Hyperlipoproteinemia enhances susceptibility to acute disseminated Candida albicans infection in low-density-lipoprotein-receptor-deficient mice

Recent studies have suggested the use of lipoproteins as an adjuvant treatment of lethal gram-negative infections. However, other important microorganisms for the etiology of sepsis, such as Candida species, grow better in lipid-rich environments. We investigated the effect of hyperlipoproteinemia on systemic candidiasis in low-density-lipoprotein-receptor-deficient (LDLR-/-) mice, in which the loss of the receptor results in a seven- to ninefold-higher plasma LDL level than that in their wild-type littermates (C57BL/6J). LDLR-/- mice died earlier, and the outgrowth of Candida albicans in the kidneys and livers of LDLR-/- mice was significantly higher compared with that of controls. After infection, circulating cytokine concentrations were significantly higher in LDLR-/- mice. In vitro, C. albicans grew better in plasma samples of LDLR-/- mice than in control plasma samples and peritoneal macrophages of LDLR-/- mice challenged with heat-killed C. albicans produced more cytokines than did those of controls. This latter phenomenon was probably due to increased binding of yeast cells to macrophages of LDLR-/- mice. These data suggest that hyperlipoproteinemia is deleterious in systemic candidiasis.     

Validation of multifrequency bioelectrical impedance analysis in monitoring fluid balance in healthy elderly subjects

BACKGROUND: Multifrequency Bioelectrical Impedance Analysis (MFBIA) is a novel method to assess body composition in elderly subjects. However, it is unclear whether MFBIA can detect changes in body water compartments in elders. We aimed to determine the within-subject variability of MFBIA and the responsiveness to a diuretic intervention in aged subjects with a stable fluid balance. METHODS: We selected 12 healthy active elderly subjects (5 male, 7 female) with a mean age of 75 years. Total body water and extracellular fluid (ECF) were measured by deuterium oxide- and potassium bromide-dilution techniques. Within-subject variability in total body MFBIA was assessed by performing four measurements at 1, 5, 50, and 100 kHz within a 2-month period. Subsequently, responsiveness of MFBIA to the ECF loss caused by oral administration of 40 mg of furosemide was determined. RESULTS: Within-subject variability in MFBIA at 1, 5, 50, and 100 kHz expressed as standard deviations was 21, 19, 14, and 14 Ohm (omega), respectively. Furosemide caused a mean weight loss of 1.8 +/- 0.6 kg, which resulted in significant increases in impedance of 57 +/- 24 omega at 1 kHz and 37 +/- 12 omega at 100 kHz (p < .001). The responsiveness of MFBIA for the diuretic intervention was best at 5 kHz (responsiveness index = 1.98). CONCLUSIONS: Within-subject variability of MFBIA was small in healthy elderly subjects with stable fluid balance. Responsiveness of MFBIA to 9% furosemide-induced ECF loss was excellent. These data support the necessity for further clinical assessment of the value of MFBIA in monitoring fluid balance in geriatric patients.     

A dilemma in analysis: issues in the serial measurement of quality of life in patients with advanced lung cancer

Despite the availability of several instruments to evaluate quality of life (QL) over time in patients with lung cancer, barriers in measurement remain. This methodological study used LCSS data (Lung Cancer Symptom Scale, a disease- and site-specific QL measure) to examine analysis methods to quantify QL where data needed for serial evaluation may be missing. Data from two large randomized trials, conducted at 30 centers, of a new combination chemotherapy regimen incorporating a new agent for patients (n = 673) with Stage III and IV non-small cell lung cancer were obtained for this study. QL had been prospectively measured at baseline, day 29, and every six weeks thereafter using the LCSS. For the slope analysis (SA) and area under the curve (AUC) analyses, an adjustment score of zero was used to indicate QL on the day of death (mortality adjustment) and each subsequent day until the end of the assessment period. Significant differences in QL, symptom scores and known prognostic factors at baseline were found in the attrition group. SA and AUC analysis allowed inclusion of 581 patients, giving an adequacy rate of 86%. By using a mortality adjustment, an additional 45 patients were included, increasing the inclusion rate to 93%. With the use of the mortality adjustment, QL was shown to decline over the interval, as opposed to rise if the adjustment had not been performed. The conclusions of the study were: (1) analysis for serial data using SA and AUC provides useful, but differing information; (2) when attrition (caused by death) is a factor, a mortality adjustment presented a more accurate assessment of QL as an endpoint; (3) more frequent evaluations of QL will capture rapid changes in patient status and reduce the attrition bias; (4) all patients should be followed until they die; and (5) QL should be given full consideration as a primary endpoint separate from survival.     

Inhibition of double-stranded RNA-dependent protein kinase PKR by vaccinia virus E3: role of complex formation and the E3 N-terminal domain

The human double-stranded RNA (dsRNA)-dependent protein kinase PKR inhibits protein synthesis by phosphorylating translation initiation factor 2alpha (eIF2alpha). Vaccinia virus E3L encodes a dsRNA binding protein that inhibits PKR in virus-infected cells, presumably by sequestering dsRNA activators. Expression of PKR in Saccharomyces cerevisiae inhibits protein synthesis by phosphorylation of eIF2alpha, dependent on its two dsRNA binding motifs (DRBMs). We found that expression of E3 in yeast overcomes the lethal effect of PKR in a manner requiring key residues (Lys-167 and Arg-168) needed for dsRNA binding by E3 in vitro. Unexpectedly, the N-terminal half of E3, and residue Trp-66 in particular, also is required for anti-PKR function. Because the E3 N-terminal region does not contribute to dsRNA binding in vitro, it appears that sequestering dsRNA is not the sole function of E3 needed for inhibition of PKR. This conclusion was supported by the fact that E3 activity was antagonized, not augmented, by overexpressing the catalytically defective PKR-K296R protein containing functional DRBMs. Coimmunoprecipitation experiments showed that a majority of PKR in yeast extracts was in a complex with E3, whose formation was completely dependent on the dsRNA binding activity of E3 and enhanced by the N-terminal half of E3. In yeast two-hybrid assays and in vitro protein binding experiments, segments of E3 and PKR containing their respective DRBMs interacted in a manner requiring E3 residues Lys-167 and Arg-168. We also detected interactions between PKR and the N-terminal half of E3 in the yeast two-hybrid and lambda repressor dimerization assays. In the latter case, the N-terminal half of E3 interacted with the kinase domain of PKR, dependent on E3 residue Trp-66. We propose that effective inhibition of PKR in yeast requires formation of an E3-PKR-dsRNA complex, in which the N-terminal half of E3 physically interacts with the protein kinase domain of PKR.     

Sex steroids do not prevent amylin-induced apoptosis in human cells

Formation of amylin-containing islet amyloid deposits may contribute to the progressive deterioration of beta cell function in non-insulin-dependent diabetes mellitus. As diabetes mellitus occurs in male, but rarely in female transgenic mice expressing human amylin in their pancreatic beta cells, it is of interest to study the influence of estradiol (E2) and testosterone (T) on amylin-induced cytotoxicity in human cells. The insulinoma cell line CM, thyroid epithelial cells (TEC) in primary culture, and nontransformed fibroblast lines were used. The occurrence of apoptotic cell death was assessed by nuclear labeling with propidium iodide. Amylin was cytotoxic on all cell types tested, but had the most pronounced effect on TEC and the weakest on the CM cell line. Although both E2 and T decreased the proportion of apoptotic cells in cultures kept in the absence of amylin, neither of the two hormones was able to counteract amylin-induced cytotoxicity. beta cell death and hyperglycemia can thus presumably not be prevented by the neutralization of amylin effects by sex steroids.