Platelets are well known for their powerful roles in vascular injury, inflammation and wound healing [1]. However, megakaryocytes, the precursor cells of platelets, have not been well known until recent decades. Megakaryocytes, a type of hematopoietic cell, undergo a series of complex processes to generate platelets, such as growth, development and maturation. At the same time, megakaryocytes possess certain cellular capabilities of immunity and inflammation [2]. In recent years, various experiments have mutually confirmed that megakaryocytes play an important role in the process of self-renewal and differentiation of HSC into multifunctional lineages ^[3][4][5][3,4,5]. In addition, as a type of intercellular communication cells, megakaryocytes also maintain the function of niches in the bone marrow [6]. Therefore, megakaryocytes play a very important role in the hematopoietic system.

Megakaryocytes release an average of 1 × 10¹¹ platelets into the blood each day. Megakaryocyte differentiation and maturation and platelet production are complex processes regulated by strict factors. This process is a hot topic in the field of hematopoiesis. In most clinical cases, thrombocytopenia is defined as a platelet count less than 100 × 10⁹/L, with clinical manifestations of splenomegaly, purpura, skin and mucous membrane hemorrhage, visceral hemorrhage, intracranial hemorrhage and even death [7]. Thrombocytopenia is a secondary complication of many diseases, such as myelodysplastic syndrome [8], leukemia [9], dengue fever [7], and parasitic infections [10]. In addition, myelosuppression caused by chemoradiotherapy also presents with a decreased platelet count [11]. To treat thrombocytopenia, platelet infusion, cytokines and TPO receptor agonists are commonly used in clinic [12]. However, platelet supply is scarce, the cost is high, the storage time of platelet is short, and it is easily contaminated by bacteria. Bloodborne infections may exist in the transfusion process, and homologous immune-related complications may also be caused. Cytokines stimulate platelet production by increasing the number of hematopoietic stem cells and blood cells. However, cytokines have many adverse reactions, including arrhythmia and acute pulmonary edema [13]. Thrombopoietin receptor agonists currently include first-generation recombinant human thrombopoietin (RhTPO) and second-generation agents (TPO-RAs), such as romiplostim and eltrombopag. Although they promote megakaryocyte differentiation and platelet production in vivo and increase the number of peripheral blood platelets, RhTPO may cross-react with endogenous TPO of patients to neutralize autoantibodies, which may result in immunogenic reactions, while romiplostim is the only TPO-RA that binds with endogenous TPO at the same site without resulting in an immunogenic reaction. However, there is a risk of thrombosis and up to 10% discontinue reaction. Both them require regular treatment and frequent monitoring. ^[14][15][16][14,15,16]. In recent years, ideas of platelet generation in vitro have been proposed, and many laboratories have successfully differentiated megakaryocytes into platelets in vitro. However, the large-scale production of platelets is still a challenging task in vitro [17].

Although the above treatment agents have significant efficacy, they have many adverse reactions and are limited in clinical application.

2. The Process of Megakaryocyte Differentiation

Megakaryocyte differentiation is a complex process in which hematopoietic stem cells differentiate into megakaryocytes, undergo terminal differentiation, and finally form proplatelets. In the classical process of hematopoietic stem cell self-renewal and directional differentiation, long-term hematopoietic stem cells differentiate into multiple multifunctional progenitor cells, then multiple multifunctional progenitor cells differentiate into myeloid progenitors and lymphoid progenitors, myeloid progenitors differentiate into megakaryocyte-erythroid progenitors and granulocyte-macrophage progenitors, and eventually differentiate into megakaryocyte progenitors and erythroid precursors ^[18][19]. Megakaryocyte progenitors eventually differentiate into megakaryocytes. After terminal differentiation and maturation, megakaryocytes eventually form proplatelets ^[19][20]. However, several recent studies have found that in addition to the ability to self-renew, long-term hematopoietic stem cells (LT-HSCs) can differentiate into short-term hematopoietic stem cells (ST-HSCs), which can differentiate into multiple multifunctional progenitors ^[20][21]. They are mainly divided into MPP1, MPP2, MPP3 and MPP4 (Figure 1). MPP1 is classified as a short-term HSC. MPP3 is largely granulocyte/macrophage-biased. MPP4 differentiates into common lymphoid progenitor cells (CLPs), which differentiate into T cells and B cells. MPP2 ^[21][22] cells are progenitor cells that tend to differentiate into common myeloid progenitors (CMPs). CMPs can differentiate into granulocyte-macrophage progenitors (GMPs) and megakaryocyte-erythroid progenitors (MEPs). MEPs differentiate into erythroid progenitors (EryPs) and megakaryocytic progenitors (MkPs). At the same time, MPP2 cells could skip the process from HSCs to CMPs and MEPs and directly differentiate into megakaryocytes. Another study reported that approximately 60% of hematopoietic stem cells are VWF+ cells in the bone marrow. In the hematopoietic microenvironment, because megakaryocytes generate platelets, they exist in vascular sinuses, and VWF + HSC have the same transcription factors and cytokines. Megakaryocytes are thought to be predisposed to induce VWF + HSC to differentiate into megakaryocytes and generate platelets [3]. Therefore, rwesearchers hypothesize that VWF+ cells have a specific tendency to differentiate into myeloid lineage cells under certain conditions. These two pathways can improve the efficiency of megakaryocytes differentiation in the hematopoietic lineage and provide an effective method for rapid platelet production in the hematopoietic system. After that, megakaryocytes will undergo multiple mitoses to undergo polyploid terminal differentiation and maturation, in which the largest polyploid nucleus reaches 128 N. Once megakaryocytes mature, they extend proplatelets into the vascular sinuses through the energy provided by the cytoskeleton and then release platelets through shear forces ^[22][23].