Translocation of CD45RA in neutrophils.

The CD45 family of high-Mr glycoprotein antigens is expressed in some molecular form on all lymphohematopoietic cells. Different cell types express various isoforms in a precisely programmed fashion. In addition to cell surface CD45 antigens, we recently demonstrated a cytoplasmic granule-associated pool of CD45RA, the highest Mr isoform, in mature neutrophils that is generally absent from the cell surface under nonstimulatory conditions. Under such conditions, the major cell surface form is CD45RO, the low-Mr isoform. In the present study, we demonstrate the ability of calcium ionophore A23187 to induce translocation of cytoplasmic CD45RA to the cell surface as well as to increase the cell surface expression of CD45RO and CD45. This process was calcium dependent and rapid, occurring within 5 min. A series of experiments using chemical antagonists of protein kinases suggest that this up-regulation may be mediated via the calmodulin system rather than via protein kinase C. Although the exact function(s) of the various isoforms of CD45 is not known, this translocation suggests a role for CD45RA in neutrophil activation.

The pathophysiology of thrombosis in cancer.

Multiple mechanisms are involved in the pathophysiology of thrombosis in cancer. It is highly likely that, in an individual patient, a multifactorial etiology is operative. The three basic mechanisms for hypercoagulability in cancer are interrelated to a degree and probably are also the mechanisms involved in metastasis in addition to thrombosis. Manipulation of coagulation in the cancer patient has the potential to prevent complications, abrogate metastasis, and potentially prolong survival.

Alterations of HLe-1 (T200) fluorescence intensity on acute lymphoblastic leukemia cells may relate to therapeutic outcome.

Quantitative differences in HLe-1 expression were studied on normal lymphocytes and lymphoblasts of patients with acute lymphoblastic leukemia (ALL). A relationship was found between quantitative antigen expression and therapeutic outcome. Alterations in fluorescence intensity (FI) were demonstrated using quantitative flow cytometric methods and the monoclonal antibody (MoAb) anti HLe-1 (T200). Lymphoblasts from patients with ALL produced FI peaks ranging form channel 17 to 112 (mean 68; n = 28), while normal lymphoid cells were at FI channel 127 +/- 3 (n = 121). Patients with the dimmest-staining lymphoblasts (channels 17-50) responded better to therapy than those with brighter-staining cells (channels 50-100). Data from this pilot study suggests that the FI of malignant lymphoblasts has implications in the clinical response to therapy.

Surface and cytoplasmic expression of CD45 antigen isoforms in normal and malignant myeloid cell differentiation.

The CD45 family contains protein tyrosine phosphatase (PTPase) activity and is expressed in one or more of its isoforms on all lymphohematopoietic cells. Considerable work has focused on CD45 expression by lymphoid cells, but minimal work has involved granulocytes. Granulocytic, or myeloid, cell differentiation is accompanied by a number of morphologic and immunophenotypic changes. This study used flow cytometric and immunocytochemical methods in conjunction with morphologic assessment to investigate the expression of CD45 isoforms during differentiation of normal and malignant granulocytic cells. On normal bone marrow cells, the quantity of surface CD45 did not change during earlier stages but did increase significantly at the terminal stages (bands and polymorphonuclear leukocytes [PMNs]). CD45RO (the low relative molecular mass [Mr] isoform) was very dimly expressed on immature cells but became increasingly brighter beginning at approximately the myelocyte stage. The high Mr isoform (CD45RA) was virtually absent from the cell surface at all stages. Only a small percentage (3-15%) of PMNs expressed surface CD45RA. However, there was a cytoplasmic pool of each isoform associated with membrane-bound granules found throughout differentiation, with remarkable increases in expression at the terminal stages. In the case of acute myeloid leukemias (AMLs), most cases expressed surface CD45RA with, or without, CD45RO, regardless of their French-American-British (FAB) classification. This appeared to be a stable process at diagnosis and relapse in individual patients and may therefore serve as a diagnostic aid. The biologic significance of this aberrant expression of CD45RA by malignant cells is unknown but raises important questions regarding the cellular processes of phosphorylation/dephosphorylation in normal and malignant cells.

Climatic forcing of carbon-oxygen isotopic covariance in temperate-region marl lakes.

Carbon and oxygen stable isotopic compositions of lacustrine carbonate from a southeastern Michigan marl lake display linear covariance over a range of 4.0% Peedee belemnite (PDB) in oxygen and 3.9% (PDB) in carbon. Mechanisms of delta 13 C-delta 18 O coupling conventionally attributed to lake closure in arid-region basins are inapplicable to hydrologically open lake systems. Thus, an alternative explanation of isotopic covariance in temperate region dimictic marl lakes is required. We propose that isotopic covariance is a direct record of change in regional climate. In short-residence-time temperate-region lake basins, summer meteoric precipitation is enriched in 18O relative to winter values, and summer organic productivity enriches epilimnic dissolved inorganic carbon in 13C. Thus, climate change toward longer summers and/or shorter winters could result in greater proportions of warm-month meteoric precipitation, longer durations of warm-month productivity, and net long-term enrichment in carbonate 18O and 13C. Isotopic covariance observed in the Michigan marl lake cores is interpreted to reflect postglacial warming from 10 to 3 ka followed by cooler mean annual temperature, a shift toward greater proportions of seasonal summer precipitation, a shortening of the winter season, or some combination of these three factors.

Oxygen isotope analysis of phosphate: improved precision using TC/EA CF-IRMS.

Oxygen isotope values of biogenic apatite have long demonstrated considerable promise for paleothermometry potential because of the abundance of material in the fossil record and greater resistance of apatite to diagenesis compared to carbonate. Unfortunately, this promise has not been fully realized because of relatively poor precision of isotopic measurements, and exceedingly small size of some substrates for analysis. Building on previous work, we demonstrate that it is possible to improve precision of delta18O(PO4) measurements using a 'reverse-plumbed' thermal conversion elemental analyzer (TC/EA) coupled to a continuous flow isotope ratio mass spectrometer (CF-IRMS) via a helium stream [Correction made here after initial online publication]. This modification to the flow of helium through the TC/EA, and careful location of the packing of glassy carbon fragments relative to the hot spot in the reactor, leads to narrower, more symmetrically distributed CO elution peaks with diminished tailing. In addition, we describe our apatite purification chemistry that uses nitric acid and cation exchange resin. Purification chemistry is optimized for processing small samples, minimizing isotopic fractionation of PO4(-3) and permitting Ca, Sr and Nd to be eluted and purified further for the measurement of delta44Ca and 87Sr/86Sr in modern biogenic apatite and 143Nd/144Nd in fossil apatite. Our methodology yields an external precision of +/- 0.15 per thousand (1sigma) for delta18O(PO4). The uncertainty is related to the preparation of the Ag3PO4 salt, conversion to CO gas in a reversed-plumbed TC/EA, analysis of oxygen isotopes using a CF-IRMS, and uncertainty in constructing calibration lines that convert raw delta18O data to the VSMOW scale. Matrix matching of samples and standards for the purpose of calibration to the VSMOW scale was determined to be unnecessary. Our method requires only slightly modified equipment that is widely available. This fact, and the demonstrated improvement in precision, should help to make apatite paleothermometry far more accessible to paleoclimate researchers.

Fluorescence intensity of immunostained cells as a diagnostic aid in lymphoid leukemias.

The standardized fluorescence intensity (FI) of immunostained normal and malignant B-cells was measured as a method of evaluating cellular heterogeneity between lymphoid malignancies. Mature B-cells from peripheral blood lymphocytes (PBL) of different individuals demonstrated characteristic peaks of FI when specific monoclonal antibodies (MoAbs) were employed in standard flow cytometric procedures. Malignant cells from patients with lymphoid leukemias demonstrated FI that differed from that of normal B-cells with various MoAbs and that differed among categories of leukemia. By using a panel of MoAbs reactive with B-cells, as well as a T200 (CD45) antigen, a scheme of malignant cell differentiation may be produced that approximates stages of normal B-cell differentiation. When the FI of malignant cells differs from that of normal cells, or when atypical peaks (or additional peaks) of FI are present in flow cytometric histograms, the investigator should be alerted to the probability of abnormal cell populations. In addition, it is frequently possible to use this information to help classify malignancies, thereby contributing to identification of small numbers of malignant B-cells in otherwise equivocal situations.

Relationship between T200 antigen expression and stages of B cell differentiation in resurgent hyperplasia of bone marrow.

Using monoclonal antibodies (MoAbs) and dual-parameter flow cytometric techniques, bone marrow mononuclear cells (MMC) from patients with resurgent hyperplasia were analyzed for their coexpression of HLe-1 (T200) and antigens normally associated with particular stages of B cell differentiation. The marrow from those with resurgent hyperplasia contained increased numbers of B cell precursors in multiple stages of differentiation compared to controls, thus providing a useful model system for studies of B cell differentiation. These studies indicate that the quantitative expression of T200 is differentiation-related on normal and malignant B cells and B cell precursors. Immature cells express low amounts of T200, while increasing levels of maturity correlated with increasing amounts of the antigen. This study increases the understanding of relationships between B cell surface antigens and T200 and further demonstrates that B cell hyperplasia occurs commonly in association with bone marrow reactive or resurgent processes. The quantitative, rather than only the qualitative, expression of T200 is therefore a useful marker of B cell differentiation in reactive hyperplasia and in further investigation of B cell malignancy.

Ifosfamide-induced renal tubular defects.

Unexpected nephrotoxicity has been described in high-dose, bolus ifosfamide (IFF) therapy. Renal injury is not thought to occur in patients receiving fractionated schedules, although microscopic hematuria from bladder irritation is not uncommon. IFF is undergoing trials in patients with malignant lymphomas, gynecologic malignancies, and advanced sarcomas and has shown promising results. This report describes renal abnormalities in four patients with malignant lymphoma receiving single-agent, fractionated IFF and suggests a proximal tubular defect in two patients who were studied in greater detail. These findings suggest an unreported and unique toxicity of IFF when given in smaller, fractionated doses.

Relationship between CD45 antigen expression and putative stages of differentiation in B-cell malignancies.

The cell-surface antigen CD45 is a complex family of high-molecular-weight glycoproteins expressed on all lymphohematopoietic cells, but not in the same molecular isoform. This antigen complex is known to exhibit protein tyrosine phosphatase (PTPase) activity and appears to have a role in regulation of cell differentiation. In that CD45 expression parallels stages of differentiation in normal bone marrow B cells, it was of interest to evaluate this process in malignant B cells. Monoclonal antibodies (MoAbs) were used to investigate the quantitative expression of CD45 and CD45RA on the B cells of lymphoid leukemias. Employing standardized flow cytometric methods, it was found that the fluorescence intensity (FI) of immunostained malignant B cells, as a reflection of the antigen content, demonstrated correlations with the putative stage of cell differentiation for malignancies at the earlier stages, but at the later stages, a progressive loss of CD45 was observed. Since this antigen family has been found to display PTPase activity, further investigation of CD45 alterations in malignancies may provide insight into potential regulatory disturbances.

In vivo upregulation of CD45RA in neutrophils of acutely infected patients.

The CD45 family of high relative molecular mass (M(r)) cell surface antigens is expressed on all lymphohematopoietic cells. Different cell types express various M(r) isoforms in a precisely programmed fashion. The cytoplasmic domain of the CD45 family possesses protein tyrosine phosphatase activity and is involved in transmembrane signaling. Under normal conditions, the major CD45 isoform on neutrophils is CD45RO, the low M(r) isoform. We recently demonstrated a cytoplasmic granule-associated pool of CD45RA, the highest M(r) isoform, in mature neutrophils that is generally absent from the cell surface under nonstimulatory conditions. Calcium ionophore A23187 induced translocation of cytoplasmic CD45RA to the cell surface and increased the cell surface expression of CD45RO and CD45. The present study demonstrates that without in vitro stimulation, there is an increase in expression of CD45RA on the surface of neutrophils from patients with systemic, acute infections, while those of patients with localized infections are only slightly increased compared to normal controls. Although the exact functions of the various isoforms of CD45 are not known, this increased expression of CD45RA suggests a role for this high M(r) isoform during in vitro neutrophil activation.

Cooler winters as a possible cause of mass extinctions at the Eocene/Oligocene boundary.

The Eocene/Oligocene boundary, at about 33.7 Myr ago, marks one of the largest extinctions of marine invertebrates in the Cenozoic period. For example, turnover of mollusc species in the US Gulf coastal plain was over 90% at this time. A temperature change across this boundary--from warm Eocene climates to cooler conditions in the Oligocene--has been suggested as a cause of this extinction event, but climate reconstructions have not provided support for this hypothesis. Here we report stable oxygen isotope measurements of aragonite in fish otoliths--ear stones--collected across the Eocene/Oligocene boundary. Palaeo-temperatures reconstructed from mean otolith oxygen isotope values show little change through this interval, in agreement with previous studies. From incremental microsampling of otoliths, however, we can resolve the seasonal variation in temperature, recorded as the otoliths continue to accrete new material over the life of the fish. These seasonal data suggest that winters became about 4 degrees C colder across the Eocene/Oligocene boundary. We suggest that temperature variability, rather than change in mean annual temperature, helped to cause faunal turnover during this transition.

Hematologic effects of heavy metal poisoning.

Heavy metal poisoning can cause a variety of hematologic disorders. Exposure to heavy metals is ubiquitous in the industrial environment and must be considered in the differential diagnosis of many types of anemia. The heavy metals most commonly associated with hematologic toxicity are arsenic and its derivative arsine, copper, gold, lead, and zinc. A few distinctive clinical features characterize the hematologic manifestations of many occult heavy metal poisonings. These features have a limited differential diagnosis. A knowledge of these clinical features can assist the astute clinician in making the correct diagnosis.

A randomized trial to compare intravenous and oral etoposide in combination with cisplatin for the treatment of small cell lung cancer.

In a randomized multi-center study, 83 patients with small cell lung cancer were randomly assigned to treatment with cisplatin 100 mg/m2 intravenously (IV) day 1 and etoposide 120 mg/m2 IV days 1, 2, and 3 or cisplatin 100 mg/m2 IV day 1 and etoposide 120 mg/m2 IV day 1 and 240 mg/m2 orally days 2 and 3. Both regimens were repeated every 4 weeks. Prior to randomization, patients were stratified by extent of disease, performance status, and gender. A total of 41 patients were randomly assigned to the parenteral treatment only regimen, and 42 patients received cisplatin and IV/oral etoposide therapy. Both treatment arms were comparable regarding patient characteristics. Limited disease (LD) patients constituted 52% and 49% of the patient population for the oral and IV etoposide regimens, respectively. The overall complete response (CR) and partial response (PR) rate was 50% (95% confidence interval [CI] 35% to 65%) for the oral etoposide regimen and 59% (95% CI 44% to 74%) for the IV etoposide regimen (P = 0.438). For both regimens, 55% of the LD patients achieved either CR or PR. Time to progression and survival were comparable for both treatment arms. Hematologic toxicity was comparable in both treatment arms, with 80% of patients experiencing grade 3 or 4 neutropenia or thrombocytopenia. Moderate to severe anemia and weight loss were more predominant with the IV than with the oral regimen.