c-Kit M541L variant is related to ineffective hemopoiesis predisposing to clonal evolution in 3D in vitro biomimetic co-culture model of bone marrow niche

Manzo, Paola; Scala, Pasqualina; Giudice, Valentina; Gorrese, Marisa; Bertolini, A.; Morini, Denise; D'Alto, Francesca; Pepe, Rita; Pedicini, Antonio; Izzo, Barbara; Verdesca, Francesco; Langella, Maddalena; Serio, Bianca; Porta, Giovanna Della; Selleri, Carmine

doi:10.1016/j.heliyon.2022.e11998

Cited by 5 publications

(4 citation statements)

References 62 publications

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“…At light microscopy examination, adherent cells had fibroblast-like spindle shapes; however, diffuse monocyte-like cells were also observed (Figure 1). The presence of these two different populations was also confirmed by flow cytometry immunophenotyping showing a mesenchymal-like population with positivity for CD90 and negativity for CD14, CD45, and HLA-DR [59,60], while a small fraction (0.2-2.6% of total nucleated cells) had an M2-macrophage-like phenotype with positivity for CD163 and CD206 (Figure 2). The presence of these two different populations was also confirmed by flow cytometry immunophenotyping showing a mesenchymal-like population with positivity for CD90 and negativity for CD14, CD45, and HLA-DR [59,60], while a small fraction (0.2-2.6% of total nucleated cells) had an M2-macrophage-like phenotype with positivity for CD163 and CD206 (Figure 2).…”

Section: Preliminary Study: Potential Clinical Relevance Of Upar Bloc...mentioning

confidence: 69%

“…The presence of these two different populations was also confirmed by flow cytometry immunophenotyping showing a mesenchymal-like population with positivity for CD90 and negativity for CD14, CD45, and HLA-DR [ 59 , 60 ], while a small fraction (0.2–2.6% of total nucleated cells) had an M2-macrophage-like phenotype with positivity for CD163 and CD206 ( Figure 2 ).…”

Section: Preliminary Study: Potential Clinical Relevance Of Upar Bloc...mentioning

confidence: 82%

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Macrophages and Urokinase Plasminogen Activator Receptor System in Multiple Myeloma: Case Series and Literature Review

Manzo

Giudice

Napolitano

et al. 2023

IJMS

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The microenvironment plays an essential role in multiple myeloma (MM) development, progression, cell proliferation, survival, immunological escape, and drug resistance. Mesenchymal stromal cells and macrophages release tolerogenic cytokines and favor anti-apoptotic signaling pathway activation, while the urokinase plasminogen activator receptor (uPAR) system contributes to migration through an extracellular matrix. Here, we first summarized the role of macrophages and the uPAR system in MM pathogenesis, and then we reported the potential therapeutic effects of uPAR inhibitors in a case series of primary MM-derived adherent cells. Our preliminary results showed that after uPAR inhibitor treatments, interleukein-6 (mean ± SD, 8734.95 ± 4169.2 pg/mL vs. 359.26 ± 393.8 pg/mL, pre- vs. post-treatment; p = 0.0012) and DKK-1 levels (mean ± SD, 7005.41 ± 6393.4 pg/mL vs. 61.74 ± 55.2 pg/mL, pre- vs. post-treatment; p = 0.0043) in culture medium were almost completely abolished, supporting further investigation of uPAR blockade as a therapeutic strategy for MM treatment. Therefore, uPAR inhibitors could exert both anti-inflammatory and pro-immunosurveillance activity. However, our preliminary results need further validation in additional in vitro and in vivo studies.

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Section: Preliminary Study: Potential Clinical Relevance Of Upar Bloc...mentioning

confidence: 69%

Section: Preliminary Study: Potential Clinical Relevance Of Upar Bloc...mentioning

confidence: 82%

Macrophages and Urokinase Plasminogen Activator Receptor System in Multiple Myeloma: Case Series and Literature Review

Manzo

Giudice

Napolitano

et al. 2023

IJMS

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“…On the other hand, biomimetic 3D cultures of stem cell represent a versatile tool for the study of myogenic commitment events ( Witt et al, 2017 ; Ergene et al, 2020 ). Indeed, BM-MSCs are frequently cultured in 3D hydrogel scaffolds, like fibrin hydrogels ( Govoni et al, 2017 ; Ciardulli et al, 2020 ; 2021 ; Manzo et al, 2022 ) that efficiently mimic SC niche ( Pollot et al, 2018 ) and may reproduce in vitro the hierarchical structure of skeletal muscle tissue. Moreover, fibrin-based scaffolds constitute a regenerative myogenic environment without fibrosis, as cells produce their own extracellular matrix (ECM) proteins and degraded fibrin excess allowing long-term skeletal muscle cell culture ( Huang et al, 2005 ; Gilbert-Honick et al, 2018 ; Matthias et al, 2018 ).…”

Section: Introductionmentioning

confidence: 99%

Peripheral blood mononuclear cells contribute to myogenesis in a 3D bioengineered system of bone marrow mesenchymal stem cells and myoblasts

Scala

Manzo

Lamparelli

et al. 2023

Front. Bioeng. Biotechnol.

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In this work, a 3D environment obtained using fibrin scaffold and two cell populations, such as bone marrow-derived mesenchymal stem cells (BM-MSCs), and primary skeletal muscle cells (SkMs), was assembled. Peripheral blood mononuclear cells (PBMCs) fraction obtained after blood filtration with HemaTrate® filter was then added to the 3D culture system to explore their influence on myogenesis. The best cell ratio into a 3D fibrin hydrogel was 1:1 (BM-MSCs plus SkMs:PBMCs) when cultured in a perfusion bioreactor; indeed, excellent viability and myogenic event induction were observed. Myogenic genes were significantly overexpressed when cultured with PBMCs, such as MyoD1 of 118-fold at day 14 and Desmin 6-fold at day 21. Desmin and Myosin Heavy Chain were also detected at protein level by immunostaining along the culture. Moreover, the presence of PBMCs in 3D culture induced a significant downregulation of pro-inflammatory cytokine gene expression, such as IL6. This smart biomimetic environment can be an excellent tool for investigation of cellular crosstalk and PBMC influence on myogenic processes.

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“…We hypothesize that augmented mutagenesis might be determined by the presence of germline predisposition to genomic instability in a context of ineffective hemopoiesis. Indeed, we have previously demonstrated that the M541L c‐Kit somatic mutation could be a loss‐of‐function alteration by reducing HSC maintenance causing an in vivo graft failure and in vitro absence of long‐term culture stability in a 3D BM model 19,20 . In this study, to elucidate pathophysiology of clonal hemopoiesis in hematological malignancies, we retrospectively investigated clinical significance of mutations in leukemia‐related genes of known pathogenetic significance and VUS in a cohort of patients with CCUS, MDS, and AML.…”

Section: Introductionmentioning

confidence: 99%

Subclones with variants of uncertain clinical significance might contribute to ineffective hemopoiesis and leukemia predisposition

Giudice

Serio

Errichiello

et al. 2023

European J of Haematology

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BackgroundSplicing modifications, genomic instability, and hypomethylation are central mechanisms promoting myelodysplasia and acute myeloid leukemia (AML). In this real‐life retrospective study, to elucidate pathophysiology of clonal hemopoiesis in hematological malignancies, we investigated clinical significance of mutations in leukemia‐related genes of known pathogenetic significance and of variants of uncertain clinical significance (VUS) in a cohort of patients with MDS and AML.MethodsA total of 59 consecutive subjects diagnosed with MDS, 48 with AML, and 17 with clonal cytopenia with unknown significance were screened for somatic mutations in AML‐related genes by next‐generation sequencing.ResultsWe showed that TET2, SETBP1, ASXL1, EZH2, RUNX1, SRSF2, DNMT3A, and IDH1/2 were commonly mutated. MDS patients also showed a high genetic complexity, especially for SETBP1. Moreover, the presence of SETBP1 wild‐type or two or more simultaneous VUS variants identified a subgroup of AML and MDS patients with better outcome, while the presence of single SETBP1 VUS variant was related to a worse prognosis, regardless TET2 mutational status.ConclusionsIn conclusions, we linked both pathogenic and VUS variants in AML‐related genes to clonal hematopoiesis; therefore, we proposed to consider those variants as prognostic markers in leukemia and myelodysplasia. However, further studies in larger prospective cohorts are required to validate our results.

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c-Kit M541L variant is related to ineffective hemopoiesis predisposing to clonal evolution in 3D in vitro biomimetic co-culture model of bone marrow niche

Cited by 5 publications

References 62 publications

Macrophages and Urokinase Plasminogen Activator Receptor System in Multiple Myeloma: Case Series and Literature Review

Macrophages and Urokinase Plasminogen Activator Receptor System in Multiple Myeloma: Case Series and Literature Review

Peripheral blood mononuclear cells contribute to myogenesis in a 3D bioengineered system of bone marrow mesenchymal stem cells and myoblasts

Subclones with variants of uncertain clinical significance might contribute to ineffective hemopoiesis and leukemia predisposition

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