H. D. Alexander

Enumeration of absolute numbers of T lymphocyte subsets in B-chronic lymphocytic leukaemia using an immunoperoxidase technique: relation to clinical stage

Summary. An immunoperoxidase technique has been used to identify and enumerate helper and suppressor T‐cell subsets, as defined by reactivity with Coulter T4 and OKT8 monoclonal antibodies in 54 patients with B chronic lymphocytic leukaemia (B‐CLL) and in the same number of matched controls. The ratio of T4+ to T8+ cells was significantly reduced in the B‐CLL group as a whole (P<0.001) and in each stage of the three clinical staging systems. There was an increase in the median absolute level of T8+ cells in the whole CLL group (P<0.001). However, subdivision of the CLL group by clinical staging systems revealed a large group (28 patients) in which median T8+ cell levels were not raised and median T4+ cell levels were low (P<0.001). There was no significant decrease in T4:T8+ ratio, increase in T8+ cells or decrease in T4+ cells with progression of clinical stage. Absolute numbers of E+ cells were significantly raised in all staging systems as were E+ T4− T8− cells (P< 0.001). A significant alteration in either of these populations with progression of clinical stage was not present.

Lactoferrin inducible monocyte cytotoxicity defective in esterase deficient monocytes

We recently demonstrated a higher incidence of monocyte esterase deficiency in patients with lymphoproliferative neoplasia and gastrointestinal carcinoma than in the normal population. Using lactoferrin to stimulate monocyte cytotoxicity we have now compared the abilities of esterase positive and esterase deficient monocytes to lyse K562 cells. The esterase deficient monocytes were from normal subjects whose monocytes have consistently failed to show cytoche‐mical esterase staining over 30–72 months and which lack specific monocyte isoenzymes. The esterase positive monocytes were from age and sex matched control subjects. Esterase positive monocytes responded to lactoferrin stimulation by a five‐fold increase in cytotoxicity whereas deficient monocytes failed to produce any response.

Hereditary monocyte esterase deficiency

Summary. An inherited cytochemical staining abnormality of monocytes is described, affecting members of three generations of one family. a‐Naphthyl acetate and a‐naphthyl butyrate esterase staining reactions have been consistently negative in 9 5% of the monocytes of the propositus and her son and in 60‐70% of the monocytes in two of four grandchildren.

Monocyte esterase deficiency in gastrointestinal cancer.

AIM--To substantiate the high incidence of monocyte esterase deficiency (MED) in gastrointestinal carcinoma already reported in a small group of patients; to compare the clinical findings in esterase deficient and esterase positive patients. METHODS--Peripheral blood smears (n = 22) or cytocentrifuge preparations (n = 52) of mononuclear cells from the peripheral blood of patients with gastrointestinal carcinoma were stained by the non-specific esterase stain (pH 5.8) using a batch technique. Samples containing > or = 85% esterase negative monocytes were identified at light microscopic examination. RESULTS--Seven of 74 patients were identified as having MED. This correlated exactly with the proportion (five of 46) found before, using an automated method, and was significantly higher than the 0.8% incidence in normal blood donors shown in that study. Comparison of the clinical details of the 12 MED patients with those of 105 esterase positive patients showed a significantly longer disease free survival in the MED cohort and increased occurrence of benign neoplasms--largely colorectal polyps--in this group also. Three patients had a borderline degree of deficiency and were excluded from comparisons, although they showed the same clinical tendencies as the MED group. CONCLUSIONS--There is a strong degree of association between monocyte esterase deficiency and gastrointestinal carcinoma. Further evidence must be sought to prove that the deficiency precedes the disease and therefore may predispose to it, or at least may identify subjects with such a predisposition. This could lead to early diagnosis and effective treatment of gastrointestinal carcinoma in a sizeable proportion of patients.

Cell sizing in chronic lymphoproliferative disorders: an aid to differential diagnosis.

AIMS: To determine if leucocyte volume distribution analysis (LVDA), obtained using a Coulter Counter Model S Plus IV, can be used to aid differentiation of chronic lymphoproliferative disorder (CLPD) subtypes. METHODS: Mean lymphocyte volume and lymphocyte distribution width were measured on each patient (n = 90) using a hard copy of an amplified LVDA histogram. The mean lymphocyte volume was taken as the mean of the values on either side of the peak at half maximum height. The lymphocyte distribution width was taken as the range of cell values between the two values used to calibrate the mean lymphocyte volume. A template showing typical histograms from commonly occurring CLPD was also produced on an acetate sheet. This was used to examine the histogram from each new patient to evaluate its usefulness as an alternative to the calculation of mean lymphocyte volume and lymphocyte distribution width. RESULTS: Mean lymphocyte volume and lymphocyte distribution width were significantly higher in B cell lymphocytic leukaemia of mixed cell type (B CLL/PL), B cell non-Hodgkins lymphoma with peripheral blood spill, hairy cell leukaemia and T cell prolymphocytic leukaemia than in B cell chronic lymphocytic leukaemia (B CLL). The mean lymphocyte volume, but not the lymphocyte distribution width, was also significantly higher in T cell chronic lymphocytic leukaemia than in B CLL. The template gave an immediate preliminary indication of possible subtype(s) of disorder and could be used as an alternative to measurement of mean lymphocyte volume and lymphocyte distribution width. CONCLUSIONS: Electronic haematology analysers producing an LVDA provide a useful, cost effective cell sizing analysis which can aid the differentiation of subtypes of CLPD.

Identification of cellular immunoglobulins in chronic lymphocytic leukaemia by immunoperoxidase staining.

An indirect immunoperoxidase technique has been used for visualisation of cellular immunoglobulins in chronic lymphocytic leukaemia. Bakers formol calcium was used as fixative. Monoclonal light and heavy chain patterns were demonstrated in 24 out of 27 cases. Only one case did not have any demonstrable immunoglobulins. The presence of alpha or gamma heavy chain immunoglobulin isotypes in leukaemic lymphocytes was found to be related to low mouse rosetting capacity (p less than 0.05).

Quantitative deficiency of monocyte-specific esterase (MSE) mRNA in monocyte esterase deficiency (MED)

Cytochemical staining of monocyte‐specific esterase (MSE) is widely used for identification of the monocytic lineage in leukaemias. Deficiency of this enzymatic activity occurs as a familial trait and the deficiency has been shown to occur with greater frequency in patients with lymphoproliferative or gastrointestinal malignant neoplastic diseases than in normal blood donors. Reverse transcriptase polymerase chain reaction (RT‐PCR), sequencing and quantification by Northern blot analysis was conducted on the MSE mRNA of 12 subjects with monocyte esterase deficiency (MED) and seven MSE‐positive subjects to examine whether mutations were present or whether the defect was quantitative. Mutations were not found in the mRNA sequences. However, MED subjects had significantly less MSE mRNA than MSE‐positive subjects (P = 0·001). These findings show that deficiency of monocyte esterase activity in MED is not as a result of the presence of inactive isoenzymes and may be owing to an abnormality in the regulation of mRNA production.

CD34+CD38-HLA-DR- correlate more closely than CD34+CD38- with engraftment post-PBSC transplantation.

Adult idiopathic autoimmune thrombocytopenic purpura is a chronic and sometimes refractory disease. Life-threatening haemorrages are rare but unpredictable, particularly in elderly patients. Corticosteroids are usually proposed as firstline therapy, but a long-term complete response (platelet count >100 × 10/l) is obtained in only about 20% of patients, and splenectomy is the treatment of choice for nonresponders, because it cures 60–80% of patients. However, the best therapy for refractory patients is a difficult challenge because the efficacy of the large number of drugs tried so far is unpredictable and often transient, and their side-effects are sometimes severe. We describe our experience of treating eight patients (five females) with dapsone. All of the patients gave their informed consent. The results of laboratory tests, including full blood counts, renal and liver function, LDH and G6PDH determinations, showed that all of the patients were platelet autoantibody positive and none had G6PDH deficiency. The mean age at the time of the start of dapsone therapy was 63 years (range 50–77). A complete response (CR) was defined as a platelet count of >100 × 10/l, a partial response (PR) as a platelet count of 50–100 × 10/l, and no response (NR) as a platelet count of <50 × 10/l. The clinical data and platelet counts of our patients are shown in Table I. Six patients had been splenectomized. The mean disease duration before treatment was 147 months (range 12–300 months). The duration of therapy ranged from to 2 to 5 months, with dapsone being given orally at a dose of 75–100 mg/d. In two patients affected by severe thrombocytopenia and with platelet counts that were constantly about 5 × 10/l, dapsone was combined with low-dose steroid treatment. CR was observed in one patient (no. 5) and PR in two (nos. 1 and 4), but dapsone had to be withdrawn because of sideeffects. We found haemolysis in patient 1 (haemoglobin values 12·8–10·5 g/dl) and a three-fold increase in transaminases in patient 4; both proved to be rapidly reversible after dapsone was stopped. No response was observed in five patients, but its administration permitted the withdrawal of steroid therapy in two (nos. 6 and 8). The mechanism of action of dapsone is unknown, although it has been suggested that the reticuloendothelial system may be blocked as a result of excessive red blood cell destruction. In comparison with other series reported in the literature (Durand et al, 1991; Godeau et al, 1993, 1997; Hernandez et al, 1995), the mean age of our patients was higher (63 years), the starting platelet count lower (mean 17 × 10/l) and the mean disease duration longer (147 months). Furthermore, a larger number had unsuccessfully undergone splenectomy (6/8 patients). Our data also confirmed that patients with severe and long-lasting ITP do not respond to dapsone, but its good tolerability and limited cost mean that it can be proposed as an alternative to conventional second-line treatments. It could be used in patients with contraindications to splenectomy or to the prolonged use of low-dose steroids, and British Journal of Haematology, 1999, 104, 641–645

Familiality of monocyte esterase deficiency in patients on continuous ambulatory peritoneal dialysis

The purpose of this study was to determine whether or not peripheral blood monocyte esterase deficiency occurring in patients on CAPD was a familial characteristic. The peripheral blood monocyte esterase status of 74 patients on CAPD was determined by a naphthyl acetate esterase staining of cytospin preparations of their mononuclear cells following separation over ficoll. The peripheral blood of first degree relatives and spouses of monocyte esterase deficiency patients was similarly investigated for the deficiency. Three patients bad monocyte esterase deficiency and familiality of the defect was demonstrated in two of their families. The third family was incompletely investigated because of lack of consent. The monocyte esterase deficiency demonstrated in this cohort of patients did not result from their renal failure but was a familial characteristic.

Early second leukaemia in a patient with successfully treated acute promyelocytic leukaemia.

The outcome in acute promyelocytic leukaemia (APL) has improved significantly since the introduction of therapy combining all trans retinoic acid (ATRA) with chemotherapy. Late sequelae of the disease have, therefore, become an important consideration in the treatment strategy. Relapse occurring after attainment of complete remission (CR) usually originates with the original clone [1]. Development of therapy-related myelodysplastic syndrome or acute leukaemia (tMDS-AML) appears rare, with most authors describing single cases or small series [2] and an incidence of only 6.5% (5 patients) in a recent series of 77 consecutive patients with APL [3]. Most patients developing tMDS-AML exhibit a clear-cut MDS phase preceding AML and latency from achievement of CR from APL of 23 months or greater [2,3]. We report a patient with APL who developed tMDS-AML with myelodysplastic features and absence of t(15;17) with a latency of only 11 months after CR from APL. Thus, poor outcome in APL may occur much earlier than has been reported, in this instance less than 1 year after CR.