Inflamed Bone Marrow Research Articles

3132 – IMPACT OF THE INFLAMED BONE MARROW NICHE ON THE PROGRESSION OF MYELOPROLIFERATIVE NEOPLASIA

Bcr/Abl-negative myeloproliferative neoplasia (MPN) is characterized by driver mutations in MPL, CALR, or JAK2, as well as an inflammatory microenvironment. The acquisition of clonal abnormalities and secondary mutations in MPN patients can progress the disease to Acute Myeloid Leukemia (AML), resulting in bone marrow (BM) fibrosis and BM failure. For these patients, treatment options fall short and do not include approaches to prevent leukemic transformation. Our group has developed an animal model for inflammation where loss of RBPJ/Notch signaling in the BM niche leads to upregulation of miR155 and an accumulation of proinflammatory cytokines. We propose to use this model (RBPJKO mouse) to study the influence of chronic inflammation on JAK2V617F-driven MPN progression. We hypothesize that chronic inflammation in the BM niche: i) promotes proliferation and genomic instability in hematopoietic cells; and ii) cooperates with preexisting genetic mutations such as JAK2V617F to promote clonal expansion and selection, accelerating disease progression. Our preliminary studies indicate that JAK2V617F hematopoietic cells drive a more aggressive disease when exposed to chronic inflammation following transplant into RBPJKO inflamed BM niches. Indeed, JAK2V617F/RBPJKO recipients exhibited accelerated neutrophilia, higher myeloid skewing and increased bone marrow cellularity and splenomegaly, when compared to JAK2V617F/RBPJWT recipients. By using alkaline comet assay and gH2Ax foci, we also demonstrate that JAK2V617F cooperates with the inflamed BM niche to promote more DNA single- and double-strand breaks in Lin- progenitors in vivo. To further study these events, we optimized an in vitro model in which WT and JAK2V617F BM Lin- progenitors are co-cultured with RBPJKO or RBPJWT BM-derived stroma. JAK2V617F cells showed increased proliferation, mitochondrial ROS accumulation, elevated 53BP1 foci and increased γH2AX MFI when exposed to the inflammatory conditions supported by RBPJKO stroma. Taken together, these data suggest that chronic inflammation can accentuate the MPN phenotype, promote replicative stress and lead to buildup of DNA-damaging ROS in JAK2V617F MPN cells. This work has the potential to provide us with new insights into the mechanisms of MPN progression and the opportunity to identify new therapeutic approaches.

Histographic analysis of oedema and fat in inflamed bone marrow based on quantitative MRI

ObjectiveTo demonstrate proof-of-concept for a quantitative MRI method using histographic analysis to assess bone marrow oedema and fat metaplasia in the sacroiliac joints.Materials and methodsFifty-three adolescents aged 12–23 with known or suspected sacroiliitis were prospectively recruited and underwent quantitative MRI (qMRI) scans, consisting of chemical shift-encoded (at 3 T) and diffusion-weighted imaging (at 1.5 T), plus conventional MRI (at 1.5 T) and clinical assessment. qMRI scans produced proton-density fat fraction (PDFF) and apparent diffusion coefficient (ADC) maps of the sacroiliac joints (SIJs), which were analysed using an in-house software tool enabling partially automated ROI definition and histographic analysis. Logistic regression and receiver operating characteristic (ROC) analyses assessed the predictive performance of ADC- and PDFF-based parameters in identifying active inflammation (oedema) and structural damage (fat metaplasia).ResultsADC-based parameters were associated with increased odds of oedema (all p < 0.05); ROC-AUC was higher for histographic parameters representing the upper end of the ADC distribution than for simple averages. Similarly, PDFF-based parameters were associated with increased odds of fat metaplasia (all p < 0.05); ROC area-under-the-curve was higher for histographic parameters representing the upper end of the PDFF distribution than for simple averages. Both ADC- and PDFF-based histographic parameters demonstrated excellent inter- and intra-observer agreement (ICC > 0.9).ConclusionsADC-based parameters can differentiate patients with bone marrow oedema from those without, whilst PDFF-based parameters can differentiate patients with fat metaplasia from those without. Histographic analysis might improve performance compared with simple averages such as the mean and median and offers excellent agreement within and between observers.Key Points• Quantitative MRI with histographic analysis can identify bone marrow oedema (an active inflammatory lesion) and fat metaplasia (a ‘chronic’ inflammatory lesion) in patients with spondyloarthritis.• The use of histographic analysis might improve the performance of quantitative MRI for detecting bone marrow oedema and fat metaplasia compared with simple averages such as the mean and median.• Bone marrow oedema and fat metaplasia are known to be of diagnostic and prognostic significance, and the proposed method could support clinical decisions around biologic (and other) therapies in spondyloarthritis.

Reacting to Inflammatory Signals

Specialized cellular niches in the bone marrow (BM) modulate critical functions of the hematopoietic stem and progenitor cells, such as self-renewal, cell-fate decisions and the balance between proliferation and differentiation. Constituents of the BM niche, such as osteoblasts, endothelial cells and macrophages, respond to inflammation with secretion of pro-inflammatory cytokines, and serve as effectors of the inflammatory circuitry; thus their excessive activation can potentially impact on the regulation of hematopoietic cells, and on the initiation and progression of myeloid malignancies. Our current understanding of the mechanisms for Bcr/Ab-negative myeloproliferative neoplasia (MPN) involves recognition of driver mutations (targeting i.e. Tet2, Jak2, Mpl, or Asxl) and of an inflammatory microenvironment. However, despite accumulating evidences on the role of inflammation and of the microenvironment in the maintenance and progression of myeloid malignancies, it is still unclear how the inflamed BM niche contributes to disease establishment and progression. Similarly, the causes leading to an inflammatory microenvironment in MPN are not fully understood. We use genetically-controlled animal model of BM inflammation to study the contribution of the inflammatory microenvironment to MPN. We found that loss of Notch signaling in the BM microenvironment results in an inflammatory state of the BM niche promoting myeloproliferation and marrow fibrosis. Using this model, we discovered that loss of Notch/RBPJ signaling leads to transcriptional upregulation of the pro-inflammatory microRNA miR-155, especially in BM endothelial and mesenchymal cells, resulting in miR-155-dependent inhibition of the NF-κB inhibitor κB-Ras1. Decreased levels of kB-Ras1 resulted in the heightened and persistent activation of NF-κB and in the increased production of NF-kB dependent pro-inflammatory cytokines, leading to uncontrolled myeloproliferation. Deletion of miR-155 in the stroma of RBPJ-/- mice prevented the development of the myeloproliferative disease, and patients affected by MPN exhibit elevated expression of miR155 in their BM. Therefore, we hypothesize that the Notch/miR155/NF-kB axis regulates the level of the inflammatory tonus in the BM niche, and that persistent deregulation of this pathway may contribute to the establishment and progression of MPN. In this presentation, we will discuss our findings and ongoing studies in the context of other recent insights on the crosstalk between tumor cells and the BM niche, and their implications for disease initiation, progression and therapeutic approaches. Disclosures No relevant conflicts of interest to declare.