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Lymph node cytology: sampling and interpretation (Proceedings)
Lymph nodes are most often aspirated only if they're enlarged, but they may also be sampled to determine if there's metastasis of a tumor. Peripheral lymph nodes are one of the easier tissues to obtain a fine needle aspirate from as this can be done relatively painlessly.
Lymph nodes are most often aspirated only if they're enlarged, but they may also be sampled to determine if there's metastasis of a tumor. Peripheral lymph nodes are one of the easier tissues to obtain a fine needle aspirate from as this can be done relatively painlessly. In many cases, a diagnosis can be made very quickly and relatively inexpensively. However, interpretation of lymph node cytology is somewhat complicated by the fact that there is overlap in the types of cells seen in the differing conditions that can cause lymph node enlargement. Sometimes, the fine needle aspirate findings could be explained by either of several processes, and histologic review of a lymph node is needed to examine the architecture of the node for a definitive diagnosis.
Obtaining the sample
Lymph node aspiration and slide preparation should be performed gently as lymphocytes, especially the less mature lymphocytes, are fragile and easily rupture. It's not at all uncommon to view lymph node cytology preparations that contain 100% ruptured cells, with naked nuclei and sometimes with streaming chromatin.
Aspiration is most often performed with a 22 gauge needle and a 6 or 12 ml syringe. While the lymph node is stabilized with one hand, the other hand performs either a suction-type aspiration or the non-suction, "tattooing" type of aspiration which may prevent the problem of too much peripheral blood contamination of the sample.
The needle is then removed from the syringe, air is drawn into the syringe, the needle is re-attached, and the sample is gently blown out onto a glass slides. Another slide is used to very gently spread out the cells without rupturing them. Slides should be air-dried and may be submitted to a laboratory or stained with an in-house stain.
Slides should be first viewed on low magnification to determine whether there are adequate cell numbers without too much peripheral blood and whether there are enough intact cells that are spread out well enough for evaluation. When good areas are identified, further microscopic examination is carried out at high power or oil immersion.
Normal lymph nodes
Aspirates from normal lymph nodes will have a heterogenous lymphocyte population. Small, mature lymphocytes will make up about 85-95% of the cells observed. These are roughly between the size of an erythrocytes and a neutrophil and they have a round nucleus that almost fills the whole cell, with only a small rim of blue cytoplasm present. Medium-sized lymphocytes are typically about 5-10% of the population. These are somewhat larger than the small, mature lymphocyte. Large lymphoblasts, that contain a distinct nucleolus typically comprise 5% or less of the cells.
Normal lymph nodes may also contain a small number of neutrophils, plasma cells, macrophages and occasional eosinophils and mast cells. Lymph node aspirates often have numerous lymphoglandular bodies, which are small, irregular but often round, granular fragments of lymphocyte cytoplasm that are characteristic of lymphoid tissue. The presence of these lymphoglandular bodies is often helpful in determining cytologically whether it is actually lymphoid tissue that has been aspirated.
Reactive or hyperplastic lymph nodes
Lymphoid reactivity or hyperplasia occurs when lymph nodes are antigenically stimulated. These nodes will also have a heterogenous lymphoid population with small, mature lymphocytes also predominating. However, the percentage of medium-sized to large lymphocytes is increased and may comprise up to 50% of the lymphoid cells, though they are usually less than 30-35%. There is also commonly an increase in the number of plasma cells. Increased numbers of mitotic figures may be seen. Reactive lymph nodes may also contain macrophages, neutrophils eosinophils and mast cells.
Lymph nodes may become inflamed when they are draining an area of infection or tissue damage. The type of inflammation present varies with the cause. The presence of increased numbers of neutrophils (> 5%) is consistent with suppurative (or neutrophilic, or purulent inflammation. This may be associated with local bacterial infection, but also with metastasis of another tumor type, or with immune-mediated disease. Eosinophilic lymphadenitis is present when there are greater than 3% eosinophils, This is often associated with hypersensitivity, but also with mast cell tumors as well as some other tumor types. The presence of increased numbers of macrophages may be called histiocytic lymphadenitis or pyogranulomatous lymphadenitis when neutrophils are also increased. Either may be associated with fungal protozoal, mycobacterial or other types of infection. When lymphadenitis is observed, a careful search for organisms should be performed.
In lymphoma, there may be a very homogenous population of lymphoid cells, though this is not always the case, especially early in disease. Most often, lymphoma is characterized by a predominance of medium-sized to large lymphocytes with nuclei that range from 2 to>3 times the diameter of an erythrocyte. These will typically make up greater than 50% of the cells present. These may be lymphoblasts, with large or distinct nucleoli but there are also lymphomas of intermediate-sized lymphocytes. There are also small cell lymphomas, especially in cats, that are difficult to diagnose cytologically, as they are composed of small, mature-appearing lymphocytes that are actually neoplastic. Neoplastic lymphocytes may have features of malignancy such as coarsely-clumped chromatin, abnormal nuclei that may be clefted or convoluted, and increased numbers of mitotic figures.
When a sample contains too many ruptured cells, care must be taken not to over-interpret the presence of large (naked) nuclei with distinct nucleoli. Only intact cells should be evaluated.
The distinction between B and T lymphocytes can't be made cytologically, for the most part, but there are some differences in prognosis that can be predicted based on the type of lymphocyte present. Immunophenotyping using primary antibodies to T and B cells can be performed on cytologic samples or histologic samples using flow cytometry, immunocytochemistry or immunohistochemistry. Also,if there is a question as to whether a population of lymphocytes is reactive or neoplastic, clonality can be determined using PCR for antigen receptor rearrangement (PARR).
Metastasis of non-lymphoid neoplasia to lymph nodes can be detected by finding cell types that would not normally be present in a lymph node or by finding significantly increased numbers (mast cells, for example) in the node. Some of the tumor types that most commonly metastasize to lymph nodes are mast cell tumors, melanomas, and carcinomas. Sarcomas less frequently are found. Diagnosis of metastatic neoplasia is assisted by finding criteria of malignancy in the metastatic cells as it's not uncommon to aspirate adjacent tissues in the process of aspirating a lymph node. In this case, non-lymphoid cell types could be erroneously interpreted as metastatic.
Some metastatic cell types often have normal cell morphology in the lymph node. Mast cells are a good example of this. The presence of a few mast cells is not diagnostic for metastasis of a mast cell tumor, but more than 3% mast cells in a lymph node aspirate is very suggestive of a neoplastic process.
Another potential problem is in diagnosis of metastatic malignant melanoma. Melanin pigment granules may be taken up by macrophages (so-called melanophages). These are round cells that contain dark blue/black pigment granules that may be interpreted as melanocytes. They may be distinguished from melanocytes if they contain other phagocytized material as well as cytoplasmic vacuoles.
It is not uncommon for a "lymph node aspirate" to contain either fat, or salivary gland cells with normal cell morphology.
The presence of immature cells of the erythroid line, myeloid line, or both are consistent with extramedullary hematopoiesis which occasionally occurs in lymph node. It could be difficult to distinguish this condition from hematopoietic neoplasia that has metastasized to a lymph node.