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Inherited thrombocytopenias
were unusually round and had altered tubulin and actin lev- els with DIAPH1 being one of the affected genes.
Finally, mention must be made of IKZF5 encoding a tran- scription factor known as Pegasus that was highlighted by whole genome sequencing of over 13,000 individuals enrolled in the National Institute for Health Research Bioresource and including 233 cases of isolated thrombocy- topenia.102 Various causal missense variants were detected in patients with isolated thrombocytopenia but mild or no bleeding; the mutations (sometimes de novo) were proposed to interfere with DNA binding and to selectively alter the expression of a series of proteins expressed in megakary- ocytes and platelets. The thrombocytopenia was mild and platelet size was normal. The platelet a-granule content was low but the platelet aggregation response was reported as inconsistent. The megakaryocytes produced fewer pro- platelets.
Thrombocytopenia, apoptosis and deficient glycosylation
Early apoptosis with phagocytosis of platelets by macrophages in the spleen or glycosylation changes fol- lowed by platelet clearance in the liver are potential causes of inherited thrombocytopenia. Although evidence for such mechanisms remains incomplete, the identification of some mutations, largely by NGS, has revived interest, particularly in deficient glycosylation.
Apoptosis
Non-syndromic mild AD thrombocytopenia but with lit- tle or no bleeding and platelets of normal size is associated with mutations in the CYCS gene encoding mitochondrial cytochrome c, a disease first reported in New Zealand.103,104 Although normal-looking megakaryocytes were present in the bone marrow, the number of naked nuclei was increased. Their presence as well as platelets in the marrow space was indicative of ectopic release. Electron microscopy revealed platelets lacking glycogen stores and the micro- tubule coil. Culture of megakaryocytes from the patients showed that abnormal and normal platelets were both released. Whether or not CYCS mutations, so far limited to three families, lead to loss of mitochondrial inner mem- brane potential and platelet phosphatidylserine expression remains speculative. Mention must also be made of a mis- sense p.E167D mutation in MASTL, coding for a micro- tubule-associated Ser/Thr kinase, in a large family first reported many years previously. The mutation was linked to moderate thrombocytopenia (with normal sized platelets), incomplete differentiation of high ploidy megakaryocytes and a propensity to mild bleeding.58,105 Recent studies on a mouse knock-in model suggest that this is a gain-of-function mutation leading to platelet survival changes and to platelets that extend unusually long pseudopods and contain hyper-stabilized microtubules.106 In mice, Mastl inhibits protein phosphatase 2, an important negative regulator of several major platelet activation path- ways. The result is an increased propensity of platelets to form aggregates, reduced platelet survival and increased signs of apoptosis.
Desialylation and other glycosylation changes
An early study showed macrothrombocytopenia and
neutropenia in a patient with a specific defect in a-2,3-sia- lylation and rapid platelet clearance by the Ashwell- Morell receptor in the liver but the gene defect was not identified.107 More recent studies using WES have linked macrothrombocytopenia with AR mutations of GNE encoding UDP-N-acetylglucosamine 2-epimerase/N- acetylmannosamine kinase, a bi-functional enzyme essen- tial for N-acetylneuraminic acid synthesis (the principle sialic acid of human platelets).108-110 Interestingly GNE mutations are also a cause of myopathy and in the above studies macrothrombocytopenia was either syndromic or isolated depending on the report. Severe menorrhagia and bleeding into the corpus luteum in affected women were noteworthy. Platelets were large and the numbers of retic- ulated (young) platelets often increased, features suggest- ing accelerated platelet clearance. Precise quantification of the platelet sialic acid deficiency and a direct measure of platelet survival have not, however, been made.
A necessary step in the synthesis of oligosaccharides is the transport of cytidine monophosphate-bound sialic acid to the Golgi apparatus. This requires a transporter protein SLC35A1, whose molecular deficiency was recently reported in two siblings with moderate thrombo- cytopenia, giant platelets, and mild bleeding.111 The macrothrombocytopenia is syndromic, featuring delayed psychomotor development, epilepsy, ataxia, microcephaly and choreiform movements. Exome sequencing highlight- ed a homozygous missense mutation in SLC35A1 in both siblings; their consanguineous parents were heterozygous. Lectin binding confirmed abnormally high b-galactose exposure on platelets from the patients and increased numbers of reticulated platelets. Bone marrow biopsies showed a high percentage of immature megakaryocytes, yet in culture megakaryocytes from the patients showed normal maturation and proplatelet formation. However quantification of platelet survival using a non-obese dia- betic/severe combined immunodeficiency mouse model confirmed that the patients’ platelets had a very short lifespan. In a separate study, severe thrombocytopenia with enlarged platelets and dysplastic megakaryocytes was the characteristic of six members of a consanguineous family in whom WES and homozygosity mapping revealed a homozygous mutation in GALE, encoding UDP-galactose-4-epimerase, an enzyme responsible for UDP-galactose inter-conversion with UDP-glucose and UDP-N-acetylgalactosamine inter-conversion with UDP- N-acetylglucosamine.112 Some individuals also showed mild anemia and febrile neutropenia while galactosemia is a feature of mutations of this gene. With predicted protein instability, the enzyme retained 40% of its normal activi- ty. Knockdown of GALE in hematopoietic cells slowed the proliferation of these cells in liquid culture; larger colonies possibly reflected an increased presence of imma- ture cells.
Macrothrombocytopenia and sitosterolemia
An unusual inherited macrothrombocytopenia with epi- genetic or environmental implications is sitosterolemia. In this condition, giant platelets associate with AR mutations in ABCG5 and ABCG8, genes which code for proteins that act as cellular transporters of plant sterols, the latter accu- mulating at high levels in blood and tissues.113 Xanthomas, premature atherosclerosis and hemolytic anemia are other
haematologica | 2020; 105(8)
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