MZLs originate in the marginal zone B-cells of lymph nodes, spleen, or mucosa-associated lymphoid tissue. The marginal zone is particularly prominent in areas of chronic exposure to antigenic stimulation, from either infectious or inflammatory sources. Exactly how chronic inflammation results in tumorigenesis is unclear and may vary among the different MZL subtypes. It is likely that there is a stepwise progression from reactive B-cell, to localized antigen-dependent tumor, to antigen independence and more aggressive phenotypes. Ultimately, abnormal B-cell clones arise, then proliferate and replace the normal B-cell population.

Cytological features of MZLs are quite variable. They can resemble germinal center centrocytes, with small to medium size, slightly irregular nuclei, moderately clumped chromatin, and inconspicuous nucleoli. Sometimes they have a monocytoid appearance with round or irregular nuclei, more abundant pale cytoplasm, and distinct cytoplasmic membranes. In some cases, they resemble closely small lymphocytes. Plasmacytoid or plasmacytic differentiation can be seen in up to 30 to 50% of cases. Scattered larger transformed cells resembling centroblasts or immunoblasts are usually present. These large cells should not constitute the majority of the cells present, become confluent, or form diffuse sheets, which would signify disease progression and transformation to large-cell lymphoma. The growth pattern of the lymphoma cells is frequently parafollicular with a marginal zone distribution. Interfollicular and diffuse areas of involvement may also be seen. In nodal MZL and mucosa-associated lymphoid tissue (MALT) lymphomas, residual reactive follicles are frequently present, which can be hyperplastic, regressed, or sometimes colonized by neoplastic cells. The typical immunophenotype of MZL is CD20+, CD79a+, CD5−, CD10−, and CD23−. CD21 can be positive. Most nodal MZLs and MALT lymphomas are sIgM+ and IgD−, while sIgG+ and sIgA+ are less common. Splenic MZL (SMZL) is typically sIgMD+. Rare cases of MZL are CD5+ and may carry a worse prognosis. Neoplastic B-cells in cases with plasmacytoid or plasmacytic differentiation express MUM1–IRF4 and monocytypic cytoplasmic immunoglobulins. Extranodal MZLs are commonly associated with chromosomal abnormalities, including t(11;18)(q21;q21), t(14;18)(q32;q21), t(1;14)(p22;q32), t(3;14)(p12;q32), and trisomy 3 or 18. Many of the genetic changes found in MZLs result in upregulation and constitutive activation of the nuclear factor kappa B (NF-kB) pathway, suggesting a vital role in its pathogenesis as well as a target for future therapies.

Due to limitations in available pathologic material, it can be challenging to differentiate MZL from other subtypes of indolent B-cell lymphomas, and “small B-cell lymphoma” is frequently reported as the diagnosis. The following subquestions discuss diagnoses that can be confused with MZL and tips to differentiate between them. A collaborative effort between clinicians and pathologists is often required to arrive at the correct diagnosis.

CLL and SLL are characterized by small lymphocytes in the peripheral blood, bone marrow, spleen, and lymph nodes. The typical immunophenotype is CD5+ and CD23+. Rare cases of CD5+ MZL can be difficult to distinguish from CLL and SLL. For those cases, dim CD20 and surface immunoglobulin expressions, as well as lack of FMC7, favor CLL or SLL. A nodal or extranodal clinical presentation with minimal marrow involvement favors a diagnosis of MZL but should not be used as the sole criterion for differentiation. Morphologically, CLL and SLL tend to exhibit more monotony and have a diffuse growth pattern. Proliferation centers (pseudo-follicles) are frequently seen, particularly in the lymph nodes. MZL is characterized by small to medium-sized lymphocytes surrounding a reactive follicle. In addition, plasmacytic and plasmacytoid differentiation is common in MZL but not CLL or SLL. Except for unusual cases, these diseases do not share common cytogenetic abnormalities, and fluorescent in situ hybridization (FISH) for common genetic anomalies associated with CLL, SLL, and MZL can be helpful in differentiating MZL from the others.

FL is the most common indolent non-Hodgkin’s lymphoma and can present similarly to MZL. However, the vast majority of FL cases are primary nodal FL, although extranodal dissemination can occur. FL is characterized by a follicular growth pattern, consisting of neoplastic germinal centers with markedly attenuated or absent mantle zones. Nodal MZL and MALT occasionally have a follicular growth pattern mimicking FL when follicular colonization is extensive. Staining for germinal center–associated markers like BCL6 and CD10, as well as BCL2 and Ki67, may also help to distinguish between FL and MZL with prominent follicular colonization. The residual germinal center B-cells are BCL6 and CD10 positive and BCL2 negative, with a very high proliferation rate highlighted by Ki67; the neoplastic MZL cells infiltrating the follicles are BCL6 and CD10 negative and BCL2 positive, with a low proliferation rate. FL with marginal zone differentiation may mimic nodal NZL. In those cases, immunostains with BCL6 and CD10 will help in the diagnostic evaluation. SMZLs usually have a micronodular growth pattern that may be mistaken for FL with splenic infiltration. However, the biphasic appearance of SMZL, coupled with the lack of BCL6 and CD10, differentiates it from FL. CD21 staining to highlight the distribution of follicular dendritic cell (FDC) meshwork is also useful in differentiating between FL and MZL (nodal and MALT types). In FL, FDC meshworks are often expanded and relatively more regular. In MZLs, the FDC meshworks can be expanded and fragmented when follicles are run over or colonized by the neoplastic MZL cells, or they can be small and tight, as seen in regressed follicles. While the majority (about 80%) of FL harbor t(14;18)(q32; 21) involving the BCL2 gene, t(14;18)(q32; 21) can also be found in a small percentage of MALT lymphomas. However, the MALT1 gene, not BCL2, is rearranged in t(14:18) in MALT lymphoma.

MCL typically presents with lymphadenopathy, but bone marrow and gastrointestinal tract involvement are very common. Morphologically, lymph nodes involved by MCL have a nodular or diffuse growth of monotonous small to medium-sized lymphocytes with irregular nuclear contours. MCL is CD5+, CD23 −/dim+, FMC7+, and CD43+, and is characterized by overexpression of cyclin D1 and the presence of t(11;14). FISH for t(11;14) is almost always positive, even in CD5− cases, making it relatively easy to distinguish from other types of lymphoma, including MZL. Cyclin D1–negative MCL may be difficult to distinguish from CD5-positive MZL. SOX11 is a useful ancillary stain in those circumstances. SOX11 is consistently positive in MCL, while it is negative in MZL.

LPL is an indolent lymphoma of small B lymphocytes with plasmacytic differentiation (i.e., plasmacytoid lymphocytes and plasma cells are also present); it tends to involve the bone marrow but can be found in lymph nodes or spleen in 15–30% of cases. A paraprotein, usually immunoglobulin M (IgM), is common but is not diagnostic of LPL and can also occur in cases of MZL with plasmacytoid differentiation. There is considerable morphologic and immunophenotypic overlap between MZL and LPL. Centrocyte-like and monocytoid cells, which can be seen in MZL, are absent in LPL. However, in bone marrow biopsies, neoplastic MZL cells appear often as small lymphocytes, and MZL with plasmacytoid and plasmacytic differentiation can be difficult to distinguish from LPL, especially when the involvement by the former is extensive. Immunohistochemistry is rarely helpful in differentiating between LPL and MZL. CD25 is more commonly positive in LPL, and CD11c is more commonly positive in MZL. Comparative genomic hybridization may help to differentiate MZL from LPL. While the two entities share deletions of 6q23 and 13q14 and gains of 3q13–q28, 6p, and 18q, gains of 4q and 8q are associated with LPL but not MZL. Recently, the L256P mutation of the MYD88 gene was reported to be present in almost all cases of LPL but was uncommon in MZL. A polymerase chain reaction (PCR) assay designed to detect and quantify this mutation identified it in 97/104 patients with Waldenström’s macroglobulinemia and 2/20 patients with SMZL. This finding, however, needs to be verified in additional patient populations before being adopted into routine clinical use.

The differential diagnoses for SMZLs include a number of small B-cell lymphomas involving the spleen that are recognized by the World Health Organization (WHO) as splenic B-cell lymphoma, unclassifiable, and hairy cell leukemia (HCL). The two best-defined provisional entities of these categories are splenic diffuse red pulp small B-cell lymphoma and hairy cell leukemia variant (HCL-v). Differentiation of these entities by peripheral blood cytology can be difficult, since they share similarities including the presence of villi. Morphologic features are more distinct among these entities. In the spleen, SMZL involves the white pulp and also the red pulp in a follicular or micronodular pattern. A biphasic cytological pattern typically observed is characterized by small round lymphocytes in the interior of the follicles surrounded by an outer zone of marginal zone cells with more abundant pale cytoplasm, admixed with scattered larger transformed cells. Splenic diffuse red pulp small B-cell lymphoma involves the red pulp with both cord and sinusoidal patterns. The neoplastic cell population is monotonous with scattered blasts, and it does not show follicular replacement, biphasic cytology, or marginal zone infiltration. An intrasinusoidal infiltration pattern is consistently seen in the bone marrow. HCL and HCL-v diffusely involve the red pulp, and the white pulp is atrophic. Immunophenotypically, SMZL cells express IgM and almost always IgD, while splenic diffuse red pulp small B-cell lymphoma, HCL, and HCL-v tend to be IgG positive. CD103 and CD11c are more frequently positive in HCL and HCL-v. Contrary to SMZL, HCL is also positive for Annexin A1, TRAP, and CD25. About 40% of SMZLs show allelic loss of chromosome 7q22–36, which is not found in splenic diffuse red pulp small B-cell lymphoma. HCL has been found to harbor a V600E mutation in the BRAF gene. This may serve as a molecular marker for HCL that could potentially distinguish it from SMZL, although it has not yet been adopted into diagnostic criteria. Although analysis of both bone marrow and peripheral blood lymphocytes is the primary means of diagnosis, occasionally the only way to acquire sufficient material for pathologic diagnosis is to perform a splenectomy. Under these circumstances, the clinician must weigh the benefits of having a precise diagnosis against the risks of the procedure.