The platelet threshold necessary to make diagnosis of thrombocytopenia has changed during years (1), and for immune thrombocytopenia (ITP) working group thrombocytopenia is defined as a condition characterized by a platelet count lower than 100,000 platelets per microliter (2).

Primary ITP is the most common cause of thrombocytopenia, having an estimated incidence of 4–9 cases out of 100,000 people per year (3–6), with about half of the pediatric cases occurring in previously healthy children.

The first part of this review described the pathogenesis of the disease, with special attention to the role of innate immune system and impairment in megakaryopoiesis (7). In this second part, we focus on the clinical aspects of ITP, particularly on the differential diagnosis and new target therapies. The process of differential diagnosis has the aim to distinguish ITP from secondary IT (caused by infections, immune defects, and other pathologies) and non-IT, particularly inherited diseases, because all these conditions require different treatments.

Therapy of ITP was historically based on the progressive use of immunoglobulins, corticosteroids, and immunosuppressive agents: following the recent advances in the study of the pathogenesis of the disease, new therapeutic targets have been identified, potentially leading to innovative therapeutic strategies.

Immune thrombocytopenia can be classified according to etiology, disease evolution and age of onset. Etiologic classification will be discussed separately, to introduce the more common forms of secondary IT.

According to disease evolution, it is possible to identify three categories of ITP: newly diagnosed ITP; persistent ITP, still present after 3 months from diagnosis; and chronic ITP, lasting 12 or more months after diagnosis (2), which represents about 20% of the total cases of ITP in childhood (8).

IT affecting young children is typically acute and self-remitting, and primary forms are the most common in this age. Adolescents IT has an intermediate phenotype between childhood-onset and adult-onset forms (9), showing a higher rate of chronicity and a greater percentage of secondary IT.

Etiologic classification divides two categories of ITP: primary ITP and secondary IT.

The primary form of IT, classically defined “idiopathic,” is often seen in childhood and triggered by non-specific viral infections (upper respiratory or gastrointestinal infections): in some cases acute infections by Epstein–Barr virus, cytomegalovirus, parvovirus, rubella, mumps, and varicella have been identified as triggers of ITP (10–12).

Secondary IT has a complex etiology, as specific infections, drugs or vaccinations and immunologic abnormalities, including immunodeficiencies, can be involved in its pathogenesis.

Other forms of thrombocytopenia occurring during childhood and adolescence could mime ITP and secondary IT, particularly when platelet reduction is the only laboratory finding. Inherited thrombocytopenias, often misdiagnosed as ITP (38), are characterized by impairment in magakaryopoiesis and include a large variety of X-linked and autosomal diseases, commonly presenting with altered platelet size (39). Among them, Wiskott–Aldrich syndrome, caused by the mutation of WAS gene on chromosome X, usually comprehends the association of thrombocytopenia with small platelets, eczema, and immunodeficiency (40).

Thrombocytopenia occurs also in acute leukemia and primary bone marrow failure syndromes as Fanconi anemia, but in these cases, the association with other cytopenias helps making differential diagnosis (41).

Also lisosomial storage disorders, as Gaucher’s and Niemann–Pick’s disease, may present thrombocytopenia at clinical onset, usually accompanied by a considerable splenomegaly (42, 43).

Other less common conditions are indicated in Table 1, which summarizes the most relevant causes of secondary IT and non-IT in childhood and adolescence.

Immune thrombocytopenia is not rarely asymptomatic, being observed during routinely laboratory evaluations. In symptomatic cases, the most common presenting feature is epistaxis (51), followed by cutaneous and mucosal minor bleeding.

Severe bleeding rates are more common in childhood compared to adult patients: a review by Neunert et al. showed that severe bleeding occurs in 20.2% of children and 9.6% of adults (52).

The most severe bleeding event is ITP-associated intracranial hemorrhage (ICH): it represents a rare cause of pediatric stroke (53), affecting only 0.4% of children with ITP (52), but complicated by elevated mortality rates, reported between 12 and 25% in recent studies (54–58). ICH, as other severe bleedings, occurs mostly in children with other features of bleeding and in patients with a platelet count lower than 10,000/μl (59) and is often preceded by a precipitant factor, like head trauma (55, 60, 61).

An anamnesis positive for previous minor bleeding is a risk factor for severe haemorrhagic events (52), while the clinical relevance of an occult hemorrhage, often identified in urinary tract, is not completely defined, as studies reported different conclusions about the association with future overt bleeding and ICH (55, 62).

Diagnosis of ITP remains one of exclusion, and differential diagnosis with secondary forms is crucial, because in these cases thrombocytopenia may be less responsive to conventional therapy, but only to the treatment of primary cause.

Interview should focus on the potential triggers of ITP (assumption of drugs, vaccines, and transfusions) and risk factors for secondary forms, as the presence of weight loss, chronic infections (HIV and HCV) and other immune-mediated disease. It is also important to investigate elements suggestive of inherited thrombocytopenia, as previous bleedings and positive familiar history.

During physical assessment, the clinician must search potential sites of bleeding (cutaneous and mucosal) and identify signs suggestive for secondary IT or other pathologies, by examining the presence of hepatosplenomegaly, abdominal masses, lymphoadenopathies, and bone pain.

Furthermore, we analyze the diagnostic the most relevant laboratory investigations in ITP, to introduce our diagnostic algorithm.

The algorithm we suggest for differential diagnosis of new onset thrombocytopenia, shown in Figure 1, is composed by one clinical step and three laboratory and radiological steps and is primarily directed to patients without acute and severe bleeding. We focused particularly on the exclusion of short-term life-threatening conditions, such as acute leukemia, lymphomas, and other neoplasia, while chronic infections and systemic autoimmune disease with partial expression are investigated in later steps. The algorithm is progressive, and therefore the investigations included in steps 3 and 4 are usually indicated only in patients with persistent and chronic ITP.

The clinical step remains fundamental: in case of severe bleeding signs, it is mandatory to treat the patient, and the required investigations are different (see Consideration on Patients with Acute Bleeding). Moreover, if anamnesis or physical assessment shows elements indicative for secondary IT (abdominal masses and adenopathies), the algorithm becomes not necessary, and the laboratory and radiological approach must start with investigations directed to confirm the etiologic hypothesis suggested by the clinical findings.

The second step comprehends laboratory exams directed to identify the conditions that more frequently cause secondary IT and non-ITP, including inherited thrombocytopenia, infections, immunodeficiency, and lymphoid malignancies.

The third step includes an abdominal echography, useful to recognize alterations in liver, spleen, and abdominal lymph nodes, not always appreciable during clinical examination. This step also considers the determination of reticulated platelet count: despite the lack of standardization of values and difficulties in interpretations, this investigation, when available, can give important information about thrombopoietic rate, and thus remains an option to consider.

The last step comprehends investigations for autoimmune diseases and chronic infections. Determination of ANA is also important to predict the evolution to a chronic form (see Prognosis and Sequelae).

About two out of three pediatric patients with ITP show a spontaneous improvement in platelet count in 6 months without necessity of medical treatment, and those remissions are usually sustained. Most of patients with newly diagnosed ITP do not show signs of bleeding, and can be managed with a “watch and see” strategy (83–86).

There is no absolute consensus about the platelet threshold necessary to start treatment in ITP: 1996 guidelines of the American Society of Hematology recommended to treat patients with a platelet count lower than 10,000/μl and minor purpura or those one with a count lower 20,000/μl and significant bleeding (87). An update published in 2011 suggested that children without bleeding or with mild bleeding should be managed only with observations, regardless of platelet count (88). Despite these recommendations, most patients with low risk of bleeding are currently treated (89).

Immune thrombocytopenia in childhood is usually self-remitting, while there is the possibility of reactivation of the disease following viral infections or other triggers. The major sequelae of acute episodes are represented by permanent neurologic damages, defined as epilepsy, cognitive and learning disorder, and paresis, that can be detected in patients surviving the acute event of ICH (55, 148).

Adolescents are more likely to develop a chronic disease: a recent work by Heitink-Pollè et al. identified some predictors of chronic ITP, including female sex, age >11, insidious onset, absence of a trigger (infection or vaccination), and ANA positivity (149).

Patients with chronic ITP can undercover secondary misdiagnosed forms, as chronic infections, CVID, or other autoimmune pathologies, suggesting that this subset of patients need continuous reevaluation and discussion of primary diagnosis. Moreover, rarely, these patients may show an evolution to SLE: since there are no specific predictors of this progression (150), a study by Panzer et al. suggested that combined assessment of ANA and anti-DsDNA may have a role in identifying subjects at higher risk (151), who need to be periodically monitored.

The centenary of ITP traced history characterized of new progressive knowledge, making it a paradigmatic model of autoimmune disease. In this issue, a second part of a revision of ITP story, we focused on ITP diagnostic approach. By combining physical examinations and laboratory findings, we designed a diagnostic algorithm, to dissect the complex diagnosis, substantially based on the exclusion of the multiple possible concurrent causes of thrombocytopenia. We described conventional therapy of ITP and focused on the new therapeutic targets.

All the authors contributed to the work presented in this paper, wrote and reviewed the paper, and provided approval of the final version.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.