Preparation and characterization of a pentaammineruthenium(III) derivative of horse heart ferricytochrome c.

A stable complex between pentaammineruthenium(III) and histidine-33 in horse heart ferricytochrome c is formed in the reaction between aquopentaammineruthenium(II) and the protein at pH 7. HPLC of the tryptic hydrolysate of the modified protein was employed to identify the pentaammineruthenium binding site. Spectroscopic measurements show that the integrity of the native structure in the vicinity of the heme c group is maintained in the ruthenium-modified protein. The reduction potentials are: heme c (Fe3+/2+), 0.26 V; Ru(NH3)5(His-33)3+/2+, 0.15 V (vs. normal hydrogen electrode).     

HIF-2alpha regulates murine hematopoietic development in an erythropoietin-dependent manner

Erythropoiesis in the adult mammal depends critically on erythropoietin, an inducible cytokine with pluripotent effects. Erythropoietin gene expression increases under conditions associated with lowered oxygen content such as anemia and hypoxia. HIF-1alpha, the founding member of the hypoxia-inducible factor (HIF) alpha class, was identified by its ability to bind and activate the hypoxia-responsive enhancer in the erythropoietin regulatory region in vitro. The existence of multiple HIF alpha members raises the question of which HIF alpha member or members regulates erythropoietin expression in vivo. We previously reported that mice lacking wild-type HIF-2alpha, encoded by the EPAS1 gene, exhibit pancytopenia. In this study, we have characterized the etiology of this hematopoietic phenotype. Molecular studies of EPAS1-null kidneys reveal dramatically decreased erythropoietin gene expression. EPAS1-null as well as heterozygous mice have impaired renal erythropoietin induction in response to hypoxia. Treatment of EPAS1-null mice with exogenous erythropoietin reverses the hematopoietic and other defects. We propose that HIF-2alpha is an essential regulator of murine erythropoietin production. Impairments in HIF signaling, involving either HIF-1alpha or HIF-2alpha, may play a prominent role in conditions involving altered hematopoietic or erythropoietin homeostasis.     

Detection of anti-HTLV-I Tax antibodies in HTLV-I enzyme-linked immunosorbent assay-negative individuals

The HTLV-I tax gene protein (Tax) is not packaged within the mature viral particle from which the proteins for the commercially available enzyme-linked immunosorbent assay (ELISA) are derived. Screening of 162 individuals within a cohort of white intravenous (IV) drug abusers, previously identified as having an increased incidence of HTLV-I infection, demonstrated that seven of them had antibodies to the HTLV-I Tax protein but tested negative in HTLV-I ELISAs and Western blots prepared from purified virion proteins. Three out of 35 individuals in other behaviorally defined high-risk groups also displayed this limited pattern of reactivity to HTLV-I proteins. The presence of the anti-HTLV- I p40/Tax antibodies was determined by radioimmunoprecipitation assay (RIPA), which also revealed low levels of anti-env reactivity. The specificity of the anti-p40 reactivity was confirmed on specific Tax ELISAs and Western blots prepared from recombinantly produced Tax. In vitro gene amplification by the polymerase chain reaction (PCR) was used to establish the presence of sequences homologous to HTLV-I proviral DNA in four/four of these HTLV-I ELISA negative, Tax ELISA/Tax western blot/RIPA positive individuals. These data suggest that the true incidence of HTLV-I infection within high-risk cohorts is greater than previously reported.     

Availability of ulipristal acetate: A secret shopper survey of pharmacies in a metropolitan area on emergency contraception

ObjectivesTo assess levonorgestrel (LNG) and ulipristal acetate (UPA) availability in pharmacies in a metropolitan area.MethodsA cross-sectional survey was conducted of all identified pharmacies within 25 miles of an urban medical center in Kansas City, KS. We categorized the pharmacies as dedicated commercial (national chains), store-associated (affiliated with a general merchandise or grocery store), or independent. We assessed LNG and UPA availability or time to availability if not currently stocked.ResultsWe contacted 165 pharmacies. Of the 165 pharmacies, few stocked UPA (12/165, 7%) whereas the majority stocked oral LNG (128/165, 78%). Dedicated commercial pharmacies were more likely to carry UPA than store-associated and independent pharmacies (11/84 [13%] vs. 1/61 [1%] vs. 0/20, respectively; P = 0.016). Most pharmacies that did not stock UPA reported that they could obtain it within 24 hours (94/153, 62%). Dedicated commercial pharmacies were most likely report the ability to obtain UPA in 24 hours (P = 0.016).ConclusionFew pharmacies stock UPA, the most effective form of oral emergency contraception. Enhanced communication between medical providers and pharmacists within current laws and regulations could enhance patient access to UPA.     

Single agent lenalidomide three times a week induces hematologic responses in AL amyloidosis patients on dialysis

The combination of lenalidomide and dexamethasone can produce hematologic responses in previously treated patients with AL amyloidosis. Because lenalidomide is primarily excreted unchanged by the kidney, adjustments to the starting dose of lenalidomide are recommended to provide appropriate drug exposure in patients with moderate or severe renal impairment and in patients on dialysis. Here, we report on a study of patients with AL amyloidosis on dialysis treated with lenalidomide at a dose of 10 mg orally three times a week. Seven patients were enrolled. All patients had received prior treatment, with 57% receiving prior high‐dose melphalan and stem cell transplantation. Two patients died before evaluation of response. The most common adverse event was infection; no thromboembolic complications were seen. One patient required dose‐modification. Hematologic responses were obtained by four of the five evaluable patients. Median overall survival was 18 months. In conclusion, adjusted dose lenalidomide was reasonably tolerated and induced sustained hematologic responses in previously treated patients with AL amyloidosis on dialysis. Am. J. Hematol. 89:706–708, 2014. © 2014 Wiley Periodicals, Inc.     

Detection of Chronic Wasting Disease in the Lymph Nodes of Free-Ranging Cervids by Real-Time Quaking-Induced Conversion

Chronic wasting disease (CWD), a transmissible spongiform encephalopathy of deer, elk, and moose, is the only prion disease affecting free-ranging animals. Since the disease was first identified in northern Colorado and southern Wyoming in 1967, new epidemic foci of the disease have been identified in 20 additional states, as well as two Canadian provinces and the Republic of South Korea. Identification of CWD-affected animals currently requires postmortem analysis of brain or lymphoid tissues using immunohistochemistry (IHC) or an enzyme-linked immunosorbent assay (ELISA), with no practical way to evaluate potential strain types or to investigate the epidemiology of existing or novel foci of disease. Using a standardized real-time (RT)-quaking-induced conversion (QuIC) assay, a seeded amplification assay employing recombinant prion protein as a conversion substrate and thioflavin T (ThT) as an amyloid-binding fluorophore, we analyzed, in a blinded manner, 1,243 retropharyngeal lymph node samples from white-tailed deer, mule deer, and moose, collected in the field from areas with current or historic CWD endemicity. RT-QuIC results were then compared with those obtained by conventional IHC and ELISA, and amplification metrics using ThT and thioflavin S were examined in relation to the clinical history of the sampled deer. The results indicate that RT-QuIC is useful for both identifying CWD-infected animals and facilitating epidemiological studies in areas in which CWD is endemic or not endemic.