Endothelial colony-forming cells from patients with chronic myeloproliferative disorders lack the disease-specific molecular clonality marker

Endothelial progenitor cells (EPCs) home from the bone marrow to the site of tissue regeneration and sustain neovascularization after acute vascular injury and upon the angiogenic switch in solid tumors. Therefore, they represent a suitable tool for cell-based therapy (CBT) in regenerative medicine and provide a novel promising target in the fight against cancer. Intracellular Ca2+ signals regulate numerous endothelial functions, such as proliferation and tubulogenesis. The growth of endothelial colony forming cells (ECFCs), which are EPCs capable of acquiring a mature endothelial phenotype, is governed by store-dependent Ca2+ entry (SOCE). This study aimed at investigating the nature and the role of VEGF-elicited Ca2+ signals in ECFCs. VEGF induced asynchronous Ca2+ oscillations, whose latency, amplitude, and frequency were correlated to the growth factor dose. Removal of external Ca2+ (0Ca2+) and SOCE inhibition with N-(4-[3,5-bis(trifluoromethyl)-1H-pyrazol-1-yl]phenyl)-4-methyl-1,2,3-thiadiazole-5-carboxamide (BTP-2) reduced the duration of the oscillatory signal. Blockade of phospholipase C-γ with U73122, emptying the inositol-1,4,5-trisphosphate (InsP3)-sensitive Ca2+ pools with cyclopiazonic acid (CPA), and inhibition of InsP3 receptors with 2-APB prevented the Ca2+ response to VEGF. VEGF-induced ECFC proliferation and tubulogenesis were inhibited by the Ca2+-chelant, BAPTA, and BTP-2. NF-κB activation by VEGF was impaired by BAPTA, BTP-2, and its selective blocker, thymoquinone. Thymoquinone, in turn, suppressed VEGF-dependent ECFC proliferation and tubulogenesis. These data indicate that VEGF-induced Ca2+ oscillations require the interplay between InsP3-dependent Ca2+ release and SOCE, and promote ECFC growth and tubulogenesis by engaging NF-κB. This novel signaling pathway might be exploited to enhance the outcome of CBT and chemotherapy.

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“…That ECFC-derived colonies belonged to endothelial lineage was confirmed as described in refs. [20,22].…”

Section: Isolation and Cultivation Of Ecfcsmentioning

confidence: 99%

Vascular Endothelial Growth Factor Stimulates Endothelial Colony Forming Cells Proliferation and Tubulogenesis by Inducing Oscillations in Intracellular Ca2+ Concentration

et al. 2011

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“…2 Because Philadelphia (Ph)-negative myeloproliferative neoplasms (MPNs) show a high incidence of vascular complications 3 and an endothelial cell dysfunction has been evidenced in patients with polycythemia vera (PV), 4 several studies explored the possibility that the neoplastic transformation in MPN could involve also the endothelial cell compartment. [5][6][7][8][9] On the whole, studies that were based on the in vitro assays for endothelial progenitors have identified JAK2 V617F mutation or specific chromosome alterations only in endothelial progenitors derived from the hematopoietic lineage (the so-called colony forming unit-endothelial cells; CFU-ECs), [5][6][7] whereas the true endothelial colony-forming cells, (E-CFCs) do not harbor genetic abnormalities. 6,7 In conflict with these findings, Sozer et al 8 found that endothelial cells isolated by microdissection from liver biopsies of patients with PV with Budd-Chiari Syndrome (BCS) exhibit the JAK2 V617F mutation.…”

Section: Introductionmentioning

confidence: 99%

“…[5][6][7][8][9] On the whole, studies that were based on the in vitro assays for endothelial progenitors have identified JAK2 V617F mutation or specific chromosome alterations only in endothelial progenitors derived from the hematopoietic lineage (the so-called colony forming unit-endothelial cells; CFU-ECs), [5][6][7] whereas the true endothelial colony-forming cells, (E-CFCs) do not harbor genetic abnormalities. 6,7 In conflict with these findings, Sozer et al 8 found that endothelial cells isolated by microdissection from liver biopsies of patients with PV with Budd-Chiari Syndrome (BCS) exhibit the JAK2 V617F mutation. In reality, granulocytes isolated from patients with MPN can harbor several genetic defects in addition to the JAK2 V617F mutation, such as the hypermethylation of suppressor of cytokine signaling (SOCS) genes [10][11][12] or the presence of a clonal pattern of proliferation.…”

Section: Introductionmentioning

confidence: 99%

Endothelial progenitor cells are clonal and exhibit the JAK2V617F mutation in a subset of thrombotic patients with Ph-negative myeloproliferative neoplasms

Teofili¹,

Martini

Iachininoto³

et al. 2011

Blood

112

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In this study we investigated whether neoplastic transformation occurring in Philadelphia (Ph)-negative myeloproliferative neoplasms (MPNs) could involve also the endothelial cell compartment. We evaluated the level of endothelial colony-forming cells (E-CFCs) in 42 patients (15 with polycythemia vera, 12 with essential thrombocythemia, and 15 with primary myelofibrosis). All patients had 1 molecular abnormality (JAK2 V617F or MPL W515K mutations, SOCS gene hypermethylation, clonal pattern of growth) detectable in their granulocytes. The growth of colonies was obtained in 22 patients and, among them, patients with primary myelofibrosis exhibited the highest level of E-CFCs. We found that E-CFCs exhibited no molecular abnormalities in12 patients, had SOCS gene hypermethylation, were polyclonal at human androgen receptor analysis in 5 patients, and resulted in JAK2 V617F mutated and clonal in 5 additional patients, all experiencing thrombotic complications. On the whole, patients with altered E-CFCs required antiproliferative therapy more frequently than patients with normal E-CFCs. Moreover JAK2 V617F -positive E-CFCs showed signal transducer and activator of transcription 5 and 3 phosphorylation rates higher than E-CFCs isolated from healthy persons and patients with MPN without molecular abnormalities. Finally, JAK2 V617F -positive E-CFCs exhibited a high proficiency to adhere to normal mononuclear cells. This study highlights a novel mechanism underlying the thrombophilia observed in MPN. (Blood. 2011;117(9):2700-2707)

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“…These studies are particularly important because Philadelphia-negative MPNs show a high incidence of vascular complications. Initial studies based on in vitro assays for endothelial progenitors showed that JAK2 V617F mutations were identified only in CFU-ECs (colonies formed by monocytoid cells with pro-angiogenic activity), but not in E-CFCs (colonies by true endothelial cells) (11). At variance with these findings, Sozer and coworkers reported that endothelial cells isolated by microdissection from liver biopsies of patients with PV with Budd-Chiari syndrome exhibit the JAK2 V617F mutation (45).…”

Section: Philadelphia-negative Mpnsmentioning

confidence: 64%

“…In addition to the possible involvement of the endothelial cell lineage in the process of clonal development of Ph − MPNs, several studies have explored the number of ECFCs in these disorders (Table 1). Basically, these studies have shown that ECFCs are increased in MF, but not in PV and ET (11,12). Furthermore, the levels of ECFCs are particularly increased in MF patients with high risk of splanchnic vein thrombosis (10) …”

Section: Philadelphia-negative Mpnsmentioning

confidence: 99%

Endothelial progenitor cells in hematologic malignancies

Testa

Saulle

Castelli

et al. 2016

Stem Cell Investig.

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Endothelial colony-forming cells from patients with chronic myeloproliferative disorders lack the disease-specific molecular clonality marker

Cited by 76 publications

References 22 publications

Vascular Endothelial Growth Factor Stimulates Endothelial Colony Forming Cells Proliferation and Tubulogenesis by Inducing Oscillations in Intracellular Ca2+ Concentration

Vascular Endothelial Growth Factor Stimulates Endothelial Colony Forming Cells Proliferation and Tubulogenesis by Inducing Oscillations in Intracellular Ca2+ Concentration

Endothelial progenitor cells are clonal and exhibit the JAK2V617F mutation in a subset of thrombotic patients with Ph-negative myeloproliferative neoplasms

Endothelial progenitor cells in hematologic malignancies

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