A 52-Year-Old Woman With an Abdominal Mass, Bilateral Pulmonary Nodules, and Mediastinal and Hilar Lymphadenopathy.

CASE PRESENTATION: A 52-year-old, nonsmoking, African-American woman with a history of obesity, hypertension, and rheumatoid arthritis was referred for workup of multiple bilateral pulmonary nodules. The pulmonary nodules were discovered incidentally while undergoing a CT scan for an abdominal mass that was radiographically diagnosed as a uterine leiomyoma. She was asymptomatic from a pulmonary standpoint without unintentional weight loss, fevers, or night sweats. Her mother and sister had a history of lung cancer. She was diagnosed with rheumatoid arthritis 5 years earlier that was controlled with adalimumab for approximately 3 years when she stopped being seen by her rheumatologist and discontinued adalimumab. During evaluation for the abdominal mass, she re-established care with a rheumatologist and was started on 40 mg prednisone daily with plans to restart adalimumab once the workup for the abdominal mass and pulmonary nodules was completed. She had undergone bariatric surgery with cholecystectomy approximately 5 years earlier, after which she experienced intentional postsurgical weight loss.

Effects of a naturally occurring amino acid substitution in bovine PrP: a model for inherited prion disease in a natural host species.

OBJECTIVE: The most common hereditary prion disease is human Creutzfeldt-Jakob disease (CJD), associated with a mutation in the prion gene resulting in a glutamic acid to lysine substitution at position 200 (E200K) in the prion protein. Models of E200K CJD in transgenic mice have proven interesting but have limitations including inconsistencies in disease presentation, requirement for mixed species chimeric protein constructs, and the relatively short life span and time to disease onset in rodents. These factors limit research on the mechanism by which the mutation drives disease development. Therefore, our objective was to provide the first assessment of cattle carrying the homologous mutation, E211K, as a system for investigating longer-term disease mechanisms. The E211K substitution was associated with a case of bovine spongiform encephalopathy from 2006. RESULTS: We assessed the molecular properties of bovine E211K prion protein, characterized the molecular genetics of a population of cattle E211K carriers (offspring of the original EK211 cow) in relation to findings in humans, and generated preliminary evidence that the impacts of copper-induced oxidative stress may be different in cattle as compared to observations in transgenic mouse models. The cattle E211K system provides the opportunity for future analysis of physiological changes over time.

Exposure of RML scrapie agent to a sodium percarbonate-based product and sodium dodecyl sulfate renders PrPSc protease sensitive but does not eliminate infectivity.

BACKGROUND: Prions, the causative agents of the transmissible spongiform encephalopathies, are notoriously difficult to inactivate. Current decontamination recommendations by the World Health Organization include prolonged exposure to 1 N sodium hydroxide or > 20,000 ppm sodium hypochlorite, or autoclaving. For decontamination of large stainless steel surfaces and equipment as in abattoirs, for example, these methods are harsh or unsuitable. The current study was designed to evaluate the effectiveness of a commercial product containing sodium percarbonate to inactivate prions. Samples of mouse brain infected with a mouse-adapted strain of the scrapie agent (RML) were exposed to a sodium percarbonate-based product (SPC-P). Treated samples were evaluated for abnormal prion protein (PrPSc)-immunoreactivity by western blot analysis, and residual infectivity by mouse bioassay. RESULTS: Exposure to a 21% solution of SPC-P or a solution containing either 2.1% or 21% SPC-P in combination with sodium dodecyl sulfate (SDS) resulted in increased proteinase K sensitivity of PrPSc. Limited reductions in infectivity were observed depending on treatment condition. A marginal effect on infectivity was observed with SPC-P alone, but an approximate 2-3 log10 reduction was observed with the addition of SDS, though exposure to SDS alone resulted in an approximate 2 log10 reduction. CONCLUSIONS: This study demonstrates that exposure of a mouse-adapted scrapie strain to SPC-P does not eliminate infectivity, but does render PrPSc protease sensitive.

Enteropathogenic Escherichia coli effector EspF interacts with host protein Abcf2.

Enteropathogenic Escherichia coli (EPEC) is a major causative agent of infant diarrhoea in developing countries. The EspF effector protein is injected from EPEC into host cells via a type III secretion system and is involved in the disruption of host intestinal barrier function. In addition, EspF is sorted to mitochondria and has a role in initiating the mitochondrial death pathway. To clarify the manner in which EspF affects host cells, we sought to identify eukaryotic EspF-binding proteins using affinity purification. Abcf2, a protein of unknown function and member of the ABC-transporter family, bound EspF in this assay. An interaction between EspF and Abcf2 was confirmed in a yeast two-hybrid system, by colocalization and by co-immunoprecipitation from EPEC-infected cells. Levels of Abcf2 were decreased in cells infected with EPEC in an EspF dose-dependent manner. Knock-down of Abcf2 expression by RNA interference increased EspF-induced caspase 9 and caspase 3 cleavage. In addition, Abcf2-knocked down cells showed increased caspase 3 cleavage upon treatment with the apoptosis inducing agent staurosporine. These results indicate that EspF induces or facilitates host cell death by targeting and interfering with the putative protective function of Abcf2.

Macrolide antibiotics modulate ERK phosphorylation and IL-8 and GM-CSF production by human bronchial epithelial cells.

Macrolide antibiotics decrease proinflammatory cytokine production in airway cells from subjects with chronic airway inflammation. However, in subjects with chronic obstructive pulmonary disease, short-term azithromycin (AZM) therapy causes a transient early increase in the blood neutrophil oxidative burst followed by a decrease in inflammatory markers with longer administration. We studied the effects of clarithromycin (CAM) and AZM on proinflammatory cytokine production from normal human bronchial epithelial (NHBE) cells. CAM decreased IL-8 over the first 6 h and then significantly increased interleukin (IL)-8 at 12-72 h after exposure (P < 0.0001). AZM also increased IL-8 at 24 and 48 h, and CAM increased granulocyte-macrophage colony-stimulating factor at 48 h. In the presence of LPS, both CAM and AZM dose-dependently increased IL-8 secretion over 24 h, but after 5 days of exposure to 10 microg/ml CAM there is suppression of IL-8 (P < 0.001). PD-98059, an inhibitor of MAP kinase/ERK kinase, inhibited CAM-induced IL-8 (P < 0.0001) and GM-CSF (P < 0.01) release. The p38 MAP kinase inhibitor SB-203580 increased CAM-induced IL-8 release (P < 0.001), and the c-jun NH2-terminal kinase inhibitor SP-600125 had no effect on IL-8. At 120 min and 6 h, CAM increased phospho-ERK1/2 (pERK) but not phospho-p38 or phospho-JNK. Over the first 90 min, CAM at 10 microg/ml inhibited pERK and then increased pERK in parallel with measured IL-8 secretion. After daily CAM exposure for 5 days, both IL-8 and pERK returned to baseline. The p38 MAP kinase inhibitor, SB-203580 increased ERK phosphorylation and IL-8 secretion. These results suggest that macrolide antibiotics can differentially modulate proinflammatory cytokine secretion in NHBE cells, in part through ERK.

Verotoxin (shiga toxin) sensitizes renal epithelial cells to increased heme toxicity: possible implications for the hemolytic uremic syndrome.

Escherichia coli-derived verotoxins (VT; Shiga toxins) are causally related to the pathogenesis of enteropathic hemolytic uremic syndrome (HUS). Profound hemolysis is a defining feature of the disease, but it is not known whether the acute intravascular release of heme proteins contributes to HUS pathology. This study examined the biologic effects of hemin and VT by means of tubular epithelial-derived ACHN cells. Hemin at concentrations >/=200 microM caused cell rounding, spike formation, and detachment that was morphologically distinct from verocytotoxicity. VT caused apoptosis at concentrations >100 pM, as demonstrated by nuclear segmentation and poly(ADP-ribose) polymerase cleavage, whereas hemin-mediated injury of ACHN cells grown in serum-containing medium lacked attributes of programmed cell death. Pretreatment of ACHN monolayers with sublethal concentrations (1 to 10 pM) of VT for 12 to 18 h led to superadditive hemin-mediated cytotoxicity. This effect was not limited to ACHN cells, but was similarly noted in microvascular endothelial cells. Heme catabolism is regulated by (inducible) heme oxygenase-1 (HO-1). VT abrogated HO-1 expression in ACHN cells. Stimulation of cells for 6 h with CdCl(2), which markedly increased HO-1 expression before the addition of VT, blunted subsequent hemin injury. In conclusion, VT augments hemin-induced toxicity in renal tubular epithelial cells that can be reversed by prior induction of HO-1. It is proposed that VT subverts the physiologic defense against heme proteins by interfering with the regulated expression of HO-1 and that this mechanism contributes to the renal pathology in patients with Escherichia coli-associated HUS.

Shiga toxin 1-induced activation of c-Jun NH(2)-terminal kinase and p38 in the human monocytic cell line THP-1: possible involvement in the production of TNF-alpha.

Shiga toxin-producing enterohemorrhagic E. coli infections cause bloody diarrhea, which may progress to life-threatening complications such as the hemolytic-uremic syndrome (HUS). HUS patients frequently have elevated levels of the proinflammatory cytokine tumor necrosis factor alpha (TNF-alpha) detectable in urine. Thus, sequelae may develop following the localized production of proinflammatory cytokines within the kidneys. A possible source of these cytokines are macrophages, which respond to the toxins by producing TNF-alpha. We have shown previously that THP-1 cells produce soluble TNF-alpha in response to the toxins, whose production requires host-cell tyrosine-kinase activity and toxin-enzymatic activity. To further examine signaling pathways involved in TNF-alpha expression, we determined that JNK1 and -2 and p38, but not ERK1 or -2, were phosphorylated following toxin exposure. Blockade of p38 activation reduced TNF-alpha production following Shiga toxin 1 treatment. Finally, we present a model of the ribotoxic stress response triggered in human macrophages by Shiga toxins.