Moshe Mittelman

Agonists to the a3 adenosine receptor induce G-CSF production via NF-κb activation: A new class of myeloprotective agents

OBJECTIVEnThe aim of this study was to evaluate the effect of CF101, a synthetic agonist to the A3 adenosine receptor (A3AR), on the production of granulocyte colony-stimulating factor (G-CSF). The ability of CF101 to act as a myeloprotective agent in chemotherapy-treated mice was tested.nnnMETHODSnCF101 was administered orally to naïve mice and its effect was studied on blood cell counts (coulter counter), serum G-CSF level (ELISA), bone marrow colony-forming cells (soft agar culture), and splenocytes ability to produce ex vivo G-CSF. Protein extract was prepared from splenocytes and Western blot analysis was carried out to evaluate expression level of key proteins. In an additional set of experiments, CF101 was administered to mice 48 hours after cyclophosphamide treatment and blood cell counts as well as serum G-CSF levels were monitored.nnnRESULTSnOral administration of CF101 to naïve mice led to the elevation of serum G-CSF levels, an increase in absolute neutrophil counts (ANC), and bone marrow colony-forming cells. Splenocytes derived from these mice produced higher G-CSF level than controls. The molecular mechanisms underlying the events prior to G-CSF production included the upregulation of NF-kappaB and the upstream kinases phosphoinositide 3-kinase (PI3K), protein kinase B/Akt (PKB/Akt), and IKK. Accelerated recovery of white blood cells and neutrophil counts were observed in cyclophosphamide-treated mice following CF101 administration.nnnCONCLUSIONnCF101 induced upregulation of the PI3K/NF-kappaB pathway leading to G-CSF production, resulting in myeloprotective effect in cyclophosphamide-treated mice.

Erythropoietin has an anti-myeloma effect - a hypothesis based on a clinical observation supported by animal studies

Recombinant human erythropoietin (rHuEpo) was introduced into clinical practice more than a decade ago, and has been found to be effective in the treatment of several types of anemia, including anemia of end‐stage renal failure and cancer‐related anemia. No study has suggested that Epo might have an effect on the biology of the disease, nor any survival advantage to cancer patients treated with Epo for anemia has been reported. Here we report six patients with advanced multiple myeloma (MM) with very poor prognostic features, whose expected survival was <6u2003months. All six patients were treated with rHuEpo for their anemia, either without any chemotherapy or very mild chemotherapy for a short time. Yet, surprisingly they lived for 45–133u2003months totally from MM diagnosis and 38–94u2003months with rHuEpo (with a good quality of life). In fact, one patient, is still alive and well, more than 8u2003yr after chemotherapy was discontinued because of a resistant aggressive disease. The course in these six MM patients led us to hypothesize that Epo might have an antineoplastic or antimyeloma effect. We proceeded and tested that hypothesis in mouse models of myeloma (Mittelman M et al., Proc Natl Acad Sci USA 98:5181,2001). In these models we confirmed that rHuEpo induced tumor regression in about 50% of the BALB/c mice inoculated with MOPC‐315 myeloma cells. We then presented evidence that the mechanism is a new immune‐mediated phenomenon, via activation of CD8+ T cells. Furthermore, evidence from the literature supports the antineoplastic effect of Epo. Epo might be used as an adjunct immune treatment in various malignant diseases, in addition to the current regimens and chemotherapeutic protocols. Fututre trials should determine the role of Epo in myeloma and cancer treatment, besides clarifying concerns about the presence of Epo receptors on some tumor cells.

A Chemically Induced Rat Model of Hemolysis with Disseminated Thrombosis

Although hemolytic anemia and thrombosis, which can be serious or even lethal, are often encountered in daily common practice., their pathogenesis has remained obscure, partially because of the absence of appropriate models. Here we present a unique chemically induced rat model of hemolytic anemia and disseminated thrombosis in which the organs developing infarction are comparable to those seen in humans. We exposed male and female Fischer F344 rats to two, three, or four daily doses of 2-butoxyethanol (BE) at 250 mg/kg body weight and examined for hemolysis and histopathological evidence of disseminated thrombosis on d 2, 3, 4, and 29. Time-course BE-related erythrocytic changes were statistically significant in both sexes. Evidence of thrombosis and infarction was seen mainly in females dosed more than once with widespread thrombotic crisis after two or three dosings, likely explicable by the more significant morphological changes in erythrocytes and hemolysis observed in this gender. We documented thrombosis and infarction in the heart, brain, Jungs, eyes, and bones. Our model with its list of target organs similar to that observed in human diseases characterized by hemolysis and thrombosis [for example, thalassemia, sickle cell disease (SCD), paroxysmal nocturnal hemoglobinuria (PNF), disseminated intravascular coagulation (DIC), thrombotic thrombocytopenic purpura (TTP), and hemolytic uremic syndrome (HUS)] suggests that it can be an excellent tool to study the pathogenesis of such complications.

Ultrastructural observations on bone marrow cells of 26 patients with myelodysplastic syndromes.

Bone marrow aspirates from 26 patients with myelodysplastic syndrome (MDS) were examined using transmission electron microscopy. The red blood cell precursors in 9 patients showed varying degrees of dyserythropoiesis including the presence of 2 or more nuclei, nuclei with bizarre shape and iron deposits in the mitochondria. The myeloid series showed a tendency to hypogranulation (5 patients) and in 2 patients there were signs of platelet phagocytosis. The monocytes had a normal ultrastructure except for one patient with chronic myelomonocytic leukemia (CMML) with transformation to acute myelo-monocytic leukemia (AMML). In this case, the monocytes were immature, with markedly convoluted nuclei and scanty heterochromatin. The lymphocytes also had a normal appearance, except for one patient in whom the lymphocytes were immature, with lobulated nuclei and suggested transformation of MDS to acute lymphoblastic leukemia. The plasma cells in 3 patients were slightly increased in number and in one of them Russell bodies were seen both in the cytoplasm and the nucleus. The megakaryocytic series showed a shift to the left and in one patient there were signs of emperipolesis. The alterations in the hematopoietic cells in patients with MDS described in the present study indicate that the electron microscope may supplement light microscopic findings and help in the establishment of a correct diagnosis. This may be also evident in those cases of MDS in which the very early stages of leukemic transformation cannot be easily detected by light microscopy.

Ocular expression of vascular cell adhesion molecule (VCAM-1) in 2-butoxyethanol-induced hemolysis and thrombosis in female rats

We demonstrated previously that exposure of rats to 2-butoxyethanol (BE) was associated with morphological changes in red blood cells, hemolytic anemia, and disseminated thrombosis and infarction in different organs including the eyes. In order to elucidate the mechanism of thrombosis formation, we examined in this study the histology and immunohistochemical expression of vascular cell adhesion molecule-1 (VCAM-1), endothelial intercellular adhesion molecule-1 (ICAM-1), and P-selectin in the eyes of the female F344 rat exposed to 2, 3, or 4 daily doses of BE/250 mg/kg body weight. In this BE hemolysis and thrombosis model, positive VCAM-1 expression occurred only in eyes of rats exposed to 3 and 4 doses and was localized in the iris (epithelium lining the posterior surface, anterior mesenchymal epithelium), ciliary processes (lining epithelium, stromal cells), and retina (hypertrophic retinal pigment epithelium). Only weak immunolabeling was seen in eyes exposed to 2 doses. The appearance of VCAM-1 immunostaining correlated with the development of thrombosis located in the same structures. No change in ICAM-1 or P-selectin expression was seen. This immunolabeling distribution suggests that VCAM-1 functions in the pathogenesis of BE-related thrombosis by promoting adhesion of erythrocytes to the endothelium.

Disseminated thrombosis-induced growth plate necrosis in rat: a unique model for growth plate arrest.

Exposure of rats to 2-butoxyethanol (BE) produces early hemolytic anemia and disseminated thrombosis. This leads to infarctions in multiple organs, including bones and cartilage. BE, administered for different durations of exposure in two separate experiments, produced metaphyseal vascular thrombosis, growth plate infarction, and partial or complete physeal growth arrest. This reproducible model may serve as a useful tool in the study of some conditions that manifest growth plate damage. The suitability of this model for investigating the pathogenesis of growth plate necrosis and as a model for potential therapy for various human growth plate disorders are discussed.

Kaposi's sarcoma associated with Castleman's disease.

To the Editor: Angiofollicular lymph node hyperplasia (Castleman’s disease) is characterized by fever, weakness, weight loss, lymphadenopathy, anemia and hyperglobulinemia (1, 2); however, the histologic picture differs from that of classical lymphoma (3-5). Kaposi’s sarcoma (KS) is an angioproliferative disease with unknown etiology (6). The disease is common in males and usually presents as a skin lesion, but may also involve visceral organs such as lungs, gastrointestinal tract and lymph nodes. We report three patients with lymphoproliferative diseases, two with Castleman’s disease and the other one with non-Hodgkin’s lymphoma, who developed HIV negative KS. The histologic evaluation revealed highly vascular hyperplasia resembling the findings in the lymph node biopsies and suggesting a possible association between KS and lymphoproliferative disorders.