JAK2 V617F tyrosine kinase mutation in cell lines derived from myeloproliferative disorders.

A mutation in the JH2 pseudokinase domain of the Janus kinase 2 gene (JAK2 V617F) has been described in chronic myeloproliferative disorders (MPD). We screened 79 acute myeloid leukemia (AML) cell lines and found five positive for JAK2 V617F (HEL, MB-02, MUTZ-8, SET-2, UKE-1), 4/5 with histories of MPD/MDS. While SET-2 expressed both mutant (mu) and wild-type (wt) JAK2, remaining positives carried homo-/hemizygous JAK2 mutations. Microsatellite analysis confirmed losses of heterozygosity (LOH) affecting the JAK2 region on chromosome 9p in MB-02, MUTZ-8 and UKE-1, but also in HEL, the only JAK2mu cell line lacking any reported MPD/MDS history. All five JAK2mu cell lines displayed cytogenetic hallmarks of MDS, namely losses of 5q or 7q, remarkably in 4/5 cases affecting both chromosomes. Our combined FISH and microsatellite analysis uncovered a novel mechanism to supplement mitotic recombination previously proposed to explain JAK2 LOH, namely chromosome deletion with/without selective JAK2mu amplification. Confirming the importance of the mutated JAK2 protein for growth and prevention of apoptosis, JAK2mu cell lines displayed higher sensitivities to JAK2 inhibition than JAK2wt cell lines. In summary, JAK2 V617F cell lines, derived from patients with history of MPD/MDS, represent novel research tools for elucidating the pathobiology of this JAK2 mutation.     

Peripartale Kardiomyopathie

Peripartum cardiomyopathy (PPCM) is a rare and potentially life-threatening heart disease of unknown etiology that is restricted to the last month of pregnancy and up to 6 months postpartum. The morphology similates that of idiopathic dilated cardiomyopathy with a rapid progression to end-stage heart failure and is associated with a high morbidity and mortality rate. PPCM is treated with standard therapy for heart failure in accordance with the current guidelines. New findings suggest that elevated oxidative stress in the postpartum phase promotes cleavage of the nursing hormone prolactin into a toxic 16-kDa fragment that affects microcirculation and cardiomyocyte metabolism. Thus prolactin blocking using bromocriptine as a more specific therapy option showed promising early clinical results and is currently being tested in larger randomized trials. Frequent symptoms such as general fatigue, dyspnea and dry cough are often misinterpreted as pregnancy-related disturbances, and diagnosis is therefore often late. Interdisciplinary management, early diagnosis and more disease-specific therapy options appear to promote cardiac recovery in PPCM patients.     

Central and regional vascular hemodynamics following intravenous milrinone in the conscious rat: comparison with dobutamine

This study examined the hemodynamic and regional vascular profile of intravenous (i.v.) milrinone during increasing doses (3, 6, 12 micrograms/kg/min, n = 8) and by intraindividual comparison of milrinone and dobutamine (n = 10) in normal conscious rats. At 3 micrograms/kg/min, Milrinone increased coronary and cerebral blood flow (radioactive microspheres 15 +/- 5 microns) (7.7-9.8 and 1.05-1.27 ml/min/g respectively, both p less than 0.05) without significant changes in systemic hemodynamics. At 6 micrograms/kg/min milrinone increased skeletal muscle blood flow (0.19-0.24 ml/min/g, p less than 0.05) along with increases in cardiac output, stroke volume, and stroke work (all p less than 0.05), while systemic vascular resistance decreased (-51%, p less than 0.05). When compared with dobutamine, milrinone caused a greater increase in cardiac output (+26% vs. +17%) and a greater reduction in systemic vascular resistance. Milrinone and dobutamine increased renal, intestinal, cerebral, and coronary flow to a similar extent, but only milrinone enhanced hepatic arterial blood flow (+26%, p less than 0.05) and tended to increase flow to skeletal muscle (+35%, p = 0.07). We conclude that milrinone exerts significant regional vasodilating effects in a conscious rat model, being most prominent in the coronary and cerebral circulations at a dosage that does not alter central hemodynamics. At higher doses, milrinone causes a balanced increase in regional blood flow including enhanced flow to skeletal muscle. The hemodynamic (particularly as compared with dobutamine) and regional vascular profile of milrinone suggests a predominant vasodilating effect in the rat. Given a similar limited response of rat and diseased human myocardium to milrinone, these findings may have important clinical implications.     

Quantitative and qualitative enzyme studies of Hodgkin's disease-derived cell lines

Any extensive analysis of Hodgkin (H) and Reed-Sternberg (RS) cells is limited by the scarcity of available material and by the common contamination with "by-stander" cells. The establishment of Hodgkin's disease-derived cell lines provides the opportunity to undertake studies in which large numbers of cells are required, as these cell lines are by definition monoclonal populations with unlimited cell growth. In this study, we analyzed the enzyme profiles of eight Hodgkin's disease cell lines (Co, Ho, Fox, HDLM-2, KM-H2, L428, L540, and L591) whereby cellular alpha-naphthyl acetate esterases, acid phosphatases, and dipeptidylpeptidase IV were examined quantitatively or qualitatively by IEF or chromatographic techniques. The results indicate that all of the H-RS cell lines examined had enzymatic features typical for lymphoid cells and, in particular, a monocyte/histiocyte origin of the cell lines could be excluded. Extrapolation of the available immunological, molecular biological, and enzymological evidence gained in vitro on cultured representatives of H-RS cells suggests a lymphoid origin for in vivo H-RS cells.     

A case of subcortical grey matter heterotopia presenting as bipolar disorder.

A case of previously diagnosed bipolar disorder was found to be associated with unilateral subcortical grey matter heterotopia, cortical hemiatrophy, midline shift, and ventriculomegaly on magnetic resonance imaging (MRI). The patient responded to pharmacotherapy with lithium carbonate and carbamazepine. This case dramatically illustrates the need for neuroimaging in psychiatric patients with apparently "functional" affective illness.     

Diagnostic procedure in osteodystrophia deformans Paget

Simultaneous scintigraphic and X-ray examination has been evaluated in 30 patients with Paget's disease for estimation of extension and activity of the disease. Scintigraphy produces a comprehensive picture of the pathophysiologic condition. When compared with X-rays the procedure is more economic with regards to cost, time and radiation dosage. Therefore, the use of radioisotopes for evaluation of the skeleton is strongly recommended for follow-up examination in Paget's disease.     

Molecular cytogenetic characterization of the mantle cell lymphoma cell line GRANTA-519

Combining fluorescence R-banding, fluorescence in situ hybridization and spectral karyotyping allowed us to precisely define chromosomal breakpoints, gains, losses and a newly detected amplification in the human mantle cell lymphoma (MCL) cell line GRANTA-519. GRANTA-519 is characterized by the t(11;14)(q13;q32) resulting in overexpression of cyclin D1, a key player in cell cycle control. Hitherto unresolved complex rearrangements involve 1p, 1q, 3cen, 9p, 11q, 12p, 12q, 16p, 17p, and 18cen. Moreover, a 4- to 6-fold gain of sequences on 18q leads to a low-level amplification of the BCL2 gene and to an overexpression of the BCL2 protein. These results provide the basis for the identification of not only candidate oncogenes responsible for MCL in gained regions, but also for the identification of putative tumor suppressor genes in commonly deleted regions like 1p22, which would eventually enable functional studies of these genes.     

A rapid and simple method for efficient coating of microtiter plates using low amounts of antigen in the presence of detergent

Bio-Beads SM-2 have previously been used for the removal of non-ionic detergents from protein solutions. Addition of Bio-Beads SM-2 to detergent solubilized antigen significantly enhanced the immobilization of antigen to microtiter wells. Depending on the incubation time used 35-45% of the applied antigen could be immobilized to the microtiter wells. Using this method and a subsequent ELISA procedure it was possible to detect monoclonal antibodies in hybridoma supernatants after coating microtiter wells with 100 microliters of a solution containing 16 ng antigen/ml in the presence of 0.01% Triton X-100.