[Analysis of glomerular anionic charge status in renal biopsy specimens of childhood minimal change nephrotic syndrome using confocal laser scanning microscopy].

Analysis of glomerular anionic charge in human renal biopsy specimens has been restricted previously to staining of sites at the electron microscopic level, which is a product that needs skills and precludes a wide observable area. The introduction of a new tool, confocal laser scanning microscopy together with FITC conjugated poly-L-lysine as a cationic tracer, which demonstrates fixed anionic sites in thin sections from routinely formalin-fixed and paraffin-embedded renal biopsy tissue, has now enabled glomerular charge at light microscopic level. In this method, the patterns of staining in tissue showing minimal change nephrotic syndrome (MCNS) indicate that the intensity of anionic charge in 4 children with heavy proteinuria was significantly less than that in 7 children without proteinuria at remission, supporting previous observations using electron microscopy. Furthermore, staining the serial sections after methylation or saponification revealed that carboxyl components such as sialic acid may be responsible for proteinuria. We anticipate that this method may facilitate the investigation of the participation of charged components in the pathogenesis of MCNS and their role in relation to glomerular proteinuria.

Immunogold labelling of cytokines in glomeruli in children with various renal diseases.

Precise localization of cytokines such as transforming growth factor-beta (TGF-beta), tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-1 and IL-6 was observed in glomeruli using immunogold electron microscopy in 21 children with various types of renal diseases. The distribution pattern of these cytokines, as well as immunoglobulins, C3c and fibrinogen (Fg), was essentially confined to the electron-dense deposits (EDDs) regardless of their location. Frequency of positive labelling of each cytokine was different among various types of renal disorder, that is, TGF-beta was found mainly in lupus nephritis (LN), membranous nephropathy and IgA nephropathy, TNF-alpha in LN, and IL-1 in Henoch-Schönlein purpura nephritis. IL-6 was detected only in 1 case of LN. TNF-alpha was also found in the cytoplasm of glomerular epithelial cells. Furthermore, in order to evaluate the relation of cytokines to mesangial expansion, extracellular matrix components such as type IV collagen, laminin and fibronectin were stained. The result was that there was no significant correlation between the signal intensity or distribution pattern of cytokines and that of extracellular matrix components. These findings indicate that these cytokines could be associated with the formation of EDDs together with immunoglobulins, C3c and Fg. The involvement of each cytokine in renal pathophysiology might also depend upon the type of renal disease. They also raise the possibility that the glomerular epithelial cells might produce or absorb TNF-alpha. However, these results did not show significant correlation between cytokine involvement and mesangial expansion.

Analysis of choline acetyltransferase protein in temperature sensitive mutant flies using newly generated monoclonal antibody.

The protein, choline acetyltransferase (ChAT; EC 2.3.1.6), was analyzed in wild-type and two different temperature-sensitive ChAT mutants of Drosophila (Cha(ts1) and Cha(ts2)) using newly generated monoclonal antibodies. In all of the three genotypes, Western blots of crude fly head extracts showed a band stained at approximately the 80-kDa position, supporting the hypothesis that these temperature-sensitive mutants were generated by point mutation in the structural gene. The staining intensity of the bands indicated that these mutants have a lesser amount of ChAT protein than wild-type, even when they are reared at the permissive temperature (18 degrees C).

[Classification of crown forms. Morphological interrelationship between upper central incisors and facial forms].

The interrelationship between tooth forms and facial outlines is represented in multivariate linear combinations. Principal Component Analysis was used in this study as an expression of morphological features. Measured data on the tooth and the face could be classified into 4 types by the first and second principal components of multivariate analysis. Of the 117 subjects, the largest group of 33 (28.9%) had of oval faces and were Class III for the face and Class II for the teeth, respectively. Of the 117 subjects, 36 showed morphological similarity between teeth and face.

Uptake of retinol by cultured fibroblasts.

Cultured fibroblasts of adult rats were used to determine whether they could take in retinol administered to the culture medium at physiological concentration. After the administration of retinol, cells were observed with a phase-contrast fluorescence light microscope (LM) and a transmission electron microscope (TEM). Retinol and retinyl fatty acyl esters (RFAE) stored in the cells were analyzed with high-performance liquid chromatography (HPLC). It was revealed that these fibroblasts could take in retinol in the medium at a concentration of 1 x 10(-7) M and store it in lipid droplets in the cytoplasm as retinyl palmitate and other RFAE.

Structure of retinular cells in a Drosophila melanogaster visual mutant, rdgA, at early stages of degeneration.

A Drosophila visual mutant rdgA has photoreceptive cells which degenerate gradually after eclosion. Fine structure of the retinular cells of rdgAKS60 and rdgAKO14 was studied during early stages of degeneration to determine the initial morphological defects. The retinular cells of these two alleles showed the following structural abnormality within 1 day after eclosion: (1) rhabdomeres were small and irregular in shape; (2) cisternae of the rough endoplasmic reticulum were more numerous than those in normal retinular cells; (3) submicrovillar cisternae were absent; and (4) lysosomes were fewer than normal. Three-dimensional reconstruction of serial sections of the ommatidia showed that the degeneration of mutant rhabdomeres proceeds more rapidly in regions remote from the nuclei. These results suggest that the process of turnover of rhabdomeric microvilli is abnormal in rdgA. We also confirmed an increase of lysosomes and destruction of cellular organelles, as reported by previous investigators at more advanced stages of degeneration.