Megakaryocytes directly regulate HSC pool size, proliferation via CXCL4

NEW ORLEANS — The location of a subset of hematopoietic stem cells in the vicinity of megakaryocytes is nonrandom, according to study findings presented at the ASH Annual Meeting and Exposition.

Researchers set out to assess the functional role of midkine in hematopoietic stem cell regulation in vivo by breeding PF4-cre mice with iDTR mice. After 7 days of delirium tremens treatment, researchers observed a 5.3-fold decrease in bone marrow midkine depletion (P<.001) and an associated 7-fold decrease in platelets (P<.0001). Depletion of midkine was associated with an 11.5-fold increase (P<.001) in the number of Lin-c-Kit+Sca-1+CD105+CD150+ HSC as the result of a 5.5-fold increase in proliferation (P<.001).

Researchers then assessed whether the increase in hematopoietic stem cell proliferation after midkine depletion affected hematopoietic stem cell function by competitive transplantations in limiting dilutions.

There were 625 hematopoietic stem cells with repopulating capacity per femur in delirium tremens-treated PF4-cre:iDTR mice — a 4.8-fold increase (P<.01) from delirium tremens-treated PF4-cre transgenic control mice. Midkine depletion did not end in aberrant hematopoiesis.

To then assess whether the effect was mediated by circulating platelets rather than bone marrow midkine, researchers injected mice with neuraminidase. A 13.9-fold decrease in blood platelets (P<.0001) was observed after neuraminidase treatment. However, neither bone marrow midkine nor hematopoietic stem cell were significantly changed; therefore, midkine regulates bone marrow hematopoietic stem cell quiescence, according to researchers.

As previously reported in the ELISA study, CXCL4 — expressed exclusively by midkine — has the greatest reduction in the bone marrow extracellular fluid upon midkine depletion. Researchers therefore hypothesized that PF4 contributed to midkine-mediated maintenance of hematopoietic stem cell quiescence.

In vitro culture experiments led researchers to observe that recombinant murine PF4 restrained hematopoietic stem cell proliferation in serum-free culture conditions (3.1-fold decrease; P<.01). Similarly co-cultures of hematopoietic stem cell with wild-type midkine, but not CXCL4-deficient Midkine, reduced hematopoietic stem cell proliferation.

In vivo, after 1 week of rmPF4 injections, a 1.5-fold reduction in hematopoietic stem cell pool was observed (P<.0002). Further, 16 weeks after transplant with bone marrow cells from PF4-treated mice, researchers observed a significant decrease in the percentage of donor-derived cells (P<.01).

When the researchers then assessed hematopoietic stem cell numbers in Pf4-/- mice vs. WT controls, Pf4-/- mice had increased hematopoietic stem cell numbers (1.5-fold; P<.01) and enhanced proliferation (1.6-fold; P<.05).

Ingmar Bruns

“Megakaryoctes, as an hematopoietic stem cell progeny, seem to control its own replenishment via a direct feedback loop, suggesting the existence of a novel hematopoietic stem cell niche formed by a cell of hematopoietic origin,” Ingmar Bruns, MD, PhD, of the Albert Einstein College of Medicine, told HemOnc Today. “These findings may add to the current pathophysiological understanding of hematological diseases that are associated with altered hematopoietic stem cell behavior and aberrant midkine numbers such as myeloproliferative diseases or myelodysplastic syndromes.”

For more information:

Bruns I. Abstract #3. Presented at: ASH Annual Meeting and Exhibition; Dec. 7-10, 2013; New Orleans.

Disclosure: The researchers report research funding from Amgen and GlaxoSmithKline.