Biomedical subjects

P H Bach

Publications and source records attributed to P H Bach.

At least 19 recordsLinked to original sources

PubMed2001

Adenine nucleotide and calpain inhibitor I protect against atractyloside-induced toxicity in rat renal cortical slices in vitro.

Atractyloside is a compound with a documented nephrotoxicity. It induces renal tubular necrosis at high doses and apoptosis at lower doses. This study investigates the potential protective effect of some chemical agents against atractyloside-induced nephrotoxicity in vitro using the precision-cut rat renal cortical slices obtained from kidneys of Wistar rats. For co-incubation experiments, slices were incubated for 3 h at 37 degrees C on a rocker platform with various chemical agents: ADP (5 mM), calpain inhibitor I (CPI, 1 mM), stevioside (STV, 2.5 mM) or probenecid (PRB, 2.5 mM) in the presence or absence of atractyloside (2 mM). For pre-incubation experiments, slices were incubated with the same chemical agents for 1 h before exposure to atractyloside. The nephrotoxic effects of atractyloside (2 mM) alone were manifested in several ways: by a marked increase in lactate dehydrogenase (LDH) and alkaline phosphatase (ALP) leakage, significant inhibition of p-aminohippurate (PAH) accumulation, marked depletion of intracellular ATP and reduced glutathione (GSH), and a significant reduction in pyruvate-stimulated gluconeogenesis. Co-incubation of slices with ADP or CPI and atractyloside completely blocked atractyloside-induced increase in LDH leakage, but not ALP leakage. Atractyloside-induced depletion of ATP and reduced gluconeogenesis was prevented by co-incubation with ADP or CPI. Furthermore, co-incubation of slices with STV and atractyloside, but not PRB, completely abolished atractyloside-induced depletion of ATP and decreased gluconeogenesis in the slices. Pre-incubation of slices with either ADP or CPI protected against atractyloside-induced increase in LDH leakage, reduced ATP and decreased gluconeogenesis. PAH uptake in the slices was inhibited by atractyloside and PRB in a time-dependent manner. While ADP and CPI were found to exert complete protection against atractyloside-induced toxicity irrespective of treatment schedule, STV is effective only under certain conditions, and PRB offer no protection at all. The results of this study demonstrate the usefulness of renal cortical slices as toxicology tool for evaluating and screening compounds for their potential protective effects, and are supportive of a role of adeninine nucleotide (ADP) and protease inhibitor (CPI) in protecting against atractyloside-induced cell injury.

Adenine Nucleotides↗

PubMed2001

Effects of the calcium channel blocker verapamil and sulphydryl reducing agent dithiothreitol on atractyloside toxicity in precision-cut rat renal cortical and liver slices.

The effects of dithiothreitol (DTT), a sulfhydryl-containing agent and verapamil (VRP), a calcium channel blocker as possible cytoprotectants against the atractyloside-induced toxicity were characterized in rat kidney and liver slices in vitro using multiple markers of toxicity. Precision-cut slices (200 microM thick) were either incubated with atractyloside (2 mM) or initially preincubated with either DTT (5 mM) or VRP (100 microM) for 30 min followed by exposure to atractyloside (2 mM) for 3 h at 37 degrees C on a rocker platform rotated at approximately 3 rpm. All of the toxicity parameters were sensitive to exposure to atractyloside, but treatment with DTT or VRP alone did not provide any indication of damage to the tissues. Preincubation of slices containing either DTT or VRP for 30 min provided total protection against atractyloside-induced increase in LDH leakage in both kidney and liver slices. Increased induction of lipid peroxidation by atractyloside in liver slices was completely abolished by DTT and VRP. Both DTT and VRP provided partial protection against atractyloside-induced inhibition of gluconeogenesis in both kidney and liver slices. Atractyloside-induced ATP depletion in both kidney and liver slices was partially abolished by VRP but not DTT. The significant depletion of GSH in the kidney slices by atractyloside was completely reversed by DTT only, while VRP alone reversed the same process in liver slices. Decreased MTT reductive capacity and significant increase in ALT leakage caused by atractyloside in liver slices was partially reversed. Complete protection was achieved with both DTT and VRP against atractyloside-induced inhibition of PAH uptake in kidney slices. These findings suggest that both DTT and VRP exert cytoprotective effects in atractyloside-induced biochemical perturbation, effects that differ in liver and kidney. The effect of these agents on atractyloside has provided us with a further understanding of the molecular mechanism of its action.

Adenosine Triphosphate↗

PubMed2001

Increased urinary uronic acid excretion in experimentally-induced renal papillary necrosis in rats.

We have evaluated the potential of urinary uronic acid measurement as an early indicator in the development of renal papillary necrosis (RPN). Urinary uronic acid was quantified with a range of other urinary biochemical parameters in rats given multiple doses of N-phenylanthranilic acid (NPAA) or mefenamic acid (MFA), each of which induces a dose-related papillary necrosis. In addition, histological examination was also carried out to confirm the development and presence of RPN. NPAA was administered to male wistar rats at p.o. doses of 100, 250, and 500 mg/kg and MFA at p.o. doses of 75, 150, and 300 mg/kg on days 1-4 and 8-11, and urine samples were collected for 16 hours each day. NPAA increased uronic acid excretion two-fold for both medium and high doses from day four. MFA increased uronic acid excretion to two and a half-fold by day 10 in the highest dose administered. Urinary creatinine was equally elevated in a dose-related manner following treatment with either NPAA or MFA. None of the other routine markers (urinary or serum) of nephrotoxicity showed any statistical changes. NPAA produced a dose- and time-related increase in excretion of uronic acid. Evidence of widespread papillary necrosis was seen histologically at the high doses of NPAA or MFA. The significant elevation of uronic acid in urine following treatment with either NPAA or MFA was well ahead of the development of RPN detectable by routine histology, suggesting that uronic acid measurement could serve as an early indicator of RPN. The assessment of urinary uronic acid may therefore provide a novel sensitive and selective marker of identifying the lesion earlier than is currently possible. An increase in urinary uronic acid following NPAA and MFA treatment supports the biochemical basis of these changes as a representative of acid mucopolysaccharides accumulation.

Animals↗

PubMed2001

Integrating metabolism and toxicity in multi-organ systems.

A wide range of experimental and predictive approaches are currently employed by the pharmaceutical industry in an attempt to avoid developing molecules with likely toxicities. Experimental systems may be adapted to handle the increasing number of molecules that require screening. The cost implication of such extensive screening is, however, immense and in silico models must therefore become the first lines of investigation to assess the metabolic and toxicological potential of new compound libraries. In silico systems have been based on the relationship between the chemical structure of novel molecules and toxicological outcomes in terms of types, but not always, locations of lesions. There should be sufficient published and in-house data available on the distribution of diverse molecules to identify the rules that govern organ, cellular and subcellular distribution of molecules (and their metabolites) as the basis of tissue and cell targeting and, more importantly, the possibility of toxicological processes. In addition, the extensive cell biology knowledge needs to be linked to current in silico systems to enhance our understanding of metabolic activation and other processes that affect likely toxicological consequences in different tissues and specific cell types. There is a need to provide a single knowledge platform that can access all current in silico products that use different datasets. The closer integration of these diverse tools will help predict other approaches to help identify the most likely toxicological potential of novel molecules, and ensure the 'least' toxic molecules are developed.

Animals↗

PubMed2000

Atractyloside nephrotoxicity: in vitro studies with suspensions of rat renal fragments and precision-cut cortical slices.

The consumption of plants containing atractyloside, a diterpenoid glycoside, causes selective proximal tubule injury leading to renal failure and death in humans. The underlying mechanisms responsible for its toxicity are still not well understood. The present study was therefore carried out to determine the mechanism and the exact sequence of events that lead to molecular toxic injury. A comparative study using renal cortical slices, suspension of freshly isolated renal proximal tubular fragments and glomeruli of male Wistar rat was made. These in vitro systems were exposed to 100-1000 mM atractyloside for 2-3 h at 37 degrees C. Atractyloside caused a significant alteration in various toxicity parameters in a concentration- and time-dependent manner in renal cortical slices and proximal tubular fragments, but not in glomeruli. The earliest change following exposure to atractyloside (1000 microM) was a significant reduction of intracellular adenosine 5'-triphosphate (ATP) content occurring within 1 h in the tubules and 2 h in slices. The significant depletion of reduced glutathione (GSH) inhibitor of p-aminohippuric (acid) (PAH) uptake and gluconeogenesis occurred simultaneously following loss of cellular energy. These events were only limited to the renal cortical slices and proximal tubular fragments. Increased severity of cellular injury resulted in cytotoxicity with the significant increase in the leakage of alkaline phosphatase (ALP) and lactate dehydrogenase (LDH) in proximal tubular fragments (occurring at 2 h) and renal cortical slices (occurring at 3 h). There were, however, no alterations in oxidized glutathione (GSSG) levels or in the ratio of GSH/GSSG. Only limited lipid peroxidation in proximal tubular fragments and glomeruli was observed at atractyloside concentrations of 500 microM and above. In all cases of toxicity, the glomeruli were unaffected. Pretreatment of slices or fragments with probenecid (1.0 mM) failed to completely abolish atractyloside toxicity. These data demonstrate dose- and time-dependent toxicity of atractyloside and clearly confirmed the proximal tubular fragments as the target tissue. Atractyloside exhibits a toxicity profile that indicates early alteration in mitochondrial function and consequently loss of cellular energy, followed by reduced metabolic function and transport processes and ultimately cell death. This appears to be the most likely mechanism by which atractyloside exerted its acute cytotoxicity. Renal cortical slices, which maintain proximal tubule and glomeruli in their anatomic relationship, responded similarly to atractyloside toxicity as the proximal tubular fragments, and might be suggested as the most suitable in vitro model system for studying the mechanisms of atractyloside toxicity as they are more likely to mirror changes seen in the whole organ.

Adenosine Triphosphate↗

PubMed1999

The role of ursodeoxycholic acid in bile acid-mediated kidney fragment toxicity.

Elevated levels of bile acids are thought to play an important role in the renal failure of patients with obstructive jaundice undergoing surgery. In contrast, ursodeoxycholic acid (UDA) is widely used to improve cholestasis and has been proposed as protective bile acids and antioxidant. The present study employs kidney fragments to determine the role of reactive oxygen species (ROS) in the mechanism of toxicity of hydrophobic bile acids and to determine the nephroprotectant properties of UDA against the hydrophobic bile acids. The hydrophobic bile acids chenodeoxycholic (200 microM) and deoxycholic acid (200 microM) significantly (P<0.05) increased lactate dehydrogenase leakage (LDH) from glomerular fragments from 2.7+/-0.4 to 5.03+/-0.23 and 4.66+/-0.37 (micromol NADH consumed/min/mg protein) for chenodeoxycholic and deoxycholic acid respectively. Preincubating the fragments with UDA (500 microM) did not prevent the leakage of LDH caused by the bile acids. The level of lipid peroxidation was not increased in fragments exposed to either ursodeoxycholic (0-500 microM), lithocholic (0-100 microM), chenodeoxycholic (0-500 microM) or deoxycholic acid (0-500 microM). Furthermore UDA (500 microM) did not prevent the increase in lipid peroxidation caused by tert-butyl hydroperoxide (0-1000 microM) in the fragments. These results suggest that hydrophobic bile acids do not cause lipid peroxidation in kidney fragments and that UDA is neither capable of preventing the loss of membrane integrity induced by hydrophobic bile acids or acting as an antioxidant in kidney fragments.

Animals↗

PubMed1998

Toxicity of atractyloside in precision-cut rat and porcine renal and hepatic tissue slices.

Atractyloside (ATR) causes acute fatal renal and hepatic necrosis in animals and humans. Precision-cut renal cortical and hepatic slices (200 +/- 15 microns) from adult male Wistar rat and domestic pigs, incubated with ATR (0.2-2.0 mM) for 3 h at 37 degrees C, inhibited pyruvate-stimulated gluconeogenesis in a concentration- and time-dependent manner. p-Aminohippurate accumulation was significantly inhibited in both rat and pig renal cortical slices from 0.2 mM ATR (p < 0.05). There was a small decrease in mitochondrial reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium to formazan in both rat and pig kidney slices, which was significant at > or = 2 mM, but no changes in liver slices from either species. However, cellular ATP was significantly depleted at > or = 0.2 mM ATR in kidney and in liver slices from both species. ATR also caused a marked leakage of lactate dehydrogenase and alkaline phosphatase from both pig and rat kidney slices at all concentrations, but only lactate dehydrogenase was significantly elevated in liver slices from both species. ATR > or = 0.5 mM caused a significant increase in lipid peroxidation, but only in liver slices of both species, and > or = 0.2 mM ATR caused a marked depletion of reduced glutathione and significant increase in oxidized glutathione in both kidney and liver slices of both species. However, GSH to GSSG ratio was only significantly altered in the liver slices, indicating that oxidative stress may be the cause of toxicity in this organ. Both rat and pig tissue slices from the same organ responded similarly to ATR, although their basal biochemistry was different. ATR toxicity to both kidney and liver showed similar patterns but it appears that the mechanisms of toxicity are different. While cytotoxicity of ATR in kidney is only accompanied with GSH depletion, that of the liver is linked to both lipid peroxidation and GSH depletion. Striated muscle slices from both species were not affected by the highest ATR concentration. This further strengthens the argument that the molecular basis of ATR, target selective toxicity, is not a measure of the interaction between ATR and mitochondria and that other factors such as selective uptake are involved. Precision-cut tissue slices show organ-specific toxicity in kidney and liver from both rat and pig and suggest different mechanisms of injury for each organ.

Adenosine Triphosphate↗

PubMed1998

Biochemistry and toxicology of the diterpenoid glycoside atractyloside.

Atractyloside (Atr) is a diterpenoid glycoside that occurs naturally in plants (many of which are used in ethnomedicines) found in Europe, Africa, South America, Asia and the far East. It is also present in animal grazing forage. Atr (and its analogues) may be present at levels as high as 600 mg/kg dried plant material. Consumption of the plants containing Atr or carboxyatractyloside (carboxyAtr) has caused fatal renal proximal tubule necrosis and/or centrilobular hepatic necrosis in man and farm animals. Although pure Atr and crude plant extracts disrupt carbohydrate homeostasis and induce similar pathophysiological lesions in the kidney and liver, it is also possible that the toxicity of Atr may be confounded by the presence of other natural constituents in plants. Atr competitively inhibits the adenine nucleoside carrier in isolated mitochondria and thus blocks oxidative phosphorylation. This has been assumed to explain changes in carbohydrate metabolism and the toxic effects in liver and kidney. Although the acute toxicity of Atr is well described, many aspects of Atr toxicity (subchronic and chronic toxicity, reproductive toxicity, mutagenicity and carcinogenicity) have not been investigated and pharmacokinetic and metabolism data are limited. In vitro proximal tubular cells are selectively sensitive to Atr, whereas other renal cell types are quite resistant. There are also differences in the response of liver and renal tissue to Atr. Thus, not all of the clinical, biochemical and morphological changes caused by Atr can simply be explained on the basis of inhibition of mitochondrial phosphorylation. The relevance to a wider human risk is shown by the presence of Atr analogues in dried roasted Coffea arabica beans (17.5 32 mg/kg). There are no data to help identify the risk of low dose chronic exposure in human coffee consumers, nor is there information on the levels of Atr or its analogues in other commonly consumed human foodstuffs.

Animals↗

PubMed1998

The role of reactive oxygen species in adriamycin and menadione-induced glomerular toxicity.

Redox cycling leading to oxidative stress has been proposed as the mechanism by which adriamycin induces glomerular toxicity in rats. The present study compares the extent of the oxidative stress and cytotoxicity induced by adriamycin to menadione (a model redox cycling quinone) in freshly isolated rat glomeruli. Adriamycin and menadione (25 microM) decreased de novo protein synthesis (measured by 3H-proline incorporation into acid-precipitable glomerular protein) by 50 and 85%, respectively, in 2 h. By contrast, menadione at 25 microM reduce glomerular membrane integrity (as assessed by lactate dehydrogenase leakage), adriamycin reduced membrane integrity at 500 microM adriamycin. Reactive oxygen species (ROS) were measured by the oxidation of dihydrodichlorofluorescein. Menadione (25 microM) and adriamycin (25 microM) increased ROS formation to 260 and 156% of controls after 30 min incubation, respectively. Oxidative stress was assessed by measuring the intracellular level of reduced glutathione (GSH) and the decrease of the NADPH/NADP- ratio which stimulates the pentose phosphate pathway (PPP): (a) menadione (25-100 microM) reduced glomerular GSH to 10-20% of controls, adriamycin (25-100 microM) had no effect; (b) menadione (10 microM) increased PPP activity 6-fold, while adriamycin (125 microM) had only a 2-fold effect. Although adriamycin and menadione generate extensive ROS and decrease protein synthesis, there was no correlation between the extent of oxidative stress and cytotoxicity in glomeruli exposed to adriamycin. These results suggest that oxidative stress may not be the primary mechanisms by which adriamycin induces selective glomerular toxicity.

Animals↗

PubMed1998

The relationship between sodium chloride concentration and bile acid cytotoxicity in cultured kidney cells.

Patients with obstructive jaundice suffer an increased incidence of mortality from post operative renal failure, which may be related to elevated circulating bile salts. This study assesses the effects of increased ionic strength (similar to that found in the kidney inner medulla) on bile salt critical micellar concentration (CMC) and cytotoxicity to renal medullary epithelial primary cultures and MDCK and NRK cell lines representing the distal and proximal tubular cells respectively. The CMC of chenodeoxycholic acid decreased from 2.86 +/- 0.07 (in isotonic Earle's Hepes buffer) to 2.30 +/- 0.07, 1.99 +/- 0.09 and 1.46 +/- 0.08 mM following the addition of 150, 250 and 500 mM NaCl. Similarly, the CMC of deoxycholic acid was reduced from 3.18 +/- 0.1 to 2.84 +/- 0.1, 2.26 +/- 0.1 and 1.79 +/- 0.09 mM by the addition of 150, 250 and 500 mM NaCl. Increasing the ionic strength of the culture medium of medullary epithelial cells by the addition of 150 mM NaCl, decreased viability by 39% (p < 0.01), 24% (p < 0.001) and 40% (p < 0.001) for lithocholic (25 microM), chenodeoxycholic (100 microM) and deoxycholic acids (100 microM), respectively. A similar increase in the ionic strength of the culture medium of MDCK cells decreased viability by 79% (p < 0.01), 46% (p < 0.01) and 15% (p < 0.01) for lithocholic (15 microM), chenodeoxycholic (100 microM) and deoxycholic (50 microM), respectively. Adding 200 mM urea to medium supplemented with 150 mM NaCl (to further increase osmolality but not ionic strength) had no effect on the cytotoxicity bile salts in MDCK cells. The addition of 150 mM NaCl to the culture medium of NRK cells resulted in a decrease viability of 15% (p < 0.01), 27% (p < 0.01) and 60% (p < 0.01) following exposure to either lithocholic (15 microM), chenodeoxycholic (50 microM) or deoxycholic acids (50 microM) respectively. These results show that increasing NaCl concentrations lowers CMC of bile salts and increase cytotoxicity in medullary epithelial primary, MDCK and NRK cells. This suggests that the high NaCl levels in the kidney inner medulla would reduce bile salt CMC such that they could damaged renal cells. This may, in part, explain the increased susceptibility of the kidney during obstructive liver disease.

Animals↗

PubMed1998

A safety assessment of fixed combinations of acetaminophen and acetylsalicylic acid, coformulated with caffeine.

Overuse and abuse of phenacetin-containing mixed analgesics has contributed to end-stage renal disease. Combination analgesics, especially those coformulated with caffeine, have been implicated as imparting a greater risk of analgesic-associated nephropathy (AAN) than single or coformulated analgesics without caffeine. This has led to a recommendation that the sale of "two plus caffeine" analgesic mixtures be reclassified from over-the-counter to prescription only availability. There is a rational basis for coformulating acetylsalicylic acid (ASA) and acetaminophen (paracetamol) as this reduces the dose of each, without altering efficacy. The coformulation of caffeine with these analgesics has a significant adjuvant effect and increases analgesic efficacy 1.4-1.6-fold. Currently available animal and human data do not support the notion that the nephrotoxic risk from coformulated ASA and acetaminophen is higher than the risk from either ASA or acetaminophen alone, in equivalent analgesic doses. There are no epidemiological data that implicate caffeine in AAN, and only limited evidence that links excessive acetaminophen usage to renal disease. There is no evidence that caffeine increases analgesics papillotoxicity directly. The presence of caffeine in mixtures of analgesics are no more addictive than other sources of caffeine. There is no evidence to suggest that adding caffeine to analgesic mixtures enhances the potential for promoting analgesic misuse in the general population. Thus distinct therapeutic benefits of ASA, acetaminophen and caffeine appear to outweigh any known risk. It is doubtful if preventing the availability of these products will significantly affect the role of analgesic abuse/overuse in end-stage renal disease. Better risk management would come from a focused educational program, developed in a close collaboration between industry, healthcare professionals and consumer organizations, such a program must warn against the potential dangers of analgesic and non-steroidal anti-inflammatory drug misuse.

Acetaminophen↗

PubMed1997