Search

Epidemiological studies related to androgens and prostate cancer (PC) have focused on serum determination of testosterone, androstenedione (A4), and DHEA, with inconsistent results. Herein, we hypothesized that differences in androgen biosynthetic and metabolic pathways, rather than differences in specific androgen concentrations, are associated with prostatic carcinogenesis. Therefore, spot urine samples from 111 incident PC cases with Gleason score at diagnosis and 227 healthy population controls, were analyzed. Urinary androgen concentrations (nanograms/milligrams of creatinine) were determined by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS). Using a factor analysis, we identified three androgen urinary excretion patterns. In a subsample, we evaluated a modification effect of the androgen receptor (AR) CAG polymorphism. Pattern I, characterized by A4 and testosterone hydroxylated metabolites (11β-OHT; 2β-OHT; 15β-OHT; 2α-OHT; 6β-OHT), was associated with high PC odds among carriers of AR gene (CAG)>19 repeats (OR: 3.67 95% CI: 1.23–11.0; P for interaction= 0.009). Conversely, higher testosterone excretion (pattern III), was marginally associated with lower (OR: 0.35 95% CI: 0.12–1.00, P for trend= 0.08) poorly differentiated PC (Gleason ≥8). No clear association was observed with pattern II (DHEA; 16α and 16β-OHT). Our results were consistent with the previous evidence which suggests that the C11-oxy backdoor pathway is important for prostatic carcinogenesis. Androgen urine excretion analysis could be useful for PC diagnosis, treatment, and prognosis; however, further studies with a larger number of samples and the urinary determination of 11-ketoandrogens are necessary.

Temephos is an organophosphorus pesticide used in control campaigns against vectors that transmit diseases, including dengue, a public health concern. The WHO classifies temephos in category III and its safe concentration (low-observable-adverse-effect level) in male rats is 100 mg/kg/day for up to 44 days. Temephos inhibits acetylcholinesterase (AChE) and is metabolized in different tissues, probably by mixed-function oxidases; one of its metabolites is bisphenol S (BPS), which is considered an endocrine disruptor. The aim of this study was to evaluate the effects of temephos on sperm function and its biotransformation in the testis, epididymis, and other tissues to explore its toxicity in rats treated with 100 mg/kg/day/5 or 7 days (gavage). AChE activity was inhibited 70% starting on day 3 and 13 or 41% mortality was observed at 5 or 7 days, respectively. After 7 days, temephos significantly decreased sperm motility (30%) and viability (10%) and increased (10%) lipoperoxidation, and the sperm DNA exhibited no damage. Temephos was distributed and metabolized in all tissues, with the highest levels observed in the adipose tissue and temephos levels were 16-fold higher in the epididymis than in the testis. Notably, BPS was observed in the testis. At 5 days, decreased sperm motility (12.5%) and viability (5.7%) were observed and sperm fertilization decreased (30%). These results suggest that temephos decreases sperm quality and fertilization capacity at recommended safe concentrations and that it is metabolized in male reproductive tissues. This pesticide places the reproductive health of exposed people at risk, suggesting the need to reevaluate its toxicity.

Temephos (Tem) is the larvicide of choice to control mosquito transmission of dengue, Zika, and chikungunya. The toxicokinetic and toxicological information of temephos is very limited. The aim of this work was to determine the toxicokinetics and dosimetry of temephos and its metabolites. Male Wistar rats were orally administered temephos (300 mg/kg) emulsified with saline solution and sacrificed over time after dosing. Temephos and its metabolites were analyzed in blood and tissues by high performance liquid chromatography-diode array detector. At least eleven metabolites were detected, including temephos-sulfoxide (Tem-SO), temephos-oxon (Tem-oxon), temephos-oxon-sulfoxide (Tem-oxon-SO), temephos-oxon-SO-monohydrolyzed (Tem-oxon-SO-OH), 4,4´-thiodiphenol, 4,4´-sulfinyldiphenol, and 4,4´-sulfonyldiphenol or bisphenol S (BPS). The mean blood concentrations of temephos were fitted to a one-compartment model for kinetic analysis. At 2 h, the peak was reached (t1/2 abs = 0.38 h), and only trace levels were detected at 36 h (t1/2 elim = 8.6 h). Temephos was detected in all tissues and preferentially accumulated in fat. Temephos-sulfone-monohydrolyzed (Tem-SO2-OH) blood levels remained constant until 36 h and gradually accumulated in the kidney. Tem-oxon was detected in the brain, liver, kidney, and fat. Clearance from the liver and kidney were 7.59 and 5.52 ml/min, respectively. These results indicate that temephos is well absorbed, extensively metabolized, widely distributed and preferentially stored in adipose tissue. It is biotransformed into reactive metabolites such as Tem-oxons, Tem-dioxons, and BPS. Tem-SO2-OH, the most abundant metabolite of temephos, could be used as an exposure biomarker for toxicokinetic modeling. These results could provide critical insight into the dosimetry and toxicity of temephos and its metabolites.

Infection with high-risk human papillomavirus (HR-HPV) is the main cause of cervical cancer (CC), but viral infection alone does not guarantee the development of this malignancy. Indeed, deficiencies of dietary micronutrients could favor cervical cancer development in individuals that harbor HR-HPV infections. The status of retinoid levels, natural and synthetic derivatives of vitamin A, is important in maintaining cellular differentiation of the cervical epithelium. Moreover, many studies show a link between deficient intake of retinoids or alteration of the retinoid receptors and CC development. In spite of this, the effect of vitamin A deficiency (VAD) in presence of HR-HPV oncoproteins on cervical carcinogenesis in vivo has not been reported. Transgenic mice expressing E6 or E7 oncoproteins (K14E6 or K14E7 mice, respectively) were used to evaluate the possible role of VAD in the development of malignant cervical lesions. The survival of the mice in VAD condition was studied, and histopathological analysis and immunohistochemical detection of molecular cancer markers such as the tumor suppressor retinoic acid receptor beta (RARβ), proliferating cell nuclear antigen (PCNA), cleaved caspase 3, and the tumor suppressor protein p16INK4A (inhibitor of CDK4) were performed. Our results show that K14E6/VAD mice showed moderate cervical dysplasia; notably, K14E7/VAD mice developed severe cervical dysplasia and cervical in situ carcinoma at an early age. VAD synergizes with HPV16E7 oncoprotein expression favoring cervical carcinogenesis in vivo.

Temephos (Tem) is an organophosphorus pesticide widely used to kill and prevent the growth of the main vectors for the transmission of dengue, zika, and chikungunya viruses. In chlorinated water, Tem is oxidized to its dioxon-sulfoxide (Tem-dox-SO), dioxon-sulfone (Tem-dox-SO2), and sulfoxide (Tem-SO) derivatives; however, these compounds are not commercially available to be used as standards and in toxicological studies. In the present study, we synthesized and characterized the Tem-oxidation products and the compound 4,4′-sulfinyldiphenol. These compounds were obtained by a simple reaction between Tem or 4,4′-thiodiphenol with sodium hypochlorite or potassium periodate, and were characterized by IR, NMR, and UPLC-HRESIMS. The in vitro evaluation of inhibitory potency of Tem-oxidized products on human red blood cell acetylcholinesterase (RBC AChE) showed that Tem-dox-SO2 was the most potent inhibitor of human RBC AChE, and its effect was more pronounced than that observed for ethyl-paraoxon, a potent typical inhibitor of AChE. An HPLC-DAD method for the analysis of metabolic products of Tem was developed, which may be useful for monitoring in biological and environmental samples. The ability of Tem-oxidized metabolites to inhibit human RBC AChE suggests that the addition of Tem to chlorinated drinking water could result in an increase in the risk of RBC AChE inhibition after exposure.

Follicle growth culminates in ovulation, which allows for the expulsion of fertilizable oocytes and the formation of corpora lutea. Bisphenol A (BPA) is present in many consumer products, and it has been suggested that BPA impairs ovulation; however, the underlying mechanisms are unknown. Therefore, this study first evaluated whether BPA alters ovulation by affecting folliculogenesis, the number of corpora lutea or eggs shed to the oviduct, ovarian gonadotropin responsiveness, hormone levels, and estrous cyclicity. Because it has been suggested (but not directly confirmed) that BPA exerts toxic effects on the fertilization ability of oocytes, a second aim was to evaluate whether BPA impacts the oocyte fertilization rate using an in vitro fertilization assay and mating. The possible effects on early zygote development were also examined. Young adult female C57BL/6J mice (39 days old) were orally dosed with corn oil (vehicle) or 50 μg/kgbw/day BPA for a period encompassing the first three reproductive cycles (12-15 days). BPA exposure did not alter any parameters related to ovulation. Moreover, BPA exposure reduced the percentage of fertilized oocytes after either in vitro fertilization or mating, but it did not alter the zygotic stages. The data indicate that exposure to the reference dose of BPA does not impact ovulation but that it does influence the oocyte quality in terms of its fertilization ability.

Vinclozolin (V) is classified as a potent endocrine disruptor. The aim of the present study was to determine the effects of V on rat liver CYP regulation and on serum levels of testosterone and estradiol during pregnancy. Pregnancy decreased the liver total CYP content by 65%, enzyme activities of MROD, PROD, and PNPH, and testosterone hydroxylation activities, as well as the protein content of CYP2A and 3A. V exposure remarkably induced the protein content and enzyme activities of CYP1A, 2A, 2B and 3A subfamilies. Testosterone and estradiol were affected in an opposite manner, provoking a 3.5-fold increase in the estradiol/testosterone ratio. These results suggest that V could regulate the hepatic CYP expression through interaction with receptors and coactivators involved in its expression and may play an important role in hormonal balance during pregnancy. In addition, the results may also contribute to understanding the toxicity of V by in utero exposure.

Hepatocellular carcinoma (HCC) arises after a long period of exposition to etiological factors that might be either independent or collectively contributing. Several rodent models resemble human HCC; however, the major limitation of these models is the lack of chronic injury that reproducibly mimics the molecular alterations as it occurs in humans. Thus, we hypothesized that chronic administration of different DEN treatments identifies the best-fit dose to induce the HCC and/or to determine whether small DEN doses act synergistically with other known hepatotoxins to induce HCC in mice. C57BL/6 J male mice were intraperitoneally injected twice a week for 6 weeks with different DEN doses ranging from 2.5 to 40 mg/kg body weight; then, selected doses (2.5, 5 and 20 mg/kg) for 6, 10, 14, and 18 weeks. We demonstrated that DEN at 20 mg/kg promoted reactive oxygen species and 4-hydroxynonenal production, cell proliferation inflammatory infiltrate, and fibrosis, which in turn induced liver cancer by week 18. These parameters were established by evaluating histopathological changes, HCC markers such as glutathione S-transferase placental-1 (Gstp1), Cytokeratin-19 (Ck19) and prostaglandin reductase-1 (Ptgr1); that of Cyp2e1, a DEN metabolizing enzyme; and the expression of the proliferation marker Ki67. While DEN at 2.5 and 5 mg/kg increased Gstp1 and Ck19, DEN at 20 mg/kg decreased them and Cyp2e1 expression and activity. In summary, our results demonstrate that DEN chronically administrated at 20 mg/kg induces the HCC, while DEN at 2.5 and 5 mg/kg could be useful in elucidating its synergistic effect with other hepatotoxic agents in mice.

Objective: To evaluate if variants in the genes CYP1A1 (T3801C and A4889G), CYP1B1 (G119T), GSTM1 (indel) and GSTT1 (indel) are associated with breast cancer (BC) among Mexican women. Materials and methods: 952 incident cases with histologically confirmed BC were matched by age (± 5 years) and zone of residence with 998 healthy population controls. Genetic variants in genes CYP1A1, CYP1B1, GSTM1 and GSTT1were genotyped by allelic discrimination and multiplex PCR. In a subsample of women, 105 markers for ancestry were determined. Results: An increased BC risk, independent of other BC risk factors, was observed among carriers of CYP1B1 G119T genotype (T/T vs. G/G: OR=1.9; 95%CI 1.4-2.5). Conclusion: Our results support the existence of genetic susceptibility for BC conferred by CYP1B1 G119T variant among Mexican women.

Vinclozolin (V) is a fungicide with antiandrogenic properties. To determine the pharmacokinetics and dosimetry of V, adult male rats were administered an oral dose of V (100 mg/kg) in corn oil and sacrificed over time after dosing. V and its metabolites were analyzed in serum and tissues by high performance liquid chromatography/diode array detector/mass spectrometer. V, 2-[[(3,5-dichlorophenyl)-carbamoyl]oxy]-2-methyl-3-butenoic acid (M1), and 3',5'-dichloro-2-hydroxy-2-methylbut-3-enanilide (M2), and five other metabolites were detected in serum and tissues. One metabolite was identified as 3',5'-dichloro-2,3,4-trihydroxy-2-methylbutylanilide (M5). The mean serum concentration data for V were fitted to a one-compartment model for kinetic analysis. At 2 h, V serum concentration peaked; whereas only trace levels were detected at 24 h (t(1/2 elim) = 3.6 h). V was detected in all tissues and preferentially accumulated in fat. M1 serum levels increased until 8 h, being at least 2-fold higher than those of V at this time, and then declined with a t(1/2) = 3.3 h. M5 was the main metabolite in serum and tissues. Serum M5 levels were 5-fold higher than V and 2-fold greater than M1 at all times. At 48 h, M5 remained the main metabolite (t(1/2 elim) = 13.1 h). Liver and kidney exhibited the highest levels of M5, V, and M1. M2 and 3,5-dichloroaniline had the lowest levels of V metabolites in serum and tissues. V is well absorbed, extensively metabolized and widely distributed. M5, the most abundant V metabolite, may be used as an exposure biomarker for pharmacokinetic modeling. These results may clarify the relationship between toxicity and tissue dose of V and its metabolites.

Cyclohexanol is a basic industrial chemical widely used because of its versatility as an industrial solvent. No studies have been conducted to evaluate the carcinogenic/co-carcinogenic hazards associated with cyclohexanol exposure. In male Fisher 344 rats liver preneoplastic lesions were induced by N-nitrosodiethylamine (150 mg/Kg) i.p., followed by the tumor promoter 2-acetylaminofluorene (2-AAF: 20 mg/kg) orally administered on three consecutive days before partial hepatectomy. The cyclohexanol administration in this hepatocarcinogenesis assay revealed that it has a strong tumor co-promoter potential. There is clear evidence that oxidative stress and the CYP2E1 are components of carcinogenesis. Although no changes in the lipid peroxidation levels were observed between treated and untreated animals, a significant increase in CYP2E1 expression was observed when cyclohexanol was administered 24 h after the last 2-AAF dose. On the other hand, levels of the proliferation markers PCNA and Ki-67 were not increased after treatment with cyclohexanol, but a marked downregulation of the Bax proapoptotic protein was found exclusively in mitochondrial extracts of animals treated with cyclohexanol. This study represents the first report of the ability of cyclohexanol-induced lesions, when administered simultaneously with 2-AAF, to potentiate the development of preneoplastic liver.

DDT and its metabolites are considered as endocrine disruptors able to promote hormone-dependent pathologies. We studied the effects of technical-grade DDT on hepatic testosterone metabolism and testosterone hydroxylase activity ratios in the rat. Male and female Wistar rats were treated by gavage with a single dose of technical-grade DDT (0, 0.1, 1, 10, and 100 mg/kg body weight) and killed 24 h later. Hepatic microsomes were incubated with [4-14C]-testosterone and the metabolites were separated by thin-layer chromatography and quantified by radio scanning. DDT increased testosterone biotransformation and modified the profile of metabolites produced in a sex-dependent manner. Males treated with a representative dose (10 mg/kg) produced relatively less androstenedione (AD), 2alpha-hydroxytestosterone (OHT), and 16alpha-OHT but higher 6beta-OHT whereas treated females produced less 7alpha-OHT and AD but higher 6beta-OHT and 6alpha-OHT than their respective controls. In both sexes DDT decreased the relative proportion of AD and increased that of 6beta-OHT suggesting that the androgen-saving pathway was affected. The testosterone 6alpha-/15alpha-OHT ratio, a proposed indicator of demasculinization, was increased in treated males. This effect was in agreement with the demasculinizing ability proposed for DDT. The effects on 6alpha-/16alpha-OHT and 6-dehydrotestosterone/16alpha-OHT ratios followed a similar tendency, with the ratio 6alpha-/16alpha-OHT being the most sensitive marker. Interestingly, these ratios were reduced in treated females suggesting that technical-grade DDT shifted testosterone hydroxylations toward a more masculine pattern. Thus, technical-grade DDT altered the hepatic sexual dimorphism in testosterone metabolism and decreased the metabolic differences between male and female rats.

The role of cytochrome P450 (CYP) and the CYP isoform involved in the activation of the widely used pesticide methyl-parathion (MePA) were investigated in rat brain extracts by measuring the effect of different CYP inhibitors on acetylcholinesterase (AChE) inhibition by MePA. Brain extracts provide a useful tool to study the activation mechanisms of organophosphorus compounds (OP) since they contain both the activating enzyme(s) and the molecular target for OP toxicity. As expected, in incubations of rat brain extract supplemented with NADPH, AChE activity was non-competitively inhibited by the presence of MePA, indicating that MePA was activated to its reactive metabolite methyl-paraoxon (MePO). Indeed, Vmaxapp decreased from 13.4 to 8.7 mol thionitrobenzoic acid (TNB)/min per mg protein. MePA activation by rat brain extracts, as measured by the AChE inhibition produced by the presence of the pesticide in the incubation, was fully prevented by previously bubbling the incubation mix with CO, by the presence of monoclonal anti-rat CYP2B1/2B2 antibodies and by the addition of phenobarbital (PB), a CYP2B substrate. Interestingly, MePA showed a greater affinity for CYP2B than PB. CYP1A1 antibodies showed no effect on MePA activation. The presence of cytochrome P450 2B (CYP2B) in the rat brain extracts was confirmed by immunoblotting. These results demonstrate indisputably the responsibility of CYP2B in MePA activation in the rat brain in vitro, suggesting that metabolic activation of OP compounds in situ might be crucial for their organ specific toxicity to the central nervous system also in vivo. © 2001 Elsevier Science Ireland Ltd. All rights reserved.

Studies of cancer chemoprevention with caffeic acid phenethyl ester (CAPE) in the resistant hepatocyte model of hepatocarcinogenesis have shown the participation of CYP drug metabolizing enzymes. To prevent neoplastic and preneoplasic lesions, we must specifically identify which CYP activities are modified in the mechanism of action of CAPE. Male Fischer-344 rats were pretreated with CAPE twelve hours before administration of diethylnitrosamine (DEN) and were sacrificed twelve hours after CAPE and twelve hours, twenty-four hours, twenty-four days, and twelve months after DEN. Other rats were treated with the CYP inhibitors α-naphthoflavone or SKF525A and sacrificed twenty-four hours and twenty-four days after DEN. Microsomes were obtained from livers to quantify protein using Western blot. Diethylnitrosamine metabolism was measured based on nitrite formation and liver histology using GGT histochemistry. Caffeic acid phenethyl ester diminished the protein levels of CYP1A2 and CYP2B1/2. The inhibition of CYP2B1/2 prevented the appearance of preneoplastic lesions. Microsomal assays demonstrated that CAPE interfered with DEN activation diminishing nitrites similar to SKF525A and probably mediated by CYP2B1/2 inhibition. A single dose of CAPE before DEN treatment reduced the appearance of tumors by 43%. These results confirmed that CAPE is a promising agent to confer chemoprotection in liver cancer and should be considered for human therapies.

Caffeic acid phenethyl ester (CAPE), a natural component of propolis, shows anticarcinogenic properties in the modified resistant hepatocyte model when administered before initiation or promotion of hepatocarcinogenesis process; however, information about the mechanism underlying this chemoprotection is limited. The aim of this work was to characterize the effect of CAPE on cytochrome P450 (CYP), which is involved in diethylnitrosamine (DEN) metabolism during the initiation stage of chemical hepatocarcinogenesis. Male Fischer-344 rats were treated as in the modified resistant hepatocyte model. Liver samples were obtained at four different times: at 12 h after pretreatment with CAPE and at 12 and 24 h and 25 days after DEN administration. Liver damage was determined by histology with hematoxylin and eosin, measurement of total CYP levels and enzyme activity, and gamma-glutamyl transpeptidase-positive (GGT+) staining of hepatocyte foci. CAPE administration prevented DEN-induced necrosis at 24 h. It also decreased O-dealkylation of 7-ethoxy-resorufin (EROD), O-dealkylation of 7-methoxyresorufin (MROD), and 7-pentoxy-resorufin activities at 12 h after its administration and EROD and MROD activities at 12 h after administration of DEN. CAPE treatment decreased GGT+ foci by 59% on day 25. Our results suggest that CAPE modifies the enzymatic activity of CYP isoforms involved in the activation of DEN, such as CYP1A1/2 and CYP2B1/2. These findings describe an alternative mechanism for understanding the ability of CAPE to protect against chemical hepatocarcinogenesis.

The effect of 0, 1, 5, 25 and 50 mg/kg body weight (b.w.) lindane, administered in corn oil by gavage, on cytochrome P450 (CYP) phenotype was investigated in the liver of ovariectomized (ovx), sham operated (sham-ope) and nonovariectomized (n/ovx) adult Wistar rats. Total hepatic microsomal CYP content and the O-dealkylation of 7-ethoxy- (EROD), 7-methoxy- (MROD), 7-pentoxy- (PROD) and 7-benzyloxy-resorufin (BROD) were assayed. In addition, CYP1A1, 2B1/2B2, 2C11 and 3A2 proteins were determined by western blot using specific anti-rat antibodies. Protein bands were visualised by chemiluminescence and their intensities were compared among groups. A statistically significant, dose-dependent increase of all parameters studied was observed in all three animal groups after lindane administration. Ovx rats, however, responded differently to lindane administration than n/ovx or sham-ope animals. At the highest doses in ovx rats, the relative liver weight was more increased and the total CYP content was less increased than in n/ovx or sham-ope animals. Moreover, the degree of induction of PROD and BROD activities was higher and that of EROD activity was lower in ovx than in either n/ovx or sham-ope animals. Accordingly, CYP2B1/2B2 protein showed the largest increase in ovx rats, whereas CYP1A protein increased more in n/ovx or sham-ope animals. CYP2C11, a cytochrome predominantly expressed in males, was also more strongly induced in ovx than in n/ovx or sham-ope animals. CYP3A2 was slightly expressed in ovx but not in n/ovx non-treated rats, although the effect of induction was clearly greater in the latter. These results, overall, indicate that ovariectomy significantly affects, both qualitatively and quantitatively, CYP expression following induction by lindane and support the anti-estrogenic effect of lindane in rats. The pathophysiological and toxicological relevance of liver CYP induction by lindane and possibly other organochlorine xenobiotics in females with a lack or deficiency of estrogen supply remains to be investigated.

Dichlorodiphenyltrichloroethane (DDT) is a well-known inducer of microsomal monooxygenase systems in rodent liver. However, little information is available on its effects on the sex-dependent regulation of CYPs preferentially affected. Therefore, our objective was to evaluate the effects of DDT on the sexual expression pattern of some hepatic P-450 isozymes. Single doses of technical DDT (0, 0.1, 1, 5, 10, or 100 mg/kg body wt) were administered by gavage to Wistar rats. The effects on CYPs 1A1, 2B11/2B2, 2C11, 2E1, 3A1, and 3A2, were assessed 24 h later by means of CYP protein content determined by Western blotting and/or enzyme activities participating in alkoxyresorufin and pnitrophenol metabolism. The highest dose induced 18-fold the expression of CYP3A2 in female rats without producing significant induction (3-fold) in males. The effects on this isozyme, which is not normally expressed in females, suggest that DDT is able to modulate sexual metabolic dimorphism, as 3A2 expression is androgen dependent. DDT did not significantly alter CYP3A1 in males, suggesting that DDT is not a pure phenobarbital (PB)-type inducer. The effects on CYP2B1/2B2 protein (19-fold) and associated enzyme activities indicated that males had a lower response threshold than females, but that the latter were able to reach a higher relative induction. The preferential induction of CYPs 2B and 3A by DDT in a sex-related manner suggest that CYP regulation could play an important role in endocrine disruption.

The ability of chromium (Cr) salts to increase metallothionein (MT) levels in rat liver, kidney and pancreas, and its relationship with the presence of toxic effects are reported here. Rats were injected subcutaneously with 0, 10, 20, 30, 40, or 50 mg K 2 Cr 2 O 7 /kg and sacrificed 24 h later. Total Cr accumulation followed a dose-dependent pattern, levels in kidney being higher than those in liver or pancreas, suggesting different tissue bioavailabilities and accumulation patterns. Cr(IV) administration resulted in a tissue-specific MT induction: pancreas and liver showed five- and 3.5-fold MT increases, respectively; no increase was observed in the kidney. A positive correlation was observed between zinc and MT concentrations in liver, and between total Cr and MT concentrations in pancreas. Serum α-amylase activity showed a dose-dependent increase starting from 20 mg/kg, whereas serum glucose levels increased at doses higher than 30 mg/kg. Serum aspartate aminotransferase and alanine aminotransferase activities were increased in a dose-dependent manner, from 20 and 30 mg/kg, respectively. Our results showed that treatment with Cr(VI) can induce MT synthesis in pancreas and suggests a subsequent binding of Cr to MT. Also, pancreas is a target organ for Cr toxicity, and the usefulness of α-amylase activity as a sensitive biomarker of Cr toxicity in human exposed populations merits further study.

Urea cycle is a hepatic metabolic pathway involving five enzymes and several intermediary metabolites and can be altered by different chemicals. To investigate the effect of arsenic, an ubiquitous hepatotoxic agent, on urea production we exposed long-term cultures of adult rat hepatocytes, which produce urea, to 1.33 and 6.67 microM arsenite for 2 weeks. In cultures exposed to 6.67 microM, urea production decreased 60-70% and cellular arginase activity decreased 30, 70 and 85% after 4, 7 and 14 days of exposure, respectively. The arginase activity released to the medium increased significantly after 4, 7 and 14 days, with a maximum value after 7 days of exposure that was 27-fold higher than that of the untreated controls. The total arginase activity also decreased 35, 52 and 82% after 4, 7 and 14 days of exposure and protein content decreased 57 and 65% after 7 and 14 days of exposure, respectively. Exposure to 6.67 microM arsenite also produced accumulation of intracytoplasmic lipid droplets, vacuolizations and enlargement of the intercellular spaces. On the other hand, exposure of hepatocytes to 1.33 microM arsenite caused an initial decrease of 20% in urea production, did not change cellular, released and total arginase activity and cellular protein content and produced accumulation of intracytoplasmic lipid droplets. These results show that long-term exposure of cultured rat hepatocytes to 6.67 microM arsenite decreases urea production, cellular and total arginase activity and protein content and increases the release of arginase into the culture medium. These alterations could be useful markers of hepatotoxicity in in vitro assays.

AIM: To study the effect of celecoxib (CXB) on diethylnitrosamine activation through the regulation of cytochrome P450 in a hepatocarcinogenesis model. METHODS: Six-week-old male Sprague-Dawley rats were randomly divided into five groups, a non-treated group (NT), a diethylnitrosamine-treated group (DEN), a DEN+CXB-treated group (DEN+CXB), and CXB 8 d-treated and CXB 32 d-treated groups. The effects of celecoxib on the enzymatic activities of CYP1A1, 2A, 2B1/2, and 2E1 were assessed in hepatic microsomes 24 h after DEN administration. Changes in CYP1A1 and CYP2B1/2 protein expression were also evaluated. The rate of DEN metabolism was measured by the production of the deethylation metabolite acetaldehyde, and the denitrosation metabolite nitrite. RESULTS: DEN+CXB administration produced a significant increase in the enzymatic activities of CYP2B1/2 and 1A1, whereas it did not change the activities of CYP2A and 2E1, compared to that of the DEN group. CXB treatment for eight days did not produce a significant effect on enzymatic activity when compared to the NT group; however, when it was administered for prolonged times (CXB 32 d group), the enzymatic activities were increased in a similar pattern to those in the DEN+CXB group. The observed increase in the enzymatic activities in the DEN+CXB group was accompanied by an increase in the CYP2B1/2 protein levels; no changes were observed in the levels of CYP1A1. In vitro, CXB increased the denitrosation of DEN, a pathway of metabolic detoxification. The addition of SKF-525A, a preferential inhibitor of CYP2B, abrogated the denitrosation of DEN. CONCLUSION: These results suggest that the mechanism of action of CXB involves enhancement of the detoxification of DEN by an increasing denitrosation via CYP2B1/2.