Oligodendrocyte gene expression in the human fetal spinal cord during the second trimester of gestation.

A comprehensive evaluation of myelination during normal human development is essential to understand the pathology of congenital diseases of white matter. The present study establishes quantitative values for normal oligodendrocyte-specific gene expression during the early stages of myelination in the human fetal spinal cord. Complementary techniques of Northern and immunoblotting were used to determine relative amounts of oligodendrocyte-specific mRNAs and proteins between 12 and 24 gestational weeks. Values were determined for myelin basic protein, 2',3'-cyclic nucleotide 3'-phosphodiesterase, and proteolipid protein. The relative amount of myelin-associated glycoprotein mRNA was also estimated. To compare gene expression between glial cell types, the relative amounts of mRNA and protein were determined for glial fibrillary acidic protein (GFAP), a cell-type specific marker for astrocytes. All oligodendrocyte-specific genes expressed similar developmental kinetics. Between 12 and 15 gestational weeks, less than a five-fold increase was detected in the expression of these genes and their protein products. Between 15 and 22 gestational weeks, the relative amounts of mRNA and protein for the myelin genes increased more than 80-fold. The kinetics of GFAP expression were similar to those of the myelin-associated genes. Absolute values for the increase in mass of the human fetal spinal cord were also obtained. These results provide data that may aid in the neuropathologic assessment and characterization of myelin disorders in the preterm, neonatal, and pediatric spinal cord.

Quantification of myelin basic protein in the human fetal spinal cord during the midtrimester of gestation.

The amount of myelin basic protein (MBP) was quantified in human fetal spinal cords from 12 to 24 gestational weeks (GW). MBP expression was determined by Northern blot, quantitative immunoblot, and immunocytochemistry. The development of compact myelin was analyzed by electron microscopy. Thirty-eight human fetal spinal cords were obtained after elective termination of intrauterine pregnancies from healthy women. Northern blot analysis showed a 15.8-fold increase in MBP mRNA between 12 and 18 GW. From 18 to 24 GW, MBP mRNA increased by 2.2-fold. The mRNA data paralleled immunoblot results that showed a 90.5-fold increase in MBP (0.147 ng/mg to 13.3 ng/mg tissue) between 12 and 18 GW and an approximately 11.5-fold increase between 18 and 24 GW (13.3 ng/mg to 154 ng/mg tissue). Immunocytochemical analysis also showed increased staining for MBP with advancing gestational age. At 12 GW, MBP immunoreactivity was observed in all three spinal cord funiculi. By 18 GW, MBP was expressed throughout the spinal cord white matter with the exception of the lateral corticospinal tracts and in the rostral levels of the fasciculus gracilis. With respect to myelin, at 12 GW, rare, noncompacted myelin lamellae were observed by electron microscopy. By 18 GW, discrete areas of compact myelin were observed in areas that showed MBP immunoreactivity, and at 24 GW, compact myelin was prominent throughout the white matter of the spinal cord. This study demonstrates a quantitative increase in MBP expression that is associated with myelin formation during the second trimester of human gestation. This information may provide normative data that can aid in the diagnosis of myelin disorders of the preterm, neonatal, and pediatric spinal cord.

Human fetal central nervous system organotypic cultures.

Many aspects of human neurodevelopment remain enigmatic. A main reason for this is, although there have been a significant number of morphologic and biochemical studies of neural tissues derived from human embryos and fetuses, this can only provide a static picture of the events at a given gestational age. Also, in vitro studies that focus on cells derived from these tissues have a limitation in that different cell types in dissociated tissue culture cannot interact in a 'normal' physiologic manner thereby, perhaps, altering their housekeeping and luxury functions. The present study focused on the development of a human explant organotypic culture model that may overcome the static limitation of the first example and permit a dynamic analysis of different cell types as they interact which may satisfy the second restriction. Because there is an array of developmental markers that define different cell phenotypes, this explant model may also provide a means of analyzing, for the first time, processes undefined in the human CNS. Human fetal CNS tissue obtained from second trimester abortuses was established in culture. The tissues were maintained for up to 12 weeks during which time they developed and differentiated. Sample cultures were harvested periodically and analyzed by light- and electron microscopy in combination with immunocytochemistry. Differentiation of neurons, astrocytes, oligodendrocytes and endothelial cells was documented using morphologic and biochemical criteria. As such, this model system may allow for the analysis of processes that occur during normal human fetal neurodevelopment and in pathologic conditions.