The NCAM-derived P2 peptide facilitates recovery of cognitive and motor function and ameliorates neuropathology following traumatic brain injury

The neural cell adhesion molecule (NCAM) plays a crucial role during development and regeneration of the nervous system, mediating neuronal differentiation, survival and plasticity. Moreover, NCAM regulates learning and memory. A peptide termed P2, corresponding to a 12-amino-acid sequence in the second immunoglobulin (Ig)-like module of NCAM, represents the natural cis -binding site for the first NCAM Ig module. The P2 peptide targets NCAM, thereby inducing a number of intracellular signaling events leading to the stimulation of neurite outgrowth and promotion of neuronal survival in vitro . The present study evaluated the effect of the P2 peptide on functional and histological outcomes following traumatic brain injury inflicted by a cortical cryogenic lesion. Lesioned rats were injected subcutaneously with P2 peptide, 5 mg/kg daily for 15 days beginning 2 h after injury. This treatment significantly improved postlesion recovery of motor and cognitive function, reduced neuronal degeneration, protected cells against oxidative stress, and increased reactive astrogliosis and neuronal plasticity in the sublesional area. P2 appeared rapidly in blood and cerebrospinal fluid after subcutaneous administration and remained detectable in blood for up to 5 h. The results suggest that P2 has therapeutic potential for the treatment of traumatic brain injury.     

Behavioral profile of mice with impaired cognition in the elevated plus-maze due to a deficiency in neural cell adhesion molecule

The elevated plus-maze (EPM) test is one of the most used tests for screening levels of anxiety in rodents. In the present study, we studied how impaired cognition due to a deficiency in the neural cell adhesion molecule (NCAM) could affect the behavior of mice in the EPM task. NCAM-knockout mice demonstrated impaired learning in both object-recognition and fear-conditioning tasks. Analysis of the behavior of mice in the EPM task using a minute-by-minute method revealed a profound influence of genotype. Wild-type mice demonstrated quick learning of the aversive properties of the open arms during the first few minutes of a single EPM task, whereas NCAM−/− mice were unable to learn the aversive properties of the open arms of EPM. Wild-type mice also demonstrated habituation to the EPM task in a test/retest paradigm whereas NCAM-knockout mice failed to habituate during the second EPM presentation. Our data show that the anxiolytic-like behavior of NCAM-knockout mice is not just related to levels of innate anxiety but also to their inability to recognize potential danger associated with the open arms of the EPM task.     

NCAM在成年大鼠不同视神经再生模型中表达的研究

目的检测神经细胞黏附分子（NCAM）在成年大鼠不同视神经再生模型中的表达，探讨NCAM分布情况与其在视神经再生中的作用。方法选择64只健康成年雄性sD大鼠，随机分成2组，自体视神经一坐骨神经吻合模型32只为实验组，自体视神经一视神经吻合模型32只作对照组。于吻合术后15天、30天、45天、60天处死动物并取出吻合段神经。利用免疫组织化学方法（SP法）和计算机图像分析技术，测定NCAM在不同吻合模型中的表达。结果NCAM在视神经一坐骨神经吻合模型表达水平高于视神经一视神经吻合模型。吻合术后早期表达水平明显高于末期。结论坐骨神经移植有助于增加NCAM表达，促进视神经再生。     

Schwann cells genetically engineered to express PSA show enhanced migratory potential without impairment of their myelinating ability in vitro

Schwann cells, the myelin-forming cells of the PNS, are attractive candidates for remyelination therapy as they can remyelinate CNS axons. Yet their integration in CNS tissue appears hampered, at least in part, by their limited motility in the CNS environment. As the polysialylated (PSA) form of NCAM regulates migration of neural precursors in the CNS and is not expressed by developing Schwann cells, we investigated whether conferring sustained expression of PSA to Schwann cells derived from postnatal rats enhances their motility. Cells were transduced with a retrovirus encoding polysialyl-transferase STX, an enzyme that synthesizes PSA on NCAM. Migration of wild type and transduced cells expressing STX or the marker gene alkaline phosphatase was examined using a gap bridging assay in dissociated cells and by grafting cells in slice cultures of postnatal brain. Migration of PSA expressing cells was significantly increased in both models, as compared to control cells, and this effect was abolished by endoneuraminidase-N stripping of PSA. PSA-positive Schwann cells retained the ability to differentiate in vitro and expressed the Krox20 and P zero myelination markers. When grafted in neonatal cerebellar slices, STX-transduced cells started to myelinate Purkinje cell axons like control cells and make myelin internodes after 2 to 3 weeks. PSA was redistributed on the cell membrane and downregulated during differentiation in pure Schwann cell cultures and slice co-cultures. Thus, migratory properties of PNS myelin-forming cells within the CNS can be enhanced without altering their differentiation program. This finding may be beneficial for the development of remyelination therapies.     

Hippocampal up-regulation of NCAM expression and polysialylation plays a key role on spatial memory

Memory formation has been associated with structural and functional modifications of synapses. Cell adhesion molecules are prominent modulators of synaptic plasticity. Here, we investigated the involvement of the cell adhesion molecules, NCAM, its polysialylated state (PSA-NCAM) and L1 in spatial learning-induced synaptic remodeling and memory storage. A differential regulation of these adhesion molecules was found in the hippocampus of rats submitted to one training session in the spatial, but not cued, version of the Morris water maze. Twenty-four hours after training, synaptic expression of NCAM and PSA-NCAM was increased, whereas L1 appeared markedly decreased. The regulation of these molecules was spatial learning-specific, except for L1 reduction, which could be attributed to swimming under stressful conditions rather than to learning. Subsequent psychopharmacological experiments were performed to address the functional role of NCAM and PSA-NCAM in the formation of spatial memories. Rats received an intracerebroventricular injection of either a synthetic peptide (C3d) aimed to interfere with NCAM function, or endoneuraminidase, an enzyme that cleaves polysialic acid from NCAM. Both treatments affected acquisition of spatial information and lead to impaired spatial memory abilities, supporting a critical role of the observed learning-induced up-regulation of synaptic NCAM expression and polysialylation on spatial learning and memory. Therefore, our findings highlight NCAM as a learning-modulated molecule critically involved in the hippocampal remodeling processes underlying spatial memory formation.     

Presence of nerve growth factor and TrkA expression in the SVZ of EAE rats: evidence for a possible functional significance

Nerve growth factor (NGF) is a well-characterized neurotrophic factor that plays a crucial role during development in the growth, differentiation, and maintenance of brain neurons as well as in the reparative response of the adult brain to neuronal damage. Recent studies have shown that acute axonal loss occurs in multiple sclerosis (MS) and its animal model, experimental allergic encephalomyelitis (EAE), and that NGF suppresses clinical symptoms of EAE in nonhuman primates. Aim of the present study was to investigate the role of NGF in the regenerative response of the adult brain to neuronal damage occurring in EAE. Using EAE rats, we have found that exogenous NGF injection and NGF deprivation (NGF autoimmunization) can act on growth and differentiation of brain precursor cells in the subventricular zone (SVZ) of EAE rats. Moreover, NGF administration in brain of EAE rats stimulates the expression of early neuronal markers on proliferating precursor cells of the SVZ. The data obtained demonstrated that NGF and its antibody affect bromodeoxyuridine (BrdU) incorporation and NGF receptor expression by SVZ progenitor cells in the brain of EAE rats.     

Macrophage-induced inflammation affects hippocampal plasticity and neuronal development in a murine model of HIV-1 encephalitis

Cognitive, behavioral, and motor impairments, during progressive human immunodeficiency virus type 1 (HIV-1) infection, are linked to activation of brain mononuclear phagocytes (MP; perivascular macrophages and microglia). Activated MPs effect a giant cell encephalitis and neuroinflammatory responses that are mirrored in severe combined immunodeficient (SCID) mice injected with human monocyte-derived macrophages (MDM). Whether activated human MDMs positioned in the basal ganglia affect hippocampal neuronal plasticity, the brain subregion involved in learning and memory, is unknown. Thus, immunohistochemical techniques were used for detection of newborn neurons (polysialylated neuronal cell adhesion molecule [PSA-NCAM]) and cell proliferation (Ki-67) to assay MDM effects on neuronal development in mouse models of HIV-1 encephalitis. Immunodeficient (C.B.-17/SCID and nonobese diabetic/SCID, NOD/SCID) and immune competent (C.B.-17) mice were injected with uninfected or HIV-1-infected MDM. Sham-operated or unmanipulated mice served as controls. Neuronal plasticity was evaluated in the hippocampal dentate gyrus (DG) at days 7 and 28. By day 7, increased numbers of Ki-67+ cells, PSA-NCAM+ cells and dendrites in DG were observed in sham-operated animals. In contrast, significant reductions in neuronal precursors and altered neuronal morphology paralleled increased microglial activation in both HIV-1-infected and uninfected MDM-injected animals. DG cellular composition was restored at day 28. We posit that activated MDM induce inflammation and diminish DG neuronal plasticity. These data provide novel explanations for the cognitive impairments manifested during advanced HIV-1 infection.     

神经细胞粘附分子(NCAM)在脑胶质瘤中的表达及其与PCNA的相关性

 目的　检测胶质瘤中神经细胞粘附分子(NCAM)和增殖细胞核抗原(PCNA)的表达状况,以探讨不同病理类型胶质瘤中二者的表达及其相关性。方法　用抗NCAM及抗PCNA单抗对8例正常脑组织和51例胶质瘤进行免疫组化检测。结果　NCAM在正常脑组织中全部阳性表达,Ⅰ～Ⅱ级胶质瘤、Ⅲ级胶质瘤、Ⅳ级胶质瘤中NCAM阳性表达分别为95.2%(20/21)、52.6%(10/19)、18.2%(2/11),阳性细胞含量随肿瘤恶性程度升高反而减少;NCAM阳性表达量与PCNA指数呈负相关(r=-0.67,P<0.01)。结论　脑胶质瘤中NCAM表达程度与胶质瘤的临床病理分级和细胞增殖活性呈负相关,它的低表达是高度恶性胶质瘤的标志之一。     

Neural cell adhesion molecule is endocytosed via a clathrin-dependent pathway

Neural cell adhesion molecule (NCAM) constitutes a group of cell surface glycoproteins that regulate cell-cell interactions in the developing and adult brain. Endocytosis is a mechanism which dynamically controls the amount of cell surface NCAM expression and may involve the rapid changes occurring in NCAM expression under certain physiological or pathological conditions. However, the endocytic pathway of NCAM is presently unknown. Using astrocytes in culture and immunofluorescence we show that NCAM is internalized and that the immunolabelling presents a high degree of colocalization with clathrin, alpha-adaptin and transferrin, suggesting that NCAM is endocytosed by a clathrin-dependent pathway. Potassium depletion which disrupts clathrin-mediated endocytosis, inhibited internalization of NCAM. Electron microscopy and immunogold studies also demonstrate that the surface of clathrin-coated vesicles are also immunolabelled for both alpha-adaptin and PSA-NCAM, the highly sialylated isoform of NCAM. Furthermore, immunoprecipation studies demonstrate that NCAM is associated with both clathrin and alpha-adaptin, a component of adaptor complex AP-2, in brain, neurons and astrocytes. These findings indicate that NCAM is mainly endocytosed via clathrin-coated vesicles, suggesting a possible mechanism that may contribute to the rapid changes in NCAM expression at the cell surface.