Megakaryocytic dysfunction in immune thrombocytopenia is linked to autophagy

Autophagy is a highly conserved biological process in eukaryotic cells. It is involved in cell development, starvation adaptation, intracellular quality control, tumour suppression, ageing, innate immunity, and other processes [1, 2]. However, both insufficient and excessive autophagy can lead to pathological conditions [3]. In recent years, experimental and clinical evidence has associated perturbations of normal autophagy processes with a number of neoplastic and autoimmune diseases [4], such as myelodysplastic syndrome (MDS) [5], chronic myelogenous leukaemia (CML) [6], systemic lupus erythematosus (SLE) [7], rheumatoid arthritis (RA) [8], multiple sclerosis (MS) [9], and aplastic anaemia (AA) [10]. In another autoimmune disease, immune thrombocytopenic purpura (ITP), autophagy plays an important role in maintaining the stemness and the microenvironment of haematopoietic stem cells [11]. Thus, on the one hand, autophagy ensures the proper differentiation of haematopoietic stem cells into megakaryocytes. On the other hand, at an early stage of megakaryocyte differentiation, induction of autophagy by inducer rapamycin or inhibitor bafilomycin A1 appears to impede megakaryocyte maturation, reduce platelet formation in bone marrow, and affect platelet function [12]. Further on, in mature megakaryocytes, autophagy deficiency induces abnormal platelet activation and function, without changing platelet number and size [13]. Accordingly, it appears that an abnormal level of autophagy causes different effects during distinct stages of cell differentiation [13]. Recently, autophagy has been demonstrated to be indispensable for normal megakaryopoiesis and platelet function in animal models with lineage-specific deletion of autophagy-related genes (ATGs) [14]. Excessive expression of mammalian target of rapamycin (mTOR) was reported in diseases related to megakaryocytes such as ITP, in which it inhibited autophagic activity and affected the differentiation of haematopoietic stem cells into megakaryocytes, the formation of megakaryocytes, and platelet function [15] Improving our understanding of autophagy will likely result in new therapeutic methods aimed at inducing autophagy-related proteins to counteract megakaryocyte/platelet disorders in clinical conditions. For example, induction of autophagy by rapamycin has already exhibited substantial therapeutic benefits in patients with ITP [16].

Autophagy, also called autophagocytosis, is a self-eating [17] and stress-induced catabolic process that delivers defective organelles and cytoplasm to the lysosome [18], and eventually forms the autolysosome. And this process also named the autophagy-lysosomal pathways (ALPs) [17]. The autophagy cytoplasmic quality control system supports the function and survival of different types of cells in most tissues of the body, for example, it provide the capability to quickly remove toxic waste and to repurpose unnecessary material [18]. Autophagy is upregulated in response to starvation, nitrogen deprivation, extra environmental and oxidative stresses, toxin and infection, DNA damage, and is downregulated in response to rapamycin treatment (Fig. 1). Based on different transport pathways and substrates in mammalian cells, autophagy can be divided into several distinct forms termed macroautophagy, microautophagy, chaperone-mediated autophagy (CMA) [19], mitophagy and aggrephagy [17], which are induced by similar stimuli and provide protection against most of diseases. Amongst these types, macroautophagy has been the most extensively studied. It is the major core of the ALPs and can be further divided into pexophagy, mitophagy, and non-selective autophagy [20].

At different stages of the autophagy pathway, a variety of ATG protein act as different function. For example, E1 enzyme ATG7 can activate autophagosome by two ubiquitin-like conjugated systems, UB1 protein ATG12 and ATG8 [21]. Briefly, autophagy consists essentially of a survival mechanism that is activated under adverse conditions to maintain cell integrity. However, paradoxically, it is also involved in a particular form of cell death called autophagic cell death or type II cell death [6].

Platelets are small anucleate cytoplasmic fragments derived from megakaryocytes, the primary physiological role is to mediate thrombosis and hemostasis [74]. Activated platelets secrete microparticles which accelerate the plaque formation by providing a new prothrombotic interface and promoting the deposition of fibrin and other blood cells at the site of thrombus formation [75, 76]. Ouseph et al. has demonstrated that basal level of autophagy process is essential for normal functioning of platelets activation and aggregation. And they further showed that platelet specific deletion of Atg7 can cause a decrease granule cargo packing ex vivo [77]. In another study, they demonstrate that starvation induced substantial autophagy (above basal level), characterized by decreased platelet aggregation, reduced calcium mobilization and granule secretion, as well as decreased adhesion to immobilized fibrinogen and eventually increased bleeding time [78]. Autophagy antagonize platelet activation by clearance certain potentially ubiquitinated proteins. These studies collectively demonstrated that basal level of autophagy in platelet is obligatory for aggregation, activation, hemostasis, and thrombosis [77, 78].

ITP is a common autoimmune-mediated bleeding disease, in which platelet membrane proteins become antigenic, stimulate the immune system to produce antibodies, and eventually result in thrombocytopenia [15]. About 50% of ITP patients have auto-antibodies that not only destroy platelets, but also impair megakaryocyte maturation and platelet production by the bone marrow [79]. In one study, a comparison of ITP mice and normal mice revealed that the former were characterised by higher frequencies of immature megakaryocytes/platelets and corresponding progenitor cells, as well as increased phagocytosis. These findings could explain the decrease in peripheral blood platelet counts observed in ITP patients [80]. McMillan and co-workers [79] previously described defective megakaryopoiesis in C-ITP (chronical immune thrombocytopenia) patients. They showed that, the addition of anti-platelet antibodies to normal megakaryocytes in liquid culture led to impaired megakaryocyte proliferation and abnormal ploidy distribution. Recently, Cao et al. [14] used an ATG7 haematopoietic conditional knockout mouse model to show that core autophagy machinery was important for normal megakaryopoiesis and platelet function. Thus, in ITP patients, the loss of autophagy prevents megakaryocyte formation and differentiation, negatively affects thrombopoiesis, and results in larger but fewer platelets, ultimately severely impairing platelet production [14]. Finally, Ouseph et al. [77] have demonstrated that autophagy is involved in the maturation of megakaryocytes and represents an important pathological condition in ITP patients (Fig. 2).

In this review, we summarized the limitations of current therapies and highlighted new treatments for ITP. The important role of autophagy in autoimmune diseases provides a new opportunity for understanding the pathogenesis of ITP. In particular, knowledge of the mechanism underlying abnormal autophagy in immature megakaryocytes may be important for the treatment of ITP patients. Similarly, autophagy induction may offer a novel therapeutic strategy against ITP or immune diseases, especially within the context of individualized treatment and disease control.

However, the study of Liu et al. demonstrated that the plasma in ITP patients induces autophagy and suppresses apoptosis [103]. And the inhibition of autophagy may be a novel treatment in further investigation. This conclusion deserves further research. For example, current results need more case validation analyses, functional verification assays and larger population-based studies to confirm [103]. And this conclusion is in contrast to our treatment of ITP autophagy induced by rapamycin. It could be interesting to explore this point and explained the role of autophagy in ITP as a balance between hyperactivity and inhibition in future.

In general, future research will benefit from focusing on targeted regulation of autophagy and related receptors, recognition mechanisms, and possible biomarkers. These will complement additional more in-depth studies on existing treatments for autophagy-based disorders.