Micrometastasis in colorectal carcinoma: a review.

Lymph node metastasis is the most important predictor of prognosis, after surgery, in colorectal carcinoma. The term "micrometastasis" has evolved from a morphological definition to one that is used with molecular-based techniques. We review the literature to evaluate the significance of detecting micrometastases in colorectal carcinoma, either by morphological or molecular techniques, and address technical difficulties encountered with both. Routine use of immunohistochemistry is not recommended as most studies show little change in staging or prognosis. Radioimmunoguided surgery may prove beneficial, but problems of false positives in benign diseases need to be addressed. Immunohistochemical detection of micrometastatic deposits in bone marrow aspirates holds the most promise for clinical practice. Molecular techniques are more sensitive than immunohistochemistry, but prognostic value needs to be determined. Molecular diagnostics can also determine genetic alterations and mutations that should improve our understanding of metastatic colon cancer and staging accuracy.

Apoptosis of gamma/delta T cells in human ehrlichiosis.

Expansion of activated T cells expressing the T-cell receptor (TCR) gamma/delta, CD45RO, and HLA-DR antigens is a prominent feature of acute infection with Ehrlichia chaffeensis in humans. The fate of these activated cells and the resolution of the gamma/delta T-cell response with return to the usual alpha/beta T-cell populations in this disease are not clearly understood. At a morphologic level, apoptotic cells are present in the peripheral blood during the acute and resolution phases of the infection. Simple culture of density gradient-separated lymphocytes from the blood of patients with acute ehrlichiosis produced cell death rapidly in the media compared to alpha/beta T cells. This loss of viability after incubation was apparently mediated by apoptosis, based on flow cytometric and morphologic analyses. The results suggest that most primed (CD45RO+) and activated (HLA-DR+) gamma/delta T cells in acute ehrlichiosis might be subject to removal from the body by programmed or apoptotic cell death.

Lymphocytosis of gamma/delta T cells in human ehrlichiosis.

The majority of T cells in peripheral blood express a T-cell receptor (TCR) comprised of alpha and beta chains. An alternate form of the TCR is comprised of gamma and delta chains. These gamma/delta T cells are associated with certain infectious lesions, and modestly elevated in peripheral blood in certain disease states. Human ehrlichiosis is characterized by hematologic abnormalities including multi-lineage cytopenias. In most cases reported, a lymphocytopenia has been present either at diagnosis, or at some time during the illness. Early in the course of antibiotic treatment (48-72 hours), the lymphocytopenia corrects itself and is rapidly followed by a lymphocytosis of T cells that express CD3, but are negative for CD4 and CD8, as well as the major form of the TCR formed by the alpha/beta heterodimer. Instead, these CD3+4-8- T cells express the gamma/delta heterodimer associated with V gamma 9 and V delta 2 chains, a population of cells usually the distinctive minority of peripheral blood T cells, but constituting the major phenotype of peripheral gamma/delta T cells.

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.

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.

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.

Acute leukemia/lymphoma of plasmacytoid T-cell type.

Plasmacytic morphologic characteristics are usually associated with cells of B-lymphocyte origin. Recently, plasmacytoid T-cells have been described in reactive lymph nodes and a rare form of lymphoma characteristically associated with myeloproliferative disorders. This report documents a case of plasmacytoid T-cell malignancy that initially presented as an acute leukemia in an elderly man with a longstanding myelodysplastic syndrome. The tumor replaced bone marrow and involved lymph nodes. Despite aggressive therapy, he died quickly of his leukemia/lymphoma. This case illustrates the need for complete cellular analysis in the diagnosis of morphologically plasmacytic malignancies and raises additional questions about the relationship of this peculiar type of T-cell to the hematopoietic marrow.

Myelodysplastic syndrome: prospective evaluation of fifty-one patients using the Dutcher scoring system.

Fifty-one patients with primary myelodysplastic syndrome were prospectively evaluated using a scoring system based on the presentation blood and bone marrow findings. Twenty-four patients (47%) evolved to acute nonlymphocytic leukemia. Stepwise regression model showed that the scoring system was the only significant variable for predicting transformation to acute leukemia (p = 0.0007, sensitivity 70.8%, specificity 77.8%). Seventy-six percent of patients with a score of 14 or greater developed acute leukemia compared to 19% with a score of 13 or less. Median survival of the entire group was 10 months. The most important prognostic factor for predicting survival was the scoring system (p = 0.0001). Survival correlated inversely with the score. This scoring system may be useful in the management of patients with myelodysplasia.

Hemolytic disease of the newborn due to anti-C and anti-G masquerading as anti-D.

After several transfusions with D-negative blood, an O Rh-negative women was apparently sensitized to the C and D antigens. In her prenatal workup, it became evident that she had in fact not been sensitized to D but to G, which initially appeared as anti-D plus anti-C. This sensitization pattern is an unusual occurrence in itself. Moreover, the fetus was affected significantly and was delivered at 32 1/2 weeks with moderate hemolytic disease of the newborn. Consideration is given to points regarding current methods of screening Rh-negative women for transfusion, the use of anti-Rh immune globulin in patients sensitized to anti-C and anti-G, medicolegal implications, and continuous attention to the risk-benefit ratio in decisions to use transfusions.

Pentazocine-induced agranulocytosis.

A 46-year-old man with pentazocine-induced agranulocytosis is described. In previously reported cases of a complete absence of mature neutrophils in the peripheral blood and bone marrow the patients had undergone marrow-depressing treatment with radiation and antineoplastic drugs. This case is unique in that the patient had complete agranulocytosis without predisposing factors.

A simplified micro ELISA procedure for the measurement of platelet-associated IgG (PAIgG).

A simplified micro ELISA procedure to measure platelet-associated IgG is described. The platelet-bound IgG first is extracted into the fluid phase by solubilizing washed platelets in 0.1% triton X-100. The solubilized IgG in the extract and IgG standards are incubated in microtiter wells previously coated with antihuman IgG. The IgG in the standards and extract bind to the solid phase antihuman IgG. The bound IgG then is measured by the addition of peroxidase labeled antihuman IgG and appropriate substrate. With this method platelets from normal controls were found to have 1.7 +/- 0.6 fg IgG/platelet (mean +/- SD). Platelets from patients with ATP had values that were two to seven times the control values. The relative advantage of this technic is discussed.

In vitro suppression of myelopoiesis by adherent murine splenocytes in experimental disseminated histoplasmosis.

Supernatant culture media obtained from adherent spleen cell preparations of mice experimentally infected with Histoplasma capsulatum yeast cells suppressed in vitro growth of human myeloid cells. This suppression was significantly greater than that obtained when splenocytes from normal mice or those inoculated with killed yeast cells were used. The use of indomethacin partially blocked this suppressive effect. A direct relationship was observed between the levels of prostaglandin E (PGE) per 10(6) adherent cells in the splenocyte preparations and the degree of in vitro myelosuppression. These findings suggest a possible mechanism for the leukopenia frequently observed in disseminated histoplasmosis.

Preservation of hematopoietic stem cells in cadaveric marrow.

CFU-C assays were performed on unselected cadavers of various postmortem intervals to determine the survival of hemopoietic stem cells in cadaveric marrows. Successful growth was achieved in eight of ten cases up to 11 hours post mortem. The two marrows in which growth was unsuccessful were from patients with underlying diseases that would have made growth impossible even if the marrow was obtained during life. Six cases in which marrow was obtained 11 to 23 hours post mortem showed no proliferative capacity. The average CFU-C in the eight cases with growth activity was 25.8 +/- 16 compared with 40.6 +/- 35 for 61 hospital patients for whom antemortem marrow cells were available for culture. These two meanings are not significantly different statistically. These findings demonstrated the persistence of hemopoietic stem cells in cadaveric marrows, showing the cadaver to be a potentially abundant source of these cells.

Significant muscle capillary basement membrane thickening in spontaneously diabetic Mystromys albicaudatus.

Skeletal muscle capillary basement membrane thickness was determined in age- and sex-matched normal and spontaneously diabetic Mystromys albicaudatus. In nondiabetic animals the average basement membrane thickness was 482.6+/-48.7 A as against 779.0+/-319.9 A in the diabetic ones. This difference was statistically significant at P less than 0.005.