Effects of substrate patterning on cellular spheroid growth and dynamics measured by gradient light interference microscopy (GLIM)

The development of three‐dimensional (3D) cellular architectures during development and pathological processes involves intricate migratory patterns that are modulated by genetics and the surrounding microenvironment. The substrate composition of cell cultures has been demonstrated to influence growth, proliferation and migration in 2D. Here, we study the growth and dynamics of mouse embryonic fibroblast cultures patterned in a tissue sheet which then exhibits 3D growth. Using gradient light interference microscopy (GLIM), a label‐free quantitative phase imaging approach, we explored the influence of geometry on cell growth patterns and rotational dynamics. We apply, for the first time to our knowledge, dispersion‐relation phase spectroscopy (DPS) in polar coordinates to generate the radial and rotational cell mass‐transport. Our data show that cells cultured on engineered substrates undergo rotational transport in a radially independent manner and exhibit faster vertical growth than the control, unpatterned cells. The use of GLIM and polar DPS provides a novel quantitative approach to studying the effects of spatially patterned substrates on cell motility and growth.     

Mafb and c-Maf Have Prenatal Compensatory and Postnatal Antagonistic Roles in Cortical Interneuron Fate and Function

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Escherichia coli AspP activity is enhanced by macromolecular crowding and by both glucose-1,6-bisphosphate and nucleotide-sugars

Escherichia coli ADP-sugar pyrophosphatase (AspP) is a "Nudix" hydrolase that catalyzes the hydrolytic breakdown of ADP-glucose linked to glycogen biosynthesis. Moderate increases of AspP activity in the cell are accompanied by significant reductions of the glycogen content. In vitro analyses showed that AspP activity is strongly enhanced by macromolecular crowding and by both glucose-1,6-bisphosphate and nucleotide-sugars, providing a first set of indicative evidences that AspP is a highly regulated enzyme. To our knowledge, AspP is the sole bacterial enzyme described to date which is activated by both G1,6P(2) and nucleotide-sugars.     

A role of lipophilic peptidoglycan-related molecules in induction of Nod1-mediated immune responses

Nod1 is an intracellular protein that is involved in recognition of bacterial molecules and whose genetic variation has been linked to several inflammatory diseases. Previous studies suggested that the recognition core of Nod1 stimulatory molecules is gamma-D-glutamyl-meso-diaminopimelic acid (iE-DAP), but the identity of the major Nod1 stimulatory molecule produced by bacteria remains unknown. Here we show that bacteria produce lipophilic molecules capable of stimulating Nod1. Analysis of synthetic compounds revealed stereoselectivity of the DAP residue and that conjugation of lipophilic acyl residues specifically enhances the Nod1 stimulatory activity of the core iE-DAP. Furthermore, we demonstrate that lipophilic molecules induce and/or enhance the secretion of innate immune mediators from primary mouse mesothelial cells and human monocytic MonoMac6 cells, and this effect is mediated through Nod1. These results provide insight into the mechanism of immune recognition via Nod1, which might be useful in the design and testing of novel immunoregulators.     

The NOD2-RICK complex signals from the plasma membrane

NOD2 plays an important role in the innate immunity of the intestinal tract. By sensing the muramyl dipeptide (MDP), a bacterial wall component, NOD2 triggers the NF-kappaB signaling pathway and promotes the release of proinflammatory cytokines such as interleukin-8. Mutations in Nod2 (1007FS, R702W, G908R) impinge on NOD2 functions and are associated with the pathogenesis of Crohn disease, a chronic inflammatory bowel disease. Although NOD2 is usually described as a cytosolic receptor for MDP, the protein is also localized at the plasma membrane, and the 1007FS mutation delocalizes NOD2 to the cytoplasm (Barnich, N., Aguirre, J. E., Reinecker, H. C., Xavier, R., and Podolsky, D. K. (2005) J. Cell Biol. 170, 21-26; McDonald, C., Chen, F. F., Ollendorff, V., Ogura, Y., Marchetto, S., Lecine, P., Borg, J. P., and Nunez, G. (2005) J. Biol. Chem. 280, 40301-40309). In this study, we demonstrate that membrane-bound versions of NOD2 and Crohn disease-associated mutants R702W and G908R are capable of responding to MDP and activating the NF-kappaB pathway from this location. In contrast, the 1007FS mutant remains unable to respond to MDP from the plasma membrane. We also show that NOD2 promotes the membrane recruitment of RICK, a serine-threonine kinase involved in NF-kappaB activation downstream of NOD2. Furthermore, the artificial attachment of RICK at the plasma membrane provokes a constitutive and strong activation of the NF-kappaB pathway and secretion of interleukin-8 showing that optimal RICK activity depends upon its subcellular localization. Finally, we show that endogenous RICK localizes at the plasma membrane in the THP1 cell line. Thus, our data suggest that NOD2 is responsible for the membrane recruitment of RICK to induce a regulated NF-kappaB signaling and production of proinflammatory cytokines.     

Which Obesity Index Best Explains Prevalence Differences in Type 2 Diabetes Mellitus?

Objective: Obesity drives the diabetes epidemic. However, it is not known which obesity index best explains variations in type 2 diabetes mellitus prevalence across populations. Research Methods and Procedures: We analyzed three cross‐sectional studies from San Antonio, TX, (Mexican‐Americans and non‐Hispanic whites, n = 2839), Mexico City (n = 2233), and Spain (n = 2161) (age range, 35 to 64 years). We used the area under the receiver operating characteristic curve (AUC) to assess performance for identifying diabetic subjects and logistic regression analysis to examine differences in diabetes prevalence. Results: AUCs for waist circumference and BMI were similar in white subjects, but the AUC for waist circumference was greater in Mexican‐origin subjects (Mexican men, 0.594 vs. 0.549, p = 0.008; and women, 0.605 vs. 0.557, p = 0.002; Mexican‐American men, 0.648 vs. 0.600, p < 0.001; and women, 0.744 vs. 0.693, p < 0.001). The AUC for waist‐to‐height ratio tended to be greater than that for waist circumference, but statistical significance was demonstrated only in Mexican women (0.628 vs. 0.613, p = 0.044), Mexican‐American women (0.774 vs. 0.758, p < 0.001), and Spanish women (0.734 vs. 0.715, p = 0.039). No obesity index was consistently superior to the others for explaining differences in diabetes prevalence among populations. Conclusions: In white and Mexican‐origin men, waist circumference may be the preferred marker for identifying diabetic subjects on account of its simplicity; in women, waist‐to‐height ratio may be better. Differences in diabetes prevalence among these populations cannot be attributed to a single measure of obesity.     

Efficient identification of critical residues based only on protein structure by network analysis

Despite the increasing number of published protein structures, and the fact that each protein's function relies on its three-dimensional structure, there is limited access to automatic programs used for the identification of critical residues from the protein structure, compared with those based on protein sequence. Here we present a new algorithm based on network analysis applied exclusively on protein structures to identify critical residues. Our results show that this method identifies critical residues for protein function with high reliability and improves automatic sequence-based approaches and previous network-based approaches. The reliability of the method depends on the conformational diversity screened for the protein of interest. We have designed a web site to give access to this software at http://bis.ifc.unam.mx/jamming/. In summary, a new method is presented that relates critical residues for protein function with the most traversed residues in networks derived from protein structures. A unique feature of the method is the inclusion of the conformational diversity of proteins in the prediction, thus reproducing a basic feature of the structure/function relationship of proteins.