Formation of a primitive ectoderm like cell population, EPL cells, from ES cells in response to biologically derived factors

The primitive ectoderm of the mouse embryo arises from the inner cell mass between 4.75 and 5.25 days post coitum, around the time of implantation. Positioned at a pivotal time in development, just prior to formation of the three germ layers of the embryo proper, the primitive ectoderm responds directly to the signals generated during gastrulation. We have identified a conditioned medium, MEDII, which caused the homogeneous conversion of ES cells to a morphologically distinct cell population, termed early primitive ectoderm-like (EPL) cells. EPL cells expressed the pluripotent cell markers Oct4, SSEA1 and alkaline phosphatase. However, the formation of EPL cells was accompanied by alterations in Fgf5, Gbx2 and Rex1 expression, a loss in chimaera forming ability, changes in factor responsiveness and modified differentiation capabilities, all consistent with the identification of EPL cells as equivalent to the primitive ectoderm population of the 5.5 to 6.0 days post coitum embryo. EPL cell formation could be reversed in the presence of LIF and withdrawal of MEDII, which suggested that EPL cell formation was not a terminal differentiation event but reflected the ability of pluripotent cells to adopt distinct cell states in response to specific factors. Partial purification of MEDII revealed the presence of two separable biological activities, both of which were required for the induction and maintenance of EPL cells. We show here the first demonstration of uniform differentiation of ES cells in response to biological factors. The formation of primitive ectoderm, both in vivo and in vitro, appears to be an obligatory step in the differentiation of the inner cell mass or ES cells into cell lineages of the embryonic germ layers. EPL cells potentially represent a model for the development of lineage specific differentiation protocols and analysis of gastrulation at a molecular level. An understanding of the active components of MEDII may provide a route for the identification of factors which induce primitive ectoderm formation in vivo.Joy Rathjen, Julie-Anne Lake, Michael D. Bettess, Jenny M. Washington, Gavin Chapman and Peter D. Rathje

Tomographic imaging and scanning thermal microscopy: thermal impedance tomography

The application of tomographic imaging techniques developed for medical applications to the data provided by the scanning thermal microscope will give access to true three-dimensional information on the thermal properties of materials on a mm length scale. In principle, the technique involves calculating and inverting a sensitivity matrix for a uniform isotropic material, collecting ordered data at several modulation frequencies, and multiplying the inverse of the matrix with the data vector. In practice, inversion of the matrix in impractical, and a novel iterative technique is used. Examples from both simulated and real data are given

Parallel computation of 3-D soil-structure interaction in time domain with a coupled FEM/SBFEM approach

The final publication is available at Springer via http://dx.doi.org/10.1007/s10915-011-9551-xThis paper introduces a parallel algorithm for the scaled boundary finite element method (SBFEM). The application code is designed to run on clusters of computers, and it enables the analysis of large-scale soil-structure-interaction problems, where an unbounded domain has to fulfill the radiation condition for wave propagation to infinity. The main focus of the paper is on the mathematical description and numerical implementation of the SBFEM. In particular, we describe in detail the algorithm to compute the acceleration unit impulse response matrices used in the SBFEM as well as the solvers for the Riccati and Lyapunov equations. Finally, two test cases validate the new code, illustrating the numerical accuracy of the results and the parallel performances. © Springer Science+Business Media, LLC 2011.Jose E. Roman and Enrique S. Quintana-Orti were partially supported by the Spanish Ministerio de Ciencia e Innovacion under grants TIN2009-07519, and TIN2008-06570-C04-01, respectively.Schauer, M.; Román Moltó, JE.; Quintana Orti, ES.; Langer, S. (2012). Parallel computation of 3-D soil-structure interaction in time domain with a coupled FEM/SBFEM approach. Journal of Scientific Computing. 52(2):446-467. doi:10.1007/s10915-011-9551-xS446467522Anderson, E., Bai, Z., Bischof, C., Demmel, J., Dongarra, J., Croz, J.D., Greenbaum, A., Hammarling, S., McKenney, A., Sorensen, D.: LAPACK User’s Guide. Society for Industrial and Applied Mathematics, Philadelphia (1992)Antes, H., Spyrakos, C.: Soil-structure interaction. In: Beskos, D., Anagnotopoulos, S. (eds.) Computer Analysis and Design of Earthquake Resistant Structures, p. 271. Computational Mechanics Publications, Southampton (1997)Appelö, D., Colonius, T.: A high-order super-grid-scale absorbing layer and its application to linear hyperbolic systems. J. Comput. Phys. 228(11), 4200–4217 (2009)Astley, R.J.: Infinite elements for wave problems: a review of current formulations and a assessment of accuracy. Int. J. Numer. Methods Eng. 49(7), 951–976 (2000)Balay, S., Buschelman, K., Eijkhout, V., Gropp, W.D., Kaushik, D., Knepley, M., McInnes, L.C., Smith, B.F., Zhang, H.: PETSc users manual. Tech. Rep. ANL-95/11 - Revision 3.1, Argonne National Laboratory (2010)Benner, P.: Contributions to the numerical solution of algebraic Riccati equations and related eigenvalue problems. Dissertation, Fak. f. Mathematik, TU Chemnitz–Zwickau, Chemnitz, FRG (1997)Benner, P.: Numerical solution of special algebraic Riccati equations via an exact line search method. In: Proc. European Control Conf. ECC 97, Paper 786, BELWARE Information Technology, Waterloo (B) (1997)Benner, P., Quintana-Ortí, E.: Solving stable generalized Lyapunov equations with the matrix sign function. Numer. Algorithms 20(1), 75–100 (1999)Benner, P., Byers, R., Quintana-Ortí, E., Quintana-Ortí, G.: Solving algebraic Riccati equations on parallel computers using Newton’s method with exact line search. Parallel Comput. 26(10), 1345–1368 (2000)Benner, P., Quintana-Ortí, E.S., Quintana-Ortí, G.: Solving linear-quadratic optimal control problems on parallel computers. Optim. Methods Softw. 23(6), 879–909 (2008)Bettess, P.: Infinite Elements. Penshaw Press, Sunderland (1992)Blackford, L.S., Choi, J., Cleary, A., D’Azevedo, E., Demmel, J., Dhillon, I., Dongarra, J., Hammarling, S., Henry, G., Petitet, A., Stanley, K., Walker, D., Whaley, R.C.: ScaLAPACK Users’ Guide. Society for Industrial and Applied Mathematics, Philadelphia (1997)Borsutzky, R.: Braunschweiger Schriften zur Mechanik - Seismic Risk Analysis of Buried Lifelines, vol. 63. Mechanik-Zentrum Technische Universität. Braunschweig (2008)Dongarra, J.J., Whaley, R.C.: LAPACK working note 94: A user’s guide to the BLACS v1.1. Tech. Rep. 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Prediction of 3D grinding temperature field based on meshless method considering infinite element

© 2018, Springer-Verlag London Ltd., part of Springer Nature. A three-dimensional numerical model to calculate the grinding temperature field distribution is presented. The finite block method, which is developed from meshless method, is used to deal with the stationary and the transient heat conduction problems in this paper. The influences of workpiece feed velocity, cooling coefficient, and the depth of cut on temperature distribution are considered. The model with temperature-dependent thermal conductivity and specific heat is presented. The Lagrange partial differential matrix from the heat transfer governing equation is obtained by using Lagrange series and mapping technique. The grinding wheel-workpiece contact area is assumed as a moving distributed square heat source. The Laplace transformation method and Durbin’s inverse technique are employed in the transient heat conduction analysis. The results of the developed model are compared with others’ finite element method solutions and analytical solutions where a good agreement is demonstrated. And the finite block method was proved a better convergence and accuracy than finite element method by comparing the ABAQUS results. In addition, the three-dimensional infinite element is introduced to perform the thermal analysis, and there is a great of advantages in the simulation of large boundary problems.The work was funded by China Scholarship Council, the Fundamental Research Funds for the Central Universities (N160306006), National Natural Science Foundation of China (51275084), and Science and technology project of Shenyang (18006001)

Colorectal Cancer Stem Cells Are Enriched in Xenogeneic Tumors Following Chemotherapy

Patients generally die of cancer after the failure of current therapies to eliminate residual disease. A subpopulation of tumor cells, termed cancer stem cells (CSC), appears uniquely able to fuel the growth of phenotypically and histologically diverse tumors. It has been proposed, therefore, that failure to effectively treat cancer may in part be due to preferential resistance of these CSC to chemotherapeutic agents. The subpopulation of human colorectal tumor cells with an ESA(+)CD44(+) phenotype are uniquely responsible for tumorigenesis and have the capacity to generate heterogeneous tumors in a xenograft setting (i.e. CoCSC). We hypothesized that if non-tumorigenic cells are more susceptible to chemotherapeutic agents, then residual tumors might be expected to contain a higher frequency of CoCSC.Xenogeneic tumors initiated with CoCSC were allowed to reach approximately 400 mm(3), at which point mice were randomized and chemotherapeutic regimens involving cyclophosphamide or Irinotecan were initiated. Data from individual tumor phenotypic analysis and serial transplants performed in limiting dilution show that residual tumors are enriched for cells with the CoCSC phenotype and have increased tumorigenic cell frequency. Moreover, the inherent ability of residual CoCSC to generate tumors appears preserved. Aldehyde dehydrogenase 1 gene expression and enzymatic activity are elevated in CoCSC and using an in vitro culture system that maintains CoCSC as demonstrated by serial transplants and lentiviral marking of single cell-derived clones, we further show that ALDH1 enzymatic activity is a major mediator of resistance to cyclophosphamide: a classical chemotherapeutic agent.CoCSC are enriched in colon tumors following chemotherapy and remain capable of rapidly regenerating tumors from which they originated. By focusing on the biology of CoCSC, major resistance mechanisms to specific chemotherapeutic agents can be attributed to specific genes, thereby suggesting avenues for improving cancer therapy

Endogenous c-Myc is essential for p53-induced apoptosis in response to DNA damage in vivo

Recent studies have suggested that C-MYC may be an excellent therapeutic cancer target and a number of new agents targeting C-MYC are in preclinical development. Given most therapeutic regimes would combine C-MYC inhibition with genotoxic damage, it is important to assess the importance of C-MYC function for DNA damage signalling in vivo. In this study, we have conditionally deleted the c-Myc gene in the adult murine intestine and investigated the apoptotic response of intestinal enterocytes to DNA damage. Remarkably, c-Myc deletion completely abrogated the immediate wave of apoptosis following both ionizing irradiation and cisplatin treatment, recapitulating the phenotype of p53 deficiency in the intestine. Consistent with this, c-Myc-deficient intestinal enterocytes did not upregulate p53. Mechanistically, this was linked to an upregulation of the E3 Ubiquitin ligase Mdm2, which targets p53 for degradation in c-Myc-deficient intestinal enterocytes. Further, low level overexpression of c-Myc, which does not impact on basal levels of apoptosis, elicited sustained apoptosis in response to DNA damage, suggesting c-Myc activity acts as a crucial cell survival rheostat following DNA damage. We also identify the importance of MYC during DNA damage-induced apoptosis in several other tissues, including the thymus and spleen, using systemic deletion of c-Myc throughout the adult mouse. Together, we have elucidated for the first time in vivo an essential role for endogenous c-Myc in signalling DNA damage-induced apoptosis through the control of the p53 tumour suppressor protein

Lawsonia intracellularis exploits β-catenin/Wnt and Notch signalling pathways during infection of intestinal crypt to alter cell homeostasis and promote cell proliferation

Lawsonia intracellularis is an obligate intracellular bacterial pathogen that causes proliferative enteropathy (PE) in pigs. L. intracellularis infection causes extensive intestinal crypt cell proliferation and inhibits secretory and absorptive cell differentiation. However, the affected host upstream cellular pathways leading to PE are still unknown. β-catenin/Wnt signalling is essential in maintaining intestinal stem cell (ISC) proliferation and self-renewal capacity, while Notch signalling governs differentiation of secretory and absorptive lineage specification. Therefore, in this report we used immunofluorescence (IF) and quantitative reverse transcriptase PCR (RTqPCR) to examine β-catenin/Wnt and Notch-1 signalling levels in uninfected and L. intracellularis infected pig ileums at 3, 7, 14, 21 and 28 days post challenge (dpc). We found that while the significant increase in Ki67+ nuclei in crypts at the peak of L. intracellularis infection suggested enhanced cell proliferation, the expression of c-MYC and ASCL2, promoters of cell growth and ISC proliferation respectively, was down-regulated. Peak infection also coincided with enhanced cytosolic and membrane-associated β-catenin staining and induction of AXIN2 and SOX9 transcripts, both encoding negative regulators of β-catenin/Wnt signalling and suggesting a potential alteration to β-catenin/Wnt signalling levels, with differential regulation of the expression of its target genes. We found that induction of HES1 and OLFM4 and the down-regulation of ATOH1 transcript levels was consistent with the increased Notch-1 signalling in crypts at the peak of infection. Interestingly, the significant down-regulation of ATOH1 transcript levels coincided with the depletion of MUC2 expression at 14 dpc, consistent with the role of ATOH1 in promoting goblet cell maturation. The lack of significant change to LGR5 transcript levels at the peak of infection suggested that the crypt hyperplasia was not due to the expansion of ISC population. Overall, simultaneous induction of Notch-1 signalling and the attenuation of β-catenin/Wnt pathway appear to be associated with the inhibition of goblet cell maturation and enhanced crypt cell proliferation at the peak of L. intracellularis infection. Moreover, the apparent differential regulation of apoptosis between crypt and lumen cells together with the strong induction of Notch-1 signalling and the enhanced SOX9 expression along crypts 14 dpc suggest an expansion of actively dividing transit amplifying and/or absorptive progenitor cells and provide a potential basis for understanding the development and maintenance of PE