The effect of radiation treatment of solid tumors on neutrophil infiltration and function: a systematic review.

Radiotherapy (RT) initiates a local and systemic immune response which can induce anti-tumor immunity and improve immunotherapy efficacy. Neutrophils are among the first immune cells that infiltrate tumors after RT and are suggested to be essential for the initial anti-tumor immune response. However, neutrophils in tumors are associated with poor outcomes and RT induced neutrophil infiltration could also change the composition of the tumor microenvironment (TME) in favor of tumor progression. To improve RT efficacy for cancer patients it is important to understand the interplay between RT and neutrophils. Here, we review the literature on how RT affects the infiltration and function of neutrophils in the TME of solid tumors, using both patients studies and preclinical murine in vivo models. In general, it was found that neutrophil levels increase and reach maximal levels in the first days after RT and can remain elevated up to three weeks. Most studies report an immunosuppressive role of neutrophils in the TME after RT, caused by upregulated expression of neutrophil indoleamine 2,3-dioxygenase 1 (IDO1) and arginase 1 (ARG1), as well as neutrophil extracellular trap (NET) formation. RT was also associated with increased reactive oxygen species (ROS) production by neutrophils, which can both improve and inhibit anti-tumor immunity. In addition, multiple murine models showed improved RT efficacy when depleting neutrophils, suggesting that neutrophils have a pro-tumor phenotype after RT. We conclude that the role of neutrophils should not be overlooked when developing RT strategies and requires further investigation in specific tumor types. In addition, neutrophils can possibly be exploited to enhance RT efficacy by combining RT with neutrophil-targeting therapies.

Single-cell transcriptomic analysis reveals heterogeneous features of myeloid-derived suppressor cells in newborns.

The transitory emergence of myeloid-derived suppressor cells (MDSCs) in infants is important for the homeostasis of the immune system in early life. The composition and functional heterogeneity of MDSCs in newborns remain elusive, hampering the understanding of the importance of MDSCs in neonates. In this study, we unraveled the maturation trajectory of polymorphonuclear (PMN)-MDSCs from the peripheral blood of human newborns by performing single-cell RNA sequencing. Results indicated that neonatal PMN-MDSCs differentiated from self-renewal progenitors, antimicrobial PMN-MDSCs, and immunosuppressive PMN-MDSCs to late PMN-MDSCs with reduced antimicrobial capacity. We also established a simple framework to distinguish these distinct stages by CD177 and CXCR2. Importantly, preterm newborns displayed a reduced abundance of classical PMN-MDSCs but increased late PMN-MDSCs, consistent with their higher susceptibility to infections and inflammation. Furthermore, newborn PMN-MDSCs were distinct from those from cancer patients, which displayed minimum expression of genes about antimicrobial capacity. This study indicates that the heterogeneity of PMN-MDSCs is associated with the maturity of human newborns.

Molecular and morphologic characterization of intraductal tubulopapillary neoplasms of pancreas with novel potentially targetable fusions.

Intraductal tubulopapillary neoplasms (ITPNs) are rare pancreatic tumors with distinct histological and molecular features. Distinction of ITPN from other pancreatic neoplasms is crucial given the known favorable prognosis and the high frequency and diversity of potentially targetable fusions in ITPN. While the histological features of ITPN are well documented, there are few reports on the cytological features, and molecular characterization of ITPN. The authors reported three cases diagnosed in their laboratory between 2016 and 2021. Clinical data, cytomorphological and histological features, with immunophenotypic and molecular characterizations of these cases are described and compared with those reported in the literature. All 3 cases were diagnosed as ITPN based on the microscopic presence of intraductal nodules composed of tightly packed small tubular glands lined by cuboidal cells lacking apparent mucin. On molecular profiling KRAS and TP53 variants were found in Case 1, FGFR2-INA fusion in Case 2, and STARD3NL-BRAF fusion was detected in Case 3. Immunohistochemistry (IHC) revealed that the neoplastic cells in Case 1 were MUC2 positive and MUC6 negative, but in Cases 2 and 3, were negative for MUC2 and positive for MUC6. These results demonstrate the immunophenotypic and molecular variabilities of histologically similar pancreatic neoplasms. The absence of alterations characteristic of more common pancreatic neoplasms should prompt the consideration of fusion studies in morphologically relevant cases. The combination of morphological, IHC, and molecular analyses is important for reliable identification of ITPN given its potential clinical management implications.

Lateral Motor Column specific expression of Sonic Hedgehog contributes to maintenance and scaling of pMN progenitor cell populations during oligodendrogenesis.

Motor neurons (MNs) and oligodendrocyte precursor cells (OPCs) emerge sequentially from the pMN precursor domain during spinal cord development. MNs diversify into muscle specific subtypes and settle in stereotypic locations in the ventral horns. In contrast, OPCs are mobile and appear to evenly populate the parenchyma. Whether earlier born MNs influence OPC production is controversial. We found that Sonic Hedgehog signaling emanating from nascent MNs of the lateral motor column is critical for maintaining a larger and more yielding pMN domain at limb levels compared to trunk levels during OPC production. Reduced Shh signaling resulted in unrecoverable diminishment of pMN domain based OPC production leaving the spinal cord impoverished of OPC. Our results suggest that production of OPC at limb levels is contingent on completion of MN production.

Self-Powered Pressure-Temperature Bimodal Sensing Based on the Piezo-Pyroelectric Effect for Robotic Perception.

Multifunctional sensors have played a crucial role in constructing high-integration electronic networks. Most of the current multifunctional sensors rely on multiple materials to simultaneously detect different physical stimuli. Here, we demonstrate the large piezo-pyroelectric effect in ferroelectric Pb(Mg1/3Nb2/3)O3-PbTiO3 (PMN-PT) single crystals for simultaneous pressure and temperature sensing. The outstanding piezoelectric and pyroelectric properties of PMN-PT result in rapid response speed and high sensitivity, with values of 46 ms and 28.4 nA kPa-1 for pressure sensing, and 1.98 s and 94.66 nC °C-1 for temperature detection, respectively. By leveraging the distinct differences in the response speed of piezoelectric and pyroelectric responses, the piezo-pyroelectric effect of PMN-PT can effectively detect pressure and temperature from mixed-force thermal stimuli, which enables a robotic hand for stimuli classification. With appealing multifunctionality, fast speed, high sensitivity, and compact structure, the proposed self-powered bimodal sensor therefore holds significant potential for high-performance artificial perception.

Single-cell analysis defines highly specific leukemia-induced neutrophils and links MMP8 expression to recruitment of tumor associated neutrophils during FGFR1 driven leukemogenesis.

BACKGROUND: Leukemias driven by activated, chimeric FGFR1 kinases typically progress to AML which have poor prognosis. Mouse models of this syndrome allow detailed analysis of cellular and molecular changes occurring during leukemogenesis. We have used these models to determine the effects of leukemia development on the immune cell composition in the leukemia microenvironment during leukemia development and progression. METHODS: Single cell RNA sequencing (scRNA-Seq) was used to characterize leukemia associated neutrophils and define gene expression changes in these cells during leukemia progression. RESULTS: scRNA-Seq revealed six distinct subgroups of neutrophils based on their specific differential gene expression. In response to leukemia development, there is a dramatic increase in only two of the neutrophil subgroups. These two subgroups show specific gene expression signatures consistent with neutrophil precursors which give rise to immature polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs). Analysis of gene expression in these precursor cells identified pathways that were specifically upregulated, the most pronounced of which involved matrix metalloproteinases Mmp8 and Mmp9, during leukemia progression. Pharmacological inhibition of MMPs using Ilomastat preferentially restricted in vitro migration of neutrophils from leukemic mice and led to a significantly improved survival in vivo, accompanied by impaired PMN-MDSC recruitment. As a result, levels of T-cells were proportionally increased. In clinically annotated TCGA databases, MMP8 was shown to act as an independent indicator for poor prognosis and correlated with higher neutrophil infiltration and poor pan-cancer prognosis. CONCLUSION: We have defined specific leukemia responsive neutrophil subgroups based on their unique gene expression profile, which appear to be the precursors of neutrophils specifically associated with leukemia progression. An important event during development of these neutrophils is upregulation MMP genes which facilitated mobilization of these precursors from the BM in response to cancer progression, suggesting a possible therapeutic approach to suppress the development of immune tolerance.

Monocytic myeloid-derived suppressor cells as an immune indicator of early diagnosis and prognosis in patients with sepsis.

BACKGROUND: Immunosuppression is a leading cause of septic death. Therefore, it is necessary to search for biomarkers that can evaluate the immune status of patients with sepsis. We assessed the diagnostic and prognostic value of low-density neutrophils (LDNs) and myeloid-derived suppressor cells (MDSCs) subsets in the peripheral blood mononuclear cells (PBMCs) of patients with sepsis. METHODS: LDNs and MDSC subsets were compared among 52 inpatients with sepsis, 33 inpatients with infection, and 32 healthy controls to investigate their potential as immune indicators of sepsis. The percentages of LDNs, monocytic MDSCs (M-MDSCs), and polymorphonuclear MDSCs (PMN-MDSCs) in PBMCs were analyzed. Sequential organ failure assessment (SOFA) scores, C-reactive protein (CRP), and procalcitonin (PCT) levels were measured concurrently. RESULTS: The percentages of LDNs and MDSC subsets were significantly increased in infection and sepsis as compared to control. MDSCs performed similarly to CRP and PCT in diagnosing infection or sepsis. LDNs and MDSC subsets positively correlated with PCT and CRP levels and showed an upward trend with the number of dysfunctional organs and SOFA score. Non-survivors had elevated M-MDSCs compared with that of patients who survived sepsis within 28 days after enrollment. CONCLUSIONS: MDSCs show potential as a diagnostic biomarker comparable to CRP and PCT, in infection and sepsis, even in distinguishing sepsis from infection. M-MDSCs show potential as a prognostic biomarker of sepsis and may be useful to predict 28-day hospital mortality in patients with sepsis.

Melatonin facts: Melatonin lacks immuno-inflammation boosting capacities at the molecular and cellular levels.

Among the properties melatonin is claimed to possess, are the immuno-inflammation inductive capacities that would be responsible of some of the paramount of activities melatonin is reported to have in most of the human pathological conditions. In the present paper, we measured the effect of melatonin on established cellular models of immuno-inflammation, and found none. The discrepancies are discussed, especially because those properties are reported at pharmacological concentration (1 µM and beyond) at which the melatonin receptors are desensitized by internalization, leading to putative non-receptor-dependent mechanism of action.

Engineering Domain Variants in 0.7Pb(Mg1/3Nb2/3)-0.3PbTiO3 Single Crystals Using High-Frequency AC Poling.

Single crystals of (001)-oriented 0.7Pb(Mg1/3Nb2/3)-0.3PbTiO3 (PMN-30PT) with a composition near the morphotropic phase boundary have attracted considerable attention due to their superior dielectric and electromechanical performance. Recently, a new alternating current (electric field) poling approach used for the enhancement of dielectric and piezoelectric properties. However, the microscopic domain variants that govern the performance, especially under high-frequency alternating current (AC) voltages, remain largely unexplored. In this work, the domain microstructure under AC poling reveals the presence of four monoclinic (MA) domain variants using a suite of scanning probe microscopy methods, and X-ray diffraction (XRD) reciprocal space mapping is tuned. It is reported on the emergence of hierarchical fine domains - needle-shaped, and 109° domain walls under applied high-frequency AC poling. Time-resolved Kelvin probe force microscopy (KPFM) reveals the charge dynamics and relaxation behavior of these needle domains and walls. The findings provide new insight and guidance to the domain engineering by high-frequency AC poling for the development of advanced transducer technology.

Exosomes promote pre-metastatic niche formation in colorectal cancer.

It is well known that colorectal cancer (CRC) has a high morbidity rate, a poor prognosis when metastasized, and a greatly shortened 5-year survival rate. Therefore, understanding the mechanism of tumor metastasis is still important. Based on the "seed and soil" theory, the concept of " premetastatic niche (PMN)" was introduced by Kaplan et al. The complex interaction between primary tumors and the metastatic organ provides a beneficial microenvironment for tumor cells to colonize at a distance. With further exploration of the PMN, exosomes have gradually attracted interest from researchers. Exosomes are extracellular vesicles secreted from cells that include various biological information and are involved in communication between cells. As a key molecule in the PMN, exosomes are closely related to tumor metastasis. In this article, we obtained information by conducting a comprehensive search across academic databases including PubMed and Web of Science using relevant keywords. Only recent, peer-reviewed articles published in the English language were considered for inclusion. This study aims to explore in depth how exosomes promote the formation of pre-metastatic microenvironment (PMN) in colorectal cancer and its related mechanisms.

Bladder-cancer-derived exosomal circRNA_0013936 promotes suppressive immunity by up-regulating fatty acid transporter protein 2 and down-regulating receptor-interacting protein kinase 3 in PMN-MDSCs.

BACKGROUND: Polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) is one of the causes of tumor immune tolerance and failure of cancer immunotherapy. Here, we found that bladder cancer (BCa)-derived exosomal circRNA_0013936 could enhance the immunosuppressive activity of PMN-MDSCs by regulating the expression of fatty acid transporter protein 2 (FATP2) and receptor-interacting protein kinase 3 (RIPK3). However, the underlying mechanism remains largely unknown. METHODS: BCa-derived exosomes was isolated and used for a series of experiments. RNA sequencing was used to identify the differentially expressed circRNAs. Western blotting, immunohistochemistry, immunofluorescence, qRT-PCR, ELISA and Flow cytometry were performed to reveal the potential mechanism of circRNA_0013936 promoting the immunosuppressive activity of PMN-MDSC. RESULTS: CircRNA_0013936 enriched in BCa-derived exosomes could promote the expression of FATP2 and inhibit the expression of RIPK3 in PMN-MDSCs. Mechanistically, circRNA_0013936 promoted the expression of FATP2 and inhibited the expression of RIPK3 expression via sponging miR-320a and miR-301b, which directly targeted JAK2 and CREB1 respectively. Ultimately, circRNA_0013936 significantly inhibited the functions of CD8+ T cells by up-regulating FATP2 through the circRNA_0013936/miR-320a/JAK2 pathway, and down-regulating RIPK3 through the circRNA_0013936/miR-301b/CREB1 pathway in PMN-MDSCs. CONCLUSIONS: BCa-derived exosomal circRNA_0013936 promotes suppressive immunity by up-regulating FATP2 through the circRNA_0013936/miR-320a/JAK2 pathway and down-regulating RIPK3 through the circRNA_0013936/miR-301b-3p/CREB1 pathway in PMN-MDSCs. These findings help to find new targets for clinical treatment of human bladder cancer.

Decreased neutrophil function in newly diagnosed multiple myeloma patients is restored with lenalidomide therapy.

OBJECTIVES: Bacterial infections are common and a major cause of morbidity and mortality in multiple myeloma (MM). We have investigated the function of polymorphonuclear leukocyte (PMN), the immune system's first line of defense against bacteria, in peripheral blood (PB) and bone marrow (BM) samples from patients with newly diagnosed MM (NDMM), smoldering MM (SMM), monoclonal gammopathy of undetermined significance (MGUS) and healthy controls. METHODS: Phagocytosis and oxidative burst in PMN cells from patients and healthy donors were investigated using PhagoTest and PhagoBurst assay. RESULTS: PMN from NDMM, SMM, and MGUS patients had reduced phagocytosis and oxidative burst ability compared with healthy controls. The dysfunction was most prominent in BM samples from MM, SMM, and MGUS patients. Importantly the reduced phagocytosis in MM patients was restored in patients on lenalidomide therapy. Consistently the ability of Escherichia coli stimulated oxidative burst in BM was reduced for the MM, SMM, and MGUS cohort in contrast to the healthy controls and the patients on lenalidomide treatment. CONCLUSION: Our results show that MM patients have neutrophil dysfunction that could contribute to susceptibility for bacterial infections and that lenalidomide therapy was associated with restored PMN function.

Synovial Calprotectin is Superior to Synovial Leukocyte Count in Excluding Chronic Periprosthetic Joint Infections, a Retrospective Cohort Study.

BACKGROUND: Synovial calprotectin is a promising biomarker for diagnosing chronic periprosthetic joint infections (PJIs), but its diagnostic value has not been directly compared to synovial leukocyte count and polymorphonuclear neutrophils. This study aimed to: (1) evaluate and compare the diagnostic accuracy between these markers in patients undergoing revision arthroplasty for chronic PJI or aseptic reasons; and (2) determine the best rule-out and rule-in test for PJI. METHODS: Synovial fluid samples from patients undergoing revision arthroplasty in hip and knee joints were collected and analyzed. Patients diagnosed with an acute PJI, patients treated with antibiotics 2 weeks prior to revision surgery, and/or patients who had active inflammatory joint disease were excluded. Periprosthetic joint infections were diagnosed based on the presence of a sinus tract and/or positive intraoperative cultures according to the European Bone and Joint Infection Society microbiological criteria. RESULTS: A total of 137 patients were included, of whom 19 (14%) were diagnosed with a PJI. Overall, synovial calprotectin had the highest diagnostic accuracy of all studied markers (area under the curve 96%). Synovial calprotectin, with a cutoff of 50 mg/L, had the highest negative predictive value of 100%. However, PMNs (> 80%) combined with a leukocyte count (> 3,000 cells/µL) showed the highest positive predictive value of an infection (positive predictive value17). CONCLUSIONS: Synovial calprotectin is the most accurate biomarker for ruling out a chronic PJI, while the combination of synovial leukocyte count and PMN is most reliable for ruling in a chronic PJI.

Can Mn:PIN-PMN-PT piezocrystal replace hard piezoceramic in power ultrasonic devices?

Mn:PIN-PMN-PT piezocrystal is investigated to determine whether its enhanced energy density makes it a candidate transducer material for power ultrasonics applications. To this end, the electromechanical and vibrational characteristics of a simple configuration of a bolted Langevin transducer (BLT) and then an ultrasonic surgical device, both incorporating Mn:PIN-PMN-PT piezocrystal, are compared with the same transducer configurations incorporating a conventional hard PZT piezoceramic commonly used in high-power ultrasonic transducers. The material properties of Mn:PIN-PMN-PT are determined using a single sample characterisation technique and these are used in finite element analysis (FEA) to design and then fabricate the BLT and ultrasonic surgical device, tuned to the first and second longitudinal modes at 20 kHz respectively. FEA is similarly used for the hard PZT versions. It is found that the superior elastic compliance of Mn:PIN-PMN-PT results in a higher radial piezo-stack deformation than the hard PZT under ultrasonic excitation of the BLT. However, the resulting longitudinal displacement amplitude of the two BLTs and two ultrasonic surgical devices is found to be equal, despite the higher figure of merit (Qkeff2) of those incorporating Mn:PIN-PMN-PT. The electrical impedance is measured at increasing excitation levels to evaluate the quality factor, Q. It is found that damping in the BLT with hard PZT is negligibly affected in the excitation range considered; however, the BLT incorporating Mn:PIN-PMN-PT exhibits a large reduction in Q. These findings indicate that, for measurements in air, the advantages of the high figure of merit of the piezocrystal material are not realised in a high-power transducer due to significantly increased damping at high excitation levels. To compare the vibrational response of the two ultrasonic surgical devices, L-C electrical impedance matching was implemented to maximise the efficiency of energy transfer from the source to the transducer under load. Results suggest that similar responses occurred for the two surgical devices in cutting tests using a low strength bone mimic material. However, the Mn:PIN-PMN-PT device exhibited better performance in cutting through higher strength ex-vivo chicken femur.

The role of equine seminal plasma derived cysteine rich secretory protein 3 (CRISP3) in the interaction between polymorphonuclear neutrophils (PMNs) and populations of viable or dead spermatozoa, and bacteria.

Breeding-induced endometritis is a physiological reaction to clear the uterus from excess spermatozoa and bacteria after breeding. Cysteine rich secretory protein 3 in seminal plasma (spCRISP3) protects spermatozoa from binding and destruction by uterine PMNs, but it is not clear if this involves all sperm and bacteria, or if it is selective to a sub-population of live sperm. The objective of this report was to determine if spCRISP3 (1) is selective in its suppression of PMN-binding to sperm based on viability of spermatozoa, (2) protects bacteria from binding to PMNs, and (3) to determine the localization pattern of spCRISP3 on viable and dead sperm. Semen was collected from five stallions and each ejaculate was divided into (1) live and (2) snap frozen (dead) sperm. Two distinct sperm populations were confirmed by DNA fragmentation and membrane integrity assays. CRISP3 was purified from pooled seminal plasma, and binding of PMNs (isolated from peripheral blood) to the two sperm populations and E. coli was evaluated with flow cytometry in the presence of spCRISP3. In addition, localization of spCRISP3 on live and dead spermatozoa was determined by immunocytochemistry. Comparisons between treatments were analyzed using a one-way-ANOVA and Bonferroni's comparison test, or Kruskal-Wallis ANOVA if not normally distributed. spCRISP3 significantly suppressed binding of PMNs to live spermatozoa (p < 0.0001) but had no effect on dead sperm or bacteria (p > 0.05). Immunocytochemistry confirmed binding of spCRISP3 to live, but not dead spermatozoa. It was concluded that a selective interaction between spCRISP3 and live spermatozoa may be part of a biological mechanism that allows safe transport of viable spermatozoa to the oviducts, while enabling dead spermatozoa and bacteria to be eliminated in a timely fashion after breeding.

Stabilizing Perovskite Pb(Mg0.33Nb0.67)O3-PbTiO3 Thin Films by Fast Deposition and Tensile Mismatched Growth Template.

Because of its low hysteresis, high dielectric constant, and strong piezoelectric response, Pb(Mg1/3Nb2/3)O3-PbTiO3 (PMN-PT) thin films have attracted considerable attention for the application in PiezoMEMS, field-effect transistors, and energy harvesting and storage devices. However, it remains a great challenge to fabricate phase-pure, pyrochlore-free PMN-PT thin films. In this study, we demonstrate that a high deposition rate, combined with a tensile mismatched template layer can stabilize the perovskite phase of PMN-PT films and prevent the nucleation of passive pyrochlore phases. We observed that an accelerated deposition rate promoted mixing of the B-site cation and facilitated relaxation of the compressively strained PMN-PT on the SrTiO3 (STO) substrate in the initial growth layer, which apparently suppressed the initial formation of pyrochlore phases. By employing La-doped-BaSnO3 (LBSO) as the tensile mismatched buffer layer, 750 nm thick phase-pure perovskite PMN-PT films were synthesized. The resulting PMN-PT films exhibited excellent crystalline quality close to that of the STO substrate.

An Update of Extracellular Vesicle Involvement in different Steps of Cancer Metastasis and Targeting Strategies.

Cancer metastasis is the deadliest event in tumorigenesis. Despite extensive research, there are still unsolved challenges regarding early metastasis detection and targeting strategies. Extracellular vesicles (EVs) and their impact on tumorigenic-related events are in the eye of current investigations. EVs represent a plethora of biomarkers and information, and they are considered key determinants in tumor progression and for tumor prognosis and monitoring. EVs are one of the key mediators for inter-cellular communications between tumor cells and their nearby stroma. They are involved in different steps of metastasis from invasion toward formation of pre-metastatic niches (PMNs), and final growth and colonization of tumor cells in desired organ/s of the target. Membrane components of EVs and their cargo can be traced for the identification of tumor metastasis, and their targeting is a promising strategy in cancer therapy. In this review, we aimed to discuss the current understanding of EV-based metastatic predilection in cancer, providing updated information about EV involvement in different metastatic steps and suggesting some strategies to hamper this devastating condition.

Surface CD52, CD84, and PTGER2 mark mature PMN-MDSCs from cancer patients and G-CSF-treated donors.

Precise molecular characterization of circulating polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) is hampered by their mixed composition of mature and immature cells and lack of specific markers. Here, we focus on mature CD66b+CD10+CD16+CD11b+ PMN-MDSCs (mPMN-MDSCs) from either cancer patients or healthy donors receiving G-CSF for stem cell mobilization (GDs). By RNA sequencing (RNA-seq) experiments, we report the identification of a distinct gene signature shared by the different mPMN-MDSC populations under investigation, also validated in mPMN-MDSCs from GDs and tumor-associated neutrophils (TANs) by single-cell RNA-seq (scRNA-seq) experiments. Analysis of such a gene signature uncovers a specific transcriptional program associated with mPMN-MDSC differentiation and allows us to identify that, in patients with either solid or hematologic tumors and in GDs, CD52, CD84, and prostaglandin E receptor 2 (PTGER2) represent potential mPMN-MDSC-associated markers. Altogether, our findings indicate that mature PMN-MDSCs distinctively undergo specific reprogramming during differentiation and lay the groundwork for selective immunomonitoring, and eventually targeting, of mature PMN-MDSCs.

Molecular and functional changes in neutrophilic granulocytes induced by nicotine: a systematic review and critical evaluation.

Background: Over 1.1 billion people smoke worldwide. The alkaloid nicotine is a prominent and addictive component of tobacco. In addition to tumors and cardiovascular disorders, tobacco consumption is associated with a variety of chronic-inflammatory diseases. Although neutrophilic granulocytes (neutrophils) play a role in the pathogenesis of many of these diseases, the impact of nicotine on neutrophils has not been systematically reviewed so far. Objectives: The aim of this systematic review was to evaluate the direct influence of nicotine on human neutrophil functions, specifically on cell death/damage, apoptosis, chemotaxis, general motility, adhesion molecule expression, eicosanoid synthesis, cytokine/chemokine expression, formation of neutrophil extracellular traps (NETs), phagocytosis, generation of reactive oxygen species (ROS), net antimicrobial activity, and enzyme release. Material and methods: This review was conducted according to the PRISMA guidelines. A literature search was performed in the databases NCBI Pubmed® and Web of Science™ in February 2023. Inclusion criteria comprised English written research articles, showing in vitro studies on the direct impact of nicotine on specified human neutrophil functions. Results: Of the 532 originally identified articles, data from 34 articles were finally compiled after several evaluation steps. The considered studies highly varied in methodological aspects. While at high concentrations (>3 mmol/l) nicotine started to be cytotoxic to neutrophils, concentrations typically achieved in blood of smokers (in the nmol/l range) applied for long exposure times (24-72h) supported the survival of neutrophils. Smoking-relevant nicotine concentrations also increased the chemotaxis of neutrophils towards several chemoattractants, elevated their production of elastase, lipocalin-2, CXCL8, leukotriene B4 and prostaglandin E2, and reduced their integrin expression. Moreover, while nicotine impaired the neutrophil phagocytotic and anti-microbial activity, a range of studies demonstrated increased NET formation. However, conflicting effects were found on ROS generation, selectin expression and release of ß-glucuronidase and myeloperoxidase. Conclusion: Nicotine seems to support the presence in the tissue and the inflammatory and selected tissue-damaging activity of neutrophils and reduces their antimicrobial functions, suggesting a direct contribution of nicotine to the pathogenesis of chronic-inflammatory diseases via influencing the neutrophil biology.

Isolation methods determine human neutrophil responses after stimulation.

Studying neutrophils is challenging due to their limited lifespan, inability to proliferate, and resistance to genetic manipulation. Neutrophils can sense various cues, making them susceptible to activation by blood collection techniques, storage conditions, RBC lysis, and the isolation procedure itself. Here we assessed the impact of the five most used methods for neutrophil isolation on neutrophil yield, purity, activation status and responsiveness. We monitored surface markers, reactive oxygen species production, and DNA release as a surrogate for neutrophil extracellular trap (NET) formation. Our results show that neutrophils isolated by negative immunomagnetic selection and density gradient methods, without RBC lysis, resembled untouched neutrophils in whole blood. They were also less activated and more responsive to milder stimuli in functional assays compared to neutrophils obtained using density gradients requiring RBC lysis. Our study highlights the importance of selecting the appropriate method for studying neutrophils, and underscores the need for standardizing isolation protocols to facilitate neutrophil subset characterization and inter-study comparisons.