Autotaxin, a secreted lysophospholipase D, is essential for blood vessel formation during development

Autotaxin (ATX), or nucleotide pyrophosphatase-phosphodiesterase 2, is a secreted lysophospholipase D that promotes cell migration, metastasis, and angiogenesis. ATX generates lysophosphatidic acid (LPA), a lipid mitogen and motility factor that acts on several G protein-coupled receptors. Here we report that ATX-deficient mice die at embryonic day 9.5 (E9.5) with profound vascular defects in yolk sac and embryo resembling the Galpha13 knockout phenotype. Furthermore, at E8.5, ATX-deficient embryos showed allantois malformation, neural tube defects, and asymmetric headfolds. The onset of these abnormalities coincided with increased expression of ATX and LPA receptors in normal embryos. ATX heterozygous mice appear healthy but show half-normal ATX activity and plasma LPA levels. Our results reveal a critical role for ATX in vascular development, indicate that ATX is the major LPA-producing enzyme in vivo, and suggest that the vascular defects in ATX-deficient embryos may be explained by loss of LPA signaling through Galpha13.     

The CpG island methylator phenotype in colorectal cancer: progress and problems

In recent years, attention has focused on the biology and potential clinical importance of the CpG island methylator phenotype (CIMP) in colorectal cancer (CRC). While it is generally well accepted that etiologically and clinically distinct subgroups exist in this disease, a precise definition of CIMP remains to be established. Here, we summarize existing literature that documents the prevalence of CIMP in CRC, with particular attention to the various methods and definitions used to classify a tumor as CIMP positive. Through a systematic review on both case-series and population based studies, we examined only original research articles reporting on sporadic CRC and/or adenomas in unselected cases. Forty-eight papers published between January 1999 and August 2011 met the inclusion criteria. We describe the use of multiple gene panels, marker threshold values, and laboratory techniques which results in a wide range in the prevalence of CIMP. Because there is no universal standard or consensus on quantifying the phenotype, establishing its true prevalence is a challenge. This bottleneck is becoming increasingly evident as molecular pathological epidemiology continues to offer possibilities for clear answers regarding environmental risk factors and disease trends. For the first time, large, unselected series of cases are available for analysis, but comparing populations and pooling data will remain a challenge unless a universal definition of CIMP and a consensus on analysis can be reached, and the primary cause of CIMP identified.     

Elevated levels of fibrinogen-derived endogenous citrullinated peptides in synovial fluid of rheumatoid arthritis patients

Introduction

Rheumatoid arthritis (RA) is an autoimmune disease characterized by inflammation of the joints and the presence of autoantibodies directed against proteins containing the non-standard arginine-derived amino acid citrulline. The protein fibrinogen, which has an essential role in blood clotting, is one of the most prominent citrullinated autoantigens in RA, particularly because it can be found in the inflamed tissue of affected joints. Here, we set out to analyze the presence of citrullinated endogenous peptides in the synovial fluid of RA and arthritic control patients.

Methods

Endogenous peptides were isolated from the synovial fluid of RA patients and controls by filtration and solid phase extraction. The peptides were identified and quantified using high-resolution liquid chromatography-mass spectrometry.

Results

Our data reveal that the synovial fluid of RA patients contains soluble endogenous peptides, derived from fibrinogen, containing significant amounts of citrulline residues and, in some cases, also phosphorylated serine. Several citrullinated peptides are found to be more abundantly present in the synovial fluid of RA patients compared to patients suffering from other inflammatory diseases affecting the joints.

Conclusions

The increased presence of citrullinated peptides in RA patients points toward a possible specific role of these peptides in the immune response at the basis of the recognition of citrullinated peptides and proteins by RA patient autoantibodies.     

A physiology-based inverse dynamic analysis of human gait using sequential convex programming: a comparative study

This paper presents an enhanced version of the previously proposed physiological inverse approach (PIA) to calculate musculotendon (MT) forces and evaluates the proposed methodology in a comparative study. PIA combines an inverse dynamic analysis with an optimisation approach that imposes muscle physiology and optimises performance over the entire motion. To solve the resulting large-scale, nonlinear optimisation problem, we neglected muscle fibre contraction speed and an approximate quadratic optimisation problem (PIA-QP) was formulated. Conversely, the enhanced version of PIA proposed in this paper takes into account muscle fibre contraction speed. The optimisation problem is solved using a sequential convex programing procedure (PIA-SCP). The comparative study includes PIA-SCP, PIA-QP and two commonly used approaches from the literature: static optimisation (SO) and computed muscle control (CMC). SO and CMC make simplifying assumptions to limit the computational time. Both methods minimise an instantaneous performance criterion. Furthermore, SO does not impose muscle physiology. All methods are applied to a gait cycle of six control subjects. The relative root mean square error averaged over all subjects, ε(RMS), between the joint torques simulated from the optimised activations and the joint torques obtained from the inverse dynamic analysis was about twice as large for SO (ε(RMS) = 86) as compared with CMC (ε(RMS) = 39) and PIA-SCP (ε(RMS) = 50). ε(RMS) was at least twice as large for PIA-QP (ε(RMS) = 197) than for all other methods. As compared with CMC, muscle activation patterns predicted by PIA-SCP better agree with experimental electromyography (EMG). This study shows that imposing muscle physiology as well as globally optimising performance is important to accurately calculate MT forces underlying gait.     

Large-scale mutagenesis in p19(ARF)- and p53-deficient mice identifies cancer genes and their collaborative networks

p53 and p19(ARF) are tumor suppressors frequently mutated in human tumors. In a high-throughput screen in mice for mutations collaborating with either p53 or p19(ARF) deficiency, we identified 10,806 retroviral insertion sites, implicating over 300 loci in tumorigenesis. This dataset reveals 20 genes that are specifically mutated in either p19(ARF)-deficient, p53-deficient or wild-type mice (including Flt3, mmu-mir-106a-363, Smg6, and Ccnd3), as well as networks of significant collaborative and mutually exclusive interactions between cancer genes. Furthermore, we found candidate tumor suppressor genes, as well as distinct clusters of insertions within genes like Flt3 and Notch1 that induce mutants with different spectra of genetic interactions. Cross species comparative analysis with aCGH data of human cancer cell lines revealed known and candidate oncogenes (Mmp13, Slamf6, and Rreb1) and tumor suppressors (Wwox and Arfrp2). This dataset should prove to be a rich resource for the study of genetic interactions that underlie tumorigenesis.     

NCAM-induced focal adhesion assembly: a functional switch upon loss of E-cadherin

Loss of expression of the cell-cell adhesion molecule E-cadherin is a hallmark of epithelial-mesenchymal transition (EMT) in development and in the progression from epithelial tumours to invasive and metastatic cancers. Here, we demonstrate that the loss of E-cadherin function upregulates expression of the neuronal cell adhesion molecule (NCAM). Subsequently, a subset of NCAM translocates from fibroblast growth factor receptor (FGFR) complexes outside lipid rafts into lipid rafts where it stimulates the non-receptor tyrosine kinase p59(Fyn) leading to the phosphorylation and activation of focal adhesion kinase and the assembly of integrin-mediated focal adhesions. Ablation of NCAM expression during EMT inhibits focal adhesion assembly, cell spreading and EMT. Conversely, forced expression of NCAM induces epithelial cell delamination and migration, and high NCAM expression correlates with tumour invasion. These results establish a mechanistic link between the loss of E-cadherin expression, NCAM function, focal adhesion assembly and cell migration and invasion.     

Subject-specific hip geometry affects predicted hip joint contact forces during gait

Hip loading affects bone remodeling and implant fixation. In this study, we have analyzed the effect of subject-specific modeling of hip geometry on muscle activation patterns and hip contact forces during gait, using musculoskeletal modeling, inverse dynamic analysis and static optimization. We first used sensitivity analysis to analyze the effect of isolated changes in femoral neck-length (NL) and neck-shaft angle (NSA) on calculated muscle activations and hip contact force during the stance phase of gait. A deformable generic musculoskeletal model was adjusted incrementally to adopt a physiological range of NL and NSA. In a second similar analysis, we adjusted hip geometry to the measurements from digitized radiographs of 20 subjects with primary hip osteoarthrosis. Finally, we studied the effect of hip abductor weakness on muscle activation patterns and hip contact force. This analysis showed that differences in NL (41-74 mm) and NSA (113-140 degrees ) affect the muscle activation of the hip abductors during stance phase and hence hip contact force by up to three times body weight. In conclusion, the results from both the sensitivity and subject-specific analysis showed that at the moment of peak contact force, altered NSA has only a minor effect on the loading configuration of the hip. Increased NL, however, results in an increase of the three hip contact-force components and a reduced vertical loading. The results of these analyses are essential to understand modified hip joint loading, and for planning hip surgery for patients with osteoarthrosis.     

In vivo analysis of Frat1 deficiency suggests compensatory activity of Frat3.

The Frat1 gene was first identified as a proto-oncogene involved in progression of mouse T cell lymphomas. More recently, FRAT/GBP (GSK-3beta Binding Protein) family members have been recognized as critical components of the Wnt signal transduction pathway. In an attempt to gain more insight into the function of Frat1, we have generated Frat1-deficient mice in which most of the coding domain was replaced by a promoterless beta-galactosidase reporter gene. While the pattern of LacZ expression in Frat1(lacZ)/+ mice indicated Frat1 to be expressed in various neural and epithelial tissues, homozygous Frat1(lacZ) mice were apparently normal, healthy and fertile. Tissues of homozygous Frat1(lacZ) mice showed expression of a second mouse Frat gene, designated Frat3. The Frat1 and Frat3 proteins are structurally and functionally very similar, since both Frat1 and Frat3 are capable of inducing a secondary axis in Xenopus embryos. The overlapping expression patterns of Frat1 and Frat3 during murine embryogenesis suggest that the apparent dispensability of Frat1 for proper development may be due to the presence of a second mouse gene encoding a functional Frat protein.