Three-dimensional ultrastructure of glomerular injury in serum sickness nephritis using the quick-freezing and deep-etching method

Abstract

The three-dimensional ultrastructure of the glomerulus in serum sickness nephritis has been investigated by the quick-freezing and deep-etching method. Compact granular immune deposits were localized in filamentous networks in the lamina densa and mesangial matrices. These constitutional fibrils with diameters of 8-15 nm, were directly attached to the immune deposits. The filamentous networks became markedly loosened around the deposits. In podocytes, reticular microfilaments with positive decoration by myosin subfragment 1 (S1) were increased in flattened foot processes and directly attached to the cell membranes. Fine filaments with diameters of 4-7 nm were undecorated by S1 and connected with actin filaments as cross-bridges. Intermediate filaments were also increased in the cell bodies and primary processes of podocytes. Connecting fibrils in lamina rara externa were partially disrupted. The immune deposits were primarily detected in the networks of lamina densa and actually destroyed the size barrier composed of filamentous networks. Moreover, the mesangial deposits also disorganized mesangial networks to probably alter mesangial flow through the matrices. Increased actin filaments in foot processes seemingly reinforced the cell membranes and the connecting fibrils in lamina rara externa, which prevented the initial detachment of podocytes from the basement membrane.

Three-Dimensional Study of Glomerular Lesions in Murine Chronic Graft-versus-Host Reaction by the Quick-Freezing and Deep-Etching Method

“…Takami et al [8] reported that mesan gial matrices in normal glomeruli had meshwork structure with jungle-gym-like labyrinth spaces, and both podocytes and endothelial cells were attached to the matrix with perpendicular fibrils. In serum sickness nephritis, immune deposits were seen in the mesangial matrix, and meshworks were disrupted around them [12]. In Masugi's nephritis, it is known that the sclerosis consisted of condensed fine fibril lar meshwork structures more thickened fibrils (type I col- lagen) were connected with [11], It is reported that mesangial deposits may induce the secretion of IL-1 or cytokines and as a result glomerular sclerosis develops [14,15].…”

“…Moreover, focal mesangiolytic changes were seen at stages later than the 7th week. These morphological findings sug gested that the deposition of the immune complex might appear during an early period of the illness in comparison with the data of ELIZA [7], Recently, three-dimensional ultrastructures of normal glomeruli have been reported by the quick-freezing and deep-etching method [8,9], Moreover, there have been other ultrastructural studies about pathological changes, such as Masugi's nephritis and serum sickness nephritis [10][11][12]. In the chronic phase of Masugi's nephritis, newly formed mesangial matrices were composed of fine fibrillar meshworks and related to the mesangial sclerosis [11].…”

“…pH 7.4, via the left ventricle under pentobarbital anesthesia. The renal cortices were cut into small pieces and washed in PB for 30 min to remove soluble proteins, as reported before [II,12], Then, they were postfixed with 0.25% glutaraldehyde in PB for 30 min. They were immersed in 10% methanol and quickly frozen in an isopentane-propane mixture (-193^C) cooled in liquid nitrogen.…”

Three-Dimensional Ultrastructure of Human Glomerular Basement Membrane by a Quick-Freezing and Deep-Etching Method

“…Recent ly, the QF-DE method was applied to clarify the threedimensional ultrastructure of the GBM in normal rats [8] or some models of renal diseases in animals [9][10][11][12][13][14], How ever, the renal tissues for our replica preparations have needed large sizes, 2x2x4 mm to obtain lots of infor mation about glomerular structures. So the QF-DE meth od has not yet been applied to biopsied materials from human kidneys.…”

“…In experimental animals, proteinuria occurred due to disruption of fibrils in the middle layer and enlargement of meshworks, which were demon strated in Masugi nephritis [9], serum sickness nephritis [10], graft-vs.-host reaction [13] and STZ-induced dia betes [14], The destruction or enlargement of meshworks in the middle layer seems to be a cause of proteinuria even in human cases. The QF-DE method has already been used for some animal models of renal diseases such as Masugi nephritis [9], serum sickness nephritis [10,11], anti-thymocyte nephritis [12], graft-vs.-host reaction [13], and STZ-induced diabetes [14]. So, we are going to apply the QF-DE method to human renal diseases, such as glo merulonephritis or diabetic nephropathy, to evaluate the mechanism of proteinuria.…”