Ocular Cytopathology

Chapter 1- Methods in Ocular Cytopathology

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CHAPTER 1

Methods in Ocular Cytology

In this chapter, methods for processing ocular cytology specimens are described. Most of the routine eye cytology specimens can be prepared by simple fixation and staining methods; although, occasionally more elaborate techniques, such as electron microscopy and immunocytochemistry, are helpful.

FIXATION AND STAINING METHODS

Each fixative and stain has advantages and disadvantages. In general, slides are either rapidly air dried or rapidly fixed in 95% ethanol. Air drying artificially expands the cells while ethanol artificially shrinks the cells. If air dried, a modified Wright or Giemsa stain should be used. (1) May-Grünwald Giemsa demonstrates excellent cytologic differentiation and is especially good for cells of hematopoietic origin. The Papanicolaou stain is excellent for squamous lesions. Hematoxylin and eosin recapitulates standard histopathology and is often preferred by those adept at surgical pathology, while Papanicolaou is preferred by those expert in exfoliative cytology. A combination of stains is helpful. The method of fixation and staining is determined by the clinical question to be answered. If allergic conjunctivitis is to be differentiated from infectious conjunctivitis, then air-dried, Giemsa-stained preparations are appropriate to differentiate eosinophils and neutrophils. If squamous dysplasia or carcinoma is suspected, then ethanol-fixed material reacted with direct fluorescent antibody is very sensitive. (2) Giemsa-stained, air-dried smears can be done as an adjunctive procedure. Example protocols for staining by May-Grünwald Giemsa, Papanicolaou, and hematoxylin and eosin are shown in Tables 1-1, 1-2, and 1-3.
References:
1. Reich C. Modified Wright stain. Am J Clin Pathol 1954:24:881.
2. Bell TA et al. Pediatrics 1984;74:224-228.

CT Scan 2.

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CT Scans of Orbit continued. This cut is taken at the level of the inferior orbit just above the nasolacrimal duct opening in the lacrimal fossa and includes the inferior orbital fissure.

ANATOMY OF THE ORBIT- CT SCANS

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Click on Picture for Enlarged View

CT scan below the orbits includes a section through the auditory canal and nose to show:
1. Maxillary sinus with the nasolacrimal duct just medial.
2. Temporalis fossa with the zygomatic arch at the arrow. Note the absence of bone on the opposite site.
3. Nasal septum.

Click on picture for enlarged view.

CT scan above the opening for the nasolacrimal duct in the lacrimal fossa and includes the inferior orbital fissure.
4. zygoma
5. inferior orbital fissure (note that the fissure is oriented medially and narrows slightly posteriorly).
6. greater wing of the sphenoid
7. nasolacrimal duct
8. inferior rectus muscle

click on picture for enlarged view.

Number 7 here demonstrates the superior orbital fissure. The most common mistake is to indicate that this is the optic canal. However, note that this opening is oriented laterally from the midline plane not medially. This is a key clue to the correct identification. Compare to the image of the CT scan that captures the optic canal below.

Click to enlarge photo

Here we see the optic canal (12 in the figure)which is of course medial to the tip superior orbital fissure and which we just capture more laterally.

Click to enlarge the photo.

In this CT image we are at the top of the orbit so that the lacrimal gland if evident (17). The real challenge here is number 15. This requires a great deal of knowledge. The structure starts lateral in the posterior orbit and moves medially. The immediate guess is that 15 is the superior oblique muscle. However, the origin for the superior oblique is medial to the optic canal, not lateral (see extraocular muscle origins). Therefore this is a critical but forgotten structure the superior ophthalmic vein which is descending to exit the superior orbital fissure in a lower plane (see venous drainage of the orbit). Correct identification of the dilated superior ophthalmic vein is key for the diagnosis of carotid-cavernous sinus fistulas.

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ANATOMY OF THE EYE FOR CYTOLOGY

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CHAPTER 2 Normal Anatomic and Cytologic Features
Accurate cytologic interpretation of ocular specimens requires a fundamental knowledge of normal ocular histology. A general overview is presented here with emphasis on those areas relevant to cytology specimens. There are numerous treatises available for more complete study of ocular anatomy (1-4). The average adult eye measures about 25 mm horizontally, 23 mm vertically, and 21 to 26 mm anterior-posteriorly (Figures 2-1 and 2-2) (5). The eye has an external approximate volume of 7.6 milliliters (ml), the aqueous has a volume of about 200 microliters, and the vitreous a volume of 4.0 ml. The eye is contained in the pear-shaped orbit that has dimensions of about 35 mm vertically, 45 mm horizontally, and 40-45 mm anteroposteriorly (6). The lacrimal gland is located superolaterally in the orbit and is divided by the orbital septum.

CONJUNCTIVA
The conjunctiva covers the posterior surface of the eyelids (palpebral conjunctiva), curves anteriorly at the fornix to reflect onto the anterior surface of the eye as the bulbar conjunctiva (Figures 2-3 and 2-4). There are subtle histologic differences in the conjunctiva of the lid margins, tarsus, fornix, and bulbar conjunctivae (3). The conjunctiva covering the lid margin and bulbar conjunctiva is a modified nonkeratinized, stratified Squamous epithelium. The tarsal and fornix conjunctiva is covered by stratified cuboidal to columnar epithelium of varying thickness (Figures 2-5). This epithelium is unusual because it retains some squamoid features, such as numerous desmosomes, yet has a microvillus surface architecture (7,8). Goblet cells are abundant over the tarsus, fornix, and specialized areas such as the plica semilunaris. Goblet cells are scarce near the lid margin and adjacent to the cornea at the limbus. Most swabs of the conjunctiva are taken from the inferior fornix and show clusters and single epithelial cells with abundant cytoplasm, eccentric nuclei, and occasional single nucleoli . Goblet cells exhibit clear vacuoles filled with mucin. The presence of keratinized epithelium in the conjunctival smear is distinctly abnormal unless the sample is taken from the caruncle or accidentally from the eyelid.

CORNEA
The cornea is covered by five to six layers of a modified stratified Squamous epithelium (Figures 2-7) (9). The basal cells are smaller and have a higher nuclear-to-cytoplasmic ratio than the other epithelial cells in the cornea. There are two to three layers of wing cells with interdigitating cytoplasmic processes connected by desmosomes to other wing cells. These attachments may explain why corneal epithelium tends to be removed in sheets. The two top layers are flattened, superficial cells with small, round nuclei and inconspicuous nucleoli. The superficial epithelial cells are normally uniform in size and shape and have many microvilli that form a microplical complex on the external surface of the cornea (10). Epithelial cells are attached to a basement membrane, beneath which lies Bowman’s layer, a specialized layer of collagen that does not regenerate after injury. The stroma is composed of lamellar sheets of collagen arranged perpendicularly. The posterior surface of the cornea is covered by Descemet’s membrane, and endothelial cells line its posterior surface (Figure 2-7). Cytologic surface smears from the normal cornea will demonstrate cohesive sheets of non-keratinized Squamous epithelium. Individual cells exhibit intermediate-size, round nuclei with bland and uniform chromatin (Figure 2-8). The presence of keratinized cells in smears from the cornea is abnormal.

References:
1. Jakobiec FA. Ocular anatomy, embryology, and teratology. Philadelphia: Harper & Row, 1982.
2. Hogan MJ, Alvarado JA, Weddell JE. Histology of the human eye. Philadelphia: W.B. Saunders, 1971.
3. Last RJ. Eugene Wolff's anatomy of the eye and orbit. Philadelphia: W.B. Saunders, 1961.
4. Fine BS, Yanoff M. Ocular histology, a text and atlas. New York: Harper & Row, 1972.
5. Stenstrom S. Untersuchungen uber die variation unk kovariation der optishen elemente des menshlickhen auges. Acta Ophthalmol 1946;26:1.
6. Duke-Elder WS. The anatomy of the visual system. In: System of ophthalmology. St Louis: CV Mosby, 1961;2:410-413.
7. Greiner JV, Covington HI, Allansmith MR. Surface morphology of the human upper tarsal conjunctiva. Am J Ophthalmol 1977;83:892-905.
8. Dark AJ, Durrant TE, McGinty F, Shortland JR, et al. Tarsal conjunctiva of the upper eyelid. Am J Ophthalmol 1974;77:555-564.
9. Hogan MJ, Alvarado JA, Weddell JE. Histology of the human eye. Philadelphia: W.B. Saunders, 1971.
10. Blumcke S, Morgenroth K Jr. The stereo ultrastructure of the external and internal surface of the cornea. J Ultrastruct Res 1967;18:502.

Ocular Cytopathology

Chapter 1- Methods in Ocular Cytopathology

CT Scan 2.

ANATOMY OF THE ORBIT- CT SCANS

ANATOMY OF THE EYE FOR CYTOLOGY

REFERENCES FINE NEEDLE ASPIRATION OF THE ORBIT

CYSTIC AND VASCULAR LESIONS

SCHWANNOMA, MENINGIOMA OF THE ORBIT

SQUAMOUS, SEBACEOUS AND BASAL CELL CARCINOMAS

WEGENER'S GRANULOMATOSIS, SARCOID, LANGERHANS HISTIOCYTOSIS

GRANULOMATOUS LESIONS OF THE ORBIT

PRIMARY LACRIMAL GLAND TUMORS

METASTATIC SMALL CELL CARCINOMA OF LUNG

MALIGNANT LYMPHOMA OF THE ORBIT

SMALL CELL TUMORS

CHAPTER 10 FNA ORBITAL LESIONS

LEUKEMIA, CHOROIDAL MELANOMA, METASTATIC CARCINOMA

CHAPTER 9- MALIGNANT NEOPLASMS- LYMPHOMA, RETINOBLASTOMA,

PARASITIC EYE INFECTION- Toxoplasmosis

VIRAL RETINITIS- Cytomegalovirus, Acute Retinal Necrosis

HELMINTHIC DISEASES- Cysticercus Cellulosae

FUNGAL ENDOPHTHALMITIS-Coccidiodomycosis, Cryptococcus

FUNGAL ENDOPHTHALMITIS, Candida, Aspergillus, Fusarium

CHAPTER 8 INTRAOCULAR INFECTIONS- Bacterial

Ch 7. Eales Disease of the Retina

Diabetic Retinopathy, Coats Disease, Proliferative Vitreoretinopathy