Reconstitution of Monocyte Subsets and PD-L1 Expression but Not T Cell PD-1 Expression in Obstructive Sleep Apnea Patients upon PAP Therapy

Obstructive sleep apnea (OSA) is characterized by nocturnal breathing intermissions resulting in oxidative stress and eventually, a low-grade systemic inflammation. The study aimed to investigate the impact of positive airway pressure (PAP) therapy on the inflammatory milieu as measured by monocyte and T cell phenotypic alterations. Participants were assessed for their OSA severity before PAP therapy and about six months later, including patient-reported outcome and therapy usage by telemetry readout. The distributions of the CD14/CD16-characterized monocyte subsets as well as the CD4/CD8-characterized effector T cell subsets with regard to their PD-1 and PD-L1 expression were analyzed by flow cytometry from blood samples. Data of 25 patients revealed a significant reconstitution of the monocyte subset distribution and a decrease in PD-L1 expression on pan-monocytes and CD8⁺ T cells without an association to initial AHI and overweight. The PD-1 expression was still increased on T cell subsets, especially on CD4⁺ TH17/22 cells. We conclude that PAP therapy might have a rapid effect on the monocyte phenotype and overall PD-L1 expression levels. However, T cell immune alterations especially on TH17/22 cells persist longer, indicating an ongoing disturbance of the adaptive immune system.     

Prognostic and Therapeutic Implications of Immune Classification by CD8+ Tumor-Infiltrating Lymphocytes and PD-L1 Expression in Sinonasal Squamous Cell Carcinoma

Sinonasal squamous cell carcinoma (SNSCC) is an aggressive tumor predominantly arising in the maxillary sinus and nasal cavities. Advances in imaging, surgical and radiotherapeutic techniques have reduced complications and morbidity; however, the prognosis generally remains poor, with an overall 5-year survival rate of 30–50%. As immunotherapy may be a new therapeutic option, we analyzed CD8⁺ tumor-infiltrating lymphocytes (TILs) and the tumor microenvironment immune type (TMIT, combining CD8⁺ TILs and PD-L1) in a series of 57 SNSCCs. Using immunohistochemistry, tissue samples of 57 SNSCCs were analyzed for expression of CD8 on TILs and of PD-L1 on tumor cells. The results were correlated to the clinical and survival data. In total, 88% (50/57) of the tumors had intratumoral CD8⁺ TILs; 19% (11/57)—CD8^high (>10%); and 39/57 (68%)—CD8^low (1–10%). PD-L1 positivity (>5%) was observed in 46% (26/57) of the SNSCCs and significantly co-occurred with CD8⁺ TILs (p = 0.000). Using univariate analysis, high intratumoral CD8⁺ TILs and TMIT I (CD8^high/PD-L1^pos) correlated with a worse survival rate. These results indicate that SNSCCs are immunogenic tumors, similar to head and neck squamous cell carcinomas. Nineteen percent of the cases were both CD8^high and PD-L1^pos and this subgroup may benefit from therapy with immune checkpoint inhibitors.     

Systemic Inflammation and the Breakdown of Intestinal Homeostasis Are Key Events in Chronic Spinal Cord Injury Patients

Our aim was to investigate the subset distribution and function of circulating monocytes, proinflammatory cytokine levels, gut barrier damage, and bacterial translocation in chronic spinal cord injury (SCI) patients. Thus, 56 SCI patients and 28 healthy donors were studied. The levels of circulating CD14^+highCD16⁻, CD14^+highCD16⁺, and CD14^+lowCD16⁺ monocytes, membrane TLR2, TLR4, and TLR9, phagocytic activity, ROS generation, and intracytoplasmic TNF-α, IL-1, IL-6, and IL-10 after lipopolysaccharide (LPS) stimulation were analyzed by polychromatic flow cytometry. Serum TNF-α, IL-1, IL-6 and IL-10 levels were measured by Luminex and LPS-binding protein (LBP), intestinal fatty acid-binding protein (I-FABP) and zonulin by ELISA. SCI patients had normal monocyte counts and subset distribution. CD14^+highCD16⁻ and CD14^+highCD16⁺ monocytes exhibited decreased TLR4, normal TLR2 and increased TLR9 expression. CD14^+highCD16⁻ monocytes had increased LPS-induced TNF-α but normal IL-1, IL-6, and IL-10 production. Monocytes exhibited defective phagocytosis but normal ROS production. Patients had enhanced serum TNF-α and IL-6 levels, normal IL-1 and IL-10 levels, and increased circulating LBP, I-FABP, and zonulin levels. Chronic SCI patients displayed impaired circulating monocyte function. These patients exhibited a systemic proinflammatory state characterized by enhanced serum TNF-α and IL-6 levels. These patients also had increased bacterial translocation and gut barrier damage.     

Norcantharidin Induces Immunogenic Cell Death of Bladder Cancer Cells through Promoting Autophagy in Acidic Culture

The acidic tumor microenvironment stands as a major obstacle to the efficient elimination of tumor cells. Norcantharidin (NCTD) is a powerful antitumor agent with multiple bioactivities. However, the effect of NCTD under acidic conditions is still unclear. Here, we report that NCTD can efficiently kill bladder cancer (BC) cells in acidic culture, and more intriguingly, NCTD can induce immunogenic cell death (ICD), thereby promoting antitumor immunity. In NCTD-treated BC cells, the surface-exposed calreticulin (ecto-CALR) was significantly increased. Consistently, co-culture with these cells promoted dendritic cell (DC) maturation. The NCTD-induced ICD is autophagy dependent, as autophagy inhibition completely blocked the NCTD-induced ecto-CALR and DC maturation. In addition, the DC showed a distinct maturation phenotype (CD80^high CD86^low) in acidic culture, as compared to that in physiological pH (CD⁸⁰ high CD86^high). Finally, the NCTD-induced ICD was validated in a mouse model. NCTD treatment significantly increased the tumor-infiltrating T lymphocytes in MB49 bladder cancer mice. Immunizing mice with NCTD-treated MB49 cells significantly increased tumor-free survival as compared to control. These findings demonstrate that NCTD could induce ICD in an acidic environment and suggest the feasibility to combine NCTD with anticancer immunotherapy to treat BC.     

CD44 Expression Intensity Marks Colorectal Cancer Cell Subpopulations with Different Extracellular Vesicle Release Capacity

Extracellular vesicles (EV) are released by virtually all cells and they transport biologically important molecules from the release site to target cells. Colorectal cancer (CRC) is a leading cause of cancer-related death cases, thus, it represents a major health issue. Although the EV cargo may reflect the molecular composition of the releasing cells and thus, EVs may hold a great promise for tumor diagnostics, the impact of intratumoral heterogeneity on the intensity of EV release is still largely unknown. By using CRC patient-derived organoids that maintain the cellular and molecular heterogeneity of the original epithelial tumor tissue, we proved that CD44^high cells produce more organoids with a higher proliferation intensity, as compared to CD44^low cells. Interestingly, we detected an increased EV release by CD44^high CRC cells. In addition, we found that the miRNA cargos of CD44^high and CD44^low cell derived EVs largely overlapped and only four miRNAs were specific for one of the above subpopulations. We observed that EVs released by CD44^high cells induced the proliferation and activation of colon fibroblasts more strongly than CD44^low cells. However, this effect was due to the higher EV number rather than to the miRNA cargo of EVs. Collectively, we identified CRC subpopulations with different EV releasing capabilities and we proved that CRC cell-released EVs have a miRNA-independent effect on fibroblast proliferation and activation.     

Distinct Immune Signatures Indicative of Treatment Response and Immune-Related Adverse Events in Melanoma Patients under Immune Checkpoint Inhibitor Therapy

To identify potential early biomarkers of treatment response and immune-related adverse events (irAE), a pilot immune monitoring study was performed in stage IV melanoma patients by flow cytometric analysis of peripheral blood mononuclear cells (PBMC). Overall, 17 patients were treated with either nivolumab or pembrolizumab alone, or with a combination of nivolumab and ipilimumab every three weeks. Of 15 patients for which complete response assessment was available, treatment responders (n = 10) as compared to non-responders (n = 5) were characterized by enhanced PD-1 expression on CD8⁺ T cells immediately before treatment (median ± median absolute deviation/MAD 26.7 ± 10.4% vs. 17.2 ± 5.3%). Responders showed a higher T cell responsiveness after T cell receptor ex vivo stimulation as determined by measurement of programmed cell death 1 (PD-1) expression on CD3⁺ T cells before the second cycle of treatment. The percentage of CD8⁺ effector memory (CD8⁺CD45RA⁻CD45RO⁺CCR7⁻) T cells was higher in responders compared to non-responders before and immediately after the first cycle of treatment (median ± MAD 39.2 ± 7.3% vs. 30.5 ± 4.1% and 37.7 ± 4.6 vs. 24.0 ± 6.4). Immune-related adverse events (irAE) were accompanied by a higher percentage of activated CD4⁺ (CD4⁺CD38⁺HLADR⁺) T cells before the second treatment cycle (median ± MAD 14.9 ± 3.9% vs. 5.3 ± 0.4%). In summary, PBMC immune monitoring of immune-checkpoint inhibition (ICI) treatment in melanoma appears to be a promising approach to identify early markers of treatment response and irAEs.     

Reduced Dendritic Cells Expressing CD200R1 in Children with Inflammatory Bowel Disease: Correlation with Th17 and Regulatory T Cells

Loss of tolerance of the adaptive immune system towards indigenous flora contributes to the development of inflammatory bowel diseases (IBD). Defects in dendritic cell (DC)-mediated innate and adoptive immune responses are conceivable. The aim of this study was to investigate the expression of the inhibitory molecules CD200R1 and their ligand CD200 on DCs, to clarify the role of the DCs in the pathogenesis of IBD. Thirty-seven pediatric IBD patients (23 with Crohn’s disease (CD) and 14 with ulcerative colitis (UC)) with mean age 13.25 ± 2.9 years were included. Fourteen age-matched healthy pediatric volunteers (five males and nine females) served as a control group (HC). The percentage of CD11c⁺ myeloid dendritic cells (mDCs) and CD123⁺ plasmacytoid DCs (pDCs) expressing CD200R1 and CD200 were evaluated in peripheral blood using flow cytometry and were correlated with routine biochemical, serological markers, serum levels of cytokines and with the percentages of circulating regulatory T cells (Treg) and CD4⁺ producing IL-17 (Th17). IBD patients showed a significant decrease in the percentage of pDCs and mDCs expressing CD200R1 compared to that of HC. Patients with UC showed increased expressions of the CD200 molecule on pDCs as compared to HC. DCs expressing CD200R1 were found to be correlated positively with Treg and negatively with TH17 and erythrocyte sedimentation rate (ESR). Our findings suggest that IBD is associated with dysregulation in the CD200R1/CD200 axis and that the decrease in DCs expressing CD200R1 may contribute to the imbalance of Th17 and Treg cells and in the pathogenesis of IBD.     

Changes in the Ratio of Tc1/Tc2 and Th1/Th2 Cells but Not in Subtypes of NK-Cells in Preeclampsia

It has been suggested that natural killer (NK) cell activity and Th1 immunity may be involved in the pathogenesis of preeclampsia. This study aimed to investigate the immunophenotypes of NK cells and type 1/type 2 immunity in both decidua and maternal peripheral blood between normal (n=11) and preeclamptic pregnant women (n=20) by flow cytometry. The results showed that no significant difference was observed between patients and controls by detecting CD56⁺CD69⁺ and CD56⁺CD94⁺ NK cells in both peripheral blood and decidua. Moreover, in preeclamptic patients, decreased percentages of Tc2 and Th2 cells and the increased ratios of Tc1/Tc2 were determined in both decidua and maternal peripheral blood. In addition, the ratio of Th1/Th2 in peripheral blood also increased. There was no significant difference of immunophenotypes of uNK cells between preeclampsia and normal pregnancy. Local decidua and systematic immunity did not correlate with each other. These results suggest that the type 1/type 2 immunity shifted to type 1 immunity including Th1 and Tc1 cells may contribute to the patho-genesis of preeclampsia.     

CD38 Correlates with an Immunosuppressive Treg Phenotype in Lupus-Prone Mice

CD38 is a transmembrane glycoprotein expressed by T-cells. It has been reported that patients with systemic lupus erythematosus (SLE) showed increased CD38⁺CD25⁺ T-cells correlating with immune activation and clinical signs. Contrariwise, CD38 deficiency in murine models has shown enhanced autoimmunity development. Recent studies have suggested that CD38⁺ regulatory T-cells are more suppressive than CD38⁻ regulatory T-cells. Thus, we have suggested that CD38 overexpression in SLE patients could play a role in regulating immune activation cells instead of enhancing it. This study found a correlation between CD38 with FoxP3 expression and immunosuppressive molecules (CD69, IL-10, CTLA-4, and PD-1) in T-cells from lupus-prone mice (B6.MRL-Fas^lpr/J). Additionally, B6.MRL-Fas^lpr/J mice showed a decreased proportion of CD38⁺ Treg cells regarding wild-type mice (WT). Furthermore, Regulatory T-Cells (Treg cells) from CD38-/- mice showed impairment in expressing immunosuppressive molecules and proliferation after stimulation through the T-cell receptor (TCR). Finally, we demonstrated an increased ratio of IFN-γ/IL-10 secretion in CD38-/- splenocytes stimulated with anti-CD3 compared with the WT. Altogether, our data suggest that CD38 represents an element in maintaining activated and proliferative Treg cells. Consequently, CD38 could have a crucial role in immune tolerance, preventing SLE development through Treg cells.     

Deregulated Expression of Immune Checkpoints on Circulating CD4 T Cells May Complicate Clinical Outcome and Response to Treatment with Checkpoint Inhibitors in Multiple Myeloma Patients

Unlike solid-tumor patients, a disappointingly small subset of multiple myeloma (MM) patients treated with checkpoint inhibitors derive clinical benefits, suggesting differential participation of inhibitory receptors involved in the development of T-cell-mediated immunosuppression. In fact, T cells in MM patients have recently been shown to display features of immunosenescence and exhaustion involved in immune response inhibition. Therefore, we aimed to identify the dominant inhibitory pathway in MM patients to achieve its effective control by therapeutic interventions. By flow cytometry, we examined peripheral blood (PB) CD4 T cell characteristics assigned to senescence or exhaustion, considering PD-1, CTLA-4, and BTLA checkpoint expression, as well as secretory effector function, i.e., capacity for IFN-γ and IL-17 secretion. Analyses were performed in a total of 40 active myeloma patients (newly diagnosed and treated) and 20 healthy controls. At the single-cell level, we found a loss of studied checkpoints’ expression on MM CD4 T cells (both effector (Teff) and regulatory (Treg) cells) primarily at diagnosis; the checkpoint deficit in MM relapse was not significant. Nonetheless, PD-1 was the only checkpoint distributed on an increased proportion of T cells in all MM patients irrespective of disease phase, and its expression on CD4 Teff cells correlated with adverse clinical courses. Among patients, the relative defect in secretory effector function of CD4 T cells was more pronounced at myeloma relapse (as seen in declined Th1/Treg and Th17/Treg cell rates). Although the contribution of PD-1 to MM clinical outcomes is suggestive, our study clearly indicated that the inappropriate expression of immune checkpoints (associated with dysfunctionality of CD4 T cells and disease clinical phase) might be responsible for the sub-optimal clinical response to therapeutic checkpoint inhibitors in MM.     

Regulatory Cell Populations in Relapsing-Remitting Multiple Sclerosis (RRMS) Patients: Effect of Disease Activity and Treatment Regimens

Multiple sclerosis (MS) is a demyelinating disease of the central nervous system (CNS) of autoimmune etiology that results from an imbalance between CNS-specific T effector cells and peripheral suppressive mechanisms mediated by regulatory cells (RC). In this research, we collected blood samples from 83 relapsing remitting MS (RRMS) patients and 45 healthy persons (HC), to assess the sizes of their RC populations, including CD4⁺CD25^highFoxp3⁺ (nTregs), CD3⁺CD4⁺HLA⁻G⁺, CD3⁺CD8⁺CD28⁻, CD3⁺CD56⁺, and CD56^bright cells, and how RC are affected by disease activity (acute phase or remission) and types of treatment (methylprednisolone, interferon, or natalizumab). In addition, we isolated peripheral blood mononuclear cells (PBMC) and cultured them with peptides mapping to myelin antigens, to determine RC responsiveness to autoantigens. The results showed decreased levels of nTregs in patients in the acute phase ± methylprednisolone and in remission + natalizumab, but HC levels in patients in remission or receiving interferon. Patients + interferon had the highest levels of CD3⁺CD4⁺HLA⁻G⁺ and CD3⁺CD8⁺CD28⁻ RC, and patients in the acute phase + methylprednisolone the lowest. Patients in remission had the highest levels of CD3⁺CD56⁺, and patients in remission + natalizumab the highest levels of CD56^bright cells. Only nTregs responded to autoantigens in culture, regardless of disease activity or treatment. The highest suppressive activity was exhibited by nTregs from patients in remission. In conclusion, in RRMS disease activity and type of treatment affect different RC populations. nTregs respond to myelin antigens, indicating that it is possible to restore immunological tolerance through nTreg induction.     

The Many Faces of CD4+ T Cells: Immunological and Structural Characteristics

As a major arm of the cellular immune response, CD4⁺ T cells are important in the control and clearance of infections. Primarily described as helpers, CD4⁺ T cells play an integral role in the development and activation of B cells and CD8⁺ T cells. CD4⁺ T cells are incredibly heterogeneous, and can be divided into six main lineages based on distinct profiles, namely T helper 1, 2, 17 and 22 (Th1, Th2, Th17, Th22), regulatory T cells (Treg) and T follicular helper cells (Tfh). Recent advances in structural biology have allowed for a detailed characterisation of the molecular mechanisms that drive CD4⁺ T cell recognition. In this review, we discuss the defining features of the main human CD4⁺ T cell lineages and their role in immunity, as well as their structural characteristics underlying their detection of pathogens.     

Identification of Surface Antigens That Define Human Pluripotent Stem Cell-Derived PRRX1+Limb-Bud-like Mesenchymal Cells

Stem cell-based therapies and experimental methods rely on efficient induction of human pluripotent stem cells (hPSCs). During limb development, the lateral plate mesoderm (LPM) produces limb-bud mesenchymal (LBM) cells that differentiate into osteochondroprogenitor cells and form cartilage tissues in the appendicular skeleton. Previously, we generated PRRX1-tdTomato reporter hPSCs to establish the protocol for inducing the hPSC-derived PRRX1⁺ LBM-like cells. However, surface antigens that assess the induction efficiency of hPSC-derived PRRX1⁺ LBM-like cells from LPM have not been identified. Here, we used PRRX1-tdTomato reporter hPSCs and found that high pluripotent cell density suppressed the expression of PRRX1 mRNA and tdTomato after LBM-like induction. RNA sequencing and flow cytometry suggested that PRRX1-tdTomato⁺ LBM-like cells are defined as CD44^high CD140B^high CD49f⁻. Importantly, other hPSC lines, including four human induced pluripotent stem cell lines (414C2, 1383D2, HPS1042, HPS1043) and two human embryonic stem cell lines (SEES4, SEES7), showed the same results. Thus, an appropriate cell density of hPSCs before differentiation is a prerequisite for inducing the CD44^high CD140B^high CD49f⁻ PRRX1⁺ LBM-like cells.     

CD4+ T-Cell Plasticity in Non-Infectious Retinal Inflammatory Disease

Non-infectious uveitis (NIU) is a potentially sight-threatening disease. Effector CD4⁺ T cells, especially interferon-γ-(IFNγ) producing Th1 cells and interleukin-17-(IL-17) producing Th17 cells, are the major immunopathogenic cells, as demonstrated by adoptive transfer of disease in a model of experimental autoimmune uveitis (EAU). CD4⁺FoxP3⁺CD25⁺ regulatory T cells (Tregs) were known to suppress function of effector CD4⁺ T cells and contribute to resolution of disease. It has been recently reported that some CD4⁺ T-cell subsets demonstrate shared phenotypes with another CD4⁺ T-cell subset, offering the potential for dual function. For example, Th17/Th1 (co-expressing IFNγ and IL-17) cells and Th17/Treg (co-expressing IL-17 and FoxP3) cells have been identified in NIU and EAU. In this review, we have investigated the evidence as to whether these ‘plastic CD4⁺ T cells’ are functionally active in uveitis. We conclude that Th17/Th1 cells are generated locally, are resistant to the immunosuppressive effects of steroids, and contribute to early development of EAU. Th17/Treg cells produce IL-17, not IL-10, and act similar to Th17 cells. These cells were considered pathogenic in uveitis. Future studies are needed to better clarify their function, and in the future, these cell subsets may in need to be taken into consideration for designing treatment strategies for disease.     

Simultaneous Genetic Ablation of PD-1, LAG-3, and TIM-3 in CD8 T Cells Delays Tumor Growth and Improves Survival Outcome

Immune checkpoint inhibitors (ICI) represented a step forward in improving the outcome of patients with various refractory solid tumors and several therapeutic regimens incorporating ICI have already been approved for a variety of tumor entities. However, besides remarkable long-term responses, checkpoint inhibition can trigger severe immune-related adverse events in some patients. In order to improve safety of ICI as well as T cell therapy, we tested the feasibility of combining T cell-based immunotherapy with genetic disruption of checkpoint molecule expression. Therefore, we generated H-Y and ovalbumin antigen-specific CD8⁺ T cells with abolished PD-1, LAG-3, and TIM-3 expression through CRISPR/Cas9 technology. CD8⁺ T cells, subjected to PD-1, LAG-3, and TIM-3 genetic editing, showed a strong reduction in immune checkpoint molecule expression after in vitro activation, while no relevant reduction in responsiveness to in vitro stimulation was observed. At the same time, in B16-OVA tumor model, transferred genetically edited OT-1 CD8⁺ T cells promoted longer survival compared to control T cells and showed enhanced expansion without associated toxicity. Our study supports the notion that antigen-specific adoptive T cell therapy with concomitant genetic disruption of multiple checkpoint inhibitory receptors could represent an effective antitumor immunotherapy approach with improved tolerability profile.     

Interferon-Beta Therapy of Multiple Sclerosis Patients Improves the Responsiveness of T Cells for Immune Suppression by Regulatory T Cells

Multiple sclerosis (MS) is an inflammatory autoimmune disease characterized by imbalanced immune regulatory networks, and MS patient-derived T effector cells are inefficiently suppressed through regulatory T cells (Treg), a phenomenon known as Treg resistance. In the current study we investigated T cell function in MS patients before and after interferon-beta therapy. We compared cytokine profile, responsiveness for Treg-mediated suppression ex vivo and evaluated reactivity of T cells in vivo using a humanized mouse model. We found that CD4⁺ and CD8⁺ T cells of therapy-naive MS patients were resistant to Treg-mediated suppression. Treg resistance is associated with an augmented IL-6 production, enhanced IL-6 receptor expression, and increased PKB/c-Akt phosphorylation. These parameters as well as responsiveness of T cells to Treg-mediated suppression were restored after interferon-beta therapy of MS patients. Following transfer into immunodeficient mice, MS T cells induced a lethal graft versus host disease (GvHD) and in contrast to T cells of healthy volunteers, this aggressive T cell response could not be controlled by Treg, but was abolished by anti-IL-6 receptor antibodies. However, magnitude and lethality of GvHD induced by MS T cells was significantly decreased after interferon-beta therapy and the reaction was prevented by Treg activation in vivo. Our data reveals that interferon-beta therapy improves the immunoregulation of autoaggressive T effector cells in MS patients by changing the IL-6 signal transduction pathway, thus restoring their sensitivity to Treg-mediated suppression.     

Temozolomide Induces the Acquisition of Invasive Phenotype by O6-Methylguanine-DNA Methyltransferase (MGMT)+ Glioblastoma Cells in a Snail-1/Cx43-Dependent Manner

Glioblastoma multiforme (GBM) recurrences after temozolomide (TMZ) treatment result from the expansion of drug-resistant and potentially invasive GBM cells. This process is facilitated by O6-Methylguanine-DNA Methyltransferase (MGMT), which counteracts alkylating TMZ activity. We traced the expansion of invasive cell lineages under persistent chemotherapeutic stress in MGMT^low (U87) and MGMT^high (T98G) GBM populations to look into the mechanisms of TMZ-induced microevolution of GBM invasiveness. TMZ treatment induced short-term, pro-invasive phenotypic shifts of U87 cells, in the absence of Snail-1 activation. They were illustrated by a transient induction of their motility and followed by the hypertrophy and the signs of senescence in scarce U87 sub-populations that survived long-term TMZ stress. In turn, MGMT^high T98G cells reacted to the long-term TMZ treatment with the permanent induction of invasiveness. Ectopic Snail-1 down-regulation attenuated this effect, whereas its up-regulation augmented T98G invasiveness. MGMT^low and MGMT^high cells both reacted to the long-term TMZ stress with the induction of Cx43 expression. However, only in MGMT^high T98G populations, Cx43 was directly involved in the induction of invasiveness, as manifested by the induction of T98G invasiveness after ectopic Cx43 up-regulation and by the opposite effect after Cx43 down-regulation. Collectively, Snail-1/Cx43-dependent signaling participates in the long-term TMZ-induced microevolution of the invasive GBM front. High MGMT activity remains a prerequisite for this process, even though MGMT-related GBM chemoresistance is not necessary for its initiation.     

Clinical and Prognostic Value of Antigen-Presenting Cells with PD-L1/PD-L2 Expression in Ovarian Cancer Patients

The latest literature demonstrates the predominant role of the programmed cell death axis (PD-1/PD-L1/PD-L2) in ovarian cancer (OC) pathogenesis. However, data concerning this issue is ambiguous. Our research aimed to evaluate the clinical importance of PD-L1/PD-L2 expression in OC environments. We evaluated the role of PD-L1/PD-L2 in OC patients (n = 53). The analysis was performed via flow cytometry on myeloid (mDCs) and plasmacytoid dendritic cells (pDCs) and monocytes/macrophages (MO/MA) in peripheral blood, peritoneal fluid (PF), and tumor tissue (TT). The data were correlated with clinicopathological characteristics and prognosis of OC patients. The concentration of soluble PD-L1 (sPD-L1) and PD-1 in the plasma and PF were determined by ELISA. We established an accumulation of PD-L1⁺/PD-L2⁺ mDCs, pDCs, and MA in the tumor microenvironment. We showed an elevated level of sPD-L1 in the PF of OC patients in comparison to plasma and healthy subjects. sPD-L1 levels in PF showed a positive relationship with Ca125 concentration. Moreover, we established an association between higher sPD-L1 levels in PF and shorter survival of OC patients. An accumulation of PD-L1⁺/PD-L2⁺ mDCs, pDCs, and MA in the TT and high sPD-L1 levels in PF could represent the hallmark of immune regulation in OC patients.