Summary Like normal hematopoietic stem cells, leukemic stem cells depend on their bone marrow (BM) microenvironment for survival, but the underlying mechanisms remain largely unknown. We have studied the contribution of nestin + BM mesenchymal stem cells (BMSCs) to MLL-AF9-driven acute myeloid leukemia (AML) development and chemoresistance in vivo . Unlike bulk stroma, nestin + BMSC numbers are not reduced in AML, but their function changes to support AML cells, at the expense of non-mutated hematopoietic stem cells (HSCs). Nestin + cell depletion delays leukemogenesis in primary AML mice and selectively decreases AML, but not normal, cells in chimeric mice. Nestin + BMSCs support survival and chemotherapy relapse of AML through increased oxidative phosphorylation, tricarboxylic acid (TCA) cycle activity, and glutathione (GSH)-mediated antioxidant defense. Therefore, AML cells co-opt energy sources and antioxidant defense mechanisms from BMSCs to survive chemotherapy.

The contribution of cell-extrinsic factors in Acute Myeloid Leukemia (AML) generation and persistence has gained interest. Bitter taste receptors (TAS2Rs) are G protein-coupled receptors known for their primary role as a central warning signal to induce aversion toward noxious or harmful substances. Nevertheless, the increasing amount of evidence about their extra-oral localization has suggested a wider function in sensing microenvironment, also in cancer settings. In this study, we found that AML cells express functional TAS2Rs. We also highlighted a significant association between the modulation of some TAS2Rs and the poor-prognosis AML groups, i.e., TP53- and TET2-mutated, supporting a potential role of TAS2Rs in AML cell biology. Gene expression profile analysis showed that TAS2R activation with the prototypical agonist, denatonium benzoate, significantly modulated a number of genes involved in relevant AML cellular processes. Functional assay substantiated molecular data and indicated that denatonium reduced AML cell proliferation by inducing cell cycle arrest in G0/G1 phase or induced apoptosis via caspase cascade activation. Moreover, denatonium exposure impaired AML cell motility and migratory capacity, and inhibited cellular respiration by decreasing glucose uptake and oxidative phosphorylation. In conclusion, our results in AML cells expand the observation of cancer TAS2R expression to the setting of hematological neoplasms and shed light on a role of TAS2Rs in the extrinsic regulation of leukemia cell functions.

Myelofibrosis (MF) is a clonal neoplasia associated with chronic inflammation due to aberrant cytokine production. Mutations in Janus Kinase-2 (JAK2), calreticulin (CALR) and myeloproliferative leukemia protein (MPL) genes have been recently associated to MF and they all activate the JAK/STAT signaling pathway. Since this pathway is essential in shaping the immune response, we investigated the role of circulating immune subsets and cytokines in 38 patients (20 carrying JAK2,13 exon-9 CALR mutation and 5 triple negative). In comparison to healthy donors, patients presented a reduced amount of circulating dendritic cells (DCs) associated with a defective ability of monocytes in differentiating into DCs. In addition, we found a reduction in circulating T-helper (Th)1 and Th17 and hypo-functional innate lymphoid cells (ILC). Results analyzed according to the mutational status showed that patients carrying JAK2 mutation had a reduction in Th17, myeloid-DCs and effector Tregs as well as increased ILC1 and cytokine producing Tregs. The CALR mutated patients revealed high ILC3 levels, reduced Th1 and their monocytes had a reduced capacity to mature into fully committed DCs. Their Tregs were also less effective in inhibiting the proliferation of autologous effector T-cells due to an increased proliferative status induced by CALR mutation. Triple negative patients presented a reduced amount of total circulating CD3, effectors Tregs and Th1 with increased ILC1. Overall, we have demonstrated that in MF different mutations lead to phenotypic and functional alterations in different immune subsets that may have a potential role in disease progression and susceptibility to infections.

In this review article, we present recent updates on the hematologic tumor microenvironment following the 3rd Scientific Workshop on the Haematological Tumour Microenvironment and its Therapeutic Targeting organized by the European School of Hematology, which took place at the Francis Crick Institute in London in February 2019. This review article is focused on recent scientific advances highlighted in the invited presentations at the meeting, which encompassed the normal and malignant niches supporting hematopoietic stem cells and their progeny. Given the precise focus, it does not discuss other relevant contributions in this field, which have been the scope of other recent reviews. The content covers basic research and possible clinical applications with the major therapeutic angle of utilizing basic knowledge to devise new strategies to target the tumor microenvironment in hematologic cancers. The review is structured in the following sections: (i) regulation of normal hematopoietic stem cell niches during development, adulthood and aging; (ii) metabolic adaptation and reprogramming in the tumor microenvironment; (iii) the key role of inflammation in reshaping the normal microenvironment and driving hematopoietic stem cell proliferation; (iv) current understanding of the tumor microenvironment in different malignancies, such as chronic lymphocytic leukemia, multiple myeloma, acute myeloid leukemia and myelodysplastic syndromes; and (v) the effects of therapies on the microenvironment and some opportunities to target the niche directly in order to improve current treatments.

Along with molecular abnormalities (mutations in JAK2, Calreticulin (CALR) and MPL genes), chronic inflammation is the major hallmark of Myelofibrosis (MF). Here, we investigated the in vitro effects of crucial factors of the inflammatory microenvironment (Interleukin (IL)-1β, Tumor Necrosis Factor (TNF)-α, Tissue Inhibitor of Metalloproteinases (TIMP)-1 and ATP) on the functional behaviour of MF-derived circulating CD34+ cells.We found that, regardless mutation status, IL-1β or TNF-α increases the survival of MF-derived CD34+ cells. In addition, along with stimulation of cell cycle progression to the S-phase, IL-1β or TNF-α ± TIMP-1 significantly stimulate(s) the in vitro clonogenic ability of CD34+ cells from JAK2V617 mutated patients. Whereas in the JAK2V617F mutated group, the addition of IL-1β or TNF-α + TIMP-1 decreased the erythroid compartment of the CALR mutated patients. Megakaryocyte progenitors were stimulated by IL-1β (JAK2V617F mutated patients only) and inhibited by TNF-α. IL-1β + TNF-α + C-X-C motif chemokine 12 (CXCL12) ± TIMP-1 highly stimulates the in vitro migration of MF-derived CD34+ cells. Interestingly, after migration toward IL-1β + TNF-α + CXCL12 ± TIMP-1, CD34+ cells from JAK2V617F mutated patients show increased clonogenic ability.Here we demonstrate that the interplay of these inflammatory factors promotes and selects the circulating MF-derived CD34+ cells with higher proliferative activity, clonogenic potential and migration ability. Targeting these micro-environmental interactions may be a clinically relevant approach.

We and others have shown that the Tissue Inhibitor of Metalloproteinases-1 (TIMP-1), a member of the inflammatory network exerting pleiotropic effects in the bone marrow (BM) microenvironment, regulates the survival and proliferation of different cell types, including normal hematopoietic progenitor cells. Moreover, TIMP-1 has been shown to be involved in cancer progression. However, its role in leukemic microenvironment has not been addressed. Here, we investigated the activity of TIMP-1 on Acute Myelogenous Leukemia (AML) cell functions. First, we found that TIMP-1 levels were increased in the BM plasma of AML patients at diagnosis. In vitro, recombinant human (rh)TIMP-1 promoted the survival and cell cycle S-phase entry of AML cells. These kinetic effects were related to the downregulation of cyclin-dependent kinase inhibitor p21. rhTIMP-1 increases CXCL12-driven migration of leukemic cells through PI3K signaling. Interestingly, activation of CD63 receptor was required for TIMP-1's cytokine/chemokine activity. Of note, rhTIMP-1 stimulation modulated mRNA expression of Hypoxia Inducible Factor (HIF)-1α, downstream of PI3K/Akt activation. We then co-cultured AML cells with normal or leukemic mesenchymal stromal cells (MSCs) to investigate the interaction of TIMP-1 with cellular component(s) of BM microenvironment. Our results showed that the proliferation and migration of leukemic cells were greatly enhanced by rhTIMP-1 in presence of AML-MSCs as compared to normal MSCs. Thus, we demonstrated that TIMP-1 modulates leukemic blasts survival, migration and function via CD63/PI3K/Akt/p21 signaling. As a “bad actor” in a “bad soil”, we propose TIMP-1 as a potential novel therapeutic target in leukemic BM microenvironment.

No abstract

IntroductionDue to their immunomodulatory properties, mesenchymal stromal cells (MSCs) have been used for auto-immune disease treatment. Crohn disease (CD) and ulcerative colitis are two major inflammatory bowel diseases (IBDs), resulting from pathological immune responses to environmental or microbial antigens. Preclinical and clinical studies have suggested that MSC-based cellular therapy hold promising potential for IBD treatment. However, open issues include the selection of the proper cell dose, the source and the optimal route of administration of MSCs for more effective results. Platelet lysate has gained clinical interest due to its efficacy in accelerating wound healing. Thus, we propose to combine the administration of MSCs with a human umbilical cord blood-derived platelet lysate (hCBPL) as a novel strategy to improve MSC-based therapy for IBD resolution.MethodsColitis was induced in 8-week-old C57BL/6J mice by daily oral administration of dextran sulphate sodium (DSS) (1.5 % w/v in tap water) for 9 days. MSCs were isolated from adipose tissue of CD patients (adCD-MSCs), expanded in proliferation medium, resuspended in hCBPL or PBS and administrated via enema for three times (1 × 106 cells/mouse/time) every other day starting on day +7 from DSS induction. The colitis evolution was evaluated by daily monitoring of body weight, stool consistency and bleeding. Histopathological analysis was performed. Inflammatory cytokine plasma levels were determined. adCD-MSCs stained with lipophilic membrane dye Nile Red, were injected in DSS mice as described above. Colon section of mice sacrificed 24 hours after last cell administration, were analyzed by confocal microscopy.ResultsWe found that adCD-MSCs could be easily isolated and expanded from CD patients. Upon injection, adCD-MSCs exerted a therapeutic effect on DSS-induced colitis. Moreover, hCBPL increased adCD-MSCs efficacy by significantly reducing colitis scores, extension of the colon inflamed area and plasma levels of inflammatory mediators. Finally, Nile Red staining of MSCs is very efficient, stable and does not impair their vitality and function. Nile Red-labelling was clearly detected in the colitic area of adCD-MSCs injected mice and it was significantly brighter in the colon sections of mice that had received adCD-MSCs/hCBPL.ConclusionsIn summary, with this study we propose a novel and promising adCD-MSC/hCBPL-based therapy for refractory IBDs.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-015-0166-2) contains supplementary material, which is available to authorized users.