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RATIONALE - Pulmonary arterial hypertension is a deadly disease of the pulmonary vasculature for which no disease-modifying therapies exist. Small-vessel stiffening and remodeling are fundamental pathological features of pulmonary arterial hypertension that occur early and drive further endovascular cell dysfunction. Bone marrow (BM)-derived proangiogenic cells (PACs), a specialized heterogeneous subpopulation of myeloid lineage cells, are thought to play an important role in pathogenesis.
OBJECTIVE - To determine whether BM-derived PACs directly contributed to experimental pulmonary hypertension (PH) by promoting small-vessel stiffening through 5-HT (serotonin 2B receptor)-mediated signaling.
METHODS AND RESULTS - We performed BM transplants using transgenic donor animals expressing diphtheria toxin secondary to activation of an endothelial-specific tamoxifen-inducible Cre and induced experimental PH using hypoxia with SU5416 to enhance endovascular injury and ablated BM-derived PACs, after which we measured right ventricular systolic pressures in a closed-chest procedure. BM-derived PAC lineage tracing was accomplished by transplanting BM from transgenic donor animals with fluorescently labeled hematopoietic cells and treating mice with a 5-HT antagonist. BM-derived PAC ablation both prevented and reversed experimental PH with SU5416-enhanced endovascular injury, reducing the number of muscularized pulmonary arterioles and normalizing arteriole stiffness as measured by atomic force microscopy. Similarly, treatment with a pharmacological antagonist of 5-HT also prevented experimental PH, reducing the number and stiffness of muscularized pulmonary arterioles. PACs accelerated pulmonary microvascular endothelial cell injury response in vitro, and the presence of BM-derived PACs significantly correlated with stiffer pulmonary arterioles in pulmonary arterial hypertension patients and mice with experimental PH. RNA sequencing of BM-derived PACs showed that 5-HT antagonism significantly altered biologic pathways regulating cell proliferation, locomotion and migration, and cytokine production and response to cytokine stimulus.
CONCLUSIONS - Together, our findings illustrate that BM-derived PACs directly contribute to experimental PH with SU5416-enhanced endovascular injury by mediating small-vessel stiffening and remodeling in a 5-HT signaling-dependent manner.

Bcl-2 family proteins reorganize mitochondrial membranes during apoptosis, to form pores and rearrange cristae. In vitro and in vivo analysis integrated with human genetics reveals a novel homeostatic mitochondrial function for Bcl-2 family protein Bid. Loss of full-length Bid results in apoptosis-independent, irregular cristae with decreased respiration. mice display stress-induced myocardial dysfunction and damage. A gene-based approach applied to a biobank, validated in two independent GWAS studies, reveals that decreased genetically determined BID expression associates with myocardial infarction (MI) susceptibility. Patients in the bottom 5% of the expression distribution exhibit >4 fold increased MI risk. Carrier status with nonsynonymous variation in Bid's membrane binding domain, Bid, associates with MI predisposition. Furthermore, Bid but not Bid associates with Mcl-1, previously implicated in cristae stability; decreased MCL-1 expression associates with MI. Our results identify a role for Bid in homeostatic mitochondrial cristae reorganization, that we link to human cardiac disease.
© 2018, Salisbury-Ruf et al.

The tumor-cell microenvironment is recognized as a dynamic place where critical cell interactions occur and play an important role in altering tumorigenesis. While many studies have investigated the effects of cellular cross-talk within distinct tumor microenvironments, these interactions have yet to be fully examined in bone. It is well-established that many common cancers metastasize to bone, resulting in the development of tumor-induced bone disease (TIBD), a multi-facetted illness that is driven by complex cell interactions within the bone marrow. Our group has previously published that myeloid progenitor cells expand in the presence of tumors in bone, aligning with the notion that myeloid cells can act as tumor promotors. Several groups, including ours, have established that transforming growth factor β (TGF-β), an abundant growth factor in bone, can regulate both TIBD and myeloid expansion. TGF-β inhibitors have been shown to increase bone volume, decrease bone destruction, and reduce but not eliminate tumor. Therefore, we hypothesize that inhibiting TGF-β will reduce myeloid expansion leading to a reduction of tumor burden in bone and osteoclast-mediated bone loss, causing to an overall reduction in TIBD. To address this hypothesis, two different mouse models of breast cancer bone colonization were pre-treated with the TGF-β neutralizing antibody, 1D11, prior to tumor inoculation (athymic: MDA-MB-231, BALB/c: 4T1) and continuously treated until sacrifice. Additionally, a genetically modified mouse model with a myeloid specific deletion of transforming growth factor beta receptor II (TGF-βRII) (TGF-βRII) was utilized in our studies. Systemic inhibition of TGF-β lead to fewer osteolytic lesions, and reduced tumor burden in bone as expected from previous studies. Additionally, early TGF-β inhibition affected expansion of distinct myeloid populations and shifted the cytokine profile of pro-tumorigenic factors in bone, 4T1 tumor cells, and bone-marrow derived macrophages. Similar observations were seen in tumor-bearing TGF-βRII mice, where these mice contained fewer bone lesions and significantly less tumor burden in bone, suggesting that TGF-β inhibition regulates myeloid expansion leading to a significant reduction in TIBD.
Published by Elsevier Inc.

Modifications of cardiolipin (CL) levels or compositions are associated with changes in mitochondrial function in a wide range of pathologies. We have made the discovery that acetaminophen remodels CL fatty acids composition from tetralinoleoyl to linoleoyltrioleoyl-CL, a remodeling that is associated with decreased mitochondrial respiration. Our data show that CL remodeling causes a shift in electron entry from complex II to the β-oxidation electron transfer flavoprotein quinone oxidoreductase (ETF/QOR) pathway. These data demonstrate that electron entry in the respiratory chain is regulated by CL fatty acid composition and provide proof-of-concept that pharmacological intervention can be used to modify CL composition.
Copyright © 2016 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

Hematopoietic stem cells (HSCs) possess long-term self-renewal capacity and multipotent differentiative capacity, to maintain the hematopoietic system. Long-term hematopoietic homeostasis requires effective control of genotoxic damage to maintain HSC function and prevent propagation of deleterious mutations. Here we investigate the role of the BH3-only Bcl-2 family member Bid in the response of murine hematopoietic cells to long-term replicative stress induced by hydroxyurea (HU). The PI3-like serine/threonine kinase, ATR, initiates the DNA damage response (DDR) to replicative stress. The pro-apoptotic Bcl-2 family member, Bid, facilitates this response to replicative stress in hematopoietic cells, but the in vivo role of this DDR function of Bid has not been defined. Surprisingly, we demonstrate that long-term HU treatment expands wild-type myeloid progenitor cells (MPCs) and HSC-enriched Lin(-)Sca1(+)Kit(+) (LSK) cells to maintain bone marrow function as measured by long-term competitive repopulating ability. Bid-/- MPCs demonstrate increased sensitivity to HU and are depleted. Bid-/- LSK cells demonstrate increased mobilization manifest by increased Bromodeoxyuridine (BrdU) incorporation. Bid-/- MPCs and LSK cells are relatively depleted, however, and bone marrow from Bid-/- mice demonstrates decreased long-term competitive repopulating ability in both primary and secondary transplants. We thus describe a survival function of Bid in hematopoiesis in the setting of chronic replicative stress.

By using flow cytometry, we analyzed myeloid nuclear differentiation antigen (MNDA) expression in myeloid precursors in bone marrow from patients with myelodysplastic syndrome (MDS) and control samples from patients undergoing orthopedic procedures. The median percentage of MNDA-dim myeloid precursors in MDS cases was 67.4% (range, 0.7%-97.5%; interquartile range, 44.9%-82.7%) of myeloid cells, with bimodal MNDA expression in most MDS samples. Control samples demonstrated a median MNDA-dim percentage in myeloid precursors of 1.2% (range, 0.2%-13.7%; interquartile range, 0.6%-2.7%), with no bimodal pattern in most samples. The area under the receiver operating characteristic curve for MNDA-dim percentage in myeloid precursors was 0.96 (P = 9 × 10(-7)). Correlation of MNDA-dim levels with World Health Organization 2008 morphologic diagnoses was not significant (P = .21), but correlation with patient International Prognostic Scoring System scores suggested a trend (P = .07). Flow cytometric assessment of MNDA in myeloid precursors in bone marrow may be useful for the diagnosis of MDS.

While a number of DNA binding transcription factors have been identified that control hematopoietic cell fate decisions, only a limited number of transcriptional corepressors (e.g., the retinoblastoma protein [pRB] and the nuclear hormone corepressor [N-CoR]) have been linked to these functions. Here, we show that the transcriptional corepressor Mtg16 (myeloid translocation gene on chromosome 16), which is targeted by t(16;21) in acute myeloid leukemia, is required for hematopoietic progenitor cell fate decisions and for early progenitor cell proliferation. Inactivation of Mtg16 skewed early myeloid progenitor cells toward the granulocytic/macrophage lineage while reducing the numbers of megakaryocyte-erythroid progenitor cells. In addition, inactivation of Mtg16 impaired the rapid expansion of short-term stem cells, multipotent progenitor cells, and megakaryocyte-erythroid progenitor cells that is required under hematopoietic stress/emergency. This impairment appears to be a failure to proliferate rather than an induction of cell death, as expression of c-Myc, but not Bcl2, complemented the Mtg16(-/-) defect.

Reduced levels of human myeloid nuclear differentiation antigen (MNDA) gene transcripts have been detected in both familial and sporadic cases of myelodysplastic syndromes (MDS). Numerous reports implicate elevated apoptosis/programmed cell death and death ligands and their receptors in the pathogenesis of MDS. MNDA and related proteins contain the pyrin domain that functions in signaling associated with programmed cell death and inflammation. We tested the hypothesis that MNDA is involved in the regulation of programmed cell death in human myeloid hematopoietic cells. Clones of K562 cells (MNDA-null) that expressed ectopic MNDA protein were established using retroviral transduction. MNDA-expressing K562 clones were resistant to tumor necrosis factor-related apoptosis inducing ligand (TRAIL)-induced apoptosis, but were not protected from programmed cell death induced with genotoxic agents or H(2)O(2). MNDA protein expression assessed in control and intermediate and high-grade MDS marrows showed several patterns of aberrant reduced MNDA. These variable patterns of dysregulated MNDA expression may relate to the variable pathophysiology of MDS. We propose that MNDA has a role regulating programmed cell death in myeloid progenitor cells, and that its down-regulation in MDS is related to granulocyte-macrophage progenitor cell sensitivity to TRAIL-induced programmed cell death.

The BCL-2 family of apoptotic proteins encompasses key regulators proximal to irreversible cell damage. The BH3-only members of this family act as sentinels, interconnecting specific death signals to the core apoptotic pathway. Our previous data demonstrated a role for BH3-only BID in maintaining myeloid homeostasis and suppressing leukemogenesis. In the absence of Bid, mice accumulate chromosomal aberrations and develop a fatal myeloproliferative disorder resembling chronic myelomonocytic leukemia. Here, we describe a role for BID in preserving genomic integrity that places BID at an early point in the path to determine the fate of a cell. We show that BID plays an unexpected role in the intra-S phase checkpoint downstream of DNA damage distinct from its proapoptotic function. We further demonstrate that this role is mediated through BID phosphorylation by the DNA-damage kinase ATM. These results establish a link between proapoptotic Bid and the DNA-damage response.