Ultrastructural localization of Tamm—Horsfall protein in human kidney using immunogold electron microscopy

Abstract

The ultrastructural localization of Tamm-Horsfall protein (THP) was studied in paraformaldehyde-fixed human renal biopsies. Pre-embedding and post-embedding immunogold labelling techniques were developed utilizing a monoclonal antibody specific for human urinary THP. With the pre-embedding technique, membrane contrast was enhanced by osmification thus allowing precise localization of gold particles. Reasonable tissue penetration of antibodies was achieved without compromising ultrastructural detail. The hydrophilic resin LR White was used for post-embedding labelling to ensure maximum penetration of antibodies. However, sections had only mild osmification and consequently localization of label was less certain. Both labelling techniques gave similar results. THP was found to be associated with two renal cell types. Epithelial cells lining the thick ascending limb of Henle's loop had gold label closely associated with the whole cell plasmalemma, with some of these cells having an apparently random distribution of label throughout the cytoplasm. Only the luminal plasmalemma of epithelial cells lining distal convoluted tubules were found to be labelled. Basolateral membranes and the cytoplasm of these cells were negative. The use of a monoclonal antibody of defined specificity combined with the two immunolabelling procedures represents a precise reliable method for studying ultrastructural localization of THP in the human kidney.

Abnormal expression and processing of uromodulin in Fabry disease reflects tubular cell storage alteration and is reversible by enzyme replacement therapy

“…Similar variability in storage has been described before (Gubler et al 1978). As in previous studies on UMOD expression in control kidney, we observed prominent expression of UMOD in the TALH and less so in the DCT in normal kidneys (Hoyer and Seiler 1979;Peach et al 1988;Schenk et al 1971;Sikri et al 1981). Thus, both processes, lysosomal storage and UMOD expression, meet in the TALH.…”

“…Abbreviations DCT distal convoluted tubule ERT enzyme replacement therapy Gb 3 globotriaosylceramide GLA !-galactosidase A MALDI-TOF matrix assisted lased desorption/ ionization-time of flight SRT substrate reduction therapy TALH thick ascending limb of Henle_s loop UAKD uromodulin-associated kidney disease UMOD uromodulin thick ascending limb of Henle_s loop (TALH), the macula densa segment and the distal convoluted tubule (DCT) (Hoyer and Seiler 1979;Peach et al 1988;Schenk et al 1971;Sikri et al 1981). Normally the synthesized protein is glycosylated, glypiated, secreted and glycosylphosphatidylinositol-anchored in the apical membrane of the polarized renal tubular epithelial cells (Kreft et al 2002;Rindler et al 1990;Serafini-Cessi et al 1993), from where it is continuously released by a specific but not yet identified protease (Cavallone et al 2001;Fukuoka and Kobayashi 2001).…”

Tamm-Horsfall protein-deficient thick ascending limbs promote injury to neighboring S3 segments in an MIP-2-dependent mechanism

“…We conclude that TAL affects the susceptibility of S3 segments to injury at least in part by regulating MIP-2 expression in a THP-dependent manner. Our findings raise the interesting possibility of a direct role of basolaterally released THP on regulating inflammation in S3 segments.CXCL2; ischemia-reperfusion; uromodulin; acute kidney injury TAMM-HORSFALL PROTEIN (THP) is a unique glycoprotein because it is exclusively expressed in the kidney, specifically in thick ascending limbs (TALs) and early distal tubules (4,15,29). Although this glycoprotein was discovered 60 years ago (37), its functions are still not fully understood (35).…”

“…CXCL2; ischemia-reperfusion; uromodulin; acute kidney injury TAMM-HORSFALL PROTEIN (THP) is a unique glycoprotein because it is exclusively expressed in the kidney, specifically in thick ascending limbs (TALs) and early distal tubules (4,15,29). Although this glycoprotein was discovered 60 years ago (37), its functions are still not fully understood (35).…”

Isolation of mouse THP gene promoter and demonstration of its kidney-specific activity in transgenic mice

“…Within the kidney, THP has been localized by immunohistochemistry and in situ hybridization to the thick ascending limb of Henle's loop (TALH) and the early distal convoluted tubules (2,3,17,27,40). At these locations, THP is believed to be membrane-anchored via its COOH-terminal glycophosphatidylinositol linkage, but the protein can be released into the urine by the action of phospholipases or proteases (7,9,15,28,34).…”