Ubiquitin Immunostaining in Thyroid Neoplasms Marks True Intranuclear Cytoplasmic Pseudoinclusions and May Help Differentiate Papillary Carcinoma from NIFTP

Abstract

Papillary thyroid carcinoma (PTC) is defined by an invasive growth pattern and classic nuclear features: enlarged, grooved, overlapping nuclei with chromatin clearing and intranuclear cytoplasmic pseudoinclusions (INCP). True INCPs are characteristic of PTC, but may infrequently be seen in noninvasive follicular thyroid neoplasms with papillary-like nuclear features (NIFTP). Nuclear abnormalities that mimic INCP ("pseudo-pseudoinclusions") are common in a variety of thyroid lesions. H&E and ubiquitin-stained whole tissue sections of classic PTC (n = 25) and NIFTP (n = 35) were evaluated. On H&E, true INCPs were present in all (100%) PTCs and absent in all NIFTPs (0%). Pseudo-pseudoinclusions were present in 13 (37%) NIFTPs. In 24 (96%) PTCs, ubiquitin was strongly expressed within INCPs. In NIFTPs, optically clear nuclei or pseudo-pseudoinclusions did not express ubiquitin (0/35). Occasionally, nuclear vacuoles in NIFTP demonstrated a marginated staining pattern, in which strong ubiquitin expression was seen at the periphery of the nucleus, but the central pale area was negative. In addition, 2 NIFTPs demonstrated intrafollicular psammomatoid calcifications which were strongly ubiquitin-positive. Psammoma bodies in PTC were ubiquitin-negative in the majority of cases. We report a previously undescribed finding: strong ubiquitin expression in true INCPs in PTC, absence of true INCPs in NIFTP, and absence of ubiquitin expression in pseudo-pseudoinclusions in NIFTP. This finding supports the difference between true INCPs (found only in PTC) and pseudo-pseudoinclusions (found in NIFTP). Using strict histologic criteria and ubiquitin immunostaining, the presence of true pseudoinclusions may exclude a diagnosis of NIFTP. Caution should be exercised when interpreting nuclear vacuoles or pseudo-pseudoinclusions.

Three‐dimensional structural analysis of papillary thyroid carcinoma nuclei with serial block‐face scanning electron microscopy (SBF‐SEM)

“…Pseudo‐pseudoinclusions may be a factor in the misdiagnosis of PTC. Immunohistochemistry of ubiquitin enabled us to distinguish between pseudo‐pseudoinclusions and INCIs 43 . There were small vacuolar structures between the INCI and the nuclear perimeter in some closed INCIs.…”

Diagnostic Utility of p62 Expression in Intranuclear Inclusions in Thyroid Core Needle Biopsy Specimens

“…There have been several attempts to distinguish NuIns from NuVas using IHC to diagnose PTC (7,15,16). Schwertheimet al (7) examined the expression of several autophagy-associated proteins in NuIns, including LC3B, ubiquitin, cathepsin D, p62/sequestosome 1, and cathepsin B.…”

Posttranslational Modifications in Thyroid Cancer: Implications for Pathogenesis, Diagnosis, Classification, and Treatment

“…In a recent study by Cracolici et al [ 44 ], the interesting observation was made that the cytoplasm in true nuclear pseudoinclusions is always positive for ubiquitin immunostaining, whereas pseudo-pseudoinclusions do not exhibit ubiquitin immunostaining. From these observations, the authors proposed that ubiquitin immunostaining may be used to differentiate true pseudoinclusions from pseudo-pseudoinclusions and that, when using this criterion for true pseudoinclusions, their presence may be used to exclude NIFTP.…”

“…Although the study by Cracolici et al [ 44 ] did not define which cytoplasmic proteins in the pseudoinclusions were ubiquitinated or the function and biological role of the ubiquitinated proteins, the observations may be of clinical importance in helping differentiate NIFTP from PTC. This would be particularly helpful in patients presenting with a Bethesda V or VI nodule and in whom the extent of surgery (i.e., lobectomy vs. total thyroidectomy) may be influenced by the degree of suspicion that the nodule is NIFTP rather than cancer.…”