Healthy Rat Kidneys Research Articles

Neuropeptide Y Is Expressed by Rat Mononuclear Blood Leukocytes and Strongly Down-Regulated during Inflammation

Neuropeptide Y (NPY), a classical sympathetic comediator, regulates immunological functions including T cell activation and migration of blood leukocytes. A NPY-mediated neuroimmune cross-talk is well conceivable in sympathetically innervated tissues. In denervated, e.g., transplanted organs, however, leukocyte function is not fundamentally disturbed. Thus, we hypothesized that NPY is expressed by blood leukocytes themselves and regulated during inflammation. NPY mRNA and peptide expression were analyzed in mononuclear leukocytes isolated from the blood vessels of healthy rat kidneys, as well as from the blood vessels of isogeneic and allogeneic renal grafts transplanted in the Dark Agouti to Lewis or in the Fischer 344 to Lewis rat strain combination. Depending on the donor strain, acute allograft rejection is either fatal or reversible but both experimental models are characterized by massive accumulation of intravascular leukocytes. Leukocytes, predominantly monocytes, isolated from the blood vessels of untreated kidneys and isografts expressed high amounts of NPY mRNA and peptide, similar to expression levels in sympathetic ganglia. During acute allograft rejection, leukocytic NPY expression drastically dropped to approximately 1% of control levels in both rat strain combinations. In conclusion, NPY is an abundantly produced and tightly regulated cytokine of mononuclear blood leukocytes.

CRBP-I in the renal tubulointerstitial compartment of healthy rats and rats with renal fibrosis

Cellular retinol-binding protein I (CRBP-I), a member of the intracellular lipid-binding protein (iLBP) superfamily, is a specific marker of quiescent stellate cells in the healthy human liver. In the diseased fibrotic/cirrhotic liver, portal and septal myofibroblasts acquire CRBP-I expression, while activated hepatic stellate cells maintain their CRBP-I expression. Here, we investigate the distribution of CRBP-I in the renal cortex of healthy rats and rats with renal fibrosis. Kidneys of healthy and adriamycin-treated rats were studied by immunohistochemistry, using antibodies against CRBP-I, desmin, vimentin and alpha-smooth muscle actin (alpha-SMA). Double stainings were done with immunofluorescence. Western blotting was performed to semi-quantify the expression levels of vimentin, desmin, alpha-SMA and CRBP-I. In the normal rat kidney, the convoluted proximal tubular epithelial cells express CRBP-I; no expression is found in the interstitium, nor in the glomeruli. In the adriamycin-induced fibrotic rat kidney, CRBP-I expression diminishes in the convoluted proximal tubular epithelial cells, whereas peritubular myofibroblasts in the interstitium acquire CRBP-I expression. In the tubulointerstitial compartment of the adriamycin-induced fibrotic rat kidney, CRBP-I is expressed in a different pattern than in the healthy rat kidney. As the convoluted proximal tubular epithelial cells dedifferentiate during fibrosis, CRBP-I expression decreases. Furthermore, de novo expression of CRBP-I is found in activated myofibroblast-like cells in the interstitium of adriamycin-treated rats. CRBP-I is therefore a useful marker to identify a subpopulation of activated/ myodifferentiated fibroblasts in the rat kidney.

CCN3 is a novel endogenous PDGF-regulated inhibitor of glomerular cell proliferation

CCN proteins affect cell proliferation, migration, attachment, and differentiation. We identified CCN3 as a suppressed gene following platelet-derived growth factor (PDGF)-BB or -DD stimulation in a cDNA-array analysis of mesangial cells. In vitro growth-arrested mesangial cells overexpressed and secreted CCN3, whereas the addition of the recombinant protein inhibited cell growth. Induction of mesangial cell proliferation by PDGF-BB or the specific PDGF beta-receptor ligand PDGF-DD led to downregulation of CCN3 mRNA, confirming the array study. Specific PDGF alpha-receptor ligands had no effect. CCN3 protein was found in arterial smooth muscle cells, the medullary interstitium, and occasional podocytes in the healthy rat kidney. Glomerular CCN3 was low prior to mesangial proliferation but increased as glomerular cell proliferation subsided during mesangioproliferative glomerulonephritis (GN). Inhibition of PDGF-B in mesangioproliferative disease led to overexpression of glomerular CCN3 mRNA. CCN3 localized mostly to podocytes in human glomeruli, but this expression varied widely in different human glomerulonephritides. Glomerular cell proliferation negatively correlated with CCN3 expression in necrotizing GN. Our study identifies CCN3 as an endogenous inhibitor of mesangial cell growth and a modulator of PDGF-induced mitogenesis.

Are renal proximal tubular epithelial cells constantly prepared for an emergency? Focus on “The proliferation capacity of the renal proximal tubule involves the bulk of differentiated epithelial cells”

a human kidney contains approximately one million functional units (the nephrons) that consist of a filter (the glomerulus) and a processing portion (the proximal, intermediate, and distal tubule). The glomeruli produce ∼180 liters of primary filtrate every day of which only 1 to 2 liters are

Tubular cell proliferation in the healthy rat kidney

We searched for morphological evidence to support the hypothesis that stem cells are responsible for renal tubular cell proliferation. The rationale of the study was that if proliferation relies on progenitors, mitotically active cells should be less differentiated than their neighbors. As the retention of the thymidine analog BrdU has been the only approach employed to identify stem cells in the kidney up to now we additionally characterized BrdU-retaining cells. Rat kidneys were fixed by perfusion. Cycling cells identified by mitotic figures or the expression of the proliferating cell nuclear antigen (PCNA) were examined by light microscopy and electron microscopy as well as immunofluorescence for four differentiation markers. Newborn rats were injected with BrdU in order to detect label-retaining cells. After a period of 8, 14 and 35 weeks the kidneys were examined for BrdU by immunofluorescence and the four differentiation markers mentioned above. All cycling cells showed the same degree of differentiation compared to non-cycling cells. Most of the detected label-retaining cells were differentiated. We conclude that cycling cells in tubules of the healthy kidney are differentiated and that the retention of label is not a criterion to identify stem cells in renal tubules.

Mesangial cell expression of proto-oncogene Ets-1 during progression of mesangioproliferative glomerulonephritis

Ets-1 is a transactivator of matrix-associated genes and key factor in neoangiogenesis. The expression of Ets-1 mRNA and protein was analyzed in healthy rat kidney and in a model for mesangioproliferative disease without and with inhibition of platelet-derived growth factor-B (PDGF-B) activity. Immunohistochemistry was performed using a specific noncrossreacting anti-Ets-1 antibody and included a counterstain with alpha-smooth muscle actin (alpha-SMA). Nuclear proteins from isolated glomeruli were analyzed by Western blotting. Changes in Ets-1 mRNA levels were detected by in situ hybridization and Northern blotting. PDGF-B antagonism was performed in nephritic rats by specific aptamers. The distribution of Ets-1-positive cells in healthy rats was heterogenous with exclusively nuclear staining of glomerular, tubular, and vascular cells. Profound changes were detected in the anti-Thy 1.1 glomerulonephritis. Nuclear Ets-1 staining was intense in mesangial cells, whereas podocyte and endothelial cell staining was unchanged. The strongest signal was seen on days 2 to 7. By Western blotting of glomerular proteins a single 52 kD band was detected in healthy rats, which was increased 4.5-fold after disease induction. At the same time a 54 kD band appeared that most likely represents phosphorylated Ets-1. Ets-1 transcripts were detected in mesangial cells that include exon IV but lack exon VII. A concordant 6.4-fold up-regulation of mRNA was detected in glomeruli. Specific PDGF-B antagonism by aptamer treatment from days 3 to 7 after disease induction led to reduced Ets-1 expression on day 7, correlating with decreased mesangial cell numbers. A temporal increase of mesangial cell Ets-1 expression correlates with mesangial cell activation in mesangioproliferative glomerulonephritis. PDGF-B may partially contribute to the increased expression.

Contrast Media-Induced Renal Tubular Vacuolization

The morphologic changes in healthy rat kidneys (n = 102) were studied 2 or 48 hours after intravenous injection of 1 or 3 g iodine (I)/kg of high-osmolality diatrizoate, low-osmolality iopromide and iohexol, or iso-osmolality iotrolan, as well as after 0.2 or 0.6 g/kg of the high-osmolality magnetic resonance contrast medium gadolinium DTPA. Physiologic saline was injected in controls. The kidneys were fixed by perfusion and the specimens were analyzed semiquantitatively by two independent observers blinded to the treatment. A statistically significant (P less than .01) cytoplasmic vacuolization was noticed in the proximal convoluted tubule cells 2 hours after injection of 3 g I/kg of diatrizoate or iopromide. Iohexol and iotrolan induced an even more significant (P less than .01) and longer-lasting vacuolization, but gadolinium DTPA did not produce lysosomal alterations. Although the vital cell organelles remained intact, reversible lysosomal alterations may represent the first structural signs of a threatening cellular injury.

The interleukin 1 gene is expressed by rat glomerular mesangial cells and is augmented in immune complex glomerulonephritis.

Cultured rat mesangial cells have features of immune effector cells that may contribute to the derangements in glomerulonephritis. Recent reports have demonstrated that mesangial cells produce a cytokine similar to interleukin 1 (IL 1). We predicted rat mesangial cells could express a gene homologous to murine macrophage IL 1. Mesangial cells were cultured by explant and were used in the fourth through sixth passages. Antibodies to desmin and fibronectin but not cytokeratin stained the cytoskeleton of all mesangial cells examined; no Ia+, leukocyte common antigen+ mononuclear phagocytes were present. RNA from mesangial cells and P388D macrophages hybridized in dot blots with a 32P-probe nick-translated from the murine IL 1 cDNA. Mesangial cells but not Swiss 3T3 fibroblasts contained IL 1 mRNA transcripts that co-migrated with the 2.0 kb message from murine P388D macrophages by Northern analysis of poly(A) RNA. In situ hybridization of cultured cells demonstrated specific hybridization of 3H-IL 1 probe to cells with the morphology of contractile mesangial cells and P388D cells but not 3T3 cells. IL 1 release is an important mediator of local inflammation and injury. Therefore we compared the expression of the IL 1 gene in total RNA from kidneys of rats with immune complex glomerulonephritis with that extracted from kidneys of healthy rats. Glomerulonephritic kidneys contain a twofold to threefold increase in IL 1 mRNA compared with normals. We conclude that rat mesangial cells express mRNA with significant homology to murine macrophage IL 1 mRNA and further suggest that local production of the potent phlogistic mediator IL 1 may be important in the pathogenesis of glomerulonephritis.