Search

Background Progressive maturation of growth plate chondrocytes drives long bone growth during endochondral ossification. Signals from the Epidermal Growth Factor Receptor (EGFR), and from Bone Morphogenetic Protein-2 (BMP2), are required for normal chondrocyte maturation. Here, we investigated cross-talk between EGFR and BMP2 signals in developing and adult growth plates. Results Using in vivo mouse models of conditional cartilage-targeted EGFR- or BMP2- loss, we show that canonical BMP signal activation is increased in the hypertrophic chondrocytes of EGFR-deficient growth plates; whereas EGFR signal activation is increased in the reserve, prehypertrophic and hypertrophic chondrocytes of BMP2-deficient growth plates. EGFR-deficient chondrocytes displayed increased BMP signal activation in vitro, accompanied by increased expression of IHH, COL10A1 and RUNX2. Hypertrophic differentiation and BMP signal activation were suppressed in normal chondrocyte cultures treated with the EGFR ligand betacellulin, effects that were partially blocked by simultaneous treatment with BMP2 or a chemical EGFR antagonist. Conclusions Cross-talk between EGFR and BMP2 signals occurs during chondrocyte maturation. In the reserve and prehypertrophic zones, BMP2 signals unilaterally suppress EGFR activity; in the hypertrophic zone, EGFR and BMP2 signals repress each other. This cross-talk may play a role in regulating chondrocyte maturation in developing and adult growth plates. This article is protected by copyright. All rights reserved.

A michrochannel plate detector has been used to image tritium-labeled protein on one- and two-dimensional electrophoresis gels. The good spatial resolution (70 microns) and high sensitivity (6.0 dpm/mm2) of the imaging system allows detection of low levels (femto moles) of labelled proteins. We are currently using the detector for identification of new targets involved in organophosphate neurotoxicity.

We describe the development of a novel detection system used for the functional imaging of proteins separated on electrophoretic gels. A microchannel plate detector is used here for real-time imaging of low levels of tritiated protein separated by two-dimensional (2-D) electrophoresis. The system employs radioisotope-free, low noise microchannel plates originally developed for photon counting in X-ray astronomy. Using the detector configuration described here, proteins were resolved on mini gels by either one or two-dimensional electrophoresis, transferred onto polyvinylidene difluoride membranes and directly imaged. Tritiated diisopropylfluorophosphate (DFP) was used as a selective label for the serine hydrolase class of enzymes and their distribution in the central nervous system was examined. This survey revealed approximately 24 protein spots by 2-D electrophoresis. We also investigated the relative sensitivity of these proteins towards DFP and found the peptidase, acylpeptide hydrolase to be the most sensitive brain protein towards this reagent. Using a number of different tritiated standards, it was found that the system can image as little as 0.1 Bq/mm 2 of tritium corresponding to 320 attomol of DFP labelled protein/mm 2 . Moreover, the system has a wide dynamic range (> 10 6 ) allowing samples of high and low activity to be quantified on the same gel.