Improved variance estimation of classification performance via reduction of bias caused by small sample size

Background  

Supervised learning for classification of cancer employs a set of design examples to learn how to discriminate between tumors. In practice it is crucial to confirm that the classifier is robust with good generalization performance to new examples, or at least that it performs better than random guessing. A suggested alternative is to obtain a confidence interval of the error rate using repeated design and test sets selected from available examples. However, it is known that even in the ideal situation of repeated designs and tests with completely novel samples in each cycle, a small test set size leads to a large bias in the estimate of the true variance between design sets. Therefore different methods for small sample performance estimation such as a recently proposed procedure called Repeated Random Sampling (RSS) is also expected to result in heavily biased estimates, which in turn translates into biased confidence intervals. Here we explore such biases and develop a refined algorithm called Repeated Independent Design and Test (RIDT).     

Behavioural fever boosts the inflammatory response in rainbow trout Oncorhynchus mykiss

Behavioural fever, manifested as an increased preferred temperature, was shown in rainbow trout Oncorhynchus mykiss following an injection of bacterial lipopolysaccharide. Simulated behavioural fever, through a 2·5° C water temperature rise following bacterial lipopolysaccharide injection, enhanced the expression of the cytokine interleukin-1β, in comparison with an untreated group held at the initial temperature. The present findings show that an important mediator in the immune response can be boosted through behavioural fever in fishes.     

Influence of chronic and acute spinal cord injury on skeletal muscle Na+-K+-ATPase and phospholemman expression in humans

Na(+)-K(+)-ATPase is an integral membrane protein crucial for the maintenance of ion homeostasis and skeletal muscle contractibility. Skeletal muscle Na(+)-K(+)-ATPase content displays remarkable plasticity in response to long-term increase in physiological demand, such as exercise training. However, the adaptations in Na(+)-K(+)-ATPase function in response to a suddenly decreased and/or habitually low level of physical activity, especially after a spinal cord injury (SCI), are incompletely known. We tested the hypothesis that skeletal muscle content of Na(+)-K(+)-ATPase and the associated regulatory proteins from the FXYD family is altered in SCI patients in a manner dependent on the severity of the spinal cord lesion and postinjury level of physical activity. Three different groups were studied: 1) six subjects with chronic complete cervical SCI, 2) seven subjects with acute, complete cervical SCI, and 3) six subjects with acute, incomplete cervical SCI. The individuals in groups 2 and 3 were studied at months 1, 3, and 12 postinjury, whereas individuals with chronic SCI were compared with an able-bodied control group. Chronic complete SCI was associated with a marked decrease in [(3)H]ouabain binding site concentration in skeletal muscle as well as reduced protein content of the α(1)-, α(2)-, and β(1)-subunit of the Na(+)-K(+)-ATPase. In line with this finding, expression of the Na(+)-K(+)-ATPase α(1)- and α(2)-subunits progressively decreased during the first year after complete but not after incomplete SCI. The expression of the regulatory protein phospholemman (PLM or FXYD1) was attenuated after complete, but not incomplete, cervical SCI. In contrast, FXYD5 was substantially upregulated in patients with complete SCI. In conclusion, the severity of the spinal cord lesion and the level of postinjury physical activity in patients with SCI are important factors controlling the expression of Na(+)-K(+)-ATPase and its regulatory proteins PLM and FXYD5.     

Placental transfer of maternally-derived IgA precludes the use of guthrie card eluates as a screening tool for primary immunodeficiency diseases

There is a need for neonatal screening tools to improve the long-term clinical outcome of patients with primary immunodeficiency diseases (PID). Recently, a PCR-based screening method for both TRECs and KRECs using Guthrie card samples has been developed. However, the applicability of these excision circle assays is limited to patients with severe T or B cell lymphopenia (SCID, XLA and A-T), whereas the most common forms of PID are not detected. Absence of serum IgA is seen in a major fraction of patients with immunological defects. As serum IgA in newborns is considered to be of fetal origin, eluates from routinely collected dried blood spot samples might thus be suitable for identification of children with PID. To assess the applicability of such screening assays, stored Guthrie card samples were obtained from 47 patients with various forms of primary immunodeficiency diseases (SCID, XLA, A-T, HIGM and IgAD), 20 individuals with normal serum IgA levels born to IgA-deficient mothers and 51 matched healthy newborns. Surprisingly, normal serum IgA levels were found in all SCID, XLA, A-T and HIGM patients and, additionally, in all those IgAD patients born to IgA-sufficient mothers. Conversely, no serum IgA was found in any of the 16 IgAD patients born by IgA-deficient mothers. Moreover, half of the IgA-sufficient individuals born by IgA-deficient mothers also lacked IgA at birth whereas no IgA-deficient individuals were found among the controls. IgA in neonatal dried blood samples thus appears to be of both maternal and fetal origin and precludes its use as a reliable marker for neonatal screening of primary immunodeficiency diseases.     

Proteomics and lipids of lipoproteins isolated at low salt concentrations in D2O/sucrose or in KBr

There is much interest in the significance of apolipoproteins and proteins that are noncovalently associated with lipoproteins. It is possible that the high ionic strength used for isolation of lipoproteins with KBr and NaI could alter the pattern of associated exchangeable proteins. Here we describe lipoprotein classes fractionation from up to 0.5 ml of serum or plasma with buffers of physiological ionic strength and pH prepared with deuterium oxide (D(2)O) and sucrose. An advantage of the D(2)O/sucrose procedure was that the lipoproteins could be directly analyzed by the techniques described without need for desalting. We compared the isolated lipoproteins with those obtained using ultracentrifugation in KBr from the same plasma pool. Electrophoretic homogeneity of the lipoproteins was very similar using the two methods, as well as their lipid composition evaluated by HPLC. Two-dimensional electrophoresis and surface-enhanced laser adsorption/ionization time-of-flight mass spectrometry indicated that the patterns of exchangeable proteins of VLDL isolated using with the two procedures were very similar. However, significant differences were found in the profiles of LDL and HDL, indicating that the D(2)O/sucrose method allowed a more complete characterization of its exchangeable apolipoproteins and proteins.     

The combination of raloxifene and epigallocatechin gallate suppresses growth and induces apoptosis in MDA-MB-231 cells

Previous studies have demonstrated that raloxifene induces apoptosis in a variety of cancer cell lines. We aimed to determine if this effect was enhanced by combining raloxifene with epigallocatechin gallate (EGCG). Results demonstrated that EGCG (25 microM) and raloxifene (1-5 microM) produced enhanced cytotoxicity toward MDA-MB-231 breast cancer cells compared to either drug alone following 7 days of treatment. The combination of 5 microM raloxifene and EGCG was the most effective as it decreased cell number by 96% of control, and time-course studies demonstrated that significant cytotoxicity began 36 h after treatment. Potential mechanisms for this effect were then investigated. Flow cytometry experiments demonstrated that apoptosis was significantly increased following 12 h of combination treatment compared to all other treatment groups. A maximal increase in the proportion of cells in the G(1)-phase of the cell cycle (116% of control) occurred following 24 h of combination treatment, 12 h after the significant increase in apoptosis, and thus was not considered to be a viable mechanism for the enhancement of apoptosis. While raloxifene was a competitive inhibitor of microsomal UDP-glucuronosyltransferase activity (K(i) of 24 microM), it did not decrease the metabolism of EGCG as the rate of disappearance of EGCG from the media was the same for cells treated with either EGCG or EGCG+raloxifene. Finally, the combination treatment reduced the phosphorylation of EGFR and AKT proteins by 21.2+/-3.3% and 31.5+/-1.7% from control, respectively. In conclusion, the synergistic cytotoxicity elicited by the combination of EGCG and raloxifene results from an earlier and greater induction of apoptosis. This is likely to be a result of reduced phosphorylation of EGFR and AKT signaling proteins.     

Root uptake of cationic amino acids by Arabidopsis depends on functional expression of amino acid permease 5

* Specific transporters mediate uptake of amino acids by plant roots. Earlier studies have indicated that the lysine histidine transporter 1 and amino acid permease 1 participate in this process, but although plant roots have been shown to absorb cationic amino acids with high affinity, neither of these transporters seems to mediate transport of L-arginine (L-Arg) or L-lysine (L-Lys). * Here, a collection of T-DNA knockout mutants were screened for alterations in Arabidopsis root uptake rates of L-Arg and it was found that only the AAP5 mutant displayed clear phenotypic divergence on high concentrations of L-Arg. A second screen using low concentrations of (15)N-labelled L-Arg in the growth media also identified AAP5 as being involved in L-Arg acquisition. * Momentaneous root uptake of basic amino acids was strongly affected in AAP5 mutant lines, but their uptake of other types of amino acids was only marginally affected. Comparisons of the root uptake characteristics of AAP5 and LHT1 mutants corroborated the hypothesis that the two transporters have distinct affinity spectra in planta. * Root uptake of all tested amino acids, except L-aspartic acid (L-Asp), was significantly affected in double AAP5*LHT1 mutants, suggesting that these two transporters account for a major proportion of roots' uptake of amino acids at low concentrations.