Clusterin is a multifunctional protein isolated from a number of tissues in several different species. In a variety of renal diseases, clusterin appears in the glomerulus and tubules in association with the membrane attack complex of complement. It is also transiently expressed after several forms of acute renal injury. In this study, we examined the expression and intrarenal distribution of clusterin following subtotal renal ablation. Male rats were subjected to either 1-1/3 nephrectomy (1-1/3 NX), uninephrectomy (UNX) or sham operation (SHAM). Two weeks after surgery, clusterin mRNA was elevated in the 1-1/3 NX group (1-1/3 NX: 1215 ± 88; UNX: 208 ± 11; SHAM: 207 ± 19 OD units; P < 0.001) Clusterin mRNA increased between 3 and 24 hours after 1-1/3 NX, plateaued, and remained elevated for at least seven weeks. The increased clusterin mRNA in 1-1/3 NX was localized to the tissue adjacent to the infarctive scar (scar 858 ± 173 vs. non-scar 98 ± 27 OD units; P < 0.001). Clusterin protein followed a similar pattern of localization being increased in most tubules and some peritubular capillaries in the peri-infarct zone. Only occasional tubules were positive for clusterin in the renal tissue distant from the scar or in the kidneys of sham operated rats. Co-localization of clusterin and C5b-9 was not detected. Evidence for apoptosis was found in the peri-infarct zone but not elsewhere in 1-1/3 NX kidney or in the normal kidney following sham operation. Infarction of 1/3 of the left kidney without contralateral nephrectomy, a maneuver which eliminates the compensatory growth, and uremia seen with 1-1/3 NX still resulted in increased clusterin mRNA in the infarcted left kidney compared to the intact right kidney (LK: 790 ± 112 vs. RK: 128 ± 25 OD units: P < 0.001), although the amount of clusterin mRNA was less than that found following 1-1/3 NX. In conclusion, persistently increased clusterin mRNA and protein was seen in the peri-infarct zone following 1-1/3 NX. This increased expression of clusterin may be playing a role in the ischemia-related apoptosis present in the scar-adjacent tissue.
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scar (S) from normal renal parenchyma (N) (non-immune rabbit serum, hematoxylin counterstain). b. Tubules in scar adjacent tissue display apoptosis (arrows) (Hematoxylin and eosin). c. Immunoperoxidase staining of a kidney of a sham operated rat shows reactivity of all epithelial cells seen in the cross section of one medullary tubule (rabbit anti-clusterin immunoperoxidase, hematoxylin counterstain), d. Most tubules in the scar-adjacent tissue show reactivity for clusterin; resulting in a band of clusterin immunoreactivity which clearly demarcated the scar tissue from the normal kidney, seen on the upper part of the figure. (Rabbit anti-clusterin immunoperoxidase, hematoxylin counterstain). e. Scar-adjacent renal parenchyma of the 1-1/3 NX kidney; reactivity for clusterin is seen in tubular epithelial cells, tubular remnants (arrows) and endothelial cells of peritubular capillaries (arrowheads). (Rabbit anti-clusterin immunoperoxidase, hematoxylin counterstain). 1. Scar-adjacent renal tissue of the 1-1/3 NX kidney; predominant apical cytoplasmic staining is seen in epithelial cells of tubule at lower left. (Rabbit anti-clusterin immunoperoxidase, hematoxylin counterstain). Clusterin was also found in casts, some of which appeared to include sloughed tubular epithelial cells. Publication of this figure in color was made possible by a grant from Marion Merrell Dow, Inc., Kansas City, Missouri, USA.
thank David Chmielewski and Stefan Kren for expert technical assis- tance, M. Tenniswood for the TRPM-2 cDNA, M. Griswold for the polyclonal rabbit anti-rat clusterin antiserum, and W. Causer for the rnurine monoclonal anti-rat C5b-9 antibody. This work was supported by a US Public Health Service Grant (AM-31437) (T.H.H.) and by a Young Investigator Grant from the National Kidney Foundation (M.E.R.). Dr. Correa-Rotter is a recipient of a Juvenile Diabetes Foundation International Postdoctoral Research Fellowship, and is also partially supported by the "Instituto Nacional de Ia Nutricidn Salvador Zubirán", Mexico City.