Effect of cDMEM media containing Ectoine on human periodontal ligament mesenchymal stem cell survival and differentiation.

BACKGROUND/AIM: Ectoine is an amino acid that can preserve osmotic balance and has more preservative activity than other osmoregulators. There are no published reports on the osmoregulators' effects on viability or differentiation of dental stem cells. The aim of this study was to investigate the effect of Ectoine as a storage media on the viability and differentiation potential of human periodontal ligament mesenchymal stem cells (hPDLMSCs). MATERIALS AND METHODS: hPDLMSCs were obtained from impacted third molar teeth. The cells were isolated, submitted to trilineage differentiation, and characterized by flow cytometer (FC). hPDLMSCs were incubated with different media containing Ectoine, complete DMEM (cDMEM), Ectoine+cDMEM, milk, and tap water for 2, 6, 12, and 24 h. The cells were analyzed by FC for viability, early-apoptosis, late apoptosis, and necrosis rates. Osteogenic and fibroblastic differentiation in hPDLMSCs were investigated by Alizarin red stain and vimentin expression. RESULTS: All treated groups showed significant decreases in cell viability after 2 h. Significant increases were detected in the number of dead cells between 2 and 12 h in all groups except the Ectoine+cDMEM group. The deposition of mineral matrix nodules was significantly higher in cells cultured with Ectoine+cDMEM compared with the other media. Higher vimentin expressions were detected in cells cultured with cDMEM and Ectoine+cDMEM media, respectively. CONCLUSIONS: Ectoine added to cDMEM media, promoted cell survival plus osteogenic and fibroblastic differentiation of hPDLMSCs.

Targeting the undruggable oncogenic KRAS: the dawn of hope.

KRAS mutations are the drivers of various cancers, including non-small cell lung cancer, colon cancer, and pancreatic cancer. Over the last 30 years, immense efforts have been made to inhibit KRAS mutants and oncogenic KRAS signaling using inhibitors. Recently, specific targeting of KRAS mutants with small molecules revived the hopes for successful therapies for lung, pancreatic, and colorectal cancer patients. Moreover, advances in gene editing, protein engineering, and drug delivery formulations have revolutionized cancer therapy regimens. New therapies aim to improve immune surveillance and enhance antitumor immunity by precisely targeting cancer cells harboring oncogenic KRAS. Here, we review recent KRAS-targeting strategies, their therapeutic potential, and remaining challenges to overcome. We also highlight the potential synergistic effects of various combinatorial therapies in preclinical and clinical trials.

Effects of different detergent-containing children's toothpastes on the viability, osteogenic and chondrogenic differentiation of human dental periodontal ligament stem cells and gingival stem cells in vitro.

BACKGROUND: Detergents are the most commonly used compounds in toothpastes due to their foaming and cleaning peoperties. This study aimed to investigate the effects of children's toothpastes with different detergent content on the viability, the osteogenic and chondrogenic differentiation potentials of human mesenchymal stem cells. METHODS: The necessary tissues for human periodontal ligament mesenchymal stem cells (hPDLMSCs) and human gingival mesenchymal stem cells (hGMSCs) isolation were obtained during extraction of 10 impacted third molar teeth. The viability of the cells stimulated with different concentratiaons of Colgate, Sensodyne, Splat, Nenedent, Perlodent toothpaste solutions and complete Dulbocco's modified eagle medium (control group) were evaluated by using the flow cytometer. In addition, the osteogenic and chondrogenic differentiation potential of human gingival and periodontal ligament mesenchymal stem cells exposed to toothpaste solutions were examined morphologically. Datas were analyzed with IBM SPSS V23. One way ANOVA test was used to determine the differences between the groups for multiple comparisons, while the Tukey post-hoc test was used for pair wise comparisons in determining which groups differed. RESULTS: A higher percentage of cell viability was detected in Control group at 20 %, 50 % and 80 % (p = 0.000) on hGMSCs. After the Control group, the highest cell viability ratios were observed in the detergent-free Splat group (p = 0.000) followed by the Sensodyne experimental group containing CABP (p = 0.000). While the cell viability rates in Nenedent group was found significantly higher than the Perlodent group at other concentrations except for 20 % concentration (p = 0.000). Colgate group had the lowest percentage of cell viability among the experimental groups at all concentrations on hPDMSCs (p = 0.000). The highest live cell ratios was detected in Control group (p = 0.000), followed by Splat and Sensodyne groups (p = 0.000). The cell viability ratios at 50 % concentration were higher in Perlodent group than Nenedent group (p = 0.000). The highest osteogenic and chondrogenic differentiation potential of mesenchymal stem cells stimulated with different toothpaste was determined in Control and Splat group. CONCLUSIONS: As a result of the findings, it was observed that toothpaste containing SLS had a more negative effect on the viability of the cells and the differentiation potentials than the other groups.

Cytotoxicity of NeoMTA Plus, ProRoot MTA and Biodentine on human dental pulp stem cells.

BACKGROUND/PURPOSE: Dental pulp stem cells (DPSCs) play a crucial role in the tissue healing process through odontoblast like cell differentiation. The aim of this study was to evaluate the biocompatibility and compare the potential invitro cytotoxic effects of NeoMTA Plus, ProRootMTA and Biodentine on human dental pulp stem cells (hDPSCs). MATERIALS AND METHODS: To assess the effects of NeoMTA Plus, ProRoot MTA and Biodentine extracts at 1st, 3rd and 7th d on hDPCs, cell populations was determined by flow cytometry using an Annexin V detection kit. The data were analyzed statistically using the Kruskal-Wallis test. A p < 0.05 was considered as statistically significant. RESULTS: All groups showed cell viability similar to that of the control group on 1st, 3rd and 7th d. Although Biodentine exhibited higher cell viability rates than the other material groups, no statistically significant differences were noted between the sampled days (p > 0.05). CONCLUSION: All materials extracts are not cytotoxic and do not induce apoptosis in the hDPSCs. These results suggest that all the tested materials can lead to positive outcomes when used as reparative biomaterials.

Reduced regulatory T cells with increased proinflammatory response in patients with schizophrenia.

AIM: To investigate whether circulating T cells including regulatory T cells (Treg) and derived cytokines contribute to the immune imbalance observed in schizophrenia. METHODS: Forty patients with schizophrenia and 40 age, sex, body mass index, education, and smoking status-matched healthy controls (HC) are included in the study. We stained cells with anti-CD14, anti-CD3, anti-CD4, anti-CD8, anti-CD19, anti-CD20, and anti-CD16/56. Peripheral blood mononuclear cells (PBMCs) were isolated and stained with the human FoxP3 kit containing anti-CD4/anti-CD25 and intracellular anti-Foxp3. PBMCs were cultured for 72 h and stimulated with anti-CD3/anti-CD28. Cytokines (IL-2, IL-4, IL-6, IL-10, IFN-γ, TNF-α, and IL-17A) were measured from the culture supernatant and plasma using the Th1/Th2/Th17 cytokine bead array kit. RESULTS: In comparison with HC, Treg percentages in schizophrenia were higher (1.17 ± 0.63 vs 0.81 ± 0.53, P = 0.005) in unstimulated but lower in the stimulated condition (0.73 ± 0.69 vs 0.97 ± 0.55, P = 0.011). Activated T cell percentages were higher in schizophrenia than HC in unstimulated (2.22 ± 0.78 vs 1.64 ± 0.89, P = 0.001) and stimulated (2.25 ± 1.01 vs 1.72 ± 1.00, P = 0.010) conditions. The culture supernatant levels of IL-6 (7505.17 ± 5170.07 vs 1787.81 ± 1363.32, P < 0.001), IL-17A (191.73 ± 212.49 vs 46.43 ± 23.99, P < 0.001), TNF-α (1557 ± 1059.69 vs 426.57 ± 174.62, P = 0.023), and IFN-γ (3204.13 ± 1397.06 vs 447.79 ± 270.13, P < 0.001); and plasma levels of IL-6 (3.83 ± 3.41vs 1.89 ± 1.14, P = 0.003) and IL-17A (1.20 ± 0.84 vs 0.83 ± 0.53, P = 0.033) were higher in patients with schizophrenia than HC. CONCLUSION: Our explorative study shows reduced level of Foxp3 expressing Treg in a stimulated condition with induced levels of proinflammatory cytokines in patients with schizophrenia.

Mechanisms of T-Cell Exhaustion in Pancreatic Cancer.

T-cell exhaustion is a phenomenon that represents the dysfunctional state of T cells in chronic infections and cancer and is closely associated with poor prognosis in many cancers. The endogenous T-cell immunity and genetically edited cell therapies (CAR-T) failed to prevent tumor immune evasion. The effector T-cell activity is perturbed by an imbalance between inhibitory and stimulatory signals causing a reprogramming in metabolism and the high levels of multiple inhibitory receptors like programmed cell death protein-1 (PD-1), cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), T cell immunoglobulin and mucin domain-containing protein 3 (TIM-3), and Lymphocyte-activation gene 3 (Lag-3). Despite the efforts to neutralize inhibitory receptors by a single agent or combinatorial immune checkpoint inhibitors to boost effector function, PDAC remains unresponsive to these therapies, suggesting that multiple molecular mechanisms play a role in stimulating the exhaustion state of tumor-infiltrating T cells. Recent studies utilizing transcriptomics, mass cytometry, and epigenomics revealed a critical role of Thymocyte selection-associated high mobility group box protein (TOX) genes and TOX-associated pathways, driving T-cell exhaustion in chronic infection and cancer. Here, we will review recently defined molecular, genetic, and cellular factors that drive T-cell exhaustion in PDAC. We will also discuss the effects of available immune checkpoint inhibitors and the latest clinical trials targeting various molecular factors mediating T-cell exhaustion in PDAC.