Megakaryocyte function and dysfunction.

More than a hundred years have passed since platelets were recognized as cells and their haemostatic functions discovered. However, the process of platelet production is still not understood. The location, the mechanism and the regulation of thrombopoiesis remain elusive. Megakaryocytes are known to be the source of platelets. Investigations of megakaryocytes have revealed their normal functions and some of the abnormalities present in various diseases which affect platelets. In recent years, new techniques of cell isolation and tissue culture have been developed and have made possible advances in characterizing megakaryocyte precursors and differentiation. The primary function of megakaryocytes is to synthesize and assemble platelet components and organelles. Although debated for a long time, new data seems to indicate that the lung may be a central locus of platelet production. The new techniques for megakaryocyte investigations have barely begun to be of use in the study of abnormal platelet production in disease.

Structural changes in the megakaryocytes of patients infected with the human immune deficiency virus (HIV-1).

Although immune mechanisms are known to be partially responsible for the thrombocytopenia of patients infected with HIV-1, an understanding of the mechanism underlying this disorder is incomplete. A casual observation that bone marrow biopsies of HIV-infected individuals seem to exhibit an unusually large number of denuded megakaryocyte nuclei (DN-MK) prompted a study comparing MK of 20 HIV-seropositive individuals with those of 10 patients with HIV-negative idiopathic thrombocytopenic purpura and 10 hematologically normal subjects. In normal marrows the number of DN-MK average 2.1 +/- 0.5 SE per 10 low power field. In patients with ITP the average number was 6.5 +/- 1.4 SEM, whereas HIV-ITP marrows had an average of 42.5 +/- 3.7 SEM. Electron microscopy of AIDS megakaryocytes exhibited ballooning of the peripheral zone to an extent not seen by us in any other myelodysplastic syndromes. These observations support the concept that the pathophysiology affecting MK/platelets in HIV-infection should not be equated with the destructive process underlying other immune thrombocytopenias.

Heterogeneity of in vitro growth pattern of megakaryocyte progenitors (CFU-M) in myeloproliferative disorders.

In groups of 26 patients with myeloproliferative disorders (MPD), 8 with chronic myelogenous leukaemia (CML); 8 with polycythaemia vera (PV); 10 with essential thrombocythaemia (ET); and 6 patients with reactive thrombocytosis (RT), we studied the growth characteristics of bone marrow CFU-M in agar culture. The bone marrows from all the patients with MPD formed so called endogenous CFU-M colonies, in the absence of PHA-LCM, that increased in a dose-dependent manner with the addition of increasing concentrations of normal human AB-citrated plasma (NH-ABCP), while the bone marrows from all the patients with RT and from healthy controls formed few or no endogenous CFU-M colonies. In MPD, the endogenous CFU-M growth was enhanced by normal T cells in a dose-dependent fashion, and was decreased with the depletion of T cells from the marrow cells. These results suggest that the formation of endogenous CFU-M colonies is caused by hypersensitivity of CFU-M in MPD to NH-ABCP, which may contain a small amount of Meg-CSF, and/or by in vitro T cell stimulation. Among MPD, the endogenous CFU-M growth in ET was significantly lower than that of other MPD patients; however, the total number of ET CFU-M grown in the presence of PHA-LCM was the highest. These data show that the bone marrow CFU-M in MPD are heterogeneous with respect to in vitro growth pattern or sensitivity to exogenous Meg-CSF.

Studies of the bone marrow in polycythemia vera and the evolution of myelofibrosis and second hematologic malignancies.

The PVSG study is unique in that it is prospective and composed of 432 patients randomized to three treatment arms. This study also provides the opportunity for serial studies of numerous sequential biopsies. Large numbers of cases with sequential biopsies covering the entire long course are essential to appreciate the full spectrum of tissue changes in this disease. The PVSG was initiated in 1967 and in mid-1985 approximately one third of the patients are alive and on protocol. For these reasons, the results must still be considered preliminary. Pretreatment biopsies from patients randomized in the PVSG have been analyzed for total cellularity, megakaryocyte concentration, and reticulin content. Considerable variation in these elements was found in these biopsies. Sequential posttreatment biopsies from these patients have also been studied and correlated with the clinical course of the disease. None of the morphologic parameters analyzed was shown to be of prognostic significance. Early in the course of PV the marrow reticulin content is almost always normal. The length of the developmental stage is unknown and the precise timing of the clinical onset may be difficult. Therefore, the 11% of patients that showed a significant increase in reticulin on initial evaluation may have had PV longer than was indicated clinically. If large numbers of sequential biopsies are studied, an increase in reticulin content can frequently be demonstrated during the active phase of the disease and before the onset of the spent phase. Currently 39 patients (9%) have developed the spent phase, or PPMM. PPMM occurred in about the same incidence in the patients treated with myelosuppressive therapy as by phlebotomy alone, the spent phase occurring in 16 patients treated by phlebotomy alone, 11 with chlorambucil, and 12 with 32P. The course of the reticulin fibrosis is slowly progressive. There is some evidence for regression in a few patients in the erythrocytotic phase, but sampling variation cannot be completely ruled out. At this time in the study, AL has developed in 37 patients (8.6%). The incidence of AL is quite low in the phlebotomy group (three cases). Presumably this represents the natural incidence in PV unmodified by therapeutic agents. The frequency is approximately equal and quite high in the chlorambucil and 32P groups. There are 19 cases in the chlorambucil-treated group and 15 in the 32P-treated group. The leukemias that developed in the PV patients occurred either de novo or following PPMM.(ABSTRACT TRUNCATED AT 400 WORDS)

Altered platelet kinetics in thrombocytopenic mice with L5178Y leukemia.

Thrombocytopenia is a consistent feature of murine leukemia L5178Y. To define the mechanism(s) associated with the decrease in platelets, serial thrombokinetic studies were performed on various days after intravenous inoculation of 10(6) L5178Y ascites cells. At the nadir of thrombocytopenia (day 5), the recovery and circulating half-time of transfused normal 51Cr-platelets was only one half of control values. The loss of circulating radioactivity (70%) at 1 hr was accounted for by splenic (40%) and hepatic (30%) accumulation of labeled platelets. However, the liver contained three times more radioactivity per milligram of tissue than the spleen. There was no increase in hepatic accumulation of 59Fe-labeled red cells under the same conditions. When 51Cr-labeled leukemic platelets (day 3) were infused into normal animals, the circulating half-time was 50% of control. Despite spleen and liver enlargement, the blood volume of the leukemic mice did not increase. The megakaryocyte concentration remained unchanged after inoculation of leukemic cells, but the diameter of megakaryocytes increased significantly from days 5 to 10. These studies show that thrombocytopenia in mice transplanted with L5178Y leukemia occurs as a result of shortened platelet survival and increased organ sequestration. The increase in hepatic platelet accumulation suggests that the mechanism of thrombocytopenia is platelet specific and is not due to passive organ pooling.

A patient with apparent idiopathic A-megakaryocytic thrombocytopenia.

A 71-year-old woman developed life-threatening thrombocytopenia which resolved (probably spontaneously), recurred, and then again resolved. The thrombocytopenia was characterized by selective aphasia of megakaryocytes, near-normal platelet survival, and very slow recovery. No underlying disorder nor any drug could be incriminated, so we must assume this patient suffered from an idiopathic and very rare form of thrombocytopenia.

The relation of thrombokinetics to bone marrow megakaryocytes in idiopathic thrombocytopenic purpura (ITP).

In 23 patients with untreated idiopathic thrombocytopenic purpura (TP), the relation of thrombokinetics to quantitative determinations of megakaryocytes in bone marrow sections was studied. The megakaryocytes were classified into maturation stages, and platelet sizes were determined. Megarkaryocyte number and volume per microliter of bone marrow were significantly higher in ITP as compared to controls. The megakaryocyte number and volume were inversely related to the peripheral platelet count. Platelet production rate was significantly increased in ITP and related to the megakaryocyte number and volume. The megakaryocytes were shifted towards more immature forms in ITP, suggesting an increased turnover rate of the expanded recognizable metakaryocyte compartment. Platelet size was significantly increased in ITP, and the mean platelet diameter was 1.6 times normal. There was a significant relationship between platelet size and platelet production rate, as well as an inverse relationship between platelet size and platelet mean life-span (MLS). There was also a significant correlation between platelet size and the proportion of young megakaryocytes.