Transcervical thymic biopsy in the immunodeficient child

With the neck in a fully extended position, a horizontal incision is made approximately 1 cm above the sternal notch. The underlying strap muscles are retracted and the dissection is continued inferiorly towards the superior mediastinum in the pretracheal plane. The innominate artery and vein may be encountered during the dissection immediately deep to the thymus.

Useful landmarks that aid in the identification of the thymus include the thyro-thymic ligaments, which are fibrous bands that connect the superior poles of the thymus to the inferior poles of the thyroid gland. The thymus can also be visually differentiated from the surrounding fatty tissue since it is a deeper tan-colored structure (Figure 1).

An incisional biopsy is obtained from one or both upper poles of the thymus. Hemostasis of the body of the thymus should be meticulous and include suture ligatures and bipolar cautery. Placement of a surgical drain to prevent hematoma formation is not routinely employed if the surgical area is dry. The retrieved thymus tissue is then placed in normal saline and dispatched immediately to the pathology and immunology laboratories.

Fourteen children underwent a transcervical thymic biopsy during the study period. Seven patients were female, and 7 were male. The median age of patients at surgery was 1 year and 6 months (range: 2 months–16 years and 8 months of age), with 9 of the children being <2 years old. The more common presenting signs and symptoms of these patients are summarized in Table 1. Other presentations included cutaneous lesions, sepsis, hepatosplenomegaly, diffuse adenopathy, hypothyroidism, bronchiectasis and anemia.

Nine patients had unilateral biopsies of the superior poles of the thymus, while 5 patients had both lobes of their thymus biopsied. In the 7 patients who did not have concurrent surgical procedures, the mean procedure time for the thymic biopsy was 68 minutes (range 48–100 minutes). A surgical drain was inserted in 9 patients. The only post-operative complication was 1 wound infection which was treated successfully with intravenous antibiotics.

The biopsy samples were subjected to histopathological and immunohistochemical analyses and are summarized in Table 2.

All patients were investigated for a putative profound T cell deficiency. Mitogenic responses to phytohemagglutinin and anti-CD3 antibody were consistently depressed in all patients. However, the number of circulating T cells, with the exception of 2 patients (p8, p14) were not markedly reduced as expected in typical Severe Combined Immune Deficiencies (SCID), hence the need for assessing the thymus. In spite of the reduced function, 4 patients had normal numbers of circulating T cells, while 8 others had reduced but not absent numbers (>1000 CD3+ cells).

Assessment of humoral immunity revealed that all patients but 2 (p5, p12) had abnormal immunoglobulin and (or) antibody levels. The genetic abnormality underlying the immunodeficiency was identified only in 3 patients.

The thymus gland was found to be normal in 5 patients (Table 2). In 2 of these 5 patients (p10, p12), the biopsy results helped to avoid bone marrow transplantation (BMT). In the other 3 patients, the pathology findings were useful in further defining the diagnosis and identifying novel immunodeficiencies.

The rest of the patients had morphological abnormalities of the thymus gland. Classic dysplasia of the thymus in SCID, with loss of architecture, depletion of lymphocytes, loss of cortico-medullary differentiation, and absence of Hassall’s corpuscles is shown in Figure 2A. Immunostaining for T cells with anti-CD3 typically demonstrates marked absence of T cells in the thymus (Figure 2B). Further, detection of cytokeratin in the SCID thymus reveals CK18 expression in the medullary spindle cell component of the thymic epithelial cells as well as cortical surface staining (Figure 3A), as compared with CK18 staining of normal thymic tissue control (Figure 3B). Immunostaining with a cocktail of antibodies for CK8 and CK18 shows strong expression in both cortical polygonal thymic epithelial cells and spindle cells (Figure 4A) as compared with the staining pattern in a normal thymus (Figure 4B). In contrast to the healthy control thymus, in which cortical thymic epithelial cells stain negatively for CK5 and CK6 and medullary thymic epithelial cells stain positively for these markers (Figure 5B), in SCID, these cells all strongly express CK5 and CK6 (Figure 5A).

Three patients had all the hallmarks of a dysplastic thymus including loss, or near loss of lobular structure and cortico-medullary distinction, replacement of thymocytes with epithelioid cells and lack of Hassall’s corpuscles. In these patients the results supported the decision to perform BMT. The other 6 patients had various degrees of thymus gland abnormalities. In 5, Hassall’s corpuscles were either absent or barely developed suggestive of a profound T cell defect. Indeed, all 5 were offered BMT. One of these patients also had bone marrow failure (p3) which was by itself an indication for BMT. In 1 patient (p3) the thymus biopsy was critical in defining the final diagnosis of a novel type of SCID, CD3δ deficiency (Dadi et al. 2003), in another (p4) changes were mild to moderate and did not warrant further intervention.