Sensitizing acute myeloid leukemia cells to induced differentiation by inhibiting the RIP1/RIP3 pathway.

Tumor necrosis factor-α (TNF-α)-induced RIP1/RIP3 (receptor-interacting protein kinase 1/receptor-interacting protein kinase 3)-mediated necroptosis has been proposed as an alternative strategy for treating apoptosis-resistant leukemia. However, we found that most acute myeloid leukemia (AML) cells, especially M4 and M5 subtypes, produce TNF and show basal level activation of RIP1/RIP3/MLKL signaling, yet do not undergo necroptosis. TNF, through RIP1/RIP3 signaling, prevents degradation of SOCS1, a key negative regulator of interferon-γ (IFN-γ) signaling. Using both pharmacologic and genetic assays, we show here that inactivation of RIP1/RIP3 resulted in reduction of SOCS1 protein levels and partial differentiation of AML cells. AML cells with inactivated RIP1/RIP3 signaling show increased sensitivity to IFN-γ-induced differentiation. RIP1/RIP3 inactivation combined with IFN-γ treatment significantly attenuated the clonogenic capacity of both primary AML cells and AML cell lines. This combination treatment also compromised the leukemogenic ability of murine AML cells in vivo. Our studies suggest that inhibition of RIP1/RIP3-mediated necroptotic signaling might be a novel strategy for the treatment of AML when combined with other differentiation inducers.

Exome sequencing of extreme clopidogrel response phenotypes identifies B4GALT2 as a determinant of on-treatment platelet reactivity.

Interindividual variability in platelet aggregation is common among patients treated with clopidogrel and both high on-treatment platelet reactivity (HTPR) and low on-treatment platelet reactivity (LTPR) increase risks for adverse clinical outcomes. CYP2C19 influences clopidogrel response but only accounts for ∼12% of the variability in platelet reactivity. To identify novel variants implicated in on-treatment platelet reactivity, patients with coronary artery disease (CAD) with extreme pharmacodynamic responses to clopidogrel and wild-type CYP2C19 were subjected to exome sequencing. Candidate variants that clustered in the LTPR subgroup subsequently were genotyped across the discovery cohort (n = 636). Importantly, carriers of B4GALT2 c.909C>T had lower on-treatment P2Y12 reaction units (PRUs; P = 0.0077) and residual platelet aggregation (P = 0.0008) compared with noncarriers, which remained significant after adjusting for CYP2C19 and other clinical variables in both the discovery (P = 0.0298) and replication (n = 160; PRU: P = 0.0001) cohorts. B4GALT2 is a platelet-expressed galactosyltransferase, indicating that B4GALT2 c.909C>T may influence clopidogrel sensitivity through atypical cell-surface glycoprotein processing and platelet adhesion.

Peripheral blood mononuclear cell-derived erythroid progenitors and erythroblasts are decreased in burn patients.

Patients with large burns suffer from anemia of critical illness. Administration of exogenous erythropoietin is ineffective, and transfusion remains the only effective treatment. We have previously shown that erythroid precursors are decreased 1 week after burn in an animal model. Therefore, we have used a two-phase liquid culture system to quantify peripheral blood mononuclear cell (PBMC) compartment-derived erythroid progenitors (EPs) in burn patients. Institutional review board approval and informed consent were obtained. Blood samples were collected at 1 to 30 days after burn, with a mean TBSA of 37.7 ± 15.8% (n = 10; 90% men; age, 46.0 ± 18 years). Four healthy volunteers served as controls. PBMCs were isolated by Ficoll-Hypaque density-gradient centrifugation and were placed in serum-free expansion medium containing cyclosporine A (1 ng/ml), granulocyte macrophage colony-stimulating factor (20 ng/ml), stem cell factor (30 ng/ml), and interleukin-3 (5 ng/ml; phase I). On day 7, cells were reseeded in serum-free expansion medium containing erythropoietin (1 U/ml), holotransferrin (0.3 mg/ml), and stem cell factor (10 ng/ml; phase II). Aliquots from the phase II culture system on day 6 were incubated with anti-CD71, CD235a, and CD36. EPs (CD71 CD36) and erythroblast subpopulations (colony-forming unit erythroids, Proerythroblasts, and intermediate erythroblasts) were identified based on the expressions of CD71 and CD235a by flow cytometry, calculated per million expanded cells, and expressed as a percentage of controls. Total EPs were significantly decreased by days 28 to 31 after the burn (19%; P < .05). Among the erythroblast subpopulations, colony-forming unit erythroids (11%; P < .004) and proerythroblasts (24%; P < .05), were decreased significantly by days 28 to 31 after the burn. PBMCs of burn patients can be used to study impaired erythropoiesis and anemia of critical illness.

Preparation and characterization of microparticles of piroxicam by spray drying and spray chilling methods.

Piroxicam, an anti-inflammatory drug, exhibits poor water solubility and flow properties, poor dissolution and poor wetting. Consequently, the aim of this study was to improve the dissolution of piroxicam. Microparticles containing piroxicam were produced by spray drying, using isopropyl alcohol and water in the ratio of 40:60 v/v as solvent system, and spray chilling technology by melting the drug and chilling it with a pneumatic nozzle to enhance dissolution rate. The prepared formulations were evaluated for in vitro dissolution and solubility. The prepared drug particles were characterized by scanning electron microscopy (SEM), differential scanning calorimeter, X-ray diffraction and Fourier transform infrared spectroscopy. Dissolution profile of the spray dried microparticles was compared with spray-chilled microparticles, pure and recrystallized samples. Spray dried microparticles and spray chilled microparticles exhibited decreased crystallinity and improved micromeritic properties. The dissolution of the spray dried microparticle and spray chilled particles were improved compared with recrystallized and pure sample of piroxicam. Consequently, it was believed that spray drying of piroxicam is a useful tool to improve dissolution but not in case of spray chilling. This may be due to the degradation of drug or variations in the resonance structure or could be due to minor distortion of bond angles. Hence, this spray drying technique can be used for formulation of tablets of piroxicam by direct compression with directly compressible tablet excipients.

Coronary bifurcation lesions.

Coronary bifurcations are prone to develop atherosclerotic plaque due to turbulent blood flow and high shear stress. These lesions amount to 15-20% of the total number of interventions. The true bifurcation lesion consist of >50% diameter obstruction of the main vessel (MV) and of the side branch (SB) in an inverted "Y" fashion. Treatment of coronary bifurcation lesions represents a challenging area in interventional cardiology but recent advances in percutaneous coronary interventions (PCI) have led to the dramatic increase in the number of patients successfully treated percutaneously. When compared with non-bifurcation interventions, bifurcation interventions have a lower rate of procedural success, higher procedural costs, longer hospitalization and a higher clinical and angiographic restenosis. Introduction of drug-eluting stents (DES) has resulted in a lower event rate and reduction of main vessel (MV) restenosis in comparison with historical controls. However, side branch (SB) ostial residual stenosis and long-term restenosis remains a problem. Although stenting the MV with provisional SB stenting seems to be the prevailing approach, in the era of DES various two-stent techniques have emerged to allow stenting of the large side branch.

Feasibility of conditioning with thymoglobulin and reduced intensity TBI to reduce acute GVHD in recipients of allogeneic SCT.

Murine studies using anti-T-cell antibodies for conditioning in allogeneic SCT demonstrate engraftment with low rates of GVHD. On the basis of this preclinical model, we conditioned 30 patients with advanced hematologic malignancies with rabbit antithymocyte globulin (ATG) and TBI, to reduce rates of fatal acute GVHD. Patients were enrolled in two sequential groups: cohort 1 received ATG 10 mg/kg in divided doses (days -4 to -1)+200 cGy TBI (n=16), and cohort 2 received ATG (days -10 to -7)+450 cGy TBI (n=14). Median donor blood chimerism for the combined group was 94, 93 and 93% in the first, second and third months after transplant. Only three developed grade II acute GVHD despite 43% of patients receiving unrelated donor transplants. One-year survival was 71+/-11 and 54+/-14%, respectively, in recipients of related and unrelated donor SCT. Donor lymphocyte infusions were needed in 12 patients for the management of relapse and for mixed donor-recipient chimerism in 4 patients. We conclude that 10 mg/kg ATG and TBI allows engraftment with a low risk of acute GVHD; however, further dose optimization of ATG is required to achieve a balance between GVHD and disease relapse.

The prevention of contrast-induced nephropathy in patients undergoing percutaneous coronary intervention.

Contrast-induced nephropathy (CIN) is a leading cause of morbidity and mortality in high-risk patients undergoing percutaneous coronary intervention (PCI) or other radiocontrast procedures. Approximately 25% of all patients selected for these procedures are at risk for its development. Patients who experience this complication have higher rates of mortality, longer hospital stays and poorer long-term outcomes. The occurrence of CIN is directly related to the number of co-existing clinical risk factors. Among the many risk factors, preexisting renal impairment, advanced age, the presence of diabetes mellitus and both the volume and type of the contrast agent administered are among the most important. While the precise pathophysiological mechanisms responsible for this condition are complex and incompletely understood, experimental studies suggest that the pathogenesis involves a combination of renal ischemia and direct tubular epithelial cell toxicity. At the present time, adequate periprocedural hydration and the selection of low-osmolar and, more recently, iso-osmolar contrasts agents are the only available tools to the operator for reducing the risk of this complication. Several other modalities, such as the use of NaHCO3 and hemofiltration, also appear promising in preventing the development of this complication. This article reviews the epidemiology, pathophysiology, and consequences of CIN. It also reviews the risk factors for the development of CIN, as well as the history of the various modalities studied in its prevention.

Assignment of the Raman spectra of some deuterated-BEDT-TTF superconductors.

We present the room-temperature Raman spectra of both the protonated and deuterated forms of kappa-(BEDT-TTF)2Cu[N(CN)2]Br, kappa-(BEDT-TTF)2Cu(NCS)2 and beta-(BEDT-TTF)2I3. Along with data for the neutral BEDT-TTF molecule these spectra are used to assign the many features in the spectra of the deuterated compounds.

Test for interlayer coherence in a quasi-two-dimensional superconductor.

Peaks in the magnetoresistivity of the layered superconductor kappa - (BEDT-TTF)2Cu(NCS)(2), measured in fields < or =45 T applied within the layers, show that the Fermi surface is extended in the interlayer direction and enable the interlayer transfer integral (t( perpendicular) approximately 0.04 meV) to be deduced. However, the quasiparticle scattering rate tau(-1) is such that Planck's over 2pi/tau approximately 6t( perpendicular), implying that kappa - (BEDT-TTF)2Cu(NCS)(2) meets the criterion used to identify interlayer incoherence. The applicability of this criterion to anisotropic materials is thus shown to be questionable.

Optimal treatment of nonaorto ostial coronary lesions in large vessels: acute and long-term results.

Percutaneous interventions of nonaorto ostial coronary lesions are usually complex, often requiring a combined approach of debulking and stenting, insertion of multiple guidewires and long procedure duration. Debulking with atherectomy device preserves side-branch patency by reducing plaque shift while coronary stenting minimizes vessel recoil and restenosis. We retrospectively evaluated the acute and long-term results of rotational atherectomy (group R, n = 94), coronary stenting (group S, n = 39), and combination of rotational atherectomy and stenting (group R-S, n = 59) in a total of 192 patients with nonaorto ostial lesions. The number of patients with diabetes mellitus and rest angina was significantly higher in groups S and R-S. Clinical success rates were high and procedural complication rates were low and comparable in all three groups. Despite the similar reference vessel size and preprocedure minimal lumen diameter (MLD), postprocedure MLD showed a trend toward larger lumen in groups S (3.15 +/- 0.18 mm) and R-S (3.21 +/- 0.16 mm). Group S had significantly higher incidence of side-branch narrowing (30.7%), requiring intervention (15.4%). At long-term follow-up (mean of 9 +/- 4 months), target lesion revascularization rate was significantly lower in groups R-S (11.9%) and S (15.4%) compared to group R (28.9%; P = 0.02). Our nonrandomized data suggest that stenting with or without rotational atherectomy provides the best long-term approach for the interventional treatment of nonaorto ostial coronary lesions. The clinical benefit and cost effectiveness of performing rotational atherectomy before stent implantation to reduce the incidence of side-branch closure requires further study.

Reduction in periprocedural enzyme elevation by abciximab after rotational atherectomy of type B2 lesions: Results of the Rota ReoPro randomized trial.

BACKGROUND: Abciximab has been shown to reduce ischemic complications and creatine kinase-myocardial band (CK-MB) elevation of both simple and complex coronary interventions. In addition to the procedural complications, one of the important mechanisms for CK-MB elevation after rotational atherectomy is an interaction between platelets and the atheromatous debris. METHODS: This study was conducted to determine whether abciximab would limit the extent of periprocedural CK-MB release after rotational atherectomy of American Heart Association/American College of Cardiology type B(2) lesions in a double-blind, randomized, placebo-controlled manner. A total of 100 lesions in 100 patients were randomized with the primary end point being a CK-MB elevation of >16 U/L. RESULTS: Procedural success was achieved in 100% in the abciximab arm compared with 98% in the placebo group with any CK-MB elevation >16 U/L of 8% in the abciximab versus 22% in the placebo group (P =.04). The peak creatine phosphokinase level (units per liter) was 102 +/- 14 versus 153 +/- 22 (P =.05) and the peak CK-MB level was 12.8 +/- 1.8 versus 24.6 +/- 3.5 (P =.06) between the abciximab and placebo groups, respectively. Slow-flow or postprocedure chest pain occurred in 14% in the abciximab group versus 30% in the placebo group (P =.04). There was 1 Q-wave myocardial infarction in the placebo arm and 1 nonhemorrhagic stroke in the abciximab group. CONCLUSIONS: Therefore the Rota ReoPro randomized trial revealed the benefit of abciximab during rotational atherectomy in reducing procedural morbidity and CK-MB elevation, and its routine use can be justified even in moderately complex lesions undergoing rotational atherectomy.

Angiogenesis in acute promyelocytic leukemia: induction by vascular endothelial growth factor and inhibition by all-trans retinoic acid.

Recent studies indicate that angiogenesis is important in the pathogenesis of leukemias, apart from its well-established role in solid tumors. In this study, the possible role of angiogenesis in acute promyelocytic leukemia (APL) was explored. Bone marrow trephine biopsies from patients with APL showed significantly increased microvessel density and hot spot density compared with normal control bone marrow biopsies. To identify the mediators of angiogenesis in APL, quantitative and functional assays were performed using the NB4 APL cell line as a model system. Conditioned media (CM) from the NB4 cells strongly stimulated endothelial cell migration. CM from the NB4 cells contained high levels of vascular endothelial growth factor (VEGF) but not basic fibroblast growth factor (bFGF). Most important, the addition of neutralizing VEGF antibodies completely inhibited the ability of NB4 CM to stimulate endothelial cell migration, suggesting that APL angiogenesis is mediated by VEGF. The effect of all-trans retinoic acid (ATRA) on APL angiogenesis was then studied. ATRA therapy resulted in a decrease in bone marrow microvessel density and hot spot density. CM from ATRA-treated APL cells did not stimulate endothelial cell migration. Finally, quantitative assays showed that ATRA treatment resulted in the abrogation of VEGF production by the NB4 cells. These results show that there is increased angiogenesis and VEGF production in APL and that ATRA therapy inhibits VEGF production and suppresses angiogenesis. The addition of specific antiangiogenic agents to differentiation therapy or chemotherapy should be explored. (Blood. 2001;97:3919-3924)

Infantile cartilaginous hamartoma of the rib. A case report.

BACKGROUND: Infantile cartilaginous hamartoma of the rib is a rare condition occurring in newborn infants, with an incidence of 1 in 3,000 (0.03%) among primary bone tumor cases. Reports of this condition so far have presented the clinical, radiologic and histopathologic features. To the best of our knowledge, reports of the cytopathologic features have not been documented. In the present case report, clinical, radiologic and cytopathologic features and differential diagnosis are enumerated. CASE: A 1-month-old, male infant presented with a chest wall mass with a clinical diagnosis of osteochondroma. On fine needle aspiration cytology, a diagnosis of infantile cartilaginous hamartoma of the rib was suggested; it was supplemented by the clinical history and radiologic findings. CONCLUSION: Although rare, this condition ought to be kept in mind while dealing with infantile chest wall masses to avoid an erroneous diagnosis of malignancy, owing to its ominous cytopathologic features.

Modulation of cytoplasmic dynein ATPase activity by the accessory subunits.

The microtubule-based motor molecule cytoplasmic dynein has been proposed to be regulated by a variety of mechanisms, including phosphorylation and specific interaction with the organelle-associated complex, dynactin. In this study, we examined whether the intermediate chain subunits of cytoplasmic dynein are involved in modulation of ATP hydrolysis, and thereby affect motility. Treatment of testis cytoplasmic dynein under hypertonic salt conditions resulted in separation of the intermediate chains from the remainder of the dynein molecule, and led to a 4-fold enhancement of ATP hydrolysis. This result suggests that the accessory subunits act as negative regulators of dynein heavy chain activity. Comparison of ATPase activities of dyneins with differing intermediate chain isoforms showed significant differences in basal ATP hydrolysis rates, with testis dynein 7-fold more active than dynein from brain. Removal of the intermediate chain subunits led to an equalization of ATPase activity between brain and testis dyneins, suggesting that the accessory subunits are responsible for the observed differences in tissue activity. Finally, our preparative procedures have allowed for the identification and purification of a 1:1 complex of dynein with dynactin. As this interaction is presumed to be mediated by the dynein intermediate chain subunits, we now have defined experimental conditions for further exploration of dynein enzymatic and motility regulation.

Managing the high-risk patient: experience with fenoldopam, a selective dopamine receptor agonist, in prevention of radiocontrast nephropathy during percutaneous coronary intervention.

Acute worsening of renal function due to contrast agents occurs in 15% to 40% of patients with baseline renal insufficiency undergoing percutaneous coronary intervention. Radiocontrast nephropathy is associated with increased morbidity, prolonged hospitalization, and higher in-hospital mortality. Our nonrandomized data suggest that in adequately hydrated patients, the dopamine-1 receptor agonist fenoldopam is a useful adjunct during PCI for prevention of RCN, reducing its incidence to less than 5%. This renoprotective effect of fenoldopam was more pronounced in diabetics, with moderate renal failure, in whom no agent has been shown so far to be beneficial.

The molecular receptive range of an odorant receptor.

An odor perception is the brain's interpretation of the activation pattern of many peripheral sensory neurons that are differentially sensitive to a wide variety of odors. The sensitivity of these neurons is determined by which of the thousand or so odor receptor proteins they express on their surface. Understanding the odor code thus requires mapping the receptive range of odorant receptors. We have adopted a pharmacological approach that uses a large and diverse pool of odorous compounds to characterize the molecular receptive field of an odor receptor. We found a high specificity for certain molecular features, but high tolerance for others-a strategy that enables the olfactory apparatus to be both highly discriminating, and able to recognize several thousand odorous compounds.

Increased bone marrow angiogenesis in B cell chronic lymphocytic leukemia.

Recent studies have shown that angiogenesis may be involved in the pathogenesis of hematopoietic malignancies, apart from its well-characterized role in the growth and metastasis of solid tumors. In this study, we quantified the degree of angiogenesis in B cell chronic lymphocytic leukemia (B-CLL) by measuring the microvessel density and hotspot density in bone marrow trephine biopsy sections with B-CLL involvement (n = 12) and compared it to normal bone marrow sections (n = 11). The B-CLL samples had a mean microvessel count/high power field (hpf) of 7.64 while the control samples had a mean microvessel count/hpf of 2.11 (P = 0.0001). The mean hotspot density in the B-CLL sections (14.83/hotspot) was also significantly higher (P = 0.0008) than the mean hotspot density in control bone marrow sections (7.09/hotspot). Both the microvessel density and hotspot density correlated positively with the clinical stage of the B-CLL patients. In a separate cohort of B-CLL patients, the median urine level of the angiogenic peptide, basic fibroblast growth factor (2216.5 pg/g, n = 14), was significantly higher (P = 0.0001) than the bFGF level in normal controls (1,084 pg/g, n = 58). These results indicate that angiogenesis may be involved in the pathogenesis of B-CLL.