Osteogenic protein-1 mRNA expression is selectively modulated after acute ischemic renal injury.

Osteogenic protein-1 (OP-1) is a morphogenetic factor highly expressed in the kidney and involved in tissue repair and development. Homozygous OP-1-deficient mice die shortly after birth due mainly to arrest of renal growth and differentiation. Because postischemic injury involves several repair mechanisms, this study examined whether kidney OP-1 mRNA expression is modulated after ischemia. Acute ischemic renal injury was achieved in rats by unilateral clamping of the renal pedicle followed by reperfusion. Rats were killed at 3, 6, 12, 24, and 48 h and 7 d after reperfusion, and kidneys were microdissected and analyzed by histology and Northern and Western blots. Changes in OP-1 mRNA were determined by measuring the ratio of OP-1/glyceraldehyde 3-phosphate dehydrogenase signals for each OP-1 transcript (4.0 and 2.4 kb) from ischemic, opposite, and sham-operated rats. The OP-1 mRNA content for transcript 4.0 kb was fivefold lower in the whole ischemic kidney compared with that in sham animals 24 h after reperfusion. In the ischemic medulla, OP-1 mRNA was strikingly downregulated 20-fold when compared with the ischemic cortex. Results for transcript 2.4 kb and for the other time points were comparable. OP-1 mRNA expression was also affected in the opposite medulla compared with the sham medulla. However, only in the ischemic medulla was the relative OP-1 content significantly lower at all time points. Similar results were obtained when analyzing OP-1 protein by Western blot at 24 h after reperfusion. Seven days after reperfusion, the levels of OP-1 mRNA returned to baseline. In conclusion, kidney OP-1 mRNA and protein are selectively downregulated in the medulla after acute ischemic renal injury. OP-1 modulation may be a key element for kidney repair.

Renal biopsy diagnosis in children presenting with haematuria.

We reviewed the clinical and renal biopsy findings in 322 children presenting during the years 1975-1996 with recurrent macro- or continuous microscopic haematuria persisting for > or =6 months, in whom non-glomerular causes were excluded. Family involvement was documented for first-degree relatives. All biopsies were examined by light microscopy, 317 by electron microscopy and 315 by immunofluorescence. Biopsies were classified as IgA nephropathy (78), Alport nephropathy (86), thin basement membrane nephropathy (TMN) (50), miscellaneous glomerulonephritis (32), hilar vasculopathy (28) and normal glomeruli (48). Although microscopic haematuria alone was more frequent in Alport nephropathy and TMN, the pattern of haematuria in individual patients did not predict histology. Of patients with familial haematuria, 79% of biopsies showed either Alport nephropathy or TMN. Hilar vasculopathy was observed both in isolation and in all abnormal histological categories.

Interactions of ultrapure bovine hemoglobin with renal epithelial cells in vivo and in vitro.

Using an ultrapurified hemoglobin (Hb) solution, we investigated the physiological effects and cellular processing of Hb in rat kidneys and in cultured opossum kidney (OK) cells. Rats infused with < 5.0 g/kg Hb showed no change in baseline serum creatinine (SCr) values (0.58 +/- 0.05 mg/dl) over 48 h, whereas transient acute renal failure followed infusion of 7.5 g/kg Hb (SCr 3.4 +/- 1.02 mg/dl, P = 0.02). Histology of Hb-infused kidneys demonstrated tubular epithelial cell injury. Renal injury was not caused by volume or oncotic load, cardiovascular effect, or ATP depletion. After Hb infusion, heme oxygenase, the rate-limiting enzyme in Hb catabolism, was induced in an organ-specific fashion. Inhibiting heme oxygenase activity with cimetidine did not alter Hb renal injury. Using OK cells, we determined that renal epithelia process Hb by fluid-phase endocytosis. Proton permeability of fluorescein Hb endosomes was unaltered compared with fluorescein dextran controls, demonstrating that Hb does not alter endosomal membrane integrity. These data suggest that Hb renal injury in rats occurs following large doses of ultrapure Hb, does not alter early steps in Hb endosomal processing by renal epithelia, and involves a mechanism that is not heme oxygenase dependent.

Attenuated varicella virus vaccine in children with renal transplants.

The long-term efficacy of varicella vaccine was studied in 34 children aged 2-18 years who were either on chronic dialysis (n = 17) or were renal transplant recipients (n = 17). Live attenuated virus (OKA line) was inoculated in a single 0.5-ml subcutaneous dose, without modification of the immunosuppressive therapy protocol for renal transplant recipients. The majority of children (85%) developed antibodies within the first 6 months, with IgG titres (enzyme-linked immunosorbent assay) greater than 1:40 (geometric mean 1:640). Of those children who were followed for longer than 2 years, 76% maintained their antibody titres. Reactions to the vaccine were minimal and the immunological protection was effective. Only 3 children developed a mild form of varicella in the post-vaccination period. We consider that seronegative children who are candidates for renal transplantation must be protected against varicella by attenuated varicella vaccination. When vaccination is performed after transplantation, no modification of immunosuppressive therapy is needed.

Permeability properties of rat renal lysosomes.

Although lysosomes maintain large pH gradients and may be subjected to significant osmotic gradients in vivo, little is known about their passive permeability properties. In recent studies, vacuolar H(+)-adenosine-triphosphatases (ATPases), such as those found in lysosomes, have been suggested to act as water channels. In addition, the erythrocyte and proximal tubule water channel CHIP28 is present on the plasma membrane of proximal tubule cells and may undergo endocytosis so that it is incorporated in lysosomes. We therefore examined water, proton, and small nonelectrolyte permeabilities in freshly purified lysosomes from rat renal proximal tubule. Lysosomes were purified by differential and Percoll gradient centrifugation. The preparation contained only lysosomes when examined by electron microscopy. Moreover, analysis by flow cytometry showed virtually all particles to be positive for acid phosphatase and cathepsin B activities. Permeabilities were measured on a stopped-flow fluorimeter by monitoring the self-quenching or pH-sensitive quenching of entrapped fluorescein derivatives. Osmotic water permeability (Pf) averaged 0.011 +/- 0.003 cm/s (n = 6), a value similar to that of biological membranes containing water channels. However, Pf was insensitive to the organic mercurial reagent p-chloromercuribenzene-sulfonate and to HgCl2 and exhibited an activation energy of 10.8 +/- 0.8 kcal/mol. These results indicate that water flux in lysosomes occurred via the lipid bilayer, and not via water channels. Addition of ATP led to lysosomal acidification (proton flux = 4.6 +/- 0.8 x 10(-11) mmol H+.s-1.cm-2), which was completely inhibited by 0.1 microM bafilomycin. Pf was insensitive to this agent as was the passive proton permeability (0.36 +/- 0.18 cm/s, n = 4). Permeabilities to small nonelectrolytes varied in proportion to the oil-water partition coefficient, confirming the applicability of Overton's rule to lysosomes. We conclude that proximal tubular lysosomes exhibit high Pf, which occurs via the lipid bilayer and not via vacuolar H(+)-ATPase.

Tubular proteinuria in steroid sensitive multi-relapsing nephrotic syndrome.

The urinary excretion of N-acetyl-beta-D-glucosaminidase (UNAG) and retinol binding protein (URBP) was studied in 65 children with steroid sensitive multirelapsing nephrotic syndrome (MRNS): 28 on cyclosporin A (CyA) therapy, 22 on prednisolone (P), 15 off-treatment and in 32 normal children to assess renal tubular damage or dysfunction. The urinary protein excretion was expressed in relation to that of creatinine (UNAG/UC in mumol pnp/h/mmol; URBP/UC in microgram/mmol). There was a weak but significantly negative correlation between age and both, UNAG/UC (r = -0.38, p < 0.01) and URBP/UC (r = -0.50, p < 0.05) in normal children, but not in nephrotics. In normals and in patients off steroids an association between these two proteins was found (r = 0.38, p < 0.05; r = 0.56, p < 0.05 respectively). Geometric mean UNAG/UC was significantly higher in nephrotics on CyA therapy (26.5 +/- 4.0), and on P (37.0 +/- 7.9) as well as in those off-treatment (16.3 +/- 3.1) compared to normal children (9.3 +/- 3.4). There was a further increase in those with raised urinary albumin: creatinine ratio (UA/UC) (> 0.1 mg/mg). URBP/UC was not increased in any of the groups of children with MRNS. Raised NAG in urine may therefore indicate active nephrotic syndrome rather than being due to the drug therapy.