Platelet factor 4 regulates megakaryopoiesis through low-density lipoprotein receptor–related protein 1 (LRP1) on megakaryocytes

Abstract

Platelet factor 4 (PF4) is a negative regulator of megakaryopoiesis, but its mechanism of action had not been addressed. Low-density lipoprotein (LDL) receptorrelated protein-1 (LRP1) has been shown to mediate endothelial cell responses to PF4 and so we tested this receptor's importance in PF4's role in megakaryopoiesis. We found that LRP1 is absent from megakaryocyte-erythrocyte progenitor cells, is maximally present on large, polyploidy megakaryocytes, and near absent on platelets. Blocking LRP1 with either receptor-associated protein (RAP), an antagonist of LDL family member receptors, or specific anti-LRP1 antibodies reversed the inhibition of megakaryocyte colony growth by PF4. In addition, using shRNA to reduce LRP1 expression was able to restore megakaryocyte colony formation in bone marrow isolated from human PF4-overexpressing mice (hPF4 High ). Further, shRNA knockdown of LRP1 expression was able to limit the effects of PF4 on megakaryopoiesis. Finally, infusion of RAP into hPF4 High mice was able to increase baseline platelet counts without affecting other lineages, suggesting that this mechanism is important in vivo. These studies extend our understanding of PF4's negative paracrine effect in megakaryopoiesis and its potential clinical implications as well as provide insights into the biology of LRP1, which is transiently expressed during megakaryopoiesis. (Blood. 2009;114:2290-2298) IntroductionAlthough the predominant cytokine regulating platelet count is thrombopoietin (TPO), during megakaryopoiesis, many other cytokines have been implicated, including interleukin-6 (IL-6), which increases TPO expression in the liver 1 ; stromal-derived factor-1, which enhances megakaryocyte chemotaxis 2 ; and IL-11, which directly stimulates megakaryocyte development. 3 A pathway by which megakaryopoiesis is auto-down-regulated has been suggested based on in vitro studies of platelet factor 4 (PF4) and later by studies of other chemokines that are also stored in ␣-granules, including the related CXC chemokines, neutrophil activating peptide-2 and IL-8, 4,5 and the more distantly related CC chemokines, regulated upon activation, normal T-cell expressed and secreted 6 and macrophage inflammatory peptide-1␣. 5,6 More recently, in vivo studies have demonstrated the importance of the PF4-negative paracrine loop under steady-state conditions and in chemotherapy-induced thrombocytopenia (CIT). 7 PF4 is a 7.8-kDa protein that is produced primarily in megakaryocytes, expressed in platelets as a tetramer, and comprises 2.5% on a molar basis of the ␣-granular releasate. 8 The biologic role(s) of PF4 is not fully understood. In addition to previous in vitro studies demonstrating an effect on megakaryocyte development, we have recently shown that PF4 can play a biologically relevant role in vivo in regulation of steady-state platelet count and in recovery after chemotherapy. 7 Unlike other chemokines that have clearly defined chemokine receptors, PF4 appears to function by binding with high affinity to glycosaminoglycans (GAG...

Downregulation of GATA1 drives impaired hematopoiesis in primary myelofibrosis

“…Figures 4 and 5). Genes that were downregulated included several that are associated with megakaryocyte maturation, such as platelet factor 4 (PF4) (12). Interestingly, as previously reported (9), we observed that GATA1 mRNA was modestly elevated or unchanged relative to control megakaryocytes ( Figure 2A).…”

Platelet factor 4 induces cell apoptosis by inhibition of STAT3 via up-regulation of SOCS3 expression in multiple myeloma

“…24,25 We first checked the expression of these receptors and found that both CXCR3B and LRP1 were expressed in the MM cell lines U266, OPM2 and NCI-H929 (data not shown). To investigate whether PF4-induced apoptosis is mediated by these receptors, we suppressed their expression in MM cells to see whether the pro-apoptotic effect of PF4 would be abolished.…”

“…24,25 We, therefore, examined whether these receptors mediated PF4 effects on apoptosis of MM cells. Knockdown of LRP1 expression, but not of CXCR3B, abolished the pro-apoptotic effect of PF4.…”

Vasoactive intestinal polypeptide suppresses proliferation of human cord blood-derived hematopoietic progenitor cells by increasing TNF-α and TGF-β production in the liver

“…This result is in accordance with those described in a previous report (Rameshwar et al, 2002) showing that VIP inhibited the proliferation of bone marrow progenitors. Although cytokines are known to be very important molecules affecting the proliferation of HSCs/HPs and are considered to be regulators of HSCs/HPs, few cytokines have been reported to inhibit hematopoiesis, such as TNF-α, TGF-β, and PF4 (Lambert et al, 2009;Sharma et al, 2011;Wang et al, 2012). In this study, we not only confirmed the inhibitory effect of VIP on the amplification of CD34 + cells purified from human cord blood samples, but also revealed that VIP upregulated the intrinsic levels of TNF-α and TGF-β 1 in CD34 + cells.…”