Isolation, characterization, and differentiation of multipotent neural progenitor cells from human cerebrospinal fluid in fetal cystic myelomeningocele.

Despite benefits of prenatal in utero repair of myelomeningocele, a severe type of spina bifida aperta, many of these patients will still suffer mild to severe impairment. One potential source of stem cells for new regenerative medicine-based therapeutic approaches for spinal cord injury repair is neural progenitor cells (NPCs) in cerebrospinal fluid (CSF). To this aim, we extracted CSF from the cyst surrounding the exposed neural placode during the surgical repair of myelomeningocele in 6 fetuses (20 to 26weeks of gestation). In primary cultured CSF-derived cells, neurogenic properties were confirmed by in vitro differentiation into various neural lineage cell types, and NPC markers expression (TBR2, CD15, SOX2) were detected by immunofluorescence and RT-PCR analysis. Differentiation into three neural lineages was corroborated by arbitrary differentiation (depletion of growths factors) or explicit differentiation as neuronal, astrocyte, or oligodendrocyte cell types using specific induction mediums. Differentiated cells showed the specific expression of neural differentiation markers (ßIII-tubulin, GFAP, CNPase, oligo-O1). In myelomeningocele patients, CSF-derived cells could become a potential source of NPCs with neurogenic capacity. Our findings support the development of innovative stem-cell-based therapeutics by autologous transplantation of CSF-derived NPCs in damaged spinal cords, such as myelomeningocele, thus promoting neural tissue regeneration in fetuses.

Diagnosis of central nervous system infection by CSF sampling of the myelomeningocele sac as an alternative to ventricular tap.

BACKGROUND: Infection significantly affects mortality and morbidity in myelomeningocele cases. Ventricular tap is the most common method performed to diagnose central nervous system (CNS) infection in myelomeningocele patients. However, the ventricular tap can cause serious trauma to the baby and to the family. Here we discuss the technique of taking a cerebrospinal fluid (CSF) sample from the sac in myelomeningocele cases. METHODS: The study comprised 24 myelomeningocele patients undergoing sac repair; 7 patients in the early period and 17 in the late period (after the first 24 hours). CSF samples were taken from the sac and via ventricular tap. In all patients' samples, cell count and cultures were compared. RESULTS: In patients who underwent early meningomyelocele repair there was no significant difference between sac and ventricular CSF. There were also no positive cultures in samples taken from both areas. In comparison, in the group submitted to late repair, the number of cells in the sac and ventricle CSF samples was over 10 in 4 of the 17 patients. The cultures from CSF samples taken from both areas were positive for E. coli. In 13 of 17 patients who were admitted to our clinic for late repair, there was no significant difference between the number of cells in the sac and in ventricular samples. Cultures taken from these patients were negative. CONCLUSION: Puncture of the sac represents a quicker and more convenient way to obtain CSF in myelomeningocele cases. In addition, there was no increase in pain to a level that would cause stress for the baby. A CSF sample can be taken from the sac for diagnosis of CNS infection in myelomeningocele patients. This method, therefore, represents a safer and more comfortable option for both the patient and doctor.

Diagnosis of central nervous system infection by CSF sampling of the myelomeningocelesac as an alternative to ventricular tap

Background. Infection significantly affects mortalityand morbidity in myelomeningocele cases. Ventriculartap is the most common method performed to diagnosecentral nervous system (CNS) infection in myelomeningocelepatients. However, the ventricular tap can causeserious trauma to the baby and to the family. Here wediscuss the technique of taking a cerebrospinal fluid(CSF) sample from the sac in myelomeningocele cases.Methods. The study comprised 24 myelomeningocelepatients undergoing sac repair; 7 patients in theearly period and 17 in the late period (after the first 24hours). CSF samples were taken from the sac and viaventricular tap. In all patients’ samples, cell count andcultures were compared.Results. In patients who underwent early meningomyelocelerepair there was no significant differencebetween sac and ventricular CSF. There were also nopositive cultures in samples taken from both areas. Incomparison, in the group submitted to late repair, thenumber of cells in the sac and ventricle CSF sampleswas over 10 in 4 of the 17 patients. The cultures fromCSF samples taken from both areas were positive for E.coli. In 13 of 17 patients who were admitted to our clinicfor late repair, there was no significant difference betweenthe number of cells in the sac and in ventricularsamples. Cultures taken from these patients were negative.Conclusion. Puncture of the sac represents a quickerand more convenient way to obtain CSF in myelomeningocelecases. In addition, there was no increase inpain to a level that would cause stress for the baby. ACSF sample can be taken from the sac for diagnosisof CNS infection in myelomeningocele patients. Thismethod, therefore, represents a safer and more comfortableoption for both the patient and doctor (AU)

Fundamentos. La infección de LCR afecta significativamentela mortalidad y morbilidad en pacientes conmielomeningocele. La punción ventricular es la técnicamás frecuentemente utilizada para detectar infección delSNC en los casos de mielomeningocele. Sin embargo, lapunción ventricular puede causar un grave estrés tantopara el niño como para la familia. En nuestro estudio,presentamos la técnica y resultados del examen de LCRobtenido a partir del saco del mielomeningocele.Pacientes y métodos. El estudio comprende 24 pacientescon mielomeningocele sometidos a cierre quirúrgicode la malformación, 7 pacientes con reparaciónprecoz y 17 con cirugía tardía (después de las primeras24 horas). Las muestras de LCR fueron obtenidas delsaco de la malformación y de los ventrículos mediantepunción ventricular. Se compararon los resultados delrecuento de células y los cultivos realizados a todos lospacientes.Resultados. En los pacientes operados precozmente,no se encontraron diferencias significativas entre elLCR lumbar y ventricular. Tampoco se obtuvieroncultivos positivos en las muestras tomadas de ambasáreas. En los operados después de las primeras 24horas, el recuento de células del LCR del saco y delventrículo fue alrededor de 10 en 4 de los 17 pacientes.Se obtuvieron cultivos positivos (E. coli) de las muestrasde LCR tomadas de ambas áreas. En 13 de 17 pacientesque fueron admitidos a nuestra clínica para reparacióntardía, no hubo una diferencia significativa entre elnúmero de células del saco y del LCR ventricular. Loscultivos de LCR obtenidos en estos pacientes fueron (..) (AU)

Potential of proton magnetic resonance spectroscopy in the evaluation of patients with tethered cord syndrome following surgery.

OBJECT: Spinal cord dysfunction is associated with an altered neuronal metabolism. The objective of this study is two-fold: 1) to compare pre- and postoperative levels of cerebrospinal fluid (CSF) metabolites in patients with spinal dysraphism and in control patients by performing proton magnetic resonance spectroscopy; and 2) to evaluate the use of magnetic resonance (MR) spectroscopy in the assessment of surgical outcomes in patients with spinal dysraphism. METHODS: The study group population was composed of patients with meningomyeloceles, lipomeningomyeloceles with tethered cord syndrome, and tethered fatty fila. All patients underwent preoperative clinical and neuroimaging (ultrasonography or MR imaging) examinations and MR spectroscopy analysis of metabolites in their CSF. Excision of the neural placode and detethering of a low-lying cord were performed with or without laminectomy. Two months postoperatively, the investigations were repeated. A comparison of pre- and postoperative CSF metabolites was performed using the Wilcoxon signed-rank test and nonparametric tests. Probability values less than 0.05 were considered significant. High levels of lactate (Lac), alanine (Ala), acetate, glycerophosphorylcholine, and choline were observed in the CSF of patients with spinal dysraphism before surgery; after surgery these levels normalized to those observed in control patients. Patients in whom cord retethering occurred could be identified by increased concentrations of Ala and Lac. CONCLUSIONS: The results highlight the potential of MR spectroscopy as a promising tool in the assessment of surgical outcomes in patients with spinal dysraphism.

Metabolite profile of cerebrospinal fluid in patients with spina bifida: a proton magnetic resonance spectroscopy study.

STUDY DESIGN: The present study was carried out to assess the metabolic differences between cerebrospinal fluid samples of patients with spina bifida and age-matched control individuals. OBJECTIVES: To study the metabolite profile of cerebrospinal fluid of patients with spina bifida using proton magnetic resonance spectroscopy, compare the levels of metabolites with controls, establish correlation of underlying neuronal dysfunction with metabolic changes in patients with spina bifida, and evaluate the potential use of this technique as an additional tool for diagnostic assessment. SUMMARY OF BACKGROUND DATA: Combination of embryopathy, stretching, ischemia, compression, and trauma is responsible for cord dysfunction in spina bifida. Changes in neuronal metabolism leads to changes in the local milieu of cerebrospinal fluid in the cord. Change in metabolite profile of cerebrospinal fluid in spina bifida in terms of increase in products of anaerobic metabolism, nerve membrane integrity, and nerve ischemia has not yet been studied. METHODS: Cerebrospinal fluid obtained from patients and control individuals were characterized using various one- and two-dimensional proton magnetic resonance spectroscopy techniques. Concentration of various metabolites was calculated using the area under the nuclear magnetic resonance peak. RESULTS: Statistically significantly higher levels of lactate, choline, glycerophosphocholine, acetate, and alanine in the cerebrospinal fluid of patients with spina bifida was observed compared with control individuals. CONCLUSIONS: Significantly higher levels of metabolites were observed in patients with spina bifida, representing a state of nerve ischemia, anaerobic metabolism, and disruption of neuronal membrane.

Correction of hindbrain herniation and anatomy of the vermis after in utero repair of myelomeningocele in sheep.

BACKGROUND/PURPOSE: In utero repair of myelomeningocele (MMC) in humans spares distal neurologic function, reverses the hindbrain herniation component of the Arnold-Chiari II malformation (ACM), and reduces the rate of postnatal shunt placement. The authors hypothesized that extravasation of cerebrospinal fluid (CSF) from the lumbar spinal cord results in herniation. This hypothesis was tested by assessing the impact of a spinal cord myelotomy on hindbrain anatomy in fetal sheep. METHODS: A MMC lesion was created surgically in 34 fetal sheep at 75 days' gestation by excision of the L1-L5 lamina, the exposed dura, and surrounding tissues. A lumbar level myelotomy was performed in 28 of the 34 fetuses to open the central canal of the spinal cord to enhance egress of CSF through the MMC defect and potentially induce hindbrain herniation. At 102 days' gestation, a repair of the MMC lesion was performed in 14 fetuses with a myelotomy. Fetuses underwent autopsy at 102, 114, 120, or 140 days' gestation. Control animals underwent 2 unrelated fetal surgical procedures at approximately 70 and 110 days' gestation. The incidence of hindbrain herniation, ventricular size, biparietal diameter, brain weight, and brain anatomy were compared between the different animal groups. RESULTS: After MMC creation, significant cerebellar tonsillar herniation was observed in 85% of fetuses that underwent creation of a myelotomy; none of the lambs without a myelotomy (n = 6) had hindbrain herniation. At autopsy, cerebellar tonsillar herniation was present at the time of MMC repair (102 days' gestation), 2 weeks after MMC repair, but was reversed 3 weeks post-MMC repair. At birth, tonsillar herniation was absent, and hindbrain anatomy was restored in 88% of the fetuses with a myelotomy that underwent fetal MMC repair. No significant differences in brain weight and ventricular size was observed between animals with and without MMC repair. CONCLUSIONS: Adding a myelotomy to the sheep model of MMC leads to hindbrain herniation that is similar to that observed in the human ACM. These experiments support the hypothesis that leakage of CSF through the exposed central canal alters the normal CSF hydrodynamics, resulting in cerebellar tonsillar herniation. Fetal MMC repair reverses hindbrain herniation and restores gross anatomy of the vermis.

MR of terminal myelocystoceles.

PURPOSE: To delineate the clinical and MR findings in children with an unusual type of spinal dysraphism, the terminal myelocystocele. Infants with a terminal myelocystocele carry a favorable neurologic prognosis if the entity is diagnosed early. Understanding the MR characteristics of this entity will allow for earlier and more accurate diagnosis. METHOD: Analysis of the medical charts and MR studies in 15 children with surgically and histologically proven myelocystocele. RESULTS: In all 15 children, MR demonstrated the primary findings of a terminal cyst of the central canal of the spinal cord which is tethered and herniated with arachnoid and cerebrospinal fluid through an area of spinal dysraphia onto the back as a mass. Of these children, 10 had additional findings (one or more) on MR of Chiari I (five cases), Chiari II (one case), cervicothoracic hydromyelia (two cases), lumbar hydromyelia (two cases), hydrocephalus (2 cases) segmentation anomalies of vertebrae (3 cases) and partial agenesis of sacrum (six cases). Of the clinical findings, all 15 children had a back mass, 10 also had cloacal exstrophy. One had imperforate anus, 10 were girls, five had ambiguous genitalia and all were neurologically intact. CONCLUSION: Children with a terminal myelocystocele present with a back mass and there is a high association with cloacal exstrophy. MR is the best noninvasive modality to diagnose all of the components of a terminal myelocystocele and the associated central nervous system findings.

The Chiari malformations: a constellation of anomalies.

The Chiari malformations form a group of abnormalities that are pathogenetically interrelated. The most important member of the group is the Chiari type II malformation, known as the Arnold-Chiari malformation. Its cardinal features are the myelomeningocele in the thoraco-lumbar spine, the venting of the intracranial cerebrospinal fluid through the central canal, the hypoplasia of the posterior fossa, the herniation of hindbrain into the cervical spinal canal, and the compressive damage to cranial nerves. Some of the abnormalities are progressive, and thus treatable. Limitation of progression may improve outcomes. The challenges to our treatment programs involve early diagnosis, delivery by Caesarean section, emergent closure of the neural plaque and prophylaxis of hydrocephalus, anticipatory prevention of the neurological compression syndromes, multidisciplinary teams, and age-appropriate interventions.

[The surgical procedure in spinal cord hernias of large size combined with hydrocephalus]. / Khirurgicheskaia taktika pri spinnomozgovykh gryzhakh bol'shikh razmerov, sochetaiushchikhsia s gidrotsefaliei.

The paper analyzes the examination and treatment of 77 patients with myelomeningoceles. It is shown that the prognosis of treatment depends on the size and content of the hernial sac, the specific features of deformities of the liquor-containing system and the severity of liquor circulatory disorders. Both specific features of myelodysplasia manifestations and those of liquor circulatory disorders should be borne in mind in choosing the optimum surgical policy for myelomeningoceles. The optimum surgical approach to treating large-sized myelomeningoceles concurrent with hydrocephalus is demonstrated to one-stage plastic surgery of the hernial hilus and drainage of the subarachnoidal space of the terminal cistern into the abdomen.

Maldevelopment of spinal cord--substance P in spina bifida.

Recent works have shown that the neuropeptide, substance P, is an important element of the central nervous system, functioning as a neurotransmitter as well as moderating neural development. In a previous communication, we provided indirect evidence that substance P played a role in the pathogenesis and pathophysiology of spina bifida by showing an elevation of CSF substance P concentrations in affected foetuses and babies. In this study, we investigated directly the occurrence of substance P in spinal cord tissues in spina bifida and compared it to that found during normal development. Using radioimmunoassay, concentrations of substance P were measured in the spinal cords of 20 normal human foetuses (aged 9-21 weeks) and 4 foetuses with spina bifida (aged 17-19 weeks). Substance P was detected in substantial amounts in the normal foetal spinal cord as early as 9 weeks' gestation, and there was progressive accumulation with age: mean concentrations +/- S.E.M. in 3 subgroups were 56.3 +/- 4.9 pmol g-1 (9-12 weeks), 59.5 +/- 7.2 pmol g-1 (13-16 weeks) and 117.3 +/- 7.2 pmol g-1 (17-21 weeks). In spina bifida, substance P was severely depleted in the dysraphic segment of the cord (mean concentration +/- S.E.M. = 22.1 +/- 2.3 pmol g-1) and moderately so in the segment above the dysraphism (mean concentration +/- S.E.M. = 47.2 +/- 4.2 pmol g-1).(ABSTRACT TRUNCATED AT 250 WORDS)

Eosinophilia in the cerebrospinal fluid of children with shunts implanted for the treatment of internal hydrocephalus.

Cerebrospinal fluid (CSF) eosinophilia was observed in 26 of 404 children after the implantation of shunts for the treatment of internal hydrocephalus. High levels of 65% and 78% were recorded in two cases, which are reported in detail. In the remaining 24 cases, CSF eosinophilia ranging between 1% and 3% was found. None of the cases with CSF eosinophilia had blood eosinophilia. The cases indicate that a reaction to the material used for the shunt should be considered along with possible parasitic infestations in patients with such findings.

Elevation of cerebrospinal fluid substance P concentrations in spina bifida.

Concentrations of substance P, a peptide believed to have neurotransmitter and possible trophic functions, were determined by radioimmune assay in the cerebrospinal fluids of ten babies and one foetus with spina bifida, and compared to those of five normal babies and ten normal foetuses. The results (mean +/- S.E.M.) were: spina bifida babies (864 +/- 357.6 fmol ml-1), normal babies (141 +/- 14.2 fmol ml-1), spina bifida foetuses (150,000 fmol ml-1), normal foetuses (22,673 +/- 8,340 fmol ml-1). Hence, in spina bifida cerebrospinal fluid substance P concentrations were grossly elevated both pre- and post-natally. The significance of these results can be considered from two aspects: there is the possibility that substance P plays a role in the development of the spinal cord which is disturbed in spina bifida; as substance P has been shown to be involved with sensory perception and autonomic reflexes, abnormal concentration of substance P in cerebrospinal fluid may aggravate the neurological deficit in spina bifida. Possibly, therapeutic manipulation of substance P, when available in the future, may be considered in these patients.

CSF polyamines in childhood.

Specimens of CSF from 76 children without neurologic disease and from 191 children with neurosurgical conditions were assayed for polyamine content. Putrescine and spermidine concentrations decreased with age. In children with intracranial tumors, polyamine concentrations in lumbar CSF were comparable with those in ventricular CSF. Putrescine level was significantly increased in children with medulloblastomas. Spermidine level was increased in children with medulloblastomas, glioblastomas, and astrocytomas. Concentrations of putrescine and spermidine were significantly increased in infants with myelomeningocele and hydrocephalus. Spermidine concentrations were significantly increased in older children with myelomeningocele, encephalocele, and hydrocephalus. Polyamines thus seem to be increased by rapid cell proliferation and by disorders affecting myelination. Since putrescine and spermidine concentrations may be of use in monitoring tumor progression, the effects of childhood growth and of hydrocephalus must be considered for children whose brain tumors are associated with hydrocephalus.

Isoelectric focusing of cerebrospinal fluid proteins in children with nontumoral hydrocephalus. A preliminary report.

The cerebrospinal fluid (CSF) protein patterns of 42 children with nontumoral hydrocephalus, 33% with associated congenital malformations (meningocele, myelomeningocele, spina bifida, monoventricular prosencephalon) were determined by isoelectric focusing and polyacrylamide gel electrophoresis. 50% of the patients showed a barrier damage pattern, 19% showed a degenerative pattern, a block pattern or a normal pattern were less frequently found. 14 children presented hydrocephalus associated with other malformations, 44% of these cases showed a degenerative pattern. Combining results from quantitative polyacrylamide gel electrophoresis and qualitative isoelectric focusing, resulted in better characterization of CSF protein abnormalities, the high resolution capacity of these techniques may give a new approach to the evaluation of the blood CSF barrier in the management of hydrocephalus.

A quantitative assessment of CSF reabsorption in infants with meningomyelocele.

The production of cerebrospinal fluid and bulk flow reabsorption were determined in four children with meningomyelocele who developed hydrocephalus. By subtracting bulk flow reabsorption from CSF production, the amount of CSF reabsorbed by alternative pathways was determined. The results of the study indicate that the majority of CSF reabsorption in these patients takes place by bulk flow. CSF reabsorption by alternative pathways, however, is significant.

Pre- and postoperative cerebrospinal fluid absorption studies in patients with myelomeningocele shunted for hydrocephalus.

Nine infants with myelomeningocele and hydrocephalus had cerebrospinal fluid formation and absorption studies before insertion of a ventriculoperitoneal shunt. Six were also tested postoperatively. The preshunt formation rate varied from 0.21-0.42 ml.min-1 and the cerebrospinal fluid absorption rate or CSF conductance (Ccsf) varied from 0.008 to 0.031 ml.min-1.mm Hg-1. The postoperative formation rate varied from 0.21 to 0.39 ml.min-1. The postoperative CSF conductance (Ct) was calculated as the sum of Ccsf and the shunt conductance (Cs). The relationship between the Ccsf and Cs curves was determined by matching the sum of these curves to the postoperative data points using the method of least-squares. The data suggests that the shunt may work directly or indirectly to establish a new resting pressure; the latter by stabilizing the sagittal sinus pressure which preoperatively increased concomitantly with the CSF pressure and thus enhanced an already-existing absorption deficit.