MalariaCount: an image analysis-based program for the accurate determination of parasitemia

Malaria is a serious global health problem and rapid, precise determination of parasitemia is necessary for malaria research and in clinical settings. Manual counting by light microscopy is the most widely used technique for parasitemia determination but it is a time-consuming and laborious process. The aim of our study was to develop an automated image analysis-based system for the rapid and accurate determination of parasitemia. We have developed, for the first time, a software, MalariaCount, that automatically generates parasitemias from images of Giemsa-stained blood smears. The potential application and robustness of MalariaCount was tested in normal and drug-treated in vitro cultures of Plasmodium falciparum. The results showed a tight correlation between MalariaCount and manual count parasitemia values. These findings suggest that MalariaCount can potentially be used as a tool to provide rapid and accurate determination of parasitemia in research laboratories where frequent, large-scale, efficient determination of parasitemia is required.     

iPS cells: a game changer for future medicine

The induced pluripotent stem cell (iPSC) technology is instrumental in advancing the fields of disease modeling and cell transplantation. We herein discuss the various issues regarding disease modeling and cell transplantation presented in previous reports, and also describe new iPSC‐based medicine including iPSC clinical trials. In such trials, iPSCs from patients can be used to predict drug responders/non‐responders by analyzing the efficacy of the drug on iPSC‐derived cells. They could also be used to stratify patients after actual clinical trials, including those with sporadic diseases, based on the drug responsiveness of each patient in the clinical trials. iPSC‐derived cells can be used for the identification of response markers, leading to increased success rates in such trials. Since iPSCs can be used in micromedicine for drug discovery, and in macromedicine for actual clinical trials, their use would tightly connect both micro‐ and macromedicine. The use of iPSCs in disease modeling, cell transplantation, and clinical trials could therefore lead to significant changes in the future of medicine.     

Allergen induced histamine release and immunoreactive-leukotriene C4 generation from leukocytes in mite sensitive asthmatic patients

Leukocytes from mite sensitive asthmatic patient were challenged with the allergen and the supernatant was assayed for histamine and immunoreactive-leukotriene C₄ (i-LTC₄). The release of histamine was quantitated by an automated fluorometric technique and i-LTC₄ was determined using a commercial radioimmunoassay kit. The results of analysis of the supernatant by high speed liquid chromatography, together with observations of modulation of the formation by agents, indicated that i-LTC₄ consisted of LTC₄ with a little amount of LTD₄. i-LTC₄ was generated as a result of basophil activation but not derived from the other cells such as monocytes and eosinophils. Allergen induced a concentration-dependent release of histamine and i-LTC₄ and the maximal release of histamine and i-LTC₄ occurred at the same dose of the allergen. At optimal concentration of the allergen, basophils produced 20.4 ± 17.9 ng of i-LTC₄/10⁶-cells (mean ± S.D., n=39) and histamine release was 55.6 ± 20.1% of total histamine. There was a significant correlation in the capacity of leukocytes to release histamine and i-LTC₄ (r=0.47, p<0.01). We found a correlation between maximal histamine release or cell sensitivity, allergen concentration for 50% histamine release, and a ratio of specific IgE to mite to total IgE in the serum, but the amount of i-LTC₄ failed to correlate significantly with the ratio. The releasability and the cell sensitivity of asthmatic patients' cells to the allergen for histamine release paralleled the severity to symptoms, but this correlation was not significant in i-LTC₄ generation.     

An investigation of intergeneric relations among Papionini monkeys based on fine hair medulla structures

A scanning electron microscopic examination of the hair medulla was performed on 12 species of Papionini monkeys in 5 genera. Their medullae can be roughly divided into two types relative to the structures. The results reinforced the phylogenetic relations found in molecular-based studies. Therefore, examination of hair medulla structures may be useful for inferring genetic affinities among the Papionini; species having similar medulla structures were closely related each other.     

Profiling of Phospho-AKT,Phospho-mTOR,Phospho-MAPK and EGFR in Non-small Cell Lung Cancer

Activation of numerous pathways has been documented in non-small cell lung cancer (NSCLC). Epidermal growth factor receptor (EGFR) has emerged as a common therapeutic target. The mitogen-activated protein kinase (MAPK) and AKT signaling pathways are downstream of EGFR and deregulated via genetic and epigenetic mechanisms in many human cancers. We evaluated selected markers in the EGFR pathway with reference to outcome. Tissues from 220 cases of NSCLC patients presented in a tissue microarray were assayed with immunohistochemistry for phosphorylated AKT, phosphorylated MAPK, phosphorylated mTOR, and EGFR and then quantified by automated image analysis. Individually, the biomarkers did not predict. Combined as ratios, p-mTOR/p-AKT, and p-MAPK/EGFR function as prognostic markers of survival (p=0.008 and p=0.029, respectively), however, no significance was found after adjustment (p=0.221, p=0.103). The sum of these ratios demonstrates a stronger correlation with survival (p<0.001) and remained statistically significant after adjustment (p=0.026). The algebraic combination of biomarkers offer the capacity to understand factors that predict outcome better than current approaches of evaluating biomarkers individually or in pairs. Our results show the sum of p-mTOR/p-AKT and p-MAPK/EGFR is a potential predictive marker of survival in NSCLC patients.     

Phylogenetic relationships ofPythium species based on ITS and 5.8S sequences of the ribosomal DNA

The sequences of ITS regions in 30 species and two groups of the genusPythium were resolved. In the phylogenetic trees, the species were generally divided into two clusters, referred to here as the F and S groups. The species in the two groups correspond in terms of their sporangial morphology, with the F group being filamentous/lobulate and the S group being spherical. Genetic divergence within the F group was lower than that within the S group. Other morphological characteristics such as oogonial structure and sexual nature appeared to be unrelated to the groupings in these trees. An alignment analysis revealed common sequences to all the species and arrangements specific to each F or S group. It was found that the ITS region was a good target in designing species-specific primers for the identification and detection ofPythium species. In the tree based on 5.8S rDNA sequences, oomycetes are distantly related to other fungi but separated from algae in Chromista.