A ten-color tube with dried antibody reagents for the screening of hematological malignancies.

INTRODUCTION: The workflow in clinical flow cytometry laboratories must constantly be reviewed to develop technical procedures that improve quality and productivity and reduce costs. Using the Beckman Coulter dry coating technology, we customized a ten-color tube with dried antibody reagents, designated the Duraclone screening tube (DST), for screening hematological malignancies. Here, we compared the applicability, clinical and numerical equivalence, and cost and time required for the technical procedures between the liquid reagents and the DST. METHODS: The DST contains CD4 + Kappa-FITC, CD8 + Lambda-PE, CD3 + CD14-ECD, CD33-PE-Cy5.5, CD20 + CD56-PE-Cy7, CD34-APC, CD19-APC-AlexaFluor700, CD10-APC-AlexaFluor750, CD5-Pacific Blue, and CD45-Krome Orange. We evaluated 20 bone marrow samples, 13 peripheral blood samples, 6 lymph node biopsy samples, 5 fine-needle aspirate samples, 5 cerebrospinal fluid samples, and 1 pleural fluid sample. RESULTS: The DST was useful for more than 60% of our samples. It was able to enumerate the majority of the populations in all types of samples with a statistically acceptable correlation with the liquid reagents. The use of the DST translated into significant time and cost savings of 15.8% and 12.3%, respectively, compared with the use of the liquid reagent. The cost was reduced by $14.36 per sample. CONCLUSIONS: The DST is an efficient solution for screening hematological malignancies with improved quality, productivity, standardization, and sustainability. These improvements could benefit patients by providing faster diagnoses using a higher quality and lower cost reagent.

[Fetal bradycardia: a retrospective study in 9 Spanish centers]. / Bradicardia fetal: estudio multicéntrico retrospectivo en 9 hospitales españoles.

OBJECTIVE: The aim of this study is to review the current management and outcomes of fetal bradycardia in 9 Spanish centers. METHODS: Retrospective multicenter study: analysis of all fetuses with bradycardia diagnosed between January 2008 and September 2010. Underlying mechanisms of fetal bradyarrhythmias were studied with echocardiography. RESULTS: A total of 37 cases were registered: 3 sinus bradycardia, 15 blocked atrial bigeminy, and 19 high grade atrioventricular blocks. Sinus bradycardia: 3 cases (100%) were associated with serious diseases. Blocked atrial bigeminy had an excellent outcome, except for one case with post-natal tachyarrhythmia. Of the atrioventricular blocks, 16% were related to congenital heart defects with isomerism, 63% related to the presence of maternal SSA/Ro antibodies, and 21% had unclear etiology. Overall mortality was 20% (37%, if terminations of pregnancy are taken into account). Risk factors for mortality were congenital heart disease, hydrops and/or ventricular dysfunction. Management strategies differed among centers. Steroids were administrated in 73% of immune-mediated atrioventricular blocks, including the only immune-mediated IInd grade block. More than half (58%) of atrioventricular blocks had a pacemaker implanted in a follow-up of 18 months. CONCLUSIONS: Sustained fetal bradycardia requires a comprehensive study in all cases, including those with sinus bradycardia. Blocked atrial bigeminy has a good prognosis, but tachyarrhythmias may develop. Heart block has significant mortality and morbidity rates, and its management is still highly controversial.

Degradabilidade in situ da matéria seca de três forrageiras tropicais nas formas in natura e ensilada / In situ dry matter degradability of three tropical forages of green chopped and ensiled forms

Estimou-se a degradabilidade ruminal da matéria seca (MS) do milho (Zea mays L.), milheto (Pennisetum glaucum (L.) R. Br.) e sorgo (Sorghum bicolor L. Moench.) utilizando-se a técnica in situ com amostras nas formas in natura e ensiladas. Amostras de 6g de cada alimento foram incubadas em triplicata no rúmen por seis, 24 e 96 horas de duas vacas secas. Estimou-se o tempo zero (t0) lavando-se os sacos em água, e foi utilizado para o cálculo da solubilidade. O delineamento experimental foi de parcelas subdivididas, sendo as forrageiras os tratamentos, e os tempos de incubação os subtratamentos. Compararam-se as médias do desaparecimento da MS por meio do teste SNK, a 5 por cento de probabilidade. Os resultados encontrados do desaparecimento da MS ( por cento) das forrageiras in natura e ensiladas nos tempos zero, seis, 24 e 96 foram respectivamente: milheto (10,07, 14,50, 20,36, 47,86; 11,64, 15,69, 21,60, 33,37), milho (12,64, 20,08, 31,77, 68,11; 13,31, 20,97, 35,31, 67,33) e sorgo (10,20, 21,55, 26,56, 58,95; 10,07, 15,10, 24,89, 44,52). A degradação potencial ( por cento) das silagens foi: milheto (36,44), milho (81,18) e sorgo (51,30).

Dry matter (DM) ruminal degradability of corn (Zea mays L.), millet (Pennisetum glaucum (L. R. Br.) and sorghum (Sorghum bicolor L. Moench.) was evaluated using in situ technique with samples in green chopped and ensiled forms. Two crossbred fistulated (live weight of 480kg) dry cows participated. Samples of six grams in each forage were incubated in the rumen for 6, 24 and 96 hours. We estimated time zero (t0) washing the bags in water and it was used to calculate solubility. The experimental design followed a randomized block design with a split plot. We compared the average of DM through the SNK test at 5 percent probability. The results of disappearance ( percent) of dry matter forages of green chopped and ensiled forms in 0, 6, 24 and 96 hours were respectively: millet (10,07, 14,50, 20,36, 47,86; 11,64, 15,69, 21,60, 33,37), corn (12,64, 20,08, 31,77, 68,11; 13,31, 20,97, 35,31, 67,33) and sorghum (10,20, 21,55, 26,56, 58,95; 10,07, 15,10, 24,89, 44,52). The potential degradation ( percent) of silages was: millet (36,44), corn (81,18) and sorghum (51,30).

Lung transplantation for patients older than 65 years: is it a feasible option?

BACKGROUND: Advanced age has been a relative contraindication to lung transplantation. However, the exact age limit for this procedure has not yet been established. The aim of this work is to present our experience with this particular group. METHODS: This retrospective review included medical charts of patients who underwent lung transplantation at our institution from January 2004 to February 2009: namely, 112 cadaveric lung transplants with 12 patients (10.7%) >65 years old. RESULTS: There were 9 male patients and the overall mean age was 68 years (range 66-72). The indications were pulmonary fibrosis in 8 and emphysema in 4 cases. Four patients had mild coronary artery disease and 4 systemic hypertension. All of the procedures were unilateral and only 2 required extracorporeal circulation. Only 5 patients received blood product transfusions intraoperatively; the mean ischemic time was 222 minutes. Four patients developed primary graft dysfunction, the mean requirement for mechanical ventilation was 30 hours, and the mean intensive care unit stay, 11 days. Postoperative complications were respiratory infections (n = 8), catheter-related infection (n = 1), atrial fibrillation (n = 2). The mean hospital stay was 28 days and the 1-year survival was 75%. CONCLUSION: Lung transplantation is a feasible option for well-selected patients with end-stage pulmonary disease who are >65 years old. Our study reinforces the modern trend for unilateral procedures in this situation.

Hyperacute rejection after single lung transplantation: a case report.

Hyperacute rejection is a well-known complication in kidney and heart transplantations. However, its occurrence in lung transplantation is extremely rare, with only 4 cases previously described. A 53-year-old female patient blood type O with end-stage chronic obstructive pulmonary disease underwent left lung transplantation. She had 2 negative pretransplantation evaluations for panel-reactive antibodies. One hour after the vascular clamps were released, progressive hypoxia developed. Fiberoptic bronchoscopy revealed an optimal bronchial anastomosis; an abundant pink frothy fluid was observed on the allograft side. Chest X ray sevealed a completely opacified left lung. Due to the low-compliance of the transplanted lung and the risk for native lung hyperinsufflation, independent mechanical ventilation was employed. Despite all measures, multiple organ failure developed and the patient died 24 hours after the procedure. A necropsy evaluation for confirmed the patency of all anastomoses and no signs of ischemia. Retrospectively, a new evaluation for panel-reactive antibodies was performed, with 24% reactivity. Complement-dependent cytotoxicity crossmatch was negative, however, a flow cytometric analysis was positive for both HLA-I (56%) and HLA-II (45%). Further investigation detected an anti-A2 in the recipient serum and the donor had an A2 antigen. Hyperacute rejection is a rare posttransplantation complication highlighted by its precocity and lethality. With the increased number of lung transplantations performed yearly, it is believed that its incidence will also rise. Therefore, prompt diagnosis and familiarity with management strategies are fundamental.

Optimization of the (32)P-Postlabeling/Thin Layer Chromatography Assay ((32)P-TLC) for In Vitro Detection of 8-Oxo-Deoxyguanosine as a Biomarker of Oxidative DNA Damage.

During the last years, much attention was focused on the measurement of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG) as a marker of oxidative DNA damage. Among various analytical techniques, the (32)P-postlabeling assay has been applied in determination of 8-oxo-dG. However, artefactual DNA oxidation could take place during the work-up procedures leading to over-estimate the level of 8-oxo-dG. In the present study, we optimized the (32)P-postlabelling assay with thin layer chromatography to measure 8-oxo-dG in standard samples of 8-oxo-dG, calf thymus DNA and primary cultured rat hepatocytes. The background levels of 8-oxo-dG in calf thymus DNA and in primary cultured rat hepatocytes were lesser than those determined by the previously described (32)P-postlabeling procedures and were in the range of those determined by chromatography methods (GC-MS, HPLC-MS).

Seasonal variations of DNA-adduct patterns in open field farmers handling pesticides.

Seasonal variations of DNA-adduct levels in peripheral blood cells were evaluated in open field farmers (n=26) by use of the 32P-postlabelling assay. Samples were collected before (sample S0) and during (sample S4) the period of intensive pesticide use. A similar sampling procedure was applied to a referent group (n=29). Exposure to pesticides was estimated via a detailed questionnaire. For the group of farmers, an increase in mean adduct level was observed during the season (mean RALS0=3.9+/-3.4 x 10(-10), mean RALS4=13.3+/-15.7 x 10(-10); p=0.008; RAL=relative adduct level). The mean adduct levels were significantly different between farmers and referents only in the S4 samples, with higher levels for farmers (p=0.02). The number of different adducts per individual was higher for farmers at S4 when compared with S0 (p=0.02) and compared with the referents at S4 (p=0.03). However, the increase of the adduct level in farmers did not seem to be attributable to the occurrence of specific new adducts in S4 as compared with S0, but was supposedly due to intensification of pre-existing spots and/or appearance of new unspecific ones. This would be in agreement with indirect genotoxic (epigenetic) effects known for several pesticides, even though a direct mechanism cannot be ruled out definitively. The implication of the pesticides used by the farmers in the modulation of DNA-adduct patterns was explored by analysis of exposure data obtained from the questionnaire.

The "bubble point" for validation of drug release or simulated absorption tests for ointments.

The aim of the present study was to design a test to ascertain the behaviour and reliability of a membrane used in drug release and simulated absorption tests in order to arrive at useful indications for simulating topical as well as gastro-intestinal absorption. The membrane can be used in two different conditions: a) as a simple porous membrane placed between the ointment and an accepting liquid phase, generally water phase; b) as a membrane soaked in a lipophilic liquid phase to simulate the horny layer between the ointment and accepting water phase. In this study the "bubble point test" was used to test the integrity of the soaking film as well as the membrane, during and after drug release and simulated absorption tests with different types of ointment. In the case of a drug release test from an ointment, the bubble point test may determine the test conditions, that is the ointment applied to either a dry or hydrated membrane. Only the use of a previously hydrated membrane can guarantee constant conditions in the in vitro model. Use of a dry membrane may lead to infiltration of liquid components of the ointment base, thus altering the contact conditions between the two phases of the cutaneous compartment model (lipogel and W/O creams). The use of a hydrated membrane may also lead to interactions between the two phases of the compartment, with osmotic exchanges between the acceptor phase and ointment sample (hydrogel, PEG gel, O/W creams). The hydrated membrane is therefore reliable only for comparison between lipophilic base ointments. In a simulated absorption test, determination of the bubble point makes it possible to ascertain the physical integrity of the lipoid liquid film immobilized by capillary action in the inner microporous structure of the membrane during the test. This condition is essential to maintain a balance between the parameters regulating the diffusion process between the different compartments of the system. The use of a lipoid-soaked membrane makes it possible to avoid interactions between the ointment sample and an aqueous acceptor phase, such as hydrosoluble bases. Since the diffusion across a lipoid film immobilised within a porous membrane depends on the drug release rate from the ointment base, the test allows a contextual evaluation of the release kinetics as well as an indication of the drug absorption possibilities through an in vitro model of the cutaneous compartment.

Relationship between tumour growth rate and carbogen-based functional MRI for a chemically induced HCC in mice.

We previously performed MRI studies of HCC (hepatocellular carcinomas) in mice showing the feasibility of measuring a carbogen effect. In the present study carbogen response of the whole tumour was compared with growth characteristics during longitudinal follow-up. HCC were chemically induced. The imaging protocol at 4.7 T comprised a fast spin-echo sequence for high-resolution screening and measurement of growth curves, and a fast gradient echo sequence allowing an entire T2*w image acquisition per respiratory cycle to perform fMRI under carbogen breathing. A new parameter, T+, the fraction of tumour voxels with increased intensity under carbogen was measured on manually defined ROIs. Twenty-two HCC were followed for 3-10 weeks. Tumours were divided into two groups, "regularly" and "irregularly" growing tumours. A linear correlation between T+ and tumour growth rate was observed only for "regularly" growing HCC. These results suggest a link between tumour growth rates and tumour fractions exhibiting signal increase upon carbogen breathing. They are compatible with observations by others that rapidly growing tumours are more hypoxic than slowly growing ones. Combined measurement of T+ and tumour growth may become a useful noninvasive follow-up approach for assessment and/or management of therapies involving vasculature-targeting and anti-proliferative drugs.

Induction of DNA synthesis in mouse liver following increases of DNA adduct levels elicited by very low cumulative doses of the genotoxic hepatocarcinogen 7H-dibenzo[c,g]carbazole.

The purpose of this work was to investigate the administration of very low but repeated doses of a genotoxic carcinogen and an eventual correlation with cellular DNA synthesis. The compound 7H-dibenzo[c,g]carbazole is a genotoxic carcinogen in the mouse liver and was administered topically at the dose of 13.35 microg per animal every 2 days to give a total of 13 applications. Animals were sacrificed 48 hours after every 2 applications until the 10th treatment, then 48 hours after every treatment. Postulated genotoxic effects such as DNA adduct formation were detected by the 32P-post labeling assay. Liver sections were examined for microscopic changes and DNA synthesis. Results showed an increase of the total DNA adduct level in the liver throughout the study with a slowing down in the level after the sixth application of the compound. This change could correspond to the onset of DNA synthesis and to the moderate hepatocellular apoptosis which was observed. The DNA synthesis, which was considered to be secondary to the cytotoxicity induced by the high level of DNA adducts altering normal cellular activity, could also be the opportunity to fix the DNA adducts into heritable mutations. These results raise the question regarding the risk assessment in humans exposed regularly to very low doses of chemicals in the environment: for non-proliferating tissue, the regular accumulation of DNA adducts could remain silent until a "threshold level" is reached from which stimulation of the DNA synthesis may fix the DNA adducts into heritable mutations, eventually leading to tumors.

Tissue-specific activities of methylated dibenzo[c,g]carbazoles in mice: carcinogenicity, DNA adduct formation, and CYP1A induction in liver and skin.

The potent multitissue carcinogen 7H-dibenzo[c,g]carbazole and nine methylated derivatives, synthesized on the basis of the positions in the parent compound that are involved in metabolism, were tested for their ability to induce sarcomas and hepatic tumors in XVIInc/Z mice. In addition, the capacity of these compounds to induce DNA adducts in skin and liver was investigated by (32)P-postlabeling analysis after their topical administration. Induction by these compounds of cytochromes P450 of the 1A family in liver and skin was investigated and correlated to their carcinogenic potential. A clear correlation was found between the tissue specificity of DNA binding and the capacity of each compound to induce skin or liver tumors. In contrast, no direct relationship was observed between the capacity of the compounds to induce cytochromes 1A1/1A2 and the tissue specificity of carcinogenesis or DNA binding in liver or skin. The results are discussed with respect to the positions of methyl groups in the 7H-dibenzo[c,g]carbazole molecule.

Ultrasonic assessment of variations in thickness of subcutaneous fat during the normal menstrual cycle.

OBJECTIVE: To verify the occurrence of natural variations in thigh and abdominal subcutaneous fat thickness related to the phases of the menstrual cycle, to assess the value of ultrasonography as a reliable method for monitoring subcutaneous fat thickness changes and to evaluate their amplitudes. METHODS: This study included 10 women (19-39 years) who menstruated regularly. None had used oral contraceptives or slimming products during the 3 months prior to the study. At cycle day 2 (CD2), CD6, CD14, CD22, CD27 and CD30 days (CD0: beginning of menstruation), the subjects were submitted to: (1) measurement of weight and thigh perimeters, (2) measurements of thigh and abdomen subcutaneous fatty tissue thickness on B-mode images acquired at 10 MHz. A protocol was designed to guarantee a reproducible repositioning during the whole time course of the study and ultrasound examinations (US) were always performed by the same trained person to avoid inter-examiner variability. RESULTS: Subcutaneous fat thicknesses decreased during the first half of the cycle and reached their lowest values at day 22 (-2.0% for the thighs; -3.3% for the abdominal region). Both thigh and abdomen subcutaneous fat reached their maximum thicknesses during menstruation with respective increases of +2.2 and +4.0%. The observed cyclic amplitude variations in the subcutaneous adipose tissue thickness accounted for 7.3% for the abdominal region and 4.1% for the thighs. CONCLUSION: Variations in adipose tissue thickness during the menstrual cycle could be quantified and monitored by US. The thickness of the thigh and abdominal hypodermis was more important during menstruation and decreased in mid-cycle with a minimum occurring 1 week after ovulation.

Kinetics of DNA adduct formation and removal in mouse hepatocytes following in vivo exposure to 5,9-dimethyldibenzo[c,g]carbazole.

5,9-Dimethyldibenzo[c,g]carbazole (DMDBC), a potent mouse hepatocarcinogen, has been shown to induce a non-linear increase in mutant frequency in the liver of the transgenic MutaMouse. To gain insight into the mechanisms underlying the mutagenicity of DMDBC in vivo, DNA damage formation and removal were monitored in mouse hepatocytes over 4-144 h after a single skin application of 10 or 90 mg/kg DMDBC. DNA adducts were measured by (32)P-post-labeling. DNA repair was assessed by: (i) the unscheduled DNA synthesis (UDS) assay, which measures [(3)H]thymidine incorporation into hepatocyte DNA undergoing excision repair; (ii) the Comet assay, which detects DNA strand breaks transiently produced between the incision and rejoining steps of the excision repair process. A plateau of approximately 400 DNA adducts/10(8) nucleotides was reached 24 h after treatment with 10 mg/kg and remained unchanged until 144 h. UDS activity was significantly induced at 15 and 24 h, while no DNA strand breaks were observed at any sampling time. These results suggest that DNA repair mechanisms were efficiently induced and the formation of a high degree of DNA damage was avoided at this dose level. Following exposure to 90 mg/kg DMDBC, the number of DNA adducts increased sharply to a maximum at 24 h ( approximately 8000/10(8) nucleotides) and then declined to approximately 500/10(8) nucleotides at 144 h. UDS activity was markedly induced from 15 to 72 h. Low levels of DNA strand breaks were observed at 24 and 48 h. The formation of large numbers of DNA adducts and the emergence of DNA strand breaks despite a strong initial induction of UDS activity suggested that DNA repair mechanisms were saturated at this dose level. This phenomenon could partly account for the non-linear induction of gene mutations previously reported in the liver of the transgenic MutaMouse.

Assessment of skin improvement treatment efficacy using the photograding of mechanically-accentuated macrorelief of thigh skin.

The assessment of the efficacy of skin improvement treatments could be obtained by several different instrumental tests. In parallel to these techniques, a visual evaluation more closely related to the consumer's considerations would be of value for the assessment of slimming treatments efficacy. A method of macrorelief scoring of skin has been developed as an alternative to usual clinical assessment. It consisted of acquiring macroscopic views of the most representative areas of women's cellulite. Photographs were taken after application of a gripping system around the thigh, used to increase the orange peel look of the skin over a restricted area (200 cm(2)) and then classified by observers according to reference groups of cellulite intensity. This publication presents the validation of this photograding method and the results of the first 2-month double-blind placebo-controlled clinical study performed on 30 subjects with this technique associated with: (i) B mode ultrasound imaging in order to correlate scorings with instrumental measurements of thigh adipose tissue thickness variations and (ii) self-evaluation questionnaire aiming at evaluating the overall appraisal and attitude towards the two products. The results clearly showed that the effects of 'slimming products' on the skin's macrorelief improvement could be evaluated objectively using this photograding technique. Moreover, the method described here allows delayed scoring of cellulite intensity from a restricted examination area independent of whole-subject appearance, thus decreasing subjectivity.

Comparison of benzo[a]pyrene DNA adduct formation in the skin of two strains of mice selected for resistance (DBA/2) or susceptibility (C3H/HeN) to contact dermatitis.

Two strains of mice were selected for resistance (DBA/2) or susceptibility (C3H/HeN) to contact dermatitis. Benzo[a]pyrene-DNA adduct formations was compared in the two mouse strains by a postlabeling procedure to determine if there was a significant effect. Results showed that adduct profiles in DBA/2 and C3H/HeN dermis were qualitatively similar. The total binding levels were higher in DBA/2 mice on the d 2 and the d 10. DNA adduct formation has been shown to inversely correlate with skin allergy induction. Data suggest that the expression of the genes responsible for the differences in responsiveness to chemical induced contact dermatitis in mouse may play an important role in benzo[a]pyrene-DNA adduct formation.

Effect of mitogenic or regenerative cell proliferation on lacz mutant frequency in the liver of MutaTMMice treated with 5, 9-dimethyldibenzo[c,g]carbazole.

The purpose of this work was to investigate the impact of cell proliferation on liver mutagenesis. The genotoxic hepatocarcinogen 5, 9-dimethyldibenzo[c,g]carbazole (DMDBC) was administered to lacZ transgenic MutaTMMice at a non-hepatotoxic dose of 10 mg/kg, which induces only a slight increase in the liver lacZ mutant frequency (MF). To determine if cell proliferation stimuli enhanced DMDBC mutagenicity, MF was analyzed in mice first receiving DMDBC 10 mg/kg, then approximately 2 weeks later, either carbon tetrachloride (CCl4, a cytotoxic agent inducing regenerative cell proliferation) or phenobarbital (PB, a mitogenic agent inducing direct hyperplasia). In preliminary studies, the extent of cell proliferation induced by CCl4, PB and DMDBC was determined in non-transgenic CD2F1 mice by means of 5-bromodeoxyuridine labeling. The labeling index was significantly increased after CCl4 and PB, while no change was detected with DMDBC. MF was then determined in MutaTMMice 28 days after initial DMDBC treatment. No increase in MF was detected in mice receiving CCl4 or PB alone. A 2- to 3-fold increase in MF was detected in mice treated with 10 mg/kg DMDBC alone. In contrast, MF was markedly increased in mice receiving DMDBC followed by proliferative treatment (15-fold with CCl4 and 25-fold with PB). These results demonstrate that expression of DMDBC-induced mutations in mouse liver largely depends on the induction of cell proliferation (by a cytotoxic or mitogenic stimulus) and illustrate that MutaTMMouse is a valuable tool to investigate the early events of liver carcinogenesis.

Kinetics of induction of DNA adducts, cell proliferation and gene mutations in the liver of MutaMice treated with 5,9-dimethyldibenzo[c,g]carbazole.

5,9-Dimethyldibenzo[c,g]carbazole (DMDBC) is a synthetic derivative of the environmental pollutant 7H-dibenzo[c,g]carbazole. DMDBC is a potent genotoxic carcinogen specific for mouse liver. Using the MutaMouse lacZ transgenic mouse model and a positive selection assay, we measured lacZ mutant frequency (MF) in the liver 28 days after a single s.c. administration of DMDBC at 3, 10, 30, 90 or 180 mg/kg. MF remained low at 3 and 10 mg/kg, but increased markedly from 30 mg/kg onwards. To investigate the reason for this non-linear response, we examined mechanisms potentially involved in mutation induction in the liver. Genotoxic effects such as DNA adduct formation were detected in 32P-post-labelling studies. Liver sections were examined for microscopic changes and cell proliferation. These parameters, and MF, were studied 2, 4, 7, 14, 21 and 28 days after a single s.c. administration of 10 or 90 mg/kg DMDBC. At 10 mg/kg, a dose found to double the MF on day 28, DNA adducts reached a level of 200-600 adducts per 10(8) nucleotides from day 4 to day 28. No changes in histology or cell proliferation were detected at this low dose. At 90 mg/kg, MF increased gradually from day 7 to day 28 (maximum 44-fold). The DNA adduct level ranged from 400 to 4500 adducts per 10(8) nucleotides on day 2, then stabilized at approximately 400 adducts per 10(8) nucleotides on day 4. An early cytotoxic effect was detected microscopically in centrilobular hepatocytes, and was followed by liver cell proliferation. These data suggest that the marked increase in MF in MutaMouse liver after treatment in vivo with DMDBC at 90 mg/kg may be explained by the induction of replicative DNA synthesis due to a cytotoxic effect, allowing the fixation of persistent DNA adducts into mutations.

Ultrasonic root-end preparation with smooth and diamond-coated tips.

The purpose of this study was to compare the effects of smooth and diamond-coated ultrasonic retrotips on the external and internal surfaces of root-end preparations with the aid of a scanning electron microscope (SEM). Forty-four mesial roots of human mandibular molars were selected. The canals were cleaned, shaped and obturated using gutta-percha and sealer. The apical portions were resected at a 45 degrees-angle bevel exposing both mesial canals and the isthmus area. The roots were then divided into two groups according to the type of root-end preparation: Group A--performed with smooth retrotips (S) and Group B--performed with diamond-coated retrotips (DC). The specimens were coded and prepared for SEM evaluation. Observations of the external surface preparation showed that the S and DC retrotips produced very well-centered cavities involving both canals and isthmus area with minimal deviations and no perforative defects. When the internal surface of the root-end preparations was evaluated, it was evident that the use of S retrotips resulted in clean canal walls with little superficial debris and smear layer. Internal canal surfaces done with DC retrotips were irregular showing patent grooves, in contrast with the more uniform, regular and smoother surfaces when S retrotips were employed.