Anemia in early congenital syphilis

The determination of blood parameters allowed us to establish that anemia develops in a number of infectious diseases, which is called "anemia of inflammation" or anemia of chronic diseases. The article presents a rare clinical case of the development of anemia of the inflammatory response, accompanied by a marked decrease in the content of hemoglobin (less than 110 g/l), hematocrit (less than 30%) and red blood cells (less than 3,0 х 1012/l), in a child with early congenital syphilis with symptoms of: specific rhinitis, papular infiltration, pemphigus, hepatosplenomegaly and damage to the nervous system. As a result of specific treatment with water-soluble benzylpenicillin salt at the rate of 100 U/kg of body weight for 28 days, and transfusion of erythrocyte mass of 15 ml/kg of child weight, there was a parallel regression of clinical manifestations of syphilis with positive dynamics of serological reactions and restoration of the number of red blood cells and the concentration of hemoglobin in the blood.

Lack of IL6 Improves Recovery from Anemia of Inflammation Which Gets Hampered in Presence of Excess Iron

Anemia of inflammation (AI) is the second most common anemia after iron deficiency anemia. The predominant regulators of AI are the cytokine interleukin 6 (IL6) and the hormone hepcidin (HAMP). IL6 is an inflammatory cytokine which also limits iron absorption by inducing HAMP, which promotes the degradation of the iron exporter ferroportin. We hypothesized that knocking down both HAMP and IL6 simultaneously will help us to understand if their mode of action in AI is uniquely limited to iron absorption and erythroid iron intake or if they also have independent roles. Henceforth, we generated IL6/HampKO (DKO) mice and, unexpectedly, observed that IL6KO mice showed the best recovery in bone marrow (BM) erythropoiesis (using flow cytometry analysis and looking at the absolute number of erythroid progenitors) after BA administration when compared to wild type (WT), HampKO and DKO mice. The best differences were observed at 14 days post BA administration. In contrast, the extramedullary erythropoiesis in the spleen was more pronounced in HampKO and DKO mice compared to WT and IL6KO animals, indicating that the mechanism impairing erythropoiesis in the BM did not affect erythroid progenitors in the spleen. These observations suggest that HAMP and IL6 proteins contribute independently to AI, with IL6 having some effect on the erythropoiesis in the BM independent from the IL6-HAMP axis leading to iron restriction. Furthermore, these observations raised the question why both HampKO and DKO mice showed reduced BM erythropoiesis compared to IL6KO animals. We investigated inflammatory cytokines and altered iron parameters as potential mediators of impaired erythropoiesis. We compared several inflammatory cytokines, including IL6, TNFa and INFg following BA administration: cytokine levels were elevated 6 hrs, reduced 48hrs after BA administration and moderately increased again two weeks later. Interestingly, among all the cytokines the levels of IL1b were significantly attenuated in IL6KO mice at day 14 compared to WT and HampKO animals. Moreover, transferrin saturation and NTBI levels were higher in HampKO and DKO animals compared to IL6KO mice. These observations strongly suggested that BM erythropoiesis is more sensitive to inflammatory insult in presence of an excess of iron, while extramedullary erythropoiesis is mildly affected and can eventually thrive under supra-physiological transferrin saturation levels. To test if increased iron affects BM erythropoiesis in presence of inflammation, we treated both WT and IL6KO mice with combination of iron dextran and BA. Both WT and IL6KO mice were treated with a combination of BA and iron at day 0 followed by alternate day of iron injections showed the poorest erythropoiesis in the BM and became rapidly sick, although the effect was significantly more pronounced in WT animals, as suggested by their survival curve. Since mycobacterium infections lead to NLPR3 inflammasome activation and Caspase1 upregulation (Marim et al. Semin Immunopathology 2017), we investigated how erythroid progenitors were affected. By flow cytometry analyses, we observed a significantly higher upregulation of the Caspase1 protein in WT and DKO mice compared to IL6KO animals. This was also reproduced by culturing WT or IL6KO BM progenitor erythroid cells in presence of mouse serum derived from WT or IL6KO mice treated with BA. Most importantly, IL6KO mice treated with BA and iron showed the highest levels of Caspase1 compared to only BA treated IL6KO mice, indicating that excess of iron abrogates the beneficial effect of IL6 deficiency on erythropoiesis under conditions of AI. Furthermore, using flow cytometry, we observed in WT mice treated with BA or BA and iron a significant increase in mitochondrial mass, which is an indicator of mitochondrial stress. The mitochondrial mass was reduced in IL6KO mice treated with BA, but again increased in IL6KO mice treated with BA and iron. We have also observed an increase of mitochondrial superoxide by confocal microscopy in WT mice compared to IL6KO mice treated with BA. Altogether, these data support a model where inflammation in presence of an excess of iron impairs BM erythropoiesis through mechanisms at least in part mediated by Caspase1 and mitochondrial dysfunction, while iron excess itself is sufficient to boost extramedullary erythropoiesis to compensate and sustain RBC production. Disclosures Vinchi: PharmaNutra: Research Funding; Vifor Pharma: Research Funding; Silence Therapeutics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Research Funding. Rivella: Ionis Pharmaceuticals: Consultancy; Meira GTx: Consultancy.

A Monoclonal Antibody Targeting ALK2 As a Potential Therapeutic Agent for Anemia of Inflammation

Background: Iron homeostasis is primarily regulated by hepcidin, a hormone predominantly expressed in the liver. Hepcidin activates the degradation of the transmembrane iron exporter ferroportin, thereby downregulating the release of iron from cells. Hepcidin expression is, at least partly, regulated in response to signaling of the type I TGF-β receptor ALK2, via SMAD2/3 phosphorylation. IL-6, which is commonly elevated in chronic kidney disease (CKD) and other inflammatory conditions, upregulates hepcidin expression and reduces serum iron bioavailability. As a result, chronic inflammatory conditions are often accompanied by secondary anemia of inflammation (AI). We have previously demonstrated that ALK2 inhibition suppressed hepcidin expression in rodents, monkeys and healthy humans. We further described that administration of a selective small molecule ALK2 kinase inhibitor (KTI-2338) reversed changes in hepcidin and iron in a mouse model of CKD, supporting the potential benefit of ALK2 inhibition in AI. Another approach to targeting ALK2 signaling is use of a neutralizing antibody. KTI-018 is a neutralizing ALK2 antibody with high affinity and selectivity for ALK2. This biologic has been demonstrated to reduce serum hepcidin and increase serum iron in healthy non-human primates. Aims: To further elucidate the specific contribution of ALK2 signaling as a driver in AI, and to determine the therapeutic potential of the antibody in this type of anemia, we assessed the effect of KTI-018 in the CKD mouse model. Methods: The study was conducted with 6-week-old male C57Bl/6 mice. Mice in the CKD cohort (CKD) were treated with once daily oral administration of adenine, a compound that metabolizes to 2,8-dihydroxyadenine, forming crystals in the proximal tubular epithelia and causing inflammation and fibrosis in the kidneys. Mice in the control cohort (healthy) received once daily oral administration of vehicle. Upon confirmation of disease, the CKD cohort was subdivided into two groups. The treatment group received twice weekly intraperitoneal treatment with KTI-018 (CKD-KTI-018), and the control group received tris-buffered saline (CKD-TBS). Healthy mice received TBS only. All mice were maintained on their assigned daily adenine or vehicle regimen. At day 53, the study was terminated and hematologic parameters, serum hepcidin, iron, and IL-6 levels were assessed. Results: After 42 days of adenine or vehicle administration, serum hepcidin, serum iron, and hematologic parameters were assessed in representative cohorts of CKD-TBS and healthy mice. The CKD-TBS cohort experienced changes associated with anemia of inflammation as compared to the healthy mice, including increased hepcidin, decreased serum iron, and decreased hematologic parameters. The differences between the healthy and CKD-TBS groups were maintained through the duration of the study. At study termination, CKD-TBS mice had increased serum IL-6 levels (218%), elevated serum hepcidin (149%), and reduced serum iron (-30%) as compared to the healthy mice. Laboratory findings characteristic of anemia were present in the CKD-TBS group, including decreased red blood cells (-6.1%), hemoglobin (-13.2%), and reticulocyte hemoglobin content (-9.3%) as compared to healthy mice. In contrast, CKD-KTI-018 mice had decreased serum hepcidin (-25%) and increased serum iron (59%) as compared to CKD-TBS mice. This restoration of serum iron corresponded to improvements in red blood cells, hemoglobin, and reticulocyte hemoglobin content, which were increased by 7.6%, 9.6%, and 6.7%, respectively, in the CKD-KTI-018 mice as compared to the CKD-TBS mice. These results demonstrate that, by decreasing serum hepcidin, KTI-018 increased the bioavailability of iron, which led to the restoration of hematologic parameters and appeared to reverse AI in mice. Discussion: In this study, a neutralizing ALK2 antibody decreased serum hepcidin, increased serum iron and consequently reversed AI in a mouse model of CKD. These results support the role of ALK2 signaling in AI and suggest that inhibition of ALK2 may be a potential treatment approach for anemia resulting from CKD and other chronic inflammatory diseases. Future studies will explore if ALK2 inhibition may prevent or treat progression of CKD itself, and the role that ALK2 inhibition may play in other chronic inflammatory conditions associated with elevated hepcidin. Disclosures Medeiros: Keros Therapeutics: Current Employment, Current equity holder in publicly-traded company. Backus: Keros Therapeutics: Current Employment, Current equity holder in publicly-traded company. Materna: Keros Therapeutics: Current Employment, Current equity holder in publicly-traded company. Fisher: Keros Therapeutics: Current Employment, Current equity holder in publicly-traded company. Lachey: Keros Therapeutics: Current Employment, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees. Seehra: Keros Therapeutics: Current Employment, Current equity holder in publicly-traded company.

Physiology and Inflammation Driven Pathophysiology of Iron Homeostasis—Mechanistic Insights into Anemia of Inflammation and Its Treatment

Anemia is very common in patients with inflammatory disorders. Its prevalence is associated with severity of the underlying disease, and it negatively affects quality of life and cardio-vascular performance of patients. Anemia of inflammation (AI) is caused by disturbances of iron metabolism resulting in iron retention within macrophages, a reduced erythrocyte half-life, and cytokine mediated inhibition of erythropoietin function and erythroid progenitor cell differentiation. AI is mostly mild to moderate, normochromic and normocytic, and characterized by low circulating iron, but normal and increased levels of the storage protein ferritin and the iron hormone hepcidin. The primary therapeutic approach for AI is treatment of the underlying inflammatory disease which mostly results in normalization of hemoglobin levels over time unless other pathologies such as vitamin deficiencies, true iron deficiency on the basis of bleeding episodes, or renal insufficiency are present. If the underlying disease and/or anemia are not resolved, iron supplementation therapy and/or treatment with erythropoietin stimulating agents may be considered whereas blood transfusions are an emergency treatment for life-threatening anemia. New treatments with hepcidin-modifying strategies and stabilizers of hypoxia inducible factors emerge but their therapeutic efficacy for treatment of AI in ill patients needs to be evaluated in clinical trials.

Causes of Anemia in Polish Older Population—Results from the PolSenior Study

Vitamin B12, folate, iron deficiency (IDA), chronic kidney disease (CKD), and anemia of inflammation (AI) are among the main causes of anemia in the elderly. WHO criteria of nutritional deficiencies neglect aging-related changes in absorption, metabolism, and utilization of nutrients. Age-specific criteria for the diagnosis of functional nutritional deficiency related to anemia are necessary. We examined the nationally representative sample of Polish seniors. Complete blood count, serum iron, ferritin, vitamin B12, folate, and renal parameters were assessed in 3452 (1632 women, 1820 men) participants aged above 64. Cut-off points for nutritional deficiencies were determined based on the WHO criteria (method-A), lower 2.5 percentile of the studied population (method-B), and receiver operating characteristic (ROC) analysis (method-C). Method-A leads to an overestimation of the prevalence of vitamin B12 and folate deficiency, while method-B to their underestimation with over 50% of unexplained anemia. Based on method-C, anemia was classified as nutritional in 55.9%. In 22.3% of cases, reasons for anemia remained unexplained, the other 21.8% were related to CKD or AI. Mild cases were less common in IDA, and more common in non-deficiency anemia. Serum folate had an insignificant impact on anemia. It is necessary to adopt the age-specific criteria for nutrient deficiency in an old population.

Anemia of inflammation in patients with colorectal cancer: Correlation with Interleukin-1, Interleukin-33 and Galectin-1

Purpose: Patients with colorectal cancer (CRC) have anemia often present as a consequence of chronic bleeding from tumor. The exact role of lL-33, Galectin-l and IL-l in the pathological genesis of anemia in colorectal cancer patients has not been elucidated yet. The main goal of this research was to analyze Gal-l, IL-l and lL-33 systemic values in anemic and non-anemic CRC patients. Methods: Concentrations of IL-33, Galectin-1 and IL-1 have been studied in blood samples of 55 CRC patients (27 without anemia and 28 with anemia). Results: CRC patients with anemia had more severe and local advanced disease compared to CRC non-anemic patients. Anemia positively correlated with higher nuclear grade, lymph and blood vessel invasion, as well as with higher TNM stage, detectable metastatic lesions in lung and liver and peritoneal carcinomatosis. Significantly higher IL-33, Gal-1 and IL-1 concentration have been found in sera of patients with CRC and detected anemia. CRC patients mostly had microcytic anemia, while ferritin values were in normal range. Analysis revealed positive mutual correlation between serum values of galectin-1, IL-1 and IL-33 in CRC patients. Level of hemoglobin negatively correlated with serum IL-33, Gal-1 and IL-1. We have analyzed the Receiver Operating Characteristic (ROC) curves of serum IL-33, Gal-1 and IL-1 showed that these cytokines can be treated as additional markers for anemia of inflammation in CRC patients. Conclusions: Predomination of Galectin-1, IL-1 and IL-33 in anemic CRC patients implicates on their potential role in anemia genesis and further development.

Vitamin D supplementation on anemia markers and the impact on PTH in patients with chronic kidney disease

Chronic Kidney Disease (CKD) is an injury that causes progressive impairment of exocrine and endocrine renal functions. A very common complication is anemia, caused by reduced erythropoietin production, iron deficiency and inflammation. Evidence demonstrates that vitamin D has effects on anemia of inflammation, through the increase in erythrocytes and decrease in pro-inflammatory cytokines. This study aims to review the effects of vitamin D supplementation on 25(OH)D2 concentrations, on anemia markers and on PTH levels. This is an integrative review carried out through the search and selection of original publications, in english and portuguese, indexed in PubMed, Web of Science and Science Direct databases belonging to the 2010-2020 range. The results pointed to 25(OH)D2 concentrations compatible with normality after vitamin D supplementation. In five studies, there was no change in hemoglobin and PTH levels, and in four studies there was a reduction in the dose of EPO or erythroid stimulating agent, attributing such effect on the role of calcitriol as a substrate for bone marrow erythropoietic cells. The study concluded that vitamin D supplementation had beneficial effects for correction of 25(OH)D2 deficiency, however, it reinforces the controversy about the behavior of vitamin D on the improvement of anemia markers and PTH levels in patients with DRC. Therefore, it is suggested that the beneficial effect of vitamin D on anemia in renal patients may be independent of PTH suppression.

UBA6 and NDFIP1 regulate the degradation of ferroportin

Hepcidin regulates iron homeostasis by controlling the level of ferroportin, the only membrane channel that facilitates export of iron from within cells. Binding of hepcidin to ferroportin induces the ubiquitination of ferroportin at multiple lysine residues and subsequently causes the internalization and degradation of the ligand-channel complex within lysosomes. The objective of this study was to identify components of the ubiquitin system that are involved in ferroportin degradation. A HepG2 cell line, which inducibly expresses ferroportin-GFP (FPN-GFP), was established to test the ability of siRNAs directed against components of the ubiquitin system to prevent BMP6- and exogenous hepcidin-induced ferroportin degradation. Of the 88 siRNAs directed against components of the ubiquitin pathway that were tested, siRNAmediated depletion of the alternative E1 enzyme UBA6 as well as the adaptor protein NDFIP1 prevented BMP6- and hepcidin- induced degradation of ferroportin in vitro. A third component of the ubiquitin pathway, ARIH1, indirectly inhibited ferroportin degradation by impairing BMP6 mediated induction of hepcidin. In mice, the AAVmediated silencing of Ndfip1 in the murine liver increased the level of hepatic ferroportin and increased circulating iron. The results suggest that the E1 enzyme UBA6 and the adaptor protein NDFIP1 are involved in iron homeostasis by regulating the degradation of ferroportin. These specific components of the ubiquitin system may be promising targets for the treatment of iron related diseases, including iron overload and anemia of inflammation.