Hematopoietic and endothelial progenitor cell trafficking in patients with myeloproliferative diseases.

BACKGROUND AND OBJECTIVES: The presence of circulating hematopoietic progenitor cells in patients with myeloproliferative diseases (MPD) has been described. However, the exact nature of such progenitor cells has not been specified until now. The aim of this work was to investigate the presence of endothelial precursor cells in the blood of patients with MPD and to assess the role of the endothelial cell lineage in the pathophysiology of this disease. DESIGN AND METHODS: Endothelial progenitor cell marker expression (CD34, prominin (CD133), kinase insert domain receptor (KDR) or vascular endothelial growth factor receptor 2 (VEGFR2), and von Willebrand factor) was assessed in the blood of 53 patients with MPD by quantitative polymerase chain reaction. Clonogenic stem cell assays were performed with progenitor cells and monocytes to assess differentiation towards the endothelial cell lineage. The patients' were divided according to whether they had essential thrombocythemia (ET, n=17), polycythemia vera (PV, n=21) or chronic idiopathic myelofibrosis (CIMF, n=15) and their data compared with data from normal controls (n=16) and patients with secondary thrombo- or erythrocytosis (n=17). RESULTS: Trafficking of CD34-positive cells was increased above the physiological level in 4/17 patients with ET, 5/21 patients with PV and 13/15 patients with CIMF. A subset of patients with CIMF co-expressed the markers CD34, prominin (CD133) and KDR, suggesting the presence of endothelial precursors among the circulating progenitor cells. Clonogenic stem cell assays confirmed differentiation towards both the hematopoietic and the endothelial cell lineage in 5/10 patients with CIMF. Furthermore, the molecular markers trisomy 8 and JAK2 V617F were found in the grown endothelial cells of patients positive for trisomy 8 or JAK2 V617F in the peripheral blood, confirming the common clonal origin of both hematopoietic and endothelial cell lineages. INTERPRETATION AND CONCLUSIONS: Endothelial precursor cells are increased in the blood of a subset of patients with CIMF, and peripheral endothelial cells bear the same molecular markers as hematopoietic cells, suggesting a primary role of pathological endothelial cells in this disease.

Mast cells in ongoing acute rejection: increase in number and expression of a different phenotype in rat heart transplants.

BACKGROUND: Mast cells (MC) are resident in healthy hearts and play important physiological and pathophysiological roles. In the transplanted heart, correlations have been found between MC number and the severity of rejection episodes, the intensity of chronic inflammation, and allograft arteriosclerotic changes. However, not much emphasis has been placed on the fact that resident donor MC, and infiltrating recipient MC do not forcedly need to share the same properties and function. To gain insight in the role of cardiac MC during acute, and ongoing acute rejection of heart transplants, we investigated MC kinetics and MC phenotype in a rat heart transplantation model. METHODS: Donor hearts from female Brown-Norway rats were transplanted to male Lewis rats. Immunosuppression was started at day 5 using ciclosporin and prednisolone. Connective tissue type MC (CTMC) were distinguished from mucosa type MC (MMC) by immunohistochemistry for rat MC protease (RMCP) -1 and -2. Expression of RMCP-1 and -2 mRNA was quantified by real time reverse transcription-polymerase chain reaction. Infiltrating Y chromosome positive MC were detected by fluorescence in situ hybridization. mRNA expression of interleukin-3 (IL-3) and of the two differentially spliced isoforms of kit ligand (KL, stem cell factor) was quantified using reverse transcription-polymerase chain reaction. RESULTS: Resident cardiac donor MC are almost exclusively CTMC and decrease in number during acute rejection. MC increase in number, and recipient MC invade the cardiac allograft during ongoing acute rejection. The phenotype of the invading MC is characterized by the expression of RMCP-2, or both RMCP-1 and RMCP-2, and thus resemble a MMC type. IL-3 mRNA is highly expressed, and the ratio of the differentially spliced mRNAs for KL-1 and KL-2 rises up to 2-fold during ongoing acute rejection. CONCLUSIONS: Our data show that MC in posttransplant hearts during ongoing acute rejection differ from MC in healthy hearts and isografts by expressing a different phenotype. Changes in IL-3 and KL expression might be responsible for the predominance of MMC over CTMC. The notion is of importance that MC in cardiac allografts may have properties and functions that differ from those in nontransplanted healthy hearts.

Quantitative expansion of structural genomic alterations in the spectrum of neuroendocrine lung carcinomas.

The pathogenesis and interrelationships of neuroendocrine lung carcinomas are not well understood. Tissue macro-arrays prepared from surgical resection specimens from 35 patients with typical carcinoid (TC), six with atypical carcinoid (AC), 13 with large cell neuroendocrine carcinoma (LCNEC), and 15 with small cell lung carcinoma (SCLC) were investigated by fluorescence in situ hybridization (FISH) and immunohistochemistry. Hybridizations with locus-specific DNA probes demonstrated a high incidence of deletion for the tumour suppressor genes p53 and retinoblastoma (Rb), and for the oncogene cyclin D1, comparable in all carcinoma types. Similarly, an increase of DNA copy number for the Her-2/neu and c-myc oncogenes was noted in all neoplasms. A more detailed quantitative analysis of the results, however, demonstrated increasing numbers of cells harbouring these genomic alterations, from low-grade TC to highly malignant SCLC, with the exception of cyclin D1 deletion. Mutations of the p53 and Rb genes, as assayed by immunohistochemical studies, were observed at high incidence in high-grade carcinomas, compared with a low incidence in the low-grade carcinomas. Conversely, in all carcinoma types, neither membrane-bound Her-2/neu nor nuclear cyclin D1 was detected. It is concluded that structural genomic alterations are frequent in neuroendocrine lung carcinomas and that their occurrence may be underestimated by immunohistochemical studies alone. The quantitative expansion of the Rb, p53, c-myc, and Her-2/neu alterations towards high-grade carcinomas suggests common pathogenetic mechanisms in the spectrum of these neoplasms.