Lysosomes Of Proximal Tubular Cells Research Articles

MO142A SINGLE-CENTER CASE SERIES OF 150 PATIENTS WITH RENAL FANCONI SYNDROME

Abstract Background and Aims Renal Fanconi syndrome (RFS) is characterized by generalized dysfunctions of renal proximal tubular (PT) transport. The causes of FS can be inherited or acquired, the latter of which mostly includes drugs, heavy metals, monoclonal light chains (LC), and primary Sj gren’s syndrome (pSS). We intended to observe the clinico-pathological features of RFS with different etiologies. Method From January 2012 to September 2018, all the patients diagnosed as RFS in our hospital were enrolled. Their clinicopathological records were retrospectively reviewed. The diagnosis of RFS was defined as existence of ≥ 3 of the following five items alone or ≥ 2 items combined with an evidence of PT damages in kidney pathology: (I) normoglycemic glycosuria; (II) generalized aminoaciduria; (III) hypophosphatemia and hyperphosphaturia; (IV) hypouricemia and hyperuricosuria; (V) proximal RTA. Results: 1. Clinical characteristics We identified 150 RFS patients, with an average age of (45.9±14.2) years old and male: female ratio 1.3:1. They presented with different degrees of PT dysfunctions and the most common were hypophosphatemia (83.2%) and aminoaciduria (80.6%). Their mean eGFR levels were 76.3 (4.5-188.6) ml/min/1.73m2 with 73.5% had proteinuria. 2. Renal pathological features 47 RFS patients received kidney biopsy and the most common pathological diagnosis was interstitial nephritis. They all presented with different degrees of proximal tubule atrophy, defective brush border, interstitial edema and fibrosis. 3. Etiology-related clinicopathologic features The most common causes of our RFS were LC (14.0%), drugs (13.3%), and pSS (9.3%). Compared to pSS associated RFS, LC associated RFS patients showed a higher prevalence of bone involvement, more severe proteinuria and less severe hypokalemia (P < 0.05) despite similar eGFR levels. Specific renal pathological features were seen in different etiology groups, including crystalline formation and increased lysosomes in PT cells under electron microscope in LC associated RFS, and CD21 positive ectopic germinal center formation in renal interstitum in parts of the pSS associated RFS. Conclusion We identified 150 RFS with different etiologies and they showed etiology-specific patterns of PT dysfunctions and renal pathologic changes.

A Vaspin\u2013HSPA1L complex protects proximal tubular cells from organelle stress in diabetic kidney disease

Proximal tubular cells (PTCs) are crucial for maintaining renal homeostasis, and tubular injuries contribute to progression of diabetic kidney disease (DKD). However, the roles of visceral adipose tissue-derived serine protease inhibitor (vaspin) in the development of DKD is not known. We found vaspin maintains PTCs through ameliorating ER stress, autophagy impairment, and lysosome dysfunction in DKD. Vaspin−/− obese mice showed enlarged and leaky lysosomes in PTCs associated with increased apoptosis, and these abnormalities were also observed in the patients with DKD. During internalization into PTCs, vaspin formed a complex with heat shock protein family A (Hsp70) member 1 like (HSPA1L) as well as 78 kDa glucose-regulated protein (GRP78). Both vaspin-partners bind to clathrin heavy chain and involve in the endocytosis. Notably, albumin-overload enhanced extracellular release of HSPA1L and overexpression of HSPA1L dissolved organelle stresses, especially autophagy impairment. Thus, vapsin/HSPA1L-mediated pathways play critical roles in maintaining organellar function of PTCs in DKD.

Specific impairment of proximal tubular cell proliferation by a monoclonal light chain responsible for Fanconi syndrome

Fanconi syndrome (FS) is a rare renal disorder featuring proximal tubule dysfunction that may occur following tubular reabsorption of a monoclonal light chain (LC), in patients with multiple myeloma. FS may precede the recognition of multiple myeloma by several years. In most cases, crystalline inclusions of monoclonal κ LCs are observed within the lysosomes of proximal tubular cells (PTCs) and probably participate in their functional alteration. To investigate the mechanism implicated in proximal tubule dysfunction, we compared the effects of κ LC-CHEB obtained from a patient with myeloma-associated FS to those of control κ LC-BON obtained from a patient without evidence of FS, on the viability and proliferation of two different PTC lines. Our data suggest that the tubular atrophy in myeloma-associated FS does not result from increased apoptosis of PTCs, but from their impaired capacity to proliferate and renew. Indeed, in vitro incubation of cultured PTCs with physiological amounts of the nephrotoxic κ LC-CHEB was sufficient to cause a depression in DNA synthesis and in cell proliferation. This effect was observed neither with control κ LC-BON nor in the absence of κ LC. The reduced turnover of PTCs may affect tubular repair and regeneration. In addition, the reduced proliferation of myeloma cells producing the same monoclonal κ LC might explain the frequent association of FS with smoldering multiple myeloma.

Proximal Tubule Cytoplasmic Fibrillary Inclusions Following Kidney Transplantation in a Patient With a Paraproteinemia

Proximal Tubule Cytoplasmic Fibrillary Inclusions Following Kidney Transplantation in a Patient With a Paraproteinemia

Administration of poly-d-glutamic acid induces proliferation of erythropoietin-producing peritubular cells in rat kidney

Erythropoietin (EPO), a 34-kDa glycoprotein, is produced predominantly by peritubular interstitial cells (PIC) in the renal cortex and is physiologically released when ambient oxygen pressure falls. However, the exact nature of EPO-producing cells in the kidney is not well understood. We discovered that brief administration of a low-molecular-weight synthetic peptide, poly-D-glutamic acid (PDG), induced prompt and robust expansion of EPO-producing PIC in rat kidney, without evidence of tubular cell necrosis/apoptosis or fibrotic reaction. Proliferating PIC in PDG-treated rats were noninflammatory, alpha-smooth muscle actin negative, and specifically expressed CD73 (ecto-5'-nucleotidase), EPO mRNA, and protein. Increased numbers of EPO-positive PIC persisted even after the cessation of PDG treatment. No erythropoietic effects of EPO were detected, potentially suggesting maintained physiological control of EPO secretion in this normoxic model. We showed previously that PDG is readily filtered and is rapidly taken up and stored in lysosomes of proximal tubular cells (PTC), resulting in an apparently nonnoxious lysosomal storage condition by virtue of its nonhydrolyzable nature (Kishore BK, Maldague P, Tulkens PM, Courtoy PJ. Lab Invest 74: 1013-1023, 1996). Based on these findings, we suggest that unknown signaling molecules, produced by PTC in response to lysosomal PDG storage, appear to specifically stimulate the proliferation of EPO-producing PIC. We conclude that this model is uniquely suited to investigate the biology of EPO production by PIC and may thus facilitate the development of novel and more economical therapies of anemias and other EPO-responsive conditions.

Protein-rich diet attenuates cyclosporin A-induced renal tubular damage in rats

Objective: The objective of the present study was to look at the effect of a protein-rich diet on cyclosporine A (CsA)-induced acute nephrotoxicity in rodents using markers of tubular damage. Design: Female Sprague-Dawley rats were conditioned to either a standard or a casein-rich diet for 2 weeks. Then, they were given CsA intraperitoneally (25 mg/kg/24 h or an equivalent volume of vehicle (Cremophor EL; Sigma Chemical Co, St. Louis, MO) for 7 days at 7 AM. Results: During CsA treatment, bodyweight, caloric consumption, water intake, and urine output were not significantly different in animals fed with the standard Rat Chow and those on the high-protein feeding. On days 1 and 7, the 24-hour urine excretion of N-acetyl-β-d-glucosaminidase (NAG) and β-galactosidase (β-GAL) were significantly (P <.001) lower in CsA-treated rats on the high-protein diet than in those on the standard Rat Chow. After 7 days of treatment with CsA, no significant difference in the renal function level was found between rats fed with the standard or the casein-rich diet. The post-necrotic cellular regeneration in renal cortex was significantly lower (p<0.001) in CsA-treated rats on the high-protein than on the standard diet. In CsA-treated rats on the standard diet, immunogold labeling showed a massive and specific concentration of the drug into lysosomes of proximal tubular cells. Contrastingly, no gold particle was found over the lysosomes of animals given the rich-protein feeding. Conclusion: In our current experimental conditions, a protective effect of high-casein diet against CsA-induced proximal tubular damage was observed in Sprague-Dawley rats. © 2003 by the National Kidney Foundation, Inc.

Ultrastructural and functional changes of the proximal tubular epithelial cells in the renal cortex from spontaneously diabetic KKAy mice.

To investigate the morphological and functional changes of epithelial cells of proximal tubules in the early stage of diabetic nephropathy, we examined the ultrastructural changes of proximal convoluted tubular cells in spontaneously diabetic KKAy mice. KKAy mice already showed significant elevation of urinary albumin excretion at 16 weeks of age. Examination revealed that lysosomes were increased in number and conspicuous in the epithelial cells of the proximal convoluted tubules from KKAy mice at 40 weeks of age when compared to control C57BL mice. Furthermore, endogenous IgG was detected in the lysosomes of proximal tubular cells from KKAy mice. These findings suggested that reabsorption activity was elevated in the proximal tubules of KKAy mice at 40 weeks of age.

Temporal variation in nephrotoxicity of low doses of isepamicin in rats

The temporal variation in the nephrotoxicity of low doses of isepamicin was studied in male Sprague-Dawley rats treated with a single daily intraperitoneal injection of saline (NaCl, 0.9%) or isepamicin (80 mg/kg of body weight) at either 0800, 1400, 2000, or 0200 h for 4 and 10 days. On day 10, the cellular regeneration (incorporation of [3H] thymidine into DNA of renal cortex) and cortical accumulation of isepamicin were significantly higher in animals treated at 1400 h than at 0200 h (P < 0.01). Immunogold labeling studies showed that isepamicin was essentially localized in the lysosomes of proximal tubular cells in all treated groups, but the density of the gold particles over the lysosomes was higher in animals treated at 1400 than at 0200 h. The results of the present study show that the renal toxicity of isepamicin was maximal at 1400 h (midlight period) and minimal at 0200 h (middark period).

Protection against gentamicin nephrotoxicity by daptomycin in nephrectomized rats

Daptomycin was previously shown to reduce gentamicin renal toxicity and this toxicity was not delayed by the concomitant injection of daptomycin (Thibault N., L. Grenier, M. Simard, M. G. Bergeron, and D. Beauchamp, Antimicrob. Agents Chemother., 38 1027–1035 (1994)). The protective effect of daptomycin against gentamicin toxicity was evaluated in 96 female Sprague-Dawley rats. Normal and nephrectomized rats were treated with saline (NaCl, 0.9%), gentamicin (30 mg/kg/12 hrs, i.p.), daptomycin (10 mg/kg/12 hrs, s.c.) or with a combination of daptomycin plus gentamicin during 4 and 10 days. On day 4, gentamicin and daptomycin cortical levels were higher in nephrectomized gentamicin-daptomycin-treated rats (p < 0.05) as compared to all other groups. The accumulation of gentamicin or daptomycin in nephrectomized gentamicin-daptomycin-treated or gentamicin-saline-treated rats was higher on day 4 (p < 0.01) than on day 10. Other parameters such as the sphingomyelinase activity in the renal cortex, the serum creatinine, and the histopathology showed significantly fewer changes in daptomycin-gentamicin-treated rats as compared to animals given gentamicin alone. On the other hand, the protection of daptomycin was less extensive in nephrectomized rats as compared to normal rats. Daptomycin and gentamicin were localized in the lysosomes of proximal tubular cells of animals treated with daptomycin and gentamicin given alone or in combination. These results suggest that daptomycin protects against gentamicin toxicity in nephrectomized rats but to a lesser extent than in normal rats.

Attenuation by daptomycin of gentamicin-induced experimental nephrotoxicity.

Previously, daptomycin was shown to reduce tobramycin nephrotoxicity in vivo (D. Beauchamp, M. Pellerin, P. Gourde, M. Pettigrew, and M. G. Bergeron, Antimicrob. Agents Chemother. 34:139-147, 1990; C. A. Wood, H. C. Finkbeiner, S. J. Kohlhepp, P. W. Kohnen, and D. C. Gilbert, Antimicrob. Agents Chemother. 33:1280-1285, 1989). Female Sprague-Dawley rats were treated with saline (NaCl, 0.9%), daptomycin (10 mg/kg of body weight every 12 h, subcutaneously), gentamicin (30 mg/kg/12 h, intraperitoneally) or with a combination of daptomycin plus gentamicin over a 10-day period. Animals were killed 4, 10, and 20 days after the end of treatment. Four days after the end of drug administration, gentamicin and daptomycin levels in the renal cortices of animals treated with the combination of daptomycin and gentamicin were significantly higher than in those of rats given gentamicin or daptomycin alone (P < 0.01). Despite the higher cortical concentrations of gentamicin, rats given the combination of gentamicin and daptomycin had less reduction in renal cortex sphingomyelinase activity, less evidence of regeneration of cellular cortical cells ([3H]thymidine incorporation into cortex DNA), lower creatinine concentration in serum, and less histopathologic evidence of injury than rats given gentamicin alone. By immunogold technique, both daptomycin and gentamicin were localized to the lysosomes of proximal tubular cells, regardless of whether animals received the drugs alone or in combination. Interestingly, myeloid body formation occurred in both those animals given gentamicin alone and those given daptomycin plus gentamicin. No significant changes were observed for all groups between 10 and 20 days after the end of therapy, suggesting that the toxicity of gentamicin was not delayed by the concomitant injection of daptomycin. The results confirm that daptomycin can attenuate experimental gentamicin nephrotoxicity.

Daptomycin may attenuate experimental tobramycin nephrotoxicity by electrostatic complexation to tobramycin.

The lipopeptidic antibiotic daptomycin is reported to reduce experimental tobramycin nephrotoxicity (D. Beauchamp, M. Pellerin, P. Gourde, M. Pettigrew and M. G. Bergeron, Antimicrob. Agents Chemother. 34:139-147, 1990; C. A. Wood, H. C. Finkbeiner, S. J. Kohlhepp, P. W. Kohnen, and D. C. Gilbert, Antimicrob. Agents Chemother. 33:1280-1285, 1989). In an attempt to explain these results, the in vivo and in vitro interactions between daptomycin and tobramycin were studied. Tobramycin alone and preincubated with negatively charged phospholipid bilayers (liposomes) was dialyzed against increasing concentrations of daptomycin in buffer at pH 5.4. A significant drop in the concentration of tobramycin was observed when daptomycin was added to the opposite half cells. Furthermore, daptomycin induced a concentration-dependent release of lipid-bound tobramycin. Gold labeling experiments showed that daptomycin could be incorporated into phospholipid layers. Female Sprague-Dawley rats were treated with daptomycin alone, with tobramycin alone, or with the combination over 2 to 10 days. Levels of daptomycin and tobramycin in serum were similar in all groups. The levels of tobramycin in the renal cortex increased significantly with time and, on day 10, reached values of 654 +/- 122 and 844 +/- 298 micrograms/g of tissue (mean +/- standard deviation; not significant) in animals treated with tobramycin and the combination of daptomycin-tobramycin, respectively. No significant difference was observed in the levels of tobramycin in the kidneys between animals treated with tobramycin or the daptomycin-tobramycin combination at any time. By contrast, daptomycin levels were significantly higher in the renal cortexes of animals treated with daptomycin-tobramycin in comparison with those in the renal cortexes of animals treated with daptomycin alone on days 6,8, and 10 (P < 0.01). For immunogold labeling studies, animals were killed 4 h after a single injection of daptomycin alone or daptomycin in combination with tobramycin. Daptomycin was found throughout the matrixes of the lysosomes of proximal tubular cells of animals treated with daptomycin alone. In animals treated with the combination of daptomycin and tobramycin, daptomycin was associated with intralysosomal myeloid bodies. Our results suggest that daptomycin might attenuate experimental aminoglycoside nephrotoxicity by interacting with the aminoglycoside, perhaps electrostatically, and thereby protecting intracellular targets of toxicity.

Subcellular distribution of daptomycin given alone or with tobramycin in renal proximal tubular cells.

Previous studies in experimental animals showed that daptomycin, a lipopeptide antibiotic, protects against aminoglycoside nephrotoxicity (C. A. Wood, H. C. Finkbeiner, S. J. Kohlhepp, P. W. Kohnen, and D. N. Gilbert, Antimicrob. Agents Chemother. 33:1280-1285, 1989; D. Beauchamp, M. Pellerin, P. Gourde, M. Pettigrew, and M. G. Bergeron, Antimicrob. Agents Chemother. 34:139-147, 1990). In order to better understand the mechanism involved in this protective effect, the subcellular distribution of daptomycin was investigated in the proximal tubular cells of animals treated with daptomycin alone or in combination with tobramycin. A first group of female Sprague-Dawley rats received a single intravenous injection of daptomycin at a dose of 100 mg/kg of body weight and were killed at 10 min, 1 h, or 24 h after the injection. Other groups of rats were treated during 10 days with saline (NaCl, 0.9%), tobramycin at dosages of 20 mg/kg/12 h, daptomycin at dosages of 10 mg/kg/12 h, or the combination tobramycin-daptomycin at the same dosages. At the time of sacrifice, the renal cortex of the right kidney of each animal was dissected, and small blocks of tissue were fixed, dehydrated, and embedded in Araldite 502 epoxy resin. The subcellular distribution of daptomycin and tobramycin was determined on ultrathin sections by immunogold labeling. Ten minutes after the injection of daptomycin alone, gold particles were seen over the brush border membrane and on the membranes of the endocytic vacuoles of proximal tubular cells. One hour after the injection, a similar distribution was seen and numerous gold particles were found over the lysosomes of proximal tubular cells. The results suggest that daptomycin might protect against aminoglycoside nephrotoxicity by interfering with the interaction between the aminoglycoside and phospholipids inside the lysosomes of proximal tubular cells.

Subcellular localization of tobramycin and vancomycin given alone and in combination in proximal tubular cells, determined by immunogold labeling

The subcellular localization of tobramycin and vancomycin in the renal cortices of rats was determined with ultrathin sections by immunogold labeling. Four groups of four rats each were treated for 10 days with saline (NaCl, 0.9%), tobramycin at dosages of 20 mg/kg of body weight per 12 h intraperitoneally, vancomycin at dosages of 25 mg/kg/12 h subcutaneously, or the combination tobramycin-vancomycin. On day 11, the animals were killed, and cubes of renal cortex were fixed overnight in phosphate-buffered glutaraldehyde (0.5%), dehydrated in ethanol, and embedded in Araldite 502 resin. Ultrathin sections were made and incubated with sheep antitobramycin antibody followed by protein A-gold (15-nm diameter) complex or rabbit antivancomycin antibody followed by gold (30-nm diameter)-labeled goat anti-rabbit antibody. For the double labeling, incubations were made on opposite sides of the grid. Tobramycin was detected over the lysosomes of proximal tubular cells, but the labeling was concentrated into small areas in the matrix of the lysosomes. Vancomycin was seen over the lysosomes of proximal tubular cells and was distributed uniformly throughout the matrix of the lysosomes. In rats treated with tobramycin-vancomycin, both drugs were still detected in lysosomes of proximal tubular cells. It is concluded that tobramycin and vancomycin accumulate in lysosomes of proximal tubular cells throughout 10 days of treatment and that vancomycin has no effect on the subcellular distribution of tobramycin.

Renal specific delivery of sulfamethoxazole in the rat by coupling to the low molecular weight protein lysozyme via an acid-sensitive linker

Sulfamethaxazole (SM) was converted to a renal specific drug targeting preparation by coupling the drug to egg-white lysozyme via an acid-sensitive cis-aconityl linker (1:1). Due to this chemical manipulation SM was rapidly distributed to the kidney. Both in vitro and in vivo data indicate that SM was uncoupled from the carrier by chemical hydrolysis in the lysosomes of proximal tubular cells, resulting in parent active drug at the target site. This concept is applicable to other drug-polypeptide conjugates which rapidly distribute to the kidney and might enable selective manipulation of renal (patho)physiology.

Subcellular distribution of gentamicin in proximal tubular cells, determined by immunogold labeling

The subcellular distribution of gentamicin in rat renal proximal tubular cells was evaluated by immunogold labeling. The distribution of the drug was monitored from 10 min to 10 days following single (40 mg/kg of body weight) and multiple (5 and 20 mg/kg/12 h) injections of gentamicin. Animals were killed on day 11, and cubes of renal cortex tissue were fixed overnight in cold phosphate-buffered glutaraldehyde (0.5%), dehydrated in ethanol, and embedded in Araldite 502 epoxy resin. Ultrathin sections were made and incubated with sheep antigentamicin and then with protein A-gold (15 nm) complex. At 10 min after a single injection, the labeling was found over the brush border membrane and over the membranes of endocytic apical vesicles of proximal tubular cells. After 1 h, a similar distribution was observed and the labeling was also seen over small lysosomes located close to the brush border membrane. At 24 h, gold particles were found over large lysosomes of proximal tubular cells. Following 10 days of treatment, lysosomes of proximal tubular cells were densely labeled with gold particles. The labeling was distributed uniformly over the lysosomes, although a lower density of labeling was observed over the myeloid bodies inside the lysosomes. Necrotic proximal tubular cells showed labeling over intact lysosomes and also in the cytoplasms of the cells, in the mitochondria, and in the nucleoli. The various control experiments demonstrated the high specificity of these results. The present immunocytochemical study better documents the subcellular disposition of gentamicin in proximal tubular cells, as previously evaluated by subcellular fractionation and autoradiography. This technique will be useful for better understanding the relationship between drug disposition and drug-induced toxicity.

Experimental studies on nephrotoxicity of aminoglycosides at low doses: Mechanisms and perspectives

The nephrotoxicity of aminoglycosides is an important consideration in the clinical use of these agents. The underlying biochemical and cytologic mechanisms have, therefore, been the subject of many extensive studies. Few experimental studies, however, have employed doses relevant to those used in clinical practice. This article focuses on data obtained using such a “low-dose” approach. Aminoglycosides accumulate specifically in the lysosomes of proximal tubular cells by a process of adsorptive pinocytosis; in the lysosomes, they induce a phospholipidosis that has been studied in vivo and in vitro (e.g., by computer-aided conformational analysis). A similar phospholipidosis is also observed in humans. The overall nephrotoxicity of aminoglycosides results from a combination of their intrinsic ability to bind to phospholipids and the extent of their renal uptake. The development of phospholipidosis is accompanied by focal necroses, tubular regeneration, and interstitial proliferation, even at low, therapeutic doses of these agents. These changes are related to the nephrotoxic potential of the aminoglycosides. Of all 2-deoxystreptamine-containing aminoglycosides tested so far, and by all criteria, amikacin is associated with the least dramatic alterations. Together, these approaches may provide a rational basis to study and compare the nephrotoxicity of aminoglycosides at clinically relevant dosages. They may also serve for the screening and design of aminoglycosides with lower toxicity than those currently available.

Recovery of cortical phospholipidosis and necrosis after acute gentamicin loading in rats

The recovery from gentamicin-induced phospholipidosis in the rat kidney cortex was characterized both morphologically and biochemically after a single 12-hr drug infusion. Total dosages administered were 10, 60, or 140 mg/kg, achieving constant serum concentrations of 3, 11, and 27 micrograms/ml, respectively. At the end of the 12-hr infusion, the cortical drug concentrations corresponding to the three dosages were 124, 450, and 993 micrograms/g of wet tissue. At the low dose (10 mg/kg), myeloid bodies were seen inside lysosomes of proximal tubular cells, along with a modest decrease of lysosomal sphingomyelinase activity. The cortical drug level declined steadily following first-order kinetics along with a disappearance of myeloid bodies and return of sphingomyelinase activity to control levels. At the high dose (140 mg/kg), we observed a sustained loss of sphingomyelinase activity (37% of controls), a subsequent increase of phospholipid concentration in the kidney cortex (up to 117% of controls 2 days after) and a prominent accumulation of myeloid bodies inside the lysosomes of proximal tubular cells (up to 4% of cell volume). Tubular regeneration and interstitial infiltration became detectable by histology and the increase of DNA synthesis as from day 1, along with an apparent reduction of the phospholipidosis at days 3 and 4. Drug cortical concentrations showed a sharp decline 2 days after infusion. An intermediate behavior was observed at 60 mg/kg. It is concluded that the proximal tubular cells behave in a fundamentally different way after gentamicin loading with low and high doses. At the low dose there is a regression of the drug-induced changes in the absence of any sign of necrosis-regeneration. Above a threshold in cortical drug concentration there is further development of the alterations leading to cell death-regeneration.

Mechanism of aminoglycoside-induced lysosomal phospholipidosis: In vitro and in vivo studies with Gentamicin and Amikacin

Gentamicin, a widely used aminoglycoside antibiotic, is concentrated in lysosomes of proximal tubular cells of the kidney, and induces therein an accumulation of myelin-like material. We show that treatment of rats with Gentamicin (10 mg/kg, 7 days) induces a loss of activity of lysosomal sphingomyelinase and phospholipase A 1, associated with an increase in the amount of total lipid phosphorus in the kidney cortex. In vitro, Gentamicin is shown by gel permeation to bind to phospholipid bilayers (liposomes) under conditions which mimic the lysosomal environment (acid pH and presence of phosphatidylinositol). The reversal of this binding by an increase in the ionic strength (> 0.04) suggests electrostatic interaction between the hydrophilic, polycationic aminoglycoside and the negatively charged phospholipids. Binding of Gentamicin impairs the hydrolysis of phosphatidylcholine present in the bilayer, by lysosomal phospholipases A 1 and A 2 from the liver or kidney. We also show that lysosomal sphingomyelinase is readily and irreversibly inactivated by liposomes in the absence of detergent. The lysosomal phospholipidosis induced by Gentamicin in the kidney, as in cultured cells [Aubert Tulkens et al., Lab. Invest. 40, 481 (1979)] appears therefore to be a direct consequence of the lysosomotropic character of this drug and its ability to inhibit therein phospholipid breakdown. Amikacin, a semi-synthetic aminoglycoside, binds more loosely to phospholipid bilayers, induces less inhibition of phospholipases in vitro and is less taken up by tubular cells in vivo. Accordingly, Amikacin does not provoke significant lysosomal phospholipidosis or loss of sphingomyelinase and phospholipase A 1 activities in vivo at the doses and time investigated (0–40 mg/kg, 7 days). Inasmuch as Amikacin is reported to be less toxic to the kidney, we suggest that lysosomal alterations are an early and significant

Effects of selenium on mercury-induced renal lesions and on subcellular mercury distribution

The effects of selenite on the renal tubular lesion caused by mercuric chloride were studied by electron microscopy using male rats injected with HgCl 2 and/or Na 2SeO 3. The protective effect of selenite on mercury-induced renal injuries was confirmed both in proximal tubular cells and in glomerulus. Mercury specifically deposited in lysosomes in proximal tubular cells of rats which received either mercury alone or mercury plus selenite, although the density of fine gold grains, which revealed the mercury deposits, appeared to be higher in the rats treated with mercury alone than in those administered mercury and selenite. These observations by electron microscopic histochemistry were consistent with the analytical data for mercury in the kidneys. The results suggest that the reduction of renal mercury toxicity by selenite should be ascribed not to the change in subcellular mercury localization but to the decrease in the level of mercury in kidneys.