Sitagliptin Induces Tolerogenic Human Dendritic Cells.

Sitagliptin, an anti-diabetic drug, is a dipeptidyl peptidase (DPP)-4/CD26 inhibitor with additional anti-inflammatory and immunomodulatory properties. In this study, we investigated for the first time the effect of sitagliptin on the differentiation and functions of human dendritic cells generated from monocytes (MoDCs) for 4 days using the standard GM-CSF/IL-4 procedure. LPS/IFN-γ treatment for an additional 24 h was used for maturation induction of MoDCs. Sitagliptin was added at the highest non-cytotoxic concentration (500 µg/mL) either at the beginning (sita 0d protocol) or after MoDC differentiation (sita 4d protocol). Sitagliptin impaired differentiation and maturation of MoDCs as judged with the lower expression of CD40, CD83, CD86, NLRP3, and HLA-DR, retention of CD14 expression, and inhibited production of IL-ß, IL-12p70, IL-23, and IL-27. In contrast, the expression of CD26, tolerogenic DC markers (ILT4 and IDO1), and production of immunoregulatory cytokines (IL-10 and TGF-ß) were increased. Generally, the sita 0d protocol was more efficient. Sitagliptin-treated MoDCs were poorer allostimulators of T-cells in MoDC/T-cell co-culture and inhibited Th1 and Th17 but augmented Th2 and Treg responses. Tolerogenic properties of sitagliptin-treated MoDCs were additionally confirmed by an increased frequency of CD4+CD25+CD127- FoxP3+ Tregs and Tr1 cells (CD4+IL-10+FoxP3-) in MoDC/T-cell co-culture. The differentiation of IL-10+ and TGF-ß+ Tregs depended on the sitagliptin protocol used. A Western blot analysis showed that sitagliptin inhibited p65 expression of NF-kB and p38MAPK during the maturation of MoDCs. In conclusion, sitagliptin induces differentiation of tolerogenic DCs, and the effect is important when considering sitagliptin for treating autoimmune diseases and allotransplant rejection.

Pomegranate Peel Extract Differently Modulates Gene Expression in Gingiva-Derived Mesenchymal Stromal Cells under Physiological and Inflammatory Conditions.

Pomegranate has shown a favorable effect on gingivitis/periodontitis, but the mechanisms involved are poorly understood. The aim of this study was to test the effect of pomegranate peel extract (PoPEx) on gingiva-derived mesenchymal stromal cells (GMSCs) under physiological and inflammatory conditions. GMSC lines from healthy (H) and periodontitis (P) gingiva (n = 3 of each) were established. The lines were treated with two non-toxic concentrations of PoPEX (low-10; high-40 µg/mL), with or without additional lipopolysaccharide (LPS) stimulation. Twenty-four genes in GMSCs involved in different functions were examined using real-time polymerase chain reaction (RT-PCR). PoPEx (mostly at higher concentrations) inhibited the basal expression of IL-6, MCP-1, GRO-α, RANTES, IP-10, HIF-1α, SDF-1, and HGF but increased the expression of IL-8, TLR3, TGF-ß, TGF-ß/LAP ratio, IDO-1, and IGFB4 genes in H-GMSCs. PoPEx increased IL-6, RANTES, MMP3, and BMP2 but inhibited TLR2 and GRO-α gene expression in P-GMSCs. LPS upregulated genes for proinflammatory cytokines and chemokines, tissue regeneration/repair (MMP3, IGFBP4, HGF), and immunomodulation (IP-10, RANTES, IDO-1, TLR3, COX-2), more strongly in P-GMSCs. PoPEx also potentiated most genes' expression in LPS-stimulated P-GMSCs, including upregulation of osteoblastic genes (RUNX2, BMP2, COL1A1, and OPG), simultaneously inhibiting cell proliferation. In conclusion, the modulatory effects of PoPEx on gene expression in GMSCs are complex and dependent on applied concentrations, GMSC type, and LPS stimulation. Generally, the effect is more pronounced in inflammation-simulating conditions.

Case report: Thyroid sarcoidosis as a rare localization of the disease: Report of two cases and review of the literature.

Sarcoidosis is a multi-systemic disease of unknown etiology that is characterized by the formation of non-necrotizing and non-caseating granulomas in affected organs. Sarcoidosis granulomas can form in any organ, but the lungs and intrathoracic lymph nodes are the most commonly affected. Thyroid involvement in sarcoidosis is very rare, with prevalence estimates of 1-4.5% in case series of autopsied patients with systemic sarcoidosis. It is extremely rare for sarcoidosis to occur solely in the thyroid gland, but it is usually associated with the involvement of other organs, primarily the lungs and lymph nodes. Typical manifestations are diffuse goiter and solitary or multiple thyroid nodules. Thyroid function remains intact in the majority of cases, but sometimes it can result in hypothyroidism or hyperthyroidism. The diagnosis can be made after fine needle aspiration cytology, but usually it is diagnosed as an incidental finding while analyzing thyroidectomy tissue or during autopsy. Oral steroids are the cornerstone of thyroid sarcoidosis management, along with specific endocrinological treatment on some occasions. Given that we found only 71 reports of thyroid sarcoidosis available in the literature, we created and analyzed a cohort of 24 patients with thyroid sarcoidosis who were described in the literature in the 21st century and reported two additional cases of thyroid sarcoidosis.

Mesenchymal Stromal Cells from Healthy and Inflamed Human Gingiva Respond Differently to Porphyromonas gingivalis.

Gingiva-Derived Mesenchymal Stromal Cells (GMSCs) have been shown to play an important role in periodontitis. However, how P. gingivalis, one of the key etiological agents of the disease, affects healthy (H)- and periodontitis (P)-GMSCs is unknown. To address this problem, we established 10 H-GMSC and 12 P-GMSC lines. No significant differences in morphology, differentiation into chondroblasts and adipocytes, expression of characteristic MSCS markers, including pericyte antigens NG2 and PDGFR, were observed between H- and P-GMSC lines. However, proliferation, cell size and osteogenic potential were higher in P-GMSCs, in contrast to their lower ability to suppress mononuclear cell proliferation. P. gingivalis up-regulated the mRNA expression of IL-6, IL-8, MCP-1, GRO-α, RANTES, TLR-2, HIF-1α, OPG, MMP-3, SDF-1, HGF and IP-10 in P-GMSCs, whereas only IL-6, MCP-1 and GRO-α were up-regulated in H-GMSCs. The expression of MCP-1, RANTES, IP-10 and HGF was significantly higher in P-GMSCs compared to H-GMSCs, but IDO1 was lower. No significant changes in the expression of TLR-3, TLR-4, TGF-ß, LAP, IGFBP4 and TIMP-1 were observed in both types of GMSCs. In conclusion, our results suggest that P-GMSCs retain their pro-inflammatory properties in culture, exhibit lower immunosuppressive potential than their healthy counterparts, and impaired regeneration-associated gene induction in culture. All these functions are potentiated significantly by P. gingivalis treatment.

Protective Effect of an Exopolysaccharide Produced by Lactiplantibacillus plantarum BGAN8 Against Cadmium-Induced Toxicity in Caco-2 Cells.

Cadmium (Cd) ranks seventh on the list of most significant potential threats to human health based on its suspected toxicity and the possibility of exposure to it. It has been reported that some bacterial exopolysaccharides (EPSs) have the ability to bind heavy metal ions. We therefore investigated the capacity of eight EPS-producing lactobacilli to adsorb Cd in the present study, and Lactiplantibacillus plantarum BGAN8 was chosen as the best candidate. In addition, we demonstrate that an EPS derived from BGAN8 (EPS-AN8) exhibits a high Cd-binding capacity and prevents Cd-mediated toxicity in intestinal epithelial Caco-2 cells. Simultaneous use of EPS-AN8 with Cd treatment prevents inflammation, disruption of tight-junction proteins, and oxidative stress. Our results indicate that the EPS in question has a strong potential to be used as a postbiotic in combatting the adverse effects of Cd. Moreover, we show that higher concentrations of EPS-AN8 can alleviate Cd-induced cell damage.

Fecal microbiota composition associates with the capacity of human peripheral blood monocytes to differentiate into immunogenic dendritic cells in vitro.

Although promising for active immunization in cancer patients, dendritic cells (DCs) vaccines generated in vitro display high inter-individual variability in their immunogenicity, which mostly limits their therapeutic efficacy. Gut microbiota composition is a key emerging factor affecting individuals' immune responses, but it is unknown how it affects the variability of donors' precursor cells to differentiate into immunogenic DCs in vitro. By analyzing gut microbiota composition in 14 healthy donors, along with the phenotype and cytokines production by monocyte-derived DCs, we found significant correlations between immunogenic properties of DC and microbiota composition. Namely, donors who had higher α-diversity of gut microbiota and higher abundance of short-chain fatty acid (SCFAs) and SCFA-producing bacteria in feces, displayed lower expression of CD1a on immature (im)DC and higher expression of ILT-3, costimulatory molecules (CD86, CD40) proinflammatory cytokines (TNF-α, IL-6, IL-8) and IL-12p70/IL-10 ratio, all of which correlated with their lower maturation potential and immunogenicity upon stimulation with LPS/IFNγ, a well-known Th1 polarizing cocktail. In contrast, imDCs generated from donors with lower α-diversity and higher abundance of Bifidobacterium and Collinsella in feces displayed higher CD1a expression and higher potential to up-regulate CD86 and CD40, increase TNF-α, IL-6, IL-8 production, and IL-12p70/IL-10 ratio upon stimulation. These results emphasize the important role of gut microbiota on the capacity of donor precursor cells to differentiate into immunogenic DCs suitable for cancer therapy, which could be harnessed for improving the actual and future DC-based cancer therapies.

Cross-Talk Between Mesenchymal Stem/Stromal Cells and Dendritic Cells.

Mesenchymal stem/stromal cells (MSCs, having both multi-potent differentiation potential and prominent immunomodulatory properties, are seen as a very powerful tool for the therapy of diseases characterized by tissue damage and/or unregulated immune responses. Dendritic cells (DCs are key immunoregulatory cells at the crossroads between immunity and tolerance, able to fine-tune the whole immune response via regulation of adaptive immunity. Therefore, untangling the complex interactions between DCs and MSCs is crucial for understanding various mechanisms involved in the pathogenesis of immune-related diseases and for the discovery of new therapeutic targets for advanced treatment procedures. From this perspective, we reviewed the data that have been obtained to date regarding the complex effects of MSCs on DC development and functions, delineating the abundant mechanisms involved in these interactions. Additionally, we have pointed out to additional mechanisms of MSC/DC cross-talk that have not been directly proven, but that could have a significant role, not only in DC functions and the maintenance of immune homeostasis, but also in migration, differentiation and the functions of MSCs. For now, much more is known about the influence of MSCs on DCs than vice versa, so more studies should be done in order to fully understand this cross-talk.

Fast dendritic cells matured with Poly (I:C) may acquire tolerogenic properties.

BACKGROUND AIMS: Because of the labor-intensive and time-consuming conventional protocols for the generation of dendritic cells (DCs) as the most promising tools for anti-cancer therapy that enable the induction of a T-helper (Th)1-mediated anti-tumor immune response, the use of short-term protocols has been proposed. However, data on the applicability of such protocols in cancer immunotherapy are quite limited. METHODS: We compared the phenotypic and functional capability of fast DCs (fDCs) differentiated for 24 h and then matured for 48 h with Poly (I:C), a strong Th1-promoting agent, with donor-matched conventional DCs (cDCs) differentiated for 5 days and matured likewise. RESULTS: Of 12 donors tested, we identified seven whose monocytes failed to develop into immunogenic DCs through the use of fDC protocol, on the basis of incomplete downregulation of CD14, low expression of CD1a and macrophage-like morphology. Such fDCs have significantly lower expression of CD83, CD86, CCR7 and CD40, weaker allo-stimulatory Th1- and Th17-polarizing capacity caused by poor production of interleukin (IL)-12p70 and IL-23 and high production of IL-10, and prominent Th2-polarizing capacity, compared with donor-matched cDCs. Furthermore, such fDCs had tolerogenic properties as judged by higher expression of indolamine dioxigenase-3, IDO-1 and IL-1ß and induction of a higher percentage of CD4(+)CD25(+)FoxP3(+) T cells. These findings correlated with increased transforming growth factor (TGF)-ß production by fDC-primed CD3(+)T cells and their stronger anti-proliferative capacity. CONCLUSIONS: We emphasize that although fDCs could probably be applied as an alternative to cDCs for cancer therapy, the fDC protocol should not be applied to donors whose DCs acquire tolerogenic capabilities.

Size-dependent effects of gold nanoparticles uptake on maturation and antitumor functions of human dendritic cells in vitro.

Gold nanoparticles (GNPs) are claimed as outstanding biomedical tools for cancer diagnostics and photo-thermal therapy, but without enough evidence on their potentially adverse immunological effects. Using a model of human dendritic cells (DCs), we showed that 10 nm- and 50 nm-sized GNPs (GNP10 and GNP50, respectively) were internalized predominantly via dynamin-dependent mechanisms, and they both impaired LPS-induced maturation and allostimulatory capacity of DCs, although the effect of GNP10 was more prominent. However, GNP10 inhibited LPS-induced production of IL-12p70 by DCs, and potentiated their Th2 polarization capacity, while GNP50 promoted Th17 polarization. Such effects of GNP10 correlated with a stronger inhibition of LPS-induced changes in Ca2+ oscillations, their higher number per DC, and more frequent extra-endosomal localization, as judged by live-cell imaging, proton, and electron microscopy, respectively. Even when released from heat-killed necrotic HEp-2 cells, GNP10 inhibited the necrotic tumor cell-induced maturation and functions of DCs, potentiated their Th2/Th17 polarization capacity, and thus, impaired the DCs' capacity to induce T cell-mediated anti-tumor cytotoxicity in vitro. Therefore, GNP10 could potentially induce more adverse DC-mediated immunological effects, compared to GNP50.

Mesenchymal stem cells from periapical lesions modulate differentiation and functional properties of monocyte-derived dendritic cells.

Immunoregulatory mechanisms within periapical lesions (PLs) are as of yet unexplored. Considering the crucial role of DCs in controlling the immune response within PLs, the immunomodulatory properties of mesenchymal stem cells (MSCs), and the colocalization of MSCs and DCs in situ, we wondered whether MSCs from PLs modulate the development and functions of DCs. Using a model of monocyte-derived DCs, we showed that PL-MSCs inhibited differentiation of DCs via soluble factors, of which IL-6 had a minor effect, but did not impair their subsequent maturation induced by pro-inflammatory cytokines. However, upon maturation such DCs favored the production of Th2/Th17 cytokines by allogenic CD4(+) lymphocytes in coculture, compared with mature DCs differentiated without PL-MSCs. PL-MSC-differentiated DCs, cultivated with pro-inflammatory cytokines and PL-MSCs, although phenotypically mature, exhibited poor allostimulatory activity, induced anergy, Th2 polarization, differentiation of suppressive CD4(+) CD25(high) CD39(+) Treg-cell subsets via IDO-1-, ILT-3-, and ILT-4-dependent mechanisms, and increased production of TGF-ß in the coculture. In contrast, DCs cultivated with PL-MSCs only during maturation stimulated proliferation and Th1 polarization of CD4(+) T cells in an IL-12-independent manner. In conclusion, PL-MSCs significantly modulate the development and functions of DCs, depending on the phase of DCs development during which the interaction occurs.

Characterization and immunosuppressive properties of mesenchymal stem cells from periapical lesions.

AIM: Mesenchymal stem cells (MSCs) isolated from healthy dental tissues are being investigated as an alternative source of MSCs for the treatment of damaged tissues and inflammatory diseases. Here we investigated whether MSCs from periapical lesions (PL-MSCs) also possess multi-lineage differentiation capacity and immunomodulatory properties. MATERIAL & METHODS: PL-MSCs, isolated by collagenase/DNAse digestion from surgically extracted PLs, were compared with MSCs from non-inflamed dental pulp (DP-MSCs) and dental follicle (DF-MSCs) for their phenotype and multi-potent differentiation potential. The anti-inflammatory and immunomodulatory effects of PL-MSCs were studied in co-culture with peripheral blood mononuclear cells (PB-MNCs) and PL-inflammatory cells (PL-ICs). RESULTS: PL-MSCs were characterized by typical MSCs phenotype, lower clonogenicity and self-renewal rate, compared to DF-MSCs and DP-MSCs. These cells possess the potential to differentiate into adipocyte-, osteoblast- and chondrocyte-like cells in vitro, which differs from that of DP-MSCs and DF-MSCs. PL-MSCs inhibited phytohemaglutinine-induced proliferation of PB-MNCs and production of IL-2, IFNγ and IL-5 in the co-culture, probably via TGF-ß-dependent mechanisms. These cells also suppressed the production of IL-1ß, IL-6, and TNF-α by PL-ICs via soluble mediators, whereas the suppression of IL-8 production required a direct cell-to-cell contact. CONCLUSION: The differentiation potential of PL-MSCs and their immunosuppressive/anti-inflammatory properties could be beneficial for the treatment of chronic periodontal diseases.