Granulocytic Cells Research Articles

New insights on extramedullary granulopoiesis and neutrophil heterogeneity in the spleen and its importance in disease.

Neutrophils are traditionally viewed as uncomplicated exterminators that arrive quickly at sites of infection, kill pathogens, and then expire. However, recent studies employing modern transcriptomics coupled with novel imaging modalities have discovered that neutrophils exhibit significant heterogeneity within organs and have complex functional roles ranging from tissue homeostasis to cancer and chronic pathologies. This has revised the view that neutrophils are simplistic butchers, and there has been a resurgent interest in neutrophils. The spleen was described as a granulopoietic organ more than 4 decades ago, and studies indicate that neutrophils are briefly retained in the spleen before returning to circulation after proliferation. Transcriptomic studies have discovered that splenic neutrophils are heterogeneous and distinct compared with those in blood. This suggests that a unique hematopoietic niche exists in the splenic microenvironment, i.e., capable of programming neutrophils in the spleen. During severe systemic inflammation with an increased need of neutrophils, the spleen can adapt by producing neutrophils through emergency granulopoiesis. In this review, we describe the structure and microanatomy of the spleen and examine how cells within the splenic microenvironment help to regulate splenic granulopoiesis. A focus is placed on exploring the increase in splenic granulopoiesis to meet host needs during infection and inflammation. Emerging technologies such as single-cell RNA sequencing, which provide valuable insight into splenic neutrophil development and heterogeneity, are also discussed. Finally, we examine how tumors subvert this natural pathway in the spleen to generate granulocytic suppressor cells to promote tumor growth.

Bone Mineral Density, C-Terminal Telopeptide of Type I Collagen, and Osteocalcin as Monitoring Parameters of Bone Remodeling in CML Patients Undergoing Imatinib Therapy: A Basic Science and Clinical Review

Background: Chronic myeloid leukemia (CML) is one of the most commonly found types of myeloproliferative neoplasms, characterized by increased proliferation of granulocytic cells without losing their differentiation ability. Imatinib, a tyrosine kinase inhibitor (TKI), can be effectively used as therapy for CML. However, Imatinib can affect bone turnover thus having clinical implications on the bones of CML patients undergoing long-term Imatinib therapy. However, parameters that can accurately describe the bone condition in CML patients receiving Imatinib still need further study. A combination of imaging techniques such as bone mineral density (BMD) and bone turnover activity markers such as C-terminal telopeptide of type I collagen (CTX-1) and osteocalcin has the potential to be used as monitoring parameters for bone density abnormalities in CML patients receiving Imatinib. Objectives: This article explains the rationale for using BMD, CTX-1, and osteocalcin as monitoring parameters of bone remodeling in CML patients receiving Imatinib. Results: First, the physiological process of bone turnover will be explained. Then, we describe the role of tyrosine kinase in bone metabolism. Next, the impact of Imatinib on BMD, CTX-1, and osteocalcin will be explained. Conclusion: The assessment of bone health of CML patients on Imatinib should include both BMD tests and bone turnover marker assays such as CTX-1 and osteocalcin.

Single-cell profiling reveals a conserved role for hypoxia-inducible factor signaling during human craniotomy infection.

Neurosurgeries complicated by infection are associated with prolonged treatment and significant morbidity. Craniotomy is a common neurosurgical procedure; however, the cellular and molecular signatures associated with craniotomy infection in human subjects are unknown. A retrospective study of over 2,500 craniotomies reveals diverse patient demographics, pathogen identity, and surgical landscapes associated with infection. Leukocyte profiling in patient tissues from craniotomy infection characterizes a predominance of granulocytic myeloid-derived suppressor cells that may arise from transmigrated blood neutrophils, based on single-cell RNA sequencing (scRNA-seq) trajectory analysis. Single-cell transcriptomic analysis identifies metabolic shifts in tissue leukocytes, including a conserved hypoxia-inducible factor (HIF) signature. The importance of HIF signaling was validated using a mouse model of Staphylococcus aureus craniotomy infection, where HIF inhibition increases chemokine production and leukocyte recruitment, exacerbating tissue pathology. These findings establish conserved metabolic and transcriptional signatures that may represent promising future therapeutic targets for human craniotomy infection in the face of increasing antimicrobial resistance.

LTBR acts as a novel immune checkpoint of tumor-associated macrophages for cancer immunotherapy.

Tumor-associated macrophages (TAMs) greatly contribute to immune checkpoint inhibitor (ICI) resistance of cancer. However, its underlying mechanisms and whether TAMs can be promising targets to overcome ICI resistance remain to be unveiled. Through integrative analysis of immune multiomics data and single-cell RNA-seq data (iMOS) in lung adenocarcinoma (LUAD), lymphotoxin β receptor (LTBR) is identified as a potential immune checkpoint of TAMs, whose high expression, duplication, and low methylation are correlated with unfavorable prognosis. Immunofluorescence staining shows that the infiltration of LTBR+ TAMs is associated with LUAD stages, immunotherapy failure, and poor prognosis. Mechanistically, LTΒR maintains immunosuppressive activity and M2 phenotype of TAMs by noncanonical nuclear factor kappa B and Wnt/β-catenin signaling pathways. Macrophage-specific knockout of LTBR hinders tumor growth and prolongs survival time via blocking TAMimmunosuppressive activity and M2 phenotype. Moreover, TAM-targeted delivery of LTΒR small interfering RNA improves the therapeutic effect of ICI via reversing TAM-mediated immunosuppression, such as boosting cytotoxic CD8+ T cells and inhibiting granulocytic myeloid-derived suppressor cells infiltration. Taken together, we bring forth an immune checkpoint discovery pipeline iMOS, identify LTBR as a novel immune checkpoint of TAMs, and propose a new immunotherapy strategy by targeting LTBR+ TAMs.

Neutrophil-specific interactome of ARHGAP25 reveals novel partners and regulatory insights

ARHGAP25, a crucial molecule in immunological processes, serves as a Rac-specific GTPase-activating protein. Its role in cell migration and phagocyte functions, affecting the outcome of complex immunological diseases such as rheumatoid arthritis, renders it a promising target for drug research. Despite its importance, our knowledge of its intracellular interactions is still limited. This study employed proteomic analysis of glutathione S-transferase (GST)-tag pulldowns and co-immunoprecipitation from neutrophilic granulocyte cell lysate, revealing 76 candidates for potential physical interactions that complement ARHGAP25’s known profile. Notably, four small GTPases (RAC2, RHOG, ARF4, and RAB27A) exhibited high affinity for ARHGAP25. The ARHGAP25–RAC2 and ARHGAP25–RHOG interactions appeared to be affected by the activation state of the small GTPases, suggesting a GTP–GDP cycle-dependent interaction. In silico dimer prediction pinpointed ARHGAP25’s GAP domain as a credible binding interface, suggesting its suitability for GTP hydrolysis. Additionally, a list of Fc receptor-related kinases, phosphatases, and three of the 14-3-3 members were identified as potential partners, with in silico predictions highlighting eight binding sites, presenting novel insight on a potential regulatory mechanism for ARHGAP25.

Tumor necrosis factor regulates leukocyte recruitment but not bacterial persistence during Staphylococcus aureus craniotomy infection

BackgroundCraniotomy is a common neurosurgery used to treat intracranial pathologies. Nearly 5% of the 14 million craniotomies performed worldwide each year become infected, most often with Staphylococcus aureus (S. aureus), which forms a biofilm on the surface of the resected bone segment to establish a chronic infection that is recalcitrant to antibiotics and immune-mediated clearance. Tumor necrosis factor (TNF), a prototypical proinflammatory cytokine, has been implicated in generating protective immunity to various infections. Although TNF is elevated during S. aureus craniotomy infection, its functional importance in regulating disease pathogenesis has not been explored.MethodsA mouse model of S. aureus craniotomy infection was used to investigate the functional importance of TNF signaling using TNF, TNFR1, and TNFR2 knockout (KO) mice by quantifying bacterial burden, immune infiltrates, inflammatory mediators, and transcriptional changes by RNA-seq. Complementary experiments examined neutrophil extracellular trap formation, leukocyte apoptosis, phagocytosis, and bactericidal activity.ResultsTNF transiently regulated neutrophil and granulocytic myeloid-derived suppressor cell recruitment to the brain, subcutaneous galea, and bone flap as evident by significant reductions in both cell types between days 7 to 14 post-infection coinciding with significant decreases in several chemokines, which recovered to wild type levels by day 28. Despite these defects, bacterial burdens were similar in TNF KO and WT mice. RNA-seq revealed enhanced lymphotoxin-α (Lta) expression in TNF KO granulocytes. Since both TNF and LTα signal through TNFR1 and TNFR2, KO mice for each receptor were examined to assess potential redundancy; however, neither strain had any impact on S. aureus burden. In vitro studies revealed that TNF loss selectively altered macrophage responses to S. aureus since TNF KO macrophages displayed significant reductions in phagocytosis, apoptosis, IL-6 production, and bactericidal activity in response to live S. aureus, whereas granulocytes were not affected.ConclusionThese findings implicate TNF in modulating granulocyte recruitment during acute craniotomy infection via secondary effects on chemokine production and identify macrophages as a key cellular target of TNF action. However, the lack of changes in bacterial burden in TNF KO animals suggests the involvement of additional signals that dictate S. aureus pathogenesis during craniotomy infection.

γ-aminobutyric acid receptor B signaling drives glioblastoma in females in an immune-dependent manner.

Sex differences in immune responses impact cancer outcomes and treatment response, including in glioblastoma (GBM). However, host factors underlying sex specific immune-cancer interactions are poorly understood. Here, we identify the neurotransmitter γ-aminobutyric acid (GABA) as a driver of GBM-promoting immune response in females. We demonstrated that GABA receptor B (GABBR) signaling enhances L-Arginine metabolism and nitric oxide synthase 2 (NOS2) expression in female granulocytic myeloid-derived suppressor cells (gMDSCs). GABBR agonist and GABA analog promoted GBM growth in females in an immune-dependent manner, while GABBR inhibition reduces gMDSC NOS2 production and extends survival only in females. Furthermore, female GBM patients have enriched GABA transcriptional signatures compared to males, and the use of GABA analogs in GBM patients is associated with worse short-term outcomes only in females. Collectively, these results highlight that GABA modulates anti-tumor immune response in a sex-specific manner, supporting future assessment of GABA pathway inhibitors as part of immunotherapy approaches.

Granulocytes and mast cells in AllergoOncology-Bridging allergy to cancer: An EAACI position paper.

Derived from the myeloid lineage, granulocytes, including basophils, eosinophils, and neutrophils, along with mast cells, play important, often disparate, roles across the allergic disease spectrum. While these cells and their mediators are commonly associated with allergic inflammation, they also exhibit several functions either promoting or restricting tumor growth. In this Position Paper we discuss common granulocyte and mast cell features relating to immunomodulatory functions in allergy and in cancer. We highlight key mechanisms which may inform cancer treatment and propose pertinent areas for future research. We suggest areas where understanding the communication between granulocytes, mast cells, and the tumor microenvironment, will be crucial for identifying immune mechanisms that may be harnessed to counteract tumor development. For example, a comprehensive understanding of allergic and immune factors driving distinct neutrophil states and those mechanisms that link mast cells with immunotherapy resistance, might enable targeted manipulation of specific subpopulations, leading to precision immunotherapy in cancer. We recommend specific areas of investigation in AllergoOncology and knowledge exchange across disease contexts to uncover pertinent reciprocal functions in allergy and cancer and allow therapeutic manipulation of these powerful cell populations. These will help address the unmet needs in stratifying and managing patients with allergic diseases and cancer.

Dual roles of CD11b+CD33+HLA-DR-/lowCD14- myeloid-derived suppressor cells with a granulocytic morphology following allogeneic hematopoietic stem cell transplantation: from inflammation promoters to immune suppressors within 90 days.

Granulocytic myeloid-derived suppressor cells (G-MDSCs) show fast recovery following allogeneic hematopoietic stem cell transplantation (allo-HSCT) constituting the major part of peripheral blood in the early phase. Although G-MDSCs mediate immune suppression through multiple mechanisms, they may also promote inflammation under specific conditions. G-MDSCs were isolated from 82 patients following allo-HSCT within 90 days after allo-HSCT, and their interactions with autologous CD3+ T-cells were examined. T-cell proliferation was assessed by flow cytometry following CFSE staining, while differentiation and interferon-γ secretion were characterized using chemokine receptor profiling and ELISpot assays, respectively. NK cell cytotoxicity was evaluated through co-culture with K562 cells. An aGVHD xenogeneic model in humanized mice was employed to study the in vivo effects of human leukocytes. Furthermore, transcriptional alterations in G-MDSCs were analyzed via RNA sequencing to investigate functional transitions. G-MDSCs promoted inflammation in the early-stage, by facilitating cytokine secretion and proliferation of T cells, as well as their differentiation into pro-inflammatory T helper subsets. At day 28, patients with a higher number of G-MDSCs exhibited an increased risk of developing grades II-IV aGvHD. Besides, adoptive transfer of G-MDSCs from patients at day 28 into humanized mice exacerbated aGvHD. However, at day 90, G-MDSCs led to immunosuppression, characterized by upregulated expression of indoleamine 2,3-dioxygenase gene and interleukin-10 secretion, coupled with the inhibition of T cell proliferation. Furthermore, transcriptional analysis of G-MDSCs at day 28 and day 90 revealed that 1445 genes were differentially expressed. These genes were associated with various pathways, revealing the molecular signatures of early post-transplant differentiation in G-MDSCs. In addition, genes linked to the endoplasmic reticulum stress were upregulated in patients without aGvHD. The acquisition of immunosuppressive function by G-MDSCs may depend on the activation of CXCL2 and DERL1 genes. Our findings revealed the alteration in the immune characteristics of G-MDSCs within the first 90 days post-allo-HSCT. Moreover, the quantity of G-MDSCs at day 28 may serve as a predictive indicator for the development of aGvHD.

Effectiveness of Use of UV Lamp in Disinfection of Additioned Vaname Shrimp (Litopenaeus vannamei) Cultivation Media Bacteria Vibrio Harveyi

One of the most promising fisheries products in Indonesia is vaname shrimp (Litopenaeus vannamei), which is also a top export comodity. From year to year the value of shrimp exports and production continues to increase. However, vaname shrimp are susceptible to bacterial and viral attacks, so countermeasures are needed, one of which is by disinfecting the cultivation media using a UV lamp. The aims of this research is to determine the level of effectiveness of UV lamps in disinfection of cultivation media. Experimental methodology with a completely randomized plan (RAL) is the method used in this research, with 4 trials and 3 replications. The results of the research conducted showed that UV lamps were able to reduce the total bacterial count to 4.69 x 108 CFU/mL at UV lamp 41 watt, reduce the total Vibrio bacteria to 30.33 x 106 CFU/mL at UV lamp 41 watt, increase the THC value to 8.83 x 106 CFU/mL cells/ml at P4, hyaline cells range from 31% – 54.67%, granulocyte cell values ​​range from 29.33% – 42%, semigranulocyte cells range from 16% – 27%, the highest AF value at P4 is 32.19%, and the lowest in P1 was 79.69%. Based on the findings, it is possible to conclude that UV lights can be used as a disinfectant of cultivation media, but the right dose or light power is needed to get maximum results.

Extracellular vesicles originating from melanoma cells promote dysregulation in haematopoiesis as a component of cancer immunoediting.

Haematopoiesis dysregulation with the presence of immature myeloid and erythroid immunosuppressive cells are key characteristics of the immune escape phase of tumour development. Here, the role of in vitro generated B16F10 tumour cell-derived extracellular vesicles (tEVs) as indirect cellular communicators, participating in tumour-induced dysregulation of haematopoiesis, was explored. The isolated tEVs displayed features of small EVs with a size range of 100-200nm, expressed the common EV markers CD63, CD9, and Alix, and had a spherical shape with a lipid bilayer membrane. Proteomic profiling revealed significant levels of angiogenic factors, particularly vascular endothelial growth factor (VEGF), osteopontin, and tissue factor, associated with the tEVs. Systemic administration of these tEVs in syngeneic mice induced splenomegaly and disrupted haematopoiesis, leading to extramedullary haematopoiesis, expansion of splenic immature erythroid progenitors, reduced bone marrow cellularity, medullary expansion of granulocytic myeloid suppressor cells, and the development of anaemia. These effects closely mirrored those observed in tumour-bearing mice and were not seen after heat inactivating the tEVs. In vitro studies demonstrated that tEVs independently induced the expansion of bone marrow granulocytic myeloid suppressor cells and B cells while reducing the frequency of cells in the erythropoietic lineage. These effects of tEVs were significantly abrogated by the blockade of VEGF or heat inactivation. Our findings underscore the important role of tEVs in dysregulating haematopoiesis during the immune escape phase of cancer immunoediting, suggesting their potential as targets for addressing immune evasion and reinstating normal hematopoietic processes.

Tregs, gMDSCs, and levels of soluble MICA as prognostic biomarkers for combined NEO-201 and pembrolizumab.

2530 Background: The humanized IgG1 monoclonal antibody NEO-201 binds to Core 1 and/or extended Core 1 O-glycans expressed by several human solid and blood tumors, as well as neutrophils, and mediates killing of cancer cells, neutrophils, regulatory T cells (Tregs) and granulocytic myeloid-derived suppressor cells (gMDSCs) via ADCC and CDC. Resistance to PD-1/PD-L1 blockade may be due to the accumulation of Tregs and gMDSCs in the tumor microenvironment (TME). NEO-201 was proven to bind and reduce the amount of circulating Tregs in cancer patients. This supports the rationale of the ongoing phase II clinical trial (NCT03476681) evaluating the activity of NEO-201 with pembrolizumab in adults with solid tumors resistant to prior checkpoint inhibitors. Furthermore, elevated serum levels of soluble MICA (sMICA) have been correlated with decreased NK cell activity. Methods: Cancer patients were treated each cycle with 3 doses of NEO-201 1.5mg/kg every 2 weeks and pembrolizumab 400mg IV every 6 weeks and imaged for response every 2 cycles. PBMCs and serum from cancer patients pre- and at multiple time points post-treatment were used to evaluate the percentage of circulating gMDSCs and Tregs (flow cytometry) and sMICA levels (ELISA). Results: NEO-201 recognizes naïve Tregs (nTregs: CD3+/CD4+/CD45RA+/Foxp3low cells) but not effector Tregs (eTregs: CD3+/CD4+/CD45RA-/Foxp3high cells). NEO-201 also binds to gMDSCs, defined as HLA-DRneg/CD33+/CD15+/ CD14neg/CD66b+ cells. In patients with durable SD, a patient with HNSCC (SD >11 months) showed >50% reduction of nTregs and >90% of gMDSCs at C3D1 (beginning of cycle 3) compared to baseline. One patient with cervical cancer (SD >8 months) showed 40% reduction of nTregs at C3D1 compared to baseline, while gMDSCs trended down to baseline levels at C3D1 after initial increase. Conversely, we observed a general uptrend of nTregs and gMDSCs in patients with PD after treatment. Median serum levels of sMICA pre-treatment were 33-fold higher in patients with PD compared to patients with SD. Levels of sMICA remained elevated in patients with PD and low in patients with SD at all time points post-treatment. Conclusions: Depletion of circulating Tregs and gMDSCs may prevent their accumulation in the TME and enhance the efficacy of pembrolizumab in subjects with tumors resistant to checkpoint inhibitors. The decrease in circulating nTregs and gMDSCs after treatment with NEO-201 and pembrolizumab was associated with durable SD. High levels of sMICA can impair ADCC mediated by NEO-201, resulting in poor clinical response. Low levels of sMICA in patients with SD, together with the reduction of both circulating nTregs and gMDSCs, could be favorable prognostic markers for clinical benefit following treatment with NEO-201 and pembrolizumab. Ongoing enrollment in this clinical trial will validate these findings in larger cohorts. Clinical trial information: NCT03476681 .

CCL9/CCR1 axis-driven chemotactic nanovesicles for attenuating metastasis of SMAD4-deficient colorectal cancer by trapping TGF-β

SMAD4 deficiency in colorectal cancer (CRC) is highly correlated with liver metastasis and high mortality, yet there are few effective precision therapies available. Here, we show that CCR1+-granulocytic myeloid-derived suppressor cells (G-MDSCs) are highly infiltrated in SMAD4-deficient CRC via CCL15/CCR1 and CCL9/CCR1 axis in clinical specimens and mouse models, respectively. The excessive TGF-β, secreted by tumor-infiltrated CCR1+-G-MDSCs, suppresses the immune response of cytotoxic T lymphocytes (CTLs), thus facilitating metastasis. Hereby, we develop engineered nanovesicles displaying CCR1 and TGFBR2 molecules (C/T-NVs) to chemotactically target the tumor driven by CCL9/CCR1 axis and trap TGF-β through TGF-β-TGFBR2 specific binding. Chemotactic C/T-NVs counteract CCR1+-G-MDSC infiltration through competitive responding CCL9/CCR1 axis. C/T-NVs-induced intratumoral TGF-β exhaustion alleviates the TGF-β-suppressed immune response of CTLs. Collectively, C/T-NVs attenuate liver metastasis of SMAD4-deficient CRC. In further exploration, high expression of programmed cell death ligand-1 (PD-L1) is observed in clinical specimens of SMAD4-deficient CRC. Combining C/T-NVs with anti-PD-L1 antibody (aPD-L1) induces tertiary lymphoid structure formation with sustained activation of CTLs, CXCL13+-CD4+ T, CXCR5+-CD20+ B cells, and enhanced secretion of cytotoxic cytokine interleukin-21 and IFN-γ around tumors, thus eradicating metastatic foci. Our strategy elicits pleiotropic antimetastatic immunity, paving the way for nanovesicle-mediated precision immunotherapy in SMAD4-deficient CRC.

Increased CD123 + HLA-DR- Granulocytes in Allergic Rhinitis and Influence of Allergens on Expression of Cell Membrane Markers.

It is reported that CD123 + HLA-DR- cells in PBMC are basophils, and CD203c, CD63, and FcεRI molecules are activation markers of basophils. However, little is known of CD123 + HLA-DR-cells in blood granulocytes. To investigate the presence of CD123 + HLA-DR- cells in the blood granulocytes and peripheral PBMC of patients with allergic rhinitis (AR), as well as the impact of allergens on the cell membrane markers of basophils. Flow cytometry was used to detect the expression of the membrane molecules. While CD123 + HLA-DR- PBMCs are representative of basophils, their presence did not significantly change in patients with AR. In contrast, both the percentage and number of CD123 + HLA-DR- granulocytes, which make up only up to 50% of basophils, were significantly increased in patients with seasonal (sAR) and perennial AR (pAR). CD63+, CD203c+, and FcεRIα+ cells within CD123 + HLA-DR- granulocytes also showed enhanced activity in patients with AR. Allergen extracts from house dust mite allergen extract (HDME) and Artemisia sieversiana wild extract further increased the number of CD123 + HLA-DR- cells in granulocytes of sAR and pAR patients, as well as in PBMCs of pAR patients. The use of CD123 + HLA-DR- granulocytes and PBMC may not be sufficient for diagnosing AR. Allergens could potentially contribute to the development of AR by influencing the number of CD123 + HLA-DR- cells, as well as the expression of CD63, CD203c, and FcεRIαin these cells.

TLR9 promotes monocytic myeloid-derived suppressor cell induction during JEV infection

Myeloid-derived suppressor cells (MDSCs) are a group of heterologous populations of immature bone marrow cells consisting of progenitor cells of macrophages, dendritic cells and granulocytes. Recent studies have revealed that the accumulation of MDSCs in the mouse spleen plays a pivotal role in suppressing the immune response following JEV infection. However, the mechanisms by which JEV induces MDSCs are poorly understood. Here, it was found that JEV infection induces mitochondrial damage and the release of mitochondrial DNA (mtDNA), which further leads to the activation of TLR9. TLR9 deficiency decreases the M-MDSCs population and their suppressive function both in vitro and in vivo. Moreover, the increase of MHCⅡ expression on antigen-presenting cells and CD28 expression on T cells in TLR9−/− mice was positively correlated with M-MDSCs reduction. Accordingly, the survival rate of TLR9−/− mice dramatically increased after JEV infection. These findings reveal the connections of mitochondrial damage and TLR9 activation to the induction of M-MDSCs during JEV infection.

Glycolysis and HIF-1α signaling are important for granulocytic myeloid-derived suppressor cell activity during Staphylococcus aureus biofilm infection

Abstract Prosthetic joint infection (PJI) is a severe complication following hip and knee arthroplasty and is reported to affect 1-2% of total arthroplasties. Staphylococcus aureus (S. aureus) contributes to most PJIs, with a higher treatment failure rate partly due to biofilm formation. Granulocytic myeloid-derived suppressor cell (G-MDSC) expansion and IL-10 secretion are critical for inducing an immunosuppressive environment that supports S. aureus persistence during PJI. Recently, S. aureus-derived lactate was shown to promote IL-10 production by G-MDSCs. However, the importance of G-MDSC metabolism in programming their anti-inflammatory activity is unknown. Single-cell RNA sequencing (scRNA-seq) of infected tissues from a mouse model of S. aureus PJI revealed enrichment of glycolysis and hypoxia pathway genes in G-MDSCs. Increased glycolytic activity in G-MDSCs was confirmed following S. aureus biofilm co-culture and attenuating glycolysis with either 2-deoxyglucose or granulocyte-specific HIF-1a deletion reduced bacterial growth during S. aureus PJI. These changes coincided with less G-MDSC suppressive activity and a transformation towards a proinflammatory phenotype. scRNA-seq of samples from PJI patients revealed heightened glycolysis and hypoxia signatures, supporting findings with the mouse PJI model. Together, these results suggest that glycolysis and HIF-1a pathways are critical for the anti-inflammatory activity of G-MDSCs during S. aureus biofilm infection.

Hematological effects for rabbits immunized by Corynebacterium pseudotuberculosis sonicated antigen

Background: Analyzing Hematological parameters is usually in use to monitor various circumstances such as infection, inflammation and anemia. For that we studied blood parameters which are in touch with immunized laboratory animals (rabbits) after immunizing these animals with Corynebacterium pseudotuberculosis then used an adjuvant of Pseudomonas aeruginosa to stimulate the immune response cells after 2 to 3 weeks of immunization. Different concentrations were used to examine the effect on the animals’ blood parameters changing.Methods: Corynebacterium pseudotuberculosis bacterium was used at current study for evaluating the effect of immunizing laboratory rabbits with two different immune stimulators working together as adjuvant on the blood picture with the aid of 12 breed rabbits from different genders, dividing them at the base of inoculation with pre-killed and sonicated cells of bacteria (antigens) onto 4 categories as following: the group-1 members of rabbits were injected with the antigen, group-2 and group-3 included rabbits injected with the two inoculations at various concentrations; final group, group-4 was the control group.Results: Thus, white blood cells known to be our main line of defenses act firstly in our body towards different infecting microorganisms, their count always was increased through the immunization correlated with other measured parameters. From the results we can see that the lymphocytes percentage had no alteration between the infected and the immunized compared to control group, while the percentage of MID that refers to the monocytes was at the same range in both infected and the immunized control rabbit’s groups. Granulocyte cells percentage, which is including heterophilic, basophilic and eosinophilic cells, was significantly decreased (P>0.05) compared with control at all groups.Conclusion: We conclude that C. pseudotuberculosis might function as a potent immunogen to notice the complete blood picture variations.Keywords: Corynebacterium pseudotuberculosis; Pseudomonas aeruginosa; Sonicated antigen; Blood parameters; Blood picture; Adjuvant

A lasso and random forest model using flow cytometry data identifies primary myelofibrosis.

Thrombocythemia (ET), polycythemia vera (PV), primary myelofibrosis (PMF), prefibrotic/early (pre-PMF), and overt fibrotic PMF (overt PMF) are classical Philadelphia-Negative (Ph-negative) myeloproliferative neoplasms (MPNs). Differentiating between these types based on morphology and molecular markers is challenging. This study aims to clarify the application of flow cytometry in the diagnosis and differential diagnosis of classical MPNs. This study retrospectively analyzed the immunophenotypes, clinical characteristics, and laboratory findings of 211 Ph-negative MPN patients, including ET, PV, pre-PMF, overt PMF, and 47 controls. Compared to ET and PV, PMF differed in white blood cells, hemoglobin, blast cells in the peripheral blood, abnormal karyotype, and WT1 gene expression. PMF also differed from controls in CD34+ cells, granulocyte phenotype, monocyte phenotype, percentage of plasma cells, and dendritic cells. Notably, the PMF group had a significantly lower plasma cell percentage compared with other groups. A lasso and random forest model select five variables (CD34+CD19+cells and CD34+CD38- cells on CD34+cells, CD13dim+CD11b- cells in granulocytes, CD38str+CD19+/-plasma, and CD123+HLA-DR-basophils), which identify PMF with a sensitivity and specificity of 90%. Simultaneously, a classification and regression tree model was constructed using the percentage of CD34+CD38- on CD34+ cells and platelet counts to distinguish between ET and pre-PMF, with accuracies of 94.3% and 83.9%, respectively. Flow immunophenotyping aids in diagnosing PMF and differentiating between ET and PV. It also helps distinguish pre-PMF from ET and guides treatment decisions.

The immunologic abnormalities in patients with paroxysmal nocturnal hemoglobinuria are associated with disease progression.

To suggest the presence of a hyperimmune state in patients, and indicate that immune system attack on glycosylphosphatidylinositol (+) (GPI+) cells while escaping GPI- cell immunity. We retrospective the immune cell subtypes in peripheral blood from 25 patients visiting Tianjin Medical University General Hospital, Tianjin, China, with classical paroxysmal nocturnal hemoglobinuria (PNH) and 50 healthy controls. The total CD3+ and CD3+CD8+ cell levels were higher in patients with PNH. The CD3+ cells are positively, correlated with lactate dehydrogenase (LDH; r=0.5453, p=0.0040), indirect bilirubin (r=0.4260, p=0.0379) and Flear- cells in monocytes (r=0.4099, p=0.0303). However, a negative correlation was observed between CD3+ cells and hemoglobin (r= -0.4530, p=0.0105). The total CD19+ cells decreased in patients, and CD19+ cells were negatively correlated with LDH (r= -0.5640, p=0.0077) and Flear- cells in monocytes (r= -0.4432, p=0.0341). Patients showed an increased proportion of total dendritic cells (DCs), with a higher proportion of myeloid DCs (mDCs) within the DC population. Moreover, the proportion of mDC/DC was positively correlated with CD59- cells (II + III types) in red cells (r=0.7941, p=0.0004), Flear- cells in granulocytes (r=0.5357, p=0.0396), and monocytes (r=0.6445, p=0.0095). Our results demonstrated that immune abnormalities are associated with PNH development.

Morphological changes in gum tissue during thiram intoxication followed by correction with resveratrol and sea buckthorn oil

Background: The widespread use of pesticides can lead to a violation of the ecological balance between the environment and the human body, provoking the development of oxidative stress, which leads to an increase in toxic forms of damage to organs and systems of the macroorganism. In order to suppress oxidative stress that develops in many pathological processes, it is advisable to use antioxidants that inhibit the formation of free radicals. The aim of the study: To study the morphological changes in gum tissues in case of thiram intoxication followed by correction with resveratrol and sea buckthorn oil. Materials and methods: The experiment was performed on 50 rats that received the fungicide thiram for 28 days, followed by correction with resveratrol or sea buckthorn oil. To assess the severity of inflammatory changes in histological preparations, morphofunctional changes in the mucosal epithelium and the cellular composition of the subepithelial layer were analysed. Fibroblastic differon cells, macrophages and inflammatory cells of granulocytes and lymphocytes were differentiated by karyological signs, and the thickness of the epithelial layer, the area occupied by the fibrous, vascular and cellular components were measured. Results: In the experimental groups “Thiram 28 days” and “Thiram+standard diet” in all rats, the development of an inflammatory reaction of the gum tissue was observed, manifested in hyperkeratosis and an increase in the thickness of the epithelium up to 173.438 microns and 230.568 microns, diffuse polymorphocellular infiltration of the subepithelial layer mainly due to granulocytes to 21.40 and 21.92, respectively, expansion of the lumen and blood filling of the vessels of the microvasculature. After the use of resveratrol and sea buckthorn oil, the thickness of the epithelial layer was restored to 114,07 and 190,03 microns, respectively, interstitial edema and cell infiltration of the subepithelial layer decreased. At the same time, due to migration and increase of fibroblastic differon cells (28.96 and 29.44), neocollagenogenesis processes and an increase in the fibrous component were observed up to 65.62 and 33.69 after application of resveratrol and sea buckthorn oil, respectively. Conclusion: The effectiveness of monotherapy with resveratrol and sea buckthorn oil in normalizing the morphology of gum tissue has been proven. At the same time, resveratrol has both an anti-inflammatory effect and a high regenerative effect.