Evaluation of the Luciferase Assay-Based In Vitro Elicitation Test for Serum IgE

ABSTRACTBackgroundAn in vitro elicitation test employing human high-affinity IgE receptor-expressing rat mast cell lines appears to be a useful method for measuring mast cell activation using a patient's IgE and an allergen; however, such cell lines are sensitive to human complements in the serum. We have recently developed a new luciferase-reporting mast cell line (RS-ATL8) to detect IgE crosslinking-induced luciferase expression (EXiLE) with relatively low quantities of serum IgE.MethodsA total of 30 patients suspected of having egg white (EW) allergy were subjected to an oral food challenge (OFC) test; then, the performances of EW-specific serum IgE (CAP-FEIA), EW-induced degranulation, and EXiLE responses in RS-ATL8 cells were compared using receiver-operating characteristic (ROC) curve analysis. The patients' sera were diluted to 1:100, which causes no cytotoxicity when sensitizing the RS-ATL8 cells for the degranulation and EXiLE tests.ResultsThe area under the ROC curves was highest in the EXiLE test (0.977), followed by CAP-FEIA (0.926) and degranulation (0.810). At an optimal cutoff range (1.648-1.876) calculated from the ROC curve of the EXiLE test, sensitivity and specificity were 0.944 and 0.917, respectively. A 95% positive predictive value was given at a cutoff level of 2.054 (fold increase in luciferase expression) by logistic regression analysis. Conclusions: In contrast to in vivo tests, the EXiLE test appears to be a useful tool in diagnosing patients suspected of having IgE-dependent EW allergy without the risk of severe systemic reactions

Tetranins: new putative spider mite elicitors of host plant defense

Summary The two‐spotted spider mite (Tetranychus urticae) is a plant‐sucking arthropod herbivore that feeds on a wide array of cultivated plants. In contrast to the well‐characterized classical chewing herbivore salivary elicitors that promote plant defense responses, little is known about sucking herbivores' elicitors. To characterize the sucking herbivore elicitors, we explored putative salivary gland proteins of spider mites by using an Agrobacterium‐mediated transient expression system or protein infiltration in damaged bean leaves. Two candidate elicitors (designated as tetranin1 (Tet1) and tetranin2 (Tet2)) triggered early leaf responses (cytosolic calcium influx and membrane depolarization) and increased the transcript abundances of defense genes in the leaves, eventually resulting in reduced survivability of T. urticae on the host leaves as well as induction of indirect plant defenses by attracting predatory mites. Tet1 and/or Tet2 also induced jasmonate, salicylate and abscisic acid biosynthesis. Notably, Tet2‐induced signaling cascades were also activated via the generation of reactive oxygen species. The signaling cascades of these two structurally dissimilar elicitors are mostly overlapping but partially distinct and thus they would coordinate the direct and indirect defense responses in host plants under spider mite attack in both shared and distinct manners

Three-dimensional Motion Analysis of Lip and Mandibular Movements during Mastication

Many aspects of the coordination of lip and mandibular movements in the process of eating have not yet been clarified. This time, aiming to objectively evaluate lip and mandibular movements when chewing, the movements of the corners of the mouth and the mandible during mastication were measured three-dimensionally and analyzed. The subjects were 20 healthy women with individual normal occlusion. The test food was a commercially-available biscuit with a weight of 1 g. With six measuring points set for the lips and pogonion, the movements at those measuring points were captured with two CCD cameras during mastication, and the resulting images were analyzed with a three-dimensional motion analysis system. The analysis result showed that X- and Z-axis movements occurred on the working-side corner of the mouth, with Z-axis movements preceding X-axis movements, while on the balancing-side corner of the mouth, X- and Z-axis movements occurred simultaneously. Data on the amount, time taken, and speed of movements measured at each anatomical landmark showed that the working-side corner of the mouth moved a greater distance at a faster pace and, therefore, in less time than that of the balancing-side corner of the mouth. This is conceivably due to the aforementioned differences in X- and Z-axis movements of the working-side and balancing-side corners of the mouth. Further comparisons and studies with expansion of the subjects to include children will be necessary

Evaluation of the Luciferase Assay-Based In Vitro Elicitation Test for Serum IgE

Background: An in vitro elicitation test employing human high-affinity IgE receptor-expressing rat mast cell lines appears to be a useful method for measuring mast cell activation using a patient's IgE and an allergen; however, such cell lines are sensitive to human complements in the serum. We have recently developed a new luciferase-reporting mast cell line (RS-ATL8) to detect IgE crosslinking-induced luciferase expression (EXiLE) with relatively low quantities of serum IgE. Methods: A total of 30 patients suspected of having egg white (EW) allergy were subjected to an oral food challenge (OFC) test; then, the performances of EW-specific serum IgE (CAP-FEIA), EW-induced degranulation, and EXiLE responses in RS-ATL8 cells were compared using receiver-operating characteristic (ROC) curve analysis. The patients' sera were diluted to 1:100, which causes no cytotoxicity when sensitizing the RS-ATL8 cells for the degranulation and EXiLE tests. Results: The area under the ROC curves was highest in the EXiLE test (0.977), followed by CAP-FEIA (0.926) and degranulation (0.810). At an optimal cutoff range (1.648-1.876) calculated from the ROC curve of the EXiLE test, sensitivity and specificity were 0.944 and 0.917, respectively. A 95% positive predictive value was given at a cutoff level of 2.054 (fold increase in luciferase expression) by logistic regression analysis. Conclusions: In contrast to in vivo tests, the EXiLE test appears to be a useful tool in diagnosing patients suspected of having IgE-dependent EW allergy without the risk of severe systemic reactions

Facilitation of brain mitochondrial activity by 5-aminolevulinic acid in a mouse model of Alzheimer's disease

The activities of mitochondrial enzymes, which are essential for neural function, decline with age and in age-related disease. In particular, the activity of cytochrome c oxidase (COX/complex IV) decreases in patients with Alzheimer's disease (AD). COX, a mitochondrial inner membrane protein complex that contains heme, plays an essential role in the electron transport chain that generates ATP. Heme synthesis begins with 5-aminolevulinic acid (5-ALA) in mitochondria. 5-ALA synthetase is the rate-limiting enzyme in heme synthesis, suggesting that supplementation with 5-ALA might help preserve mitochondrial activity in the aged brain. We administered a diet containing 5-ALA to triple-transgenic AD (3xTg-AD) model mice for 6 months, starting at 3 months of age. COX activity and protein expression, as well as mitochondrial membrane potential, were significantly higher in brains of 5-ALA-fed mice than in controls. Synaptotagmin protein levels were also significantly higher in 5-ALA-fed mice, suggesting improved preservation of synapses. Although brain A levels tended to decrease in 5-ALA-fed mice, we observed no other significant changes in other biochemical and pathological hallmarks of AD. Nevertheless, our study suggests that daily oral administration of 5-ALA could preserve mitochondrial enzyme activities in the brains of aged individuals, thereby contributing to the preservation of neural activity

Warming to human-skin temperature with moderate humidity significantly reduces the survival rate of human pathogenic bacteria on dry surfaces of a 96-well plastic plate.

The experiment was performed in a thermo-hygrostat incubator. Various combinations of temperature (T) (25°C -37°C) and humidity (M) (45%-90%) were adjusted by the controller installed in the incubator. See the Materials and Methods. A. PCA scatter plot showing the impact on the survival rate of E. coil of temperature (25°C–37°C) and humidity (45%–90%). PC1-factor loading shows ˗0.646 (T: temperature), ˗0.288 (M: humidity), and 0.707 (S: the survival rate of E. coli). PC2-factor loading shows ˗0.408 (T) and 0.913 (M). Six runs were performed for each combination of matrices. B. Comparison of the survival rate of E. coli on dry surfaces (18 h/0 h) with 45%–90% humidity at 25°C. Bars (n = 6) show the average ± SD. *, pC. Comparison of the survival rate of E. coli on dry surfaces (18 h/0 h) with 45%–90% humidity at 37°C. Bars (n = 6) show the average ± SD. *, p (PDF)</p

Low temperature and humidity in hospitals increases the survival rate of bacteria on high-touch dry surfaces.

A. PCA scatter plot showing four groups (Groups 1–4) with distinct environmental conditions [temperature (T: T1–3), humidity (M: M1–3), and number of people (N: N1–3)] in three hospitals [designated H, K, and M]. PC1-factor loading shows 0.248(T1)/ 0.275(T2)/ 0.297(T3), ˗0.419(M1)/ ˗0.398(M2)/ ˗0.424(M3), and ˗0.306(N1)/ ˗0.265(N2)/ ˗0.312(N3) (N: number of people at that time). PC2-factor loading shows ˗0.425(T1)/ ˗0.415(T2)/ ˗0.383(T3) and ˗0.404(N1)/ ˗0.392(N2)/ ˗0.423(N3). As specified in the text, the data (66 sets) published in our previous manuscript were reused [19]. “O” ward, Obstetrics. “S” ward, Surgery. “I” ward, Internal medicine. “%,” cumulative contribution rates for PC1 and PC2. See S1 Table. B. Comparison of the number of live bacteria (CFU) on dry surfaces between Groups 1–4. Bars [Group 1 (n = 108), Group 2 (n = 135), Group 3 (n = 144), and Group 4 (n = 189)] show the average ± SD. *, pC. Comparison of the amount of ATP (as an index for the frequency of “human contact”) on dry surfaces between Groups 1–4. Bars [Group 1 (n = 117), Group 2 (n = 135), Group 3 (n = 157), and Group 4 (n = 189)] show the average ± SD.</p