Microproteinuria in children with vesicoureteric reflux

Microproteinuria was assessed by the measurement of albumin, retinol binding protein (RBP) and creatinine concentrations in random midstream urine samples using a single enzyme linked immunoassay (ELISA) in 36 children with vesicoureteric reflux (VUR) and 36 control patients. Infection was excluded by culture and microscopy of the specimens of urine. No patient was hypertensive. Albumin excretion increased in patients with increasing severity of VUR and with renal scarring. Similar findings were observed with RBP excretion. The results show that glomerular and tubular handling of proteins is altered in VUR. The degree of microproteinuria correlates well with the severity of the VUR and is evidence of tubular dysfunction. The effects of medical management and anti-reflux surgery on microproteinuria require further evaluation.     

Model of Epstein-Barr virus infection of human thymocytes: expression of viral genome and impact on cellular receptor expression in the T- lymphoblastic cell line, HPB-ALL

Infection of B lymphocytes and epithelial tissue by Epstein-Barr virus (EBV) is associated with malignancy and autoimmunity. The cellular receptor for EBV has been identified as CD21 (CR2). A molecule, which is biochemically and immunologically similar to B-cell CD21, has been identified on a subpopulation of immature thymocytes, suggesting a role for this molecule in the regulation of T-cell development and further suggesting that immature T cells might be susceptible to EBV infection. A growing body of literature now documents the presence of EBV in tumors of T-cell origin. We have evaluated the susceptibility of the human immature T cell line, HPB-ALL, to infection by EBV. Electron microscopy studies showed a rapid internalization of virus by HPB cells. Southern blotting showed the intracellular presence of linear EBV genomes, and components of the virus replicative cycle were identified. Expression of the BamHI Z region of the genome, encoding the nuclear protein, ZEBRA, which is strictly associated with productive infection in B cells, was detected in HPB-ALL cells. A spliced variant of Z, RAZ, was also identified. Cell surface expression of EBV late antigens was observed to occur transiently. Infection of HPB cells was also accompanied by altered expression of T-cell surface molecules involved in antigen recognition, a process critical to normal development of the T-cell repertoire. Delineation of the outcome of T- cell infection by EBV may lead to a better understanding of the role of this virus in autoimmune processes and malignancy.     

The reliability of height and height velocity in the assessment of growth (the Wessex Growth Study)

Both biochemical and auxological measurements can be used to assess growth. Quality control in routinely reported in laboratory studies, but the reproducibility of height measurements, and the height velocity data derived from them, is seldom considered. We have previously established our error and in this report we examine its implications for the screening of short children and subsequent monitoring of their growth. The 95% confidence interval for height for a 5 year old observed to be on the 3rd centile for height, spanned the 2nd-4th centile. However, the confidence interval for a 12 month height velocity appropriate to such a child spanned the 8th-52nd centiles, the lower limit pathological and the upper more than satisfactory. A single height velocity even over 12 months therefore lacks the precision to provide a reliable index of current growth in short children. Furthermore, serial height velocity calculations on a cohort of 78 short normal children showed no significant correlation from year to year, suggesting that velocity is also unable to predict future growth. Although the proportion of this cohort of short children lying beneath the 25th centile for velocity remained constant from year to year, the identity of the individuals comprising that proportion changed, a phenomenon which could be largely accounted for by the random error associated with height velocity. Our data suggest that, by the time a trend in abnormal velocity is reliably established, a deviation from the height centiles is clearly evident. Although velocity charts are attractive in concept, they seem to be no more discriminating than height charts in practice, and may be clinically deceptive unless interpreted with great care.     

Expression of GD3 ganglioside by developing rat cerebellar Purkinje cells in situ

GD3 is a major ganglioside of the immature vertebrate CNS, and its expression is suggested to be characteristic of immature neuroectodermal cells. Using immunocytochemistry on cryostat sections of developing rat cerebellum with a monoclonal antibody specific for GD3, we have found that GD3 begins to be expressed on the plasma membrane of Purkinje cell bodies and dendrites beginning at postnatal day 7. Staining became brighter as the dendritic tree of the cells enlarged. As the Purkinje cells began to mature in different folia, they became GD3+, until by 15 days postnatal all Purkinje cells were GD3+. Positive staining of the dendritic tree was still present in the adult cerebellum. Using a monoclonal antibody 7-8D2, which recognizes cerebellar granule cells and their axons (the parallel fibres), and polyclonal antibodies against a synaptic vesicle component synaptophysin, double-immunofluorescence staining together with anti-GD3 antibodies suggested that the appearance of GD3 immunoreactivity did not correlate either with the ingrowth of parallel fibres or the presence of their synapses on Purkinje cell dendrites. However, comparison with earlier morphological studies showed that the appearance of GD3 immunoreactivity correlated well with the formation of climbing fibre synapses on Purkinje cell dendrites and the onset of the rapid expansion of the dendritic tree. These results are in keeping with the idea that elevated GD3 concentrations are found in certain cell types during periods of rapid growth or high metabolic activity but also show that this is not only restricted to immature cells.     

Oligodendroglial progenitor cells but not oligodendroglia divide during normal development of the rat cerebellum

Summary To identify the stage in the life cycle of oligodendroglia at which they are mitotic during normalin vivo development [³H]thymidine autoradiography has been combined with immunocytochemistry on frozen sections of rat cerebellum. A panel of antibodies have been used that recognize antigens expressed by oligodendroglia at different stages of differentiation from progenitor to mature cell. It has been demonstrated that of the cells of the Oligodendroglial lineage only the progenitors, identified by their expression of the ganglioside G_D3, were seen to incorporate [³H]thymidine at all the developmental stages tested. Only few dividing G_D3-positive cells were observed in the subventricular zones of the fourth-ventricle. The greatest number of dividing G_D3-positive progenitors in the rat cerebellum was observed in the folia at postnatal day 7. Silver grains were never observed over cells that could be distinguished as oligodendroglia by their expression of galactocerebroside, 23-cyclic nucleotide 3-phosphohydrolase, or myelin basic protein. Mitotic astroglia were observed at all stages and could be clearly distinguished by their expression of glial fibrillary acidic protein and glutamine synthetase. When animals were injected with [³H]thymidine at postnatal day 7 and killed at 1-day intervals radiolabel was first observed in galactocerebrosidepositive and 23-cyclic nucleotide 3-phosphohydrolase-positive oligodendroglia at day 9 and in myelin basic protein-positive cells at day 10–11, 3-days after the last cell division. Thus, we have demonstrated for the first time usingin situ immunocytochemical techniques, a mitotic glial progenitor cell that is known to give rise to oligodendroglia bothin vivo andin vitro.     

Down-regulation of GAP-43 During Oligodendrocyte Development and Lack of Expression by Astrocytes In Vivo: Implications for Macroglial Differentiation

The discovery of molecular markers which are selectively expressed during the development of specific classes of rat central nervous system macroglia has greatly advanced our understanding of how these cells are related. In particular, it has been shown in tissue culture that oligodendrocytes and some astrocytes (type-2) may be derived from a common progenitor cell (O-2A progenitor). However, the existence of type-2 astrocytes in vivo has yet to be unequivocally established. Recently, it has been reported that the neural-specific growth-associated protein-43 (GAP-43, otherwise known as B-50, F1, pp46 and neuromodulin) may be expressed by cells of the O-2A lineage in vitro. We set out to examine the cellular specificity of GAP-43 in O-2A progenitors and their descendants in vitro and in vivo. Using a polyclonal antiserum against a GAP-43 fusion protein we have shown the presence of immunoreactive GAP-43 in the membranes of bipotential O-2A glial progenitor cells and type-2 astrocytes by Western blotting and immunocytochemistry of cells in culture. In contrast to previous studies, double labelling with mature oligodendrocyte markers showed that GAP-43 is down-regulated during oligodendrocyte differentiation in vitro. Immunohistochemical staining of sections of developing rat brain demonstrated the same developmental regulation of GAP-43, suggesting that oligodendrocytes only express GAP-43 at immature stages. In addition, normal and reactive astrocytes in tissue sections were not labelled with GAP-43.     

PreCimp: Pre‐collapsing imputation approach increases imputation accuracy of rare variants in terms of collapsed variables

Imputation is widely used for obtaining information about rare variants. However, one issue concerning imputation is the low accuracy of imputed rare variants as the inaccurate imputed rare variants may distort the results of region‐based association tests. Therefore, we developed a pre‐collapsing imputation method (PreCimp) to improve the accuracy of imputation by using collapsed variables. Briefly, collapsed variables are generated using rare variants in the reference panel, and a new reference panel is constructed by inserting pre‐collapsed variables into the original reference panel. Following imputation analysis provides the imputed genotypes of the collapsed variables. We demonstrated the performance of PreCimp on 5,349 genotyped samples using a Korean population specific reference panel including 848 samples of exome sequencing, Affymetrix 5.0, and exome chip. PreCimp outperformed a traditional post‐collapsing method that collapses imputed variants after single rare variant imputation analysis. Compared with the results of post‐collapsing method, PreCimp approach was shown to relatively increase imputation accuracy about 3.4–6.3% when dosage r² is between 0.6 and 0.8, 10.9–16.1% when dosage r² is between 0.4 and 0.6, and 21.4 ～ 129.4% when dosage r² is below 0.4.