Axillary lymph node dissection on the run?

The standard approach of performing a completion axillary lymph node dissection (cALND) after a positive sentinel node for breast cancer patients is no longer generally accepted. This study applied the criterion of a 27% risk of having residual positive lymph nodes calculated by the MD Anderson nomogram to perform a cALND. This 27% cut-off is based on the number of positive non-sentinels in the Z0011 trial. A cohort of 166 cN0, sentinel positive breast cancer patients was used to validate the MD Anderson nomogram. ROC (Receiver Operating Characteristic) analysis shows an AUC (Area Under the Curve) of 0.76 and an optimal cut-off at 34% risk of positive non- SLNs (sensitivity 86%, specificity 57%). The 27% cut-off has a sensitivity of 88% and a specificity of 41% to detect positive non-sentinels. In a second cohort (N= 114) the 27% cut-off criterion was prospectively applied and appeared to be practice changing. Although we take minimal risk to leave disease behind (2/166 patients >3 positive nodes), 30.7 % in the first cohort and 54.4 % of the patients in the second cohort could be spared a cALND. The Z0011 criteria would have had more impact, omitting 90% of the cALND, but leaves more disease behind. The impact of leaving disease behind on survival remains unanswered but is awaited by long term follow up of large prospective cohort studies.

Smad-interacting protein 1 affects acute and tonic, but not chronic pain.

BACKGROUND: Smad-interacting protein 1 (also named Zeb2 and Zfhx1b) is a transcription factor that plays an important role in neuronal development and, when mutated, causes Mowat-Wilson syndrome (MWS). A corresponding mouse model carrying a heterozygous Zeb2 deletion was comprehensively analysed in the German Mouse Clinic. The most prominent phenotype was the reduced pain sensitivity. In this study, we investigated the role of Zeb2 in inflammatory and neuropathic pain. METHODS: For this, we tested mutant Zeb2 animals in different models of inflammatory pain like abdominal constriction, formalin and carrageenan test. Furthermore, we studied the pain reactivity of the mice after peripheral nerve ligation. To examine the nociceptive transmission of primary sensory dorsal root ganglia (DRG) neurons, we determined the neuronal activity in the spinal dorsal horn after the formalin test using staining of c-Fos. Next, we characterized the neuronal cell population in the DRGs and in the sciatic nerve to study the effect of the Zeb2 mutation on peripheral nerve morphology. RESULTS: The present data show that Zeb2 is involved in the development of primary sensory DRG neurons, especially of C- and Aδ fibres. These alterations contribute to a hypoalgesic phenotype in inflammatory but not in neuropathic pain in these Zeb2(+/-) mice. CONCLUSION: Our data suggest that the under-reaction to pain observed in MWS patients results from a reduced responsivity to nociceptive stimulation rather than an inability to communicate discomfort.

Contrast-enhanced nanofocus computed tomography images the cartilage subtissue architecture in three dimensions.

We describe a non-destructive imaging method, named contrast-enhanced nanofocus X-ray computed tomography (CE-nanoCT), that permits simultaneously imaging and quantifying in 3D the (sub)tissue architecture and (biochemical) composition of cartilage and bone in small animal models at a novel contrast and spatial resolution. To demonstrate the potential of this novel methodology, a newborn mouse was scanned using CE-nanoCT. This allowed simultaneously visualising the bone and cartilage structure much like the traditional alcian blue-alizarin red skeletal stain. Additionally, it enabled a 3D visualisation at such a high spatial image resolution that internal, micro-scale structures could be digitally dissected and evaluated for size, structure and composition. Ex vivo treatment with papain, that is known to specifically remove the non-calcified cartilage layer but keep the calcified cartilage intact, proved CE-nanoCT to be applicable to visualise the subdivisions within the hyaline cartilage of the articular joint of mice. The quantitative power of CE-nanoCT in vivo was evaluated using a mouse model for osteoarthritis (OA), where OA-like cartilage lesions are induced by meniscus destabilisation surgery. The thickness of both the non-calcified and calcified cartilage layer in the knee joint of such mice was visualised and quantified in 3D and compared to unaffected mice. Finally, to show that different forms of cartilage and tissue combinations can be distinguished using CE-nanoCT, different cartilaginous body parts of the mouse were imaged. In conclusion, CE-nanoCT can provide novel insights in preclinical research by quantifying in a non-destructive 3D manner pathological differences, in particular in developing mice, newborns or adults.

Transforming growth factor beta signalling in vitro and in vivo: activin ligand-receptor interaction, Smad5 in vasculogenesis, and repression of target genes by the deltaEF1/ZEB-related SIP1 in the vertebrate embryo.

The identification and characterization of components of the transforming growth factor beta (TGFbeta) signalling pathway are proceeding at a very fast pace. To illustrate a number of our activities in this field, we first summarize our work aiming at the selection from a large collection of single residue substitution mutants of two activin A polypeptides in which D27 and K102, respectively, have been modified. This work has highlighted the importance of K102 and its positive charge for binding to activin type II receptors. Activin K102E, which did not bind to high-affinity receptor complexes, may be a valuable beta chain, when incorporated in recombinant inhibin to unambiguously detect novel inhibin binding sites at the cell surface. We then illustrate how Smad5 knockout mice and an overexpression approach with a truncated TGFbeta type II receptor in the mouse embryo can contribute to the identification of a novel TGFbeta-->TbetaRII/ALK1-->Smad5 pathway in endothelial cells in the embryo proper and the yolk sac vasculature. We conclude with a summary of our results with a Smad-interacting transcriptional repressor but focus on its biological significance in the vertebrate embryo.

Mice with a homozygous gene trap vector insertion in mgcRacGAP die during pre-implantation development.

In a phenotypic screen in mice using a gene trap approach in embryonic stem cells, we have identified a recessive loss-of-function mutation in the mgcRacGAP gene. Maternal protein is present in the oocyte, and mgcRacGAP gene transcription starts at the four-cell stage and persists throughout mouse pre-implantation development. Total mgcRacGAP deficiency results in pre-implantation lethality. Such E3.5 embryos display a dramatic reduction in cell number, but undergo compaction and form a blastocoel. At E3.0-3.5, binucleated blastomeres in which the nuclei are partially interconnected are frequently observed, suggesting that mgcRacGAP is required for normal mitosis and cytokinesis in the pre-implantation embryo. All homozygous mutant blastocysts fail to grow out on fibronectin-coated substrates, but a fraction of them can still induce decidual swelling in vivo. The mgcRacGAP mRNA expression pattern in post-implantation embryos and adult mouse brain suggests a role in neuronal cells. Our results indicate that mgcRacGAP is essential for the earliest stages of mouse embryogenesis, and add evidence that CYK-4-like proteins also play a role in microtubule-dependent steps in the cytokinesis of vertebrate cells. In addition, the severe phenotype of null embryos indicates that mgcRacGAP is functionally non-redundant and cannot be substituted by other GAPs during early cleavage of the mammalian embryo.