Прегледај по Аутор "Mihajlović, Dušan"

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    Fecal microbiota composition associates with the capacity of human peripheral blood monocytes to differentiate into immunogenic dendritic cells in vitro
    (2021) Radojević, Dušan; Tomić, Sergej; Mihajlović, Dušan; Tolinački, Maja; Pavlović, Bojan; Vučević, Dragana; Bojić, Svetlana; Golić, Nataša; Čolić, Miodrag; Đokić, Jelena
    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.
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    Functionalization-dependent effects of cellulose nanofibrils on tolerogenic mechanisms of human dendritic cells
    (Dove Medical Press, 2018) Tomić, Sergej; Ilić, Nataša; Kokol, Vanja; Gruden-Movsesijan, Alisa; Mihajlović, Dušan; Bekić, Marina; Sofronić-Milosavljević, Ljiljana; Čolić, Miodrag; Vučević, Dragana
    Background: Cellulose nanofibrils (CNF) are attractive nanomaterials for various biomedical applications due to their excellent biocompatibility and biomimetic properties. However, their immunoregulatory properties are insufficiently investigated, especially in relation to their functionalization, which could cause problems during their clinical application. Methods: Using a model of human dendritic cells (DC), which have a central role in the regulation of immune response, we investigated how differentially functionalized CNF, ie, native (n) CNF, 2,2,6,6-tetramethylpiperidine 1-oxyl radical-oxidized (c) CNF, and 3-aminopropylphosphoric acid-functionalized (APAc) CNF, affect DC properties, their viability, morphology, differentiation and maturation potential, and the capacity to regulate T cell-mediated immune response. Results: Nontoxic doses of APAcCNF displayed the strongest inhibitory effects on DC differentiation, maturation, and T helper (Th) 1 and Th17 polarization capacity, followed by cCNF and nCNF, respectively. These results correlated with a specific pattern of regulatory cytokines production by APAcCNF-DC and their increased capacity to induce suppressive CD8+CD25+IL-10+ regulatory T cells in immunoglobulin-like transcript (ILT)-3- and ILT-4-dependent manner. In contrast, nCNF-DC induced predominantly suppressive CD4+CD25hiFoxP3hi regulatory T cells in indolamine 2,3-dioxygenase-1-dependent manner. Different tolerogenic properties of CNF correlated with their size and APA functionalization, as well as with different expression of CD209 and actin bundles at the place of contact with CNF. Conclusion: The capacity to induce different types of DC-mediated tolerogenic immune responses by functionalized CNF opens new perspectives for their application as well-tolerated nanomaterials in tissue engineering and novel platforms for the therapy of inflammatory T cell-mediated pathologies.
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    Phosphonate-Modified Cellulose Nanocrystals Potentiate the Th1 Polarising Capacity of Monocyte-Derived Dendritic Cells via GABA-B Receptor
    (Dove Medical Press, 2022) Bekić, Marina; Vasiljević, Miloš; Stojanović, Dušica; Kokol, Vanja; Mihajlović, Dušan; Vučević, Dragana; Uskoković, Petar; Čolić, Miodrag; Tomić, Sergej
    Purpose: Phosphonates, like 3-AminoPropylphosphonic Acid (ApA), possess a great potential for the therapy of bone tumours, and their delivery via cellulose nanocrystals (CNCs) seems a promising approach for their increased efficacy in target tissues. However, the immunological effects of CNC-phosphonates have not been investigated thoroughly. The main aim was to examine how the modification of CNCs with phosphonate affects their immunomodulatory properties in human cells. Methods: Wood-based native (n) CNCs were modified via oxidation (ox-CNCs) and subsequent conjugation with ApA (ApA-CNCs). CNCs were characterised by atomic force microscopy (AFM) and nanoindentation. Cytotoxicity and immunomodulatory potential of CNCs were investigated in cultures of human peripheral blood mononuclear cells (PBMCs) and monocyte-derived dendritic cells (MoDCs)/T cells co-cultures by monitoring phenotype, cytokines production, allostimulatory and Th/Treg polarisation capacity. Results: AFM showed an increase in CNCs' thickens, elasticity modulus and hardness during the modification with ApA. When applied at non-toxic doses, nCNCs showed a tolerogenic potential upon internalisation by MoDCs, as judged by their increased capacity to up-regulate tolerogenic markers and induce regulatory T cells (Treg), especially when present during the differentiation of MoDCs. In contrast, ox- and ApA-CNCs induced oxidative stress and autophagy in MoDCs, which correlated with their stimulatory effect on the maturation of MoDCs, but also inhibition of MoDCs differentiation. ApA-CNC-treated MoDCs displayed the highest allostimulatory and Th1/CTL polarising activity in co-cultures with T cells. These effects of ApA-CNCs were mediated via GABA-B receptor-induced lowering of cAMP levels in MoDCs, and they could be blocked by GABA-B receptor inhibitor. Moreover, the Th1 polarising and allostimulatory capacity of MoDCs differentiated with ApA-CNC were largely preserved upon the maturation of MoDCs, whereas nCNC- and ox-CNC-differentiated MoDCs displayed an increased tolerogenic potential. Conclusion: The delivery of ApA via CNCs induces potent DC-mediated Th1 polarisation, which could be beneficial in their potential application in tumour therapy.
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    Production of immunoregulatory cytokines in clinically asymptomatic periapical lesions depends on the lesions size
    (Ministry of Defance, Serbia, 2019) Duka, Miloš; Gazivoda, Dragan; Marković, Milan; Majstorović, Ivana; Tatomirović, Željka; Mihajlović, Dušan; Čolić, Miodrag
    Bacground/Aim. Development of periapical lesions (PLs) involves a complex cross-talk between pathogenic microor­ganisms from the root canal and host immune mechanisms. In these processes proinflammatory cytokines are involved in stimulation of inflammation and osteodestructive mecha­nisms, whereas anti-inflammatory cytokines, with the im­munoregulatory functions, have the opposite effects. How this balance is controlled is still the subject of numerous studies. The aim of this study was to examine whether the local production of interleukin (IL)-10, IL-27 and trans­forming growth factor (TGF)-β in human asymptomatic PLs depends on the lesion size and how levels of these im­munoregulatory cytokines correlate with the cellular compo­sition of PLs. Methods. The study was conducted on 30 PLs which were collected at the Clinic for Stomatology of the Military Medical Academy, Belgrade, Serbia. The PLs were divided according to their size into small- and large-size lesions (n = 12 and n = 18, respectively). The inflam­matory cells (PL-ICs) were isolated from PLs and cultivated for 24 hours in culture medium supplemented with phorbol myristate acetate and Ca2+ ionophore. Cytospins were proc­essed for immunocytology by using monoclonal antibodies to cell subsets. The levels of cytokines in culture super­natants were determined by the ELISA method. Statistical analysis was performed using the Student t-test and the Spearman’s correlation test. The values of p < 0.05 were considered to be significant. Results. The levels of IL-10 and TGF-β were significantly higher in the PL-ICs cultures of large-size lesions than in small ones (p < 0.01 and p < 0.05, respectively). In contrast, the levels of IL-27 were higher in the cultures of small-size lesions than in small ones (p < 0.05). Although the total number of PL-ICs and the proportion of mononuclear phagocytes were higher in the large-size PLs (p < 0.01 and p < 0.05, respectively), their main composition was not significantly different between the groups. The proportions of B cells/plasma cells (CD19/CD38+ cells), CD8+ T cells and CD14+ cells were significantly higher in the large-size PLs (p < 0.005; p < 0.05; p < 0.05, respectively). In contrast, the proportion of total T cells (CD3+ cells) was higher in the small-size lesions (p < 0.05). No statistically significant correlation was found between the levels of these cytokines and the composi­tion/phenotype of PL-ICs. Conclusion. This study sug­gests that IL-10, IL-27 and TGF-β may play different roles in suppression of unwanted immune/inflammatory reac­tions in asymptomatic PLs, depending on the extension of pathological process as judged by the size of lesions.
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    Sitagliptin Induces Tolerogenic Human Dendritic Cells
    (MDPI, 2023) Drakul, Marija; Tomić, Sergej; Bekić, Marina; Mihajlović, Dušan; Vasiljević, Miloš; Rakočević, Sara; Ðokić, Jelena; Popović, Nikola; Bokonjić, Dejan; Čolić, Miodrag
    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-y treatment for an additional 24 h was used for maturation induction of MoDCs. Sitagliptin was added at the highest non-cytotoxic concentration (500 g/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- , 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- ) 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- + 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.