IC can be used to demonstrate the morphology of normal and abnormal conjunctival cells. Cytology report was positive in 77% of histopathology reports in the moderate dysplasia to microinvasive carcinoma group. There were no false positives.
IC is accurate in predicting the diagnosis of OSSN. Correlation rate of IC with histological findings was accurate in 80% of cases, poor in 12% of cases, and not correlated in 8% of cases. There were no false positives.
Intraepithelial OSSN or corneal/conjunctival intraepithelial neoplasia (142) Invasive OSSN or SCC (23)
The cytomorphology of OSSN is described in detail. For intraepithelial lesions, these include (1) keratinized dysplastic cells often accompanied by hyperkeratosis (55%), (2) syncytial-like groupings (35%), and (3) nonkeratinized dysplastic (10%) cells. Meanwhile, invasive cases had a tendency to be more highly keratinized and to have a greater degree of inflammation than the keratinized high grade intraepithelial cases but it was not possible to confidently predict invasion on IC. Sensitivity of IC in diagnosing OSSN was 78% overall but was lower (70%) when the lesion was invasive by histopathology.
OSSN (50) including SCC (20), dysplasia (20), squamous papilloma (4), and nondysplastic changes of the epithelia (6) Pigmented lesions (5) including nevus (4) and malignant melanoma (1)
Compared with histologic findings, IC had a high positive predictive value (PPV) of 97.4% and a fair negative predictive value (NPV) of 52.9%, making it a good screening tool but inadequate gold standard. Moreover, IC is less sensitive for keratotic lesions and invasive disease.
Cytological features related to malignancy were applied to determine an index score that best differentiates invasive SCC from preinvasive lesions. With an index score of ≥4.25, sensitivity was 95%, specificity was 93%, PPV was 95%, and NPV was 93% for predicting SCC. However, in two preinvasive lesions (one AK lesion and one CCIN lesion), IC sampling was not sufficiently deep to reach atypical cells and presents a limitation to diagnose disease affecting deeper tissue.
Pterygia without atypical cells (19) Pterygia with associated OSSN (13)
IC showed high agreement with histopathology in detecting unsuspected OSSN in pterygia patients (sensitivity 92%, specificity 94%, PPV 92%, and NPV 94%).
Toluidine blue 1% is a good tool for the diagnosis of OSSN and premalignant lesions but the intensity of staining does not correlate with the degree of malignancy (sensitivity 100%, specificity 50%, PPV 73%, and NPV 100%).
Malignant lesions including invasive SCC (16) and severe dysplasia (17) Benign or premalignant lesions (42)
Methylene blue 1% can exclude malignant lesions but cannot replace histopathology as the gold standard (sensitivity 97%, specificity 50%, PPV 60%, and NPV 95%).
Toluidine blue 0.05% is a good screening tool but not a good diagnostic tool due to a high frequency of false positives (sensitivity 92%, specificity 31%, PPV 41%, and NPV 88%).
20 MHz high frequency ultrasound demonstrates deep involvement of tumor into the sclera, globe, or orbit and is a useful tool for tumors with deep invasion. It is of otherwise limited utility in tumors without deep extension.
Conjunctival intraepithelial neoplasia and SCC (11)
20 and/or 50 MHz high frequency ultrasound helps delineate tumor thickness, shape, and internal reflectivity and is particularly helpful in determining if the tumor has extended into the sclera, eye, and orbit.
IVCM findings of OSSN lesions were described. Structural findings included large areas of superficial cell debris and/or keratin debris accompanied by syncytial-like groupings, loss of the normal structure of the conjunctival epithelium and/or the corneal basal epithelium layer, papillomatous organization, large fibrovascular structures, and fine vessels perpendicular to the tumor surface. Marginal findings included subepithelial (pre-Bowman) space involvement in 4 cases, irregular healthy tissue infiltration at the lateral edge of the lesion in 2 cases, and abrupt demarcation between neoplastic cells and normal epithelium in 8 cases. In vivo cytomorphologic findings included cellular anisocytosis, pleocytosis, and anisonucleosis, enlarged nuclei with high nuclear to cytoplasmic ratio, high reflective cytoplasm, and indistinct cytoplasmic borders.
Corneal/conjunctival intraepithelial neoplasia (CCIN) (4) Control (4; 2 with limbal stem cell deficiency, 1 with suspicious limbal lesion, 1 with diffuse keratoconjunctival proliferation)
This study defined IVCM features of CCIN, which included (1) hyperreflective pleomorphic cells of varying shapes and sizes, (2) the edge of hyperreflective CCIN lesion contrasting with the darker and smaller normal cells, (3) “starry night sky” pattern of the basal layer produced by prominent nucleoli, and (4) absence of subbasal corneal nerves within areas involved by CCIN compared to nonaffected regions. These IVCM findings were found to be highly correlated with histologic features.
IVCM demonstrated cellular anisocytosis and enlarged nuclei with high nuclear to cytoplasmic ratio in conjunctival intraepithelial neoplasia while nests were partially formed by isolated keratinized, binucleated, and actively mitotic dysmorphic epithelial cells in carcinoma in situ and ocular surface squamous carcinoma. The IVCM characteristics were similar to histopathologic findings.
OSSN (26) Benign conjunctival lesions (18) Age matched controls with normal conjunctiva (57)
IVCM was able to reliably distinguish normal conjunctiva from conjunctival lesions. However, IVCM was unable to reliably differentiate OSSN from benign conjunctival lesions due to an overlap in IVCM features between the two conditions (kappa = 0.44, 95% CI 0.32–0.57). IVCM has a low sensitivity (38.5%) and moderate specificity (66.7%) for distinguishing OSSN from benign conjunctival lesions.
Ultra-high resolution (UHR) OCT demonstrated a thickened hyperreflective epithelium and abrupt transition from normal to hyperreflective epithelium in all CCIN cases. After medical treatment and clinical resolution, 4 cases demonstrated normal epithelial configuration on UHR-OCT while 3 cases showed residual disease that was clinically invisible. Continuation of treatment resulted in complete resolution of residual lesions on UHR-OCT.
UHR-OCT findings of OSSN and pterygia were well correlated to histopathologic findings. Both UHR-OCT and histopathologic specimens of OSSN demonstrated a thickened layer of epithelium, often with an abrupt transition from normal to neoplastic tissue. Meanwhile, both diagnostic modalities for pterygia showed a normal thin epithelium with underlying thickening of the subepithelial mucosal layers. Mean epithelial thickness on UHR-OCT for OSSN (346 μm) was significantly higher than for pterygia (101 μm) (). UHR-OCT had a high sensitivity (94%) and specificity (100%) for differentiating OSSN from pterygia.
UHR-OCT and histopathologic findings were closely correlated for all the examined ocular lesions. Specifically for OSSN, the epithelial layer was severely thickened and hyperreflective with an abrupt transition between normal and affected epithelium. In large lesions, a shadow from the hyperreflective epithelium may obscure the inferior border. The subepithelial layer was uninvolved in OSSN.
OSSN (21) Pterygia or pinguecula (24) Lymphoma (3) Pigmented conjunctival lesions (18) Salzmann nodular degeneration (6) Normal (10)
Commercially available HR-OCT is also capable of differentiating various lesions based on optical signs. Specifically in OSSN, HR-OCT shows epithelial thickening and hyperreflectivity.
