Transforming growth factor-alpha acting at the epidermal growth factor receptor reduces infarct volume after permanent middle cerebral artery occlusion in rats.
Transforming growth factor-alpha (TGF-alpha) is a ligand for the epidermal growth factor (EGF) receptor (EGFR), and is more abundant than EGF in the brain. The authors studied whether administration of exogenous TGF-alpha into the brain can protect neurons against ischemia in a model of permanent middle cerebral artery (MCA) occlusion in the rat, and whether any effect of TGF-alpha was mediated by EGFR by administering 4,5-dianilinophthalimide (DAPH), a protein-tyrosine kinase inhibitor with high selectivity for EGFR. Rats received either TGF-alpha (10 or 25 ng), DAPH (100 ng), DAPH plus TGF-alpha (25 ng), or vehicle in the ipsilateral first ventricle. Drugs were administered twice: 30 minutes before and 30 minutes after MCA occlusion, and infarct volume was evaluated 24 hours later. Transforming growth factor-alpha at the dose of 25 ng caused a statistically significant reduction of infarct volume (60%) in relation to ischemic rats administered vehicle. This reduction was no longer seen when TGF-alpha was administered in combination with DAPH. The present results show that TGF-alpha can protect neurons from ischemic damage, and that this effect is mediated by EGFR. It is suggested that activation of EGFR-mediated intracellular signalling pathways contributes to the survival of neural cells susceptible to ischemic injury. (+info)
Phthalascidin, a synthetic antitumor agent with potency and mode of action comparable to ecteinascidin 743.
A series of totally synthetic molecules that are structurally related to the marine natural product ecteinascidin 743 (Et 743) has been prepared and evaluated as antitumor agents. The most active of these, phthalascidin, is very similar to Et 743 with regard to in vitro potency and mode of action across a variety of cell types. The antiproliferative activity of phthalascidin (IC50 = 0.1-1 nM) is greater than that of the agents Taxol, camptothecin, adriamycin, mitomycin C, cisplatin, bleomycin, and etoposide by 1-3 orders of magnitude, and the mechanism of action is clearly different from these currently used drugs. Phthalascidin and Et 743 induce DNA-protein cross-linking and, although they seem to interact with topoisomerase (topo) I (but not topo II), topo I may not be the primary protein target of these agents. Phthalascidin and Et 743 show undiminished potency in camptothecin- and etoposide-resistant cells. Phthalascidin is more readily synthesized and more stable than Et 743, which is currently undergoing clinical trials. The relationship of chemical structure and antitumor activity for this class of molecules has been clarified by this study. (+info)
Responses of plant vascular systems to auxin transport inhibition.
To assess the role of auxin flows in plant vascular patterning, the development of vascular systems under conditions of inhibited auxin transport was analyzed. In Arabidopsis, nearly identical responses evoked by three auxin transport inhibitor substances revealed an enormous plasticity of the vascular pattern and suggest an involvement of auxin flows in determining the sites of vascular differentiation and in promoting vascular tissue continuity. Organs formed under conditions of reduced auxin transport contained increased numbers of vascular strands and cells within those strands were improperly aligned. In leaves, vascular tissues became progressively confined towards the leaf margin as the concentration of auxin transport inhibitor was increased, suggesting that the leaf vascular system depends on inductive signals from the margin of the leaf. Staged application of auxin transport inhibitor demonstrated that primary, secondary and tertiary veins became unresponsive to further modulations of auxin transport at successive stages of early leaf development. Correlation of these stages to anatomical features in early leaf primordia indicated that the pattern of primary and secondary strands becomes fixed at the onset of lamina expansion. Similar alterations in the leaf vascular responses of alyssum, snapdragon and tobacco plants suggest common functions of auxin flows in vascular patterning in dicots, while two types of vascular pattern alterations in Arabidopsis auxin transport mutants suggest that at least two distinct primary defects can result in impaired auxin flow. We discuss these observations with regard to the relative contributions of auxin transport, auxin sensitivity and the cellular organisation of the developing organ on the vascular pattern. (+info)
Using a radioalloster to test predictions of the cooperativity model for gallamine binding to the allosteric site of muscarinic acetylcholine M(2) receptors.
The muscarinic M(2) receptor contains an orthosteric and an allosteric site. Binding of an allosteric agent may induce a shift alpha of the equilibrium dissociation constant K(D) of a radioligand for the orthosteric site. According to the cooperativity model, the K(A) of alloster binding is expected to be shifted to an identical extent depending on whether the orthosteric site is occupied by the orthoster or not. Here, the novel radioalloster [(3)H]dimethyl-W84 (N,N'-bis[3-(1,3-dihydro-1, 3-dioxo-4-methyl-2H-isoindol-2-yl)propyl]-N,N,N',N'-tetramethyl-1, 6-hexanediaminium diiodide) was applied to directly measure the K(A) shift induced for the prototype allosteric modulator gallamine by binding of N-methylscopolamine (NMS) to the orthosteric site of porcine heart M(2) receptors (4 mM Na(2)HPO(4), 1 mM KH(2)PO(4), pH 7.4; 23 degrees C; data are means +/- S.E.). First, in the common way, the concentration-dependent inhibition by gallamine of [(3)H]NMS equilibrium binding was measured and analyzed using the cooperativity model, which yielded for the affinity of gallamine binding at free receptors a pK(A)= 8.35 +/- 0.09 and a cooperativity factor alpha = 46 (n = 5). The dissociation constant for gallamine binding at NMS-occupied receptors was predicted as p(alpha. K(A)) = 6.69. Labeling of the allosteric site by [(3)H]dimethyl-W84 allowed the measure of competitive displacement curves for gallamine. The K(i) for gallamine at free receptors amounted to pK(i,-NMS) = 8.27 +/- 0.39 (n = 5), which is in line with the prediction of the cooperativtiy model. In the presence of 1 microM NMS, to occupy the orthosteric site, gallamine displaced [(3)H]dimethyl-W84 with pK(i, +NMS) = 6.60 +/- 0.19 (n = 3). Thus, the NMS-induced pK(i) shift amounted to 47, which matches the predicted value of alpha = 46. These results validate the cooperativity model. (+info)
Auxin is required for leaf vein pattern in Arabidopsis.
To investigate possible roles of polar auxin transport in vein patterning, cotyledon and leaf vein patterns were compared for plants grown in medium containing polar auxin transport inhibitors (N-1-naphthylphthalamic acid, 9-hydroxyfluorene-9-carboxylic acid, and 2,3,5-triiodobenzoic acid) and in medium containing a less well-characterized inhibitor of auxin-mediated processes, 2-(p-chlorophynoxy)-2-methylpropionic acid. Cotyledon vein pattern was not affected by any inhibitor treatments, although vein morphology was altered. In contrast, leaf vein pattern was affected by inhibitor treatments. Growth in polar auxin transport inhibitors resulted in leaves that lacked vascular continuity through the petiole and had broad, loosely organized midveins, an increased number of secondary veins, and a dense band of misshapen tracheary elements adjacent to the leaf margin. Analysis of leaf vein pattern developmental time courses suggested that the primary vein did not develop in polar auxin transport inhibitor-grown plants, and that the broad midvein observed in these seedlings resulted from the coalescence of proximal regions of secondary veins. Possible models for leaf vein patterning that could account for these observations are discussed. (+info)
Targeted antioxidant properties of N-[(tetramethyl-3-pyrroline-3-carboxamido)propyl]phthalimide and its nitroxide metabolite in preventing postischemic myocardial injury.
We investigated the cardioprotective efficacy of a new compound based on 2,2,5,5-tetramethyl-3-pyrroline-3-carboxamide (TPC-NH). Biochemical studies using electron paramagnetic resonance (EPR) spectroscopy suggest that TPC-NH is a scavenger of reactive oxygen species. In vitro cellular studies show that TPC-NH protects isolated cardiomyocytes against oxidative damage caused by superoxide radicals. Ex vivo EPR studies on the isolated rat heart indicate that the TPC-NH is metabolically oxidized to the nitroxide form. Studies were also performed in the isolated rat heart model to measure the efficacy of TPC-NH and its metabolites in preventing postischemic reperfusion injury. Serial measurements of contractile function were performed on hearts subjected to ischemia-reperfusion. Hearts were either untreated or treated with 50 microM TPC-NH or with its metabolites for 1 min before ischemia and during the first 5 min of reflow. TPC-NH showed marked protection with a more than 3-fold increased recovery of contractile function compared with control hearts, whereas its oxidative metabolites exhibited significant but lower protection. Thus, TPC-NH and, to a lesser extent, its oxidation metabolites exhibit potent membrane-targeted antioxidant action and exert marked protection against myocardial injury in the postischemic heart. (+info)
Dual-mode regulation of hair growth cycle by two Fgf-5 gene products.
As the result of alternative mRNA splicing, Fgf-5, the gene encoding fibroblast growth factor-5, translates to both long and short forms of the protein, respectively, designated fibroblast growth factor-5 and fibroblast growth factor-5S. We previously showed that localization of fibroblast growth factor-5 and the level of fibroblast growth factor-5S in murine skin are hair-cycle dependent. In this study, we examined the effect of fibroblast growth factor-5 and fibroblast growth factor-5S on the hair growth cycle in mice. Once the anagen phase of the hair growth cycle was induced in the dorsal skin by depilation during telogen, and effects of subcutaneous injection of fibroblast growth factor-5 and fibroblast growth factor-5S into the affected region were analyzed. We found that fibroblast growth factor-5 inhibited hair growth during anagen and promoted the transition from anagen to catagen. Interestingly, whereas fibroblast growth factor-5S alone exerted no effect on hair growth, it significantly inhibited the catagen-promoting activity of fibroblast growth factor-5 when the two proteins were injected simultaneously. Because neither fibroblast growth factor-5 nor fibroblast growth factor-5S affected skin thickness, it is postulated that changes in skin thickness during hair cycle are separately regulated by factors other than those regulating hair and follicle growth. The present results, together with our earlier findings that fibroblast growth factor-5-producing cells gather around dermal papillae during catagen, whereas fibroblast growth factor-5S is abundantly expressed in the hair follicles only during the latter half of anagen, suggests that the mouse hair growth cycle is regulated by the two Fgf-5 gene products acting in concert: fibroblast growth factor-5 induces catagen, whereas fibroblast growth factor-5S antagonizes this activity during anagen. (+info)
Auxin regulates the initiation and radial position of plant lateral organs.
Leaves originate from the shoot apical meristem, a small mound of undifferentiated tissue at the tip of the stem. Leaf formation begins with the selection of a group of founder cells in the so-called peripheral zone at the flank of the meristem, followed by the initiation of local growth and finally morphogenesis of the resulting bulge into a differentiated leaf. Whereas the mechanisms controlling the switch between meristem propagation and leaf initiation are being identified by genetic and molecular analyses, the radial positioning of leaves, known as phyllotaxis, remains poorly understood. Hormones, especially auxin and gibberellin, are known to influence phyllotaxis, but their specific role in the determination of organ position is not clear. We show that inhibition of polar auxin transport blocks leaf formation at the vegetative tomato meristem, resulting in pinlike naked stems with an intact meristem at the tip. Microapplication of the natural auxin indole-3-acetic acid (IAA) to the apex of such pins restores leaf formation. Similarly, exogenous IAA induces flower formation on Arabidopsis pin-formed1-1 inflorescence apices, which are blocked in flower formation because of a mutation in a putative auxin transport protein. Our results show that auxin is required for and sufficient to induce organogenesis both in the vegetative tomato meristem and in the Arabidopsis inflorescence meristem. In this study, organogenesis always strictly coincided with the site of IAA application in the radial dimension, whereas in the apical-basal dimension, organ formation always occurred at a fixed distance from the summit of the meristem. We propose that auxin determines the radial position and the size of lateral organs but not the apical-basal position or the identity of the induced structures. (+info)