Steady state erythropoiesis produces new erythrocytes at a constant rate to replace senescent erythrocytes removed in the spleen and liver. Inflammation caused by infection or tissue damage skews bone marrow hematopoiesis, increasing myelopoiesis at the expense of steady state erythropoiesis. To compensate for the loss of production, stress erythropoiesis is induced. Stress erythropoiesis is highly conserved between mouse and human but utilizes a different strategy than steady state erythropoiesis. Inflammatory signals promote the proliferation of immature stress erythroid progenitors (SEPS), which in response to erythropoietin and other signals transition to stress erythroid progenitors committed to differentiation. Here we show that TNFα dependent signaling increases ROS in SEPs during the proliferation stage, however, blocking ROS production impairs their later differentiation. In addition to TNFα, nitric oxide dependent signaling drives the proliferation of stress erythroid progenitors and production of nitric oxide must be decreased so that the progenitor cells can differentiate. As progenitor cells transition to differentiation, increased production of the anti-inflammatory metabolite itaconate activates Nfe2l2 or Nrf2, which inhibits Nos2 expression, leading to decreased nitric oxide production. Mutation of Irg1, the enzyme that catalyzes the production of itaconate, causes a delayed recovery from inflammatory anemia induced by heat killed Brucella abortus . Loss of itaconate-dependent activation of Nrf2 is rescued in vivo by IL-10, which leads to activation of Nrf2 and differentiation. These data show that the differentiation of stress erythroid progenitors relies on a switch to an anti-inflammatory metabolism and increased expression of pro-resolving cytokines. Key points 1. The transition to differentiation of stress erythroid progenitors requires anti-inflammatory signals. 2. The anti-inflammatory metabolite itaconate and IL-10 increase Nrf2 activity to promote the differentiation of stress erythroid progenitors.

Inflammation skews bone marrow hematopoiesis increasing the production of myeloid effector cells at the expense of steady-state erythropoiesis. A compensatory stress erythropoiesis response is induced to maintain homeostasis until inflammation is resolved. In contrast to steady-state erythroid progenitors, stress erythroid progenitors (SEPs) utilize signals induced by inflammatory stimuli. However, the mechanistic basis for this is not clear. Here we reveal a nitric oxide (NO)-dependent regulatory network underlying two stages of stress erythropoiesis, namely proliferation, and the transition to differentiation. In the proliferative stage, immature SEPs and cells in the niche increased expression of inducible nitric oxide synthase ( Nos2 or iNOS ) to generate NO. Increased NO rewires SEP metabolism to increase anabolic pathways, which drive the biosynthesis of nucleotides, amino acids and other intermediates needed for cell division. This NO-dependent metabolism promotes cell proliferation while also inhibiting erythroid differentiation leading to the amplification of a large population of non-committed progenitors. The transition of these progenitors to differentiation is mediated by the activation of nuclear factor erythroid 2-related factor 2 (Nfe2l2 or Nrf2). Nrf2 acts as an anti-inflammatory regulator that decreases NO production, which removes the NO-dependent erythroid inhibition and allows for differentiation. These data provide a paradigm for how alterations in metabolism allow inflammatory signals to amplify immature progenitors prior to differentiation. Key points Nitric-oxide (NO) dependent signaling favors an anabolic metabolism that promotes proliferation and inhibits differentiation. Activation of Nfe2l2 (Nrf2) decreases NO production allowing erythroid differentiation.

Infection and tissue damage induce inflammation, which increases myelopoiesis at the expense of steady state erythropoiesis. Stress erythropoiesis is induced to compensate for the loss of erythroid output until the inflammation is resolved and bone marrow erythropoiesis can resume. Steady state erythropoiesis constantly produces erythrocytes, while stress erythropoiesis generates a bolus of new erythrocytes through the rapid expansion of immature progenitor cells which is followed by the synchronous differentiation of progenitors. We hypothesized that the proliferation of early progenitor cells and their transition to differentiation is regulated by changes in metabolism.

Bone marrow medullary erythropoiesis is primarily homeostatic. It produces new erythrocytes at a constant rate, which is balanced by the turnover of senescent erythrocytes by macrophages in the spleen. Despite the enormous capacity of the bone marrow to produce erythrocytes, there are times when it is unable to keep pace with erythroid demand. At these times stress erythropoiesis predominates. Stress erythropoiesis generates a large bolus of new erythrocytes to maintain homeostasis until steady state erythropoiesis can resume. In this review, we outline the mechanistic differences between stress erythropoiesis and steady state erythropoiesis and show that their responses to inflammation are complementary. We propose a new hypothesis that stress erythropoiesis is induced by inflammation and plays a key role in maintaining erythroid homeostasis during inflammatory responses.

Inflammation caused by infection or tissue damage has a profound effect on hematopoiesis. Pro-inflammatory cytokines skew hematopoiesis to increase the production of myeloid effector cells, but this change comes with a cost as erythropoiesis is inhibited by these signals. In order to compensate for the decrease in steady state erythropoiesis, stress erythropoiesis is induced. This process is distinct from steady sate erythropoiesis and shares many properties with other stem cell mediated tissue regenerative responses. It is conserved between mouse and human. Unlike steady state erythropoiesis which constantly produces new erythrocytes, the strategy of stress erythropoiesis is to produce a bolus of erythrocytes that will maintain homeostasis until steady state erythropoiesis can resume. Previous work showed that immature stress erythroid progenitors rapidly proliferate but do not differentiate, which generates a transient amplifying population of progenitors. These progenitors develop in close contact with macrophages in structures known as erythroblastic islands (EBIs). Erythropoietin (Epo) dependent signaling in EBI macrophages changes the signals made by the macrophages from those promoting proliferation (Wnt and Hedgehog) to those promoting differentiation (PGJ2 and PGE2). Here we report that PGE2 dependent mobilization of intracellular calcium activates the Perk kinase (Eif2ak3) mediated integrated stress pathway in stress erythroid progenitors. This pathway acts to increase the expression of amino acid transporters and tRNA synthases resulting in the activation of the mTorC1 complex. These two pathways drive the commitment to erythroid differentiation and are required for the generation of new erythrocytes that will maintain homeostasis until steady state erythropoiesis can resume. Inflammation caused by infection or tissue damage has a profound effect on hematopoiesis. Pro-inflammatory cytokines skew hematopoiesis to increase the production of myeloid effector cells, but this change comes with a cost as erythropoiesis is inhibited by these signals. In order to compensate for the decrease in steady state erythropoiesis, stress erythropoiesis is induced. This process is distinct from steady sate erythropoiesis and shares many properties with other stem cell mediated tissue regenerative responses. It is conserved between mouse and human. Unlike steady state erythropoiesis which constantly produces new erythrocytes, the strategy of stress erythropoiesis is to produce a bolus of erythrocytes that will maintain homeostasis until steady state erythropoiesis can resume. Previous work showed that immature stress erythroid progenitors rapidly proliferate but do not differentiate, which generates a transient amplifying population of progenitors. These progenitors develop in close contact with macrophages in structures known as erythroblastic islands (EBIs). Erythropoietin (Epo) dependent signaling in EBI macrophages changes the signals made by the macrophages from those promoting proliferation (Wnt and Hedgehog) to those promoting differentiation (PGJ2 and PGE2). Here we report that PGE2 dependent mobilization of intracellular calcium activates the Perk kinase (Eif2ak3) mediated integrated stress pathway in stress erythroid progenitors. This pathway acts to increase the expression of amino acid transporters and tRNA synthases resulting in the activation of the mTorC1 complex. These two pathways drive the commitment to erythroid differentiation and are required for the generation of new erythrocytes that will maintain homeostasis until steady state erythropoiesis can resume.

Anemic stress induces stress erythropoiesis, which rapidly generates new erythrocytes to restore tissue oxygenation. Stress erythropoiesis is best understood in mice where it is extramedullary and occurs primarily in the spleen. However, both human and mouse stress erythropoiesis use signals and progenitor cells that are distinct from steady-state erythropoiesis. Immature stress erythroid progenitors (SEPs) are derived from short-term hematopoietic stem cells. Although the SEPs are capable of self-renewal, they are erythroid restricted. Inflammation and anemic stress induce the rapid proliferation of SEPs, but they do not differentiate until serum erythropoietin (Epo) levels increase. Here we show that rather than directly regulating SEPs, Epo promotes this transition from proliferation to differentiation by acting on macrophages in the splenic niche. During the proliferative stage, macrophages produce canonical Wnt ligands that promote proliferation and inhibit differentiation. Epo/Stat5-dependent signaling induces the production of bioactive lipid mediators in macrophages. Increased production of prostaglandin J2 (PGJ2) activates peroxisome proliferator-activated receptor γ (PPARγ)-dependent repression of Wnt expression, whereas increased production of prostaglandin E2 (PGE2) promotes the differentiation of SEPs.

Low serum Se is independently associated with anemia in elderly population, dialysis patients, sickle-cells patients, and hypothyroidism patients. Previous work from our laboratory showed that dietary Se deficiency in mice showed mild anemia indicating activation of stress- erythropoietic mechanisms. Unlike steady state erythropoiesis that is primarily responsible for homeostasis to produce new erythrocytes at a constant rate, stress erythropoiesis predominates when the bone marrow cannot generate sufficient erythrocytes. During such a process, short-term reconstituting hematopoietic stem cells (CD34+Kit+Sca1+Linneg) migrate to the spleen leading to the proliferation and differentiation of stress-erythroid progenitors (SEPs). These cells lead to stress burst forming unit- erythroid cells (BFU-E) followed by terminal differentiation to erythrocytes. Recent studies demonstrate deficits in selenoproteins block the expansion and development of stress BFU-E with defects in terminal differentiation. Analysis of selenoprotein expression showed that selenoprotein W (SelenoW) was highly expressed in developing SEPs. CRISPR-Cas9 knockout of SelenoW blocked the proliferation of immature SEPs in murine and human stress erythropoiesis cultures demonstrating a central role for SelenoW in stress erythropoiesis. Using the two-culture system to generate SEPs, selenoprotein N (SelenoN) expression increased as the progenitors transition from self-renewing progenitors to form committed erythroid progenitors. SelenoN-/- mice showed significantly slower erythroid recovery following phenylhydrazine (PHZ)-induced acute hemolytic anemia. As in the muscle satellite cells where SelenoN regulates cellular Ca2+ signaling, SelenoN may also regulate Ca2+ signaling in SEPs to modulate pathways of differentiation. In summary, these data suggest that multiple selenoproteins, including SelenoN and SelenoW, coordinately regulate stress erythropoiesis.

Background A novel avian-origin influenza A (H7N9) virus emerged and spread among humans in Eastern China in 2013. Prophylactic treatment with antibiotics and probiotics for secondary infection is as important as antiviral treatment. This study aims to assess the ability of probiotic treatment to restore internal homeostasis under antibiotic pressure and to reduce/ameliorate the risk of secondary infections resulting from infection with the H7N9 virus. Methods This is a retrospective study in archival samples. Between April 1 and May 10, 2013, 113 stool, sputum, and blood specimens were collected and analyzed by denaturing gradient gel electrophoresis (DGGE) to determine the composition of the patient microbiomes. Microbial diversity was calculated using Gel-Pro analyzer and Past software. Cluster analysis of DGGE pattern profiles was employed to create a phylogenetic tree for each patient, and multidimensional scaling (MDS) and principal component analysis (PCA) were performed to visualize relationships between individual lanes. Results Five patients had secondary infections, including Klebsiella pneumonia, Acinetobacter baumanii and Candida albicans infection. The DGGE profiles of fecal samples obtained at different time points from the same individual were clearly different, particularly for patients with secondary infections. Shannon's diversity index and evenness index were lower in all infected groups compared to the control group. After B. subtilis and E. faecium or C. butyricum administration, the fecal bacterial profiles of patients who had not been treated with antibiotics displayed a trend of increasing diversity and evenness. C. butyricum failed to reduce/ameliorate secondary infection in H7N9-infected patients, but administration of B. subtilis and E. faecium appeared to reduce/ameliorate secondary infection in one patient. Conclusion H7N9 infection might decrease intestinal microbial diversity and species richness in humans. C. butyricum failed to reduce/ameliorate secondary infection in H7N9-infected patients. B. subtilis and E. faecium may also play a role in reducing/ameliorating secondary infection in these patients.

Tissue hypoxia induces a systemic response designed to increase oxygen delivery to tissues. One component of this response is increased erythropoiesis. Steady-state erythropoiesis is primarily homeostatic, producing new erythrocytes to replace old erythrocytes removed from circulation by the spleen. In response to anemia, the situation is different. New erythrocytes must be rapidly made to increase hemoglobin levels. At these times, stress erythropoiesis predominates. Stress erythropoiesis is best characterized in the mouse, where it is extramedullary and utilizes progenitors and signals that are distinct from steady-state erythropoiesis. In this report, we use an in vitro culture system that recapitulates the in vivo development of stress erythroid progenitors. We identify cell-surface markers that delineate a series of stress erythroid progenitors with increasing maturity. In addition, we use this in vitro culture system to expand human stress erythroid progenitor cells that express analogous cell-surface markers. Consistent with previous suggestions that human stress erythropoiesis is similar to fetal erythropoiesis, we demonstrate that human stress erythroid progenitors express fetal hemoglobin upon differentiation. These data demonstrate that similar to murine bone marrow, human bone marrow contains cells that can generate BMP4-dependent stress erythroid burst-forming units when cultured under stress erythropoiesis conditions.

Liver cirrhosis occurs as a consequence of many chronic liver diseases that are prevalent worldwide. Here we characterize the gut microbiome in liver cirrhosis by comparing 98 patients and 83 healthy control individuals. We build a reference gene set for the cohort containing 2.69 million genes, 36.1% of which are novel. Quantitative metagenomics reveals 75,245 genes that differ in abundance between the patients and healthy individuals (false discovery rate < 0.0001) and can be grouped into 66 clusters representing cognate bacterial species; 28 are enriched in patients and 38 in control individuals. Most (54%) of the patient-enriched, taxonomically assigned species are of buccal origin, suggesting an invasion of the gut from the mouth in liver cirrhosis. Biomarkers specific to liver cirrhosis at gene and function levels are revealed by a comparison with those for type 2 diabetes and inflammatory bowel disease. On the basis of only 15 biomarkers, a highly accurate patient discrimination index is created and validated on an independent cohort. Thus microbiota-targeted biomarkers may be a powerful tool for diagnosis of different diseases.

A novel influenza A (H7N9) virus of avian origin emerged in eastern China in the spring of 2013. This virus causes severe disease in humans, including acute and often lethal respiratory failure. As of January 2014, 275 cases of H7N9-infected patients had been reported, highlighting the urgency of identifying biomarkers for predicting disease severity and fatal outcomes. Here, we show that plasma levels of angiotensin II, a major regulatory peptide of the renin-angiotensin system, are markedly elevated in H7N9 patients and are associated with disease progression. Moreover, the sustained high levels of angiotensin II in these patients are strongly correlated with mortality. The predictive value of angiotensin II is higher than that of C-reactive protein and some clinical parameters such as the PaO2/FiO(2) ratio (partial pressure of arterial oxygen to the fraction of inspired oxygen). Our findings indicate that angiotensin II is a biomarker for lethality in flu infections.