"Home-brew" FISH assay shows higher efficiency than BCR-ABL dual color, dual fusion probe in detecting microdeletions and complex rearrangements associated with t(9;22) in chronic myeloid leukemia

We carried out fluorescence in situ hybridization (FISH) studies on 18 Ph+ chronic myeloid leukemia (CML) cases with chromosome 22 genomic deletions with the Vysis BCR-ABL dual-color/dual-fusion probe (BCR-ABL DC/DF) to compare the hybridization patterns obtained with this approach to those obtained with the "home brew" BAC/PAC system. Our results are the following: chromosome 22 microdeletions less than 400 kilobases (Kb) were not detected by the BCR DC/DF probe; FISH analysis with the BCR DC/DF probe in cases bearing chromosome 22 microdeletions ranging from 400 to 700 Kb produced a faint signal on the der(9); and the BCR-ABL DC/DF FISH pattern was comparable to the one obtained by the home brew probe in the presence of a 900-Kb chromosome 22 microdeletion. Our home-brew FISH system represents an accurate method for revealing a subset of CML patients with der(9) microdeletions.     

Endometrioid-like yolk sac and Sertoli-Leydig cell tumors in a carrier of a Y heterochromatin insertion into 1qh region: a causal association?

We describe the case of a young woman showing yolk sac tumors (YST) and a Sertoli-Leydig cell tumor (SLCT) in the right ovary, with recurrences in the right adnexum and with hepatic metastasis. To our knowledge, YST and SLCT have never been described as components of the same tumor or reported as associated in the same patient. The patient's karyotype showed the presence of Y chromosome inserted into the 1qh region; the inserted region corresponded to Yq12 heterochromatin. LOH analysis revealed 1p36 paternal allele loss in the proband tumor, thus supporting a germ cell origin for the tumor. The presence of Y heterochromatin in 1qh DNA might induce disturbances in the normal regulation of oncogenes located in 1q.     

Influence of estrogens and antiestrogens on the expression of selected hormone-responsive genes

Estrogen exerts a primary regulatory role on a wide variety of physiological processes in different tissues and organs. Agonistic ad antagonistic compounds are widely used in human health and, therefore, a deep understanding of their mechanisms of action at the molecular level is mandatory. The effect of 17beta-estradiol and three antiestrogenic drugs, comprising two selective estrogen receptor modulator (SERM, 4-OH-tamoxifen, Raloxifene) and the pure antiestrogen ICI 182,780, on genome-wide gene expression levels was evaluated in breast carcinoma cell lines by DNA microarray analysis. Different clusters of genes, showing specific coregulation patterns, were found. First, several groups of genes displaying temporal-specific up- or down-regulation were characterized. Second, clusters of genes responding to different antiestrogenic drugs in either antagonstic or agonistic fashion, were found. Genes responding specifically to antiestrogens, but not to estrogen, were also identified. In addition, each individual compound exhibited a very specific gene regulation. Bioinformatic analysis was applied to the regulatory sequences of different groups of genes and confirmed that specific pathways and secondary responses are activated at each temporal point and in response to different compounds. Our results underline the complexity of genomic responses to estrogen in breast cancer cells and strongly suggest that the molecular characterization of estrogen agonists and antagonists used in human therapy should be carefully studied.     

Composite-to-composite microtensile bond strength in the repair of a microfilled hybrid resin: effect of surface treatment and oxygen inhibition

PURPOSE: To compare the 24-h microtensile bond strength of a microfilled hybrid composite to the same material after mechanical and/or chemical treatment and assess the effect of oxygen inhibition on the composite-composite bond. MATERIALS AND METHODS: Forty composite cylinders of Gradia Direct Anterior (GC) were prepared and stored 24 h prior to the following surface treatments: 50-microm aluminum oxide air abrasion and 37% phosphoric acid etching (group 1); hydrochloric acid and 6.9% hydrofluoric acid etching (group 2); diamond bur roughening and 37% phosphoric acid etching (group 3); diamond bur roughening (group 4). In all groups, Prime & Bond NT (Dentsply De Trey) was applied and light cured in air or under a nitrogen atmosphere, prior to layering a buildup of the repairing resin composite. Microtensile bond strength measurements were performed. Data were statistically analyzed with two-way ANOVA and Tukey's test (alpha = 0.05). RESULTS: The curing atmosphere did not significantly influence the interfacial strength (p < 0.05). Surface treatment significantly affected the composite-composite bond (p > 0.05). Air abrasion, regardless of curing atmosphere, resulted in the strongest bond (p < 0.05). The other treatments were comparable. CONCLUSION: Air abrasion and the application of a bonding agent offer satisfactory bond strengths for composite repair. The oxygen inhibition layer on a light-cured adhesive is not crucial to the success of the 24-h composite-composite bond.     

Lymph node blast crisis in chronic myeloid leukemia mimicking T-immunoblastic lymphoma.

BACKGROUND: Chronic myeloid leukemia arises from a somatic mutation in a pluripotent stem cell. It generally terminates with a blastic crisis (BC). One third of BC are lymphoid, and most have a pre-B phenotype. Few cases of T-lymphoid BC have been reported. Here we describe a lymph node blast crisis mimicking T-immunoblastic lymphoma. METHODS: Bone marrow and lymph nodes were histologically examined by standard methods and by an immunoperoxidase technique. Cytogenetic studies were also performed on lymph node and blood cells. Analysis of T-cell receptor genes and BCR rearrangements were performed on DNA extracted from both frozen bone marrow and lymph-node cells. RESULTS: Lymph-node histology showed an infiltration by large lymphoid blasts, consistent with a diagnosis of immunoblastic lymphoma. Blast cells were CD2, CD7, TDT positive, and negative for myeloid and mature lymphoid antigens. The Ph1 chromosome was found in both bone marrow and lymph-node cells. BCR rearrangement was found in the DNA from both bone marrow and lymph-node cells. TCR genes were not rearranged. DISCUSSION: The present study provides strong evidence that the lymph-node blast crisis of CML can assume the morphological appearance of immunoblastic lymphoma and may retain the immunological phenotype and genetic features of early T cells with BCR rearrangements.     

p11 modulates L-DOPA therapeutic effects and dyskinesia via distinct cell types in experimental Parkinsonism

The reduced movement repertoire of Parkinson’s disease (PD) is mainly due to degeneration of nigrostriatal dopamine neurons. Restoration of dopamine transmission by levodopa (L-DOPA) relieves motor symptoms of PD but often causes disabling dyskinesias. Subchronic L-DOPA increases levels of adaptor protein p11 (S100A10) in dopaminoceptive neurons of the striatum. Using experimental mouse models of Parkinsonism, we report here that global p11 knockout (KO) mice develop fewer jaw tremors in response to tacrine. Following L-DOPA, global p11KO mice show reduced therapeutic responses on rotational motor sensitization, but also develop less dyskinetic side effects. Studies using conditional p11KO mice reveal that distinct cell populations mediate these therapeutic and side effects. Selective deletion of p11 in cholinergic acetyltransferase (ChAT) neurons reduces tacrine-induced tremor. Mice lacking p11 in dopamine D2R-containing neurons have a reduced response to L-DOPA on the therapeutic parameters, but develop dyskinetic side effects. In contrast, mice lacking p11 in dopamine D1R-containing neurons exhibit tremor and rotational responses toward L-DOPA, but develop less dyskinesia. Moreover, coadministration of rapamycin with L-DOPA counteracts L-DOPA–induced dyskinesias in wild-type mice, but not in mice lacking p11 in D1R-containing neurons. 6-OHDA lesioning causes an increase of evoked striatal glutamate release in wild type, but not in global p11KO mice, indicating that altered glutamate neurotransmission could contribute to the reduced L-DOPA responsivity. These data demonstrate that p11 located in ChAT or D2R-containing neurons is involved in regulating therapeutic actions in experimental PD, whereas p11 in D1R-containing neurons underlies the development of L-DOPA–induced dyskinesias.Parkinson’s disease (PD) is characterized by a progressive degeneration of dopaminergic neurons projecting from substantia nigra pars compacta (SNc) to striatum, eventually resulting in bradykinesia, rigidity, resting tremor, and postural imbalance (1). Dopamine replacement strategies are effective for many motor symptoms. At early stages, MAO-B inhibitors or dopamine D2 receptor agonists may provide sufficient symptomatic relief, but as the disease progresses essentially all patients will require treatment with levodopa (L-DOPA) (1). Following decarboxylation, L-DOPA is converted to dopamine acting on both dopamine D1 and D2 receptors. D1 and D2 receptors are segregated in striatonigral and striatopallidal pathway neurons, respectively (2). Activation of D1 and D2 receptors causes synergistic stimulation of locomotion, explaining the stronger anti-Parkinsonian action of L-DOPA compared with selective D2 receptor agonists (1, 2). Whereas dopaminergic agents often successfully treat bradykinesia and rigidity in PD, additional therapy with anticholinergic agents is sometimes required for optimal treatment of resting tremor (1). Moreover, the therapeutic effect of L-DOPA is gradually shortened as disabling dyskinetic side effects emerge (3). There is no licensed treatment against L-DOPA–induced dyskinesias (LIDs).Targeting the serotonergic, glutamatergic, and/or cholinergic systems have been reported to counteract LIDs (3). p11 (i.e., S100A10) is a member of the S100 EF-hand protein family, which increase the levels of distinct serotonin (5-HT_1BR and 5-HT₄R) and glutamate (mGluR5) receptors at the cell surface, resulting in enhanced effects on cell signaling via these receptors (4, 5). p11 is widely expressed in the brain with particularly high levels in cholinergic neurons (6–9). p11 is regulated by a variety of stimuli and therapies, most notably antidepressants (4). p11 is strongly up-regulated in the striatum following repeated treatment with L-DOPA in the 6-OHDA–lesioning model of experimental parkinsonism (10). p11 can therefore influence several pathways implicated in LIDs.In the present study, we examined global and cell-specific conditional p11 knockout (KO) mice in experimental Parkinsonism models to identify the brain circuitries that mediate p11-dependent therapeutic effects of L-DOPA as well as LIDs.     

Aberrant methylation of DAP-kinase in therapy-related acute myeloid leukemia and myelodysplastic syndromes

Death-associated protein kinase (DAP-kinase), a proapoptotic serine/threonine kinase, is a candidate tumor suppressor gene. We studied the methylation status of DAP-kinase of 194 bone marrow samples from 160 patients with acute myeloid leukemia (AML) and 34 with a myelodysplastic syndrome (MDS) at the time of initial diagnosis by polymerase chain reaction (PCR). Hypermethylation of DAP-kinase was present in 27.5% (44 of 160) of AML and in 47% (16 of 34) of MDS specimens and significantly correlated to loss of DAP-kinase expression (P =.008). It was significantly more frequent in AML secondary to therapy for other malignancies (s-AML; 14 of 29, 48.3%), as compared to de novo AML (30 of 131, 22.9%, P =.01). DAP-kinase hypermethylation in AML was associated with myelodysplastic changes in the bone marrow at the time of the initial diagnosis (P =.002) and with the presence of cytogenetic abnormalities (P =.02). Alteration in the apoptotic response due to the loss of DAP-kinase function may be an early event in the transformation pathway to secondary leukemia via myelodysplasia.     

Low-Dose Acetazolamide in the Treatment of Premenstrual Dysphoric Disorder: A Case Series

The treatment of premenstrual dysphoric disorder (PMDD) is far from satisfactory, as there is a high proportion of patients who do not respond to conventional treatment. The antidiuretic sulfonamide, acetazolamide, inhibits carbonic anhydrase and potentiates GABAergic transmission; the latter is putatively involved in PMDD. We therefore tried acetazolamide in a series of women with intractable PMDD. Here, we describe a series of eight women diagnosed with DSM-IV-TR PMDD, five of whom had comorbidity with a mood disorder and one with an anxiety disorder, who were resistant to treatment and responded with symptom disappearance after being added-on 125 mg/day acetazolamide for 7-10 days prior to menses each month. Patients were free from premenstrual symptoms at the 12-month follow-up. We suggest that acetazolamide may be used to improve symptoms of PMDD in cases not responding to other treatments. GABAergic mechanisms may be involved in counteracting PMDD symptoms.     

Vitamin D in “early” primary Sjögren’s syndrome: does it play a role in influencing disease phenotypes?

Beyond its well-established role in the maintenance of mineral homeostasis, 25-OH-vitamin D deficiency seems to be involved in the development and severity of several autoimmune diseases. To date, contrasting data have been reported regarding the presence of hypovitaminosis D in primary Sjögren’s syndrome (pSS). To assess the prevalence of hypovitaminosis D in pSS at an early stage of the disease and to evaluate its impact on pSS clinical manifestations and disease activity, unselected consecutive subjects with recent onset dry mouth and/or dry eyes who underwent a comprehensive diagnostic algorithm for pSS (AECG criteria) were prospectively included in the study. The levels of 25[OH]-D3 were measured by monoclonal antibody immunoradiometric assay. Conditions of 25[OH]-D3 severe deficiency, deficiency, and insufficiency were defined as levels <10, <20, and 20–30 ng/ml, respectively, and their frequencies were investigated in pSS patients and controls. The levels of 25[OH]-D3 were also correlated with patients’ demographic, clinical, and serologic features. Seventy-six consecutive females were included: 30/76 patients fulfilled the AECG criteria for pSS. The remaining 46/76 patients represented the control group. No statistical differences were found in the serum levels of 25[OH]-D3 between pSS patients [median levels = 20 ng/ml (IQR 9.3–26)] and controls [median levels = 22.5 ng/ml (IQR 15.6–33)]. In particular, the frequency of 25[OH]-D3 severe deficiency was not significantly different in patients with pSS when compared to controls (23 vs. 17.4 %, p value = 0.24). We found a significant correlation between serum 25[OH]-D3 levels and white blood cells count (r = 0.29, p = 0.01). More specifically, leukocytopenia was significantly associated with 25[OH]-D3 severe deficiency, being documented in 40 % of the subjects with a 25[OH]-D3 severe deficiency and in 11 % of the subjects without a severe vitamin D deficiency (p = 0.02). We did not observe any further association or correlation between hypovitaminosis D and pSS glandular and extra-glandular features. Although the role of hypovitaminosis D in pSS pathogenesis remains controversial, the results of this study encourage the assessment of vitamin D in specific pSS subsets that could mostly benefit from a supplementation.