Differentiation of human embryonic stem cells and human induced pluripotent stem cells into steroid-producing cells.

Although there have been reports of the differentiation of mesenchymal stem cells and mouse embryonic stem (ES) cells into steroid-producing cells, the differentiation of human ES/induced pluripotent stem (iPS) cells into steroid-producing cells has not been reported. The purpose of our present study was to establish a method for inducing differentiation of human ES/iPS cells into steroid-producing cells. The first approach we tried was embryoid body formation and further culture on adherent plates. The resultant differentiated cells expressed mRNA encoding the steroidogenic enzymes steroidogenic acute regulatory protein, 3β-hydroxysteroid dehydrogenase, cytochrome P450-containing enzyme (CYP)-11A1, CYP17A1, and CYP19, and secreted progesterone was detected in the cell medium. However, expression of human chorionic gonadotropin was also detected, suggesting the differentiated cells were trophoblast like. We next tried a multistep approach. As a first step, human ES/iPS cells were induced to differentiate into the mesodermal lineage. After 7 d of differentiation induced by 6-bromoindirubin-3'-oxime (a glycogen synthase kinase-3β inhibitor), the human ES/iPS cells had differentiated into fetal liver kinase-1- and platelet derived growth factor receptor-α-expressing mesodermal lineage cells. As a second step, plasmid DNA encoding steroidogenic factor-1, a master regulator of steroidogenesis, was introduced into these mesodermal cells. The forced expression of steroidogenic factor-1 and subsequent addition of 8-bromoadenosine 3',5'-cyclic monophosphate induced the mesodermal cells to differentiate into the steroidogenic cell lineage, and expression of CYP21A2 and CYP11B1, in addition to steroidogenic acute regulatory protein, 3β-hydroxysteroid dehydrogenase, CYP11A1, and CYP17A1, was detected. Moreover, secreted cortisol was detected in the medium, but human chorionic gonadotropin was not. These findings indicate that the steroid-producing cells obtained through the described multistep method are not trophoblast like; instead, they exhibit characteristics of adrenal cortical cells

Noise robust 2D bird localization via sound using microphone arrays

Birds in the wild are difficult to localize, because their sizes tend to be small, they move swiftly, and they are often visually occluded. However, their location information is crucial for ethological studies on birds' behaviour. Recently, automating the process has been studied as a hot topic, where spatial sensors and sensor networks are commonly used. To avoid the visual occlusion problem, many studies focus on acoustic signal processing by applying microphone arrays and perform 1D azimuth localization through bird songs. In this study, we perform 2D sound source localization in the Cartesian coordinates using azimuths from multiple microphone arrays. To estimate the exact bird's location, we calculate the intersection points of these azimuth lines. Although this approach is simple and easy to be implemented, it has two main issues. One is that even small noise interference in azimuth values results in corrupting the localization data. This leads to a problem, where the intersection points between the azimuth lines do not intersect in one point for a single bird, but in several points. This proves difficulty in estimating the exact location of each bird. Especially in a far-field application, even small noise corruption leads to large localization errors. The other issue is that in the bird's natural habitat, elements such as leaves, grass and rivers are natural noise sources. It is difficult to extract the bird songs in such a noisy environment. We propose an algorithm involving statistic methods, sound feature analysis and machine learning. Based on this approach, a noise robust bird localization system has been established. We have performed numerous simulations to further understand the limitations of the system. Based on the results we have also derived the system's design guidelines, describing how the results change depending on the number of microphone arrays, signal-to-noise ratio, bird's distance from the devices, array's transfer function, type of the singing bird and specific parameter settings used in the algorithms. Such detailed guidelines support interested researchers in creating a similar system, which can contribute to ethological researches

Crystal structure of a Ca2+-dependent regulator of flagellar motility reveals the open-closed structural transition

Sperm chemotaxis toward a chemoattractant is very important for the success of fertilization. Calaxin, a member of the neuronal calcium sensor protein family, directly acts on outer-arm dynein and regulates specific flagellar movement during sperm chemotaxis of ascidian, Ciona intestinalis. Here, we present the crystal structures of calaxin both in the open and closed states upon Ca2+ and Mg2+ binding. The crystal structures revealed that three of the four EF-hands of a calaxin molecule bound Ca2+ ions and that EF2 and EF3 played a critical role in the conformational transition between the open and closed states. The rotation of α7 and α8 helices induces a significant conformational change of a part of the α10 helix into the loop. The structural differences between the Ca2+- and Mg2+-bound forms indicates that EF3 in the closed state has a lower affinity for Mg2+, suggesting that calaxin tends to adopt the open state in Mg2+-bound form. SAXS data supports that Ca2+-binding causes the structural transition toward the closed state. The changes in the structural transition of the C-terminal domain may be required to bind outer-arm dynein. These results provide a novel mechanism for recognizing a target protein using a calcium sensor protein

Omega-3 Polyunsaturated Fatty Acids Suppress the Cystic Lesion Formation of Peritoneal Endometriosis in Transgenic Mouse Models

Omega-3 polyunsaturated fatty acids (omega-3 PUFAs) play a role in controlling pathological inflammatory reactions. Endometriosis is characterized by the presence of endometrial tissue on the peritoneum and an exaggerated inflammatory environment around ectopic tissues. Here peritoneal endometriosis was reproduced using a mouse model in which murine endometrial fragments were inoculated into the peritoneal cavity of mice. Fat-1 mice, in which omega-6 can be converted to omega-3 PUFAs, or wild type mice, in which it cannot, were used for the endometriosis model to address the actions of omega-3 PUFAs on the development of endometriotic lesions. The number and weight of cystic endometriotic lesions in fat-1 mice two weeks after inoculation were significantly less than half to those of controls. Mediator lipidomics revealed that cystic endometriotic lesions and peritoneal fluids were abundant in 12/15-hydroxyeicosapentaenoic acid (12/15-HEPE), derived from eicosapentaenoic acid (EPA), and their amount in fat-1 mice was significantly larger than that in controls. 12/15-Lipoxygenase (12/15-LOX)-knockout (KO) and control mice with or without EPA administration were assessed for the endometriosis model. EPA administration decreased the number of lesions in controls but not in 12/15-LOX-KO mice. The peritoneal fluids in EPA-fed 12/15-LOX-KO mice contained reduced levels of EPA metabolites such as 12/15-HEPE and EPA-derived resolvin E3 even after EPA administration. cDNA microarrays of endometriotic lesions revealed that Interleukin-6 (IL-6) expression in fat-1 mice was significantly lower than that in controls. These results suggest that both endogenous and exogenous EPA-derived PUFAs protect against the development of endometriosis through their anti-inflammatory effects and, in particular, the 12/15-LOX-pathway products of EPA may be key mediators to suppress endometriosis

A Proteomic Approach for the Diagnosis of ‘Oketsu’ (blood stasis), a Pathophysiologic Concept of Japanese Traditional (Kampo) Medicine

‘Oketsu’ is a pathophysiologic concept in Japanese traditional (Kampo) medicine, primarily denoting blood stasis/stagnant syndrome. Here we have explored plasma protein biomarkers and/or diagnostic algorithms for ‘Oketsu’. Sixteen rheumatoid arthritis (RA) patients were treated with keishibukuryogan (KBG), a representative Kampo medicine for improving ‘Oketsu’. Plasma samples were diagnosed as either having an ‘Oketsu’ (n = 19) or ‘non-Oketsu’ (n = 29) state according to Terasawa's ‘Oketsu’ scoring system. Protein profiles were obtained by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS) and hierarchical clustering and decision tree analyses were performed. KBG treatment for 4 or 12 weeks decreased the ‘Oketsu’ scores significantly. SELDI protein profiles gave 266 protein peaks, whose expression was significantly different between the ‘Oketsu’ and ‘non-Oketsu’ states. Hierarchical clustering gave three major clusters (I, II, III). The majority (68.4%) of ‘Oketsu’ samples were clustered into one cluster as the principal component of cluster I. The remaining ‘Oketsu’ profiles constituted a minor component of cluster II and were all derived from patients cured of the ‘Oketsu’ state at 12 weeks. Construction of the decision tree addressed the possibility of developing a diagnostic algorithm for ‘Oketsu’. A reduction in measurement/pre-processing conditions (from 55 to 16) gave a similar outcome in the clustering and decision tree analyses. The present study suggests that the pathophysiologic concept of Kampo medicine ‘Oketsu’ has a physical basis in terms of the profile of blood proteins. It may be possible to establish a set of objective criteria for diagnosing ‘Oketsu’ using a combination of proteomic and bioinformatics-based classification methods

Collimator for Variable Sensitivity and Spatial Resolution Without the Need for Exchange

A new design of collimator is proposed that has variable sensitivity and spatial resolution, eliminating the need for exchanging collimators in a gamma camera. Using Monte Carlo simulations, the present article evaluates the shielding of undesirable gamma rays in a parallel-hole collimator. It consists of a number of layers of rectangular holes. These layers consist of alternately stacked fixed and movable collimators. In high-resolution mode, the movable collimators are shifted by half the aperture pitch along the diagonal direction. The first collimator (type A) has 50 layers with fixed thicknesses of 1.2 mm. The second collimator (type B) has 25 layers with a thickness of 1.0 mm on the object side and 25 layers with a thickness of 1.4 mm on the opposite side. The third collimator (type C) has 20 layers with non-uniform thicknesses. The ratios of the maximum artificial peak to the main-peak are calculated for point-source responses. The ratios for types A, B, and C collimators are 0.78, 0.08, and 0.03, respectively. The same performance for shielding undesirable gamma rays is achieved in the type C collimator as for a conventional collimator

Cows painted with zebra-like striping can avoid biting fly attack

Experimental and comparative studies suggest that the striped coats of zebras can prevent biting fly attacks. Biting flies are serious pests of livestock that cause economic losses in animal production. We hypothesized that cows painted with black and white stripes on their body could avoid biting fly attacks and show fewer fly-repelling behaviors. Six Japanese Black cows were assigned to treatments using a 3 × 3 Latin-square design. The treatments were black-and-white painted stripes, black painted stripes, and no stripes (all-black body surface). Recorded fly-repelling behaviors were head throw, ear beat, leg stamp, skin twitch, and tail flick. Photo images of the right side of each cow were taken using a commercial digital camera after every observation and biting flies on the body and each leg were counted from the photo images. Here we show that the numbers of biting flies on Japanese Black cows painted with black-and-white stripes were significantly lower than those on non-painted cows and cows painted only with black stripes. The frequencies of fly-repelling behaviors in cows painted with black-and-white stripes were also lower than those in the non-painted and black-striped cows. These results thus suggest that painting black-and-white stripes on livestock such as cattle can prevent biting fly attacks and provide an alternative method of defending livestock against biting flies without using pesticides in animal production, thereby proposing a solution for the problem of pesticide resistance in the environment

Gene expression profiling of HiMAC-irradiated normal human fibroblasts by HiCEP

Gene expression profiling of normal human fibroblasts irradiated with heavy ion particles by a novel analysis methodFujimori, A., Suetomi,K., Kojima, A., Fang, Y-Q., Egusa, A., Takahashi, S., Okayasu, R.HiCEP (High-coverage expression profiling) is a novel comprehensive analysis method which is based on DNA finger printing and PCR amplification. It enables to detect any altered gene expression among 60-70% of all the actually transcribed genes in any eukaryotic cells and tissues. We previously applied HiCEP to a primary culture of normal human fibroblasts and observed gene expression responding to X-ray at the very low dose (10 mGy). As the result of screening approximately 23,000 transcripts, we have identified a set of CXC chemokines (CXCL1, CXCL2, CXCL6 and CXCL8) up-regulated by the 10 mGy X-rays in the normal human fibroblasts (HFLIII) (Cancer Res 2005; 65: 10159-10163). Those genes have hardly been expected from the previous studies using the higher (>100 mGy) doses of radiation. Our observation indicated that different molecular mechanisms are involved in the response to ionizing radiation with different doses /dose rates, suggesting that different cellular responses could be induced by ionizing radiation with different LETs. Accelerated heavy ion particles (at high LET) provide promising effects for radiotherapy of certain types of malignancy, however, the molecular basis of its advantage to gamma rays is not fully understood.This time, we applied HiCEP to normal human fibroblasts (HFL III) irradiated with high-LET radiation generated in our institute. More than 40 genes were found to be up-regulated in the irradiated cells by 3 folds within 4 hrs post-irradiation of carbon ion at 2Gy (70 keV/micro m). Those included the DNA damage-inducible genes (CDKN1A, CyclinG, Gadd45a), and also some unexpected genes, both predicted and unpredicted from the current public databases.53th Annual Meeting of the Radiation Research Societ