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-β, 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-β) 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-β+ 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.

Dapsone is a sulfone used in treating inflammatory skin conditions. Despite its widespread dermatological use, the pharmacological actions of dapsone remain poorly understood. Here, we examined how different aspects of neutrophil functions are affected by dapsone. Peripheral blood neutrophils from healthy donors were stimulated with phorbol-12-myristate-13-acetate (PMA), N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP), or calcium ionophore (CaI) or primed with cytokines prior to stimulation, in the presence of different concentrations of dapsone (from 10 to 50 µg/mL), followed by analyses of their survival, phenotype, and functional properties. We found that dapsone at the concentration of 50 μg/mL induced a significant neutrophil apoptotic rate during 6 h and 18 h, while other concentrations were well tolerated compared to control non-treated cells. However, dapsone significantly decreased the induced oxidative burst of neutrophils at all non-cytotoxic concentrations. Additionally, dapsone showed a dose-dependent suppression of NETosis in activated neutrophils. The production of IL-8 by dapsone-treated neutrophils was decreased under both stimulated (fMLP) and primed (TNF-α/fMLP) conditions. Moreover, dapsone inhibited the expression of CD11b/CD18, CD66, and CD89 and reversed or significantly mitigated the downregulation of CD16, CD32, CD181, CD88, and CD62L on neutrophils after priming and fMLP stimulation. In conclusion, our results indicate the complexity of dapsone actions on neutrophil functions, extending previous knowledge on the suppression of oxidative burst and IL-8 production upon neutrophils’ activation. Suppressed NETosis and modulation of marker expression associated with different neutrophil functions under inflammatory conditions are new findings, not recognized previously.

Dysfunction of neutrophils in patients with cystic fibrosis (CF) is best characterized in bronchoalveolar lavage (BAL), whereas peripheral blood neutrophils are less examined, and the results are contradictory, especially in younger populations. Therefore, this work aimed to study functional and phenotypic changes in circulating neutrophils in children with CF. The study included 19 CF children (5–17 years) and 14 corresponding age-matched healthy children. Isolated neutrophils were cultured either alone or with different stimuli. Several functions were studied: apoptosis, NET-osis, phagocytosis, and production of reactive oxygen species (ROS), neutrophil elastase (NE), and 11 cytokines. In addition, the expression of 20 molecules involved in different functions of neutrophils was evaluated by using flow cytometry. CF neutrophils showed reduced apoptosis and lower production of NE and IL-18 compared to the healthy controls, whereas IL-8 was augmented. All of these functions were further potentiated after neutrophil stimulation, which included higher ROS production and the up-regulation of CD11b and IL-10 expression. NET-osis was higher only when neutrophils from moderate–severe CF were treated with Pseudomonas aeruginosa, and the process correlated with forced expiratory volume in the first second (FEV1). Phagocytosis was not significantly changed. In conclusion, circulating neutrophils from children with CF showed fewer impaired changes in phenotype than in function. Functional abnormalities, which were already present at the baseline levels in neutrophils, depended on the type of stimuli that mimicked different activation states of these cells at the site of infection.

Introduction. GLP-1R agonists (GLP-1RAs) are a type of anti-hyperglycemic medications used for the management of type 2 diabetes mellitus (T2DM). In addition to glucose-lowering effects, GLP-1R agonists (GLP-1RAs) also provide further advantages by promoting weight loss and controlling blood pressure. Furthermore, GLP-1RAs have been reported for their therapeutic benefits in neurological, cardiovascular, endocrine, and metabolic diseases. Emerging evidence from many clinical and experimental studies, as well as in vitro researches, suggests that GLP-1RA agonists may reduce inflamma tion and modify the immunological response, but the underlying molecu lar mechanisms remain unclear. In this research, we examined the effect of GLP-1RA on the cytotoxicity, proliferative activity, and cytokine production of human peripheral blood mononuclear cells (PBMCs) in vitro. Methods. The peripheral blood mononuclear cells (PBMCs) cultures from healthy volunteers were used. GLP-1RA (DMB), in different concentrations, was added to the cell cultures. A flow cytometry assay was used to assess cell viability, while MTT and CFSE dye dilution assays were used to quantify proliferative activity. Cytokine levels were measured using a sandwich en zyme-linked immunosorbent assay (ELISA). Results. The cytotoxicity data demonstrated that only the highest DMB concentration (1000 nM) decreased the metabolic activity (viability) of PBMCs (p<0.05) in comparison to untreated cultures, while concentra tions of 200 nM (p<0.05) and 1000 nM (p<0.001) of GLP-1RA increased the proportion of late apoptotic and necrotic cells (p<0.001). The prolif eration of PBMCs was significantly decreased in the presence of GLP-1RA at a 100 nM concentration when compared to the control (p<0.05). The same concentration of GLP-1RA significantly reduced the production of IL-6, IL-1β, and TNF-α. Conclusion. Our results suggested that GLP-1RA (DMB), at a non-toxic concentration of 100 nM, inhibited the proliferation and production of pro-inflammatory cytokines by human PBMCs in vitro. These findings open the perspective to study the immune response in detail in future experiments.

Introduction. Knowledge of orbital anthropometry is essential for reconstructive surgery, ophthalmology and forensic identification, yet normative multislice computed tomography (MSCT) data for this region are scarce. The aim was to determine the principal orbital parameters in adults without orbital pathology and to examine sex, age and side differences. Method. In this retrospective study, 154 orbits in 77 adults (51 men, 26 women; aged 24-86 years) were measured on three-dimensional reconstructions from a 64-slice MSCT scanner. Orbital height, width, volume, orbital index and the interorbital and biorbital distances were recorded and compared by sex, age group and side. Results. Men had significantly greater orbital width, volume and biorbital and interorbital distances than women bilaterally (p ≤ 0.009), whereas orbital height did not differ by sex. The right orbital index was higher in women (90.02 ± 6.45 versus 86.36 ± 7.26; p = 0.034). In the older group the right orbit showed greater height, volume and orbital index (p ≤ 0.031), with no significant age effect on the left. The right orbit was significantly wider and had a higher orbital index than the left (p ≤ 0.011), while orbital volume was symmetrical (p = 0.099). Conclusion. Orbital morphometry is sex-, age- and side-dependent, and the MSCT-derived values reported here provide population-specific reference data for clinical and forensic use.

Sitagliptin is a dipeptidyl peptidase-4 (DPP-4) inhibitor used to treat type 2 diabetes. However, several studies have demonstrated its anti-inflammatory and immunomodulatory properties. The aim of this study was to investigate the effect of sitagliptin on the functional and phenotypic properties of human neutrophils under normal (NG, 5.5 mM)- and high (HG, 22 mM)-glucose conditions in vitro. Neutrophils were pretreated with varying concentrations of sitagliptin and stimulated with phorbol-12-myristate-13-acetate (PMA), N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP), calcium ionophore (CaI), or opsonized zymosan (OpZym). Survival, phenotypic, and functional characteristics were then assessed. Our results showed that sitagliptin was non-cytotoxic to neutrophils even at very high concentrations. It decreased the production of reactive oxygen species (ROS) and neutrophil extracellular traps (NETs), generally following a stimulus- and concentration-dependent pattern. The effect was more pronounced under HG conditions. Furthermore, sitagliptin showed a significant ROS-scavenging effect in a cell-free system. It also rapidly altered the expression of surface markers in both resting and fMLP-stimulated neutrophils, typically upregulating CD10, CD16, CD62L, CD63, CD88, CD89, and PD-L1, and downregulating CD11b/CD18, CD66b, and CD182, a phenotype consistent with a dampened, less-primed activation state of these cells. In conclusion, sitagliptin exhibited marked antioxidative/ROS-scavenging activity in neutrophil cultures and induced a coordinated shift in neutrophil phenotype, accompanied by suppression of NETosis under both NG and HG conditions. Collectively, these data support the view that neutrophils may constitute an additional cellular target contributing to sitagliptin’s anti-inflammatory and immunomodulatory profile.