Increased Inflammasome Activation Is Associated with Aging and Chronic Myelomonocytic Leukemia Disease Severity.

Aging causes chronic low-grade inflammation known as inflamm-aging. It is a risk factor for several chronic disorders, including chronic myelomonocytic leukemia (CMML), a hematological malignancy that is most prevalent in older people. Recent studies suggest a critical role for the NLRP3 (NOD-, LRR- and pyrin domain-containing protein 3) inflammasome in inflamm-aging. However, the mechanisms involved in NLRP3 activation in aging and its involvement in CMML progression are not fully understood. In this study, we report that aging increases IL-1ß production upon NLRP3 activation in human CD14+ monocytes. Interestingly, we found that the TLR1/2 agonist Pam3CSK4 directly activates the NLRP3 inflammasome in monocytes from older but not from younger healthy donors. Furthermore, we observed a dichotomous response to NLRP3 inflammasome activation in monocytes from a small cohort of CMML patients, and some patients produced high levels of IL-1ß and some patients produced low levels of IL-1ß compared with older healthy donors. Intriguingly, CMML patients with heightened NLRP3 activation showed increased treatment dependency and disease severity. Collectively, our results suggest that aging causes increased sensitivity to NLRP3 inflammasome activation at a cellular level, which may explain increased inflammation and immune dysregulation in older individuals. Furthermore, NLRP3 inflammasome activation was dysregulated in a small cohort of CMML patients and was positively correlated with disease severity.

JAK2-mutant hematopoietic cells display metabolic alterations that can be targeted to treat myeloproliferative neoplasms.

Increased energy requirement and metabolic reprogramming are hallmarks of cancer cells. We show that metabolic alterations in hematopoietic cells are fundamental to the pathogenesis of mutant JAK2-driven myeloproliferative neoplasms (MPNs). We found that expression of mutant JAK2 augmented and subverted metabolic activity of MPN cells, resulting in systemic metabolic changes in vivo, including hypoglycemia, adipose tissue atrophy, and early mortality. Hypoglycemia in MPN mouse models correlated with hyperactive erythropoiesis and was due to a combination of elevated glycolysis and increased oxidative phosphorylation. Modulating nutrient supply through high-fat diet improved survival, whereas high-glucose diet augmented the MPN phenotype. Transcriptomic and metabolomic analyses identified numerous metabolic nodes in JAK2-mutant hematopoietic stem and progenitor cells that were altered in comparison with wild-type controls. We studied the consequences of elevated levels of Pfkfb3, a key regulatory enzyme of glycolysis, and found that pharmacological inhibition of Pfkfb3 with the small molecule 3PO reversed hypoglycemia and reduced hematopoietic manifestations of MPNs. These effects were additive with the JAK1/2 inhibitor ruxolitinib in vivo and in vitro. Inhibition of glycolysis by 3PO altered the redox homeostasis, leading to accumulation of reactive oxygen species and augmented apoptosis rate. Our findings reveal the contribution of metabolic alterations to the pathogenesis of MPNs and suggest that metabolic dependencies of mutant cells represent vulnerabilities that can be targeted for treating MPNs.

Parvovirus B19 Passive Transmission by Transfusion of Intercept® Blood System-Treated Platelet Concentrate.

BACKGROUND: Pathogen reduction methods for blood components are effective for a large number of viruses though less against small, non-enveloped viruses such as Parvovirus B19 (B19V). This article describes the passive transmission by transfusion of two B19V-contaminated pooled platelet concentrates (PCs) which were treated with the Intercept® blood pathogen reduction system. CASE REPORTS: Two transfusion cases of B19V-contaminated Intercept-treated pooled PCs were described. Due to the analysis delay, the PCs were already transfused. The viral content of each donation was 4.87 × 10(10) IU/ml in case 1and 1.46 × 10(8) IU/ml in case 2. B19V (52 IU/ml) was detected in the recipient of the case 1 PC, whereas no virus could be detected in the case 2 PC recipient. A B19V IgM response and a transient boost of the underlying B19V IgG immune status and was observed in recipient 1. Recipient of the case 2 PC remained B19V IgG- and IgM-negative. B19V DNA sequence and phylogenetic analysis revealed a 100% homology between donor and recipient. CONCLUSION: This report describes passive B19V transmission by a PC with very high B19 viral load which elicited a transient boost of the B19V immunity, but not by a PC with a lower B19V content, suggesting that there is a B19 viral load threshold value at which B19V inactivation is exceeded.

Ribonuclease inhibitor and angiogenin system regulates cell type-specific global translation.

Translation of mRNAs is a fundamental process that occurs in all cell types of multicellular organisms. Conventionally, it has been considered a default step in gene expression, lacking specific regulation. However, recent studies have documented that certain mRNAs exhibit cell type-specific translation. Despite this, it remains unclear whether global translation is controlled in a cell type-specific manner. By using human cell lines and mouse models, we found that deletion of the ribosome-associated protein ribonuclease inhibitor 1 (RNH1) decreases global translation selectively in hematopoietic-origin cells but not in the non-hematopoietic-origin cells. RNH1-mediated cell type-specific translation is mechanistically linked to angiogenin-induced ribosomal biogenesis. Collectively, this study unravels the existence of cell type-specific global translation regulators and highlights the complex translation regulation in vertebrates.

Immune-Mediated Thrombotic Thrombocytopenic Purpura Following mRNA-Based COVID-19 Vaccine BNT162b2: Case Report and Mini-Review of the Literature.

Introduction: An increasing number of case reports have associated vaccinations against coronavirus disease 2019 (COVID-19) with immune-mediated thrombotic thrombocytopenic purpura (iTTP), a very rare but potentially life-threatening thrombotic microangiopathy, which leads to ischemic organ dysfunction. Thrombus formation in iTTP is related to a severe deficiency of the specific von Willebrand-factor-cleaving protease ADAMTS13 due to ADAMTS13 autoantibodies. Methods: We present a case of iTTP following exposure to the mRNA-based COVID-19 vaccine BNT162b2 (Comirnaty®, Pfizer-BioNTech). In addition, we review previously reported cases in the literature and assess current evidence. Results: Apart from our case, twenty cases of iTTP occurring after COVID-19 vaccination had been published until the end of November 2021. There were 11 male and 10 female cases; their median age at diagnosis was 50 years (range 14-84 years). Five patients (24%) had a preexisting history of iTTP. Recombinant adenoviral vector-based vaccines were involved in 19%, mRNA-based vaccines in 81%. The median onset of symptoms after vaccination was 12 days (range 5-37), with 20 cases presenting within 30 days. Treatment included therapeutic plasma exchange in all patients. Additional rituximab, caplacizumab, or both these treatments were given in 43% (9/21), 14% (3/21), and 24% (5/21) of cases, respectively. One patient died, despite a prolonged clinical course in one patient, all surviving patients were in clinical remission at the end of the observational period. Conclusion: Clinical features of iTTP following COVID-19 vaccination were in line with those of pre-pandemic iTTP. When timely initiated, an excellent response to standard treatment was seen in all cases. ADAMTS13 activity should be determined pre-vaccination in patients with a history of a previous iTTP episode. None of the reported cases met the WHO criteria for assessing an adverse event following immunization (AEFI) as a consistent causal association to immunization. Further surveillance of safety data and additional case-based assessment are needed.

LRR-protein RNH1 dampens the inflammasome activation and is associated with COVID-19 severity.

Inflammasomes are cytosolic innate immune sensors of pathogen infection and cellular damage that induce caspase-1-mediated inflammation upon activation. Although inflammation is protective, uncontrolled excessive inflammation can cause inflammatory diseases and can be detrimental, such as in coronavirus disease (COVID-19). However, the underlying mechanisms that control inflammasome activation are incompletely understood. Here we report that the leucine-rich repeat (LRR) protein ribonuclease inhibitor (RNH1), which shares homology with LRRs of NLRP (nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain containing) proteins, attenuates inflammasome activation. Deletion of RNH1 in macrophages increases interleukin (IL)-1ß production and caspase-1 activation in response to inflammasome stimulation. Mechanistically, RNH1 decreases pro-IL-1ß expression and induces proteasome-mediated caspase-1 degradation. Corroborating this, mouse models of monosodium urate (MSU)-induced peritonitis and lipopolysaccharide (LPS)-induced endotoxemia, which are dependent on caspase-1, respectively, show increased neutrophil infiltration and lethality in Rnh1 -/- mice compared with wild-type mice. Furthermore, RNH1 protein levels were negatively related with disease severity and inflammation in hospitalized COVID-19 patients. We propose that RNH1 is a new inflammasome regulator with relevance to COVID-19 severity.

Inflammasome Activation in Myeloid Malignancies-Friend or Foe?

Myeloid malignancies including myelodysplastic syndromes, myeloproliferative neoplasms and acute myeloid leukemia are heterogeneous disorders originating from mutated hematopoietic stem and progenitor cells (HSPCs). Genetically, they are very heterogeneous and characterized by uncontrolled proliferation and/or blockage of differentiation of abnormal HSPCs. Recent studies suggest the involvement of inflammasome activation in disease initiation and clonal progression. Inflammasomes are cytosolic innate immune sensors that, upon activation, induce caspase-1 mediated processing of interleukin (IL) -1-cytokine members IL-1ß and IL-18, as well as initiation of gasdermin D-dependent pyroptosis. Inflammasome activation leads to a pro-inflammatory microenvironment in the bone marrow, which drives proliferation and may induce clonal selection of mutated HSPCs. However, there are also contradictory data showing that inflammasome activation actually counteracts leukemogenesis. Overall, the beneficial or detrimental effect of inflammasome activation seems to be highly dependent on mutational, environmental, and immunological contexts and an improved understanding is fundamental to advance specific therapeutic targeting strategies. This review summarizes current knowledge about this dichotomous effect of inflammasome activation in myeloid malignancies and provides further perspectives on therapeutic targeting.

Proviral integration site for Moloney murine leukemia virus 1, but not phosphatidylinositol-3 kinase, is essential in the antiapoptotic signaling cascade initiated by IL-5 in eosinophils.

BACKGROUND: Eosinophil differentiation, activation, and survival are largely regulated by IL-5. IL-5-mediated transmembrane signal transduction involves both Lyn-mitogen-activated protein kinases and Janus kinase 2-signal transducer and activator of transcription pathways. OBJECTIVE: We sought to determine whether additional signaling molecules/pathways are critically involved in IL-5-mediated eosinophil survival. METHODS: Eosinophil survival and apoptosis were measured in the presence and absence of IL-5 and defined pharmacologic inhibitors in vitro. The specific role of the serine/threonine kinase proviral integration site for Moloney murine leukemia virus (Pim) 1 was tested by using HIV-transactivator of transcription fusion proteins containing wild-type Pim-1 or a dominant-negative form of Pim-1. The expression of Pim-1 in eosinophils was analyzed by means of immunoblotting and immunofluorescence. RESULTS: Although pharmacologic inhibition of phosphatidylinositol-3 kinase (PI3K) by LY294002, wortmannin, or the selective PI3K p110delta isoform inhibitor IC87114 was successful in each case, only LY294002 blocked increased IL-5-mediated eosinophil survival. This suggested that LY294002 inhibited another kinase that is critically involved in this process in addition to PI3K. Indeed, Pim-1 was rapidly and strongly expressed in eosinophils after IL-5 stimulation in vitro and readily detected in eosinophils under inflammatory conditions in vivo. Moreover, by using specific protein transfer, we identified Pim-1 as a critical element in IL-5-mediated antiapoptotic signaling in eosinophils. CONCLUSIONS: Pim-1, but not PI3K, plays a major role in IL-5-mediated antiapoptotic signaling in eosinophils.

The effects of intravenous iron supplementation on fatigue and general health in non-anemic blood donors with iron deficiency: a randomized placebo-controlled superiority trial.

We investigated whether intravenous iron supplementation improves fatigue and general health in non-anemic repeat adult blood donors with iron deficiency (ferritin ≤ 50 µg/L). Of 1,487 potentially eligible participants, 203 were randomly assigned to a single intravenous dose of 800 mg iron-carboxymaltose and 202 to placebo; 393 participants completed the trial. At 6 to 8 weeks after intervention, self-rated mean fatigue scores (numeric rating scale from 1-10, primary outcome) were 3.9 ± 1.8 in the iron supplementation group and 4.0 ± 2.2 in the placebo group, showing no group difference (p = 0.819). Pre-specified subgroup analyses of gender, ferritin < 25 µg/L and fatigue ≥ 4 points, as well as exploratory analyses of lower ferritin cut-offs did not reveal any between-group differences. In terms of secondary outcomes, the mean differences were 114.2 µg/L for ferritin (95% CI 103.1-125.3) and 5.7 g/L for hemoglobin (95% CI 4.3-7.2) with significantly higher values in the iron supplementation group. No group differences were observed for different measures of general well-being and other clinical and safety outcomes. Intravenous iron supplementation compared with placebo resulted in increase of ferritin and hemoglobin levels in repeat blood donors with low iron stores, yet had no effect on fatigue and general well-being.

Ribonuclease inhibitor 1 regulates erythropoiesis by controlling GATA1 translation.

Ribosomal proteins (RP) regulate specific gene expression by selectively translating subsets of mRNAs. Indeed, in Diamond-Blackfan anemia and 5q- syndrome, mutations in RP genes lead to a specific defect in erythroid gene translation and cause anemia. Little is known about the molecular mechanisms of selective mRNA translation and involvement of ribosomal-associated factors in this process. Ribonuclease inhibitor 1 (RNH1) is a ubiquitously expressed protein that binds to and inhibits pancreatic-type ribonucleases. Here, we report that RNH1 binds to ribosomes and regulates erythropoiesis by controlling translation of the erythroid transcription factor GATA1. Rnh1-deficient mice die between embryonic days E8.5 and E10 due to impaired production of mature erythroid cells from progenitor cells. In Rnh1-deficient embryos, mRNA levels of Gata1 are normal, but GATA1 protein levels are decreased. At the molecular level, we found that RNH1 binds to the 40S subunit of ribosomes and facilitates polysome formation on Gata1 mRNA to confer transcript-specific translation. Further, RNH1 knockdown in human CD34+ progenitor cells decreased erythroid differentiation without affecting myelopoiesis. Our results reveal an unsuspected role for RNH1 in the control of GATA1 mRNA translation and erythropoiesis.

RhoH is a negative regulator of eosinophilopoiesis.

Eosinophils are frequently elevated in pathological conditions and can cause tissue damage and disease exacerbation. The number of eosinophils in the blood is largely regulated by factors controlling their production in the bone marrow. While several exogenous factors, such as interleukin-5, have been described to promote eosinophil differentiation, comparatively little is known about eosinophil-intrinsic factors that control their de novo generation. Here, we report that the small atypical GTPase RhoH is induced during human eosinophil differentiation, highly expressed in mature blood eosinophils and further upregulated in patients suffering from a hypereosinophilic syndrome. Overexpression of RhoH increases, in a Rho-associated protein kinase-dependent manner, the expression of GATA-2, a transcription factor involved in regulating eosinophil differentiation. In RhoH-/- mice, we observed reduced GATA-2 expression as well as accelerated eosinophil differentiation both in vitro and in vivo. Conversely, RhoH overexpression in bone marrow progenitors reduces eosinophil development in mixed bone marrow chimeras. These results highlight a novel negative regulatory role for RhoH in eosinophil differentiation, most likely in consequence of altered GATA-2 levels.

Catapult-like release of mitochondrial DNA by eosinophils contributes to antibacterial defense.

Although eosinophils are considered useful in defense mechanisms against parasites, their exact function in innate immunity remains unclear. The aim of this study is to better understand the role of eosinophils within the gastrointestinal immune system. We show here that lipopolysaccharide from Gram-negative bacteria activates interleukin-5 (IL-5)- or interferon-gamma-primed eosinophils to release mitochondrial DNA in a reactive oxygen species-dependent manner, but independent of eosinophil death. Notably, the process of DNA release occurs rapidly in a catapult-like manner--in less than one second. In the extracellular space, the mitochondrial DNA and the granule proteins form extracellular structures able to bind and kill bacteria both in vitro and under inflammatory conditions in vivo. Moreover, after cecal ligation and puncture, Il5-transgenic but not wild-type mice show intestinal eosinophil infiltration and extracellular DNA deposition in association with protection against microbial sepsis. These data suggest a previously undescribed mechanism of eosinophil-mediated innate immune responses that might be crucial for maintaining the intestinal barrier function after inflammation-associated epithelial cell damage, preventing the host from uncontrolled invasion of bacteria.