Thrombin: a neuronal cell modulator.
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Biomedical subjects
V L Turgeon
Publications and source records attributed to V L Turgeon.
Pigment epithelium-derived factor promotes the survival and differentiation of developing spinal motor neurons.
Pigment epithelium-derived factor (PEDF) is a member of the serine protease inhibitor (serpin) superfamily that has been shown previously to promote the survival and/or differentiation of rat cerebellar granule neurons and human retinoblastoma cells in vitro. However, in contrast to most serpins, PEDF has no inhibitory activity against any known proteases, and its described biological activities do not appear to require the serpin-reactive loop located toward the carboxy end of the polypeptide. Because another serpin, protease nexin-1, has been shown to promote the in vivo survival and growth of motor neurons, the authors investigated the potential neurotrophic effects of PEDF on spinal cord motor neurons in highly enriched cultures and in vivo after injury. Here, it is shown that native bovine and recombinant human PEDF promoted the survival and differentiation (neurite outgrowth) of embryonic chick spinal cord motor neurons in vitro in a dose-dependent manner. A truncated form of PEDF that lacks approximately 62% of the carboxy end of the polypeptide comprising the homologous serpin-reactive loop also exhibited neurotrophic activities similar to those of the full-length protein. Furthermore, the data here showed that PEDF was transported retrogradely and prevented the death and atrophy of spinal motor neurons in the developing neonatal mouse after axotomy. These results indicate that PEDF exerts trophic effects on motor neurons, and, together with previous reports, these findings suggest that this protein may be useful as a pharmacologic agent to promote the development and maintenance of motor neurons. J. Comp. Neurol. 412:506-514, 1999. Published 1999 Wiley-Liss, Inc.
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Activation of the protease-activated thrombin receptor (PAR)-1 induces motoneuron degeneration in the developing avian embryo.
Several studies have shown that both neuronal and glial cells express functional thrombin receptors as well as prothrombin transcripts. Recently, we (and others) have shown that alpha-thrombin induces apoptotic cell death in different neuronal cell types, including motoneurons, in culture. Thrombin-induced effects on different cells are mediated through the cell surface protease-activated thrombin receptor, PAR-1. Furthermore, it has been shown that, in contrast to thrombin, which induces proteolysis of other proteins besides its receptor, the thrombin receptor agonist peptide, serine-phenylalanine-leucine-leucine-arginine-asparagine-proline (SFLLRNP), is only known to activate this receptor. However, whether activation of the thrombin receptor in vivo affects the development of spinal cord motoneurons is not known. Here, we show that treatment with a synthetic SFLLRNP peptide induced a dose-dependent degeneration and death of spinal motoneurons both in highly enriched cultures and in the developing chick embryo in vivo. However, cotreatment with caspase inhibitors completely abolished SFLLRNP-induced motoneuron death both in vitro and in vivo. These results suggest that developing motoneurons express functionally active PAR-1 whose activation leads to cell death through stimulation of the caspase family of proteins. Our findings also suggest a novel and deleterious role for PAR-like receptors in the central nervous system, different from their previously known functions in the vascular and circulatory system.
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Prevention of thrombin-induced motoneuron degeneration with different neurotrophic factors in highly enriched cultures.
Previous reports have shown that neuronal and glial cells express functionally active thrombin receptors. The thrombin receptor (PAR-1), a member of a growing family of protease activated receptors (PARs), requires cleavage of the extracellular amino-terminus domain by thrombin to induce signal transduction. Studies from our laboratory have shown that PAR-1 activation following the addition of thrombin or a synthetic thrombin receptor activating peptide (TRAP) induces motoneuron cell death both in vitro and in vivo. In addition to increasing motoneuron cell death, PAR- 1 activation leads to decreases in the mean neurite length and side branching in highly enriched motoneuron cultures. It has been suggested that motoneuron survival depends on access to sufficient target-derived neurotrophic factors through axonal branching and synaptic contacts. However, whether the thrombininduced effects on motoneurons can be prevented by neurotrophic factors is still unknown. Using highly enriched avian motoneuron cultures, we show here that alone, soluble chick skeletal muscle extracts (CMX), brain-derived neurotrophic factor (BDNF), ciliary neurotrophic factor (CNTF), and glial cell line-derived neurotrophic factor (GDNF) significantly increased motoneuron survival compared to controls, whereas nerve growth factor (NGF) did not have a significant effect on motoneuron survival. Furthermore, cotreatment with muscle-derived agents (i.e., CMX, BDNF, GDNF) significantly prevented the death of motoneurons induced by alpha-thrombin. Yet, non-muscle-derived agents (CNTF and NGF) had little or no significant effect in reversing thrombin-induced motoneuron death. CMX and CNTF significantly increased the mean length of neurites, whereas NGF, BDNF, and GDNF failed to enhance neurite outgrowth compared to controls. Furthermore, CMX and CNTF significantly prevented thrombin-induced inhibition of neurite outgrowth, whereas BDNF and GDNF only partially reversed thrombin-induced inhibition of neurite outgrowth. These findings show differential effects of neurotrophic factors on thrombin-induced motoneuron degeneration and suggest specific overlaps between the trophic and stress pathways activated by some neurotrophic agents and thrombin, respectively.
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Thrombin perturbs neurite outgrowth and induces apoptotic cell death in enriched chick spinal motoneuron cultures through caspase activation.
Increasing evidence indicates several roles for thrombin-like serine proteases and their cognate inhibitors (serpins) in normal development and/or pathology of the nervous system. In addition to its prominent role in thrombosis and/or hemostasis, thrombin inhibits neurite outgrowth in neuroblastoma and primary neuronal cells in vitro, prevents stellation of glial cells, and induces cell death in glial and neuronal cell cultures. Thrombin is known to act via a cell surface protease-activated receptor (PAR-1), and recent evidence suggests that rodent neurons express PAR-1. Previously, we have shown that the thrombin inhibitor, protease nexin-1, significantly prevents neuronal cell death both in vitro and in vivo. Here we have examined the effects of human alpha-thrombin and the presence and/or activation of PAR-1 on the survival and differentiation of highly enriched cultures of embryonic chick spinal motoneurons. We show that thrombin significantly decreased the mean neurite length, prevented neurite branching, and induced motoneuron death by an apoptosis-like mechanism in a dose-dependent manner. These effects were prevented by cotreatment with hirudin, a specific thrombin inhibitor. Treatment of the cultures with a synthetic thrombin receptor-activating peptide (SFLLRNP) mimicked the deleterious effects of thrombin on motoneurons. Furthermore, cotreatment of the cultures with inhibitors of caspase activities completely prevented the death of motoneurons induced by either thrombin or SFLLRNP. These findings indicate that (1) embryonic avian spinal motoneurons express functional PAR-1 and (2) activation of this receptor induces neuronal cell degeneration and death via stimulation of caspases. Together with previous reports, our results suggest that thrombin, its receptor(s), and endogenous thrombin inhibitors may be important regulators of neuronal cell fate during development, after injury, and in pathology of the nervous system.
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The role of thrombin-like (serine) proteases in the development, plasticity and pathology of the nervous system.
There is increasing evidence suggesting that members of the serine protease family, including thrombin, chymotrypsin, urokinase plasminogen activator, and kallikrein, may play a role in normal development and/or pathology of the nervous system. Serine proteases and their cognate inhibitors have been shown to be increased in the neural parenchyma and cerebrospinal fluid following injury to the blood brain barrier. Zymogen precursors of thrombin and thrombin-like proteases as well as their receptors have also been localized in several distinct regions of the developing or adult brain. Thrombin-like proteases have been shown to exert deleterious effects, including neurite retraction and death, on different neuronal and non-neuronal cell populations in vitro. These effects appear to be mediated through cell surface receptors and can be prevented or reversed with specific serine protease inhibitors (serpins). Furthermore, we have recently shown that treatment with protease nexin-1 (a serpin that inhibits thrombin-like proteases) promotes the survival and growth of spinal motoneurons during the period of programmed cell death and following injury. Taken together, these observations suggest that thrombin-like proteases play a deleterious role, whereas serpins promote the development and maintenance of neuronal cells. Thus, changes in the balance between serine proteases and their cognate inhibitors may lead to pathological states similar to those associated with some neurodegenerative diseases such as Alzheimer's disease. The present review summarizes the current state of research involving such serine proteases and speculates on the possible role of these thrombin-like proteases in the development, plasticity and pathology of the nervous system.
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