Probenecid protects against In vivo acetaminophen-induced nephrotoxicity in male Wistar rats

Renal effects of acetaminophen (APAP) were studied in rats pretreated with probenecid to analyze whether acute APAP-induced nephrotoxicity could be related to a probenecid-sensitive transport system for APAP or its S-derived conjugates. The administration of probenecid (200 mg/kg b.wt. i.p.) 30 min before APAP administration (1000 mg/kg b.wt. i.p.) improved urine flow rate and protected against the alterations on glomerular filtration rate and urea and creatinine plasma levels induced by APAP. Fewer epithelial cells and granular casts and a decrease in the urinary excretion of protein and glucose were observed in rats pretreated with probenecid. Probenecid pretreatment promoted an elevation in the urinary 16-hr excretion of APAP and a diminution in the plasma levels attained by APAP. These results suggest that protection afforded by probenecid in vivo could be a consequence of the inhibition of APAP S-conjugate renal uptake and/or an increase in APAP renal clearance. The effects of APAP in presence of probenecid were studied with the isolated perfused kidney model. Perfusion with probenecid (0.1 mM) before APAP (10 mM) did not change APAP direct renal effects, APAP urinary excretion, or APAP renal clearance relative to glomerular filtration rate. Our results suggest that protection afforded by probenecid in vivo could be the result of the inhibition of the uptake of nephrotoxic APAP metabolites and/or a diuresis-induced enhanced APAP renal excretion.     

In vitro and in vivo antineoplastic effects of orthovanadate

A liquid chromatographic method for the determination of the macrolide antibiotics, roxithromycin and clarithromycin, in plasma is described. The method is fully automated, employing on-line solid-phase extraction for sample clean-up, using the Prospekt unit. Plasma samples, mixed with internal standard, were injected onto exchangeable CN cartridges. After washing, the compounds were eluted and transferred to a C18 analytical column for separation and electrochemical detection. Clarithromycin was used as internal standard when assaying roxithromycin and vice versa. The recovery of the solid-phase extraction method was 90% and higher, and the relative standard deviation was about 3%. The limit of quantitation was 0.5 mumol/l when 25 microliters of plasma was injected. Comparison with a liquid-liquid extraction method for sample clean-up showed good agreement.     

In vitro and in vivo protection against kainate-induced excitotoxicity by melatonin

In this study, the protective effect of melatonin against kainate (KA)-induced neurotoxicity was evaluated in vitro and in vivo. In rat brain synaptosomes, KA-induced oxidative stress was measured as shown by significant increases in both the basal generation of reactive oxygen species (ROS), assessed by a fluorescent method, and lipid peroxidation, evaluated as malondialdehyde (MDA) levels. Melatonin decreased, in a concentration-dependent manner, KA-induced lipid peroxidation. The intrinsic fluorescence of melatonin molecule hindered the evaluation of its protective effect against KA-induced ROS generation. However, melatonin was able to reduce FeSO4/ascorbate-induced ROS generation. The melatonin protective effect was confirmed by in vivo experiments: 73% of rats injected with KA (10 mg/kg i.p.) died within 5 days; melatonin administration i.p. significantly reduced mortality of the animals. The present results suggest that melatonin might be considered a pharmacological agent for the treatment of neurodegenerative pathologies.     

In vitro and in vivo activities of trybizine hydrochloride against various pathogenic trypanosome species

Trybizine hydrochloride [O,O'-bis(4,6-diamino-1,2-dihydro-2, 2-tetramethylene-s-triazine-1-yl)-1,6-hexanediol dihydrochloride] was active in vitro against the sleeping sickness-causing agents Trypanosoma brucei subsp. rhodesiense and T. brucei subsp. gambiense; against a multidrug-resistant organism, T. brucei subsp. brucei; and against animal-pathogenic organisms Trypanosoma evansi, Trypanosoma equiperdum, and Trypanosoma congolense; but not against the intracellular parasites Trypanosoma cruzi and Leishmania donovani. Cytotoxic effects against mammalian cells were observed at approximately 10(6)-fold higher concentrations than those necessary to inhibit T. brucei subsp. rhodesiense. Trybizine hydrochloride was able to eliminate T. brucei subsp. rhodesiense and T. brucei subsp. gambiense in an acute rodent model with four intraperitoneal doses of 0.25 mg kg of body weight-1 or four doses of 1 mg kg-1, respectively, or with four oral doses of 20 mg kg-1. The compound expressed activity against suramin-resistant T. evansi strains in mice. However, these concentrations were not sufficient to cure mice infected with multidrug-resistant T. brucei subsp. brucei. A late-stage rodent model with central nervous system involvement could not be cured, indicating that trybizine may not pass the blood-brain barrier in sufficient quantities.     

In vitro and in vivo efficacy of the triazole TAK-187 against Cryptococcus neoformans

Multiple isolates of Cryptococcus neoformans, including those with fluconazole resistance, were tested to assess the in vitro activity of the new triazole TAK-187. MICs of TAK-187 were at least eightfold lower than those of fluconazole, and fungicidal concentrations for most isolates were 4 microg/ml or less. TAK-187 also was evaluated as intermittent therapy using two dosages in a rabbit model of experimental cryptococcal meningitis. Compared to daily treatment with fluconazole, as little as two doses of TAK-187 given 7 days apart were found to be effective. Plasma and cerebrospinal fluid TAK-187 concentrations were many times higher than MICs and fungicidal concentrations. Based upon its therapeutic efficacy and long half-life in the rabbit model, TAK-187 should be investigated for intermittent dosing in treatment or suppression of cryptococcal infections in humans.     

In vitro and in vivo activity of two Pt(IV) salts against leishmania donovani

The activities of 8 platinum drug complex salts were determined against Leishmania donovani promastigotes. The three most active salts were selected: [PtIVBr6]H2 (pentamidine); [PtIVBr6]H2 (stilbamidine), and [PtIVCl6]H2 (2-piperazinyl(1) ethyl amine), which induced growth-inhibition rates of more than 50% at 24 h of treatment and at the maximum dosage tested. The cytotoxicity assays on the macrophage cell line J-774 showed high cytotoxicity for the salt [PtIVBr6]H2 (stilbamidine) with a percentage of specific 51Cr release of 58.2% at 24 h of incubation and 100 microg/ml. Meanwhile, assays of the other compounds showed practically no cytotoxicity. The salt [PtIVBr6]H2 (pentamidine) notably inhibited the incorporation of 3H-thymidine in the treated parasites. The ultrastructural alterations observed in the flagellates treated with the salts [PtIVCl6]H2 (2-piperazinyl(1)ethyl amine) and [PtIVBr6]H2 (pentamidine) suggest that both act preferentially at the nuclear level and at the kinetoplast-mitochondrion complex. Both compounds showed a high in vivo activity in parasitized Wistar rats.     

In vivo and in vitro antiinflammatory activity of saikosaponins

Buddlejasaponin I and saikosaponin 1 and 2, biologically active compounds from Scrophularia scorodonia and Bupleurum rigidum respectively, exert potent in vivo antiinflammatory effects on mouse ear edema induced by phorbol myristate acetate (PMA). The effects of these compounds on swelling and other inflammatory parameters are described. In screening for in vitro effects of saikosaponins on cellular systems generating cyclooxygenase (COX) and lipoxygenase (LOX) metabolites, we observed that most saikosaponins showed a significant effect. The action is more marked on LOX metabolite LTC4. Our data support the inhibition of arachidonic acid metabolism as one of the biochemical mechanisms that might be the rationale for the putative antiphlogistic activity of these saikosaponins.     

Efficacy of artemisinin and mefloquine combinations against Plasmodium falciparum. In vitro simulation of in vivo pharmacokinetics

The activity of artemisinin in combination with mefloquine was tested in vitro against a chloroquine-sensitive (F32) strain of Plasmodium falciparum. A method of repetitive dosing and extending the culture observation period to 28-30 days was used to mimic the in vivo pharmacokinetic situation. Plasmodium falciparum was exposed to artemisinin from 10(-8) to 10(-5) M, mefloquine from 3 x 10(-9) to 10(-5) M and their combinations. The exposure time for artemisinin was 3 hours twice daily and for mefloquine 24 hours. The drug-dosing duration was 3 days. Neither artemisinin nor mefloquine alone provided radical clearance of P. falciparum, even when maximum concentrations (10(-5) M) were applied. The antiparasitic activity of artemisinin and mefloquine were significantly higher when dosed alone. Effective concentrations for different degrees of inhibition (EC 50, 90 and 99) of both artemisinin and mefloquine respectively were significantly lower when used in combination. At concentrations normally reached in vivo, this effect was clearly synergistic (P = 0.016) Our in vitro model of intermittent dosing of artemisinin and mefloquine combinations for 3 days provides significant evidence of positive interaction between the two compounds. Lower combination concentrations around the MIC-values for the individual compounds showed synergistic effect, and high concentrations showed additive effect. This indicates that such drug combinations may provide radical clearance at concentrations lower than those required for single-drug treatment.     

In vitro and in vivo calcification of vascular bioprostheses

Efficacy of different chemical treatments on calcification of vascular graft in vitro and in vivo was studied. Culture medium-filled rat aortas were separately treated in 0.2% glutaraldehyde and epoxy compound, and photooxidized in 0.01% methylene blue for a shorter period (group 1). Another group of rat aortas were separately treated in the same chemicals for a longer period (group 2). All fresh and treated aortas of both groups were cultured for 21 days in an organ culture medium and implanted (except for group 1) in weanling rats for five months. Histology and immunohistochemistry revealed that differently treated aortas of group 1 grow and calcify, and the smooth muscle cells between elastin fibers are the primary site of calcium deposition. In contrast, differently treated aortas of group 2 neither grew, nor did calcify in the medium except the epoxy compound cross-linked aorta of group 2 which did not grow but did calcify. Untreated aorta did not calcify. All fresh and differently treated aortic homografts calcified severely in rats. Our whole arterial segment-calcification system would be useful for analyzing the molecular and cellular mechanisms of both bioprosthetic and atherosclerotic calcification of vascular graft. New anticalcification technique is the only hope for better outcome of future vascular bioprostheses.     

In vitro and in vivo effects of activated macrophage supernatant on distal limb wounds of ponies

OBJECTIVE: To determine whether monokines produced by activated rabbit peritoneal macrophages can inhibit development of exuberant granulation tissue formation in distal limb wounds in ponies. DESIGN: Randomized block. ANIMALS: 5 castrated male ponies, 2 to 6 years old and weighing 140 to 190 kg. PROCEDURE: In vitro activity of cell-free rabbit peritoneal macrophage supernatant was determined after incubation of fibroblasts from the flank and the distal portion of limbs of horses and ponies. Tritiated thymidine was then added, and after reincubation, radioactivity was measured. After creation of a 4-cm2, full-thickness wound on the mid dorsal aspect of each metacarpus and metatarsus of each pony, in vivo activity of the macrophage supernatant was evaluated. Biopsy specimens were collected at random sites near a border of each wound at 4, 6, and 10 weeks after creation of the wounds. Treatment effects were evaluated on the basis of presence of exuberant granulation tissue requiring excision, number of times that excision was required, total area of the wound, epithelialized area, area of granulation bed, and histologic evaluation of the biopsy specimens. RESULTS: The macrophage supernatant effectively inhibited proliferation of equine fibroblasts in vitro. No significant in vivo treatment effects were found among the 4 treatment groups. CONCLUSION AND CLINICAL RELEVANCE: Monokines from stimulated rabbit peritoneal macrophages may have potential for improving wound healing in horses and ponies because of their effective inhibition in vitro of equine fibroblast proliferation.     

The myelotoxicity of chloramphenicol: in vitro and in vivo studies: I. In vitro effects on cells in culture

1 Chloramphenicol is used extensively in non-industrialized countries for the treatment of life-threatening infections because it is cheap and effective, despite its known hemotoxicity and linkage to fatal aplastic anaemia. It is important to define the mechanism of toxicity so that means can be devised to ameliorate the toxic effects in order to produce safer usage. 2 Chloramphenicol, at concentrations from 5 mM to 2 mM initiated apoptosis in dividing cells from a monkey kidney-derived cell line and in haematopoietic progenitor cells from human neonatal cord blood. 3 Growth of progenitor cells was suppressed at concentrations of chloramphenicol which would be considered less than therapeutic during patient treatment. 4 These effects could be ameliorated in progenitor cells by co-culture with the antioxidant mercaptoethylamine and in monkey kidney cells by co-culture with vitamin C. 5 This is the first report of apoptosis in chloramphenicol toxicity and suggests a possible link between a metabolic event i.e. the production of free radicals; a morphological effect, apoptosis; and a clinical effect, bone marrow suppression and aplastic anaemia.     

In vivo and in vitro assessment of barium sulphate suspensions

In vitro methods of testing the efficiency of barium sulphate suspensions in delineating mucosal detail using canine cadaveric stomachs have been described in the literature. In this study a comparison is made between in vitro and in vivo tests in the stomach and small intestine of dogs, using several brands of barium sulphate. The results indicate that there is considerable variation in the behavior of these suspensions between the in vitro and in vivo tests particularly in the stomach. It is our view that in vitro tests of this sort are of little value for assessing the relative advantages and disadvantages of these suspensions in demonstrating mucosal detail.     

In vitro and in vivo hematopoietic activities of Betafectin PGG-glucan

Betafectin PGG-glucan is a novel beta-(1,3)glucan that has broad-spectrum anti-infective activities without cytokine induction. Here we report that PGG-glucan also has both in vitro and in vivo hematopoietic activities. In vitro studies with bone marrow target cells from the C3H/HeN mouse revealed that although PGG-glucan alone had no direct effect on hematopoietic colony-forming cell (CFC) growth, when combined with granulocyte colony-stimulating factor (CSF) or granulocyte-macrophage CSF, it increased CFC numbers 1.5- to 2.0-fold over those obtained with CSFs alone. Bone marrow cells cultured for high-proliferative-potential CFCs in the presence of interleukin (IL)-1, IL-3, macrophage CSF, and stem cell factor (SCF), or cultured for erythroid burst-forming units in the presence of IL-3, SCF, and erythropoietin, also exhibited enhanced growth in the presence of PGG-glucan. The synergistic effect of PGG-glucan was specific and could be abrogated by anti-PGG-glucan antibody. The ability of PGG-glucan to modulate hematopoiesis in vivo was evaluated in myelosuppressed rodents and primates. C3H/HeN female mice were intravenously administered saline solution or PGG-glucan (0.5 mg/kg) 24 hours before the intraperitoneal administration of cyclophosphamide (200 mg/kg), and the recovery of bone marrow cellularity and granulocyte-macrophage progenitor cells was evaluated on days 4 and 8 after cyclophosphamide treatment. At both time points, enhanced hematopoietic recovery was observed in PGG-glucan-treated mice compared with saline-treated control mice. In a final series of in vivo experiments, we evaluated the ability of therapeutically administered PGG-glucan to enhance hematopoietic recovery in cyclophosphamide-treated cynomolgus monkeys. Monkeys received intravenous infusions of cyclophosphamide (55 mg/kg) on days 1 and 2, followed on days 3 and 10 by intravenous infusion of PGG-glucan (0.5, 1.0, or 2.0 mg/kg). Compared with those in saline-treated monkeys, accelerated white blood cell recovery and a reduction in the median duration of neutropenia were observed in PGG-glucan-treated monkeys. These studies illustrate that PGG-glucan has both in vitro and in vivo hematopoietic activities and that this agent may be useful in the prevention and/or treatment of chemotherapy-associated myelosuppression.     

In vitro and in vivo immunopharmacological profile of Sch 40120

Sch 40120 (10-(3-chlorophenyl) - 6,8,9,10- tetrahydrobenzo [b] [1,8] naphthyridin-5 (7H)-one) is a leukotriene inhibitor that is also a potent inhibitor of acute inflammatory responses in rodent systems. In the present study, we have evaluated the effects of this drug on immune function as well as its activity in models of immune mediated chronic inflammatory disease. Sch 40120 was particularly effective in suppressing T cell proliferative responses in vitro. Antigen-specific and poly-clonally-induced in vitro antibody responses were also inhibited by the drug. However, the in vivo potency of Sch 40120 in suppressing immune responses and in inhibiting the pathological changes seen in rodent models of autoimmune disease (EAE and adjuvant arthritis) was somewhat less than that previously observed in models of acute inflammation. Nevertheless, the spectrum of activities exhibited by Sch 40120 suggests that it will be particularly useful in the treatment of psoriasis where T lymphocytes have been implicated in the development of disease and leukotrienes appear to have a role in the persistence of psoriatic plaques.     

Role of cytochrome P-450 as a source of catalytic iron in cisplatin-induced nephrotoxicity

BACKGROUND: Iron plays a role in free radical-mediated tissue injury, including cisplatin-induced nephrotoxicity. However, the source of iron (catalyzing free radical reactions) is not known. We examined the role of cytochrome P-450 as a source of catalytic iron in cisplatin-induced nephrotoxicity both in vivo and in vitro. METHODS: Cisplatin-induced acute renal failure was produced in rats by intraperitoneal injection of cisplatin (10 mg/kg body wt). Piperonyl butoxide, a cytochrome P-450 inhibitor, was administered intraperitoneally (400 mg/kg body wt twice at 48-hr intervals) prior to cisplatin injection. The effects of cisplatin in the absence or presence of piperonyl butoxide on the belomycin-detectable iron, cytochrome P-450 content in the kidney, and renal functional and histological changes were evaluated. In an in vitro study, the effect of cytochrome P-450 inhibitors, cimetidine or piperonyl butoxide, on cisplatin-induced cytotoxicity and catalytic iron release from LLC-PK1 cells was examined. RESULTS: In cisplatin-treated rats, there was a marked decrease in the cytochrome P-450 content specifically in the kidney, accompanied by increased bleomycin-detectable iron content in the kidney. Piperonyl butoxide prevented cisplatin-induced loss of cytochrome P-450 as well as the increase of bleomycin-detectable iron in the kidney, along with both functional and histological protection. Both cimetidine and piperonyl butoxide prevented cisplatin-induced increase in bleomycin-detectable iron and cytotoxicity in LLC-PK1 cells. Treatment of cimetidine did not affect cellular uptake of cisplatin. CONCLUSION: Cytochrome P-450, a group of heme proteins, may serve as a significant source of catalytic iron in cisplatin-induced nephrotoxicity.     

In vitro and in vivo biologic effects of interleukin-3 (IL-3) in follicular low-grade lymphoma

We have taken an enhancer trap approach to identify genes that are expressed in hematopoietic cells and tissues of Drosophila. We conducted a molecular analysis of two P-element insertion strains that have reporter gene expression in embryonic hemocytes, strain 197 and vikingICO. This analysis has determined that viking encodes a collagen type IV gene, alpha2(IV). The viking locus is located adjacent to the previously described DCg1, which encodes collagen alpha1(IV), and in the opposite orientation. The alpha2(IV) and alpha1(IV) collagens are structurally very similar to one another, and to vertebrate type IV collagens. In early development, viking and DCg1 are transcribed in the same tissue-specific pattern, primarily in the hemocytes and fat body cells. Our results suggest that both the alpha1 and alpha2 collagen IV chains may contribute to basement membranes in Drosophila. This work also provides the foundation for a more complete genetic dissection of collagen type IV molecules and their developmental function in Drosophila.     

In vivo and in vitro macrophage activation by systemic adjuvants

Six systemic adjuvants of immunity were tested for their ability to induce macrophage activation. Four of them: living BCG, hydrosoluble extracts from BCG (HIU II) and from M.smegmatis (IPM), and lipopolysaccharide from E.coli (LPS), when administered to normal mice render macrophages non-specifically cytotoxic for tumor cells in vitro. The intensity of this phenomenon varied according to the route and time of adjuvant administration. In contrast, lentinan extracted from Lentinus edodes, and levamisole which is a synthetic chemical compound, depressed macrophage cytotoxic potential. BCG, IPM and LPS were shown to have a direct action on macrophages. After in vitro exposure to these agents, the cytotoxic potential of normal macrophages was greatly increased. Levamisole was unable to stimulate this macrophage function directly in vitro. On the other hand, such a macrophage activation has been induced in vitro when normal macrophages were cultivated in the presence of MIF coming from the supernatant of human lymphoblastoid cell lines.     

In vitro and in vivo effects of cisplatin and etoposide in combination on small cell lung cancer cell lines

The effects of cisplatin (CDDP) and etoposide (ETP) in combination were evaluated in vitro and in vivo using small cell lung cancer cell lines. The combination effects in vitro were investigated using isobologram analysis. Used together, CDDP and ETP showed a synergistic effect against cell growth on only 1 cell line (SBC-3), additive effects on 6 (SBC-2, SBC-5, Lu130, Lu134AH, Lu135T and H69) and an antagonistic effect on 1 (SBC-1). In the in vivo experiment, nude mice were inoculated with SBC-1, SBC-3 and SBC-5 cells. Two or 5 mg/kg CDDP and 10 or 30 mg/kg ETP were administered intraperitoneally alone and simultaneously in combination to nude mice. The in vivo effects of the combination were determined by comparing the observed growth ratio in mice treated with the combination with the expected value of this ratio calculated based on the assumption that the effects of the drugs were simply additive. According to this definition, synergistic effects were observed against all 3 tumors. Thus, the in vivo and in vitro effects differed. The toxicity of the combination therapy, which was analyzed by estimating the body weight change of mice, was no higher than that of CDDP or ETP alone. These results suggest that the excellent clinical effects of CDDP and ETP combination therapy may be attributable not to drug interaction at the cellular level but to the feasibility of combined use of them at full doses without overlapping side effects.