Pain assessment during conscious sedation for cervical cancer high-dose-rate brachytherapy.

BACKGROUND: This observational study set out to evaluate the effectiveness of conscious sedation anesthesia for pain control during high-dose-rate (hdr) brachytherapy using a ring-and-tandem applicator system for patients with cervical cancer. METHODS: At the time of initiation of the hdr cervical cancer brachytherapy program at our institution, patients received a detailed symptom assessment during the procedures. Brachytherapy was carried out using a Smit sleeve, together with a ring-and-tandem applicator. Midazolam and an opioid-hydromorphone, morphine, or fentanyl-were the main agents used to achieve conscious sedation. RESULTS: From January 2009 to October 2010, 20 patients (median age: 45 years) underwent 57 procedures. All patients received chemoradiation with curative intent. The median duration of the procedures was 1.4 hours, and no significant cardiovascular events were noted. The total dose of intravenous midazolam used ranged from 0.5 mg to 8.5 mg (median: 2.5 mg). The total dose of intravenous morphine equivalent used ranged from 2.5 mg to 60 mg (median: 8 mg). The mean and median pain scores during the procedures were 1.4 and 1.1 respectively. Brief moments of moderate to severe incidental pain were noted at the time of certain events during the procedure-specifically during insertion of the ring-and-tandem applicator. The maximal pain score during the entire procedure ranged from 0 to 10 (median: 4.7). The period of recovery from conscious sedation was relatively brief (median discharge time: 1 hour). CONCLUSIONS: We were able to demonstrate that patients undergoing hdr brachytherapy for cervical cancer can achieve good pain control with conscious sedation.

Synthesis and biological activity of atrial natriuretic factor analogues: effect of modifications to the disulfide bridge.

A series of atrial natriuretic factor (ANF) analogues with modifications to the disulfide bridge and lacking the exocyclic N-terminal sequence was synthesized. The native cystine residue was substituted by isofunctional deamino carba, beta,beta-dimethyl carba and dehydro dicarba spanners that bridge residues 106 and 120. The compounds were prepared by segment condensation coupling using the base-labile (9-fluorenylmethyl)carboxyl protecting group. Biological evaluation revealed that the exocyclic N-terminal segment of ANF is not necessary for expression of high biological activity. The compounds retained high affinity for ANF receptors in bovine adrenal zona glomerulosa cells and were found to be potent antihypertensive and diuretic agents, indicating that the native disulfide bridge can be mimicked by isosteric spanning residues. It was noted that the reported analogues, unlike the endogenous hormone, show marked reduced inhibitory activity on PGE1-stimulated aldosterone secretion from adrenal zona glomerulosa cells. This lack of inhibition may be a contributing element to the low saluresis in spite of the high level of diuresis observed with some analogues.

Peptidomimetic inhibitors of herpes simplex virus ribonucleotide reductase: a new class of antiviral agents.

We have been investigating a new class of antiviral compounds effective against herpes simplex virus (HSV) in vitro and in vivo. Antiviral activity results from inhibition of HSV ribonucleotide reductase (RR). The inhibitors are designed as mimics of the RR small subunit C-terminus, a region essential for RR subunit association and consequently enzymatic activity. Inhibition results from specific binding of the inhibitor to the HSV RR large subunit thereby preventing subunit association. This report details the structure--activity studies that lead to the indentification of BILD 1263, a potent inhibitor of HSV RR subunit association (IC50, 0.2 nM) that also inhibits the replication of HSV types 1 and 2 in cell culture (EC50, 3 and 4 microM) and reduces the severity of HSV-1-induced keratitis in a murine ocular model. The discovery of inhibitors with in vitro antiviral results from a combination of improving inhibitor potency in a RR binding assay and modifying inhibitor physicochemical properties. The importance and possible role of the new structural modifications introduced into this inhibitor series is discussed.

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.

Nephrotoxicity of amphotericin B in rats: effects of the time of administration.

Amphotericin B is a potentially nephrotoxic agent used for the treatment of severe mycoses and numerous fungal infections. Temporal variation in the nephrotoxicity of amphotericin B was studied in rats maintained on a light-dark period of 14 hrs of light and 10 hrs of darkness (light on: 06h00). Subgroups of animals were treated with a single daily i.p. dose of either 5% dextrose or amphotericin B (10 mg/kg/day) given at either 07h00, 13h00, 19h00 or 01h00 for 4 and 10 days. On day 4, no significant difference was observed in any parameter studied. On day 10, the cellular regeneration ([3H]-thymidine incorporation into DNA of renal cortex)(p<0.01), BUN levels (p<0.05), serum creatinine (p<0.05), and accumulation of amphotericin B in the renal cortex (p<0.05) were significantly higher when animals were treated with similar subcellular localization of amphotericin B in the proximal tubular cells of the renal cortex. These results showed a temporal variation in the nephrotoxicity of amphotericin B (peak toxicity occurred at 07h00) which is different from that of other nephrotoxic antibiotics such as aminoglycosides.

Nephrotoxicity of low doses of tobramycin in rats: effect of the time of administration.

The circadian and the circannual variations of the nephrotoxicity of tobramycin were studied in female Sprague-Dawley rats. Animals were maintained on a light-dark period of 14/10 hrs (light on: 06h00 to 20h00). They were injected once daily for 4 and 10 days with saline or tobramycin at a dose of 40 mg/kg/day i.p. at either 08h00, 14h00, 20h00 and 02h00, in April 1991, July 91, October 91, January 92. In April 91, tobramycin injected at 14h00 during 10 days induced a significant increase of [3H]-thymidine incorporation into DNA of renal cortex as compared to other groups (p < 0.01): toxicity was highest at 14h00 and lowest at 02h00. No temporal change was observed in the renal cortical accumulation of tobramycin, and in serum creatinine after the 4 or 10 days of treatment. In experiments done in April, July and October 1991 and in January 1992, no circannual variation was found in tobramycin cortical levels but peaks of toxicity were observed at 02h00 in April and October 1991 and at 14h00 in July 1991 and January 1992. There was no linear correlation between the toxicity and the tobramycin accumulation in the renal cortex (r = 0.21). The data suggest that the circadian changes in tobramycin toxicity are due to temporal changes in the susceptibility of renal cells to tobramycin.

Day-night treatment difference of tobramycin serum and intrarenal drug distribution and nephrotoxicity in rats: effects of fasting.

The effects of short-term food deprivation on the serum and renal distribution and nephrotoxicity of tobramycin were studied in female Sprague-Dawley rats maintained on a 14-h light/10-h dark cycle (light on: 06:00). For the distribution study, a single injection of tobramycin (40 mg/kg, i.p.) was administered at 14:00 or 02:00 to normally fed animals or to animals fasted for 12 h before tobramycin injection; these treatment times correspond to the peak and trough of tobramycin nephrotoxicity as previously determined in other studies. The serum and cortical levels of tobramycin were significantly higher 60, 120, and 240 min after the injection in fasted animals treated at 02:00 compared with normally fed animals treated at the same time (p < 0.05). In animals injected at 14:00, similar levels of tobramycin were measured in both fasted and fed rats. In the nephrotoxicity study, female Sprague-Dawley rats were fasted for 12 h before and 24 h after the timed single injection of tobramycin (150 mg/kg, i.p.). The 24-h urinary excretion of beta-galactosidase was significantly higher in fasted animals treated at 02:00 than in fed rats treated at the same time of day. Seventy-two hours following tobramycin injection, serum creatinine levels and cortical levels of tobramycin were significantly higher in fasted rats treated at 14:00 than at 02:00 and in fed rats treated at 14:00. These data suggest that a short period of food deprivation modulates the temporal variations of tobramycin nephrotoxicity.

At the crossroads of my life--a holistic prevention and health promotion program.

A number of women experience difficulties during the climacteric related to the major change they are undergoing (i.e. physical, psychological, social and familial). These difficulties are often linked to negative attitudes and misunderstanding concerning the phenomena involved. This article describes a holistic prevention and health promotion program aimed at women aged 40 to 55. The program, in the form of an awareness-raising group, addresses the physical and psychological aspects of this period in a women's life, and presents participants with avenues to greater autonomy and better overall health. The organizational and evaluation aspects of the program are also presented.

Benefits of distributed HIS/RIS-PACS integration and a proposed architecture.

The tight integration of the Hospital Information System/Radiology Information System (HIS/RIS) and the Picture Archive Communication System (PACS) has become a priority in modern healthcare delivery systems. Traditional paper-based systems are being replaced by gateway style interface engines. Gateways provide a tightly integrated link between the HIS/RIS and the PACS, increasing productivity by automating many mundane clerical tasks associated with paper-based systems. A centralized gateway, however, represents a processing bottleneck and single point of failure. A self-monitoring distributed gateway architecture that replicates essential services increases the fault tolerance and the overall availability of the gateway, while providing reduced, consistent transaction times. A possible distributed architecture is proposed as a means to realize the advantages of a distributed architecture.

Current developments in departmental PACS for ultrasound.

1. INTRODUCTION TO DEPARTMENTAL PACS. Full or partial Departmental PACS is generally taken to mean an image management system focused on serving the needs of a specific modality or modality application. It will provide a modality specific means of image acquisition, specialized redisplay of images, distribution, and local long term storage of images. A Departmental PACS can be considered in isolation or as a component in a distributed Radiology PACS which consists of one or more departmental work groups on a back bone, potentially with shared resources. 2. DEPARTMENTAL PACS Issues Implementation of a Departmental PACS requires an in-depth knowledge of departmental clinical practice and work flow in all affected areas in the department, including patient intake, image collection, data routing, retrieval of previous image data, reporting, and long term data management and storage. Optimization of modality specific image display systems requires significant involvement from representative physician users. System architectures and user interfaces must be flexible enough to support the span of variation in clinical practice encountered in the site. A departmental PACS should offer a variety of "open" communications interfaces, both local and wide area, recognizing that outreach efforts are often driven by specific imaging departments. Interfaces to other departmental PAC systems and other information systems must be considered in order to facilitate institutions developing "Best of Breed" PACS systems. As hospitals move toward the integrated electronic medical record, means need to exist for a client process launched from a physician desktop to acquire images and/or reports from a departmental system. At minimum, HIS/RIS interfaces need to be considered to minimize re-keying of data and reduce data entry errors. 3. DESIGN OBJECTIVES FOR ALI ULTRAPACS. The key objectives were to design a product which could function either as a free standing PACS or as a departmental subnet on a larger PACS backbone, one which could function in a local or mixed local and wide area environment and one which could provide a cost effective implementation based on currently available technologies. 4. IMPLEMENTATION STRATEGIES. In order to attain the product design objectives, ALI made the following critical implementation decisions: 1) to build the UltraPACS application as a suite of separable UNIX processes based on a message passing client server model; 2) to host the application and operating system on a Digital Equipment Corporation PC using an Intel microprocessor because of the competition and broad variety of suppliers in the PC arena; and 3) to use NEXTSTEP as the UNIX variant of choice because of NEXTSTEPUs strong object orientation, superb development tools, and the excellent integration between the Graphical User Interface level and the underlying operating system layers. 5. CONCLUSION. ALI is convinced that a departmental approach to PACS offers significant advantages over monolithic PACS in several key areas including: optimization for modality specific clinical practice and workflow, cost effectiveness, the potential for an institution to implement PACS in a stepwise fashion, starting with the department with the potential for the greatest savings, and the capability for an institution to build a "best of breed" solution for large scale PACS.

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.

Kinetic studies on the inhibition of isopeptidase T by ubiquitin aldehyde.

Isopeptidase T (IPaseT) can hydrolyze isopeptide bonds of polyubiquitin (polyUb) chains, simple C-terminal derivatives of Ub, and certain peptides. We recently reported that IPaseT is regulated by ubiquitin (Ub); while submicromolar Ub activates, higher concentrations inhibit this enzyme [Stein et al. (1995) Biochemistry 34, 12616]. To explain these observations, we proposed a model for IPaseT involving two binding sites for Ub. According to the model, the two sites are adjacent to one another and are the extended active site that binds two Ub moieties of a polyUb chain. The "activation site" binds the Ub that donates Lys to the isopeptide bond. The "inhibition site" is adjacent and binds the Ub that donates the C-terminal Gly to the isopeptide bond. We now report that the interaction of IPaseT with the C-terminal aldehyde of Ub (Ub-H) is also modulated by Ub. In the absence of Ub, Ub-H inhibits IPaseT with a Ki of 2.3 nM, while at 0.6 microM Ub, where the "activation site" is occupied, Ki is less than 0.1 nM. At high Ub concentrations, where both the "activation" and "inhibition" sites are occupied, IPaseT cannot bind Ub-H. We also determined the kinetics of inhibition of IPaseT by Ub-H. In the absence of Ub, a two-step mechanism is followed. In the first step, Ub-H slowly combines with IPaseT to form a relatively weak complex (K1 = 260 nM) that slowly isomerizes to the final, stable complex that accumulates in the steady-state (k2 = 2 x 10(-3) s-1; k-2 = 0.02 x 10(-3) s-1). In contrast, Ub-activated IPaseT is inhibited by Ub-H through a three-step process. In the first step, Ub-H rapidly combines with IPaseT to form a complex (K1 = 10 nM) that slowly isomerizes to a second, more stable complex (k2 = 18 x 10(-3) s-1; k-2 = 1.5 x 10(-3) s-1). In the third step, the second complex converts to the final complex (k3 = 1.5 x 10(-3) s-1; k-3 < 0.2 x 10(-3) s-1). To unify the results of this study with our previous results on catalysis, we propose that binding of Ub either to catalytic transition states or to tetrahedral inhibition intermediates liberates more free energy than binding of Ub to the reactant state of IPaseT and that IPaseT can utilize this binding energy to stabilize both of these tetrahedral species. The overall effect is a Ub-induced increase in catalytic efficiency or inhibitory potency.

Effectiveness and toxicity of gentamicin in an experimental model of pyelonephritis: effect of the time of administration.

Temporal variations in the renal toxicity of aminoglycosides have been reported for experimental animals as well as for humans. In fact, maximal renal toxicity of aminoglycosides was observed when the drug was given during the rest period, while a lower toxicity was observed when the drug was injected during the activity period. The aim of the present study was to evaluate temporal variations in the effectiveness and renal toxicity of gentamicin in an experimental model of pyelonephritis in rats. The experiments were carried out with female Sprague-Dawley rats (185 to 250 g). They had free access to food and water throughout the study and were maintained on a 14-h light-10-h dark cycle. Animals were divided into four groups corresponding to the respective time of induction of pyelonephritis and treatment: 0700, 1300, 1900, and 0100 h. Pyelonephritis was induced by a direct inoculation of Escherichia coli (10(7) to 10(8) CFU) in the left kidney. Animals were treated for 3 and 7 days with a single daily dose of gentamicin (20 and 40 mg/kg of body weight, respectively) or saline (NaCl, 0.9%) at either 0700, 1300, 1900, or 0100 h. Animals treated at 0100 h for 3 days with gentamicin (20 mg/kg) showed a significantly lower number of bacteria in their kidneys than did all other groups (P < 0.01). After 7 days of treatment, the efficacy, evaluated by the log CFU per gram of tissue and by the percentage of sterilized kidneys, was also higher when gentamicin was administered at 0100 h. The beta-galactosidase and the N-acetyl-beta-D-glucosaminidase activities were significantly higher in urine of rats given gentamicin at 1300 h than in urine of rats treated at another time of day (P < 0.05). Gentamicin injected at 1300 h induced a significantly greater increase of [3H]thymidine incorporation into DNA of renal cortex (P < 0.01), a significantly greater inhibition of sphingomyelinase activity (P < 0.05), and significantly more histopathological lesions than the same dose injected at another time of the day. Creatinine and blood urea nitrogen levels in serum were significantly higher (P < 0.05) and the creatinine clearance was significantly lower (P < 0.05) when gentamicin was injected at 1300 h than when it was injected at another time of day. Our data suggest temporal variations in both the toxicity and the effectiveness of gentamicin, the drug being more effective and less toxic when injected during the activity period of the animals.

Effect of fasting on temporal variation in the nephrotoxicity of amphotericin B in rats.

Evidence for temporal variation in the nephrotoxicity of amphotericin B was recently reported in experimental animals. The role of food in these variations was determined by studying the effect of a short fasting period on the temporal variation in the renal toxicity of amphotericin B. Twenty-eight normally fed and 28 fasted female Sprague-Dawley rats were used. Food was available ad libitum to the fed rats, while the fasted animals were fasted 12 h before and 24 h after amphotericin B injection to minimize stress for the animals. Water was available ad libitum to both groups of rats, which were maintained on a 14-h light, 10-h dark regimen (light on at 0600 h). Renal toxicity was determined by comparing the levels of excretion of renal enzyme and the serum creatinine and blood urea nitrogen (BUN) levels at the time of the maximal (0700 h) or the minimal (1900 h) nephrotoxicity after the intraperitoneal administration of a single dose of dextrose (5%; control group) or amphotericin B (50 mg/kg of body weight; treated group) to the rats. The nephrotoxicities obtained after amphotericin B administration at both times of day were compared to the nephrotoxicities observed for time-matched controls. In fed animals, the 24-h urinary excretion of N-acetyl-beta-D-glucosaminidase and beta-galactosidase was significantly higher when amphotericin B was injected at 0700 and 1900 h. The excretion of these two enzymes was reduced significantly (P < 0.05) in fasting rats, and this effect was larger at 0700 h (P < 0.05) than at 1900 h. The serum creatinine level was also significantly higher (P < 0.05) in fed animals treated at 0700 h than in fed animals treated at 1900 h. Fasting reduced significantly (P < 0.05) the increase in the serum creatinine level, and this effect was larger in the animals treated at 0700 h. Similar data were obtained for BUN levels. Amphotericin B accumulation was significantly higher (P < 0.05) in the renal cortexes of fed rats than in those of fasted animals, but there was no difference according to the time of injection. These results demonstrated that fasting reduces the nephrotoxicity of amphotericin B and that food availability is of crucial importance in the temporal variation in the renal toxicity of amphotericin B in rats.

Ceftriaxone protects against tobramycin nephrotoxicity.

The effect of ceftriaxone on tobramycin-induced nephrotoxicity was investigated. Female Sprague-Dawley rats were treated during 4 and 10 days with saline (NaCl, 0.9%), ceftriaxone at a dose of 100 mg/kg of body weight/12 h subcutaneously, tobramycin at doses of 40 and 60 mg/kg/12 h intraperitoneally, or the combination ceftriaxone-tobramycin. Creatinine levels in serum were significantly higher in animals treated with tobramycin alone given at 60 mg/kg/12 h during 10 days, compared with control animals (P < 0.01) or animals receiving the combination tobramycin-ceftriaxone (P < 0.01). After 10 days of treatment, ceftriaxone did not accumulate in renal tissue but did reduce the renal intracortical accumulation of tobramycin (P < 0.05). Tobramycin given alone at either 40 or 60 mg/kg/12 h induced a significant inhibition of sphingomyelinase activity compared with control animals (P < 0.05). However, this enzyme activity was significantly less inhibited when tobramycin was injected in combination with ceftriaxone (P < 0.05). Ceftriaxone alone had no effect on the activity of this enzyme. The [3H]thymidine incorporation into the DNA of renal cortex was also significantly lower in animals treated with tobramycin-ceftriaxone compared with animals receiving tobramycin alone (P < 0.05). The 24-h urinary excretion of beta-galactosidase was significantly reduced in animals treated with the combination tobramycin-ceftriaxone compared with the administration of tobramycin alone at 40 and 60 mg/kg/12 h after 5 and 10 days (P < 0.05). Histologically, ceftriazone induced very few cellular alterations and reduced considerably the presence of typical signs of tobramycin nephrotoxicity. This investigation demonstrated that ceftriaxone protects animals against tobramycin-induced nephrotoxicity.

Multiplex PCR assays for the detection of clinically relevant antibiotic resistance genes in staphylococci isolated from patients infected after cardiac surgery. The ESPRIT Trial.

Multiresistant staphylococci (82 Staphylococcus aureus and 114 coagulase-negative staphylococci) were characterized by testing with rapid multiplex polymerase chain reaction (PCR) assays for species identification and detection of associated antibiotic resistance genes. These 196 staphylococci were isolated from 149 adult patients who developed wound infection after elective coronary artery bypass grafts and/or valve surgery. The multiplex PCR assays allowed identification of the most common staphylococcal species with S. aureus- and Staphylococcus epidermidis-specific primers as well as the detection of the erythromycin resistance genes ermA, ermB, ermC and msrA, the aminoglycoside resistance gene aac(6')-aph(2"), the oxacillin resistance gene mecA and the penicillin resistance gene blaZ. There was a very good correlation between the genotypic analysis by PCR and the phenotype determined by standard methods of susceptibility testing and identification of staphylococcal species: 100% for erythromycin resistance, 98.0% for gentamicin resistance, 99.0% for oxacillin resistance, 100% for penicillin resistance and 100% for S. aureus and S. epidermidis species identification. This study suggests that the incidence and distribution of the tested clinically relevant antibiotic resistance genes in staphylococci associated with infections after cardiac surgery do not differ from those in strains from other infections. These multiplex PCR assays may be used as diagnostic tools to replace or complement standard methods of susceptibility testing and identification of staphylococci.

Effects of fasting on temporal variations in nephrotoxicity of gentamicin in rats.

Evidence for temporal variations in the nephrotoxicity of low doses of aminoglycosides were recently shown by using specific and sensitive parameters of renal toxicity. The aim of the present study was to evaluate the effect of a short period of fasting on the temporal variations in the renal toxicity of gentamicin. Twenty-eight normally fed (i.e., food and water were available ad libitum throughout the experiment) female Sprague-Dawley rats (weight, 175 to 220 g) and 28 fasted rats (i.e., only water was available during a 12-h fast before and a 24-h fast after gentamicin injection) were used. The animals were synchronized on a 14-h light, 10-h dark cycle (lights on at 0600 h) for 1 week before gentamicin administration. In July 1993, each group of animals was treated with a single intraperitoneal injection of saline (NaCl, 0.9%) or gentamicin (150 mg/kg of body weight) at either the peak (1400 h) or the trough (0200 h) of the previously determined toxicity. On day 1, the 24-h urinary excretion of beta-galactosidase, N-acetyl-beta-D-glucosaminidase, and gamma-glutamyltransferase was significantly higher in normally fed animals treated with gentamicin at 1400 h than in their time-matched controls and in normally fed animals treated at 0200 h (P < 0.01), which had normal levels of these enzymes. By contrast, the urinary excretion of these enzymes was significantly higher in both groups of gentamicin-treated, fasted rats than in their time-matched control groups (P < 0.01), reaching levels similar to those measured in normally fed rats treated at 1400 h. The accumulation of gentamicin was significantly lower in the renal cortex of normally fed rats treated at 0200 h than in rats treated at 1400 h (P < 0.05), but this time-dependent difference was not found in fasted rats treated at 0200 and 1400 h. Immunogold labeling done on ultrathin sections and observed by electron microscopy showed a similar subcellular localization of gentamicin in normally fed and fasted rats treated at either 1400 or 0200 h. These results suggest that the feeding period is of crucial importance in the temporal variations of the nephrotoxicity of gentamicin in rats.

Attenuation of gentamicin-induced nephrotoxicity in rats by fleroxacin.

The effect of fleroxacin on gentamicin-induced nephrotoxicity was evaluated with female Sprague-Dawley rats. Animals were injected during 4 or 10 days with saline (NaCl; 0.9%), gentamicin alone at doses of 10 and 40 mg/kg of body weight/12 h (subcutaneously), fleroxacin alone at a dose of 25 mg/kg/12 h (intraperitoneally), or the combination gentamicin-fleroxacin in the same regimen. Gentamicin induced a dose- and time-dependent renal toxicity as evaluated by gentamicin cortical levels, sphingomyelinase activity in the renal cortex, histopathologic and morphometric analysis, blood urea nitrogen and serum creatinine levels, and cellular regeneration ([3H]thymidine incorporation into DNA of cortical cells). The extent of these changes was significantly reduced when gentamicin was given in combination with fleroxacin. Although the mechanisms by which fleroxacin reduces the nephrotoxic potential of gentamicin are unknown, we propose that the fleroxacin-gentamicin combination enhances exocytosis activity in proximal tubular cells, as suggested by the higher excretion of urinary enzymes and lower cortical levels of gentamicin observed in animals treated with the combination fleroxacin-gentamicin compared with those treated with gentamicin alone. The protective effect of fleroxacin on gentamicin nephrotoxicity should be investigated further.

Temporal changes of pharmacokinetics, nephrotoxicity, and subcellular distribution of tobramycin in rats.

The present study was designed to determine the temporal changes in tobramycin nephrotoxicity during the dark and the light periods of the day and to look for the mechanisms of such changes. Female Sprague-Dawley rats (9 to 11 weeks old) were housed in a 14-h-light-10-h-dark cycle (lights on 0600 to 2000 h). A bolus of tobramycin (60 mg/kg of body weight) was intravenously injected into a first group of 15 rats, at either 1400 or 0200 h. Six blood samples were taken from each rat, 30 to 210 min after the bolus injection. The total clearance of the drug was reduced during the rest period (1400 h) of rats compared with the activity period (0200 h) (P = 0.0007). Another group of 99 rats was given intraperitoneally a single dose of tobramycin (40 mg/kg), and renal cortices were collected 2 to 222 h after injection. The cortical drug levels were always higher in animals injected at 1400 h than in those injected at 0200 h. A last group of 32 rats was used in the studies of tobramycin (30 mg/kg/day, once daily for 10 days, intraperitoneally) nephrotoxicity and subcellular distribution. Weight gain in the rats receiving tobramycin (both 1400 and 0200 h) was significantly (P = 0.028) less than that in the controls. Nephrotoxicity, indicated by the incorporation of [3H]thymidine into cortical DNA and urinary excretion of N-acetyl-beta-D-glucosaminidase, was significantly higher in animals treated at 1400 h than in those treated at 0200 h. No difference in the subcellular distribution of tobramycin was observed. The data indicate that the reduction in the clearance of tobramycin during the rest period is in part responsible for the higher nephrotoxicity in rats.

Mechanistic studies on the inactivation of the proteasome by lactacystin: a central role for clasto-lactacystin beta-lactone.

Lactacystin is a Streptomyces metabolite that inhibits cell cycle progression and induces differentiation in a murine neuroblastoma cell line. The cellular target of lactacystin is the 20 S proteasome, also known as the multicatalytic proteinase complex, an essential component of the ubiquitin-proteasome pathway for intracellular protein degradation. In aqueous solution at pH 8, lactacystin undergoes spontaneous hydrolysis to yield N-acetyl-L-cysteine and the inactive lactacystin analog, clasto-lactacystin dihydroxy acid. We have studied the mechanism of lactacystin hydrolysis under these conditions and found that it proceeds exclusively through the intermediacy of the active lactacystin analog, clasto-lactacystin beta-lactone. Conditions that stabilize lactacystin (and thus prevent the transient accumulation of the intermediate beta-lactone) negate the ability of lactacystin to inactivate the proteasome. Together these findings suggest that lactacystin acts as a precursor for clasto-lactacystin beta-lactone and that the latter is the sole species that interacts with the proteasome.