Bacterial abscesses are an important clinical and radiological mimic of neoplastic lesions. A mature bacterial abscess typically has a central necrotic region, a rim of reactive tissue with fibrosis and a peripheral zone of gliotic brain tissue with oedema. Smear cytology findings will depend on which region of an abscess is sampled; however, neutrophils tend to be prominent in all areas (Fig. 31.3A). The central zone will contain pus. The reactive rim will contain newly formed capillaries and macrophages in addition to acute inflammatory cells. A sampling of the surrounding reactive tissue may not be diagnostic.

Fig. 31.3 (A) Smear of the wall of a bacterial abscess, showing infiltration of neutrophils (toluidine blue). (B) Smear of cerebral tuberculosis showing epithelioid macrophages (H&E). (C) Smear of cerebral tuberculosis showing multinucleated giant cell (toluidine blue).

Tuberculosis is an important cause of infection of the central nervous system. Suspected cases should not usually undergo frozen section, due to the risk to laboratory staff; however, smear cytology with appropriate health and safety precautions may be undertaken. Necrotic tissue, epithelioid macrophages, and multinucleated giant cells may be seen (Fig. 31.3B,C). Epithelioid macrophages may resemble the epithelial cells of a metastatic carcinoma.⁷

Parasitic cysts may contain fragments of the parasite which may be of diagnostic utility intraoperatively. The cyst fluid in such cysts will usually contain macrophages and eosinophils.

Toxoplasmosis, due to infection by the parasite Toxoplasma gondii, results in a necrotising infection, usually in immunocompromised patients. On imaging, it produces ring-enhancing lesions that may be mistaken for a neoplasm. On smear, the edge of the lesion may show reactive gliosis and contain chronic inflammatory cells. More central smears also include necrotic debris and neutrophils. If found, the bradyzoites, swollen cells containing numerous Toxoplasma organisms, are diagnostic.

Fungal infection may produce a granulomatous pattern of inflammation. The presence of a mixed inflammatory infiltrate including multinucleated giant cells in a smear should raise the possibility of fungal infection. Although typically the organism will be identified on paraffin sections rather than cytological specimens, fungal elements may be seen on smear. Fungi that infect the central nervous system may produce hyphae (Aspergillus, Mucor), pseudohyphae (Candida), or yeasts (Cryptococcus).⁷ In histoplasmosis the organisms are characteristically found within macrophages.

The most frequently seen viral infection on intraoperative smear is progressive multifocal demyelinating leukoencephalopathy (see below).

Patients with suspected prion infection should not routinely undergo intraoperative neuropathological assessment, and the tissue should be handled with appropriate decontamination protocols.

Glioblastoma, also known as glioblastoma multiforme is, as its name suggests, a tumour characterised by a wide variety of morphologies, both between different tumours, and within the same tumour. It is defined as a WHO grade IV, diffusely infiltrating astrocytic tumour with nuclear pleomorphism, mitotic activity and necrosis and/or vascular endothelial hyperplasia.

Glioblastomas range from tumours composed of small cells with high nuclear/cytoplasmic ratio and relatively little specific evidence of differentiation (Fig. 31.6A), to giant cell forms with abundant GFAP-positive cytoplasm (Fig. 31.6B). Many are somewhere between these two extremes, consisting of moderate sized cells with elongated nuclei set within a fibrillary stroma (Fig. 31.6C). On intraoperative smear the full range of morphology seen on histology can be represented. Polarisation may help to reveal a fibrillary stroma, which should have a finer texture than the vessel associated collagen, which may be more prominent. Mitotic activity, necrosis and vascular endothelial hyperplasia are all supportive features (Fig. 31.6D); however, such features may not be seen, which may be due to sampling artifact, and their absence is not diagnostic of a low-grade tumour. In some cases it may only be possible to diagnose an astrocytic tumour, with grading awaiting paraffin sections.

Fig. 31.6 Different examples highlighting the range of morphology that may be seen in smears of glioblastoma. (A) A tumour composed of relatively small cells. (B) A tumour containing large neoplastic cells. (C) A typical glioblastoma showing atypical elongated nuclei set within a fibrillary stroma. (D) A glioblastoma with vascular endothelial hyperplasia (toluidine blue).

Grade II diffusely infiltrating astrocytomas are usually slowly growing tumours in young adults. They tend to transform to anaplastic astrocytoma or glioblastoma.

There are several morphological variants of diffuse astrocytoma: fibrillary astrocytoma, gemistocytic astrocytoma and protoplasmic astrocytoma. The most common form is fibrillary astrocytoma, in which astrocytic cells with mildly atypical nuclei and poorly discernible cytoplasm are set within a fibrillary stroma (Fig. 31.7A). The stroma is often microcystic. Mitotic activity is minimal, and necrosis and vascular endothelial hyperplasia absent. On smear, the fibrillary stroma is prominent. The cellularity may be only slightly greater than that of normal brain, and in some cases it may be necessary to wait for paraffin sections to confirm neoplasia.

Fig. 31.7 (A) Smear of a diffuse fibrillary astrocytoma (H&E). (B) Smear of gemistocytic astrocytoma showing neoplastic astrocytes with abundant eosinophilic cytoplasm (H&E).

Gemistocytic astrocytomas contain prominent astrocytes with abundant eccentric eosinophilic cytoplasm. This subtype of astrocytoma, while remaining grade II, has a more aggressive natural history than other types of grade II astrocytoma. The gemistocytic cells are prominent in smears (Fig. 31.7B).

Protoplasmic astrocytoma is a rare subtype composed of astrocytes with monomorphous rounded nuclei and scanty cytoplasm with few processes, set within a stroma rich in mucoid microcysts.²

Anaplastic astrocytoma, WHO grade III, shows features of a diffusely infiltrating astrocytic tumour, set within a fibrillary stroma; however, mitotic activity is more prominent than in a grade II astrocytoma (Fig. 31.7). The WHO Classification is vague about the mitotic rate cut-off between grade II and III astrocytic tumours (which may be reasonable as the distinction is an arbitrary definition within a spectrum of malignancy). Finding any identifiable mitotic figures in a smear should raise concern regarding atypia, and if more than one or two mitoses are seen, I would regard this as favouring anaplasia. Necrosis and vascular endothelial hyperplasia (in the absence of previous intervention) are not present by definition.

Pilocytic astrocytoma is a slowly growing, frequently cystic WHO grade I tumour most frequently found in children and adolescents. The posterior fossa is a common site of origin. The tumour tends to have a biphasic architecture, with areas of differing cellularity. This may be reflected in the smear. The neoplastic cells of pilocytic astrocytoma have fine bipolar processes. The nuclei are usually relatively monomorphous and have a delicate chromatin pattern;⁵ however, in longstanding lesions nuclear pleomorphism may be prominent, and is not of prognostic significance.

Rosenthal fibres are eosinophilic on H&E stained smears, and stain blue with toluidine blue. They are frequent in pilocytic astrocytomas. Granular eosinophilic bodies are also supportive of the diagnosis of a circumscribed astrocytoma (Fig. 31.8).

Fig. 31.8 Smear of pilocytic astrocytoma showing bipolar astrocytic cells with minimal nuclear atypia, and a granular eosinophilic body (Toluidine blue).

The vascular pattern of a pilocytic astrocytoma may cause confusion. Although in diffusely infiltrating astrocytomas vascular endothelial hyperplasia is an indicator of a grade IV diagnosis, in pilocytic astrocytoma it is of no prognostic significance.

Pleomorphic xanthoastrocytoma (PXA) is a rare circumscribed WHO grade II tumour found predominantly in children and adolescents. The lesion is usually superficial and supratentorial. Always consider PXA in the differential of a young patient with a superficial glioma. It is easy to mistake for a malignant astrocytic tumour, as the astrocytes are large and pleomorphic. The cells may have foamy cytoplasm, but this may be inconspicuous. Eosinophilic granular bodies may be seen. A useful feature to help avoid overdiagnosing such tumours is the discrepancy between the pleomorphic cytology and the paucity of mitotic activity.

Subependymal giant cell astrocytoma (SEGA) is a rare WHO grade I periventricular tumour found in patients with tuberous sclerosis. The neoplastic cells are large and intermediate in morphology between astrocytes and neurones.¹¹ These cells will be conspicuous on intraoperative smear.

Primitive neuroectodermal tumours (PNETs) are WHO grade IV tumours predominantly found in children. The most frequent subtype is the cerebellar medulloblastoma; however, supratentorial PNETs are also found, and PNETs are occasionally seen in adults.

PNETs are highly cellular, malignant tumours, with immunocytochemical and sometimes morphological evidence of primitive neuronal differentiation. They may also concurrently show evidence of glial differentiation.

PNETs usually smear as densely cellular monolayers with oval or carrot-shaped nuclei and scanty cytoplasm (Fig. 31.10).¹³ The chromatin pattern is finely granular, and the nuclei are moderately pleomorphic. Mitotic activity may be prominent, and necrosis may be seen. Neuroblastic rosettes may be identified.

Fig. 31.10 Smear of medulloblastoma.

(Courtesy of Dr T S Jacques, Institute of Child Health, London.)

Given the cerebellar location of many PNETs great care should be taken not to confuse cerebellar granular cell neurones with neoplastic medulloblastoma cells. The cells of a medulloblastoma are usually larger than cerebellar granule cell neurones, more pleomorphic, and mitotically active.

In older adult patients metastatic small cell carcinoma and small cell glioblastoma may produce a similar appearance, and are statistically much more frequent than PNET in this age group.

Tumours partially or exclusively composed of neurons with a mature appearance are typically found in the cerebral cortex of young patients with seizures. The neurons are usually large, atypical, and may be binucleate. In gangliocytomas the neoplastic ganglion cells are the only neoplastic component of the tumour. Gangliogliomas contain an additional neoplastic glial component. Calcification and a perivascular chronic inflammatory reaction are often seen. There is frequent subarachnoid involvement of the tumour, which can induce a fibrous response, making smearing difficult.

It may be difficult on a smear to differentiate between an astrocytic tumour infiltrating grey matter and a ganglioglioma. Often the cytoplasm of the neoplastic neurons will be damaged by the smearing process, leaving large bare nuclei with prominent nucleoli. If preserved, the cytoplasm of the ganglion cells may be abundant, and abnormal processes may be seen.

There is a rare malignant variant of ganglioglioma in which there is frank anaplasia of the neoplastic glial component (anaplastic ganglioglioma, WHO grade III).

Central neurocytoma is a tumour composed of sheets of monomorphous small neuronal cells resembling granular cell neurons with rounded nuclei, delicate chromatin, and scanty cytoplasm set within a variably abundant neuropil stroma.¹⁶ It is usually found in the lateral ventricles, is associated with a good prognosis, and is a WHO grade II lesion.

Central neurocytomas tend to spread evenly on smearing, and the monomorphic nature of the cells is prominent. Cytoplasm is inconspicuous.

Although there may be rosette-like fibrillary areas, the neuronal nature of the lesion may be easier to appreciate with immunocytochemical staining than on morphological grounds. The morphology may closely resemble oligodendroglioma.

Dysembryoplastic neuroepithelial tumour (DNET) is a benign tumour characteristically found in the temporal cortex of children and younger adults with partial seizures. The characteristic feature of the tumour is the ‘specific glioneuronal element’, in which parallel bundles of axons are surrounded by cells with small rounded nuclei, resembling oligodendrocytes. ‘Floating neurons’ are present in the matrix between these columns. Many cases are associated with cortical dysplasia in adjacent cortex.

DNET has been divided into simple and complex forms depending on the architecture. The simple form shows the glioneuronal element, without additional cellular elements. The complex form includes both the glioneuronal element and glial nodules. The tumour may include areas with astrocytic, oligodendroglial and/or neuronal differentiation.

Given the vast range of architecture and morphology that can be associated with the diagnosis of DNET, the diagnosis may be difficult to make on small biopsies. The differential diagnosis may include oligodendroglioma, astrocytic tumours and ganglioglioma.

The situation is even more difficult on intraoperative smear. In an appropriate clinical and radiological context finding a population of small, round oligodendroglial-like cells associated with spaces containing ‘floating neurons’ on a smear would be supportive of a diagnosis of DNET.¹⁷ Great caution should be exercised in making such a diagnosis in the absence of supportive clinical/radiological features. It is very difficult to reliably establish intraoperatively whether a neurone is a ‘floating neurone’ in a DNET, or whether the neurone is an entrapped non-neoplastic neurone at the edge of a diffusely infiltrating glioma.