Platelet-derived growth factor protects neurons against gp120-mediated toxicity

The human immunodeficiency virus (HIV)-1 envelope glycoprotein gp120 has been implicated in mediating neuronal apoptosis, a hallmark feature of HIV-associated dementia (HAD). Mitigation of the toxic effects of gp120 could thus be a potential mechanism for reducing HIV toxicity in the brain. In this study the authors hypothesized that neurotrophic factor, such as platelet-derived growth factor (PDGF), could protect the neurons against gp120-mediated apoptosis. SH-SY5Y cells treated with gp120 exhibited increased cell death when measured by lactate dehydrogenase (LDH) and deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL) assay, with concomitant loss of neurites and increased cell rounding. Pretreatment with PDGF-BB, however, reduced gp120-associated neurotoxicity and rescued the neurite outgrowth. Additionally, gp120-mediated activation of caspase-3 was also significantly reduced in cells pretreated with PDGF-BB. Antiapoptotic effects of PDGF-BB were also confirmed by monitoring levels of anti- and proapoptotic genes, Bcl-xL and Bax, respectively. Furthermore, PDGF-mediated protection against gp120 involved the phosphoinositide (PI) 3-kinase/Akt pathway. Taken together these findings lead us to suggest that PDGF-BB could be considered as a therapeutic agent that can mitigate gp120-mediated neurotoxicity in HAD.     

Platelet-derived growth factor induces chemotaxis of neuroepithelial stem cells

The ability of differentiating cells to migrate within the developing central nervous system (CNS) depends on extrinsic guidance signals, some of which are growth factors. In this study we have investigated the chemotactic response of cultured stem cells from the embryonic rat cortex to platelet-derived growth factor (PDGF). Nestin-positive stem cells from the developing CNS can be maintained and expanded in vitro under serum-free conditions in the presence of basic fibroblast growth factor (bFGF). Northern blot analysis of PDGF receptor expression revealed both α- and β-receptors on bFGF-treated neural stem cells. Both PDGF-AA and PDGF-BB readily induced directed migration of cultured neuroepithelial cells as measured in a microchemotaxis assay. Blocking of the migratory response was achieved by incubation with PDGF isoform-specific antibodies. More than 90% of the migrating cells were nestin-positive and incorporation of BrdU was also seen suggesting the cells to be immature and not yet committed to a specific cell lineage. These findings suggest a role for PDGF in cell migration in the developing cortex. J. Neurosci. Res. 53:521–530, 1998. © 1998 Wiley-Liss, Inc.     

Platelet-derived growth factor is mitogenic for O-2Aadult progenitor cells.

We report that platelet-derived growth factor (PDGF) is a potent mitogen for oligodendrocyte type-2 astrocyte (O-2A) progenitor cells derived from the optic nerves of adult rats. Moreover, O-2Aadult progenitors cultured in PDGF express the range of properties we have described previously for O-2Aadult progenitors cultured in the presence of type-1 astrocytes. Similarly, previous studies have demonstrated that PDGF is able to mimic the influence of type-1 astrocytes on O-2Aperinatal progenitors. Specifically, O-2Aadult progenitors and O-2Aperinatal progenitors exposed to PDGF express differences in average cell cycle time (59 +/- 5 h for O-2Aadult progenitors versus 20 +/- 6 h for O-2Aperinatal progenitors), average rate of migration (4.1 +/- 0.6 microns h-1 versus 24.6 +/- 5.4 microns h-1), morphology (unipolar versus bipolar), and antigenic phenotype (04+ vimentin- versus 04- vimentin+). Thus, our present results indicate that a single signalling molecule secreted by type-1 astrocytes produces markedly different cellular behaviours in two related O-2A progenitor populations.     

Platelet-derived growth factor in human glioma

Platelet-derived growth factor (PDGF) is a 30 kDa protein consisting of disulfide-bonded dimers of A- and B-chains. PDGF receptors are of two types, α- and β-receptors, which are members of the protein-tyrosine kinase family of receptors. The receptors are activated by ligand-induced dimerization, whereby the receptors become phosphorylated on tyrosine residues. These form attachment sites for signalling molecules, which inter alia activate the Ras · Raf pathway. PDGF has important functions in development and is required for a proper timing of oligodendrocyte differentiation. The v-sis oncogene of simian sarcoma virus (SSV) is a retroviral homolog of the B-chain gene, and induces transformation by an autocrine activation of PDGF receptors at the cell surface. SSV induces malignant glioma in experimental animals, suggesting a role for autocrine PDGF in glioma development. PDGF and PDGF receptors are frequently coexpressed in human glioma cell lines. Specific and nonspecific PDGF antagonists block the growth of some glioma cell lines in vitro and in vivo, suggesting that autocrine PDGF is involved in transformation and tumorigenesis. In situ studies of human gliomas show overexpression of α-receptors in glioma cells of high-grade tumors. In a few cases, overexpression is caused by receptor amplification. Since high-grade glioma cells also express the PDGF A-chain, an autocrine activation of the α-receptor may drive the proliferation of glioma cells in vivo. © 1995 Wiley-Liss, Inc.     

Nerve growth factor receptor-immunoreactive neurons within the developing human cortex.

A monoclonal antibody recognizing the p75 receptor for nerve growth factor (NGF) was used to assess the immunohistochemical expression of NGF receptors within the developing human neo-, limbic, and paralimbic cortices as well as the hippocampal complex. Between embryonic weeks 16 and 26, a transient population of neurons located within the upper and lower subplate zones of the neo-, limbic, and paralimbic cortices expressed the receptor for NGF. In contrast, NGF receptor-immunoreactive neurons were only observed in the upper subplate zone of the entorhinal cortex at embryonic week 40 (term), a staining pattern not observed in a 5-year-old specimen. The expression of NGF receptor-immunoreactive neurons within the upper subplate zone between embryonic weeks 16 and 40 was characterized by a dense band of immunoreactive neurons and neuropil. These neurons were bipolar with basal and apically directed neurites. NGF receptor-immunoreactive neurons were also scattered throughout the lower subplate zone and underlying white matter between embryonic weeks 19 and 26. These neurons were multipolar, with less apically directed neurites. NGF receptor-immunoreactive subplate neurons displayed a topographic distribution with the heaviest concentration found within limbic and paralimbic cortices as well as association neocortex. In contrast, light to moderate NGF receptor-immunoreactivity was seen in sensory-motor cortex. Within the hippocampal complex, only a few lightly stained NGF receptor-immunoreactive neurons were seen within the fimbria, hilar region of the dentate gyrus, and subiculum. The expression of NGF receptor-immunoreactivity increased within the subplate zone of the pre- and parasubiculum culminating in intense entorhinal cortex staining. As the entorhinal cortex merged with the developing inferior temporal association cortex, there was a marked reduction in staining intensity. In contrast to those in the subplate zone, neurons within the germinal zone and cortical plate were NGF receptor immunonegative at all times examined. The presence of NGF receptors in the subplate zone suggests that neurotrophins such as NGF play an important role in the transient viability of these neurons as well as in the guidance of cortical afferent inputs into topographically organized regions of the cerebral cortex.     

Platelet-derived growth factor receptors of mouse central nervous system cells in vitro

This study evaluates the distribution of receptors for platelet-derived growth factor (PDGF) on central nervous cells maintained in vitro using colloidal gold-labeled immunocytochemical markers at the electron microscopic level. Platelet-derived growth factor receptors were found to be sparsely distributed over the surface of type 1 astrocytes, apparent type 2 astrocytes, and neurons. Receptors appeared to be preferentially associated with filopodia-like extensions of the cell membrane. The existence of functional receptors was confirmed using the impermeant, water-soluble affinity cross-linking agent bis(sulfosuccinimidyl) suberate to covalently link radiolabeled PDGF to its receptor. The PDGF/receptor complexes could also be immunoprecipitated with the same antibody used in immunocytochemical experiments. The improved resolution of these techniques allows definitive identification of PDGF receptors on cultured mammalian central nervous system cells other than oligodendrocytes. These data expand the range of possible roles of PDGF during nervous system development. Receptors for PDGF are likely to play a key role in the differentiation of cells in the central nervous system. © 1995 Wiley-Liss, Inc.     

Platelet-derived growth factor is a survival factor for PSA-NCAM + oligodendrocyte pre-progenitor cells

Mature oligodendroglia, which synthesize and express lipids and proteins characteristic of myelin, are generated from precursor cells which are formed in germinal matrix, then migrate widely through the neuraxis. We now demonstrate that these precursor cells can be recognized at a very early stage by their surface expression of polysialylated neural cell adhesion molecules (PSA-NCAM), and only later bind anti ganglioside antibodies that had previously been used to recognize “O-2A” oligodendroglial precursor cells. PSA-NCAM expression by these cells is likely to be of functional significance, since a recent study demonstrated that O-2A cells become immobile when stripped of PSA-NCAM. Platelet-derived growth factor (PDGF) proved to be a survival factor for these PSA-NCAM + cells, and in a defined medium, PDGF was sufficient to ensure maturation of immunopurified PSA-NCAM + cells to oligodendroglia. © 1995 Wiley-Liss, Inc.     

Platelet-derived growth factor inhibits platelet activation in heparinized whole blood.

We previously have demonstrated that human platelets have functionally active platelet-derived growth factor alpha-receptors. Studies with gel-filtered platelets showed that an autocrine inhibition pathway is transduced through this tyrosine kinase receptor during platelet activation. The physiological significance of this inhibitory effect of platelet-derived growth factor on gel-filtered platelets activation is, however, not known. In the present study, we investigated whether platelet-derived growth factor inhibits platelet activation under more physiological conditions in heparinized whole blood, which represents a more physiological condition than gel-filtered platelets. Using flow cytometric assays, we demonstrate here that platelet-derived growth factor inhibits thrombin-, thrombin receptor agonist peptide SFLLRN-, and collagen-induced platelet aggregation and shedding of platelet-derived microparticles from the platelet plasma membrane during platelet aggregation in stirred heparinized whole blood. The inhibitory effect of platelet-derived growth factor was dose dependent. However, under nonaggregating conditions (no stirring), we could not demonstrate any significant effect of platelet-derived growth factor on thrombin- and thrombin receptor agonist peptide-induced platelet surface expression of P-selectin. Our results demonstrate that platelet-derived growth factor appears to be a true antithrombotic agent only under aggregating conditions in heparinized whole blood.     

Platelet-derived growth factor and regulation of Schwann cell proliferation in vivo.

To examine the role of platelet-derived growth factor (PDGF) in the in vivo regulation of Schwann cell proliferation, steady-state levels of mRNAs encoding PDGF A and B chains, and PDGF alpha and beta receptors were measured in immature and adult rat sciatic nerves and in cultured rat Schwann cells. PDGF B chain and PDGF beta receptor mRNAs are present in immature rat sciatic nerves and to a lesser extent in adult rat nerves. Short-term cultures of neonatal rat Schwann cells express PDGF beta receptor mRNA, but not PDGF B chain mRNA, and are stimulated to synthesize DNA by addition of PDGF BB to the medium. These data indicate that PDGF BB is a developmentally regulated paracrine growth factor for rat Schwann cells. Very long-term cultures of rat Schwann cells, which have lost normal dependence on exogenous growth factors, express PDGF B chain mRNA as well as mRNAs encoding the PDGF alpha and beta receptors, suggesting that, under these circumstances, PDGF BB also act as an autocrine growth factor. PDGF A chain mRNA is present in both immature and adult rat sciatic nerves and is expressed by primary and secondary cultures of rat Schwann cells as well. However, because the abundance of PDGF alpha receptor mRNA is very low in rat Schwann cells, PDGF AA is not likely to be a significant autocrine growth factor for rat Schwann cells.     

Platelet-derived growth factor receptor is expressed by cells in the early oligodendrocyte lineage

We report the loaclization of PDGFRα mRNA (PDGFRα) in phenotypically defined cells during the first postanatal week of rat forebrain development. Using a method of combined immunocytochemistry and in situ hybridization we have demonstrated the cellular colocalization of PDGFRα mRNA with GD₃ ganglioside or 04 sulfatide, phenotypic markers of oligodendrocytes, in the gray and white matter of the dorsal cerebral cortex at all ages studies. Population analysis of the PDGFRα+/G₃+ and PDGFRα+/analysis of the PDGFRα+/G_{D 3}+and PDGFRα+/04+ cells revealed that three populations express PDGFRα: G_{D 3}+, G_{D 3}+/04+, and 04+, corresponding to two lineage stages, progenitor and preoligodendrocyte, in oligodendrocyte development. Immature oligodendrocytes, identifice by galactocerebroside immunoreactivity, did not express detectable levels of PDGFR α mRNA. Post-mitotic neurons, identifice by immunoperodxidase localization of the 68 kD neurofilament, and astrocytes identified by S-100 or GFAP immunoreactivity were also negative for PDGFRα mRNA occured in oligodendrocyte cell populations which are post-migratory and proliferative, but which do not express myelin proteins characteristic of post-mitotic oligodendrocytes. © 1994 Wiley-Liss, Inc.     

Nerve growth factor increases choline acetyltransferase activity in developing basal forebrain neurons

Nerve growth factor (NGF) is a neuronotrophic protein. Its effects on developing peripheral sensory and sympathetic neurons have been extensively characterized, but it is not clear whether NGF plays a role during the development of central nervous system neurons. To address this point, we examined the effect of NGF on the activity of neurotransmitter enzymes in several brain regions. Intracerebroventricular injections of highly purified mouse NGF had a marked effect on the activity of choline acetyltransferase (ChAT), a selective marker of cholinergic neurons. NGF elicited prominent increases in ChAT activity in the basal forebrain of neonatal rats, including the septum and a region which contains neurons of the nucleus basalis and substantia innominata. NGF also increased ChAT activity in the hippocampus and neocortex, terminal regions for the fibers of basal forebrain cholinergic neurons. In analogy with the response of developing peripheral neurons, the NGF effect was shown to be selective for basal forebrain cholinergic cells and to be dose-dependent. Furthermore, septal neurons closely resembled sympathetic neurons in the time course of their response to NGF. These observations suggest that endogenous NGF does play a role in the development of basal forebrain cholinergic neurons.     

Nerve growth factor regulates synaptophysin expression in developing trigeminal ganglion neurons in vitro

The role of neuronal growth factors in synaptic maturation of sensory neurons, including trigeminal ganglion (TG) neurons, remains poorly understood. Here, we show that nerve growth factor (NGF) regulates the intracellular distribution of the synaptic vesicle protein synaptophysin (Syp) in newborn rat TG neurons in vitro. While reducing the number of Syp-positive cell bodies, NGF dramatically increases Syp immunoreactivity in both proximal and distal segments of the neurite. Intriguingly, the increase in Syp immunoreactivity occurs only in neuron-enriched cultures, in which the number of non-neuronal cells is significantly reduced. Together, our data indicate that NGF is a candidate molecule involved in early postnatal maturation of TG neurons, including control of presynaptic assembly, and thereby formation of synaptic connections.     

Platelet-derived growth factor-BB pretreatment attenuates excitotoxic death in cultured hippocampal neurons

Neuronal excitotoxic death results from excess stimulation by elevated levels of extracellular glutamate acting on N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors. While excitotoxicity is typically attenuated by using glutamate receptor antagonists, we report here that neuronal deaths induced directly by brief exposures to glutamate or NMDA were both attenuated by preincubation with platelet-derived growth factor-BB (PDGF-BB). The neuroprotection was concentration and time dependent; preincubation for at least 24 h with a minimum of 10 ng/mL of PDGF-BB was required for maximal neuroprotective effect. The NMDA receptor antagonist MK-801 also afforded partial protection, and when MK-801 was used with PDGF-BB, neuronal survival was comparable to that of untreated controls. When protection of inhibitory and excitatory neurons by PDGF treatment was compared, the excitatory neurons appeared to be selectively protected. The present results demonstrate that PDGF pretreatment can protect neurons from direct glutamate-induced excitotoxicity in vitro and suggests that PDGF might possibly function as a neuroprotective agent in potential therapeutic applications.     

血小板源生长因子和脑源性神经营养因子促进NG2蛋白聚糖阳性神经祖细胞向神经元分化

目的探讨血小板源生长因子(PDGF)和脑源性神经营养因子(BDNF)对NG2蛋白聚糖阳性神经祖细胞(NG2细胞)分化的作用。方法根据文献方法从成年大鼠海马原代培养NG2细胞,7d后添加10~40ng/m lPDGF和/或BDNF继续培养7d,以免疫荧光双重染色法鉴定细胞性质。结果10ng/m l PDGF和BDNF联合处置明显增加Map2a/b、GAD67和GABA表达水平。结论PDGF和BDNF联合处置促进NG2细胞向神经元(特别是GABA能抑制性中间神经元)分化。     

Platelet-derived growth factor delays oligodendrocyte differentiation and axonal myelination in vivo in the anterior medullary velum of the developing rat

The AA dimeric form of platelet-derived growth factor (PDGF-AA) is implicated in the differentiation of cells of the oligodendrocyte lineage, which express PDGF receptors of the alpha subunit type (PDGF-αR). In the present study, we show that a single injection of PDGF-AA into the cerebrospinal fluid of neonatal rats delays oligodendrocyte differentiation and interrupts the progress of myelination in the anterior medullary velum (AMV), a white matter tract roofing the IVth ventricle of the brain. PDGF-AA or saline was injected intrathecally in postnatal day (P) 7 rats, and the AMV was subsequently removed and immunolabelled with the oligodendrocyte-specific antibody Rip, at P9, P12, and P21, corresponding to postinjection days (PID) 2, 5, and 14. At P9 (PID2), myelination was retarded in PDGF-AA-treated rats as opposed to saline-treated controls but progressed rapidly after P12 (PID5). Quantification supported the qualitative observations that PDGF-AA mediated an acute decrease in the number of Rip+ oligodendrocytes at P9–12, which largely recovered by P21, suggesting that PDGF-AA may have delayed recruitment of myelinating oligodendrocytes. However, the definitive number of Rip+ oligodendrocytes in the AMV was not increased, suggesting that its action as a promoter of early oligodendrocyte survival may not ultimately affect the definitive number of myelinating oliogdendrocytes in vivo. We discuss the possibilities that excess PDGF-AA may have acted on early oligodendrocytes (precursors or preoligodendrocytes) to either (1) delay their differentiation by maintaining them in the cell cycle or (2) accelerate their differentiation, which may result in premature cell death in the absence of synchronised survival signals. This study supports a role for PDGF-AA in the timing of oligodendrocyte differentiation in vivo, as has been shown in vitro. J. Neurosci. Res. 48:588–596, 1997. © 1997 Wiley-Liss Inc.     

Electron microscopic immunocytochemical localization of nerve growth factor in developing mouse olfactory neurons

The immunocytochemical localization of nerve growth factor (NGF) in the embryonic mouse has been examined using correlative light and electron microscope procedures. In flat-embedded thick sections, primary sensory neurons of the olfactory pathway were clearly visible by their high level of NGF-like immunoreactivity. When ultrathin sections were examined under the electron microscope, the HRP reaction product present in the nasal epithelium was confined to the cytoplasm of the chemoreceptor cells, including the dendrites projecting into the nasal cavity. The axons projecting to the olfactory bulb and axon bundles within the bulb were also stained specifically for NGF. The reaction product did not appear to be associated with any particular subcellular structure. Neither the columnar supporting cells nor the basal precursor cells showed any evidence of immunoreactivity. No evidence was found for the presence of NGF in cells such as epithelial or glial cells within the immediate environment of the receptor neurons. These first subcellular localization studies indicate the presence of high levels of NGF in neurons not known to be sensitive to the trophic molecule.     

Nerve growth factor and chronic ethanol treatment alter calcium homeostasis in developing rat septal neurons

Chronic ethanol treatment (CET) during development produces cellular adaptations resulting in tolerance to the acute effects of ethanol (EtOH). The objectives of this study were to determine whether CET during the prenatal period (PCET) followed by a period of in vitro CET (PCET-CET) altered intracellular calcium [Ca(2+)](i) and produced tolerance to acute EtOH treatment (AET), and whether nerve growth factor (NGF) modulated the effects of PCET-CET in cultured developing rat septal neurons. Fetuses were obtained from EtOH-fed and sucrose-fed (diet-control) female rats. Neurons from PCET fetuses were cultured in the presence of NGF (+NGF) and 200 mg/dl (mg %) EtOH and diet-control cultures received NGF and no EtOH. PCET and diet-control cultures were then divided into two groups, +NGF and -NGF (withdrawn from NGF), and exposed acutely to one of five doses of EtOH during stimulation with potassium (K(+)) chloride. [Ca(2+)](i) was measured using fura-2. PCET-CET did not affect resting [Ca(2+)](i). PCET-CET decreased and acute EtOH withdrawal increased overall K(+)-stimulated changes in [Ca(2+)](i), but only in +NGF PCET neurons. Reducing the level of EtOH from 200 to 100 mg % decreased overall K(+)-stimulated [Ca(2+)](i) in -NGF PCET neurons. The effects of PCET-CET or PCET-CET combined with NGF on overall K(+)-stimulated changes in [Ca(2+)](i) occurred mostly in the early and middle phases of the K(+)-response. NGF reduced overall K(+)-stimulated changes in [Ca(2+)](i) in PCET neurons during EtOH withdrawal and during AET with 200 mg % EtOH and increased overall K(+)-stimulated changes in [Ca(2+)](i) during AET with 400 and 800 mg % EtOH. There was no effect of NGF on overall K(+)-stimulated changes in [Ca(2+)](i) in diet-control neurons with the exception that NGF-treatment decreased overall K(+)-stimulated changes in [Ca(2+)](i) during AET with 400 mg % EtOH. The effects of AET on overall K(+)-stimulated changes in [Ca(2+)](i) mostly occurred in +NGF PCET neurons. In conclusion, CET during development of the brain could adversely affect Ca(2+)-dependent functions such as neuronal migration, neurite outgrowth, and synaptogenesis in neurons even in the presence of neurotrophin support.     

Platelet-derived growth factor promotes repair of chronically demyelinated white matter

In multiple sclerosis, remyelination becomes limited after repeated or prolonged episodes of demyelination. To test the effect of platelet-derived growth factor-A (PDGF-A) in recovery from chronic demyelination we induced corpus callosum demyelination using cuprizone treatment in hPDGF-A transgenic (tg) mice with the human PDGF-A gene under control of an astrocyte-specific promoter. After chronic demyelination and removal of cuprizone from the diet, remyelination and oligodendrocyte density improved significantly in hPDGF-A tg mice compared with wild-type mice. In hPDGF-A tg mice, oligodendrocyte progenitor density and proliferation values were increased in the corpus callosum during acute demyelination but not during chronic demyelination or the subsequent recovery period, compared with hPDGF-A tg mice without cuprizone or to treatment-matched wild-type mice. Proliferation within the subventricular zone and subcallosal zone was elevated throughout cuprizone treatment but was not different between hPDGF-A tg and wild-type mice. Importantly, hPDGF-A tg mice had reduced apoptosis in the corpus callosum during the recovery period after chronic demyelination. Therefore, PDGF-A may support oligodendrocyte generation and survival to promote remyelination of chronic lesions. Furthermore, preventing oligodendrocyte apoptosis may be important not only during active demyelination but also for supporting the generation of new oligodendrocytes to remyelinate chronic lesions.     

Platelet-derived growth factor signals play critical roles in differentiation of longitudinal smooth muscle cells in mouse embryonic gut

Abstract  In the development of mouse gut, longitudinal smooth muscle cells (LMC) and interstitial cells of Cajal (ICC) originate from common precursor cells expressing c-Kit. Recently, some gastrointestinal stromal tumours, which develop from smooth muscle layers of the gut and have gain-of-function mutations of c-kit , have been reported to have gain-of-function mutations of platelet-derived growth factor (PDGF) receptor α gene. These data raise the possibility that PDGF signalling might be involved in the development of LMC. Therefore, we examined the expression pattern of the PDGF signal family of embryonic gut by immunohistochemistry and in situ hybridization, and investigated the role of PDGF signals in the development of smooth muscle layers in mouse gut using a new organ culture system. During embryonic development, the circular muscle layer expressed PDGF-A, enteric neurons expressed PDGF-B and common precursor cells of LMC and ICC expressed both PDGF receptor α and β. The selective PDGF receptor inhibitor AG1295 suppressed the differentiation of LMC in gut explants. We conclude that PDGF signals play critical roles in the differentiation of LMC in mouse embryonic gut.