Assessment of Neural Cell Adhesion Molecule (NCAM) in Autistic Serum and Postmortem Brain

Studies have identified structural abnormalities in areas of the autistic brain, with a pattern suggesting that a neurodevelopmental anomaly took place. Neural cell adhesion molecule (NCAM), which is involved in development of the central nervous system, was previously shown to be decreased in the serum of autistic individuals. In the present study, we measured NCAM protein in the sera from controls, patients with autism, siblings of autistic patients, and individuals with other neurologic disorders, but found no significant differences. We also measured NCAM protein in autistic postmortem brain samples and found the longest isoform, NCAM-180, to be significantly decreased. In addition, we investigated the mRNA expression of NCAM in these brain samples using cDNA microarrays and RT-PCR. Results show that NCAM mRNA levels are not altered in autism.     

Increased Intermale Aggression and Neuroendocrine Response in Mice Deficient for the Neural Cell Adhesion Molecule (NCAM)

Mice deficient for the neural cell adhesion molecule (NCAM) show morphological and behavioural abnormalities in the adult form, including a reduced size of the olfactory bulb, reduced exploratory behaviour, and deficits in spatial learning. Here we report increased aggressive behaviour of both homozygous (NCAM -/–) and heterozygous (NCAM +/–) male mutant mice towards an unfamiliar male intruding into their home cage. While plasma testosterone concentrations did not differ between genotypes before or after behavioural testing, corticosterone levels were higher in mutant residents than in wild-type (NCAM +/+) residents 30 min after encountering the intruder. Levels of c-fos mRNA, analysed to monitor neuronal activation, were similar in primary output structures of the olfactory bulb in NCAM-deficient and NCAM +/+ mice, but were increased in brain areas of the limbic system in both NCAM –/– and NCAM +/– mutant mice after the behavioural test. These results indicate that abnormalities in social behaviour correlate with enhanced neuronal activity in limbic brain areas and result in increased social stress in NCAM-deficient mice.     

Expression of the Highly Polysialylated Neural Cell Adhesion Molecule During Postnatal Myelination and Following Chemically Induced Demyelination of the Adult Mouse Spinal Cord

We have investigated the expression of the highly polysialylated neural cell adhesion molecule in the mouse spinal cord during postnatal myelination and in the adult after chemically induced demyelination. By double immunohistochemistry, using a monoclonal antibody (anti-Men B) which specifically recognizes polysialic acid (PSA) units on neural cell adhesion molecule (N-CAM), and an anti-myelin basic protein, a caudorostral gradient of expression of PSA-NCAM was observed at postnatal day 1 (P1), which was inversely related to the gradient of myelination. At P7, PSA-NCAM labelling decreased relative to P1. In white matter, this decrease was correlated with the progression of myelination. PSA-NCAM immunoreactivity persisted in as yet unmyelinated structures, i.e. the corticospinal tract, the dorsomedial part of the ventral funiculus and the lateral funiculi, and decreased with the onset of myelination of these structures at P15. In the adult, PSA-NCAM expression remained in discrete structures, i.e. Iaminae I and II of the dorsal horn and lamina X around the central canal. The ependymal cells and the astrocyte endfeet under the meninges were also labelled. In addition, PSA-NCAM expression was reinduced on various cells and structures after lysolecithin-induced demyelination of the adult mouse spinal cord. At early times after demyelination, PSA-NCAM was expressed on glial cells around the lesion but also at a distance from this zone. Seven days after injection, cellular PSA-NCAM expression was found around but also within the lesion. This expression was totally abolished 15 days after injection. Double immunohistochemistry for PSA and cell-specific markers showed that the cells which expressed PSA-NCAM after demyelination were oligodendrocyte precursors, reactive astrocytes and Schwann cells. PSA-NCAM re-expression on all cell types was transient and ceased when myelin repair was accomplished. The spatial and temporal regulation of PSA-NCAM expression during development and after demyelination suggests a role for PSA-NCAM in glial plasticity during the myelination and remyelination processes.     

神经细胞黏附分子在正常老化大鼠黑质致密部多巴胺能神经细胞的表达

目的：观察神经细胞黏附分子（NCAM）和酪氨酸羟化酶（TH）在大鼠黑质致密部（SNc）多巴胺（DA）能神经细胞表达的变化,探讨其发生机制。方法：健康雄性SD大鼠24只,分为成年组（4—5月龄）和老年组（≥24月龄）,取中脑黑质,分别进行TH和NCAM的免疫组织化学染色及免疫印迹检测蛋白表达,显微镜下计数免疫组化染色阳性神经细胞,灰度分析电泳条带。结果：SNc的DA能神经细胞几乎都表达NCAM,老年大鼠SNcTH阳性细胞总数及蛋白量均无减少（P〉0．05）,但尾侧段TH阳性细胞减少（P〈0．05）;NCAM在阳性细胞总数及蛋白量均减少（P〈0．05）,但各段阳性细胞的减少无统计学意义（P〉0．05）。结论：正常老化后大鼠的SNcDA能神经细胞DA合成降低,并且NCAM可能参与了这种DA的合成下降。     

Expression of the Neural Recognition Molecule L1 by Cultured Neural Cells is influenced by K+ and the Glutamate Receptor Agonist NMDA

To investigate the influence of neuronal activity on the expression of neural recognition molecules, cultures of neural cell lines and dissociated cells of early postnatal mouse cerebellum were maintained in the presence of elevated concentrations of K+ and the glutamate agonist N-methyl-d-aspartate (NMDA). Levels of expression of the neural adhesion molecules L1 and N-CAM at the cell surface were measured by an enzyme-linked immunosorbent assay. Expression of L1 was up-regulated in neuroblastoma N2A cells after 1 day of maintenance in 40 and 60 mM K+, but not in phaeochromocytoma PC12 cells. Expression levels of N-CAM and antigens recognized by the monoclonal antibody A2B5 or by polyclonal antibodies to crude membrane fractions of liver were not significantly altered by elevated K+ concentrations in these two cell lines. In monolayer cultures of early postnatal mouse cerebellum, an increase of 60% in expression of L1, but not N-CAM or A2B5, was seen at 20 and 40 mM K+. This increase in L1 expression was specifically inhibitable by the Ca2+ channel blocker nicardipine. NMDA at a concentration of 100 microM increased levels of L1, but not of N-CAM. This increase was inhibitable by the NMDA antagonists 2-amino-5-phosphonovalerate and MK-801, but not significantly by the kainate/quisqualate antagonist 6-cyano-7-nitroquinoxaline-2,3-dione. The increase in L1 expression at higher K+ concentrations was not inhibitable by the NMDA antagonists, indicating that the K+-mediated increase in L1 expression is not due to release of glutamate by cerebellar neurons. These observations indicate that compounds influencing neuronal membrane properties, and thus neuronal excitability, are capable of regulating the expression of L1. In a more general context, these findings suggest that previously observed changes in synaptic connectivity in situ, resulting from activity-dependent fine tuning of neuronal morphology, may be mediated by alterations in the expression of recognition molecules.     

Molecular Association of the Neural Adhesion Molecules L1 and N-CAM in the Surface Membrane of Neuroblastoma Cells is Shown by Chemical Cross-linking

The two neural cell adhesion molecules L1 and N-CAM could be shown to be associated in the surface membrane of cultured neuroblastoma cells by chemical cross-linking with 3,3'-dithiobis(sulphosuccinimidyl-propionate) and subsequent immunopurification and precipitation using antibodies to L1 and N-CAM. Glycoproteins recognized in neuroblastoma cells by antibodies to mouse liver membranes were not chemically cross-linked to L1 or N-CAM. These observations suggest that a molecular association between the two molecules may be the basis for their functional cooperativity (Kadmon et al., 1990a,b, J. Cell Biol., 110, 193-208; 209-218).     

L1 Cell Adhesion Molecule is Expressed by Noradrenergic but not Adrenergic Chromaffin Cells: A Possible Major Role for L1 in Adrenal Medullary Design

The adrenal medulla of higher animals is constituted of homotypic groups of chromaffin cells secreting either adrenalin or noradrenalin. Since not all chromaffin cells are individually innervated by fibres of the splanchnic nerve, this tissue characteristic is crucial to the physiological function of the gland. In an attempt to analyse differences between these chromaffin cell types which might underlie the establishment of this tissue pattern, we examined the expression of the adhesion molecule L1 in this gland by immunocytochemistry at the optical and ultrastructural levels in rats. L1, an adhesion molecule abundant in the central nervous system, was found to be present in the adrenal medulla of adults; it was strongly expressed on innervating axons and their surrounding Schwann cells and also on a subpopulation of chromaffin cells. The nature of these chromaffin cells was examined by immunocytochemistry using antibodies against the catecholamine-synthesizing enzyme phenylethanolamine N -methyltransferase (PNMT), which are capable of distinguishing between adrenergic and noradrenergic cells. Immunofluorescence labelling of sequential frozen sections demonstrated that chromaffin cells which express L1 do not express PNMT; conversely, L1 was not detected in any chromaffin cells expressing PNMT. Ultrastructural immunocytochemistry confirmed the existence of two non-overlapping populations of chromaffin cells. It is concluded that, in the adrenal medulla, noradrenergic but not adrenergic chromaffin cells express this adhesion molecule. These data, together with our previous observations that all chromaffin cells express the neural cell adhesion molecule, NCAM, suggest that L1, in cooperation with NCAM, could be responsible for the association of noradrenergic cells in the form of homotypic aggregates segregated from groups of adrenergic cells within the adrenal medulla.     

Lateral Mobility of the Cell Adhesion Molecule L1 Within the Surface Membrane of Morphologically Undifferentiated and Differentiated Neuroblastoma Cells

Lateral mobility and localization in the surface membrane of the adhesion molecule L1 was studied in morphologically undifferentiated and differentiated neuroblastoma cells to gain insight into its possible association with the different molecular forms of N-CAM. In undifferentiated cells, the fraction of mobile L1 molecules is high and similar to that of N-CAM 140. Upon long-term morphological differentiation, the fraction of mobile L1 molecules is reduced by a factor of three and is similar to that of N-CAM 180, the predominant molecular form of N-CAM in differentiated neuroblastoma cells. Comparable to N-CAM 180, L1 is also preferentially accumulated at contact sites between these cells as seen by indirect immunofluorescence. These observations raise the question of whether at least part of the L1 molecules may be directly or indirectly (e.g. via N-CAM 180) linked to the cytoskeleton, thus stabilizing cell contacts between differentiated cells.     

进展性缺血性脑卒中患者细胞因子及黏附分子的研究

目的探讨白细胞介素-1(IL-1)I、L-6、肿瘤坏死因子(TNF)、可溶性细胞间黏附分子-1(sICAM-1)和血管细胞黏附分子-1(sVCAM-1)在进展性缺血性脑卒中发病中的作用。方法收集108例首次发病且于起病24 h内入院的脑梗死患者作为研究对象,检测其外周血IL-1I、L-6、TNF、sVCAM-1、sICAM-1水平。进展性缺血性脑卒中的诊断根据神经功能缺损(SSS)评分判断。结果108例脑梗死患者中有29例(26.9%)于7 d内发展为进展性脑卒中,其入院时外周血IL-1I、L-6、TNFs、VCAM-1s、ICAM-1水平明显高于无进展的脑梗死患者,且皮层梗死患者外周血IL-1I、L-6、TNF、sVCAM-1、sICAM-1水平高于皮层下梗死患者(P<0.01)。经Logistic回归分析显示,IL-1I、L-6、TNF、sVCAM-1s、ICAM-1是进展性缺血性脑卒中的独立危险因素,且其外周血浓度与体温、血糖、纤维蛋白原浓度以及梗死灶大小呈正相关。结论细胞因子及黏附分子不仅参与脑梗死后急性期反应,而且其外周血水平增高与进展性缺血性脑卒中密切相关,可能在进展性缺血性脑卒中的发病过程中具有重要作用。     

The Adhesion Molecule on Glia (AMOG) Is Widely Expressed by Astrocytes in Developing and Adult Mouse Brain

Adhesion molecule on glia (AMOG) is a 45 - 50 kD cell surface glycoprotein structurally similar to the Na, K-ATPase beta-subunit and associated with the catalytic subunit of this enzyme. Previous immunofluorescence results had suggested that AMOG is transiently expressed on Bergmann glia during mouse cerebellar development, and antibody-inhibition results have implicated it in the migration of granule neurons. We report that, while AMOG mRNA is detected in Bergmann glia during the migratory period, this astrocyte derivative continues to express AMOG mRNA at similar levels in adult mice suggesting a functional role for AMOG in adulthood. Evidence from RNA and protein blot analyses that AMOG is present before birth, increasing about ten fold in adult mouse brain and cerebellum is also provided. RNA blot analysis of astrocyte-enriched cell populations and in situ hybridization results show that astrocytes synthesize AMOG mRNA in all regions of the developing and adult brain. In the adult, AMOG mRNA is more abundant in grey than white matter and, among grey matter regions, highest in cerebellar cortex. These results indicate a relationship between density of neuronal elements and AMOG expression. It is further speculated that AMOG is part of a Na,K-ATPase complex expressed preferentially by astrocytes in mouse brain.     

Evidence for Cis Interaction and Cooperative Signalling by the Heat-stable Antigen Nectadrin (murine CD24) and the Cell Adhesion Molecule L1 in Neurons

L1 is a transmembranal hornophilic cell adhesion molecule of the immunoglobulin superfamily expressed by neural and lymphoid cells. The heat-stable antigen (HSA, murine CD24) nectadrin is a highly and heterogeneously glycosylated glycophosphatidylinositol-linked differentiation antigen of haematopoietic and neural cells. L1 and nectadrin have been shown to mediate cell adhesion and intracellular Ca²⁺ signals in neurons and B lymphoblasts, respectively. Here we show that nectadrin is co-expressed with L1 in murine cerebellar granule cell neurons and neuroblastoma N2A cells. Purified nectadrin bound to L1 with an apparent binding ratio of five nectadrin molecules to one L1 molecule at saturation. Binding between nectadrin and purified N-CAM was not observed. In co-capping experiments nectadrin co-redistributed with L1 and N-CAM. Since in these cells N-CAM and L1 cohere by cis binding nectadrin appears to join the L1-N-CAM complex through binding to L1. Antibodies to each L1 and nectadrin evoked small increases in the intracellular Ca²⁺ concentration. However, when both antibodies were added together or in tandem to the cells, a strong intracellular Ca²⁺ signal was measured that was at least 6- and 10-fold stronger than the signal separately induced by L1 and nectadrin antibodies respectively. Such a cooperative effect was not observed in B lymphoblasts, using the same antibodies, or in neurons, using a combination of L1 and Thy-1 antibodies. Both the weak Ca²⁺ signal mediated by L1 alone and the enhanced signal jointly triggered by antibodies to L1 and nectadrin were inhibited by phorbol 12-myristate 13-acetate and were not significantly affected by Ni²⁺ and Cd²⁺ cations, suggesting that they are related to one another and involve recruitment of intracellular Ca²⁺. Nectadrin therefore appears to join a functional complex of neuronal adhesion molecules and to potentiate the signal transduction pathway of L1, possibly in response to neuron-neuron contact formation.     

Expression of four immunoglobulin superfamily adhesion molecules (L1, Nr-CAM/Bravo,neurofascin/ABGP,and N-CAM)in the developing mouse spinal cord

To identify cell adhesion molecules (CAMs) expressed by mammalian motoneurons, we applied the polymerase chain reaction to a murine motor neuron-like cell line, NSC-34. Using primers derived from a group of Ll-related CAMs, we cloned two alternatively spliced forms of mouse L1, which differ by a 12-base-pair insert, plus putative murine orthologs of the chicken cell adhesion molecules Nr-CAM/Bravo and neurofascin. All four mRNAs are expressed in NSC-34 cells, but only neurofascin and the insert-minus form of Ll are expressed in its neuroblastoma parent, N18TG2. Analysis of RNA in neonatal tissues reveals expression largely restricted to the brain and spinal cord. In situ hybridization histochemistry of spinal cord shows that motoneurons express L1, Nr-CAM, and neurofascin as well as N-CAM. Ll and N-CAM RNAs are detected throughout the period studied (from embryonic day,[E]11 to postnatal day [P]28), whereas Nr-CAM is expressed only at early ages (<E15) and neurofascin is predominantly expressed postnatally. Moreover, each CAM is expressed by distinct subsets of neighboring cells and at distinct times. For example, Nr-CAM mRNA is present in floor plate cells of embryonic spinal cord, whereas neurofascin is expressed by a subset of glia postnatally. Finally, we show that each CAM has a distinct spatiotemporal pattern of expression in dorsal root ganglia. © 1995 Wiley-Liss, Inc.     

Glial contribution to seizure: K activation of (Na, K)-ATPase in bulk isolated glial cells and synaptosomes of epileptogenic cortex

Glial and neuronal (Na⁺, K⁺)-ATPase have dissimilar apparent affinities for K⁺ ions. Glial (Na⁺, K⁺)-ATPase is maximally activated by 20 mM K⁺ while neuronal (Na⁺, K⁺)-ATPase is maximally stimulated by 3–5 mM K⁺. Because this glial property may play an important role in the clearance of [K⁺]₀ during seizures, we investigated the K⁺ activation of (Na⁺, K⁺)-ATPase within bulk isolated glial cells and synaptosomes isolated from epileptogenic brains. The primary focus (F), the homolateral brain area around the focus (PF) and the mirror (M) or secondary focus induced by freezing lesions were studied.Results show that both glial and synaptosomal enzyme activities in the epileptogenic state change in comparison with controls, i.e. sham-operated cats. In F and M., glial enzyme decreased reaction velocities between 3 and 18 mM K⁺. In PF, maximum velocities increased in glial (Na⁺, K⁺)-ATPase. These results indicate that in actively firing epileptogenic tissue (F, M), glial (Na⁺, K⁺)-ATPase decreased rate reactions while the catalytic activity was increased in cortical tissues surrounding the focus. These phenomena appeared early, i.e. 1–3 days after production of the freezing lesion, and was associated with a sharp decrease in absolute levels of enzyme activity.Synaptosomal (Na⁺, K⁺)-ATPase from controls always exhibited a saturation curve at 3–6 mM K⁺. Synaptosomal enzyme activities in the primary (F) lesion increased slightly 24 h after lesion production, then progressively decreased 3 days after lesion production. No significant changes were seen in M and PF.     

Effects of the HIV-1 Envelope Glycoprotein gp120 in Cerebellar Cultures. [Ca2+]i Increases in a Glial Cell Subpopulation

The various types of cells present in cultures prepared from the postnatal rat cerebellum, identified by their gross morphology and immunocytochemistry, were loaded with the specific dye fura-2 and analysed individually for [Ca²⁺]_i changes induced by the HIV-1 envelope glycoprotein gp120 and a variety of other treatments. In granule neurons [Ca²⁺]_i increases were induced by high KCl and glutamate (mainly through the NMDA receptor) while in type-1 astrocytes this effect was observed after serotonin, carbachol and also quisqualate. In contrast, administration of gp120 was always without effect in these cells. Type-2 astrocytes (an arborized cell type responsive to agonists targeted to the glutamatergic AMPA and cholinergic receptors) were also most often unresponsive to the viral glycoprotein. However, among the cells exhibiting the arborized phenotype, a subpopulation (-13%) responded to gp120 with conspicuous [Ca²⁺]_i increases sustained by both release from intracellular stores and influx across the plasma membrane. These responses to the viral protein did not involve activation of either voltage-gated Ca²⁺ channels or glutamatergic receptors. Although not yet conclusively identified by specific cytochemical markers, the gp120-responsive cells resemble type-2 astrocytes and differ from neurons and type-1 astrocytes both in gross phenotype and in a number of receptor/channel properties: positivity to AMPA and cholinergic agonists; negativity to NMDA, serotonin and high KCl. From these results it is concluded that a subpopulation of glial cells is affected by gp120. The role of these cells in HIV brain infection and damage requires further studies to be precisely established.     

Neural (N-) cadherin,a synaptic adhesion molecule,is induced in hippocampal mossy fiber axonal sprouts by seizure

Aberrant mossy fiber sprouting and synaptic reorganization are plastic responses in human temporal lobe epilepsy, and in pilocarpine-induced epilepsy in rodents. Although the morphological features of the hippocampal epileptic reaction have been well documented, the molecular mechanisms underlying these structural changes are not understood. The classic cadherins, calcium-dependent cell adhesion molecules, are known to function in development in neurite outgrowth, synapse formation, and stabilization. In pilocarpine-induced status epilepticus, the expression of N-cadherin mRNA was sharply upregulated and reached a maximum level (1- to 2.5-fold) at 1- to 4 weeks postseizure in the granule cell layer and the pyramidal cell layer of CA3. N-cadherin protein was correspondingly increased and became concentrated in the inner molecular layer of the dentate gyrus, consistent with the position of mossy fiber axonal sprouts. Moreover, N-cadherin labeling was punctate; colocalized with definitive synaptic markers, and partially localized on polysialated forms of neural cell adhesion molecule (PSA-NCAM)-positive dendrites of granule cells in the inner molecular layer. Our findings show that N-cadherin is likely to be a key factor in responsive synaptogenesis following status epilepticus, where it functions as a mediator of de novo synapse formation.     

Association of the Neuronal Cell Adhesion Molecule (NrCAM) Gene Variants with Personality Traits and Addictive Symptoms in Methamphetamine Use Disorder

Objective

1) To investigate the relationship between NrCAM polymorphisms and methamphetamine abuse in an ethnically homogenous Korean population. 2) To further support our findings by investigating the association among NrCAM gene variants, certain personality traits, and addictive symptoms of methamphetamine abusers.

Methods

Thirty-seven male methamphetamine abusers (age=43.3±7.8) and30 non-users (16 men, 14 women; age=59.8±10.4) were recruited. Ten single nucleotide polymorphisms (SNPs) in the NrCAM gene were assayed to compare genotype distributions between the 2 groups. Personality characteristics were measured using the Temperament and Character Inventory (TCI) and the NEO Personality Inventory, Revised (NEO PI-R). Addictive symptoms were assessed using the Diagnostic Interview for Genetic Studies (DIGS) and reviews of the subject''s medical records.

Results

Among the 10 SNPs in the NrCAM gene, the frequency of the TA genotype at rs1990162 was significantly lower in methamphetamine abusers compared to non-users (p=0.042). In the 3 NrCAM gene SNPs (rs381318, rs2072546, and rs6954366), the distribution of genotypes and alleles were significantly associated with some traits in the TCI and NEO PI-R. Genotypes and alleles at 5 gene SNPs (rs2142325, rs381318, rs1269621, rs1269634, and rs1990162) were associated with certain addictive symptom dimensions in the patients.

Conclusion

These findings support the idea that NrCAM is associated with genetic susceptibility of methamphetamine abuse and is also associated with certain personality characteristics that may increase disturbed addictive behavior.     

The Role of the PI3K Pathway in the Regeneration of the Damaged Brain by Neural Stem Cells after Cerebral Infarction

Neurologic deficits resulting from stroke remain largely intractable, which has prompted thousands of studies aimed at developing methods for treating these neurologic sequelae. Endogenous neurogenesis is also known to occur after brain damage, including that due to cerebral infarction. Focusing on this process may provide a solution for treating neurologic deficits caused by cerebral infarction. The phosphatidylinositol-3-kinase (PI3K) pathway is known to play important roles in cell survival, and many studies have focused on use of the PI3K pathway to treat brain injury after stroke. Furthermore, since the PI3K pathway may also play key roles in the physiology of neural stem cells (NSCs), eliciting the appropriate activation of the PI3K pathway in NSCs may help to improve the sequelae of cerebral infarction. This review describes the PI3K pathway, its roles in the brain and NSCs after cerebral infarction, and the therapeutic possibility of activating the pathway to improve neurologic deficits after cerebral infarction.     

Schwann cell differentiation in Charcot-Marie-Tooth disease type 1A (CMT1A): normal number of myelinating Schwann cells in young CMT1A patients and neural cell adhesion molecule expression in onion bulbs

Charcot-Marie-Tooth disease type 1A (CMT1A) is a common hereditary demyelinating neuropathy caused by a duplication of the gene for the myelin protein PMP22, resulting in overexpression of PMP22 in young patients. Although genetically well defined, the pathogenesis of the hereditary demyelinating neuropathy CMT1A is still unclear. Homology of PMP22 cDNA to the growth arrest-specific gene gas3 and experiments in vitro showing decreased proliferation in PMP22-overexpressing Schwann cells suggest a role of PMP22 in Schwann cell differentiation. Furthermore, overexpression of PMP22 in fibroblasts induces programmed cell death. In this report we applied morphometrical methods using electron micrographs and immunohistochemistry to further characterise Schwann cells in CMT1A nerve biopsy samples from CMT1A patients. We show that the total number of PMP22-expressing Schwann cells, i.e. Schwann cells that are in a 1:1 relationship with axons, was not reduced in sural nerve biopsy samples from six young CMT1A patients. We excluded non-specific secondary Schwann cell proliferation. Thus, in young CMT1A patients with increased PMP22 overexpression there seems to be no evidence for altered initial Schwann cell proliferation in achieving a 1:1 relationship to axons prior to the process of de- and remyelination. Further, using electron microscopy we found no evidence for apoptosis of Schwann cells in CMT1A . However, we provide additional support for an abnormal Schwann cell phenotype in CMT1A by showing the expression of neural cell adhesion molecule immunoreactivity in onion bulbs. Thus, the role of PMP22 in cell growth and differentiation does not lead to an altered number of myelinating Schwann cells but to altered Schwann cell differentiation in CMT1A. Received: 23 September 1996 / Revised: 28 November 1996, 31 January 1997, 2 April 1997 / Accepted: 3 April 1997     

Genetic variants in Cell Adhesion Molecule 1 (CADM1): A validation study of a novel endothelial cell venous thrombosis risk factor

Introduction

In a protein C deficient family, we recently identified a candidate gene, CADM1, which interacted with protein C deficiency in increasing the risk of venous thrombosis (VT). This study aimed to determine whether CADM1 variants also interact with protein C pathway abnormalities in increasing VT risk outside this family.

Materials and methods

We genotyped over 300 CADM1 variants in the population-based MEGA case-control study. We compared VT risks between cases with low protein C activity (n = 194), low protein S levels (n = 23), high factor VIII activity (n = 165) or factor V Leiden carriers (n = 580), and all 4004 controls. Positive associations were repeated in all 3496 cases and 4004 controls.

Results

We found 22 variants which were associated with VT in one of the protein C pathway risk groups. After mutual adjustment, six variants remained associated with VT. The strongest evidence was found for rs220842 and rs11608105. For rs220842, the odds ratio (OR) for VT was 3.2 (95% CI 1.2-9.0) for cases with high factor VIII activity compared with controls. In addition, this variant was associated with an increased risk of VT in the overall study population (OR: 1.5, 95% CI 1.0-2.2). The other variant, rs11608105, was not associated with VT in the overall study population (OR: 1.0, 95% CI 0.8-1.1), but showed a strong effect on VT risk (OR: 21, 95% CI 5.1-88) when combined with low protein C or S levels.

Conclusions

In a population-based association study, we confirm a role for CADM1 variants in increasing the risk of VT by interaction with protein C pathway abnormalities.     

The Critical Period for Repair of CNS of Neonatal Opossum (Monodelphis domestica) in Culture: Correlation with Development of Glial Cells, Myelin and Growth-inhibitory Molecules

A comparison was made of neurite growth across spinal cord lesions in the isolated central nervous system (CMS) of newborn opossums (Monodelphis domestica) at various stages of development. The aim was to define the critical period at which growth after injury ceases to occur, with emphasis on growth-inhibitory proteins, myelin and glial cells. In postnatal opossums 3–6 days old (P3–6), repair was observed 5 days after lesions were made in culture at the cervical level (C7) by crushing with forceps. Through-conduction of action potentials was re-established and axons stained by Oil grew into and beyond the crush. In a series of 66 animals 29 showed repair. In 28 animals at P11–12 with comparable lesions repair was observed in five preparations. At P13–14, the CMS was still viable in culture, but none of the 25 preparations examined showed any axonal growth into the crush or conduction through it. The rostra-caudal gradient of development permitted lesions to be made in mature cervical and immature lumbar regions of P11–12 spinal cord. Growth across crushes occurred in lumbar but not in cervical segments of the same preparation. The development of glial cells and myelin was assessed by electron microscopy and by staining with specific antibodies (Rip-1 and myelin-associated glycoprotein) in cervical segments of neonatal P6–14 opossums. At P8, oligodendrocytes and thin myelin sheaths started to appear followed at P9 by astrocytes stained with antibody against glial fibrillary acidic protein. By P14, astrocytes, oligodendrocytes and well-developed myelin sheaths were abundant. The cervical crush sites of P12 cords contained occasional astrocytes but no oligodendrocytes. Specific antibodies (IN-1) to neurite growth-inhibiting proteins (NI-35/250) associated with oligodendrocytes and myelin in the rat CNS cross-reacted with opossum proteins. Assays using the spreading of 3T3 fibroblasts and IN-1 showed that by P7 inhibitory proteins became apparent, particularly in the hindbrain and cervical spinal cord. The concentrations of NI-35/250 thereafter increased and became abundant in the adult opossum. Our finding of a well-defined critical period, encompassing only 5 days, in CNS preparations that can be maintained in culture offers advantages for analysing mechanisms that promote or prevent CNS repair.