Hereditary Benign Intraepithelial Dyskeratosis: An Evaluation of Diagnostic Cytology

Hereditary benign intraepithelial dyskeratosis (HBID) (Mendelial Inheritance of Man [MIM] identification 127600) is a rare autosomal dominant disorder characterized by elevated epithelial plaques located on the ocular and oral mucous membranes. The condition, which was first documented by von Sallmann and Paton,1,2 occurs almost exclusively in individuals who reside in the northeastern section of North Carolina. Most affected persons are members of the Native American Haliwa-Saponi tribe (formerly referred to as Haliwa Indians) or are descendents of them. The ethnic group takes the Haliwa portion of its name from Halifax and Warren counties in North Carolina, where most of the population live. Saponi denotes the “Red Earth People,” one of the original ancestral tribes from which the people descended. Currently, approximately 3000 members are enrolled in the clan. Rare cases of HBID have been described outside of North Carolina. One such individual lived in Philadelphia her entire life but was of Haliwa-Saponi ancestry.3,4 Two individuals from Texas were diagnosed with HBID, but their ancestry remains uncertain.5 

HBID is characterized by bilateral elevated white to grayish epithelial plaques on the exposed perilimbal conjunctiva, sometimes with encroachment of the cornea (Figure 1). The conjunctival leukoplakic lesions are readily moveable over the underlying tissue, and the remainder of the ocular examination is typically unremarkable. The epibulbar blood vessels are commonly hyperemic and have given rise to the common colloquial term for HBID: red eye disease. This bloodshot appearance has caused discrimination against some affected individuals in job employment and in social interactions. Persons not familiar with HBID frequently assume from the appearance of the eyes that alcohol, drug use, or an ocular infection is responsible for the affliction. Clinically, ocular involvement ranges from mild asymptomatic lesions to extensive plaques that involve much of the cornea and bulbar conjunctiva and impaired vision.6 

HBID has an apparent seasonal variation, becoming worse in the spring and summer and subsiding in the cooler weather of autumn. According to affected individuals and their family members, the plaques can shed spontaneously, but this has never been documented photographically. Although the ocular lesions of HBID can resemble malignant neoplasms clinically, and they invariably recur following surgical excision, they do not invade the underlying tissue and remain localized to the conjunctiva and cornea. Furthermore, malignant transformation has never been documented.

The histologic features of surgically excised lesions of HBID include hyperplastic stratified squamous epithelium with an overlying multilayered parakeratotic mantle containing rounded cells with dense cytoplasm and pyknotic nuclei. A mild to moderate lymphoplasmacytic infiltrate within the subepithelial stroma beneath the affected epithelium is typical (Figure 2, A).1–9 Scattered throughout the hyperplastic epithelium from the basal layer to the surface are rounded isolated cells with pyknotic or crenated nuclei surrounded by dense eosinophilic cytoplasm, lending the appearance defined as intraepithelial dyskeratosis in the term HBID (Figure 2, B). The adjacent conjunctival epithelium may vary from normal to a thickened stratified squamous epithelium with or without goblet cells. An ultrastructural study documented the presence of vesicular bodies and cytoplasmic tonofilaments and an absence of cellular interdigitations and desmosomes in dyskeratotic cells.10 

Field trips to the Haliwa-Saponi Native American tribe and to another large family in northeast North Carolina with allegedly no Native American ancestry were undertaken. During these field trips, all family members were examined after providing written informed consent according to a Duke University Institutional Review Board–approved protocol. The conjunctiva was briefly brushed with a CytoSoft Brush (1 CP-5B, Medical Packaging Corp, Camarillo, Calif) for cytologic examination in consenting participants after instilling 1% xylocaine topical local anesthetic. The specimens were smeared on glass slides, fixed with Shannon Cell Fixx Spray Fixative (Thermo Electron Corp, Watham, Mass), and stained with the Papanicolaou method.

Cytologic preparations were obtained from the conjunctiva of 40 individuals (Table), and 123 slides were prepared from this material for microscopic evaluation. A total of 26 individuals were female, 14 were male, and their ages ranged from 5 to 67 years. As previously described, patients were clinically classified by a board-certified ophthalmologist as “definite” HBID (conjunctival erythema in both eyes and plaque >2 mm greatest dimension in both eyes), “probable” HBID (conjunctival erythema in both eyes and plaque >2 mm in a single eye), “unknown” (conjunctival erythema with or without plaque in either eye, or status not possible to determine by clinical examination), or “normal” (no conjunctival abnormality).11 A total of 15 individuals were clinically classified as definite HBID, 4 as probable HBID, 5 as unknown but suspected HBID, and 16 as normal conjunctiva. All cytologic preparations were evaluated in a masked format without knowledge of the clinical information. Genotyping was performed by the fluorescent allele static scanning technique on DNA extracted from blood obtained by venipuncture as previously documented.11 

The cytologic preparations from 15 patients with a clinical impression of definite HBID disclosed rounded squamous epithelial cells with dense homogenous orange cytoplasm and hyperchromatic oval or crenated nuclei (Figure 3, A and B). Cellular halos or cytoplasmic inclusions were not identified. Small lymphocytes were occasionally seen (Figure 3, C). All 15 patients with clinically definite HBID demonstrated 3 alleles for chromosome 4q35 markers.

Three of 4 subjects with a clinical impression of probable HBID manifested the aforementioned cytologic features of HBID, and 1 was cytologically normal. The 3 with the cytologic features of HBID also demonstrated 3 alleles for chromosome 4q35, and the cytologically normal individual did not.

Of 5 patients with unknown but suspected HBID, 3 had cytologic features of HBID, and 2 were cytologically normal. Those with the cytologic features of HBID exhibited 3 alleles for chromosome 4q35 markers, and the 2 with normal cytology did not.

Cytologic brushes of the conjunctiva of the 16 individuals with a clinically normal conjunctiva disclosed squamous cells with round to oval nuclei with a delicate chromatin pattern and lightly eosinophilic or transparent cytoplasm. These normal cells were generally polygonal or elongated, unlike the rounded cells seen in HBID. Unlike the malignant cells of squamous cell carcinoma, the ratios of nuclear to cytoplasmic areas in the cells of HBID were low, and the cells did not have the atypical nuclei of malignant neoplasms with irregular membranes or prominent nucleoli. All clinically normal patients were negative for the chromosome 4q35 markers. Thus, the correlation was 100% between the cytology and genetic findings.

HBID is a clinicopathologic entity that exhibits characteristic clinical, histopathologic, and genetic features. The typical clinical features are confirmed histologically in severe cases that affect vision and whereby the lesions are surgically excised. Although at the current time there are no recommended indications to use cytology and genetic studies for diagnostic purposes, hypothetically they might serve as possible alternatives if there were to be individuals in the future with either an uncertain diagnosis or in whom an excisional biopsy might not be indicated. Our findings indicate that HBID can be diagnosed in an appropriate clinical setting when the characteristic cytologic abnormalities are present, and confirmation of the characteristic cytology was supported in all cases by the presence of the genetic abnormality involving duplication in chromosome 4q35.

The HBID gene has been mapped to the telomeric region of chromosome 4 (4q35) with a peak logarithm of the odds (LOD) score of (ẑ) = 8.97. All affected individuals have had 3 alleles for 2 tightly linked markers, D4S1652 and D4S2390, suggesting that HBID is caused by a genetic duplication at this locus.11 A candidate gene in the region of the duplication is the large (15-kb) human homolog of the Drosophila fat gene, an epithelial gene that can promote abnormal epithelial cell proliferation in Drosophila12,13 and which is thought to function in mammalian cell communication. Even though affected members of the Haliwa-Saponi tribe and persons with no apparent Native American ancestry with HBID do not share a known common ancestor, the finding of an identical duplicated haplotype in these individuals strongly suggests a common ancestral founder.

The hereditary, benign, and intraepithelial components of HBID are well established, and although the responsible gene has yet to be identified, the disorder appears to involve the process of dyskeratosis, which is the abnormal or premature keratinization of individual keratinocytes that results in the formation of cells with a dense hypereosinophilic cytoplasm. Dyskeratocytes can be seen in a variety of benign and malignant processes, including condylomas and squamous cell carcinoma.

HBID shares some cytologic similarities with the genetically unrelated, autosomal dominant Darier-White disease (MIM 124200) and the white sponge nevus of Cannon (MIM 193900), which may also manifest dyskeratotic cells in benign oral mucosa. In contrast to HBID, however, Darier-White disease and the white sponge nevus of Cannon both spare the conjunctiva. Darier-White disease is characterized by suprabasal acantholysis of the epidermis, with the formation of suprabasal clefts. The dyskeratotic cells display central, homogenous, basophilic, pyknotic nuclei surrounded by a clear halo, with a shell of basophilic dyskeratotic material adjacent to the halo.2,8,14–16 The dyskeratocytes are referred to as corps ronds when they are localized within the stratum spinosum, and as grains when they are located within the stratum corneum. Although one report described the occasional finding of perinuclear halos in HBID,2 we have not observed them in any of our cytologic preparations from affected persons. The white sponge nevus of Cannon is characterized by surface parakeratosis and hydropic swelling of the epithelial cells that extends into the rete ridges but spares the basal layer.17,18 The white sponge nevus cells exhibit perinuclear and paranuclear dense eosinophilic intracytoplasmic inclusions composed of disorganized aggregates of cytokeratin filaments and clearing of peripheral cytoplasm.19