Diffuse large B-cell lymphoma with a novel translocation involving BCL6.

Translocation of the BCL6 gene is one of the most common chromosomal changes seen in diffuse large B-cell lymphoma, primarily affecting the 5' regulatory region which is usually replaced with the sequences of the translocation partner. This translocation involves both immunoglobulin and nonimmunoglobulin gene partners, the former being the more common. Here we report a case of diffuse large B-cell lymphoma with immunophenotypic features of a germinal center type, involving translocation of BCL6 to chromosome 11 and partnering with one or more nonimmunoglobulin genes. To our knowledge this novel translocation in the context of an activated B-cell type diffuse large B-cell lymphoma has not been previously described in the literature. We speculate about the putative partner gene involved in the translocation and the pathophysiological significance of the translocation.

The t(8;22) in chronic myeloid leukemia fuses BCR to FGFR1: transforming activity and specific inhibition of FGFR1 fusion proteins.

This report describes 2 patients with a clinical and hematologic diagnosis of chronic myeloid leukemia (CML) in chronic phase who had an acquired t(8;22)(p11;q11). Analysis by fluorescence in situ hybridization (FISH) and reverse transcription-polymerase chain reaction (RT-PCR) indicated that both patients were negative for the BCR-ABL fusion, but suggested that the BCR gene was disrupted. Further FISH indicated a breakpoint within fibroblast growth factor receptor 1 (FGFR1), the receptor tyrosine kinase that is known to be disrupted in a distinctive myeloproliferative disorder, most commonly by fusion to ZNF198. RT-PCR confirmed the presence in both cases of an in-frame messenger RNA fusion between BCR exon 4 and FGFR1 exon 9. Expression of BCR-FGFR1 in the factor-dependent cell line Ba/F3 resulted in interleukin 3-independent clones that grew at a comparable rate to cells transformed with ZNF198-FGFR1. The growth of transformed cells was inhibited by the phosphatidylinositol 3-kinase inhibitor LY294002, the farnesyltransferase inhibitors L744832 and manumycin A, the p38 inhibitors SB202190 and SB203580 but not by the MEK inhibitor PD98059. The growth of BaF3/BCR-FGFR1 and BaF3/ZNF198-FGFR1 was not significantly inhibited by treatment with STI571, but was inhibited by SU5402, a compound with inhibitory activity against FGFR1. Inhibition with this compound was associated with decreased phosphorylation of ERK1/2 and BCR-FGFR1 or ZNF198-FGFR1, and was dose dependent with an inhibitory concentration of 50% of approximately 5 microM. As expected, growth of BaF3/BCR-ABL was inhibited by STI571 but not by SU5402. The study demonstrates that the BCR-FGFR1 fusion may occur in patients with apparently typical CML. Patients with constitutively active FGFR1 fusion genes may be amenable to treatment with specific FGFR1 inhibitors.

Liposarcoma in the bone marrow: a terminal event.

We report a case of metastatic liposarcoma in the bone marrow with a rapidly fatal course. In view of the poor prognosis and paucity of clinical and imaging findings in patients with high-grade liposarcoma that is metastatic to bone marrow, we propose that bone marrow examination should be performed during the patient's initial evaluation as well as follow-up examinations.

Cytodiagnosis of a primary effusion lymphoma. A case report.

BACKGROUND: Primary effusion lymphoma (PEL), commonly described in AIDS patients, is a unique subset of lymphoma in which the neoplastic lymphocytes proliferate exclusively in serous cavities. CASE: A 27-year-old male, HIV positive for five years and with multiple opportunistic infections in the past, was admitted for sudden-onset shortness of breath caused by a pleural effusion. Cytologic examination of the pleural fluid revealed medium to large atypical lymphocytes with a high mitosis rate, suspicious for lymphoma. Further diagnostic tests, such as immunophenotypic analysis and cytogenetic and molecular studies, confirmed the diagnosis of PEL. CONCLUSION: Cytopathologists and cytotechnologists should be aware of this new entity since additional studies are required for a definitive diagnosis.

Biotechnology at low Reynolds numbers.

The shrinking of liquid handling systems to the micron and submicron size range entails moving into the area of small Reynolds numbers. The fluid dynamics in this regime are very different from the macroscale. We present an intuitive explanation of how the different physics of small Reynolds numbers flow, along with microscopic sizes, can influence device design, and give examples from our own work using fluid flow in microfabricated devices designed for biological processing.

Deformation and flow of red blood cells in a synthetic lattice: evidence for an active cytoskeleton.

We introduce the use of microfabrication techniques to construct on a silicon wafer a synthetic capillary bed with 2.5- to 4-micron (mu)-wide channels. Establishment of a fluid pressure gradient allowed us to observe simultaneously using optical microscopy hundreds of cells flowing through the bed at physiological speeds. We find a large distribution of mobilities among red cells flowing through the structure; smaller channels provide a greater impedance to flow than larger ones, indicating that kinetic drag variations provide the origin of the distribution. The mobility of a particular cell is not correlated with the cell diameter but appears to be inversely correlated with intracellular calcium concentration of the cell, as determined by fluorescence of the calcium-binding dye fluo-3 AM. Also, we are able to use the parallel processing nature of our arrays to observe isolated events where the rigidity of the red cell seems to change suddenly over several orders of magnitude as it blocks a channel in the array.

Acute agranular CD4-positive natural killer cell leukemia. Comprehensive clinicopathologic studies including virologic and in vitro culture with inducing agents.

BACKGROUND: A 63-year-old male presented with fever, a subcutaneous nodule, gingival hypertrophy, lacrimal gland enlargement, and no lymphadenopathy or hepatosplenomegaly, but had anemia, thrombocytopenia, and peripheral blood (PB) plus bone marrow (BM) involvement by leukemic cells. There was minimal response to multiagent chemotherapy and local radiotherapy, with a survival of 6.5 months from disease diagnosis. METHODS: The PB and/or BM leukemic cells were evaluated using electron microscopy (EM), immunohistochemistry, flow-cytometric immunophenotyping, cytochemistry, cytogenetics, Southern blot analysis for gene rearrangement and Epstein-Barr virus (EBV), polymerase chain reaction for EBV and human herpes virus-6 (HHV-6), and in vitro culturing with inducing agents. RESULTS: The leukemic cells were agranular and monocytoid, with a hairy cell-like bone marrow biopsy infiltrate. Myeloperoxidase (MPO) and alpha-naphthyl butyrate esterase staining was negative, and periodic acid-Schiff staining was positive by light microscopy. Electron microscopy showed MPO negativity and a lack of parallel tubular arrays. The immunophenotype was CD3-, CD56+, CD4+, CD8-, CD15+, TCR1-, and TCR2-, with germline immunoglobulin and T-cell receptor genes and an abnormal karyotype (44XY, 5q-, -13, 13q+, -15). No genomic material for EBV or HHV-6 was detected. Cell cultures with butyrate and N,N-hexamethylene bis-acetamide suggested the possible induction of tumor cells to express a T-cell immunophenotype. CONCLUSION: A case of clonal acute natural killer (NK) cell leukemia with an unusual morphology (agranular) and unique phenotype (CD3-, CD56+, CD4+, CD15+) is presented. Unlike as in other acute NK leukemias, EBV was negative; there was no evidence of HHV-6. The tumor cell, after culturing with differentiating agents, may have been induced to express a T-cell immunophenotype.

Terminal deoxynucleotidyl transferase activity in acute leukemia: a study of 100 cases comparing an immunoperoxidase (PAP) vs immunofluorescent method.

A comparison between the immunofluorescent (IF) method for terminal deoxynucleotidyl transferase (TdT) activity and the immunoperoxidase (IP) method by peroxidase-anti-peroxidase (PAP) technique was done for 100 cases of acute leukemia. For the acute lymphoblastic leukemias (ALL) there was agreement in 93% of the cases. However, the IP method detected 51/55 (93%) TdT+ cases versus 47/55 (85%) by the IF method. For the acute nonlymphocytic leukemias (ANLL), there was an agreement in 89% of the cases. The IP method detected 8/36 (22%) TdT-positive cases while IF detected 4/36 (11%) positive cases. If a figure of 10% TdT+ cells is considered significant in the marrow of the ANLLs, then the IP method would detect eight additional cases for a total of 16/36 (44%) TdT+ cases. This latter figure questions the ability of the IP TdT assay as a single test adequately to determine the lineage of a cell line. It may be rather that TdT is a marker that is expressed in a stem cell.

Morphometric study of megakaryocytes size and prognosis in adults with acute non-lymphocytic leukemia.

Morphometric analysis of megakaryocyte cellular and nuclear size in bone marrow biopsies of 28 cases of acute non-lymphocytic leukemia at diagnosis and 15 controls was performed. Median megakaryocyte cell diameter, median area, and perimeter were less than 95% control range (less than 18.5 microns, less than 263 microns2, less than 64 microns) in 5/5 of induction failures. Attainment of complete remission was significantly greater in those with median megakaryocyte diameter greater than or equal to 18.5 microns (p less than 0.01), median area greater than or equal to 263 microns (p less than 0.001) and perimeter greater than or equal to 64 microns (p less than 0.001). Prolonged complete remission was correlated with normal megakaryocyte size with the median megakaryocyte area, median diameter, and perimeter within or greater than the reference range (greater than or equal to 18.5 microns, greater than or equal to 263 microns2, greater than or equal to 64 microns, p less than 0.05) in 6/7 cases with continuous remissions greater than 3 yr. Measurements of megakaryocyte size may be useful in predicting induction failure and possibly the likelihood of prolonged complete remission in adults.

Prolymphocytic transformation of chronic lymphocytic leukemia. A report of three cases and review of the literature.

Three cases of prolymphocytic transformation of chronic lymphocytic leukemia (PLL-trs CLL) are described, and a review of the literature yields 15 additional cases. PLL-trs CLL may be recognized when two populations of cells are present and the number of prolymphocytes are greater than 15% admixed with the small round lymphocytes of CLL. Surface immunoglobulin intensity appears to be greater on the prolymphocytes than on the small lymphocyte in the same case, suggesting a clonal differentiation. At the time of transformation, the spleen is generally larger than at the time of diagnosis of CLL, and lymphadenopathy is variable. Transformation is thought to be associated with increasing refractoriness to treatment and shorter survival, although most of the deaths occurred in clinical Stages III and IV, known to have a poor prognosis. Similar features between PLL-trs CLL and so-called de novo PLL also suggest an evolutionary process. However, additional morphologic and immunologic studies in transforming CLL cases are necessary to answer this question.

Bone marrow response to chemotherapy in acute lymphocytic leukaemia and acute non-lymphocytic leukaemia.

Histopathologic changes in core bone marrow biopsies were reviewed in 33 patients with acute leukaemia during chemotherapy to compare the changes in acute lymphocytic leukaemia (ALL) with acute non-lymphocytic leukaemia (ANLL). Cellular, stromal, and bony changes were evaluated with regard to diagnosis and time of biopsy from initiation of chemotherapy. A significant difference was noted in the plasma cell response. Plasmacytosis was present in 19/19 cases of ANLL, but in only 2/14 cases of ALL. Cellular depletion was also significantly less frequent in ALL. Other stromal changes such as haemorrhage, dilatation of sinusoids and fat regeneration, as well as osteoblastic bone activity occurred with similar frequencies in all cases of treated acute leukaemia. Fibrosis, necrosis, and serous atrophy were uncommon. Differing chemotherapeutic regimens and differing patient ages were both correlated with the plasma cell response, but not with the difference in cellular depletion.

Heterogeneous acute leukemias.

We have reported two cases of Philadelphia-chromosome-negative acute leukemia with heterogeneous populations of leukemic cells and reviewed six cases from the literature. The cases were identified by several methods, including morphology, cytochemistry, enzymatic analysis, cytogenetic analysis, and electron microscopy. Predisposing factors were probably present in three cases and may partially account for the seemingly increasing incidence, as has been the experience with other secondary leukemias. The prognosis was variable, but most of the complete remissions were seen only when lymphocytic and nonlymphocytic antileukemic chemotherapeutic regimes were used. The occurrence of these unusual acute leukemias strongly supports the concept of a lymphoid-myeloid stem cell.

A prospective study of nosocomial infections in a chronic care facility.

To elucidate the epidemiology of nosocomial infections occurring in nursing homes and chronic care facilities, the authors undertook a prospective study of patients requiring two different levels of nursing care. The overall rate of infection was higher on the intermediate care ward than on the nursing home ward (1.35 versus 0.67 infections/100 patient care days). Pneumonias and symptomatic urinary tract infections accounted for 49 per cent of all infections. Eight of ten cases of pneumonia occurring on the nursing home ward were diagnosed in the winter months, and no case was diagnosed in the summer months. Resistance to gentamicin, tobramycin, ampicillin, and trimethoprim-sulfa was common among organisms causing symptomatic urinary tract infections.

Prostatic acid phosphatase as measured with two radioimmunoassay kits in the detection of prostatic adenocarcinoma.

Serum prostatic acid phosphatase concentration was measured with two commercially available radioimmunoassay kits. Results were compared with histological evidence of prostatic adenocarcinoma obtained at autopsy in 33 patients. The serum assay did not differentiate significantly (p greater than 0.1) between patients with adenocarcinoma and those without. We conclude that the test, at least as performed by use of these kits, is of little value in the detection of occult disease.

Antigenemia detected by enzyme-linked immunosorbent assay in rabbits with systemic candidiasis.

To aid in the rapid, accurate, and early diagnosis of systemic and invasive Candida infections, a double-antibody sandwich enzyme-linked immunosorbent assay was developed for the detection of circulating Candida antigens. The sensitivity for purified mannan was 0.1 to 1 ng/ml. No reactivity was seen with a variety of other fungal and bacterial antigen preparations. The assay was evaluated in normal or immunosuppressed rabbits with varying degrees of systemic candidiasis. All rabbits given infecting doses of 10(7) or 10(8) organisms were positive for Candida antigens with increasing titers until death. Rabbits given 10(6) organisms were variably and transiently positive. All control infected and uninfected rabbits were negative. This sensitive and specific technique detects Candida antigens in the serum of infected rabbits before widespread organ involvement occurs and should prove extremely valuable in the early diagnosis of systemic candidiasis.

Evaluation and comparison of two assays for detection of immunity to rubella infection.

Two commercially available rapid screening tests, Rubacell (Abbott Laboratories; passive hemagglutination) and FIAX (International Diagnostic Technology; indirect immunofluorescence) were compared with a standard hemagglutination inhibition assay for detection of immunity to rubella infection. In tests of approximately 300 sera, both rapid assays were specific and sensitive and showed a high predictive value of a positive result. Within-run reproducibility studies were excellent for both tests; however, Rubacell was superior to FIAX with respect to time-cost analysis.

New kinetic procedure for the measurement of conjugated bilirubin with a centrifugal analyzer.

A procedure has been developed for the automated measurement of conjugated bilirubin in serum with a centrifugal analyzer. The conjugated bilirubin is measured by a fixed time kinetic method which monitors the reaction between conjugated bilirubin and diazotized sulfanilic acid at 550 nm. Results are calculated based on a comparison of the reagent blank-corrected absorbance changes between 15 seconds and 75 seconds for sample vs changes in an empirical standard. The standard used is N-(1-naphthyl) ethylene diamine dihydrochloride (NEDC) which reacts with diazotized sulfanilic acid at a rate comparable to conjugated bilirubin. The standard is calibrated by comparison with a modified Jendrassik and Grof procedure using a serum blank-corrected centrifugal analyzer reference method. The method is linear to 150 mg per 1 with a sensitivity of 3.0 milliabsorbance units per 1.0 mg per 1 of conjugated bilirubin using a 35 mul sample volume. Within-run precision is 1 percent for elevated concentrations of bilirubin. Hemolysis introduces a negative interference, the nature of which is discussed.

Centrifugal analyzer method for total bilirubin in serum by use of diazotized 2-chloroaniline-5-sulfonic acid.

We describe two centrifugal analyzer methods for measuring total bilirubin in serum. Diazotized 2-chloroaniline-5-sulfonic acid is used in both. In the first procedure, ethylene glycol and methanol are used as an accelerator solvent; results correlate well with those by a Jendrassik and Grof method adapted to the centrifugal analyzer, but there is considerable hemolysis interference. In the second method, dimethyl sulfoxide is used with the ethylene glycol/methanol solvent and almost all hemolysis interference is eliminated. For either method, a 15-microliter sample is required. In the second method, instrument response is linearly related to concentration to 250 mg/L and the within-run precision (CV) is about 1%.

Munchausen syndrome by proxy: definition of factitious bleeding in an infant by 51Cr labeling of erythrocytes.

The Munchausen syndrome by proxy is a phenomenon in which symptoms of a disease are fabricated by some person other than the patient. This report describes and 8-week-old infant with repetitive bleeding episodes, presumably originating from the upper respiratory tract. Extensive investigations, including angiography, several endoscopies under general anesthesia, and reinfusion of the infant's red cells labeled with 51Cr followed by pulmonary and upper airway scanning, failed to reveal the source of bleeding. Within two weeks after initiation of the 51Cr studies, radioactivity of facial blood from two separate bleeding episodes did not exceed background counts. Simultaneous examination of the infant's capillary blood, however, showed moderate to marked radioactivity. The Rh subtype of the facial blood was cc, whereas the infant's type was Cc. These findings indicated that the facial blood was factitious in origin. No further "bleeding" occurred after this information was presented to the parents. This case represents an unusual example of the Munchausen syndrome by proxy. Awareness of this entity can prevent potentially harmful investigations. Documenting its occurrence and sharing the information with parents in a nonaccusatory manner may prevent future harm to the patient.