Irvin A. Lampert

Apoptosis is a Common Histopathological Finding in Myelodysplasia: the Correlate of Ineffective Haematopoiesis

Twenty-three bone marrow biopsies from patients with myelodysplastic syndromes (MDS) with typical histological features were studied, and evidence of apoptosis of erythroid and immature myeloid precursors was seen by light microscopy in each case. This was quantified and found to be significantly greater than that seen in ten normal bone marrow biopsies (p < 0.01). There was no difference in the degree of apoptosis seen in the various sub-groups of MDS proposed by the French-American-British (FAB) co-operative group. These results show apoptosis of haemopoietic precursors to be a characteristic histological finding in MDS, representing intramedullary death of these cells, a factor that may play a role in the pathogenesis of the cytopenias seen in this condition.

Promiscuous expression of Epstein-Barr virus genes in Burkitt's lymphoma from the central African country Malawi

Primary BL in Malawian children has a very high frequency association, approaching 100%, with the human herpesvirus EBV. A detailed study carried out on viral gene expression in these tumours, using both fresh material and methanol‐fixed FNAs, showed, contrary to prediction, that most belong to a variant “class II” latency category, with lytic cycle–related genes also expressed. That is, in addition to EBNA1 expression, membrane proteins (LMP1/2A), immediate early (BZLF1) and early (IR2 and IR4) genes, a putative viral oncogene (BARF1), CST (BART) antisense transcripts and the viral bcl‐2 homologue are expressed in a high proportion of the BLs. Most, but not all, express the small viral (EBER) RNAs. Two other significant observations were made: (i) in addition to expression of cellular cytokine (IL‐10) transcripts in all tumours investigated, the normally silent viral IL‐10 homologue was expressed in some tumours; (ii) whereas EBNA1 expression from its restricted Qp promoter was generally observed, the nonrestricted Cp/Wp promoter was also active in some tumours. Viral gene expression in the Malawian [endemic (e)] BLs appears to be more promiscuous than predicted from other studies, but expression accords with the cytopathologic picture of eBLs as a rapidly proliferating cell population accompanied by considerable necrosis, and a clinically diverse disease. A small‐scale study of relapse Malawian BLs revealed a different picture of viral association, more akin to systemic BL than eBL, where EBV appears to be absent or present only at very low levels. The significance of these findings is considered.

STI571 (Imatinib Mesylate) Reduces Bone Marrow Cellularity and Normalizes Morphologic Features Irrespective of Cytogenetic Response

The tyrosine kinase inhibitor STI571 (imatinib mesylate, Gleevec) is an effective treatment for chronic myeloid leukemia (CML). We examined bone marrow samples from 53 patients with CML who were receiving STI571 in 3 multicenter phase 2 trials to assess morphologic changes and cytogenetic response to this drug. In most patients with initially increased blasts, the bone marrow blast count rapidly decreased during STI571 therapy. Reductions in cellularity, the myeloid/erythroid ratio (commonly with relative erythroid hyperplasia), and reticulin fibrosis (if present pretreatment) also were seen in most patients, resulting in an appearance resembling normal marrow in many cases. Eighteen patients (34%) had some degree of cytogenetic response. Surprisingly, these striking morphologic changes occurred irrespective of any cytogenetic response to STI571. Thus, STI571 seems to affect the differentiation of CML cells in vivo, causing even extensively Philadelphia chromosome-positive hematopoiesis to exhibitfeatures resembling normal hematopoiesis.

High expression of CD23 in the proliferation centers of chronic lymphocytic leukemia in lymph nodes and spleen

Chronic lymphocytic leukemia (CLL) is a B-cell neoplasm composed of a heterogeneous mixture of cells, including small lymphocytes, prolymphocytes, and large transformed cells; these last cells appear to represent the proliferating compartment. CLL cells express, in addition to B cell markers, the transmembrane receptor CD23. CD23 functions as the receptor for IgE and also appears to play a role in controlling the growth and proliferation of lymphocytes. Its level of expression among the different cells in CLL has not been examined. In this study, we show that CD23 expression is much higher in the large transformed CLL cells than in the small lymphoid population. This may provide an explanation for the observed correlation between a circulating CD23 cleavage product (soluble CD23) and prognosis in CLL. In addition, we have shown that proliferation in splenic CLL occurs preferentially in the white pulp zones, even in cases in which both the white and red pulp are extensively infiltrated.

Optimal processing of bone marrow trephine biopsy: the Hammersmith Protocol

Specimens of bone marrow trephine biopsy (BMT) are transported and fixed in acetic acid–zinc–formalin fixative, decalcified in 10% formic acid–5% formaldehyde and processed with other specimens to paraffin-wax embedding. Sections, 1-μm-thick, are cut by experienced histotechnologists and used for haematoxylin and eosin, Giemsa, reticulin silver and other histological stains. Further, all immunohistochemical procedures used in the laboratory, including double immunostaining, can be used on these sections with no or minimal modifications. About 10 000 BMT specimens have been analysed using this procedure since 1997 and diseases involving the bone marrow have been classified successfully. More recently, standardised polymerase chain reaction-based analysis and mRNA in situ hybridisation studies have been conducted. Excellent morphology with good antigen, DNA and RNA preservation is offered by the Hammersmith Protocol.

In vivo evidence for apoptosis in the bone marrow in systemic lupus erythematosus

An increase in leucocyte apoptosis and impaired clearance of apoptotic cells has been observed in patients with systemic lupus erythematosus (SLE). Apoptotic cells are likely to be a key source of autoantigens in SLE as they express many of the nuclear autoantigens (in surface blebs and apoptotic bodies) that are relevant to this disease. The clearance of apoptotic cells is usually a rapid process, such that few cells are usually seen in the extracellular environment in vivo. We report a case in which multiple apoptotic bodies were observed in the bone marrow of a patient with SLE that was complicated by an immune-mediated pancytopenia. We have subsequently examined the frequency of apoptotic cells, identified morphologically, and by caspase-3 staining in bone-marrow trephine samples taken from patients with SLE over a 10-year period of follow-up. A high proportion of bone marrows contained apoptotic debris. The novel demonstration of apoptotic bodies in vivo in patients with SLE is unusual and supports the notion that the marrow may be a target organ in the disease. Their abundance is also consistent with the hypothesis that normal clearance mechanisms are defective and/or overwhelmed in SLE.

The histopathology of prolymphocytic leukaemia with particular reference to the spleen: a comparison with chronic lymphocytic leukaemia

The histological features of prolymphocytic leukaemia are described in spleen, lymph nodes, liver and bone marrow from nine cases. Eight patients had B‐PLL one had T‐PLL. These features were compared with those of six cases of B chronic lymphocytic leukaemia (B‐CLL) with massive splenomegaly. Both PLL and B‐CLL showed enlargement of the white pulp with nodule formation and diffuse infiltration of the red pulp. In both PLL and CLL larger cells were found in the white pulp nodules often concentrated on the periphery producing a bizonal appearance. The extent of involvement of the white pulp was greater in PLL than in uncomplicated CLL. However, this was not so in two cases of B‐CLL in prolymphocytoid transformation in which the heavy replacement of the white pulp by large cells without a bizonal arrangement had effaced the red pulp. The cytological differences between PLL and CLL were best appreciated in splenic red pulp. The cells of CLL were small lymphocytes with clumped chromatin, those of PLL were larger with bigger nuclei, often indented in some cases, and distinct nucleoli. The pattern of infiltration in the bone marrow, liver and lymph nodes was similar in PLL and CLL. In conclusion, PLL can be distinguished from CLL by morphological and immunological features. The distinction is important clinically, because the survival of PLL is shorter than that of CLL.

Selective sparing of enterochromaffin cells in graft versus host disease affecting the colonic mucosa

Graft versus host disease affecting the large bowel causes destruction of the crypt epithelium. There is a selective sparing of enterochromaffin cells in the majority of cases. As a consequence, single as well as small clumps of enterochromaffin cells are to be seen in the sites formerly occupied by the destroyed crypt epithelium. The reason for this phenomenon is unclear, but it may be related to the fact that the enterochromaffin cells are end‐stage and non‐proliferating cells. This is useful diagnostically. However, cytotoxic drugs or irradiation must be excluded as the cause of the mucosal damage to bowel as there are theoretical reasons to expect that a similar phenomenon will be seen after these forms of therapy.

Histopathology of the spleen in T-cell large granular lymphocyte leukemia and T-cell prolymphocytic leukemia - A comparative review

We review retrospectively the spleen histology in 8 patients with T-cell large granular lymphocyte (LGL) leukemia and 4 with T-cell prolymphocytic leukemia (T-PLL) to identify characteristic patterns of involvement and to distinguish such patterns from those described in other low grade B- and T-cell malignancies. Moderate splenic enlargement with red pulp expansion due to lymphocytic infiltration was characteristic of LGL leukemia. Abnormal lymphocytes expressed cytotoxic granule proteins and were consistently CD45RO and CD5 negative in contrast to normal red pulp T cells. This infiltration respected anatomic boundaries with encroachment but no invasion of white pulp areas. Unlike in hairy cell leukemia, the main differential diagnosis for red pulp lymphocytosis, the white pulp was not only preserved in T-cell LGL leukemia but showed germinal center hyperplasia with expansion of the mantle zones. By comparison, T-PLL spleens showed marked red pulp lymphoid infiltration by medium-sized cells with irregular nuclei and prominent eosinophilic nucleoli. T-PLL lymphocytes, unlike LGLs, were more invasive, infiltrating the spleen capsule as well as white pulp areas. T-cell prolymphocytes did not express cytotoxic granule proteins or NK-cell markers, were CD5+, CD45RO+ like normal spleen T cells, were CD2+, CD3+, CD45+, CD43+, TCRβ+, but CD25−, CD30−, ALK-1−, TRAP−, DBA44−, and TdT−. Expression of CD4 and CD8 in these cells mirrored that of circulating T-PLL cells. These observations on the morphologic and immunohistochemical appearances of the spleen in T-cell LGL leukemia and T-PLL may aid diagnosis of these uncommon T-cell disorders, particularly T-cell LGL leukemia, where presentation may be cryptic and where unique pathognomonic features, are absent.

Expression of Epstein-Barr virus lytically related genes in African Burkitt's lymphoma: correlation with patient response to therapy.

A study on the Epstein‐Barr virus (EBV)–associated malignancy (endemic) Burkitts lymphoma (BL) was initiated on fine‐needle‐aspiration biopsies from 46 proven BL cases in Malawi. Gene expression that might correlate with patient serology (where high levels of antibodies to lytically related genes are commonly observed) was explored. In two‐thirds of the cases, we identified the EBV BZLF1 replication activator intermediate early protein ZEBRA in varying quantities and to varying extents in cells by immuno‐cytochemistry. The early lytic‐cycle gene transcript BHLF1 was assessed positively by solid‐phase hybridisation in over half of the same tumours. Evidence of transcription of these genes was confirmed on a smaller number of surgically removed fresh biopsies by RT‐PCR. We asked whether our findings, which are generally counter to the established notion that EBV gene expression in BLs is restricted to the latent function, EBNA1, might offer some explanation for the differential responses to chemotherapy observed among African patients. Where the duration of follow‐up was sufficient to assign the cases (37 in number) to one of 3 catagories, namely, complete, partial or no response, a significant correlation between expression of the viral function ZEBRA and a positive patient response to treatment was found. Lack of this was associated with poor prognosis. Clinical data and EBV gene expression results support the postulate of subgroups of African BLs, the intermediate early antigen providing a marker of potential use in patient management. Int. J. Cancer 81:6–11, 1999.