Determination of Serum Arginase-1 Concentrations and Serum Arginase Activity for the Non-Invasive Diagnosis of Endometriosis.

Backgroud: Endometriosis remains a diagnostic challenge, both clinically and economically, affecting 6% to 15% of women of child-bearing potential. We have attempted to determine whether testing serum concentrations and activity of arginase isoenzymes could be useful for the non-invasive diagnosis of endometriosis. Methods: This study involved 180 women (105 endometriosis subjects-study group B; 22 subjects with other benign gynaecological conditions-control group 1-K1, both undergoing surgery; and 53 healthy subjects without features of endometriosis-control group 2-K2). Results: Preoperative and postoperative arginase-1 (Arg-1) concentrations were significantly higher in patients, as compared with the control groups K1 (p < 0.0001 and p = 0.0005, respectively) and K2 (both p < 0.0001). Similarly, arginase activity was significantly higher in patients than in the control group K1 before surgery and higher than in both control groups after surgery. No significant differences in either Arg-1 concentrations or arginase activity were noted between the operated control group K1 and the non-operated control group K2. A significant postoperative decrease in Arg-1 concentration was observed within both patient (p < 0.0001) and control group K1 (p = 0.0043). Diagnostic performance was assessed using the receiver operating characteristic (ROC) method. The threshold for differentiation between endometriosis patients and healthy non-operated controls was 42.3 ng/mL, with a sensitivity of 90% and specificity of 81%. For differentiation of patients and operated controls with benign gynaecological conditions, the threshold was 78.4 ng/mL, with a sensitivity of 61% and specificity of 95%. Conclusions: We, therefore, conclude that Arg-1 serum concentrations and arginase activity could be considered potential biomarkers for endometriosis but require further studies on larger cohorts of patients.

The Relationship between Fine Particle Matter (PM2.5) Exposure and Upper Respiratory Tract Diseases.

PM2.5 is one of the most harmful components of airborne pollution and includes particles with diameters of less than 2.5 µm. Almost 90% of the world's population lives in areas with poor air quality exceeding the norms established by the WHO. PM2.5 exposure affects various organs and systems of the human body including the upper respiratory tract which is one of the most prone to its adverse effects. PM2.5 can disrupt nasal epithelial cell metabolism, decrease the integrity of the epithelial barrier, affect mucociliary clearance, and alter the inflammatory process in the nasal mucosa. Those effects may increase the chance of developing upper respiratory tract diseases in areas with high PM2.5 pollution. PM2.5's contribution to allergic rhinitis (AR) and rhinosinusitis was recently thoroughly investigated. Numerous studies demonstrated various mechanisms that occur when subjects with AR or rhinosinusitis are exposed to PM2.5. Various immunological changes and alterations in the nasal and sinonasal epithelia were reported. These changes may contribute to the observations that exposure to higher PM2.5 concentrations may increase AR and rhinosinusitis symptoms in patients and the number of clinical visits. Thus, studying novel strategies against PM2.5 has recently become the focus of researchers' attention. In this review, we summarize the current knowledge on the effects of PM2.5 on healthy upper respiratory tract mucosa and PM2.5's contribution to AR and rhinosinusitis. Finally, we summarize the current advances in developing strategies against PM2.5 particles' effects on the upper respiratory tract.

International Society for Extracellular Vesicles Workshop. QuantitatEVs: multiscale analyses, from bulk to single extracellular vesicle.

The 'QuantitatEVs: multiscale analyses, from bulk to single vesicle' workshop aimed to discuss quantitative strategies and harmonized wet and computational approaches toward the comprehensive analysis of extracellular vesicles (EVs) from bulk to single vesicle analyses with a special focus on emerging technologies. The workshop covered the key issues in the quantitative analysis of different EV-associated molecular components and EV biophysical features, which are considered the core of EV-associated biomarker discovery and validation for their clinical translation. The in-person-only workshop was held in Trento, Italy, from January 31st to February 2nd, 2023, and continued in Milan on February 3rd with "Next Generation EVs", a satellite event dedicated to early career researchers (ECR). This report summarizes the main topics and outcomes of the workshop.

Immunosuppressive Extracellular Vesicles as a Linking Factor in the Development of Tumor and Endometriotic Lesions in the Gynecologic Tract.

Both gynecological tumors and endometriosis require for their development a favorable environment, termed in the case of tumors a "pre-metastatic niche" and in case of endometriosis a "pro-endometriotic niche". This is characterized by chronic inflammation and immunosuppression that support the further progression of initial lesions. This microenvironment is established and shaped in the course of a vivid cross-talk between the tumor or endometrial cells with other stromal, endothelial and immune cells. There is emerging evidence that extracellular vesicles (EVs) play a key role in this cellular communication, mediating both in tumors and endometriosis similar immunosuppressive and pro-inflammatory mechanisms. In this review, we discuss the latest findings about EVs as immunosuppressive factors, highlighting the parallels between gynecological tumors and endometriosis. Furthermore, we outline their role as potential diagnostic or prognostic biomarkers as well as their future in therapeutic applications.

PD-L1 CAR effector cells induce self-amplifying cytotoxic effects against target cells.

BACKGROUND: Immune checkpoint inhibitors and chimeric antigen receptor (CAR)-based therapies have transformed cancer treatment. Recently, combining these approaches into a strategy of PD-L1-targeted CAR has been proposed to target PD-L1high tumors. Our study provides new information on the efficacy of such an approach against PD-L1low targets. METHODS: New atezolizumab-based PD-L1-targeted CAR was generated and introduced into T, NK, or NK-92 cells. Breast cancer MDA-MB-231 and MCF-7 cell lines or non-malignant cells (HEK293T, HMEC, MCF-10A, or BM-MSC) were used as targets to assess the reactivity or cytotoxic activity of the PD-L1-CAR-bearing immune effector cells. Stimulation with IFNγ or with supernatants from activated CAR T cells were used to induce upregulation of PD-L1 molecule expression on the target cells. HER2-CAR T cells were used for combination with PD-L1-CAR T cells against MCF-7 cells. RESULTS: PD-L1-CAR effector cells responded vigorously with degranulation and cytokine production to PD-L1high MDA-MB-231 cells, but not to PD-L1low MCF-7 cells. However, in long-term killing assays, both MDA-MB-231 and MCF-7 cells were eliminated by the PD-L1-CAR cells, although with a delay in the case of PD-L1low MCF-7 cells. Notably, the coculture of MCF-7 cells with activated PD-L1-CAR cells led to bystander induction of PD-L1 expression on MCF-7 cells and to the unique self-amplifying effect of the PD-L1-CAR cells. Accordingly, PD-L1-CAR T cells were active not only against MDA-MD-231 and MCF-7-PD-L1 but also against MCF-7-pLVX cells in tumor xenograft models. Importantly, we have also observed potent cytotoxic effects of PD-L1-CAR cells against non-malignant MCF-10A, HMEC, and BM-MSC cells, but not against HEK293T cells that initially did not express PD-L1 and were unresponsive to the stimulation . Finally, we have observed that HER-2-CAR T cells stimulate PD-L1 expression on MCF-7 cells and therefore accelerate the functionality of PD-L1-CAR T cells when used in combination. CONCLUSIONS: In summary, our studies show that CAR-effector cells trigger the expression of PD-L1 on target cells, which in case of PD-L1-CAR results in the unique self-amplification phenomenon. This self-amplifying effect could be responsible for the enhanced cytotoxicity of PD-L1-CAR T cells against both malignant and non-malignant cells and implies extensive caution in introducing PD-L1-CAR strategy into clinical studies.

PRDX-1 Supports the Survival and Antitumor Activity of Primary and CAR-Modified NK Cells under Oxidative Stress.

Oxidative stress, caused by the imbalance between reactive species generation and the dysfunctional capacity of antioxidant defenses, is one of the characteristic features of cancer. Here, we quantified hydrogen peroxide in the tumor microenvironment (TME) and demonstrated that hydrogen peroxide concentrations are elevated in tumor interstitial fluid isolated from murine breast cancers in vivo, when compared with blood or normal subcutaneous fluid. Therefore, we investigated the effects of increased hydrogen peroxide concentration on immune cell functions. NK cells were more susceptible to hydrogen peroxide than T cells or B cells, and by comparing T, B, and NK cells' sensitivities to redox stress and their antioxidant capacities, we identified peroxiredoxin-1 (PRDX1) as a lacking element of NK cells' antioxidative defense. We observed that priming with IL15 protected NK cells' functions in the presence of high hydrogen peroxide and simultaneously upregulated PRDX1 expression. However, the effect of IL15 on PRDX1 expression was transient and strictly dependent on the presence of the cytokine. Therefore, we genetically modified NK cells to stably overexpress PRDX1, which led to increased survival and NK cell activity in redox stress conditions. Finally, we generated PD-L1-CAR NK cells overexpressing PRDX1 that displayed potent antitumor activity against breast cancer cells under oxidative stress. These results demonstrate that hydrogen peroxide, at concentrations detected in the TME, suppresses NK cell function and that genetic modification strategies can improve CAR NK cells' resistance and potency against solid tumors.

The Tumor Microenvironment-A Metabolic Obstacle to NK Cells' Activity.

NK cells have unique capabilities of recognition and destruction of tumor cells, without the requirement for prior immunization of the host. Maintaining tolerance to healthy cells makes them an attractive therapeutic tool for almost all types of cancer. Unfortunately, metabolic changes associated with malignant transformation and tumor progression lead to immunosuppression within the tumor microenvironment, which in turn limits the efficacy of various immunotherapies. In this review, we provide a brief description of the metabolic changes characteristic for the tumor microenvironment. Both tumor and tumor-associated cells produce and secrete factors that directly or indirectly prevent NK cell cytotoxicity. Here, we depict the molecular mechanisms responsible for the inhibition of immune effector cells by metabolic factors. Finally, we summarize the strategies to enhance NK cell function for the treatment of tumors.

Small extracellular vesicles containing arginase-1 suppress T-cell responses and promote tumor growth in ovarian carcinoma.

Tumor-driven immune suppression is a major barrier to successful immunotherapy in ovarian carcinomas (OvCa). Among various mechanisms responsible for immune suppression, arginase-1 (ARG1)-carrying small extracellular vesicles (EVs) emerge as important contributors to tumor growth and tumor escape from the host immune system. Here, we report that small EVs found in the ascites and plasma of OvCa patients contain ARG1. EVs suppress proliferation of CD4+ and CD8+ T-cells in vitro and in vivo in OvCa mouse models. In mice, ARG1-containing EVs are transported to draining lymph nodes, taken up by dendritic cells and inhibit antigen-specific T-cell proliferation. Increased expression of ARG1 in mouse OvCa cells is associated with accelerated tumor progression that can be blocked by an arginase inhibitor. Altogether, our studies show that tumor cells use EVs as vehicles to carry over long distances and deliver to immune cells a metabolic checkpoint molecule - ARG1, mitigating anti-tumor immune responses.