The effect of filling technique on the cuspal strain, polymerization shrinkage stress, enamel crack formation and depth of cure of restored molars.

OBJECTIVE: Evaluate the effect of different restorative filling techniques on the residual shrinkage stress (ShrS), cuspal strain (CS), depth of cure (DC), and enamel crack formation (Ec) in molars with MOD restorations. METHODS: Post-gel shrinkage, elastic modulus, compressive and diametral tensile strength of the Filtek One Bulk Fill composite were calculated. Sixty molars with MOD preparations were restored using four filling techniques: Bulk; Horizontal; Oblique; Natural enamel and dentin substitution (NEDS) technique. CS was measured using a strain gauge (n = 10). The DC (n = 5) was measured using Knoop hardness. Shrinkage stress/strain was analyzed using 3D finite element analysis. The Ec analysis was carried out by transillumination. Two-way ANOVA with repeated measures and Tukey's HSD test (α = 0.05) was performed for the CS data. Two-Way ANOVA and Tukey's HSD test was performed for the DC data (α = 0.05). RESULTS: CS was higher at the lingual cusp for the horizontal and NEDS technique. No statistical difference was found between the buccal and lingual CS values for the Bulk (p = 0.367) or Oblique techniques (p = 0.192). CS values were lower for the Bulk. More enamel cracks were found for the Bulk. DC was lower at 4 mm regardless the filling technique. The Horizontal showed the highest ShrS values. The Bulk generated the lower ShrS values. SIGNIFICANCE: A Bulk technique caused the lowest shrinkage stress/strain. An Oblique technique yielded the best balance between stress, strain and crack formation. NEDS technique is a good alternative to decrease the number of increments while maintaining the stress levels nearby the Oblique technique.

Steroid hormones and first trimester vascular remodeling.

Successful implantation and placentation require neoangiogenesis and the remodeling of the uterine spiral arteries. Progesterone and estradiol control various of the placental functions, but their role in vascular remodeling remains controversial. Therefore, this chapter aims to summarize the current knowledge regarding the role of steroid hormones in the uteroplacental vascular remodeling during the first trimester of gestation.

Evaluation of α5ß1 integrin as a candidate marker for fertility in bull sperm samples.

With the progressive increase in the use of reproductive biotechnologies in the cattle industry, like artificial insemination and in vitro embryo production, the accurate determination of fertilizing competence of cryopreserved sperm samples is an essential issue. The routine methodology to assess bull sperm quality relies primarily on count, viability and motility of spermatozoa. However, these parameters do not tightly predict the reproductive success of samples. Therefore, identification of complementary markers of sperm functionality to strengthen the predictability of traditional spermogram is desirable to improve livestock reproduction practices. Previous results from our laboratory indicated that α5ß1 integrin plays a key role in bovine sperm function and mediates their interaction with the female reproductive tract. Thus, this study aimed to investigate whether the localization of α5ß1 held a correlation with fertilizing ability of bovine cryopreserved semen samples. Firstly, we assessed the quality of samples from six different bulls (A-F). We determined motility and viability of sperm samples after thawing and selection. Additionally, we measured the capacitation state of the samples by chlortetracycline (CTC) assay in the presence or absence of heparin, as an indicator of their responsiveness to a capacitating stimulus. Based on these assays, samples were classified being A the bull with the lowest quality and F the bull with the highest quality. Then, we studied the presence and localization of α5ß1 integrin. This protein showed a distribution pattern in the acrosomal (A), post-acrosomal (P) and acrosomal + post-acrosomal (A + P) regions with different localization percentages among the studied samples. Next, we determined the fertilizing ability of the samples in in vitro fertilization (IVF) assays and performed correlation analyses between IVF outcome and the routine spermogram parameters or α5ß1 integrin localization patterns. When the percentage of cells showing α5ß1 integrin was compared to fertilization rate, no correlation was observed. However, the presence of α5ß1 integrin in P and A + P regions (PA pattern), positively correlated with IVF rate (p < 0.05). These results suggest that while routine semen analyses failed to predict sperm reproductive competence, integrin localization in post-acrosomal region (PA pattern) showed a positive correlation with IVF outcome, thus posing an attractive marker to predict more accurately the reproductive performance of an individual.

Nitric oxide synthase 1 and cyclooxygenase-2 enzymes are targets of muscarinic activation in normal and inflamed NIH3T3 cells.

OBJECTIVE: Fibroblasts are sentinel cells that could serve as intermediaries in the immune reaction in the inflammatory process. In this work, we investigate the action of the muscarinic agonist carbachol (CARB) on the expression and function of nitric oxide synthase (NOS) and cyclooxygenase (COX) in fibroblasts under normal or inflammatory conditions. METHODS: The normal fibroblast cell line, 3T3, from NIH swiss mouse embryo, was used. The inflammatory milieu was mimicked with lipopolysaccharide (LPS) (10 ng/ml) plus interferon gamma (IFNgamma) (0.5 ng/ml). Nitric oxide (NO) and prostaglandin E(2) (PGE(2)) production were measured by Griess reagent and radioimmunoassay, respectively. NOS, COX, and nuclear transcription factor kappa B (NF-kappaB) were studied by Western blot. RESULTS: CARB increased NO synthesis by 57 +/- 5%, while a 150 +/- 10% increase in NO liberation was triggered by LPS plus IFNgamma treatment. CARB added to LPS plus IFNgamma potentiated NO synthesis by 227 +/- 19%. CARB also upregulated NOS1 protein expression via NF-kappaB activation. In addition CARB and LPS plus IFNgamma stimulated PGE(2) synthesis by 72 +/- 9 and 42 +/- 4%, respectively, while CARB added to LPS plus IFNgamma treated cells produced a synergism in PGE(2) liberation (130 +/- 12%) via COX-2. CONCLUSION: Activation of muscarinic acetylcholine receptors can mimic mild inflammatory conditions or can deepen pre-existing inflammation, establishing a fine-tuned set-up on fibroblasts that in turn could be alerting the immune system.

Real-time PCR in clinical practice: a powerful tool for evaluating Leishmania chagasi loads in naturally infected dogs.

The performance of the less expensive SYBR-Green-based PCR assay, for quantifying Leishmania chagasi in smears of bone-marrow aspirates from naturally infected, mongrel dogs, was recently compared with that of a similar PCR based on TaqMan chemistry. Aspirates were obtained from 36 infected dogs and examined for parasites by direct examination, culture, and quantitative PCR (qPCR) using specific primers (based on the parasite's kinetoplast DNA), DNA extracted from a smear, and either the SYBR-Green or TaqMan chemistries. Every aspirate smear was found PCR-positive for L. chagasi (whether the assay employed SYBR Green or TaqMan) but only 74% of the aspirates were found positive by culture and only 33% by direct, microscopical examination. There was no evidence of PCR inhibition when the DNA was collected from smears, and the parasite loads estimated using the SYBR-Green PCR were almost identical to those estimated using the TaqMan PCR (r=0.99). As a method for quantifying parasite loads in dogs infected with L. chagasi (and, probably, other mammals infected with other leishmanial parasites), PCR based on SYBR Green may therefore be an appropriate and inexpensive alternative to PCR based on TaqMan, and a reliable clinical tool.

Fibronectin induces capacitation-associated events through the endocannabinoid system in bull sperm.

Mammalian ejaculated spermatozoa must undergo a series of changes in the female reproductive tract, collectively called capacitation, in order to fertilize the oocyte. We reported that fibronectin (Fn), a glycoprotein from the extracellular matrix, and anandamide (AEA), one of the major members of the endocannabinoid family, are present in the bovine oviductal fluid and regulate bull sperm function. Also, AEA induces bovine sperm capacitation, through CB1 and TRPV1 receptors. In this work, we investigated if Fn induces bovine sperm capacitation thought the activation of the endocannabinoid system in this process. We incubated sperm with Fn (100 µg/ml) and/or capsazepine, a TRPV1 antagonist (0.1 µM) and some events related to sperm capacitation such as LPC-induced acrosome reaction, sperm-release from the oviduct, induction of PKA phosphorylated substrates (pPKAs) and protein tyrosine phosphorylation (pY) and nitric oxide (NO) production were assessed. Also, we studied the activity of fatty acid amide hydrolase (FAAH), the enzyme that degrades AEA. We found that Fn, via α5ß1 integrin, induced capacitation-associated events. Also, Fn stimulated signaling pathways associated to capacitation as cAMP/PKA and NO/NO synthase. Moreover, Fn decreased the FAAH activity and this correlated with sperm capacitation. Capsazepine reversed fibronectin-induced capacitation, and pPKAs and NO levels. The incubation of spermatozoa with R-methanandamide (1.4 nM), a stable analogue of AEA, increased cAMP and pPKAs levels. The presence of H89 (50 µM) or KT5720 (100 nM) (PKA inhibitors) prevented AEA-induced capacitation. In addition, R-methanandamide and capsaicin (0.01 µM), a TRPV1 agonist, increased NO production via the PKA pathway. These results indicate that Fn, through α5ß1, supports capacitation in bovine spermatozoa. This effect is dependent on the activation of TRPV1 through cAMP/PKA and NO signaling pathways. We propose that Fn could be considered as a new agent that promotes sperm capacitation in bull sperm. Our findings contribute to better understand the significance of Fn signaling in the capacitating events that lead to successful fertilization and embryo development in mammals including humans.

Anandamide regulates lipopolysaccharide-induced nitric oxide synthesis and tissue damage in the murine uterus.

In women, the association between chronic marijuana smoking and early miscarriage has long been known. Anandamide, a major endocannabinoid, mimics some of the psychotropic, hypnotic and analgesic effects of Delta(9)-tetrahydrocannabinol, the psychoactive component of marijuana. The uterus contains the highest concentrations of anandamide yet discovered in mammalian tissues and this suggests that it might play a role in reproduction. The production of small amounts of nitric oxide (NO) regulates various physiological events including implantation and myometrial relaxation, but in an inflammatory setting such as sepsis, NO has toxic effects as it is a free radical. The results presented in this study indicate that anandamide modulates NO production induced by lipopolysaccharide (LPS) in an in-vitro murine model. It was shown that LPS-induced NO synthesis and tissue damage were mediated by anandamide, as a cannabinoid receptor type I antagonist could block the effect of LPS (P < 0.001). This endotoxin inhibited anandamide uterine degradation (P < 0.05) and increased the expression of one of its synthesizing enzymes (P < 0.05). Contrary to the known anti-inflammatory and protective effects, in this model anandamide seems to act as a pro-inflammatory molecule modulating the production of NO induced by LPS. This proinflammatory effect of anandamide may be implicated in pathological reproductive events such as septic abortion.

Influence of Helicobacter pylori infection on the expression of MLH1 and MGMT in patients with chronic gastritis and gastric cancer.

The aim of the present study was to evaluate the influence of Helicobacter pylori on MLH1 and MGMT mRNA levels in patients with chronic gastritis and gastric cancer. The study included 217 patients, of which 26 were uninfected, 127 had chronic gastritis and were H. pylori-positive, and 64 had gastric cancer. Bacterial genotypes were evaluated by polymerase chain reaction (PCR), and the expression levels of MLH1 and MGMT were determined by quantitative real-time PCR and immunohistochemistry. There was an association between infection with cagA, vacA s1m1 strains and gastric cancer development. When the gastric epithelium and associated inflammation were examined for expression of MLH1 and MGMT, an overall increase in expression was observed. On the other hand, these levels decrease significantly among gastric cancer patients. The loss of MLH1 and MGMT expression in gastric cancer patients suggests that it is not an early event in H. pylori-associated gastric carcinogenesis.

Dual effect of anandamide on rat placenta nitric oxide synthesis.

Anandamide (AEA) has been reported to have pleiotropic effects on reproduction, but the mechanism by which it exerts these effects is unclear. The aim of this study is to characterize rat placental endocannabinoid system and to analyze the possible functional role of AEA in the regulation of NO levels in rat placenta during pregnancy. We found that cannabinoids receptors (CB1 and CB2), FAAH and TRPV1 were expressed in chorio-allantoic placenta. NOS activity peaked at day 13 and decreased with progression of pregnancy. Both exogenous and endogenous AEA significantly decreased NOS activity. Although pre-incubation with AM251 (CB1 antagonist) or AM630 (CB2 antagonist) had no effect, co-incubation with both antagonists induced NOS activity. Furthermore, pre-incubation with exogenous AEA and both antagonists resulted in the induction of placental NOS activity and this effect was reverted with capsazepine (selective TRPV1 antagonist). Additionally, the enhanced NO synthesis caused by capsaicin was abrogated by co-treatment with capsazepine, illustrating that NOS activity could be modulated by TRPV1. Finally, the inhibition of TRPV1 receptor by capsazepine caused a significant fall in NOS activity. These data support the concept that AEA modulates NO levels by two independent pathways: (1) diminishing the NOS activity via CBs; and (2) stimulating NO synthesis via TRPV1. We hypothesized that AEA have an important implication in the normal function of placental tissues.

Artificial sweetener saccharin disrupts intestinal epithelial cells' barrier function in vitro.

SCOPE: Consumption of non-nutritive sweeteners (NNS) is a dietary practice used by those who wish to lose weight or by patients on a sugar-restricted diet such as those with DM2. Although these substances are safe, possible biological interactions with the digestive tract, particularly in relation to intestinal permeability, have not been studied. Thus, the current work sought to investigate the action of different NNS on intestinal permeability using an in vitro Caco-2 cell model. METHODS AND RESULTS: Caco-2 cells were incubated with acesulfame K, aspartame, saccharin, or sucralose at equimolar concentrations. Acesulfame K, aspartame, and sucralose did not disrupt monolayer integrity in the cells. However, saccharin increased paracellular permeability and decreased transepithelial electrical resistance (TEER) via a non-cytotoxic mechanism. The levels of the tight junction protein claudin-1 were reduced in Caco-2 cells that had previously been exposed to saccharin. The inhibition of nuclear factor-κB (NF-κB) was able to prevent the reduction in TEER induced by saccharin treatment. Thalidomide, as an inhibitor of ubiquitin ligase, was able to prevent the decrease in claudin-1 protein expression and the TEER reduction in Caco-2 cells. CONCLUSIONS: Saccharin disrupts monolayer integrity and alters paracellular permeability in a Caco-2 cell monolayer model, via a mechanism involving NF-κB activation, resulting in the ubiquitination of the tight junction protein claudin-1. Saccharin consumption may potentially alter the intestinal integrity in humans.

Effect of Helicobacter pylori infection and acid blockade by lansoprazole on clarithromycin bioavailability.

The effect of proton pump inhibitors and Helicobacter pylori infection on the bioavailability of antibiotics is poorly understood. We determined the effects of 5-day oral administration of 60 mg lansoprazole on the bioavailability of clarithromycin in individuals with and without H. pylori infection. Thirteen H. pylori-infected and 10 non-infected healthy volunteers were enrolled in a study with an open-randomized two-period crossover design and a 21-day washout period between phases. Plasma concentrations of clarithromycin in subjects with and without lansoprazole pre-treatment were measured by liquid chromatography coupled to a tandem mass spectrometer. Clarithromycin Cmax and AUC0-10 h were significantly reduced after lansoprazole administration. In addition, lansoprazole treatment of the H. pylori-positive group resulted in a statistically significant greater reduction in Cmax (40 vs 15%) and AUC0-10 h (30 vs 10%) compared to lansoprazole-treated H. pylori-negative subjects. Thus, treatment with lansoprazole for 5 days reduced bioavailability of clarithromycin, irrespective of H. pylori status. This reduction, however, was even more pronounced in H. pylori-infected individuals.

Molecular basis of pyruvate kinase deficiency among Tunisians: description of new mutations affecting coding and noncoding regions in the PKLR gene.

INTRODUCTION: Pyruvate kinase (PK) deficiency is one of the most common hereditary nonspherocytic hemolytic anemias worldwide with clinical manifestations ranging from mild to severe hemolysis. However, investigation of this enzymopathy is lacking in Tunisia. We report here a pioneer investigation of PK deficiency among Tunisian cases referred to our laboratory for biological analysis of unknown cause of hemolytic anemia. METHODS: Two hundred and fifty-three patients with unknown cause of hemolytic anemia have been addressed to our laboratory in order to investigate for red blood cells genetic disorders. Red cell enzyme activities were measured by standard methods, and molecular analysis was performed by DNA sequencing. The interpretation of mutation effect and the molecular modeling were performed by using specific software. RESULTS: Six different PKLR mutations were found (c.966-1G>T; c.965+1G>A; c.721G>T; c.1163C>A; c.1456C>T; c.1537T>A), among which four are described for the first time. Genotype-phenotype correlations for the novel missense mutations were investigated by three-dimensional structure analysis. CONCLUSION: This study provides important data of PK deficiency among Tunisians. It might be followed by a large neonatal screening to determine the spectrum of PK mutations and identify potential deficient patients for an early medical follow-up.

17-beta-Estradiol upregulates COX-2 in the rat oviduct.

We investigated the regulation of cyclooxygenase-2 (COX-2) by 17-beta-estradiol (E2) in the rat oviduct. We observed that COX-2 is expressed mainly in proestrous and estrous stages, periods under estrogenic influence. While exogenous administration of E2 (1 microg/rat) significantly increased COX-2 protein levels, progesterone did not modify it. COX-2 was mainly localized on oviductal epithelial cells from estrogenized rat. Induction of COX-2 expression by E2 was partially reverted by tamoxifen (1 mg/rat), an E2 receptor antagonist. Estradiol treatment also increased prostaglandins (PGs) synthesis: 6-keto-PGF(1alpha) (40%), a stable metabolite of prostacyclin (PGI2), PGF(2alpha) (40%) and PGE2 (50%). Tamoxifen completely suppressed this enhancement. In order to discriminate which isoform of COX was implicated in the stimulatory effect of E2 on PGs synthesis, oviducts were preincubated with meloxicam (Melo: 10(-9)M) or NS-398 (10(-7)M), two selective COX-2 inhibitors. Both Melo and NS-398 abolished the increase of PGs synthesis stimulated by E2. All together, these data indicate that E2 could upregulate COX-2 expression and activity in the rat oviduct and that the stimulatory effect of E2 may be receptor-mediated.

Nitric oxide (NO) inhibits prostaglandin E2 9-ketoreductase (9-KPR) activity in human fetal membranes.

Nitric oxide (NO) synthesized by fetal membranes may act either directly inhibiting myometrium contractility or indirectly interacting with tocolytic agents as prostaglandins (PGs). Here we examined if NO could modulate prostaglandin E(2) 9-ketoreductase (9-KPR) activity in human fetal membranes (HFM). 9-KPR is the enzyme that converts PGE(2) into PGF(2alpha), the main PGs known to induce uterine contractility at term. Chorioamnion explants obtained from elective caesareans were incubated with aminoguanidine (AG), an iNOS inhibitor, or NOC-18, a NO donor. NOC-18 (2mM) increased PGE(2) production and diminished PGF(2alpha) synthesis in HFM. AG presented the opposite effect. When we evaluated the activity of 9-KPR by the conversion of [(3)H]-PGE(2) into [(3)H]-PGF(2alpha) and 13,14-dihidro-15-keto prostaglandin F(2alpha) (the PGF(2alpha) metabolite), we found that NOC-18 inhibited 9-KPR activity. Interestingly, AG did not elicit any effect on 9-KPR but l-NAME, a non-selective NOS inhibitor, significantly increased its activity. Our data suggests that exogenous NO inhibits 9-KPR activity in HFM, thus modulating the synthesis of important labor mediators as PGF(2alpha).

Detection of new pathogenic mutations in patients with congenital haemolytic anaemia using next-generation sequencing.

INTRODUCTION: Congenital haemolytic anaemia (CHA) refers to a group of genetically heterogeneous disorders, mainly caused by changes in genes encoding globin chains, cytoskeletal proteins and red cell enzymes, in which accurate diagnosis can be challenging with conventional techniques. METHODS: To set-up a comprehensive assay for detecting mutations that could improve aetiological diagnosis, we designed a custom panel for sequencing coding regions from 40 genes known to be involved in the pathogenesis of CHA, using the Ion Torrent™ (Thermo Fisher Scientific, S.L. Waltham, MA, USA) Personal Genome Machine (PGM) Sequencer. A control group of 16 samples with previously known mutations and a test group of 10 patients with unknown mutations were included for assay validation and application, respectively. RESULTS: In the test group, we identified pathogenic mutations in all cases: four patients had novel mutations in genes related to membrane defects (SPTB, ANK1, SLC4A1 and EPB41), four were homozygous or compound heterozygous for mutations in genes related to enzyme deficiencies (GPI, TPI1 and GSS), one had a mutation in the HBB gene and another presented a homozygous mutation in the ADAMTS13 gene. CONCLUSIONS: Ion PGM sequencing with our custom panel is a highly efficient way to detect mutations causing haemolytic anaemia, including new variations. It is a high-throughput detection method that is ready for application in clinical laboratories.

Effect of in vivo administration of epidermal growth factor on prostaglandin production and NOS activity in term rat placentae. Possible participation of placental EGF receptors.

Many authors hypothesize that the epidermal growth factor (EGF) is involved in the onset of labor. Previous reports from our laboratory showed that intrauterine administration of EGF delays the beginning of labor. The aims of this study were: 1) to analyze the effect of intrauterine administration of 500 ng EGF on placental prostaglandins and nitric oxide, and 2) to characterize the expression of EGF receptors (EGF-R) in pregnant rat placentae. Saline solution (sham group) and 500 ng EGF (EGF-treated group) were administered via intrauterine injection on day 21 of gestation, and both groups of animals were sacrificed on day 22 (sham rats delivered on day 22). Results showed that EGF treatment: 1) inhibited the production of prostaglandin E (p<0.001) and F(2alpha) (p<0.01), 2) increased the synthesis of nitric oxide (p<0.001), and 3) reduced the expression of cyclooxygenase-II, the enzyme responsible for PG synthesis. Placentae were found to express EGF-R and its activated form, and the expressions of both forms were higher at mid and term pregnancy. Hence, EGF is a very interesting molecule for studying the regulation of placental prostaglandin and nitric oxide production related to the parturition process.

New once-daily, highly effective rescue triple therapy after multiple Helicobacter pylori treatment failures: a pilot study.

BACKGROUND: Helicobacter pylori treatment failure is a growing problem in daily practice. AIM: To determine the efficacy of the combination of rabeprazole, levofloxacin and furazolidone as a rescue therapy. METHODS: Duodenal ulcer patients previously submitted, without success, to at least two H. pylori treatment regimens were included. Gastroscopy (urease test, histological examination and culture) and (13)C-urea breath test were performed. All patients received a combination of rabeprazole 20 mg, levofloxacin 500 mg and furazolidone 200 mg (two tablets) administered in a single dose in the morning for 10 days. Clinical examination and a new (13)C-urea breath test were performed 90 days after therapy. RESULTS: Twelve patients (eight females and four males), mean age 43 (30-58) years were included. Two patients failed to complete the treatment because of nausea and vomiting. Ten patients completed the study and took all the medications as advised. Culture was obtained in six patients: 100 and 83% of the samples were sensitive to furazolidone and levofloxacin, respectively. Per-protocol and intention-to-treat eradication rates were 100 and 83% (P = 0.019). CONCLUSIONS: the combination of rabeprazole, levofloxacin and furazolidone in a single daily dose for 10 days constitutes a highly-effective and low-cost alternative as a third-line therapy in patients infected with H. pylori.

Plasma hydroxy-metronidazole/metronidazole ratio in hepatitis C virus-induced liver disease.

It has been suggested that the measurement of metronidazole clearance is a sensitive method for evaluating liver function. The aim of this study was to evaluate the usefulness of plasma hydroxy-metronidazole/metronidazole ratios as indicators of dynamic liver function to detect changes resulting from the various forms of chronic hepatitis C virus (HCV) infection. A total of 139 individuals were studied: 14 healthy volunteers, 22 healthy, asymptomatic, consecutive anti-HCV-positive HCV-RNA negative subjects, 81 patients with chronic hepatitis C (49 with moderate/severe chronic hepatitis and 34 with mild hepatitis), and 20 patients with cirrhosis of the liver. HCV status was determined by the polymerase chain reaction. Plasma concentrations of metronidazole and its hydroxy-metabolite were measured by reverse-phase high-performance liquid chromatography with ultraviolet detection in a blood sample collected 10 min after the end of a metronidazole infusion. Anti-HCV-positive HCV-RNA-negative individuals demonstrated a significantly reduced capacity to metabolize intravenously infused metronidazole compared to healthy individuals (0.0478 +/- 0.0044 vs 0.0742 +/- 0.0232). Liver cirrhosis patients also had a reduced plasma hydroxy-metronidazole/metronidazole ratio when compared to the other groups of anti-HCV-positive individuals (0.0300 +/- 0.0032 vs 0.0438 +/- 0.0027 (moderate/severe chronic hepatitis) vs 0.0455 +/- 0.0026 (mild chronic hepatitis) and vs 0.0478 +/- 0.0044 (anti-HCV-positive, HCV-RNA-negative individuals)). These results suggest an impairment of the metronidazole metabolizing system induced by HCV infection that lasts after viral clearance. In those patients with chronic hepatitis C, this impairment is paralleled by progression of the disease to liver cirrhosis.