Orofacial clefts lead to increased pro-inflammatory cytokine levels on neonatal oral mucosa.

Orofacial clefts (OFC) are frequent congenital malformations characterized by insufficient separation of oral and nasal cavities and require presurgical infant orthopedics and surgical interventions within the first year of life. Wound healing disorders and higher prevalence of gingivitis and plaque levels are well-known challenges in treatment of children with OFC. However, oral inflammatory mediators were not investigated after birth using non-invasive sampling methods so far. In order to investigate the impact of OFC on oral cytokine levels, we collected tongue smear samples from 15 neonates with OFC and 17 control neonates at two time points (T), T0 at first consultation after birth, and T1, 4 to 5 weeks later. The samples were analyzed using multiplex immunoassay. Overall, we found significantly increased cytokine levels (TNF, IL-1ß/-2/-6/-8/-10) in tongue smear samples from neonates with OFC compared to controls, especially at T0. The increase was even more pronounced in neonates with a higher cleft severity. Further, we detected a significant positive correlation between cleft severity score and distinct pro-inflammatory mediators (GM-CSF, IL-1ß, IL-6, IL-8) at T0. Further, we found that breast-milk (bottle) feeding was associated with lower levels of pro-inflammatory cytokines (IL-6/-8) in neonates with OFC compared to formula-fed neonates. Our study demonstrated that neonates with OFC, especially with high cleft severity, are characterized by markedly increased inflammatory mediators in tongue smear samples within the first weeks of life potentially presenting a risk for oral inflammatory diseases. Therefore, an inflammatory monitoring of neonates with (severe) OFC and the encouragement of mother to breast-milk (bottle) feed might be advisable after birth and/or prior to cleft surgery.

Leukemia derived dendritic cell (DCleu) mediated immune response goes along with reduced (leukemia-specific) regulatory T-cells.

The blastmodulatory Kit-M, composed of granulocyte-macrophage colony-stimulating-factor (GM-CSF) and Prostaglandin E1 (PGE1), is known to convert myeloid leukaemic blasts (from AML patients) into leukaemia derived dendritic cells (DCleu), which activate immunoreactive cells to gain antileukemic/leukaemia-specific activity. In this study we had a special focus on the influence of Kit-M treated, DC/DCleu containing patients'whole blood (WB, n = 16) on the provision of immunosuppressive regulatory T-cells. We could confirm that Kit-M significantly increased frequencies of (mature) dendritic cells (DC) and DCleu from leukemic whole blood (WB) without induction of blast proliferation. After mixed lymphocyte culture (MLC) with patients' T-cells we confirmed that DCleu mediated leukemia-specific responses- going along with activated and leukemia-specific T- and NK-cells in an intracellular cytokine staining assay (ICS) and a degranulation assay (Deg)- resulted in an increased anti-leukemic cytotoxicity (Cytotoxicity Fluorolysis Assay = CTX). We could demonstrate that (leukemia-specific) CD4+ and CD8+ regulatory T-cell population (Treg) decreased significantly after MLC compared to controls. We found significant positive correlations of leukemia-specific CD3+CD4+ cells with frequencies of (mature) DCleu. Achieved anti-leukemic cytotoxicity correlated significantly positive with leukemia-specific CD3+CD8+ cells and significantly negatively with (leukemia-specific) Treg. In summary we demonstrate that immunesuppressive (leukemia-specific) regulatory T-cells are significantly downregulated after Kit-M triggered MLC- going along with a (reinstalled) antileukemic reactivity of the immune system (as demonstrated with functional assays ICS, Deg, CTX).

Dendritic Cell-Triggered Immune Activation Goes along with Provision of (Leukemia-Specific) Integrin Beta 7-Expressing Immune Cells and Improved Antileukemic Processes.

Integrin beta 7 (ß7), a subunit of the integrin receptor, is expressed on the surface of immune cells and mediates cell-cell adhesions and interactions, e.g., antitumor or autoimmune reactions. Here, we analyzed, whether the stimulation of immune cells by dendritic cells (of leukemic derivation in AML patients or of monocyte derivation in healthy donors) leads to increased/leukemia-specific ß7 expression in immune cells after T-cell-enriched mixed lymphocyte culture-finally leading to improved antileukemic cytotoxicity. Healthy, as well as AML and MDS patients' whole blood (WB) was treated with Kit-M (granulocyte-macrophage colony-stimulating factor (GM-CSF) + prostaglandin E1 (PGE1)) or Kit-I (GM-CSF + Picibanil) in order to generate DCs (DCleu or monocyte-derived DC), which were then used as stimulator cells in MLC. To quantify antigen/leukemia-specific/antileukemic functionality, a degranulation assay (DEG), an intracellular cytokine assay (INTCYT) and a cytotoxicity fluorolysis assay (CTX) were used. (Leukemia-specific) cell subtypes were quantified via flow cytometry. The Kit treatment of WB (compared to the control) resulted in the generation of DC/DCleu, which induced increased activation of innate and adaptive cells after MLC. Kit-pretreated WB (vs. the control) led to significantly increased frequencies of ß7-expressing T-cells, degranulating and intracellular cytokine-producing ß7-expressing immune cells and, in patients' samples, increased blast lysis. Positive correlations were found between the Kit-M-mediated improvement of blast lysis (vs. the control) and frequencies of ß7-expressing T-cells. Our findings indicate that DC-based immune therapies might be able to specifically activate the immune system against blasts going along with increased frequencies of (leukemia-specific) ß7-expressing immune cells. Furthermore, ß7 might qualify as a predictor for the efficiency and the success of AML and/or MDS therapies.

Expression of Surface-associated 82 kDa proMMP-9 in Lymphatic Leukemia Blast Cells Differentially Correlates With Prognosis.

BACKGROUND/AIM: Matrix metalloproteinases (MMPs) degrade extracellular matrix and process regulatory proteins. Recently, a membrane-bound 82kDa variant of proMMP-9 identified on myeloid blasts was shown to be associated with prognosis. PATIENTS AND METHODS: To investigate the role of 82kDa proMMP-9 with acute lymphoblastic leukemia (ALL) and chronic lymphoid leukemia (CLL), we performed flow-cytometry analysis of expression on ALL blasts (n=18) and CLL lymphocytes (n=21) from blood and correlated data with clinical parameters. RESULTS: In ALL, mature B-linear blasts expressed higher levels of 82kDa proMMP-9 compared to T-linear blasts. Elevated levels of 82kDa proMMP-9 were found in elderly patients and at patients with relapse. No correlation was observed on blood cells and extramedullary disease. In CLL, the 82kDa proMMP-9 expression did not correlate with any of the clinical parameters. CONCLUSION: Our findings suggest that higher levels of 82kDa proMMP-9 expression on blast cells may correlate with a more unfavorable ALL-subtype. Further studies are required to clarify the prognostic role of the 82kDa pro-MMP-9 expression.

Influence of Natural Killer Cells and Natural Killer T Cells on Periodontal Disease: A Systematic Review of the Current Literature.

Natural killer (NK) cells, as members of the innate immune system, and natural killer T (NKT) cells, bridging innate and adaptive immunity, play a prominent role in chronic inflammatory diseases and cancerogenesis, yet have scarcely been examined in oral diseases. Therefore, systematic research on the latest literature focusing on NK/NKT cell-mediated mechanisms in periodontal disease, including the time period 1988-2020, was carried out in MEDLINE (PubMed) using a predetermined search strategy, with a final selection of 25 studies. The results showed that NK cells tend to have rather proinflammatory influences via cytokine production, cytotoxic effects, dendritic-cell-crosstalk, and autoimmune reactions, while contrarily, NKT cell-mediated mechanisms were proinflammatory and immunoregulatory, ranging from protective effects via B-cell-regulation, specific antibody production, and the suppression of autoimmunity to destructive effects via cytokine production, dendritic-cell-crosstalk, and T-/B-cell interactions. Since NK cells seem to have a proinflammatory role in periodontitis, further research should focus on the proinflammatory and immunoregulatory properties of NKT cells in order to create, in addition to antibacterial strategies in dental inflammatory disease, novel anti-inflammatory therapeutic approaches modulating host immunity towards dental health.