Inhibition of Megakaryocyte Development in the Bone Marrow Underlies Dengue Virus-Induced Thrombocytopenia in Humanized Mice

Abstract

A characteristic clinical feature of dengue virus infection is thrombocytopenia, though its underlying mechanism is not definitively determined. By adoptive transfer of human CD34؉ fetal liver cells into immunodeficient mice, we have constructed humanized mice with significant levels of human platelets, monocytes/macrophages, and hepatocytes. Infection of these mice with both lab-adapted and clinical strains of dengue virus induces characteristic human hematological changes, including transient leukopenia and thrombocytopenia. We show that the specific depletion of human platelets is not mediated by antibodies in the periphery or reduced production of human thrombopoietin in the liver but reduction of human megakaryocytes and megakaryocyte progenitors in the bone marrow of the infected mice. These findings identify inhibition of platelet production in the bone marrow as a key mechanism underlying dengue-induced thrombocytopenia and suggest the utility of the improved humanized mouse model in studying dengue virus infection and pathogenesis in a human cell context. Dengue is an acute febrile illness caused by dengue virus (DENV), which is spread through mosquito vectors. Dengue manifests in a wide range of clinical symptoms and is usually accompanied by hematological changes, such as leukopenia and thrombocytopenia in mild cases and plasma leakage, hemorrhage, or organ impairment, such as liver damage, in severe cases (1). Even after decades of research, the cause of thrombocytopenia or platelet drop during dengue disease is still unclear.Platelets are small (2 to 3 m), anucleated cells which play a major role in homeostasis and coagulation. They are derived from megakaryocytes, which are large (30-to 100-m), nucleated, polyploid cells. Megakaryocytes differentiate from hematopoietic stem cells in the bone marrow (BM) (2, 3) through progenitor cells known as megakaryocytic CFU (CFU-MK) that express CD34 and CD41 (4). Thrombopoietin (TPO) is the principal cytokine that drives the expansion and differentiation of the progenitors to megakaryocytes (3, 5) and is produced mainly by hepatocytes in the liver (6).Two hypotheses have been proposed to explain thrombocytopenia during DENV infection: clearance of platelets from periphery and loss of platelet production in the BM. The peripheral mechanism is thought to involve antibody-mediated depletion where the antibody-opsonized DENV binds to platelets, which are then cleared by activated immune cells (7,8). Another possibility is that antibodies to the viral nonstructural protein (NS1) crossreact with autoantigens expressed on platelets, which binds and tags them for clearance (9, 10). Alternatively, platelet production in the BM could be suppressed, although the evidence supporting this hypothesis is lacking due to difficulties in obtaining BM biopsy specimens from acute dengue patients. One report shows that the BM was hypocellular early during dengue disease but later became hypercellular, as if recovered from acute suppression (11). There is also evidence that DENV i...

Utility of Humanized BLT Mice for Analysis of Dengue Virus Infection and Antiviral Drug Testing

“…Consistent with previous studies (21,32), we also observed a significant decrease in circulating total platelets, i.e., thrombocytopenia, another hallmark of DENV infection in humans. Thrombocytopenia is fre- quently observed among dengue fever patients, and two mechanisms have been proposed to explain it: antibody-mediated depletion of platelets from peripheral blood (45)(46)(47)(48)(49) and deficient platelet production by megakaryocytes in the bone marrow (25,50). The latter hypothesis is supported by a study that demonstrated that human megakaryocytes from the bone marrow were highly permissive for DENV infection in vitro, suggesting that megakaryocytes may also be targeted in vivo, thus providing a plausible mechanism for thrombocytopenia during acute infection in humans (50).…”

Mountain high and valley deep: epigenetic controls of pluripotency and cell fate

“…In particular, its efficiency remains low (Mikkelsen et al, 2008;Pasque et al, 2012;Gaspar-Maia et al, 2013;Sridharan et al, 2013;Nashun et al, 2015), the extreme stability of adult somatic cell epigenetic signature makes iPSCs prone to errors (Plath and Lowry, 2011), and the use of DNA constructs and the subsequent possibility of exogenous sequence integration preclude their clinical use for safety concerns (Stadtfeld et al, 2008;Kim et al, 2009;Zhou and Freed, 2009;Seki et al, 2010). In order to circumvent these limits, smallmolecule compounds have been used to modulate the epigenetic state by inhibiting and/or activating, in a reversible way, specific signaling pathways (Huangfu et al, 2008;Ichida et al, 2009;Li et al, 2011;Hou et al, 2013).…”

Viral infections in mice with reconstituted human immune system components

“…Dengue virus infection was performed by intraperitoneal, subcutaneous of intravenous injection into HIS mice [62][63][64][65][66]. In most instances dengue serotype 2 viruses were used.…”

“…Dengue virus infection was performed by intraperitoneal, subcutaneous of intravenous injection into HIS mice [62][63][64][65][66]. In most instances dengue serotype however, is dengue hemorrhagic fever after heterologous infection with another dengue virus serotype and it is thought that heterologous antibodies enhance infection by the second phenotype to cause this severe disease [67].…”