra virgin olive could have a beneficial effect on tissue regeneration by modulating fibroblast 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 mg/mL clindamycin or 400 mg/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 k B ligand (RANKL), type I collagen (Col -I), bone morphogenetic proteins 2 and 7 (BMP-2 and BMP-7), TGF-I31 and TGF-I3 receptors (TGF-I3R1, TGF-I3R2, and TGFI3R3), and vascular endothelial growth factor (VEGF). Results: Treatment with 150 mg/mL clindamycin significantly increased the gene expression of TFG-01, TGF-0R1, TGF-0R2, TGF-0R3, RUNX-2, Col -1, OSX, OSC, BMP-2, BMP-7, ALP, VEGF, and RANKL by HOBs. Treatment with 400 mu g/mL amoxicillin significantly increased the gene expression of TGF-0 R1, Col -I, OSC, RANKL, and OPG alone. Conclusion: These findings suggest that 150 mu g/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. 2023 Association for Dental Sciences of the Republic of China. Publishing services by Elsevier B.V. This is an open access article under the CC BY -NC -ND license (http://creativecommons. org/licenses/by-nc-nd/4.0/).

Background: Bisphenol A (BPA) and its analogs (BPF, BPS, and BPAF) are recognized for inducing detrimental effects on various tissues, including bone. Objectives: The aim of this study is to investigate their impact on information and repair processes, specifically focusing on vascular endothelial growth factor (VEGF), transforming growth factor β1 (TGF-β1), and the receptors for transforming growth factor β (TGFR1, TGFR2, and TGFR3). Methods: Human osteoblasts isolated through primary culture from bone samples of healthy volunteers were subjected to cultivation in the presence of various dosage levels (10−5, 10−6, or 10−7 M) of BPA, BPF, BPS, or BPAF for 24 h. Gene expressions of RANKL, OPG, TGF-β1, TGFR1, TGFR2, TGFR3, and VEGF were analyzed by real-time polymerase chain reaction (RT-PCR). All experiments included untreated cells as controls. Results: Expressions of RANKL and OPG were dose-dependently downregulated by the presence of all tested bisphenols (BPs) except for BPAF, whose presence upregulated OPG expression at all three doses. TGF-β1 expression was downregulated by all BP treatments, and TGF-β1 receptor expression was also downregulated as a function of the BP and dose. VEGF expression was downregulated in the presence of BPF and BPAF at all three doses and in the presence of BPA at the two higher doses (10−5, and 10−6 M), but it was not changed by the presence of BPS at any dose. Conclusions: The inhibition of both RANKL and OPG by the BPs, with a higher %inhibition of RANKL than of OPG, appears to rule out BP-induced activation of osteoclastogenesis via RANKL/RANK/OPG. Nevertheless, the effect of the BPs on the expression by osteoblasts of TGF-β1, TGF-β receptors, and VEGF indicates that these compounds can be responsible for major molecular changes in this cell population, contributing to their adverse effects on bone tissue.

Consequences of the disease produced by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have led to an urgent search for preventive and therapeutic strategies. Besides drug treatments, proposals have been made for supplementation with biomolecules possessing immunomodulatory and antioxidant properties. The objective of this study was to review published evidence on the clinical usefulness of supplementation with vitamin D, antioxidant vitamins (vitamin A, vitamin E, and vitamin C), melatonin, lactoferrin and natural products found in food (curcumin, luteolin, ginger, allicin, magnesium and zinc) as supplements in SARS-CoV-2 infection. In general, supplementation of conventional treatments with these biomolecules has been found to improve the clinical symptoms and severity of the coronavirus disease (COVID-19), with some indications of a preventive effect. In conclusion, these compounds may assist in preventing and/or improving the symptoms of COVID-19. Nevertheless, only limited evidence is available, and findings have been inconsistent. Further investigations are needed to verify the therapeutic potential of these supplements.

ObjectiveTo 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.MethodsExperimental 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-β1, VEGF, TGF-βR1, TGF-βR2, and TGF-βR3 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).ResultsTDg-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.SignificanceTDg-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.

Bone effects attributed to bisphenols (BPs) include the inhibition of growth and differentiation. This study analyzes the effect of BPA analogs (BPS, BPF, and BPAF) on the gene expression of the osteogenic markers RUNX2, osterix (OSX), bone morphogenetic protein-2 (BMP-2), BMP-7, alkaline phosphatase (ALP), collagen-1 (COL-1), and osteocalcin (OSC). Human osteoblasts were obtained by primary culture from bone chips harvested during routine dental work in healthy volunteers and were treated with BPF, BPS, or BPAF for 24 h at doses of 10(-5), 10(-6), and 10(-7) M. Untreated cells were used as controls. Real-time PCR was used to determine the expression of the osteogenic marker genes RUNX2, OSX, BMP-2, BMP-7, ALP, COL-1, and OSC. The expression of all studied markers was inhibited in the presence of each analog; some markers (COL-1; OSC, BMP2) were inhibited at all three doses and others only at the highest doses (10(-5) and 10(-6) M). Results obtained for the gene expression of osteogenic markers reveal an adverse effect of BPA analogs (BPF, BPS, and BPAF) on the physiology of human osteoblasts. The impact on ALP, COL-1, and OSC synthesis and therefore on bone matrix formation and mineralization is similar to that observed after exposure to BPA. Further research is warranted to determine the possible contribution of BP exposure to the development of bone diseases such as osteoporosis.

Bisphenol A (BPA) analogs, like BPA, could have adverse effects on human health including bone health. The aim was to determine the effect of BPF, BPS and BPAF on the growth and differentiation of cultured human osteoblasts. Osteoblasts primary culture from bone chips harvested during routine dental work and treated with BPF, BPS, or BPAF for 24 h at doses of 10-5, 10-6, and 10-7 M. Next, cell proliferation was studied, apoptosis induction, and alkaline phosphatase (ALP) activity. In addition, mineralization was evaluated at 7, 14, and 21 days of cell culture in an osteogenic medium supplemented with BP analog at the studied doses. BPS treatment inhibited proliferation in a dose-dependent manner at all three doses by inducing apoptosis; BPF exerted a significant inhibitory effect on cell proliferation at the highest dose alone by an increase of apoptosis; while BPAF had no effect on proliferation or cell viability. Cell differentiation was adversely affected by treatment with BPA analogs in a dose-dependent, observing a reduction in calcium nodule formation at 21 days. According to the results obtained, these BPA analogs could potentially pose a threat to bone health, depending on their concentration in the organism.

Fibroblasts contribute to maintaining tissue integrity and homeostasis and are a key cell population in wound healing. This cell population can be stimulated by some bioactive compounds such as extra virgin olive oil (EVOO) polyphenols. The aim of this study was to determine the effects of hydroxytyrosol (htyr), tyrosol (tyr), and oleocanthal (ole) phenolic compounds present in EVOO on the proliferation, migration, cell cycle, and antigenic profile of cultured human fibroblasts. CCD-1064Sk human fibroblast cells were treated for 24 h with each polyphenol at doses ranging 10−5 to 10−9 M. Cell proliferation was evaluated using the MTT spectrophotometric technique, migration capacity by culture insert assay, and cell cycle and antigenic profile with flow cytometry. Cell proliferation was significantly increased by treatment with all compounds. The highest increases followed treatments with htyr or tyr at doses of 10−5 or 10−6 M and with ole at 10−6 and 10−7 M, and these compounds and doses were used for assays of antigenic profile, cell cycle, and migration. During the first few hours after treatment, increased fibronectin and α-actin expressions and greater cell migration were observed, with no cell cycle changes. In conclusion, these in vitro results suggest that phenolic compounds in EVOO might contribute to wound healing through action on fibroblasts related to tissue regeneration.

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 (TiDoxNPs and Ti-DexNPs). Undoped NPs and uncovered Ti discs were used as control. Human MG-63 osteoblastlike 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-& beta;1, TGF-& beta;R1 and TGF-& beta;R2). 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.

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-β1 and TGF-β receptors (TGF-βR1, TGF-βR2, and TGF-βR3), 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.

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-β1, VEGF, TGF-βR1, TGF-βR2, and TGF-βR3. 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-β1, VEGF, TGF-βR1, TGF-βR2, and TGF-βR3 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.

(1) Background: Bisphenol A (BPA) is an endocrine disruptor that is widely present in the environment and exerts adverse effects on various body tissues. The objective of this study was to determine its repercussions on bone tissue by examining its impact on selected functional parameters of human osteoblasts. (2) Methods: Three human osteoblast lines were treated with BPA at doses of 10−5, 10−6, or 10−7 M. At 24 h post-treatment, a dose-dependent inhibition of cell growth, alkaline phosphatase activity, and mineralization was observed. (4) Results: The expression of CD54 and CD80 antigens was increased at doses of 10−5 and 10−6 M, while the phagocytic capacity and the expression of osteogenic genes (ALP, COL-1, OSC, RUNX2, OSX, BMP-2, and BMP-7) were significantly and dose-dependently reduced in the presence of BPA. (5) Conclusions: According to these findings, BPA exerts adverse effects on osteoblasts by altering their differentiation/maturation and their proliferative and functional capacity, potentially affecting bone health. Given the widespread exposure to this contaminant, further human studies are warranted to determine the long-term risk to bone health posed by BPA.

Background Antiseptics are used for the cleansing of acute or chronic wounds to eliminate micro-organisms from the wound bed. However, they have effects on the skin cells. Aim To determine the effects of hexetidine, povidone-iodine (PI), undecylenamidopropyl-betaine/polyhexanide (UBP), chlorhexidine, disodium eosin and hydrogen peroxide on human skin fibroblasts. Methods CCD-1064Sk cells were treated with hexetidine, PI, UBP, chlorhexidine, disodium eosin or hydrogen peroxide. Spectrophotometry was used to measure cell viability and flow cytometry was used to study apoptosis and necrosis after the treatment. In vitro wound scratch assays were performed to determine the gap closure. Results All antiseptics significantly reduced the viability of human skin fibroblasts compared with controls. The percentage wound closure was lower with hexetidine, PI and UBP. The scratch assay could not be measured after treatments with chlorhexidine, disodium eosin or hydrogen peroxide, owing to their cytotoxicity. The apoptosis/necrosis experiments evidenced a significant reduction in viable cells compared with controls. An increased percentage of apoptotic cells was observed after treatment with all antiseptics. Compared with controls, the percentage of necrotic cells was significantly increased with all antiseptics except for hexetidine. Conclusion The proliferation, migration and viability of human skin fibroblasts are reduced by treatment with hexetidine, PI, UBP, chlorhexidine, disodium eosin and hydrogen peroxide.

Garlic is one of the most widely employed condiments in cooking. It has also been used since ancient times in traditional plant-based medicine, largely based on its organosulfur compounds. The objective of this study was to provide updated information on the biological and therapeutic garlic properties. Garlic has been found to possess important biological properties with high therapeutic potential, which is influenced by the mode of its utilization, preparation, and extraction. It has been attributed with antioxidant, anti-inflammatory, and immunomodulatory capacities. Garlic, in particular its organosulfur compounds, can maintain immune system homeostasis through positive effects on immune cells, especially by regulating cytokine proliferation and expression. This may underlie their usefulness in the treatment of infectious and tumor processes. These compounds can also offer vascular benefits by regulating lipid metabolism or by exerting antihypertensive and antiaggregant effects. However, further clinical trials are warranted to confirm the therapeutic potential of garlic and its derivatives.

Polymeric membranes are employed in guided bone regeneration (GBR) as physical barriers to facilitate bone in-growth. A bioactive and biomimetic membrane with the ability to participate in the healing and regeneration of the bone is necessary. The aim of the present study was to analyze how novel silicon dioxide composite membranes functionalized with zinc or doxycycline can modulate the osteoblasts’ proliferation, differentiation, and expression of selected antigenic markers related to immunomodulation. Nanostructured acrylate-based membranes were developed, blended with silica, and functionalized with zinc or doxycycline. They were subjected to MG63 osteoblast-like cells culturing. Proliferation was assessed by MTT-assay, differentiation by evaluating the alkaline phosphatase activity by a spectrophotometric method and antigenic phenotype was assessed by flow cytometry for selected markers. Mean comparisons were conducted by one-way ANOVA and Tukey tests (p < 0.05). The blending of silica nanoparticles in the tested non-resorbable polymeric scaffold improved the proliferation and differentiation of osteoblasts, but doxycycline doped scaffolds attained the best results. Osteoblasts cultured on doxycycline functionalized membranes presented higher expression of CD54, CD80, CD86, and HLA-DR, indicating a beneficial immunomodulation activity. Doxycycline doped membranes may be a potential candidate for use in GBR procedures in several challenging pathologies, including periodontal disease.

Abstract Study question We aimed to analyze the effects of decidualization on the expression of chemokines that attract abortogenic T cells by human DSCs. Summary answer Decidualization inhibits the expression of chemokines that attract Th1 and Tc1 cells by DSCs, thereby preventing the arrival of abortogenic T cells into the decidua. What is known already Decidual stromal cells (DSCs) are the most abundant cells in the human decidua, the tissue that constitutes the maternal component of the placenta. Numerous evidences confirm that DSCs play a key role in maternal-fetal immune tolerance. In normal pregnancy, DSCs undergo a process of differentiation (decidualization) under the effect of progesterone and other pregnancy hormones. Decidualized DSCs become rounded and secrete prolactin, IL–15 and other factors. In the mouse, it has been observed that during pregnancy, DSCs inhibit the expression of chemokines that attract abortogenic Th1 and Tc1 cells from blood to the decidua. Study design, size, duration We compared the expression of CXCR3-binding chemokines by undifferentiated and decidualized human DSCs. We also compared the capacity of these cells to attract activated Th1 and Tc1 cells in vitro. Ten DSC lines were obtained from elective vaginal terminations of first-trimester pregnancies (6–11 weeks). Donors were healthy women aged 20–30 years. Informed consent was obtained from each donor. This study was approved by the Research and Ethics Committee of the University of Granada. Participants/materials, setting, methods Decidual stromal cell lines were established as previously described. These lines were decidualized with progesterone and cAMP in vitro. The expression of chemokines by these cells was studied by RT-PCR. Peripheral blood lymphocytes were activated with PHA, anti-CD28 and IL–2. As a consequence of this activation, CXCR3+ Th1 and Tc1 cells were produced. We used a migration assay in Transwell chambers to study the capacity of DSCs to attract these activated T cells. Main results and the role of chance We observed that those chemokines that bind to CXCR3, a chemokine receptor detected in activated Th1 and Tc1 cells, were not expressed by either undifferentiated and decidualized DSCs (CXCL9) or their expression was inhibited in decidualized DSCs (CXCL10 P < 0.01, CXCL11 P < 0.05). We found that conditioned media of undifferentiated DSCs decreased the migration of CXCR3+ activated T cells (Th1 and Tc1 cells) (P < 0.05), and this effect was even stronger with conditioned media of decidialized DSCs P < 0.001). These results demonstrated that decidualization of DSCs during pregnancy inhibits the expression of chemokines that attract Th1 and Tc1 cells by DSCs, thereby preventing the arrival of abortogenic T cells into the decidua. Limitations, reasons for caution This is an in vitro study due to the impossibility of performing an in vivo study in humans for ethical reasons. Wider implications of the findings: Several publications have shown that DSCs have a therapeutic effect in various Th1-associated diseases. Our results explain this effect and suggest the extension of the use of these cells in the treatment of this type of diseases. Trial registration number Not applicable