Subacute sclerosing panencephalitis (SSPE)

Subacute sclerosing panencephalitis is a rare progressive neurological disorder of childhood and early adolescence caused by persistent measles virus. The diagnosis is based upon characteristic clinical manifestations, periodic EEG discharges, raised antibody titre against measles/SSPE in the plasma and cerebrospinal fluid and increase of gamma-globulins in the cerebrospinal fluid. Histopathological examination shows neuronal loss, astrogliosis, demyelination, infiltration of inflammatory cells, and intranuclear inclusions in neurons, oligodendrocytes and astrocytes. In most cases nucleocapsids are detected by electron microscopy. Although treatment is still undetermined, combination of intrathecal high-dose interferon-alpha and intravenous ribavirin administered at an early stage of SSPE seems to be effective.     

Electron microscopic finding of eccrine sweat gland epithelial cells in a patient with Krabbe disease

A 13 month old boy was found to have severely reduced β-galactocerebrosidase activity suggesting infantile Krabbe disease. Clinically, the patient showed a progressive neurological deterioration with white-matter disease on radiological study. Axillary skin biopsy was performed to support the diagnosis. On electron microscopy, needle-like inclusions, which are the typical finding seen in the cytoplasm of astrocytes and Schwann cells in the classic infantile form, were present in eccrine sweat gland epithelial cells. This method is useful for diagnosis when nerve biopsy and biochemical analysis are not readily available.     

Brain Defects in Infants with Potter Syndrome (Oligohydramnios Sequence)

Defects of neuronal migration were detected in the brains of five unrelated infants with Potter syndrome (oligohydramnios sequence). These consisted of abnormal lamination of cerebral cortex, white matter heterotopias, and meningeal and molecular zone neuronal-glial ectopias. Besides, various other brain anomalies were sometimes found. They comprised one or more of the followings: abnormal gyration patterns (gyral fusion, cerebellar microgyria), cerebellar granule and Purkinje cell heterotopias, brain stem heterotopias, adysplasia of basal ganglia, gliosis, mineralization, and hydrocephalus. Detailed investigations, using standard neuropathologic stains, immunohistochemical and Golgi methods, and a new electron microscopic histochemical technique that we applied to study the developing human brain, suggest that migration defects of neurons are caused by an abnormality in their glial guides, the radial glial fibers, during the period of cortical histogenesis. We hypothesize that abnormally and precociously induced radial glial transformation into astrocytes is the pathogenic mechanism for the defects in neuronal migration. The etiological factor(s) that precipitates such abnormal glial transformation seems to be heterogeneous. Its relation to the Potter anomaly is discussed.     

The germinative zone produces the most cortical astrocytes after neuronal migration in the developing mammalian brain.

The origin of astrocytes of the mouse neocortex during the fetal and early postnatal periods as determined by immunocytological, autoradiographic, electron microscopic and antimitotic methods is described. Most astrocytes destined for the white matter and the infragranular cortical layers are derived from the transformation of radial glial cells between P0 and P10 with an inside-out pattern. This cell metamorphosis is not directly preceded by mitosis and involves the activation of the radial glial lysosomal apparatus. In opposition to recent hypotheses, our findings suggest that most astrocytes destined for the supragranular cortical layers are produced in the germinative zone after the migration of the infragranular neurons and themselves migrate afterwards to the upper cortex between E16 and the first postnatal days. These astrocytes do not display an intermediate stage of the radial glial cell and do not participate in the pattern of appearance of the deeper astrocytes. This second step of astrocytogenesis is a condition for normal cytoarchitectonic development and the maintenance of the supragranular layers, since the deprivation of the astrocytic equipment of the supragranular layers by an antimitotic drug drastically reduces the number of supragranular neurons.     

胰岛素样生长因子-1对新生大鼠神经细胞氧化损伤保护作用的研究

目的 探讨氧化应激状态下,胰岛素样生长因子-1（IGF-1）对新生大鼠神经细胞氧化损伤的保护作用。方法 原代培养新生大鼠大脑皮层神经元、少突胶质细胞及星形胶质细胞,不同浓度H2O2（0~60 μM）诱导氧化应激细胞模型,LDH 法检测各种神经细胞损伤程度,MTT 法检测各种神经细胞活力;不同浓度H2O2（0~80 μM）诱导氧化应激神经元细胞模型,Western blot 检测IGF-1（25 ng/mL）施加前后神经元细胞内Akt 的磷酸化水平。结果 与未经H2O2 处理组相比,不同浓度H2O2 处理细胞24 h 后,大脑皮层神经元、少突胶质细胞及星形胶质细胞损伤程度均有升高、细胞活力均有降低;但神经元变化更为显著,与少突胶质细胞和星形胶质细胞相比,差异有统计学意义（均P2O2 处理大脑皮层神经元5 min 后,相比于未经H2O2 处理组,可见Akt 磷酸化水平呈H2O2 浓度依赖性降低（均P2O2 导致的神经元细胞Akt 磷酸化,与未经H2O2 处理组比较,差异无统计学意义（P>0.05）,但是对高浓度H2O2 导致的Akt 磷酸化作用无明显效果,其磷酸化水平均低于未经H2O2 处理组（均P2O2 处理1 h 后,再加入25 ng/mL 的IGF-1,IGF-1 处理前后Akt 磷酸化水平均已恢复至未经H2O2 处理时的水平（均P>0.05）。结论 大脑皮层神经元对H2O2 诱导的氧化应激损伤较其他神经细胞敏感;IGF-1 对皮层神经元氧化应激损伤具有保护作用。     

Development of Cerebellar Hypoplasia in Jaundiced Gunn Rats: A Morphological Study

Abstract The developmental process of cerebellar hypoplasia was investigated in homozygous (j/j) Gunn rats (born from heterozygous (+/j) mothers) with hereditary hyperbilirubinemia by light microscopy of Epon-embedded sections and Golgi preparations, electron microscopy and immunofluorescence with anti-SlOO protein immune serum. No abnormalities were found in the cerebellar cortex at postnatal day 1. But at day 3, some Purkinje cells contained whorled membranous inclusions, enlarged Golgi cisternae and vesicles, and/or dilated endoplasmic reticulum. Membranous inclusions filled the cytoplasm of a large number of Purkinje cells at day 10. These severely damaged Purkinje cells died and disappeared by day 30. The remaining Purkinje cells contained only a few membranous inclusions at day 30. At the adult stage, membranous inclusions were no longer detected in the remaining Purkinje cells. In these Purkinje cells, the dendrites were poorly developed and primitive synaptic junctions with parallel fibers were occasionally found on the dendritic shafts even at the adult stage. The perisomatic processes were often present on the remaining Purkinje cell soma from day 18 to the adult stage and were in synaptic contact with climbing fibers. The external granular layer was thinner in j/j than in +/j rats without hyperbilirubinemia at days 12 and 15 and disappeared at day 18 in j/j rats, i.e., earlier in j/j than in +/j rats. The molecular layer was thinner at day 10 and became slightly thicker thereafter. Cells in the molecular layer were scarce after day 12. The internal granular layer was thinner after day 12. An insufficient number of cells in the internal granular layer was apparent after day 12. Bergmann glia cells were hypertrophied after day 10 and filledthe spaces where Purkinje cells had disappeared. Subsequently, Bergmann fibers and astrocytes in the internal granular layer were hypertrophied after day 15. Thus, in the developmental process of cerebellar hypoplasia in j/j Gunn rats, Purkinje cell damage preceded the histogenetic abnormalities of the cortex including astrocytic changes.     

The usefulness of bone marrow aspiration in the diagnosis of Niemann-Pick disease type C in infantile liver disease.

BACKGROUND: Niemann-Pick disease type C (NPC) is a fatal, autosomal recessive lysosomal storage disease which may present in infancy with cholestatic jaundice and/or hepatosplenomegaly. In cholestatic patients with splenomegaly, a bone marrow aspirate has been advocated as a relatively accessible tissue to demonstrate storage phenomena. Typically in patients with NPC, macrophages with abnormal cholesterol storage, so called foam cells, can be detected in the bone marrow. AIM: To review our experience of bone marrow aspiration in children with NPC presenting with infantile liver disease. METHODS: A retrospective analysis of 11 consecutive children (8 males) from Birmingham Children's Hospital with NPC presenting with infantile liver disease was undertaken. The diagnosis of NPC was confirmed in all cases by demonstrating undetectable or low rates of cholesterol esterification and positive filipin staining for free cholesterol in cultured fibroblasts. RESULTS: The median age at presentation was 1.5 months (range 0.5-10). Bone marrow aspirates showed storage cells in only 7/11 cases. Bone marrow aspirates which had storage cells were undertaken at a median age of 11 months while those with no storage cells were undertaken at median age 2.3 months. The overall sensitivity of bone marrow aspirates for detecting storage cells in children presenting with infantile liver disease was 64%; however, for children who had bone marrow aspirates in the first year of life it was only 57%. CONCLUSIONS: The sensitivity of bone marrow aspirate for the diagnosis of NPC disease in patients presenting with infantile liver disease was lower than previously reported. Where NPC is suspected clinically, definitive investigations should be undertaken promptly. There is a need to develop sensitive screening methods for NPC in children presenting with infantile liver disease.     

缺氧缺血对人类胚胎脑室管膜下区神经细胞亚群的影响

目的 研究缺氧缺血对妊娠中期人类胚胎脑室管膜下区(subventricular zone,SVZ)神经细胞亚群的影响.方法 取即刻解离的妊娠中期人类胚胎脑SVZ细胞,分缺氧缺血组(HI组)和对照组短时培养.以氧糖缺失法(OGD)制备缺氧缺血损伤细胞模型.培养前以细胞成活率评价损伤程度,培养后以细胞特异性标志蛋白nestin、MAP2、GFAP、PDGFRa及RCA120的抗体通过免疫荧光细胞化学法分别鉴定神经干细胞(NSCs)、神经元、星形胶质细胞、少突胶质祖细胞及小胶质细胞,比较其百分含量.结果 HI组的细胞成活率(63.41%±0.06%),明显低于正常组(98.9%±0.01%)(P＜0.001),短时培养后HI组中细胞亚群中含量最高的是GFAP(+)的星形胶质细胞56.48%±0.03%,其次为神经干细胞NSCs 22.47%±0.03%而PDGFRa(+)的少突胶质祖细胞含量最低;在对照组中最高则为MAP2(+)的神经元48.81%±0.03%,其次为GFAP(+)的星形胶质细胞32.31%±0.03%.含量最少的为小胶质细胞1.15%±0.01%.结论 妊娠中期人类胚胎脑SVZ含有NSCs、神经元、星形胶质细胞、少突胶质祖细胞和小胶质细胞,缺氧缺血对SVZ神经细胞损伤明显,不同细胞对缺氧缺血损伤的耐受性不同:NSCs、星形胶质细胞对缺氧缺血损伤的耐受性相对强于神经元、少突胶质祖细胞.     

Lipid peroxidation, caspase-3 immunoreactivity, and pyknosis in late-gestation fetal sheep brain after umbilical cord occlusion

Umbilical cord occlusion (UCO), a known risk factor for perinatal brain damage, causes severe fetal asphyxia leading to oxidative stress, lipid peroxidation, and cell death. We have determined the effects of two 10-min UCO on the distribution of the lipid peroxidation marker 4-hydroxynonenal (4-HNE) and the activated form of the apoptosis marker caspase-3 in the brains of late-gestation fetal sheep. UCO caused asphyxia, hypertension, and bradycardia, but these parameters normalized 2 h after the occlusion. At postmortem, 48 h after the second UCO there were significantly higher numbers of 4-HNE-positive cells in all layers of the hippocampus and cerebellum, the parietal cortex, substantia nigra, caudate nucleus, putamen, and thalamus compared with control brains. 4-HNE immunoreactivity was also found in white matter tracts of the subcallosal bundle, external medullary lamina, reticular thalamic nucleus, and cerebellar fiber tracts only in UCO brains. Double-labeling identified these cells as predominantly neurons and astrocytes, with oligodendrocytes showing lower levels of 4-HNE immunoreactivity. After UCO, the number of caspase-3-immunopositive cells was increased significantly in the hippocampal CA1, molecular layer and dentate gyrus, ventrolateral thalamic nucleus, substantia nigra, putamen, and cerebellar granular and molecular layers compared with controls. Double-labeling revealed caspase-3 immunoreactivity was mainly in neurons, and to lesser extent in astrocytes and oligodendrocytes. Pyknotic cell numbers were significantly increased in hippocampal CA1 and CA3, parietal cortex, caudate nucleus, putamen, and cerebellar Purkinje cells after UCO. These data indicate that brief asphyxia induces widespread lipid peroxidation involving all cell types of the fetal brain and apoptosis in both neurons and glia.     

Inflammatory mediators and neonatal brain damage

Inflammatory mediators are multifunctional cytokines that play important roles both in normal central nervous system (CNS) development and in the response of the brain to diverse forms of injury. Interleukin (IL)-1beta, tumor necrosis factor-alpha and IL-6 are among the best-characterized early-response cytokines. Recent data suggest that they may be synthesized and secreted by several CNS cell types, including microglia, astrocytes and neurons. Biological effects of these cytokines that could influence the progression of injury in the brain include stimulating the synthesis of other cytokines and neuronal injury mediators such as nitric oxide synthase, inducing leukocyte infiltration and the expression of adhesion molecules, influencing glial gene expression and damaging oligodendrocytes. In the immature brain, proinflammatory cytokines might lead to white matter damage during prenatal intrauterine infection and contribute to progressive neuronal damage in acute brain injury evoked by cerebral hypoxia-ischemia. Interrupting the proinflammatory cascade might limit the extent of irreversible injury.     

Maternal corticosteroid therapy and the fetal brain in experimental hyaline membrane disease

The possible acute deleterious effects of maternal glucocorticoid administration on the fetal nervous system and the pathologic significance of sudanophilic lipids in glial cells were studied in the premature pigtail monkey (macaca mulatta). At 72, 48, and 24 h before delivery at 135 +/- 1 d gestation, dams were treated with either 4 mg dexamethasone or saline. After delivery, respiratory function of each fetus was determined and supported. The animals were sacrificed at 3 h of age. Brain weights were similar in the two groups. Dark, shrunken, pyknotic neurons were present in the hippocampus of three treated and two control animals. Lipid-containing glial cells were present in all animals. Neither appeared to be related to steroid treatment or to the degree of respiratory distress. We conclude that short-term glucocorticoid therapy in doses analogous to those used in humans for the prevention of hyaline membrane disease does not result in acute neuronal damage. Sudanophilic lipid accumulation in glial cells is not always abnormal and must be distinguished from glial fatty metamorphosis.     

大鼠胚胎神经干细胞移植治疗类脊髓栓系综合征的实验研究

目的将胎鼠的神经干细胞移植到脊髓栓系综合征大鼠的病变脊髓中，观察治疗效果。方法从孕17d大鼠胚胎脊髓中分离、培养神经干细胞并诱导分化，通过免疫组化技术研究证实其特性。制作大鼠脊髓栓系模型，3d后将未分化的神经干细胞注人病变脊髓内，2个月后观察大鼠的运动功能，处死大鼠，研究移植后的干细胞在体内存活、迁移及分化情况。结果实验中分离、培养的神经干细胞能够被诱导分化成神经元、星形胶质细胞和少突胶质细胞。脊髓栓系大鼠移植干细胞后运动功能改善，神经元数目增多，显著好于未移植组。免疫组化方法证实移植后的干细胞在体内可以存活、迁移、分化成神经细胞。结论大鼠胚胎神经干细胞体内、体外均具有多向分化潜能，移植神经干细胞是治疗脊髓栓系神经变性的一种有效途径。     

Ultrastructural features of gaucher disease treated with enzyme replacement therapy presenting as mesenteric mass lesions

The classical ultrastructural features of Gaucher disease include large numbers of intracytoplasmic, membrane-bound lysosomal inclusions containing characteristic tubular structures on an electron-lucent background, representing the periodic acid schiff (PAS)--positive Gaucher cells identifiable on light microscopy. Following enzyme replacement therapy (ERT), many of the manifestations of the condition are ameliorated, but persistent mesenteric lymphadenopathy has been reported, the ultrastructural features of which previously have not been described. Two children, aged 4 and 8 years old, respectively, both presented with persistent abdominal lymphadenopathy whilst receiving ERT for Gaucher disease. Needle core biopsies were carried out, that demonstrated collections of macrophages and only scattered storage-type cells on light microscopy. PAS staining was negative in one case and only focally positive in the other. Electron microscopic examination, however, confirmed the cells represented macrophages, the cytoplasm of which contained scattered abnormal inclusions containing occasional twisted tubular structures of the type reported in classic Gaucher disease. ERT in Gaucher disease appears to reduce accumulation of the metabolic products at many sites. But for uncertain reasons, abdominal lymphadenopathy may occur containing macrophages that do not form granulomas or classic Gaucher cells on light microscopy. These probably represent incomplete clearance, incomplete/partial enzyme replacement, or possibly an unusual response to a relatively small amount of storage material.     

碱性成纤维细胞生长因子对胚胎大鼠神经干细胞增殖分化的影响

目的探讨分离、培养及鉴定胚胎大鼠神经干细胞(NSCs)方法，观察碱性成纤维细胞生长因子(bFGF)对NSCs增殖、分化的影响。方法从大鼠胚胎脑组织中分离出NSCs，用bRGF和胎牛血清诱导其增殖和分化，予BrdU以标记分裂细胞，采用免疫细胞化学鉴定NSCs和分化神经细胞。结果大鼠胚胎脑NSCs在无细胞因子和胎牛血清的培养基中无新生细胞形成，但能在bFGF和血清诱导下形成克隆，并产生nestin和BrdU阳性细胞，贴壁后分化为神经元和星形胶质细胞。结论该方法从大鼠胚胎大脑分离出的细胞具有NSCs特性，即自我更新和多向分化潜能;bFGF是NSCs增殖必须的丝裂原。     

Physically handicapped school leavers.

A 7 year old child perinatally infected with HIV who died from progressive muscular paralysis and central nervous respiratory failure is described. Cytomegalovirus (CMV) prophylaxis with a special intravenous CMV hyper-immunoglobulin had been successfully conducted for more than four years. Macroscopic and microscopic immunohistochemical examination of the spinal cord revealed a diffuse CMV infiltration of the entire myelon. CMV infected cells were identified as astrocytes, oligodendrocytes, neurons, macrophages, ependymal, endothelial, and Schwann cells. Other organs had no signs of CMV infection. Central nervous spinal CMV infection was most probably due to insufficient penetration of the blood-brain barrier by the CMV hyper-immunoglobulin. In suspicious cases early spinal magnetic resonance imaging (1.5 tesla) combined with an examination of urine and cerebrospinal fluid for CMV is recommended.     

Cytomegalovirus myelitis in perinatally acquired HIV

A 7 year old child perinatally infected with HIV who died from progressive muscular paralysis and central nervous respiratory failure is described. Cytomegalovirus (CMV) prophylaxis with a special intravenous CMV hyper-immunoglobulin had been successfully conducted for more than four years. Macroscopic and microscopic immunohistochemical examination of the spinal cord revealed a diffuse CMV infiltration of the entire myelon. CMV infected cells were identified as astrocytes, oligodendrocytes, neurons, macrophages, ependymal, endothelial, and Schwann cells. Other organs had no signs of CMV infection. Central nervous spinal CMV infection was most probably due to insufficient penetration of the blood-brain barrier by the CMV hyper-immunoglobulin. In suspicious cases early spinal magnetic resonance imaging (1.5 tesla) combined with an examination of urine and cerebrospinal fluid for CMV is recommended.     

EFFECTS OF ETHANOL ON GLIAL CELL PROLIFERATION: RELEVANCE TO THE FETAL ALCOHOL SYNDROME

Exposure to ethanol during pregnancy is detrimental to brain development. Individuals affected by the Fetal Alcohol Syndrome present a number of central nervous system dysfunctions including microencephaly and mental retardation. Studies on the mechanisms of ethanol's developmental neurotoxicity have focused on its interaction with neurons; however, emerging evidence is suggesting that ethanol can significantly affect glial cells as well. A number of in vitro studies have shown that ethanol can inhibit the proliferation of various glial cells (mostly primary astrocytes or astrocytoma cells) at relatively high concentrations (100-200 mM). On the other hand, proliferation induced by some, but not all mitogens, is inhabited by low concentrations (10-50 mM) of ethanol. These inhibitory effects of ethanol may contribute to its developmental neurotoxicity observed following in vivo exposure. Animal models have indeed shown that ethanol causes microencephaly when given during the brain growth spurt, a period of brain development characterized by astroglial proliferation and maturation.     

The usefulness of bone marrow aspiration in the diagnosis of Niemann-Pick disease type C in infantile liver disease

Background

Niemann–Pick disease type C (NPC) is a fatal, autosomal recessive lysosomal storage disease which may present in infancy with cholestatic jaundice and/or hepatosplenomegaly. In cholestatic patients with splenomegaly, a bone marrow aspirate has been advocated as a relatively accessible tissue to demonstrate storage phenomena. Typically in patients with NPC, macrophages with abnormal cholesterol storage, so called foam cells, can be detected in the bone marrow.

Aim

To review our experience of bone marrow aspiration in children with NPC presenting with infantile liver disease.

Methods

A retrospective analysis of 11 consecutive children (8 males) from Birmingham Children''s Hospital with NPC presenting with infantile liver disease was undertaken. The diagnosis of NPC was confirmed in all cases by demonstrating undetectable or low rates of cholesterol esterification and positive filipin staining for free cholesterol in cultured fibroblasts.

Results

The median age at presentation was 1.5 months (range 0.5–10). Bone marrow aspirates showed storage cells in only 7/11 cases. Bone marrow aspirates which had storage cells were undertaken at a median age of 11 months while those with no storage cells were undertaken at median age 2.3 months. The overall sensitivity of bone marrow aspirates for detecting storage cells in children presenting with infantile liver disease was 64%; however, for children who had bone marrow aspirates in the first year of life it was only 57%.

Conclusions

The sensitivity of bone marrow aspirate for the diagnosis of NPC disease in patients presenting with infantile liver disease was lower than previously reported. Where NPC is suspected clinically, definitive investigations should be undertaken promptly. There is a need to develop sensitive screening methods for NPC in children presenting with infantile liver disease.     

Ultrastructural features of nerve fascicles and basal lamina abnormalities in Hirschsprung's disease.

Although the pathogenesis of Hirschsprung's disease (HD) is not completely resolved, both the absence of nerve cells and the hypertrophy of nerve fascicles within the aganglionic colonic segment have been attributed to an abnormal intestinal microenvironment. Studies on animal models for HD revealed an altered ultrastructure of ingrowing nerve fascicles and abnormalities of basal laminae (BL). Therefore, the purpose of this study was to examine the ultrastructure of hypertrophied nerve fascicles in human HD with special reference to structural abnormalities of BL. Colonic specimens were obtained from patients with HD (n = 10) and controls (n = 5) and processed for electron-microscopical examination. Hypertrophied nerve fascicles were characterized by a prominent perineural sheath surrounded by large amounts of collagen bundles, a collagen-filled endoneurium, vasa nervorum and abundant glial cells of extraenteric ultrastructure, which were arranged in mono- or oligoaxonal units and frequently displayed different stages of myelination. As these ultrastructural characteristics resembled typical features of extrinsic nerves and were similar to those observed in subserosal nerves, the prominent intramural nerve fascicles were considered to be of extraenteric origin. Most likely their overabundance contributes to the functional obstruction of the terminal colon. Morphological abnormalities of BL encountered in the aganglionic colonic segment consisted of an extensive multilamination of BL surrounding glial processes and an irregular thickening of BL surrounding perineurocytes and smooth muscle cells of the muscularis mucosae. Similar alterations in BL have also been described in inherited peripheral and diabetic autonomic neuropathies and attributed to reactivated schwann cells. Thus, the overproduction of BL material within the hypertrophied nerve fascicles in HD may reflect an increased activity of proliferating glial cells. Since the smooth muscle cells of the muscularis mucosae showed abnormalities of BL similar to those observed in murine models for HD, it is suggestive that also in human HD the aganglionic colon is affected by a disturbed intestinal micro-environment impairing the neuronal colonisation and promoting the ingrowth of extrinsic nerves. The ultrastructurally observed alterations in BL of both neuronal and non-neuronal cells, as well as the increased amount of perineural and endoneural collagen provide further evidence that extracellular matrix components are abnormally distributed and overproduced within the bowel wall of patients affected by HD.