The effect of cephaloridine on the stability of rat kidney lysosomes.

The administration of cephaloridine to rats caused a decrease in the excretion of acid phosphatase into the urine. The antibiotic itself had no effect on urinary acid phosphatase and inhibitors or proteolytic enzymes were not present in the urine from treated rats. Cephaloridine may therefore be stabilizing the lysosomal membrane in vivo and experiments with isolated lysosomes confirm this hypothesis. The lysosomal integrity was followed by measuring the acid phosphatase activity and the light scattering properties of the particles. A good correlation was obtained between these parameters in the case of thermal disruption and progesterone induced lysis of the lysosomes and low concentrations of cephaloridine (0.1-1.0 mmol/1) protected the lysosomes against this form of damage.     

The effect of surfactants on the aggregation state of amphotericin B

We have studied the effect of two surfactants, one non-ionic, lauryl sucrose (LS) and the other ionic, sodium deoxycholate (DOC), on the aggregation state of amphotericin B (AmB) and its selectivity towards ergosterol and cholesterol. It is shown that the addition of these surfactants has very similar effects on the AmB micelles. Below the critical micellar concentration of the surfactants, mixed micelles with AmB are first formed as a result of the penetration of the surfactant molecules into the AmB micelles. At higher concentrations of the surfactant molecules, the micellar structure is completely destroyed and AmB is found as monomers in solution. When the concentration of the surfactant is further increased, micelles of the surfactant molecules are built up, AmB remaining in monomeric form. However, the critical micellar concentration of LS is modified by the presence of AmB in solution, while that of DOC is not affected, thereby indicating that the interactions of AmB with LS are stronger than those of DOC with AmB. We also show that both surfactants enhance the selectivity of the AmB binding to sterols at exactly the concentrations of the surfactants which induce the monomerization of the antibiotic. It is observed that the maximal selectivity is found at a concentration of the surfactants corresponding to their particular CMC in presence of the antibiotic.     

The effect of amphotericin B on the ultrastructure of Prototheca species

Prototheca zopfii and Prototheca wickerhamii strains were exposed to subinhibitory concentrations of the antimycotic amphotericin B, and the effect of the treatment on their ultrastructure was assessed. The results revealed ultrastructural changes in the treated cells, expressed by swelling of mitochondria, degradation of cell organelles, accumulation of microbody like structures, lipid droplets and starch granules in the cytoplasm, and changes in the inner layer of the cell wall.     

Natriuretic effect of digoxin-like immunoreactive substance on dog kidney

We previously reported that digoxin-like immunoreactive substance (DLIS) was found only in the blood of those dialysis patients who were hypertensive and had high systemic vascular resistance. In order to determine whether the DLIS was a marker for the natriuretic hormone, renal infusion studies were carried out in anesthetized dogs. When ultrafiltrates from patients with high blood DLIS levels were infused into the renal artery of one kidney there was a significant increase in the fractional excretion of sodium (FE Na) from its baseline value. Further, the FE Na of these kidneys were significantly higher than the FE Na noted for the contralateral kidneys which were simultaneously infused with ultrafiltrates obtained from dialysis patients lacking DLIS activity in their blood. We conclude that the DLIS is or represents a marker for natriuretic hormone. Since the natriuresis noted was independent of renal plasma flow and glomerular filtration rate and since the fractional excretion of potassium was not influenced by the infusion, we believe that DLIS is different from atrial natriuretic factor.     

The heterogeneity of rat kidney lysosomes revealed by immunoelectron microscopic staining for cathepsins B and H

The heterogeneity of lysosomes was studied by analyzing the immunostaining behavior of cathepsins B and H in rat kidney proximal tubule cells. Rat kidneys were fixed by perfusion and embedded in Lowicryl K4M. A protein A-gold technique was applied to serial sections and a double labeling technique to conventional sections. By analyzing the immunostaining behavior of cathepsins B and H in the same lysosomes which were cut into separate sections, four types of lysosomes were found: Type 1 positive for both proteinases; type 2 strongly positive for cathepsin B, but weakly or negative for cathepsin H; type 3 strongly positive for cathepsin H, but weakly or negative for cathepsin B; and type 4 negative for both proteinases. The double labeling by two different sizes of the protein A-gold probes showed these four types of lysosomes. The results indicate that there exists the lysosomal heterogeneity of the proteinase content in the kidney proximal tubule cells.     

The heterogeneity of rat kidney lysosomes revealed by immunoelectron microscopic staining for cathepsins B and H

Summary The heterogeneity of lysosomes was studied by analyzing the immunostaining behavior of cathepsins B and H in rat kidney proximal tubule cells. Rat kidneys were fixed by perfusion and embedded in Lowicryl K4M. A protein A-gold technique was applied to serial sections and a double labeling technique to conventional sections. By analyzing the immunostaining behavior of cathepsins B and H in the same lysosomes which were cut into separate sections, four types of lysosomes were found: Type 1 positive for both proteinases; type 2 strongly positive for cathepsin B, but weakly or negative for cathepsin H; type 3 strongly positive for cathepsin H, but weakly or negative for cathepsin B; and type 4 negative for both proteinases. The double labeling by two different sizes of the protein A-gold probes showed these four types of lysosomes. The results indicate that there exists the lysosomal heterogeneity of the proteinase content in the kidney proximal tubule cells.     

Alteration of membrane permeability by amphotericin B

Amphotericin B (AmB) increased unidirectional Na transport and net transcellular sodium movements across the skin of the frog, Rana pipiens, when added to the solution bathing the corium side, but not from the outer epidermal surface. The AmB response was prevented with pretreatment with amiloride, ouabain and mucosal sodium substitution. Alteration in pH markedly reduced the permeability changes induced by AmB. AmB did not interfere with the increase in sodium transport induced by antidiuretic hormone. The present study demonstrates that AmB interacts with the skin of the frog, Rana pipiens, from the corium side specifically increasing transepithelial sodium transport. The increase in transport apparently occurs through the existing sodium pathway.     

Lipid composition and effect of amphotericin B on yeast cells of Paracoccidioides brasiliensis

Yeast cells of Paracoccidioides brasiliensis strain SN, were obtained for analysis of lipid composition. Total lipids, phospholipids, sterols, and qualitative sterols and fatty acid composition were determined. Such analysis were made on cells cultured in the presence or absence of amphotericin B and on non proliferating cell suspensions exposed to the antibiotic. Marked alterations in lipid contents were observed in this different conditions. The major alterations were the reduction of total lipids, sterols, and palmitoleic acid in both, proliferating and non proliferating antibiotic exposed cells. The effect of amphotericin B was evaluated also in terms of viability and release of intracellular substances, at different times of exposure. The minimal inhibitory concentration (MIC) determined for that strain of this fungus was 0.2 g/mL.     

Mitigation of amphotericin B nephrotoxicity by mannitol.

Renal transplant recipients are susceptible to a number of fungal infections amenable to therapy with amphotericin B, but azotaemia is an almost invariable sequel to the use of this agent. As intravenous mannitol has been shown to minimize nephrotoxicity induced by amphotericin B in dogs we treated four kidney transplant recipients who had systemic fungal infections with mannitol and amphotericin B. None showed significant reduction in renal function though a mild metabolic acidosis did develop.     

The effect of amphotericin B on the permeability of lipid bilayers to divalent trace metals

In this study amphotericin B released the divalent trace metals Zn²⁺, Co²⁺, Cu²⁺, Ni²⁺, Mn²⁺, Fe²⁺, Cd²⁺ and Pb²⁺ from multilamellar liposomes containing cholesterol. This observation is consistent with amphotericin B channels being permeable to these metals, and it is proposed, therefore, that the antibiotic may be useful in investigating the metabolism of these elements.     

The effect of amphotericin B on the water and nonelectrolyte permeability of thin lipid membranes

This paper reports the effects of amphotericin B, a polyene antibiotic, on the water and nonelectrolyte permeability of optically black, thin lipid membranes formed from sheep red blood cell lipids dissolved in decane. The permeability coefficients for the diffusion of water and nonelectrolytes (P_DDi) were estimated from unidirectional tracer fluxes when net water flow (J_w) was zero. Alternatively, an osmotic water permeability coefficient (P_f) was computed from J_w when the two aqueous phases contained unequal solute concentrations. In the absence of amphotericin B, when the membrane solutions contained equimolar amounts of cholesterol and phospholipid, P_f was 22.9 ± 4.6 µsec^-1 and P _DDHDH2O was 10.8 ± 2.4 µsec^-1. Furthermore, P_DDi was < 0.05 µsec^-1 for urea, glycerol, ribose, arabinose, glucose, and sucrose, and σ_i, the reflection coefficient of each of these solutes was one. When amphotericin B (10^-6 M) was present in the aqueous phases and the membrane solutions contained equimolar amounts of cholesterol and phospholipid, P _DDHDH2O was 18.1 ± 2.4 µsec^-1; P_f was 549 ± 143 µsec^-1 when glucose, sucrose, and raffinose were the aqueous solutes. Concomitantly, P_DDi varied inversely, and σ_i directly, with the effective hydrodynamic radii of the solutes tested. These polyene-dependent phenomena required the presence of cholesterol in the membrane solutions. These data were analyzed in terms of restricted diffusion and filtration through uniform right circular cylinders, and were compatible with the hypothesis that the interactions of amphotericin B with membrane-bound cholesterol result in the formation of pores whose equivalent radii are in the range 7 to 10.5 A.     

Inhibition of photosynthetic electron transport by amphotericin B

By assaying partial reactions of the photosynthetic electron transport system using thylakoids from spinach as well as from the algae Bumilleriopsis, Dunaliella , and Anabaena , it was demonstrated that the polyene antibiotic amphotericin B has no specific effect on plastocyanin. Pretreating spinach and algal thylakoids with this antibiotic decreased photosystem-II as well as photosystem-I activity regardless of whether the membranes contained plastocyanin or cytochrome c-553. Different sensitivity of cell-free electron transport activity against this antibiotic was observed due to the species used. With Dunaliella , the photosystem-II region was inhibited more strongly than photosystem-I, while Bumilleriopsis chloroplasts – although not containing plastocyanin – exhibited a stronger inhibition of the photosystem-I region. Apparently, amphotericin B mainly solubilizes redox compounds that form connecting pools in the photosynthetic electron transport chain.     

Inhibition of photosynthetic electron transfer by amphotericin B

The polyene antibiotic amphotericin B inhibits photosynthetic electron transfer by Class II maize mesophyll chloroplasts, from water to FeCN, DCIP and diquat but not to plastocyanin. Photosystem 1 activity is also inhibited by amphotericin B, but ferredoxin-NADP reductase activity is not affected. The activity of all the photosynthetic electron transfer systems inhibited by amphotericin B can be restored by the addition of carrier amounts of plastocyanin. The results suggest that amphotericin B inhibits photosynthetic electron transfer by acting only at the plastocyanin site in the chain, and that the primary site of reduction of FeCN and DCIP from water by Class II chloroplasts lies on the reducing side of photosystem 1.     

The effect of tubular damage by mercuric chloride on kidney function and some urinary enzymes in the dog

Alkaline and acid phosphatases, β-glucosidase, β-galactosidase, N-acetyl-β-glucosaminidase and lactate dehydrogenase were monitored in the urine and serum of dogs with renal tubular damage induced by a series of increasing doses of mercuric chloride. Isocitrate dehydrogenase, glutamate oxaloacetate transaminase and glutamate pyruvate transaminase were assayed in the serum. The functional capacity of the kidney was determined by creatinine, inulin and p-aminohippurate (PAH) clearances and tubular maxima tests. Serum urea levels, total protein in the urine and the specific gravity of the urine were monitored throughout the study. The extent and location of the kidney damage was determined by histological investigation. Evidence is presented that the assay of urinary alkaline and acid phosphatase is the most sensitive method of detecting renal tubular damage in the dog. The clearance of [¹⁴C]-propranolol was compared before and after the administration of mercuric chloride. In the presence of tubular damage the blood half-life of propranolol and the rate of excretion of metabolites in the urine were increased.     

Influence of amphotericin B deoxycholate or amphotericin B colloidal dispersion on renal tubule epithelium in rat

Amphotericin B deoxycholate (AmB) or Amphotericin B colloidal dispersion (ABCD) are used in clinics for the treatment of systemic fungal infections. The goal of our study was to compare the nephrotoxicity of these drugs in rat kidney. The effects of AmB and ABCD on the ultrastructure of the epithelium of renal tubules were studied and evaluated using morphometric and statistical methods. Two groups of 3 animals were established: group 1 was treated with AmB desoxycholate and group 2, to which ABCD was applied. AmB caused more than ABCD ultrastructural changes in the cytoplasm of the epithelial cells: damage to mitochondria, vacuolation of cytoplasm, and increased values of volume density of peroxisomes. However, we failed to observe significant differences in morphology and density of the other cell organelles. The proximal tubules seemed to be more sensitive to the nephrotoxic influence of both formulas than the distal tubules of rat kidney. Although, AmB causes more severe damage than ABCD, both drugs cause damage to renal tubuli.