ACE inhibition and ANG II receptor blockade improve glomerular size-selectivity in IgA nephropathy.
(9/2888)
Protein trafficking across the glomerular capillary has a pathogenic role in subsequent renal damage. Despite evidence that angiotensin-converting enzyme (ACE) inhibitors improve glomerular size-selectivity, whether this effect is solely due to ANG II blocking or if other mediators also play a contributory role is not clear yet. We studied 20 proteinuric patients with IgA nephropathy, who received either enalapril (20 mg/day) or the ANG II receptor blocker irbesartan (100 mg/day) for 28 days in a randomized double-blind study. Measurements of blood pressure, renal hemodynamics, and fractional clearance of neutral dextran of graded sizes were performed before and after 28 days of treatment. Both enalapril and irbesartan significantly reduced blood pressure over baseline. This reduction reached the maximum effect 4-6 h after drug administration but did not last for the entire 24-h period. Despite transient antihypertensive effect, proteinuria was effectively reduced by both treatments to comparable extents. Neither enalapril nor irbesartan modified the sieving coefficients of small dextran molecules, but both effectively reduced transglomerular passage of large test macromolecules. Theoretical analysis of sieving coefficients showed that neither drug affected significantly the mean pore radius or the spread of the pore-size distribution, but both importantly and comparably reduced the importance of a nonselective shunt pathway. These data suggest that antagonism of ANG II is the key mechanism by which ACE inhibitors exert their beneficial effect on glomerular size-selective function and consequently on glomerular filtration and urinary output of plasma proteins. (+info)
A physiological barrier distal to the anatomic blood-brain barrier in a model of transvascular delivery.
(10/2888)
BACKGROUND AND PURPOSE: Osmotic disruption of the blood-brain barrier (BBB) provides a method for transvascular delivery of therapeutic agents to the brain. The apparent global delivery of viral-sized iron oxide particles to the rat brain after BBB opening as seen on MR images was compared with the cellular and subcellular location and distribution of the particles. METHODS: Two dextran-coated superparamagnetic monocrystalline iron oxide nanoparticle contrast agents, MION and Feridex, were administered intraarterially in rats at 10 mg Fe/kg immediately after osmotic opening of the BBB with hyperosmolar mannitol. After 2 to 24 hours, iron distribution in the brain was evaluated first with MR imaging then by histochemical analysis and electron microscopy to assess perivascular and intracellular distribution. RESULTS: After BBB opening, MR images showed enhancement throughout the disrupted hemisphere for both Feridex and MION. Feridex histochemical staining was found in capillaries of the disrupted hemisphere. Electron microscopy showed that the Feridex particles passed the capillary endothelial cells but did not cross beyond the basement membrane. In contrast, after MION delivery, iron histochemistry was detected within cell bodies in the disrupted hemisphere, and the electron-dense MION core was detected intracellularly and extracellularly in the neuropil. CONCLUSION: MR images showing homogeneous delivery to the brain at the macroscopic level did not indicate delivery at the microscopic level. These data support the presence of a physiological barrier at the basal lamina, analogous to the podocyte in the kidney, distal to the anatomic (tight junction) BBB, which may limit the distribution of some proteins and viral particles after transvascular delivery to the brain. (+info)
Comparison of ultrasmall particles of iron oxide (USPIO)-enhanced T2-weighted, conventional T2-weighted, and gadolinium-enhanced T1-weighted MR images in rats with experimental autoimmune encephalomyelitis.
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BACKGROUND AND PURPOSE: Ultrasmall particles of iron oxide (USPIO) constitute a contrast agent that accumulates in cells from the mononuclear phagocytic system. In the CNS they may accumulate in phagocytic cells such as macrophages. The goal of this study was to compare USPIO-enhanced MR images with conventional T2-weighted images and gadolinium-enhanced T1-weighted images in a model of experimental autoimmune encephalomyelitis (EAE). METHODS: Nine rats with EAE and four control rats were imaged at 4.7 T and 1.5 T with conventional T1- and T2-weighted sequences, gadolinium-enhanced T1-weighted sequences, and T2-weighted sequences obtained 24 hours after intravenous injection of a USPIO contrast agent, AMI-227. Histologic examination was performed with hematoxylin-eosin stain, Perls' stain for iron, and ED1 immunohistochemistry for macrophages. RESULTS: USPIO-enhanced images showed a high sensitivity (8/9) for detecting EAE lesions, whereas poor sensitivity was obtained with T2-weighted images (1/9) and gadolinium-enhanced T1-weighted images (0/9). All the MR findings in the control rats were negative. Histologic examination revealed the presence of macrophages at the site where abnormalities were seen on USPIO-enhanced images. CONCLUSION: The high sensitivity of USPIO for macrophage activity relative to other imaging techniques is explained by the histologic findings of numerous perivascular cell infiltrates, including macrophages, in EAE. This work supports the possibility of intracellular USPIO transport to the CNS by monocytes/macrophages, which may have future implications for imaging of human inflammatory diseases. (+info)
A substituted dextran enhances muscle fiber survival and regeneration in ischemic and denervated rat EDL muscle.
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Ischemia and denervation of EDL muscle of adult rat induce a large central zone of degeneration surrounded by a thin zone of peripheral surviving muscle fibers. Muscle regeneration is a complex phenomenon in which many agents interact, such as growth factors and heparan sulfate components of the extracellular matrix. We have shown that synthetic polymers, called RGTA (as regenerating agents), which imitate the heparan sulfates, are able to stimulate tissue repair when applied at the site of injury. In crushed muscles, RGTA were found to accelerate both regeneration and reinnervation. In vitro, RGTA act as protectors and potentiators of various heparin binding growth factors (HBGF). It was postulated that in vivo their tissue repair properties were due in part to an increase of bioavailability of endogenously released HBGF. In the present work, we show that ischemic and denervated EDL muscle treated by a unique injection of RGTA differs from the control after 1 wk in several aspects: 1) the epimysial postinflammatory reaction is inhibited and the area of fibrotic tissue among fibers is reduced; 2) the peripheral zone, as measured by the number of intact muscle fibers, was increased by more than twofold; and 3) In the central zone, RGTA enhances the regeneration of the muscle fibers as well as muscle revascularization. These results suggest that RGTA both protects muscle fibers from degeneration and preserves the differentiated state of the surviving fibers. For the first time it is demonstrated that a functionalized polymeric compound can prevent some of the damage resulting from muscle ischemia. RGTA may therefore open a new therapeutic approach for muscle fibrosis and other postischemic muscle pathologies. (+info)
Studies on the uptake, metabolism, and release of endogenous and exogenous chemicals by use of the isolated perfused lung.
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The isolated perfused lung is a valuable tool for studying many lung functions. The kinds of information one can obtain from the isolated perfused lung are illustrated by examples from our studies on the uptake, accumulation, and metabolism of endogenous and exogenous chemicals. (+info)
Assay of intercellular adhesiveness using cell-coated Sephadex beads as collecting particles.
(14/2888)
A simple, rapid and precise method, based on a previous method, for measuring relative rates of intercellular adhesion is described. DEAE-Sephadex beads were treated with nitrocellulose in order to allow cells to grow on their surfaces. Balb/c 3T3 and Balb/c 3T12 cells were used to characterize the assay. They formed confluent cell layers on nitrocellulose-treated DEAE-Sephadex. These cell-coated beads were employed to collect 32P-labelled cells from single cell suspensions. Since they formed statistically uniform, large collecting surfaces, the collection of labelled cells was markedly improved as compared to the original assay. The cell-coated beads collected a large percentage of the labelled cells in a short time. The percentage of cells collected was independent of the concentration of labelled cells in the assay mixture, and the collection was linear for approximately 60 min. The variability between replicate assays was usually +/- 5%. The assay allows the rapid and precise determination of intercellular adhesion in large numbers of individual samples. These features make it useful to screen for effects of different treatments on intercellular adhesions. (+info)
Modified LDL-mediated increases in endothelial layer permeability are attenuated with 17 beta-estradiol.
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-Current research suggests that estrogen may have primary effects on the artery wall. To investigate the mechanisms of female sex hormone actions in the artery wall, we used an isolated, perfused, rat carotid artery model to examine the effects of estradiol on the rates of accumulation of normal (N-LDL) and minimally modified (MM-LDL) low density lipoprotein in ovariectomized rats. N-LDL, MM-LDL, and oxidized LDL (OX-LDL) were fluorescently labeled and perfused into individual arteries. The rate of LDL accumulation was measured by quantitative fluorescence microscopy before and after treatment with estradiol (1 nmol/L, 272 pg/mL). Estradiol had no effect on the rate of N-LDL accumulation (45+/-12 versus 48+/-15 ng cholesterol per cm2 per h). However, estradiol significantly decreased the rate of MM-LDL (240+/-48 versus 160+/-48 ng cholesterol per cm2 per h; P<0.05) and OX-LDL (191+/-53 versus 112+/-36 ng cholesterol per cm2 per h; P<0.05) accumulation. Further experiments showed that perfusion of unlabeled MM-LDL (100 microgram/mL) increased endothelial layer permeability when the rate of accumulation of a water-soluble, fluorescently labeled, reference molecule (64 000-molecular weight dextran) was determined before and after perfusion of MM-LDL (319+/-96 versus 510+/-191 ng per cm2 per h, n=6 arteries; P<0.05). Estradiol prevented the expected increase in the rate of dextran accumulation when perfused with MM-LDL (control, 415+/-49 ng per cm2 per h and MM-LDL+estradiol, 415+/-160 ng per cm2 per h). Our studies show that estradiol prevents compromise of the endothelial barrier mediated by MM-LDL and attenuates accumulation of MM-LDL in the artery wall. (+info)
Perforin-dependent nuclear entry of granzyme B precedes apoptosis, and is not a consequence of nuclear membrane dysfunction.
(16/2888)
Killer lymphocytes utilize the synergy of a membranolytic protein, perforin, and the serine protease granzyme B (grB) to induce target cell apoptosis, however the mechanism of this synergy remains incompletely defined. We have previously shown that perforin specifically induces the redistribution of cytoplasmic grB into the nucleus of dying cells, however a causal role for nuclear targeting of grB in cell death has not been demonstrated. In the present study, we used confocal laser scanning microscopy (CLSM) to determine whether the nuclear accumulation of fluoresceinated (FITC-) grB precedes or is a consequence of apoptosis. Two distinct and mutually exclusive cellular responses were observed in FDC-P1 cells: (i) up to 50% of the cells rapidly accumulated FITC-grB in the nucleus (maximal at 7 min; t1/2 of 2 min) and underwent apoptosis; (ii) the remaining cells took up FITC-grB only into the cytoplasm, and escaped apoptosis. Under these conditions, DNA fragmentation was not observed for at least 13 min, indicating nuclear accumulation of grB preceded the execution phase of apoptosis. Furthermore, nuclear import of grB proceeded through an intact nuclear membrane, as the nuclei of cells whose cytoplasm was pre-loaded with 70 kDa FITC-dextran excluded dextran for up to 90 min while still undergoing apoptosis in response to perforin and grB. These findings indicated that perforin-induced nuclear accumulation of grB precedes apoptosis, and is not a by-product of caspase-induced nuclear membrane degradation. The cell membrane lesions formed by perforin in these experiments were not large enough to permit a 13 kDa protein (yeast cdk p13suc) access into the cytoplasm, but an 8 kDa protein (bacterial azurin) was able to equilibrate between the cytosol and the exterior. Therefore, transmembrane pores large enough to allow passive diffusion of grB (32 kDa) into the cell are not necessary for apoptosis. Rather, a perforin-dependent signal results in a redistribution of grB from the cytoplasm to the nucleus, where it may contribute to the nuclear changes associated with apoptosis. (+info)