Hematopoietic Response Research Articles

Nicotinic alpha 7 receptor expression and modulation of the lung epithelial response to lipopolysaccharide.

Nicotine modulates multiple inflammatory responses in the lung through the nicotinic acetylcholine receptor subtype alpha7 (α7). Previously we reported that α7 modulates both the hematopoietic and epithelium responses in the lung to the bacterial inflammogen, lipopolysaccharide (LPS). Here we apply immunohistochemistry, flow cytometry and RNA-Seq analysis of isolated distal lung epithelium to further define α7-expression and function in this tissue. Mouse lines were used that co-express a bicistronic tau-green fluorescent protein (tGFP) as a reporter of α7 (α7G) expression and that harbor an α7 with a specific point mutation (α7E260A:G) that selectively uncouples it from cell calcium-signaling mechanisms. The tGFP reporter reveals strong cell-specific α7-expression by alveolar macrophages (AM), Club cells and ATII cells. Ciliated cells do not express detectible tGFP, but their numbers decrease by one-third in the α7E260A:G lung compared to controls. Transcriptional comparisons (RNA-Seq) between α7G and α7E260A:G enriched lung epithelium 24 hours after challenge with either intra-nasal (i.n.) saline or LPS reveals a robust α7-genotype impact on both the stasis and inflammatory response of this tissue. Overall the α7E260A:G lung epithelium exhibits reduced inflammatory cytokine/chemokine expression to i.n. LPS. Transcripts specific to Club cells (e.g., CC10, secretoglobins and Muc5b) or to ATII cells (e.g., surfactant proteins) were constitutively decreased in in the α7E260A:G lung, but they were strongly induced in response to i.n. LPS. Protein analysis applying immunohistochemistry and ELISA also revealed α7-associated differences suggested by RNA-Seq including altered mucin protein 5b (Muc5b) accumulation in the α7E260A:G bronchia, that in some cases appeared to form airway plugs, and a substantial increase in extracellular matrix deposits around α7E260A:G airway bronchia linings that was not seen in controls. Our results show that α7 is an important modulator of normal gene expression stasis and the response to an inhaled inflammogen in the distal lung epithelium. Further, when normal α7 signaling is disrupted, changes in lung gene expression resemble those associated with long-term lung pathologies seen in humans who use inhaled nicotine products.

Persistent injury-associated anemia: the role of the bone marrow microenvironment

BackgroundThe regulation of erythropoiesis involves hematopoietic progenitor cells, bone marrow stroma, and the microenvironment. Following severe injury, a hypercatecholamine state develops that is associated with increased mobilization of hematopoietic progenitor cells to peripheral blood and decreased growth of bone marrow erythroid progenitor cells that manifests clinically as a persistent injury-associated anemia. Changes within the bone marrow microenvironment influence the development of erythroid progenitor cells. Therefore, we sought to determine the effects of lung contusion, hemorrhagic shock, and chronic stress on the hematopoietic cytokine response. Materials and methodsBone marrow was obtained from male Sprague–Dawley rats (n = 6/group) killed 7 d after lung contusion followed by hemorrhagic shock (LCHS) or LCHS followed by daily chronic restraint stress (LCHS/CS). End point polymerase chain reaction was performed for interleukin-1β, interleukin-10, stem cell factor, transforming growth factor-β, high-mobility group box-1 (HMGB-1), and B-cell lymphoma-extra large. ResultsSeven days following LCHS and LCHS/CS, bone marrow expression of prohematopoietic cytokines (interleukin-1β, interleukin-10, stem cell factor, and transforming growth factor-β) was significantly decreased, and bone marrow expression of HMGB-1 was significantly increased. B-cell lymphoma-extra large bone marrow expression was not affected by LCHS or LCHS/CS (naïve: 44 ± 12, LCHS: 44 ± 12, LCHS/CS: 37 ± 1, all P > 0.05). ConclusionsThe bone marrow microenvironment was significantly altered following severe trauma in a rodent model. Prohematopoietic cytokines were downregulated, and the proinflammatory cytokine HMGB-1 had increased bone marrow expression. Modulation of the bone marrow microenvironment may represent a therapeutic strategy following severe trauma to alleviate persistent injury-associated anemia.

The Role of Serotonin beyond the Central Nervous System during Embryogenesis.

Serotonin, or 5-hydroxytryptamine (5-HT), is a well-known neurotransmitter that plays vital roles in neural activities and social behaviors. Clinically, deficiency of serotonin is linked with many psychiatric disorders. Interestingly, a large proportion of serotonin is also produced outside the central nervous system (CNS). There is increasing evidence demonstrating important roles of serotonin in the peripheral tissues. Here, we will describe the multiple biological functions of serotonin in hematopoietic system, such as development of hematopoietic stem and progenitor cells (HSPCs), differentiation of hematopoietic cells, maintenance of vascular system, and relationship with hematological diseases. The roles of serotonin in inflammatory responses mediated by hematopoietic cells as well as in liver regeneration are also discussed. Our recent understandings of the impact of serotonin on hematopoietic system, immune responses, and tissue regeneration support utilization of serotonin as a potential therapeutic target for the treatment of hematological diseases and organ repair in clinic.

Single-dose intravenous iron infusion or oral iron for treatment of fatigue after postpartum haemorrhage: arandomized controlled trial.

To evaluate the clinical efficacy of a single-dose intravenous infusion of iron isomaltoside compared with current treatment practice with oral iron measured by physical fatigue in women after postpartum haemorrhage. Single-centre, open-label, randomized controlled trial. Participants received intravenous iron (n=97) or oral iron (n=99), and completed the Multidimensional Fatigue Inventory and Edinburgh Postnatal Depression Scale, and haematological and iron parameters were measured. Primary outcome was the aggregated change in physical fatigue score from baseline to 12weeks postpartum. The difference in physical fatigue score was -0·97 (95% CI: -1·65; -0·28, P=0·006) in favour of intravenous iron, but did not meet the predefined difference of 1·8. Across visits, we found statistically significant differences in fatigue and depression scores, as well as in haematological and iron parameters, all in favour of intravenous iron. There were no serious adverse reactions. A single dose of intravenous iron was associated with a statistically significant reduction in aggregated physical fatigue within 12weeks after postpartum haemorrhage compared to standard medical care with oral iron below the prespecified criteria of clinical superiority. As patient-reported outcomes improved significantly and intravenous iron resulted in a fast hematopoietic response without serious adverse reactions, intravenous iron may be a useful alternative after postpartum haemorrhage if oral iron is not absorbed or tolerated.

The Role of the Bone Marrow Stromal Compartment in the Hematopoietic Response to Microbial Infections.

Continuous production of blood cells unfolds within a complex three-dimensional tissue scaffold established by highly organized stromal cell networks of mesenchymal, neural, and vascular origin inside bone marrow (BM) cavities. Collectively, stromal cells have been shown to serve two principal roles; first as primary participants of bone remodeling and metabolism and second as master regulators of different stages of blood cell development and production. Indeed, ample evidence demonstrates that stromal cells can sense and integrate systemic signals to shape hematopoietic responses and that these regulatory mechanisms are subverted in multiple pathologic conditions. Microbial infections are stressors that elicit potent inflammatory reactions and induce substantial alterations of hematopoietic output. Whether the cellular components of the BM stromal microenvironment are targeted by infections and participate in infection-induced hematopoiesis has not been investigated in sufficient detail to date. In this manuscript, we provide a succinct updated overview of the different cell populations that are currently known to form BM stroma. We discuss experimental evidence demonstrating that different stromal components are actively damaged or functionally altered by pathogens and/or ensuing inflammatory signals and review how these effects are known to contribute to the hematologic manifestations observed during infections.

Divergent Functions of TLR2 on Hematopoietic and Nonhematopoietic Cells during Chronic Mycobacterium tuberculosis Infection.

We have reported that TLR2 is crucial for host resistance against chronic Mycobacterium tuberculosis infection; however, which cell types are key players in this response remain unknown. This led us to decipher the relative contribution of TLR2 on nonhematopoietic and hematopoietic cells in resistance against chronic M. tuberculosis infection in mice infected with M. tuberculosis Erdman. Consistent with our previous report, at 8 wk of infection, TLR2 knockout (TLR2KO)→TLR2KO bone marrow chimeric mice exhibited increased bacterial burden, disorganized accumulation of lymphocytes and mononuclear cells, and extensive pulmonary immunopathology compared with wild-type (WT)→WT chimeric mice. Bacterial burden and pulmonary immunopathology of chimeric mice lacking TLR2 in the hematopoietic compartment (TLR2KO→WT) was comparable to TLR2KO mice. In contrast, chimeric mice deficient in TLR2 in the nonhematopoietic compartment (WT→TLR2KO) exhibited a marked attenuation in granulomatous inflammation compared with WT mice. Although the latter mice did not exhibit improved pulmonary bacterial control, significant reductions in bacterial burden in the draining lymph nodes, spleen, and liver were observed. These findings establish that the TLR2-mediated hematopoietic response promotes stable control of pulmonary bacterial burden and granuloma integrity, whereas TLR2 signaling on nonhematopoietic cells may partly facilitate granulomatous inflammation and bacterial dissemination.

Post-transplant intravenous laronidase augmentation for Hurler syndrome: anti-drug antibody, biochemical and donor hematopoietic graft response

Post-transplant intravenous laronidase augmentation for Hurler syndrome: anti-drug antibody, biochemical and donor hematopoietic graft response

CXCL12-Abundant Reticular (CAR) Cells Are Key Regulators for a Sustained Hematopoietic Response during Chronic Inflammation

The blood system is established and maintained by hematopoietic stem and progenitor cells (HSPCs) that are located in the bone marrow (BM) where they are surrounded by different types of hematopoietic and non-hematopoietic cells that together form a complex supportive microenvironment. It is well established that the microenvironment delivers a plethora of extrinsic signals that help to maintain stemness of HSCs and guide steady-state HSPC differentiation. The BM microenvironment also likely is a key player during infection and inflammation-induced demand-adapted hematopoiesis (e.g. Boettcher et al. Blood 2014; Manz and Boettcher Nat Immunol 2014).We set out to study how the BM microenvironment supports the hematopoietic system in adaptation to severe systemic inflammation using lipopolysaccharide (LPS) or polyinosinic-polycytidylic acid (poly(I:C)) to mimic gram-negative bacterial or viral infection, respectively. Based on expression of CD45 and Ter119 in conjunction with Sca1, CD31, Tie2 and CD140b, we were able to flow-cytometrically define and isolate several cell populations enriched for distinct non-hematopoietic cell types in mouse BM, i.e. mesenchymal stromal cells (MSCs), CXCL12-abundant reticular cells (CAR cells), arteriolar endothelial cells (AECs) and sinusoidal endothelial cells (SECs).Gene expression analyses for a number of prototypic inflammatory cytokines and chemokines revealed that interleukin-6 (IL-6) is highly significantly and specifically up-regulated in CAR cells during LPS stimulation. Extensive experimentation involving Il6 expression analyses from unseparated total tissue extracts (bone marrow, spleen, lung, heart, liver, kidney), and IL-6 levels in plasma and BM lysates from reciprocal BM chimeric Il6-/- mice (Il6-/-→WT, WT→ Il6-/-) as well as conditional Il6-/- mice (Lepr-Cre;Il6 flox/flox) demonstrated that CAR cells act as a primary source of IL-6 during LPS stimulation.To test the putative functional consequences of IL-6 secretion from CAR cells during LPS-induced inflammation, we studied the hematopoietic response in LPS-treated WT and Il6-/- mice. While there was neither a relative nor absolute difference in immunophenotypically defined HSPCs, myeloid progenitors or mature myeloid lineages following short-term LPS treatment (2 injections over 3 days), we observed a significant decrease in absolute numbers of cells along the myeloid lineage from the earliest HSPCs to the mature myeloid cells upon chronic-repetitive LPS stimulation (9 injections over 21 days) in Il6-/- mice compared to WT mice. Moreover, the total number of colony-forming units granulocyte (CFU-G) was significantly reduced in the BM of LPS-treated Il6-/- mice compared to WT mice. We observed the identical impaired hematopoietic responses in reciprocal BM chimeric Il6-/- mice and conditional Il6-/- mice (Lepr-Cre;Il6 flox/flox) corroborating that CAR cell-derived IL-6 is crucial for mounting an appropriate hematopoietic response upon LPS stimulation.To address the question whether CAR cells may directly be able to sense Toll-like receptor (TLR) ligands (e.g. LPS is recognized by TLR4) we compared expression of Tlr1 - Tlr9 in MSCs, SECs, AECs, and CAR cells isolated from BM of WT mice to that found in pooled conventional and plasmacytoid dendritic cells (cDCs / pDCs) known to express different TLRs. Interestingly, the different BM niche cells expressed several TLRs at surprisingly high levels. To test whether IL-6 secretion following LPS stimulation is dependent on Tlr4-expressing CAR cells we generated reciprocal chimeric BM mice with a Tlr4-deficiency in either hematopoietic or non-hematopoietic cells. Our data demonstrated that Tlr4 expression on both hematopoietic and non-hematopoietic cells is equally contributing to IL-6 secretion during short-term LPS stimulation. However, following chronic-repetitive LPS stimulation, IL-6 secretion is completely dependent on Tlr4 expression in non-hematopoietic cells strongly suggesting that CAR cells sense and secrete IL-6 in that setting.In summary, our data reveal that CAR cells play an essential role for a sustained hematopoietic response during chronic LPS treatment. IL-6 released from CAR cells acts as a critical growth stimulus for the early HSPC compartment to provide a reservoir of highly proliferative myeloid progenitors in order to counterbalance inflammation-driven cell consumption. DisclosuresNo relevant conflicts of interest to declare.

Pharmacokinetics and Exposure-Response of Luspatercept in Patients with Beta-Thalassemia: Preliminary Results from Phase 2 Studies

▪Background: In β-thalassemia, imbalanced production of α and β globin chains in erythroid precursors inhibits late-stage erythroid differentiation, leading to ineffective erythropoiesis and anemia. Luspatercept is a modifiedactivin receptor type IIB-IgG Fc fusion protein that promotes terminal erythroid differentiation and mitigates ineffective erythropoiesis in murine β-thalassemia. In phase 2 studies, luspatercept led to long-term increases in hemoglobin (Hb) levels and reduction in transfusion burden in patients (pts) with β-thalassemia.Aims: To characterize the pharmacokinetics (PK) of luspatercept, and to explore the exposure-response relationship for efficacy and safety in pts with β-thalassemia, thereby informing selection of the starting dose for phase 3 studies of luspatercept in β-thalassemia.Methods: PK, safety, and efficacy data were collected from two phase 2 studies (base and extension). In the base study, luspatercept was administered once every 3 weeks by subcutaneous injection to sequential cohorts for up to 5 doses. The base study included a dose-finding phase (at fixed doses ranging from 0.2 mg/kg to 1.25 mg/kg) and an expansion cohort (at a starting dose of 0.8 mg/kg followed by individual dose titration up to 1.25 mg/kg). Pts completing the base study were eligible to enroll in an extension study, where they continued to receive luspatercept every 3 weeks for up to 24 months. Pts who had treatment interruption for ≥ 3 months before enrolling in the extension study received a starting dose of 0.8 mg/kg (followed by dose titration) and were treated as "new" pts in the exposure-response analysis. The main exposure endpoint was area under the luspatercept serum concentration-time curve (AUC). Clinical endpoints included Hb increase, transfusion reduction, and drug-related adverse events (AEs) in weeks 1-15. Responders were defined as non-transfusion-dependent (NTD) pts with a mean Hb increase ≥ 1.0 g/dL/15 weeks, and transfusion-dependent (TD) pts with a transfusion reduction ≥ 33%/15 weeks.Results: As of July 20, 2016, preliminary data were available for 89 pts, including 49 NTD pts (baselineHb 6.5-9.8 g/dL) and 40 TD pts (baseline transfusion burden 4-18 units/12 weeks). Median age was 37 years (range 20-62), and 47% were female. A total of 49 pts were eligible for individual dose titration, with ~71% having at least 1 dose escalation (to 1 mg/kg) and 27% having 2 dose escalations (up to 1.25 mg/kg) within the first 5 cycles.Luspatercept PK was adequately described by a 1-compartment PK model with linear absorption and elimination. The half-life of luspatercept in serum was~10 days across doses. Body weight positively correlated with luspatercept clearance and its volume of distribution. TD pts had~23% lower volume of distribution than NTD pts.In NTD pts, higher luspatercept serum AUC correlated with a greater increase in Hblevels (P < 0.01). Most TD pts had a reduction in transfusion burden; however, an exposure-dependent trend was not apparent for relative or absolute reduction in units transfused, possibly due to the narrow exposure range tested. The median AUC was similar for NTD responders and TD responders (~100d·μg/mL vs ~120d·μg/mL). As AUC increased, the frequency of responders increased for NTD pts, TD pts, and the 2 groups combined (Figure). Population PK simulation predicted that a starting dose of 1.0 mg/kg luspatercept would result in > 90% of NTD pts and ~50% of TD pts achieving the median AUC concentration observed in responders. By contrast, < 50% pts (NTD or TD) were predicted to achieve the target AUC at the 0.8 mg/kg dose.Grade 2-3 drug-related AEs (all types) were more frequent with higher AUC (P < 0.05). In both NTD and TD pts, bone and muscle pain were the most common AEs; however, these AES did not have a significant relationship with luspatercept serum exposure.Conclusions: Higher luspatercept serum exposure correlated with greater erythroid hematopoietic response as well as more frequent grade 2-3 related AEs. Exposure-response modeling and PK simulation support a phase 3 starting dose of 1.0 mg/kg and intra-patient dose escalation up to 1.25 mg/kg according to pts' response. A phase 3 study of luspatercept in regularly transfused adults with β-thalassemia is ongoing (BELIEVE study; ClinicalTrials.gov NCT02604433). [Display omitted] DisclosuresChen:Celgene Corporation: Employment, Equity Ownership. Laadem:Celgene Corporation: Employment, Equity Ownership. Wilson:Acceleron Pharma: Employment, Equity Ownership. Zhang:Acceleron Pharma: Employment. Sherman:Acceleron Pharma: Employment, Equity Ownership, Patents & Royalties. Ritland:Celgene Corporation: Employment, Equity Ownership. Attie:Acceleron Pharma: Employment, Equity Ownership.

Pharmacokinetics and Exposure-Response of Luspatercept in Patients with Anemia Due to Low- or Intermediate-1-Risk Myelodysplastic Syndromes (MDS): Preliminary Results from Phase 2 Studies

Background: Luspatercept is a modified ActRIIB-IgG Fc fusion protein that corrects ineffective erythropoiesis. In phase 2 studies, luspatercept treatment led to long-term increases in hemoglobin (Hb) levels and reduction in transfusion burden in patients (pts) with IPSS Low- or Intermediate-1-risk MDS.Aims: To characterize the pharmacokinetics (PK) of luspatercept and explore the exposure-response relationship for efficacy and safety in pts with MDS, thereby informing selection of the starting dose for phase 3 studies of luspatercept in MDS.Methods: PK, safety, and efficacy data were collected from two phase 2 studies (base and extension). In the base study, luspatercept was administered once every 3 weeks by subcutaneous injection to sequential cohorts for up to 5 doses. The base study included a dose-finding phase (at fixed doses from 0.125 to 1.75 mg/kg), and an expansion cohort (at a starting dose of 1.0 mg/kg followed by individual dose titration up to 1.75 mg/kg). Pts completing the base study were eligible to enroll in an extension study, where they continued to receive luspatercept every 3 weeks for up to 24 months. Pts who had treatment interruption for ≥ 3 months before enrolling in the extension study received a starting dose of 1.0 mg/kg (followed by dose titration) and were treated as "new" pts in the exposure-response analysis. The main exposure endpoint was area under the luspatercept serum concentration−time curve (AUC). Clinical endpoints included Hb increase, transfusion reduction, and drug-related adverse events (AEs) in weeks 1-15. Responders were defined as pts achieving erythroid hematologic improvement (HI-E) per IWG criteria, i.e. Hb increase ≥ 1.5 g/dL for 8 weeks in low transfusion burden (LTB) pts, and transfusion reduction ≥ 4 RBC units/8 weeks in high transfusion burden (HTB) pts.Results: As of July 20, 2016, preliminary data were available for 66 pts: 22 LTB pts (baseline Hb 6.4-10.1 g/dL) and 44 HTB pts (baseline transfusion burden 4-18 units/8 weeks). Median age was 72 years (range 27-90); 41% were female. A total of 39 pts were eligible for individual dose titration; of these, ~49% had ≥ 1 dose escalation (to 1.33 mg/kg) and ~15% had 2 dose escalations (to 1.75 mg/kg) in the first 3 months.Luspatercept PK was adequately described by a 1-compartment PK model with linear absorption and elimination. Half-life of luspatercept in serum was ~10-14 days across doses. Body weight positively correlated with luspatercept clearance and its volume of distribution. Baseline transfusion burden (LTB vs HTB) and erythropoietin (EPO) level (10-4,752 U/L) had no significant effect on luspatercept PK.In LTB pts who were transfusion-free on treatment, higher luspatercept AUC correlated with greater Hb increase (P < 0.01). In HTB pts, AUC correlated with reduced transfusion units in pts with baseline EPO ≤ 500 U/L (P< 0.01) but not in pts with baseline EPO > 500 U/L. Median AUC was 148 d·µg/mL in LTB responders and 185 d·µg/mL in HTB responders. Luspatercept AUC also correlated with frequency of IWG HI-E responders for LTB pts, HTB pts (baseline EPO ≤ 500 U/L), and the 2 groups combined. In pts requiring transfusion (≥ 2 units/8 weeks) with baseline EPO ≤ 500 U/L, baseline transfusion burden was a significant predictor of achieving transfusion independence (TI) ≥ 8 weeks, and higher luspatercept AUC was associated with greater TI rate after accounting for baseline transfusion burden. Thus, individualized dosing based on baseline transfusion burden may increase the likelihood of achieving TI in HTB pts. Population PK simulation predicted that the starting dose resulting in 90% of LTB pts and 50% of HTB pts achieving efficacious AUC for HI-E would be 1 mg/kg and 1.1 mg/kg, respectively; higher doses would result in a higher proportion of pts achieving efficacious AUC. There was no significant relationship between severity and frequency of drug-related AEs and luspatercept serum exposure.Conclusions: Higher luspatercept serum exposure correlated with greater erythroid hematopoietic response for both LTB and HTB pts. Exposure-response modeling and PK simulation support a phase 3 starting dose of 1.0 mg/kg and intra-patient dose escalation up to 1.75 mg/kg according to erythroid hematopoietic response. A phase 3 study of luspatercept in regularly transfused ring sideroblast positive patients with lower-risk MDS according to IPSS-R criteria is ongoing (MEDALIST study; ClinicalTrials.gov NCT02631070). DisclosuresChen:Celgene Corporation: Employment, Equity Ownership. Laadem:Celgene Corporation: Employment, Equity Ownership. Wilson:Acceleron Pharma: Employment, Equity Ownership. Zhang:Acceleron Pharma: Employment. Sherman:Acceleron Pharma: Employment, Equity Ownership, Patents & Royalties. Ritland:Celgene Corporation: Employment, Equity Ownership. Attie:Acceleron Pharma: Employment, Equity Ownership.

Rapid and direct megakaryocyte-lineage differentiation from HSCS hallmark hematopoietic system response to stress

Rapid and direct megakaryocyte-lineage differentiation from HSCS hallmark hematopoietic system response to stress

LEPR-expressing cells are key regulators for a sustained hematopoietic response during chronic inflammation

The blood system is established and maintained by hematopoietic stem and progenitor cells (HSPCs) that are located in the bone marrow (BM) where they are surrounded by different types of supportive cells. Accumulating evidence suggests that important extrinsic regulatory mechanisms have evolved to maintain stemness of HSCs and guide HSPC differentiation into more mature blood cell lineages. We set out to study how the BM microenvironment supports the hematopoietic system in adaptation to inflammation. Therefore wild-type (WT) mice were stimulated with LPS or polyIC to mimic gram-negative bacterial or viral infection, respectively. Based on expression of CD45 and Ter119 to exclude hematopoietic cells in conjuction with Sca1, CD31 and CD140b, we are able to flow-cytometrically define several non-hematopoietic cell types in mouse BM: so-called PαS cells which were shown to have mesenchymal stem cell characteristics, a stromal cell population enriched for CXCL12-abundant reticular cells (CARs), and endothelial cells (ECs). These different cell types were isolated and their gene expression profiles were determined and compared to steady-state WT mice. We found that Il6 is significantly and specifically up-regulated during LPS stimulation by the cell population enriched for CARs. This could be validated using qPCR and by injecting BM chimeric mice (Wt>Wt, Wt>Il6-/-, Il6-/->Wt, Il6-/->Il6-/-) with PBS and LPS. We could show that IL6 protein is almost exclusively produced by non-hematopoietic cells. We then generated cell type-specific Il6-deficient mice by crossing LepR-Cre mice with IL6fl/fl mice, and could confirm that IL6 is mainly produced by non-hematopoietic cells that are enriched for CARs. To functionally study the regulatory role of IL6 on hematopoiesis during inflammation, WT and Il6-/- mice were injected with PBS or LPS over 3 weeks every other day. We analyzed phenotypically defined hematopoietic stem and progenitor cells as well as more mature myeloid cell types, and observed a significant difference in absolute cell numbers of these cell types in LPS-treated WT mice compared to LPS-treated Il6-/- mice. We could observe the same significant difference in absolute cell numbers when we repeated the experiment with LepR-Cre; IL6fl/fl mice and their LepR-Cre negative littermates. Our preliminary data suggests that CAR cell produced IL6 is essential for a sustained myelopoietic response during chronic LPS treatment. IL6 may act as a proliferative stimulus for the early HSPC compartment during infection to provide a reservoir of highly proliferative myeloid progenitors in order to counterbalance inflammation-driven cell losses.

Hematopoietic stem cell response to inflammation.

Hematopoietic stem cells (HSCs) have unique functional properties, including self-renewal and multi-lineage differentiation potential, and are thought to be fully responsible for lifelong hematopoiesis. However, recent studies have shown that HSCs divide much more slowly than thought, and, therefore, that daily hematopoiesis is maintained not by HSCs but by hematopoietic progenitors with limited self-renewal. When hematopoietic stress such as an infection occurs, hematopoietic production is at high demand at the site of infection. To meet hematopoietic needs, HSCs are also presumably recruited to orchestrate hematopoiesis. The beneficial and detrimental effects of inflammation on HSC function and the associated hematopoietic regulation are discussed herein, by summarizing recent findings.

Digitalization of a non-irradiated acute myeloid leukemia model.

BackgroundComputer-aided, interdisciplinary researches for biomedicine have valuable prospects, as digitalization of experimental subjects provide opportunities for saving the economic costs of researches, as well as promoting the acquisition of knowledge. Acute myeloid leukemia (AML) is intensively studied over long periods of time. Till nowaday, most of the studies primarily focus on the leukemic cells rather than how normal hematopoietic cells are affected by the leukemic environment. Accordingly, the conventional animal models for AML are mostly myeloablated as leukemia can be induced with short latency and complete penetrance. Meanwhile, most previous computational models focus on modeling the leukemic cells but not the multi-tissue leukemic body resided by both leukemic and normal blood cells. Recently, a non-irradiated AML mouse model has been established; therefore, normal hematopoietic cells can be investigated during leukemia development. Experiments based on the non-irradiated animal model have monitored the kinetics of leukemic and (intact) hematopoietic cells in multiple tissues simultaneously; and thus a systematic computational model for the multi-tissue hematopoiesis under leukemia has become possible.ResultsIn the present work, we adopted the modeling methods in previous works, but aimed to model the tri-tissue (peripheral blood, spleen and bone marrow) dynamics of hematopoiesis under leukemia. The cell kinetics generated from the non-irradiated experimental model were used as the reference data for modeling. All mathematical formulas were systematically enumerated, and model parameters were estimated via numerical optimization. Multiple validations by additional experimental data were then conducted for the established computational model. In the results, we illustrated that the important fact of functional depression of hematopoietic stem/progenitor cells (HSC/HPC) in leukemic bone marrow (BM), which must require additional experiments to be established, could also be inferred from our computation model that utilized only the cell kinetics data as the input.ConclusionThe digitalized AML model established in the present work is effective for reconstructing the hematopoiesis under leukemia as well as simulating the hematopoietic response to leukemic cell expansion. Given the validity and efficiency, the model can be of potential utilities in future biomedical studies; additionally, the modeling method itself can be also applied elsewhere.Electronic supplementary materialThe online version of this article (doi:10.1186/s12918-016-0308-x) contains supplementary material, which is available to authorized users.

A Molecular Profile of the Endothelial Cell Response to Ionizing Radiation.

Ionizing radiation exposure can cause acute radiation sickness (ARS) by damaging the hematopoietic compartment. Radiation damages quiescent hematopoietic stem cells (HSCs) and proliferating hematopoietic cells, resulting in neutropenia, thrombocytopenia and increased risk for long-term hematopoietic dysfunction and myelodysplasia. While some aspects of the hematopoietic response to radiation injury are intrinsic to hematopoietic cells, the recovery of the HSC pool and overall hematopoiesis is also dependent on signals from bone marrow endothelial cells (BM ECs) within the HSC vascular niche. The precise mechanisms through which BM ECs regulate HSC regeneration remain unclear. Characterization of the altered EC gene expression that occurs in response to radiation could provide a roadmap to the discovery of EC-derived mechanisms that regulate hematopoietic regeneration. Here, we show that 5 Gy total-body irradiation substantially alters the expression of numerous genes in BM ECs within 24 h and this molecular response largely resolves by day 14 postirradiation. Several unique and nonannotated genes, which encode secreted proteins were upregulated and downregulated in ECs in response to radiation. These results highlight the complexity of the molecular response of BM ECs to ionizing radiation and identify several candidate mechanisms that should be prioritized for functional analysis in models of hematopoietic injury and regeneration.

Hepcidin Response to Iron Therapy in Patients with Non-Dialysis Dependent CKD: An Analysis of the FIND-CKD Trial

Hepcidin is the key regulator of iron homeostasis but data are limited regarding its temporal response to iron therapy, and response to intravenous versus oral iron. In the 56-week, open-label, multicenter, prospective, randomized FIND-CKD study, 626 anemic patients with non-dialysis dependent chronic kidney disease (ND-CKD) and iron deficiency not receiving an erythropoiesis stimulating agent were randomized (1:1:2) to intravenous ferric carboxymaltose (FCM), targeting higher (400–600μg/L) or lower (100–200μg/L) ferritin, or to oral iron. Serum hepcidin levels were measured centrally in a subset of 61 patients. Mean (SD) baseline hepcidin level was 4.0(3.5), 7.3(6.4) and 6.5(5.6) ng/mL in the high ferritin FCM (n = 17), low ferritin FCM (n = 16) and oral iron group (n = 28). The mean (SD) endpoint value (i.e. the last post-baseline value) was 26.0(9.1),15.7(7.7) and 16.3(11.0) ng/mL, respectively. The increase in hepcidin from baseline was significantly smaller with low ferritin FCM or oral iron vs high ferritin FCM at all time points up to week 52. Significant correlations were found between absolute hepcidin and ferritin values (r = 0.65, p<0.001) and between final post-baseline increases in both parameters (r = 0.70, p<0.001). The increase in hepcidin levels over the 12-month study generally mirrored the cumulative iron dose in each group. Hepcidin and transferrin saturation (TSAT) absolute values showed no correlation, although there was an association between final post-baseline increases (r = 0.42, p<0.001). Absolute values (r = 0.36, p = 0.004) and final post-baseline increases of hepcidin and hemoglobin (p = 0.30, p = 0.030) correlated weakly. Baseline hepcidin levels were not predictive of a hematopoietic response to iron therapy. In conclusion, hepcidin levels rose in response to either intravenous or oral iron therapy, but the speed and extent of the rise was greatest with intravenous iron targeting a higher ferritin level. However neither the baseline level nor the change in hepcidin was able to predict response to therapy in this cohort.

Terminal differentiation induction as DNA damage response in hematopoietic stem cells by GADD45A.

Hematopoietic stem cells (HSCs) sustain lifelong blood cell regeneration by balancing their ability for self-renewal with their ability to differentiate into all blood cell types. To prevent organ exhaustion and malignant transformation, long-lived HSCs, in particular, must be protected from exogenous and endogenous stress, which cause severe DNA damage. When DNA is damaged, distinct DNA repair mechanisms and cell fate controls occur in adult HSCs compared with committed cells. Growth arrest and DNA damage-inducible 45 alpha (GADD45A) is known to coordinate a variety of cellular stress responses, indicating the molecule is an important stress mediator. So far, the function of GADD45A in hematopoietic stem and progenitor cells is controversial and appears highly dependent on the cell type and stress stimulus. Recent studies have analyzed its role in cell fate decision control of prospectively isolated HSCs and have revealed unexpected functions of GADD45A, as discussed here. The upregulation of GADD45A by DNA damage-causing conditions results in enhanced HSC differentiation, probably to efficiently eliminate aberrant HSCs from the system. These findings, in concert with a few studies on other stem cell systems, have led us to propose DNA damage-induced differentiation as a novel DNA damage response mechanism in stem cells that circumvents the fatal consequences of cumulative DNA damage in the stem cell compartment.

Adiponectin Enhances Antibacterial Activity of Hematopoietic Cells by Suppressing Bone Marrow Inflammation

Obesity has been shown to increase the morbidity ofinfections, however, the underlying mechanisms remain largely unknown. Here we demonstrate thatobesity caused adiponectin deficiency in the bone marrow (BM), which led to an inflamed BM characterized by increased tumor necrosis factor (TNF) production from bone marrow macrophages. Hematopoietic stem and progenitor cells (HSPCs) chronically exposed to excessive TNF in obese marrow aberrantly expressed cytokine signaling suppressor SOCS3, impairing JAK-STAT mediated signal transduction and cytokine-driven cell proliferation. Accordingly, both obese and adiponectin-deficient mice showed attenuated clearance of infected Listeria monocytogenes, indicating that obesity or loss of adiponectin is critical for exacerbation of infection. Adiponectin treatment restored the defective HSPC proliferation and bacterial clearance of obese and adiponectin-deficient mice, affirming the importance of adiponectin against infection. Taken together, our findings demonstrate that obesity impairs hematopoietic response against infections through a TNF-SOCS3-STAT3 axis, highlighting adiponectin as a legitimate target against obesity-related infections.

Sexually dimorphic myeloid inflammatory and metabolic responses to diet-induced obesity.

It is well known in clinical and animal studies that women and men have different disease risk as well as different disease physiology. Women of reproductive age are protected from metabolic and cardiovascular disease compared with postmenopausal women and men. Most murine studies are skewed toward the use of male mice to study obesity-induced metabolic dysfunction because of similar protection in female mice. We have investigated dietary obesity in a mouse model and have directly compared inflammatory responses in males and females. In this review we will summarize what is known about sex differences in diet-induced inflammation and will summarize our data on this topic. It is clear that sex differences in high-fat diet-induced inflammatory activation are due to cell intrinsic differences in hematopoietic responses to obesogenic cues, but further research is needed to understand what leads to sexually dimorphic responses.

203 - Fetal Mesenchymal Stem Cells Exhibit a Wnt-dependent Protective Action on Radiation-induced Hematopoietic Failure

For adult allogeneic bone marrow transplant, myeloablative or chemotherapy is essential used to make space for donor cells and prevent graft rejection. This conditioning regimen is associated with significant morbidity and mortality of 10–15%, and may cause by damage of bone marrow microenvironment for donor stem cell implantation. Our previously data had showed that co-transplanted murine fetal mesenchymal stem cells from amniotic fluid (AFSCs)or placenta (PDSCs) with allogeneic hematopoietic stem cells enhance the hematopoietic cell recovery rate. However, little is known about the action mechanism(s) of AFSC on hematopoietic regeneration. The experimental methodology design of this study is mainly divided into two parts, first part is to verify the hematopoietic supporting and survival activity of AFSCs in inflamed environment. The second part is in vivo evaluated the protective effects of AFSCs systemic administration on hematopoietic regeneration after total body irradiation. AFSCs showed highly resistance to irradiation, CsA and apoptotic inducer, also exhibit anti-apoptotic genes included survivin, MCl-1, XIAP etc. Results of cobblestone formation assay and colony-unit forming assay have shown that AFSCs enhance multiple hematopoietic lineage development. Systemically administrated of AFSCs into irradiated mice have shown that AFSCs alleviate radiation-induced hematopoietic toxic responses and decreased inflammatory damages. Mechanistic, MSCs injection elevated the differentiate turnover rate of bone marrow hematopoietic stem cells into myeloid- and lymphoid- lineages. The Wnt-blocking experiments have proven that radiation rescue action of AFSCs is Wnt-dependent. Conclusions our data revealed that systemically administered fetal MSCs provoke a protective mechanism for supporting hematopoietic reconstitution after radiation exposure. Hence, AFSCs administration may serve as fast and effective first-line treatment cell source to radiation-induced hematopoietic failure.