Stem cell intervention ameliorates epigallocatechin-3-gallate/lipopolysaccharide-induced hepatotoxicity in mice.

Stem cells are identified as a novel cell therapy for regenerative medicine because of their ability to differentiate into many functional cell types. We have shown earlier a new model of hepatotoxicity in mice by administering (1500 mg/kg) epigallocatechin-3-gallate (EGCG) intragastric (IG) for 5 days after a single intraperitoneal dose (6 mg/kg) of lipopolysaccharide (LPS). In this study, we aimed to study the effect of intrahepatic (IH) injection of mouse embryonic stem cells (MESCs) on the hepatotoxicity induced by EGCG/LPS in mice. Mice were administered EGCG/LPS and rested for 3 days. MESCs were obtained from American Type Culture Collection and cultured in vitro for 4 days. Stem cells were injected IH. Seven days later, a single dose of LPS (6 mg/kg) followed by daily doses of IG administration of EGCG were re-administered for 5 days. At the end of the experiment, blood samples were collected for analysis of biochemical parameters associated with liver. Results showed that the group of mice that were administered MESCs prior to EGCG/LPS showed lower levels of alanine amino transferase, alkaline phosphatase, and bilirubin, higher albumin/globulin ratio, and less remarkable histopathological lesions. Also, that group of mice showed less expression of oxidative stress biomarkers (oxidized low-density lipoprotein Ox.LDL and chemokine CXCL16), less expression of nuclear protein receptors (retinoic acid receptor and retinoid X receptor), and less expression of inflammatory biomarkers (tumor necrosis factor α and transforming growth factor ß1) compared with other groups of mice that were not given MESCs. In conclusion, MESCs can ameliorate EGCG/LPS-induced hepatotoxicity in mice.

Germ cell mutagenicity of topoisomerase I inhibitor topotecan detected in the male mouse-dominant lethal study.

To investigate the ability of topotecan, a topoisomerase I-targeting anticancer drug, to induce dominant lethal mutations in male mouse germ cells, males were treated with single doses of 3, 6 and 12 mg/kg topotecan. Each male was mated at 4-day intervals to virgin females for a total of nine 4-day mating intervals. The two highest doses of topotecan are shown to be mutagenic in post-meiotic cells. The greatest effect occurred in those cells which were in the early-spermatid stage at the time of exposure. Mice treated with 12 mg/kg topotecan showed an additional peak of dominant lethal induction in mature sperm during the first 4-day matings after treatment. The mutagenic effects were directly correlated with free radicals accumulation as an obvious increase in the generation reactive oxygen species and 8-hydroxydeoxyguanosine was noted in animals treated with 6 and 12 mg/kg topotecan. Treatment of male mice with N-acetylcysteine, a free radical scavenger, significantly protected mice from topotecan-induce dominant lethality. Moreover, N-acetylcysteine had no antagonizing effect on topotecan-induce topoisomerase-I inhibition. Our study provides evidence that topotecan is a germ cell mutagen and its effect is more pronounced during the post-meiotic stages through a mechanism that may involves increases in DNA oxidative stress.

Toward gene therapy of uterine fibroids: targeting modified adenovirus to human leiomyoma cells.

BACKGROUND: To circumvent the paucity of the primary adenovirus (Ad5) receptor and the non-specific Ad5 tropism in the context of uterine leiomyoma cells, Ad5 modification strategies would be beneficial. METHODS: We screened several modified adenoviruses to identify the most efficient and selective virus toward human leiomyoma cells to be used as candidate for delivering therapeutic genes. We propagated: wild-type Ad5-luc, fiber-modified viruses: ad5 RGD-luc, Ad5-Sigma-luc, Ad5/3-luc and Ad5-CAV2-luc, as well as transcriptional targeted viruses: ad5 survivin-luc, Ad5-heparanase-luc, Ad5-MSLN-CRAD-luc and Ad5-SLPI-luc, on 293 cells and purified them by double CsCL density centrifugation. Then we transfected primary cultures of human leiomyoma cells derived from fibroids of four different patients, telomerase-immortalized human leiomyoma cell line (huLM), telomerase-immortalized normal human myometrial cell line (HM9) and immortalized normal human liver cells (THLE3) with the viruses at 5, 10 and 50 plaque-forming units (PFU)/cell. After 48 h, luciferase activities were measured and normalized to the total cellular protein content. RESULTS: Ad5-RGD-luc and Ad5-CAV2-luc, Ad5-SLPI-luc and Ad5-MSLN-CRAD-luc at 5, 10 and 50 pfu/cell showed significantly higher expression levels of luciferase activity in both primary and immortalized human leiomyoma cells when compared with Ad5-Luc. Additionally, these modified viruses demonstrated selectivity toward leiomyoma cells, compared with myometrial cells and exhibited lower liver cell transduction, compared with Ad5-luc, at the same dose levels. CONCLUSIONS: Ad5-CAV2-luc, Ad5-RGD-luc, Ad5-SLPI-luc and Ad5-MSLN-CRAD-luc are promising delivery vehicles in the context of leiomyoma gene therapy.

Orthovanadate increased the frequency of aneuploid mouse sperm without micronucleus induction in mouse bone marrow erythrocytes at the same dose level.

The objective of the current study was to investigate the ability of orthovanadate to induce aneuploidy in mouse sperm and micronuclei in mouse bone marrow cells at the same dose levels. The BrdU-incorporation assay was performed to test if the chemical treatment altered the duration of the meiotic divisions. It was found that orthovanadate (25mg/kg bw) treatment did not cause meiotic delay. To determine the frequencies of hyperhaploid and diploid sperm, male mice were treated by intraperitoneal (i.p.) injection with 5, 15 or 25mg/kg bw orthovanadate and sperm were sampled from the Caudae epididymes 22 days later. Fluorescence in situ hybridization (FISH) was performed with DNA-probes for chromosomes 8, X or Y. Significant increases in the frequencies of total hyperhaploid sperm (p<0.01) were found with 15 and 25mg/kg bw orthovanadate, indicating induced non-disjunction during male meiosis. The dose-response was described best by a linear equation. Orthovanadate did not significantly increase the frequencies of diploid sperm at any of the three doses tested, indicating that no complete meiotic arrest occurred. Orthovanadate was investigated also by the micronucleus test at i.p. doses of 1, 5, 15 or 25mg/kg bw, followed by bone marrow sampling 24h after treatment. None of the orthovanadate doses caused a significant increase in the rates of micronuclei (MN). Since the results show that orthovanadate induced non-disjunction during male meiosis without an accompanying induction of MN in bone marrow erythrocytes under the present experimental conditions and doses, it is concluded that male germ cells (meiosis) are more sensitive to the aneugenic effects of orthovanadate than somatic cells (mitosis). However, induction of micronuclei was reported in the literature with orthovanadate, vanadylsulfate and ammonium metavanadate, which contradicts the notion that vanadium compounds might be unique germ cell aneugens.

Influence of p-coumaric acid on doxorubicin-induced oxidative stress in rat's heart.

The therapeutic value of doxorubicin (DOX) as anticancer antibiotic is limited by its cardiotoxicity. The implication of natural phenolic acids in the prevention of many pathologic diseases has been reported. Herein, the ability of p-coumaric (PC) acid, a member of phenolic acids, to protect rat's heart against DOX-induced oxidative stress was investigated. Three main groups of albino rats were used; DOX, PC, and PC plus DOX-receiving animals. Corresponding control animals were also used. DOX was administered i.p. in a single dose of 15mgkg(-1). PC alone, in a dose of 100mgkg(-1), was orally administered for five consecutive days. In PC/DOX group, rats received PC 5 days prior to DOX. DOX-induced high serum levels of lactic dehydrogenase (LDH) and creatine phosphokinase (CPK), were reduced significantly by PC administration, compared to DOX-receiving rats. Pretreatment with PC ameliorated the cardiac content of glutathione (GSH), and superoxide dismutase (SOD) & catalase (CAT) activities, compared to DOX-receiving rats. On the other hand, accumulation of cardiac content of MDA significantly decreased following PC pretreatment, compared to DOX-treated rats. The data presented here indicate that PC protects rats hearts against DOX-induced oxidative stress in the heart. It may be worthy to consider the usefulness of PC as adjuvant therapy in cancer management.

Etoposide and merbarone are clastogenic and aneugenic in the mouse bone marrow micronucleus test complemented by fluorescence in situ hybridization with the mouse minor satellite DNA probe.

The topoisomerase II (topo II) inhibitors etoposide (VP-16) and merbarone (MER) were investigated with the in vivo micronucleus test (MN test) combined with fluorescence in situ hybridization (FISH) using the mouse minor satellite DNA probe to discriminate MN of clastogenic and aneugenic origin. All experiments were performed with male (102/ElxC3H/El) F1 mice bred in the mouse colony of the GSF Research Center. The sample size per experimental group was five animals and 2,000 polychromatic erythrocytes (PCE) were scored per animal from coded slides in the conventional MN test. A separate set of coded slides was used for the FISH analysis. All treatments consisted of single intraperitoneal injections. Colchicine (COL, 3 mg/kg) and mitomycin (MMC, 1 mg/kg) were used as a positive control aneugen and clastogen, respectively, and these compounds produced the expected responses. A dose of 1 mg/kg VP-16 induced 3.44% MNPCE (compared to the concurrent solvent control of 0.37%, P < 0.001) and of these 39.9% (1.4% MNPCE) showed one or more fluorescent signals. MER (7.5-60 mg/kg) increased the MNPCE frequencies in a dose-dependent manner, with 15 mg/kg being the lowest positive dose. At the highest dose of 60 mg/kg of MER, a total of 4.26% MNPCE were found (compared to 0.31% in the concurrent solvent control, P < 0.001) and of these 46.2% (2.0% MNPCE) contained one or more fluorescent signals. The data demonstrate that VP-16 and MER induced both clastogenic and aneugenic events despite their different modes of topo II inhibition.

HPV - Relato de um caso clínico / HPV - a case report

Relato de caso de paciente de 16 anos de idade, com diagnóstico de infecçäo cérvico vaginal por Papilomavírus Humano (HPV), sugerido inicialmente através de citologia oncótica pela presença evidente de coilócitos, critério patognomônico para HPV e posteriormente comprovado por Captura Híbrida a presença de HPV de alto potencial oncogênico. Realizou-se também colposcopia, onde se observaram lesöes em colo e vulva características do mesmo agente, contudo o estudo histopatológico demonstrou alteraçöes displásicas, sem entretanto caracterizar presença de qualquer agente específico

Molecular cytogenetic analysis in mouse sperm of chemically induced aneuploidy: studies with topoisomerase II inhibitors.

The ability of two topoisomerase II (topo II) inhibitors, etoposide (VP-16) and merbarone (MER), to induce meiotic delay and aneuploidy in mouse spermatocytes was investigated. The progression from meiotic divisions to epididymal sperm was determined by injecting male mice with 5-bromo-2'-deoxyuridine (BrdU) and treating the animals 13 days later with the test chemicals. At 20-24 days after treatment, BrdU-containing sperm were identified with a FITC-labelled anti-BrdU antibody and green fluorescent sperm were scored with a laser scanning cytometer (LSC). It was found that VP-16 (50mg/kg) treatment induced a meiotic delay of about 24h. A significant reduction of BrdU-labelled sperm was observed at 22 days compared to the controls (VP-16 group: 14.20%; controls: 41.10%, P<0.001). At 23 and 24 days, there were no significant differences between the VP-16 and the control groups. MER (80 mg/kg) treatment did not cause meiotic delay. To determine the frequencies of hyperhaploid and diploid sperm, male mice were treated with 12.5, 25 and 50mg/kg VP-16 or 15, 30 and 60 mg/kg MER. Sperm were sampled from the Caudae epididymes 24 days after VP-16 treatment or 22 days after MER treatment. Significant increases above the concurrent controls in the frequencies of total hyperhaploid sperm were found after treatment with 25, 50mg/kg VP-16 (0.074 and 0.122% versus 0.052%) and after treatment with 60 mg/kg MER (0.098% versus 0.044%). Furthermore, significant increases in the frequencies of diploid sperm were found after treatment of mice with all three doses of VP-16 (0.024, 0.032 and 0.056% versus 0.004 and 0.00%, respectively) and with 30 and 60 mg/kg MER (0.022 and 0.05% versus 0.004 and 0.002%, respectively). All dose responses could be expressed by linear equations. The results indicate that cancer patients may stand transient risk for siring chromosomally abnormal offspring after chemotherapy with these topo II inhibitors.

The impact of ofloxacin on rat testicular DNA: application of image analysis.

Ofloxacin induces its antibacterial action mainly by inhibition of DNA gyrase which is equivalent to topoisomerase II in mammalian cells. The present study was focused on the impact of ofloxacin on rat testicular DNA ploidy in a dose-response relationship using an image analysis technique on testicular sections following Fuelgen DNA staining. Sperm count and motility as well as sperm head abnormality tests were also investigated. Ofloxacin was given p.o. in doses of 36, 72 and 360 mg kg(-1) day(-1) for 15 consecutive days. The animals were then left to live safely to day 60 from starting the experiment to give them a chance to complete the cycle of spermatogenesis. Results revealed that ofloxacin significantly decreased the percentage of haploid cells in a dose-dependent manner with concomitant increase in the percentage of diploid cells. Sperm count and motility was also markedly decreased in a dose-dependent fashion. Sperm head abnormality showed no marked change following ofloxacin treatment. In conclusion, ofloxacin induced a marked disturbance in rat testicular DNA ploidy, which may be explained on the basis of cross-reactivity to topoisomerase II.

Evaluation of cisplatin combined with ondansetron in Ehrlich ascites carcinoma in vitro and in vivo.

AIMS AND BACKGROUND: Nausea and vomiting occur in the majority of patients receiving cisplatin (CDDP) chemotherapy. Ondansetron, a new 5-HT3 receptor antagonist, has been used effectively to control CDDP-induced nausea and vomiting. This study examined the potential of ondansetron to interfere with CDDP antitumor activity and toxicity in Ehrlich ascites carcinoma (EAC). METHODS: The influence of ondansetron on CDDP cytotoxicity was evaluated using EAC cells in culture. In addition, the influence of ondansetron pretreatment on CDDP-induced antitumor activity and host tissue toxicity was studied in EAC-bearing mice. RESULTS: Ondansetron (0.25 microM) enhanced CDDP (0-32 microM) cytotoxicity against EAC cells in vitro. In EAC-bearing mice ondansetron (0.2 mg/kg,ip) administered 1 h before CDDP (7 mg/kg, ip) did not modify the antitumor activity of CDDP. CDDP (7 mg/kg, ip) single treatment induced significant increases in blood urea nitrogen (2-fold) and serum creatinine (2.5-fold) and significant decreases in hematocrit (25%) and white blood cell count (39%) compared to saline treatment. Mice receiving ondansetron 1 h before CDDP showed no significant enhancement of CDDP-induced nephrotoxicity or myelosuppression compared to those pretreated with saline receiving the same dose of CDDP. CONCLUSIONS: This study suggests that the use of ondansetron to control CDDP-induced nausea and vomiting does not affect CDDP antitumor efficacy.

Differential alteration of cisplatin cytotoxicity and myelotoxicity by the paclitaxel vehicle cremophor EL.

Cremophor EL (CR), the paclitaxel vehicle, has previously been reported to alter the pharmacokinetics and/or pharmacodynamics of some anticancer drugs including paclitaxel. Several experimental and clinical studies suggested that cisplatin (CDDP) in combination with paclitaxel results in less hematological toxicity than anticipated. To reveal the role of CR in this important pharmacological interaction, we evaluated the interaction of CR with CDDP in vitro and in vivo using experimental Ehrlich ascites carcinoma (EAC) tumor. CR (1 microg/ml) significantly enhanced the in vitro cytotoxicity of CDDP in cultured EAC cells. This enhancement was not associated with a parallel increase in CDDP cellular uptake. In tumor-bearing mice, CR (2.5 ml/kg, i.v.) given in combination with CDDP (7 mg/kg, i.v.) did not significantly change CDDP pharmacokinetics, antitumor activity or nephrotoxicity. On the other hand, CDDP-induced hematological toxicity was significantly reduced by CR. This protective effect was related to CR-induced inhibition of cellular CDDP accumulation in bone marrow. This study presents evidence that CR may play an important role in the pharmacological interaction between CDDP and paclitaxel. The present data may suggest formulation of CDDP with CR for systemic treatment. Further studies are yet necessary to establish the clinical value of CR as a modifier for CDDP therapeutic index.

p53-degradation by HPV-16 E6 preferentially affects the removal of cyclobutane pyrimidine dimers from non-transcribed strand and sensitizes mammary epithelial cells to UV-irradiation.

Nucleotide excision repair (NER), the most versatile and ubiquitous mechanism for DNA repair, operates to remove many types of DNA base lesions. We have studied the role of p53 function in modulating the repair of DNA damage following UV irradiation in normal and p53-compromised human mammary epithelial cells (HMEC). The effect of UV-induced DNA damage on cellular cytotoxicity and apoptosis was determined in conjunction with global, gene- and strand-specific repair. Cytotoxicity studies, using clonogenic survival and MTT assays, showed that HPV-16 E6-expressing HMEC were more UV sensitive than p53-WT cell lines. High apoptotic index obtained with p53-compromised cells was in conformity to both the low clonogenic survival and the low cellular viability. No discernible differences in the formation of initial UV-induced cyclobutane pyrimidine dimers (CPD) were observed in the cell lines of varying p53 functional status. However, the extent and the rate of damage removal from genome overall were highest for p53-WT cells. Further examination of strand-specific repair in the p53 gene revealed that the removal of CPD in the non-transcribed strand (NTS) was slower in p53-compromised cells compared to the normal p53-WT cell lines. These results suggest that loss of p53 function, in the absence of other genetic alterations, decreased both overall amount of CPD repaired and their removal rate from the genome. Additionally, normal function of p53 is required for the repair of the NTS, but not of the transcribed strand (TS) in genomic DNA in human epithelial cells. Thus, failure of quantitative removal of CPD by global genomic repair (GGR), due to loss of p53 function, causes the enhanced UV sensitivity and increased damage-induced apoptosis via a p53-independent pathway. Nevertheless, recovery of cells from UV damage requires normal p53 function and efficient GGR.

Selective agonists of retinoic acid receptors: comparative toxicokinetics and embryonic exposure.

Three biologically active synthetic retinoids were investigated that bind selectively to retinoic acid receptors RARs (alpha, beta and gamma). The retinoids were previously demonstrated to have different teratogenic effects in the mouse in terms of potency and regioselectivity. The teratogenic potency rank order (alpha >beta >gamma) was found to be more or less compatible with the receptor binding affinities and transactivation potencies of the retinoid ligands to their respective receptors. The RARalpha agonist (Am580; CD336) induced a wide spectrum of malformations; CD2019 (RARbeta agonist) and especially CD437 (RARgamma agonist) produced more restricted defects. In the current study we tried to address whether the differences in teratogenic effects are solely related to binding affinity and transactivation differences or also due to differences in embryonic exposure. Therefore, transplacental kinetics of the ligands were assessed following administration of a single oral dose of 15 mg/kg of either retinoid given to NMRI mice on day 11 of gestation. Am580 was rapidly transferred to the embryo resulting in the highest embryonic exposure [embryo to maternal plasma area under the time vs concentration curve (AUC)(0-24 h )ratio (E/M) was 1.7], in accordance with its highest teratogenic potency. The low placental transfer of CD2019 (E/M of 0.3) was compatible with its lower teratogenic potential. Of major interest was the finding that the CD437, though being least teratogenic, exhibited considerable embryonic exposure (E/M of 0.6). These findings suggest that both the embryonic exposure and receptor binding transactivation selectivity are crucial determinants of the teratogenicity of these retinoid ligands.

Adverse testicular effects of some quinolone members in rats.

In the last few years, a marked decrease in male fertility has been reported. Environmental factors were recently suspected for this effect. Among those factors is the misuse of drugs and in particular antibiotics. Quinolones are a group of antibacterial agents with broad-spectrum activity. Testicular impairment of some quinolone members is controversial; a matter which stimulated our attention to investigate the adverse testicular effects of the most familiar quinolone members, namely: ofloxacin, ciprofloxacin and pefloxacin. They were given to rats in doses of 72, 135 and 72 mg kg(-1) day(-1) p.o., respectively, for 15 consecutive days. Ofloxacin was also used to establish a dose-response relationship in doses of 36, 72 and 360 mg kg(-1) day(-1) p.o. for 15 consecutive days. Results revealed that ofloxacin, ciprofloxacin and pefloxacin reduced testicular LDH-X activity by 39.8%, 62.7% and 60.7%, respectively. Moreover, sperm count, motility and daily sperm production were markedly decreased. Ofloxacin induced a dose-dependent decrease in testicular LDH-X activity, sperm count and motility. Furthermore, daily sperm production showed a marked reduction which amounted to 26.1% and 40. 0% following administration of ofloxacin (72, 360 mg kg(-1) day(-1) x 15 days), respectively. Moreover, administration of ofloxacin resulted in marked testicular histopathological changes. It is concluded that, ofloxacin, ciprofloxacin and pefloxacin significantly impaired both testicular function and structure in rats.

The influence of thymoquinone on doxorubicin-induced hyperlipidemic nephropathy in rats.

The effect of thymoquinone (TQ), the main constituent of the volatile oil of Nigella sativa seeds, on the nephropathy and oxidative stress induced by doxorubicin (DOX) in rats was investigated. A single intravenous injection of DOX (6 mg/kg) induced a severe nephrotic syndrome (after 5 weeks) associated with hypoalbuminemia, hypoproteinemia, elevated serum urea, hyperlipidemia, and a high urinary excretion of protein, albumin and N-acetyl-beta-D-glucosaminidase (NAG). In the kidney, DOX induced a significant increase in total triglycerides (TG), total cholesterol (TC), and lipid peroxides and a significant decrease in non-protein sulfhydryl (NPSH) content and catalase (CAT) activity. Treatment of rats with TQ (10 mg/kg per day) supplemented with the drinking water for 5 days before DOX, and daily thereafter, significantly lowered serum urea, TG, and TC. Similarly, TG, TC and lipid peroxides in the kidneys of TQ-treated rats were decreased significantly compared with DOX alone. Moreover, NPSH content and CAT activity in the kidneys of TQ-treated DOX group were significantly elevated compared with DOX alone. Treatment with TQ significantly suppressed DOX-induced proteinuria, albuminuria, and urinary excretion of NAG. The results confirm the involvement of free radicals in the pathogenesis of nephropathy induced by DOX. Likewise, the study demonstrates the high antioxidant potential of TQ and its marked effect on the suppression of DOX-induced nephropathy. The data suggest that TQ might be applicable as a protective agent for proteinuria and hyperlipidemia associated with nephrotic syndrome.

Prevention of cisplatin-induced nephrotoxicity by methimazole.

Nephrotoxicity is a dose-limiting factor in the use of cisplatin against solid tumours. Methimazole, an antithyroid drug containing a free SH group, has a nephroprotective potential against chemically-induced nephrotoxicity. We tried to explore the nephrotoxic effect of the experimentally therapeutic dose of cisplatin (7 mg kg(-1), i.p.), particularly on the nuclear level of kidney cells in male albino rats, as well as the possible protective effect of methimazole. Furthermore, the drug interaction regarding the oncolytic effect of cisplatin was examined in Ehrlich ascites carcinoma (EAC)-bearing mice. A single dose of cisplatin caused kidney damage, 6 days after injection, manifested by 219% increase in serum creatinine, 384% increase in blood urea nitrogen and 170% increase in kidney content of lipid peroxides. Kidney DNA showed clear fragmentations detected by gel electrophoresis. However, kidney reduced glutathione was unchanged at that time period. Histological examination of kidney confirmed the toxic effect of cisplatin. Methimazole (40 mg kg(-1), i.p., 30 min before cisplatin injection) significantly protected the kidney from the nephrotoxic effect of cisplatin as judged from the biochemical parameters investigated as well as the histopathological examination. On the other hand, the survival data in EAC-bearing mice treated with both drugs indicated the persistence of an effective cytotoxic action. This study points to a promising use of this combination and necessitates further experimental and clinical studies.

Renal metabolism of acrylonitrile to cyanide: in vitro studies.

Acrylonitrile (VCN) is a widely used industrial chemical. The present work examines the mechanism of its renal toxicity. In renal centrifugal fractions from Sprague-Dawley rats, the metabolism of VCN to cyanide (CN(-)) was highest in the microsomal fraction and required a NADPH-generating system in the presence of magnesium ions for maximum activity. This biotransformation of VCN to CN(-)was characterised with respect to time (15 min), microsomal protein concentration (3 mg ml(-1)), pH (7.5) and temperature (37 degrees C). The V(max)of the reaction was 118.2 pmol CN(-)mg(-1)protein min(-1)and K(m)was 160.2 micromol VCN. Activation of VCN to CN(-)was markedly increased in microsomes obtained from phenobarbital (PB), ethanol, 4-methylpyrazole and 3-methylcholanthrene-treated rats by 161.5, 89.6, 71.0 and 50.2%, respectively. Addition of SKF 525-A (5x10(-4)m) or benzimidazole (2 m m) to the incubation mixtures significantly inhibited VCN metabolism by 66.6 and 78.8%, respectively. VCN metabolism to CN(-)was enhanced significantly by the addition of 10 m m of glutathione (GSH), l -cysteine, d -penicillamine, cysteamine or 2-mercaptoethanol to 389.5, 886.5, 611. 1, 145.5 and 384.0% of control, respectively. These findings indicate that VCN is metabolised in the kidney via cytochrome P-450-dependent mixed function oxidase system. 1999 Academic Press.

Effect of bromocriptine on uterine contractility in near-term pregnant rats.

Treatment of pregnant albino rats at gestation day 9 with the dopamine agonist, bromocriptine, in a dose of 0.7 mg kg-1 day-1, i.p. for 11 days produced a significant increase in the normal uterine contractions both in vitro and in vivo. The increase in frequency (F), amplitude (A) and area under the curve (AUC) in the in vitro experiment amounted to 35, 80 and 58%, respectively; while the increase in F and A in the in vivo experiment was 36 and 25%, respectively, in comparison with the corresponding control group. Addition of oxytocin (5x10(-12)-4x10(-11) m) to the uterus isolated from rats pretreated with bromocriptine resulted in marked uterotonic effect (24, 35 and 49% increase in F; 25, 35 and 46% increase in A and 42, 62 and 122% increase in AUC of contractions). Also, the in vivo experiment showed that an injection of oxytocin at the time of investigation (0.125-1.0 I.U. kg-1, i.v.) into rats pretreated with bromocriptine caused a marked increase in F (33, 40 and 81%) and A (33, 37 and 75%) of uterine contractions compared to the values of bromocriptine-treated animals. These results indicate that bromocriptine should be used with caution during pregnancy. In addition, this must be considered when using oxytocin during delivery of females pretreated with bromocriptine.

Possible functional immunotoxicity of acrylonitrile (VCN).

Acrylonitrile (vinyl cyanide, VCN), an environmental pollutant, has been shown to be an animal and human carcinogen particularly for the GIT. In a previous work done in our laboratory, VCN induced immunosuppressive effects as indicated by a decrease in plaque forming cell (PFC) response to SRBCs (sheep red blood cell) immunization, a marked depletion of spleen lymphocyte subsets by flow cytometric analysis as well as bacterial translocation of the normal flora leading to brachial lymph node abscess. This work was carried out to evaluate the systemic and/or local immunotoxic potential of VCN. Acrylonitrile (2.7 mg kg-1 day-1) was given to CD-1 mice once daily for 5, 10 and 15 days. Immunohistochemical assessment of the number of cells capable of producing IgA in different intestinal compartments (duodenum, jejunum and ileum) revealed a significant decrease following VCN treatment. On the contrary, Bromodeoxyuridine (BrdU) incorporation in gut epithelial cells (duodenum and ileum) showed a significant increase in the same VCN-treated groups of animals. On the other hand, [3H]thymidine uptake was significantly decreased in splenocytes stimulated with phytohemaglutinin (PHA), Concanavalin-A (Con-A) and Lipopolysaccharide (LPS) and derived from animals treated with VCN. The effects of VCN were started after 5 days and increased up to 15 days of daily treatment in most of the investigated parameters. The results suggested that VCN has a profound immunosuppressive effect either systemically or locally which could be a contributing factor in its GIT carcinogenicity.

Identification of metallothionein receptors in human astrocytes.

Metallothionein (MT) isoforms are low molecular weight (6000-7000 Da) zinc binding proteins containing 60-68 amino acid residues, 25-30% cysteine, no aromatic amino acids, and binding between 5-7 g zinc/mol of protein. Since the synthesis of MT is induced by endotoxin, cytokines, and glucocorticoids, MT is now considered to be an acute phase protein protecting against oxygen radicals and oxidative damages caused by inflammation, tissue injury, and stress to the central nervous system. By postulating that a specific mechanism must exist to foster the induction of MTs I and II by numerous and diversified factors, we searched for and identified for the first time, MT receptors on U373MG cell membrane preparations, by using fluoresceinated MT I isoform probe; and by employing cysteine, glutathione, and four MT isoforms to determine high affinity and specific binding. MT receptors revealed a Kd value of 0.84 nM and a Bmax of 99.82 fmol/mg protein. Moreover, MT receptors were found in greater density on the surface of aggregated astrocytes. We postulate that conditions or agents generating reactive oxygen species may influence the expression of MT receptors.