Label-free imaging and identification of typical cells of acute myeloid leukaemia and myelodysplastic syndrome by Raman microspectroscopy.

In clinical practice, the diagnosis and classification of acute myeloid leukaemia (AML) and myelodysplastic syndrome (MDS) start from the manual examination of stained smears of bone marrow (BM) and peripheral blood (PB) by using an optical microscope. This step is subjective and scarcely reproducible. Therefore, the development of subjective and potentially automatable methods for the recognition of typical AML/MDS cells is necessary. Here we have used Raman spectroscopy for distinguishing myeloblasts, promyelocytes, abnormal promyelocytes and erhytroblasts, which have to be counted for a correct diagnosis and morphological classification of AML and MDS. BM samples from patients affected by four different AML subtypes, mostly characterized by the presence of the four subpopulations selected for this study, were analyzed. First, each cell was scanned by acquiring 4096 spectra, thus obtaining Raman images which demonstrate an accurate description of morphological features characteristic of each subpopulation. Raman imaging coupled with hierarchical cluster analysis permitted the automatic discrimination and localization of the nucleus, the cytoplasm, myeloperoxidase containing granules and haemoglobin. Second, the averaged Raman fingerprint of each cell was analysed by multivariate analysis (principal component analysis and linear discriminant analysis) in order to study the typical vibrational features of each subpopulation and also for the automatic recognition of cells. The leave-one-out cross validation of a Raman-based classification model demonstrated the correct classification of myeloblasts, promyelocytes (normal/abnormal) and erhytroblasts with an accuracy of 100%. Normal and abnormal promyelocytes were distinguished with 95% accuracy. The overall classification accuracy considering the four subpopulations was 98%. This proof-of-concept study shows that Raman micro-spectroscopy could be a valid approach for developing label-free, objective and automatic methods for the morphological classification and counting of cells from AML/MDS patients, in substitution of the manual examination of BM and PB stained smears.

Molecular characterization of a novel endonuclease (Xib) and possible involvement in lysosomal glycogen storage disorders.

We cloned and partially characterized a human endonuclease (Xib) which shows sequence homologies to pancreatic DNase I but an enzymatic activity closer to DNase II. We report on the structural differences found between Xib and other recently cloned human DNases. Fluores cence microscopy analysis of transiently transfected cells with Xib::pEGFP constructs indicate that the protein is located in the cytoplasm and possibly anchored to a membrane, as deduced from a hydrophobic amino acid stretch present at the C-terminal end. Xib is overexpressed in muscle and cardiac tissues and is alternately spliced in several normal and neoplastic cells. In situ hybridization studies using human cardiac and muscle biopsies indicate accumulation of Xib transcript in the vacuoles of muscle cells from patients affected by vacuolar myopathy as acid maltase deficiency; however, no point mutations were detected in their DNA.

[Castleman's disease. A clinical case]. / La malattia di Castleman. Un caso clinico.

Castleman's disease is an uncommon lymph node syndrome. It is usually a lymphoid hamartoma, rarely systemic. We report a case of Castleman's multicentric disease observed in our hospital and review the literature. In this work we review prognosis, follow-up and therapy of 46 multicentric disease reported in the literature.

[Exfoliative cytology of psoriasis: effects of cyclosporin A]. / Citologia esfoliativa della psoriasi: effetti della ciclosporina A.

The cytological modifications of the horny layer cell population in psoriatic subjects during treatment with Cyclosporin A are described. They are studied by means of the cytoexfoliative method. The value of this technique is discussed, using it either for diagnosis, or for evaluating therapeutic efficacy of various treatments. The cytological findings are respectively compared with those of other psoriasis therapies, such as Methotrexate and RO 10-9359 (etretinate).