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Neonatal Platelet Disorders
Key Points
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The most common cause of mild to moderate, early-onset thrombocytopenia in well-appearing neonates is placental insufficiency, frequently manifesting as small-for-gestational status at birth. This thrombocytopenia resolves spontaneously, usually within 10 days, and carries good prognosis. Thrombocytopenia in sick infants is usually associated with sepsis or necrotizing enterocolitis (NEC) and requires prompt intervention.
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Neonates with platelet counts <50 × 10 9 /L in the first day of life, particularly if well appearing, should be screened for neonatal alloimmune thrombocytopenia (NAIT). Random donor platelet transfusions (± IVIG) are the first line of therapy for these infants, unless HPA-1b1b and 5a5a platelets are immediately available for use (as is the case in some European countries, or if a prior child was affected). In those cases, these platelets are the preferred first line treatment.
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The risk of bleeding in thrombocytopenic neonates is multifactorial and is not related to the severity of the thrombocytopenia. Gestational age <28 weeks, postnatal age <10 days, and a diagnosis of NEC are more important predictors of bleeding than the platelet count.
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Historically, there has been significant variability in platelet transfusion thresholds used in the NICU. The recently published large multicenter PlaNeT-2 trial found a higher incidence of death or major bleeding among neonates randomized to receive platelet transfusions for platelet counts <50 × 10 3 /mcL compared to those transfused only for platelet counts <25 ×10 3 /mcL, supporting the hypothesis that platelet transfusions might be harmful to neonates.
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A subsequent risk stratification analysis of neonates enrolled in PlaNeT-2 demonstrated that infants with a high baseline risk of bleeding and mortality benefit from the low platelet transfusion threshold as much as (or more than) low-risk neonates.
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Most platelet function defects seen in the NICU are acquired, and are related to medications, medical conditions (i.e., uremia), or medical interventions (i.e., ECMO, therapeutic hypothermia). The most severe platelet function defects, which can present with bleeding in neonatal life, are Bernard-Soulier syndrome, caused by deficiency of glycoprotein Ib (the vWF receptor), and Glanzmann thrombasthenia, caused by a deficiency of glycoprotein IIb/IIIa (the fibrinogen receptor) on the platelet surface.
Fetal and Neonatal Platelet Production
A mounting body of evidence arising over the last decade has clearly demonstrated that there are substantial morphological and biological differences between fetal/neonatal and adult megakaryocytes and platelets. Developmental stage-specific differences are ontogenetically important because they allow the fetus to maintain stable platelet counts while the blood volume is rapidly expanding in a time characterized by exceptionally rapid growth. The complex process of platelet production can be represented as consisting of four main steps: (1) the production of thrombopoietic factors, mainly thrombopoietin (TPO), (2) the proliferation of megakaryocyte progenitors, (3) the differentiation and maturation of megakaryocytes through a unique process of endomitosis, and finally (4) the production and release of platelets into the circulation.
Studies culturing megakaryocyte progenitors derived from term and preterm umbilical cord blood, fetal blood (18 to 22 weeks’ gestation), or fetal bone marrow have documented that fetal/neonatal megakaryocyte progenitors proliferate at a significantly higher rate as compared to their adult counterparts. Additionally, fetal and neonatal megakaryocytes are substantially smaller and have lower ploidy levels than adult megakaryocytes. Contrary to adult megakaryocytes, which mature as their ploidy level increases, neonatal megakaryocytes are fully mature and capable of platelet production despite their small size and low ploidy. This dissociation between proliferation, polyploidization, and cytoplasmic maturation is a hallmark feature of neonatal megakaryopoiesis. The net result of this process is the production of large numbers of low-ploidy but highly mature megakaryocytes, with which fetuses and neonates populate their rapidly expanding bone marrow space and blood volume, while maintaining normal platelet counts. As developmental processes are easily disturbed, however, sick neonates, and particularly very low birth weight infants (birth weight <1500 g), are at high risk of thrombocytopenia.
Platelet Counts During Development and Reference Ranges
In 2009, Wiedmeier et al. published the largest study on neonatal platelet counts conducted to date, which included approximately 47,000 infants delivered between 22 and 42 weeks’ gestation. This study showed that platelet counts at birth increased with advancing gestational age ( Fig. 68.1 ) by approximately 2 × 10 9 /L for each week of gestation. Importantly, while the mean platelet count was ≥200 × 10 9 /L even in the most preterm infants, the 5th percentile was 104 × 10 9 /L for those ≤32 weeks’ gestation, and 123 × 10 9 /L for late-preterm and term neonates (see Fig. 68.1 ). These findings suggested that different definitions of thrombocytopenia should be applied to preterm infants. In that regard, however, it is important to emphasize that the reference ranges in that study were generated by eliminating the top 5% and the bottom 5% of all available values, rather than excluding values based on diagnoses. Thus, these should be considered “epidemiological reference ranges” rather than “normal ranges.” Nevertheless, these data suggest that platelet counts between 100 and 150 × 10 9 /L might be more frequent among otherwise healthy extremely preterm infants than among full term neonates or older children/adults.
Thrombocytopenia in neonates (as in adults) has traditionally been defined as a platelet count <150 × 10 9 /L and has been classified as mild (100 to 150 × 10 9 /L), moderate (50 to 99 × 10 9 /L), and severe (<50 × 10 9 /L). However, consistent with the data by Wiedmeier et al., platelet counts in the 100 to 149 × 10 9 /L range are more common among healthy neonates than adults. The incidence of thrombocytopenia in neonates varies significantly, depending on the population studied. Based on the traditional definitions, large studies in unselected populations of live-births (including healthy and sick neonates) established an overall incidence of neonatal thrombocytopenia of 0.7% to 0.9%. However, when focusing on neonates admitted to the NICU, the incidence of thrombocytopenia is much higher, ranging from 18% to 35%. The incidence of thrombocytopenia is also inversely correlated to the gestational age, so that the most immature neonates are the most frequently affected: platelet counts <150 × 10 9 /L were found at least once during the hospital stay in 70% of infants with a birth weight <1000 g.
Platelet Function and Primary Hemostasis
Multiple studies evaluating platelet adhesion, aggregation, and activation have shown that neonatal platelets are hyporesponsive in vitro to most agonists compared with adult platelets, and this low level of reactivity is more pronounced in preterm infants. Platelet aggregation studies demonstrated that platelets from neonatal (full term) cord blood were less responsive than adult platelets to agonists such as adenosine diphosphate (ADP), epinephrine, collagen, thrombin, and thromboxane analogues. Similar results were obtained in flow cytometric platelet activation studies, which showed decreased expression of surface activation markers in neonatal platelets stimulated with thrombin, ADP, and epinephrine. Different mechanisms account for the hyporeactivity of neonatal platelets to various agents: (1) the hyporesponsiveness to epinephrine is due to fewer α2-adrenergic receptors, the binding sites for epinephrine ; (2) the reduced response to collagen likely results from impaired calcium mobilization, although recent studies also showed a mildly reduced expression of GPVI (the collagen receptor) coupled with an intracellular signaling defect ; (3) the decreased response to thromboxane results from differences in signaling downstream from the receptor in neonatal platelets ; and (4) the decreased responsiveness to thrombin is related to reduced expression of PAR-1 and PAR-4 in neonatal platelets. Recently, developmental differences have also been described in regard to platelet inhibitory pathways, specifically a hypersensitivity of neonatal platelets to the inhibitory effects of prostaglandin E1 (PGE 1 ) during ADP- and collagen-induced platelet aggregation.
Surprisingly, while the hypofunctional platelet phenotype in vitro would predict a bleeding tendency, healthy full-term neonates have normal to enhanced primary hemostasis, compared to older children or adults. Bleeding times (BTs) in healthy term neonates are shorter than bleeding times in adults. Similarly, studies using the Platelet Function Analyzer (PFA-100, an in vitro test of primary hemostasis that measures the time it takes to occlude a small aperture, or Closure Time) found that cord blood samples from term neonates exhibited shorter closure times (CTs) than samples from older children or adults. The results of these studies suggest that there is an enhanced platelet/vessel wall interaction in full-term neonates, likely related to their higher hematocrits, higher mean corpuscular volumes, and higher concentrations of von Willebrand factors (particularly its ultralong polymers), all factors that, when combined, effectively counteract the neonatal platelet hyporeactivity. Taken together, the available evidence strongly suggests that the in vitro platelet hyporeactivity of healthy full-term infants is an integral part of a carefully balanced and well-functioning neonatal hemostatic system, rather than a developmental deficiency.
These compensatory mechanisms might be less well developed in preterm infants, whose platelets are also more hyporeactive than those of full-term infants, leading to longer BTs and therefore a less balanced and probably more vulnerable hemostatic system. Specifically, BTs performed on the first day of life were longer in preterm compared with term infants, with neonates <33 weeks’ gestation exhibiting the longest BTs. Saxonhouse et al. found that PFA-100 CTs from non-thrombocytopenic neonates were inversely correlated to gestational age in both cord blood and neonatal peripheral blood samples obtained on the first day of life. Importantly, however, while these BTs and CTs were longer in preterm compared to term neonates, they were still near or within the normal range for adults, suggesting that healthy preterm neonates also have adequate primary hemostasis. Data regarding how disease processes perturb this delicate system, particularly in the preterm neonate, are lacking.
In vitro studies using flow-cytometry or the cone and platelet analyzer showed that the neonatal platelet function improves significantly and nearly normalizes by 10 to 14 days, even in preterm infants. Consistent with this, Del Vecchio et al. found that, by day of life 10, all infants had shorter BTs than at birth, and early gestational age-related differences had disappeared. Moreover, little or no further shortening of BTs occurred between days 10 and 30. While no causal association has been demonstrated, this period overlaps with the period of highest risk of bleeding among preterm NICU patients, namely during the first 10 days of life.
Neonatal Thrombocytopenia
When evaluating a thrombocytopenic neonate, the first step to narrow the differential diagnosis is to classify the thrombocytopenia as either early onset (within the first 72 hours of life) or late onset (after 72 hours of life), and to determine whether the infant is clinically ill or well. Importantly, infection/sepsis should always be considered near the top of the differential diagnosis (regardless of the time of presentation and the infant’s appearance), as any delay in diagnosis and treatment can have life-threatening consequences.
Early-Onset Thrombocytopenia ( Fig. 68.2 , Table 68.1 )
The most frequent cause of early-onset thrombocytopenia in a well-appearing neonate is placental insufficiency, seen in infants born to mothers with pregnancy-induced hypertension/preeclampsia, and in those with intrauterine growth restriction (IUGR). This thrombocytopenia is always mild to moderate, presents immediately or shortly after birth, reaches a nadir on day of life 4, and resolves within 10 days. In a recent large cohort study, approximately one third of small for gestational age (SGA) infants had thrombocytopenia (<150 × 10 9 /L) during the first week of life, compared to only 10% of non-SGA gestational age-matched infants. This type of thrombocytopenia was associated with low mortality (2%), as long as there was no identified cause for the SGA other than placental insufficiency (i.e., genetic syndrome or congenital infection). If an SGA otherwise non-dysmorphic infant with mild to moderate thrombocytopenia remains clinically stable and the platelet count normalizes within 10 days, no further evaluation is necessary. However, if the thrombocytopenia becomes severe and/or persists >10 days, further investigation is indicated.
Table 68.1
Classification of Neonatal Thrombocytopenia By Time of Presentation
| Early-Onset Thrombocytopenia |
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| Late-Onset Thrombocytopenia |
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| Can Present Both Early and Late |
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ATRUS , Amegakaryocytic thrombocytopenia with proximal radio-ulnar synostosis; CMV , cytomegalovirus; DIC , disseminated intravascular coagulation; IUGR , intrauterine growth restriction; NAIT , neonatal alloimmune thrombocytopenia; NEC , necrotizing enterocolitis; TAR , thrombocytopenia-absent radii syndrome; TORCH , toxoplasmosis, other, rubella, cytomegalovirus, herpes simplex virus infection complex.
Severe early-onset thrombocytopenia in an otherwise healthy infant should trigger suspicion for an immune-mediated thrombocytopenia, either autoimmune (if the mother is also thrombocytopenic) or alloimmune (if the mother has a normal platelet count). These varieties of thrombocytopenia are discussed in detail below. Early-onset thrombocytopenia of any severity in an ill-appearing term or preterm neonate should prompt evaluation for sepsis, congenital viral or parasitic infections, or disseminated intravascular coagulation (DIC). DIC is most frequently associated with sepsis but can also be secondary to birth asphyxia.
In addition to these considerations, the affected neonate should be carefully examined for any radial and thumb abnormalities (suggestive of thrombocytopenia-absent radii [TAR] syndrome, amegakaryocytic thrombocytopenia with radio-ulnar synostosis, or Fanconi anemia). The inability to rotate the forearm on physical examination, in the presence of severe early-onset thrombocytopenia, suggests the rare diagnosis of congenital amegakaryocytic thrombocytopenia with proximal radio-ulnar synostosis (ATRUS). Dysmorphic features on physical exam should warrant investigation for other genetic disorders associated with early-onset thrombocytopenia, most commonly trisomy 21, trisomy 18, trisomy 13, Turner syndrome, Noonan syndrome, and Jacobsen syndrome.
The presence of hepato- and/or splenomegaly is suggestive of viral infection, although it can also be seen in hemophagocytic syndrome and liver disease from different etiologies. Other diagnoses, such as renal vein thrombosis, Kasabach-Merritt syndrome, and inborn errors of metabolism (mainly propionic acidemia and methylmalonic acidemia) should be considered and evaluated for based on specific clinical indications (i.e., hematuria in renal vein thrombosis, presence of a vascular tumor in Kasabach-Merritt syndrome).
Late-Onset Thrombocytopenia ( Fig. 68.3 , see Table 68.1 )
The most common causes of thrombocytopenia of any severity presenting after 72 hours of life are sepsis (bacterial or fungal) and necrotizing enterocolitis (NEC). Affected infants are usually ill-appearing and have other signs suggestive of sepsis and/or NEC. However, it is important to keep in mind that thrombocytopenia can be the first presenting sign of these processes and can precede clinical deterioration. Appropriate treatment (i.e., antibiotics, supportive respiratory and cardiovascular care, bowel rest in case of NEC, and surgery in case of surgical NEC) usually improves the platelet count in 1 to 2 weeks, although in some infants the thrombocytopenia persists for several weeks. The reasons underlying this prolonged thrombocytopenia are unclear.
If bacterial/fungal sepsis and NEC are ruled out, viral infections such as herpes simplex virus, cytomegalovirus (CMV), or enterovirus should be considered. These viral infections are frequently accompanied by abnormal liver enzymes. If the infant has or has recently had a central venous or arterial catheter, thromboses should be part of the differential diagnosis, although thromboses typically only cause thrombocytopenia if the thrombus is enlarging or is infected. Finally, drug-induced thrombocytopenia, while rare in neonates, should be considered if the infant is clinically well, other potential etiologies have been ruled out, and he/she is receiving heparin, antibiotics (penicillins, cephalosporins, metronidazole, vancomycin, or rifampin), indomethacin, famotidine, cimetidine, phenobarbital, or phenytoin, among others. Other less common causes of late-onset thrombocytopenia include inborn errors of metabolism and Fanconi anemia (rare).
Novel tools to evaluate platelet production, which aid in the evaluation of thrombocytopenia of unclear etiology, have been recently developed, and are likely to become widely available to clinicians in the near future. Among those, the immature platelet fraction (IPF) measures the percentage of newly released platelets (<24 hours). The IPF can be measured in a standard hematological cell counter (Sysmex 2100 XE Hematology Analyzer) as part of the complete cell count, and can help differentiate thrombocytopenias associated with decreased platelet production from those with increased platelet destruction, in a manner similar to the use of reticulocyte counts to evaluate anemia. Thus, an elevated IPF would suggest platelet consumption (as in neonatal alloimmune thrombocytopenia [NAIT] or DIC), while a decreased IPF would be consistent with a hyporegenerative thrombocytopenia, as in bone marrow suppression or failure.
Reference intervals for the IPF% and the absolute IPF (also known as the immature platelet count, or IPC) have been evaluated in healthy adults and full term neonates, using umbilical cord blood samples ( Table 68.2 ), as well as in nonthrombocytopenic NICU patients (IPF% = 4.1 ± 1.8). In the largest IPF study published to date, MacQueen et al. examined 24,372 platelet counts and IPFs from 9172 term and preterm neonates 0 to 90 days old. Data from nonthrombocytopenic infants in this cohort were used to generate age-specific reference intervals for IPF% and IPC (calculated as IPF% × platelet count) ( Fig. 68.4 ). As seen in the figure, the IPF at the time of birth was higher in preterm infants and decreased through gestation until 32 weeks, at which time it stabilized at full-term values (similar to adult values). Postnatally, the IPF increased progressively over the first 2 weeks of life and returned to baseline by 1 month in infants of all gestational ages. This study also assessed IPF percentages in neonates with thrombocytopenia, and found significantly higher values in neonates with consumptive etiologies compared to those with thrombocytopenia secondary to decreased production ( Table 68.3 ). Thus, when available, the IPF can help differentiate rare congenital thrombocytopenias (usually low) from NAIT or other consumptive disorders (usually elevated). Other studies have shown the usefulness of the IPF to predict platelet recovery in neonates. In patients with NEC and severe thrombocytopenia, a low absolute IPF has also been associated with a poor prognosis and high mortality.
Table 68.2
Reference Intervals for Platelet Counts and Immature Platelet Fractions (IPF) in Healthy Adults and Term Neonates (Umbilical Cord Blood)
A-IPF , Absolute immature platelet fraction; IPF , immature platelet fraction.
| Healthy Individuals | ||||
|---|---|---|---|---|
| Total ( n = 2152) | Men ( n = 1252) | Women ( n = 900) | Umbilical Cord Blood ( n = 133) | |
| Platelet counts (×10 9 /L) | ||||
| Reference interval | 162–347 | 161–338 | 164–360 | 191–392 |
| Lower limit (95% CI) | 160–164 | 158–164 | 160–169 | 168–208 |
| Upper limit (95% CI) | 340–353 | 326–344 | 351–372 | 364–447 |
| %-IPF (%) | ||||
| Reference interval | 0.5–3.3 | 0.5–3.1 | 0.5–3.4 | 0.7–3.8 |
| Lower limit (95% CI) | 0.5–0.5 | 0.5–0.6 | 0.5–0.5 | 0.7–0.9 |
| Upper limit (95% CI) | 3.2–3.4 | 3.0–3.3 | 3.3–3.5 | 3.0–3.8 |
| A-IPF (×10 9 /L) | ||||
| Reference interval | 1.25–7.02 | 1.30–6.80 | 1.21–7.15 | 1.94–9.69 |
| Lower limit (95% CI) | 1.19–1.30 | 1.20–1.41 | 1.10–1.27 | 1.66–2.58 |
| Upper limit (95% CI) | 6.75–7.24 | 649–7.16 | 6.9–7.48 | 7.96–10.57 |
From Ko YJ, Kim H, Hur M, et al. Establishment of reference interval for immature platelet fraction. Int J Lab Hematol . 2013;35(5):528–533.
Table 68.3
Immature Platelet Fraction Values in Neonates With Thrombocytopenia of Different Etiologies
From MacQueen BC, Christensen RD, Henry E, et al. The immature platelet fraction: creating neonatal reference intervals and using these to categorize neonatal thrombocytopenias. J Perinatol . 2017;37(7):836.
| Mechanism of Thrombocytopenia | N | IPF% |
|---|---|---|
| Hypoproliferative * | 92 | 10.4 ± 2.9 |
| Consumptive ** | 98 | 20.9 ± 7.9 |
| Both | 76 | 17.9 ± 5.9 |
| Indeterminate *** | 14 | 12.8 ± 8.1 |
* Small for gestational age, birth asphyxia, or a syndrome associated with hypoproliferative thrombocytopenia.
** Immune-mediated, necrotizing enterocolitis (NEC), sepsis, or disseminated intravascular coagulation (DIC).
Immune Thrombocytopenia
Immune thrombocytopenia occurs due to the passive transfer of antibodies from the maternal to the fetal circulation. There are two distinct types of immune-mediated thrombocytopenia: (1) NAIT and (2) autoimmune thrombocytopenia.
Neonatal Alloimmune Thrombocytopenia
Epidemiology
NAIT is the most common underlying cause of early-onset severe thrombocytopenia, with an incidence among liveborn neonates of 0.5 to 1.5 per 1000 births. The true incidence of the disease is likely higher, however, since the milder cases might go undetected and the most severe cases lead to intrauterine death. Intrauterine death or intracranial hemorrhage (ICH) may occur as early as at 14 to 16 weeks of gestation, resulting in a relatively high incidence of intrauterine ICH (>10%). The overall incidence of ICH (prenatal and postnatal) is particularly high in this population, affecting up to 20% of infants with NAIT and potentially leading to lifelong consequences. ICH may occur during the first pregnancy and has a recurrence risk close to 100% in subsequent pregnancies in the absence of prenatal treatment.
Pathophysiology
In NAIT, the antibody is produced in the mother against a specific human platelet antigen (HPA) present in the fetus but absent in the mother. The antigen is inherited from the father of the fetus. The antigens responsible for NAIT are caused by single nucleotide polymorphisms on any of the main glycoproteins located on the platelet surface, particularly GPIIb/IIIa. The first platelet antigen was identified in 1959 by von Loghem et al. and was designated Zw-a (later PLA1). The initial nomenclature for these antigens came from the name of the patients, leading to confusion in the field. In 1990, a simplified system for HPA nomenclature was described, in which each antigen was given a HPA number. Antigens were numbered chronologically, according to the date of their initial report. The bi-allelic antigens were given an alphabetic designation of “a” or “b” in the order of their frequency (higher frequency for “a”). Thus, the Zw-a/PLA1 antigen was named HPA-1, with its two serological forms designated as HPA-1a for the common form, and HPA-1b for the less common form (the latter corresponding to PLA2). Currently, there are at least 33 HPA antigens identified. The frequency of each antigen varies within ethnic groups: in Caucasians, antibodies to HPA-1a are the major cause of NAIT, followed by HPA-5a and, less frequently, HPA-9b, HPA-3a and b, and HPA-15. Antibodies to HPA-4b are the predominant cause of NAIT in the Japanese population.
The anti-HPA antibody produced in the maternal serum crosses the placenta and reaches the fetal circulation, leading to platelet destruction, apoptosis of early megakaryocyte progenitors (therefore decreased platelet production), and thrombocytopenia.
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