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Bone morphogenetic protein (BMP)-evoked reorientation and chemotaxis of cells occurs with rapid onset and involves events local to the cell membrane. The signaling pathways underlying these rapid processes likely diverge from those mediating classical transcriptional responses to BMPs but it remains unclear how BMP receptors are utilized to generate distinct intracellular mechanisms. We show that BMP7-evoked chemotaxis of monocytic cells depends on the activity of canonical type II BMP receptors. Although the three canonical type II BMP receptors are expressed in monocytic cells, inhibition of receptor subunit expression by RNAi reveals that ActRIIA and BMPRII, but not ActRIIB, are each essential for BMP7-evoked chemotaxis but not required individually for BMP-mediated induction. Furthermore, the chemotactic response to BMP7 does not involve canonical Smad4-dependent signaling but acts through PI3K-dependent signaling, illustrating selective activation of distinct intracellular events through differential engagement of receptors. We suggest a model of a BMP receptor complex in which the coordinated activity of ActRIIA and BMPRII receptor subunits selectively mediates the chemotactic response to BMP7.

BackgroundBone morphogenetic protein (BMP)7 evokes both inductive and axon orienting responses in dorsal interneurons (dI neurons) in the developing spinal cord. These events occur sequentially during the development of spinal neurons but in these and other cell types such inductive and acute chemotactic responses occur concurrently, highlighting the requirement for divergent intracellular signaling. Both type I and type II BMP receptor subtypes have been implicated selectively in orienting responses but it remains unclear how, in a given cell, divergence occurs. We have examined the mechanisms by which disparate BMP7 activities are generated in dorsal spinal neurons.ResultsWe show that widely different threshold concentrations of BMP7 are required to elicit the divergent inductive and axon orienting responses. Type I BMP receptor kinase activity is required for activation of pSmad signaling and induction of dI character by BMP7, a high threshold response. In contrast, neither type I BMP receptor kinase activity nor Smad1/5/8 phosphorylation is involved in the low threshold orienting responses of dI axons to BMP7. Instead, BMP7-evoked axonal repulsion and growth cone collapse are dependent on phosphoinositide-3-kinase (PI3K) activation, plausibly through type II receptor signaling. BMP7 stimulates PI3K-dependent signaling in dI neurons. BMP6, which evokes neural induction but does not have orienting activity, activates Smad signaling but does not stimulate PI3K.ConclusionsDivergent signaling through pSmad-dependent and PI3K-dependent (Smad-independent) mechanisms mediates the inductive and orienting responses of dI neurons to BMP7. A model is proposed whereby selective engagement of BMP receptor subunits underlies choice of signaling pathway.

Background Oxygen therapy, using supraphysiological concentrations of oxygen (hyperoxia), is routinely administered to patients who require respiratory support including mechanical ventilation (MV). However, prolonged exposure to hyperoxia results in acute lung injury (ALI) and accumulation of high mobility group box 1 (HMGB1) in the airways. We previously showed that airway HMGB1 mediates hyperoxia-induced lung injury in a mouse model of ALI. Cholinergic signaling through the α7 nicotinic acetylcholine receptor (α7nAChR) attenuates several inflammatory conditions. The aim of this study was to determine whether 3–(2,4 dimethoxy-benzylidene)-anabaseine dihydrochloride, GTS-21, an α7nAChR partial agonist, inhibits hyperoxia-induced HMGB1 accumulation in the airways and circulation, and consequently attenuates inflammatory lung injury. Methods Mice were exposed to hyperoxia (≥99% O2) for 3 days and treated concurrently with GTS-21 (0.04, 0.4 and 4 mg/kg, i.p.) or the control vehicle, saline. Results The systemic administration of GTS-21 (4 mg/kg) significantly decreased levels of HMGB1 in the airways and the serum. Moreover, GTS-21 (4 mg/kg) significantly reduced hyperoxia-induced acute inflammatory lung injury, as indicated by the decreased total protein content in the airways, reduced infiltration of inflammatory monocytes/macrophages and neutrophils into the lung tissue and airways, and improved lung injury histopathology. Conclusions Our results indicate that GTS-21 can attenuate hyperoxia-induced ALI by inhibiting extracellular HMGB1-mediated inflammatory responses. This suggests that the α7nAChR represents a potential pharmacological target for the treatment regimen of oxidative inflammatory lung injury in patients receiving oxygen therapy.

Hemodynamic variability in patients with atrial fibrillation may originate from a direct influence of the variations in RR intervals on myocardial contractility. With the aid of a computer the serial autocorrelation function and the histogram of the RR intervals of patients with atrial fibrillation receiving no medication were produced. The RR intervals were randomly distributed and the histograms rather skew. Next, random rhythms with histograms matching those of the patients were produced with a radioactive source and a GeigerMiiller counter. These rhythms were used to stimulate isolated perfused rat hearts to study the relationship between the RR interval and a number of contractile parameters. The ECG and the isotonic contractions were digitized and processed by the computer. Serial crosscorrelation coefficients were computed between the RR interval on the one hand, and contraction height, contraction area, and maximum of the first derivative of the contraction on the other hand. The zero order crosscorrelation coefficient between the RR interval and contraction height was 0.6, between the RR interval and contraction area 0.8, and between the RR interval and maximum of first derivative 0.5. Although small, the first and second to approximately the tenth order coefficients were definitely negative. It is concluded that during a random rhythm the contractile parameters of an isolated heart are strongly related to the preceding RR interval. It is conceivable that this relation contributes to the variability of hemodynamic parameters during atrial fibrillation in man. • Atrial fibrillation is a common rhythm disturbance of the heart in man. During this type of arrhythmia the ventricles beat very irregularly and, in general, each contraction differs from the preceding one and from the next. ADDITIONAL KEY WORDS

BackgroundIn dorsal spinal neurons and monocytes, bone morphogenetic protein (BMP)7 activates distinct transduction pathways, one leading to inductive specification and the other to axon orientation and chemotaxis. BMP7-evoked induction, also stimulated by the closely related BMP6, acts through a Smad cascade, leading to nuclear signaling, and is not BMPR subunit selective. Orientation is evoked by BMP7, but not by BMP6, through PI3K-dependent cytoskeletal activation mediated by the type II BMPRs, ActRIIA and BMPRII and is independent of the Smad cascade. The responses can be stimulated concurrently and suggest that BMP7, but not BMP6, can selectively activate BMPR subunits that engage the divergent paths. Although structural and biochemical analyses of selected BMP/BMPR interfaces have identified key regions of interaction, how these translate into function by related BMPs is poorly understood. To determine the mechanisms underlying the distinct activities of BMP7 and the disparate properties of BMP7 and BMP6 in spinal cord development, we have performed a family-wide structure/function analysis of BMPs and used the information to predict and test sites within BMPs that may control agonist properties, in particular the ability of a BMP to orient axons, through interactions with BMPRs.ResultsWe demonstrate that whereas all BMPs can induce dorsal neurons, there is selectivity in the ability also to orient axons or evoke growth cone collapse. The degree to which a BMP orients is not predictable by overall protein similarity with other BMPs but comparison of sequences of potent and weakly orienting BMPs with that of the non-orienting BMP6 revealed three candidate positions within the BMPs at which the amino acid residues may confer or obstruct orienting ability. Residue swapping analysis has identified one residue, Gln48 in BMP6, that blocks axon orienting ability. Replacing Gln48 with any of the amino acids present at the equivalent residue position in the orienting subset of BMPs confers orienting activity on BMP6. Conversely, swapping Gln48 into BMP7 reduces orienting ability. The inductive capacity of the BMPs was unchanged by these residue swaps.ConclusionsThe results suggest that the presence of the Gln48 residue in BMP6 is structurally inhibitory for BMP/BMPR interactions that result in the activation of intracellular signaling, leading to axon orientation. Moreover, since residue 48 in BMP7 and the corresponding residue in BMP2 are important for type II BMPR binding, our results provide a basis for a mechanistic understanding of the diverse activities of BMPs in spinal cord development.

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The tyrosine phosphatase SHP-2 has been implicated in a variety of signaling pathways, including those mediated by neurotrophins in neurons. To examine the role of SHP-2 in the development of sympathetic neurons, we inhibited the function of SHP-2 in transgenic mice by overexpressing a catalytically inactive SHP-2 mutant under the control of the human dopamine beta-hydroxylase promoter. Expression of mutant SHP-2 did not influence the survival, axon initiation, or pathfinding abilities of the sympathetic neurons. However, mutant SHP-2 expression resulted in an overproduction of sympathetic fibers in sympathetic target organs. This was due to interference with SHP-2 function, as overexpression of wild type SHP-2 had no such effect. In vitro, NGF-dependent neurite growth was inhibited in neurons expressing mutant SHP-2 but not in those expressing wild type SHP-2. Mutant (but not wt) SHP-2 expression also inhibited NGF-stimulated ERK activation. The NGF-dependent survival pathway was less affected than the neurite growth pathway. Our results suggest that NGF-regulated axon growth signals, and to a lesser degree survival signals, are mediated through a SHP-2-dependent pathway in sympathetic neurons. The increased sympathetic innervation in target tissues of neurons expressing mutant SHP-2 may result from interference with normal "stop" signals dependent on signaling by gradients of NGF.