A rapid and sensitive method for measuring monooxygenase activities in hepatocytes cultured in 96-well plates

Summary Measurement of biotransformation activities in cells is of great importance for drug metabolism and toxicologic studies. It is currently done by measuring the enzymatic activities in partially purified microsomes. In the present work we report on a rapid, easy, sensitive, and reproducible fluorimetric assay for quantifying cytochrome P450-dependent monooxygenase activities (P450IA1, P450IIB1) in hepatocytes cultured in 96-well plates. The procedure involves the direct determination of enzymatic activities in intact hepatocytes while avoiding cell homogenization, thereby permitting use of a the reduced number of cells and allowing cultured cells to be used in later experiments. Substrates (7-ethoxyresorufin, 7-pentoxyresorufin) are added to culture medium and metabolized by hepatocytes. After enzymatic deconjugation, the fluorescent resorufin present in culture medium is quantified by means of a microplate fluorimetric reader. Major advantages of this technique, as compared to other available methods, are: a) no cell disruption is required; b) activity can be measured with a very small number of cells; c) rapid processing time; and d) possibility of performing repeated assays with the same cell monolayer.     

Antiparasitic properties of homoallylamines and related compounds

A study of some antiparasitic properties of several homoallylamines and related tetrahydroquinolines and quinolines, previously described, was carried out using in vitro activity assays against the epimastigote form of Trypanosoma cruzi and against Trichomonas vaginalis. Unspecific cytotoxicity against murine macrophages was also studied. Although the antichagasic and trichomonacidal activities are not comparable to those of the standard drugs, nifurtimox and metronidazole, some of the compounds exhibit an interesting specific antiparasitic activity.     

Molecular biology in prostate cancer

Summary Genes involved in cancer generation are usually tumor suppressors and oncogenes. Progressive genetic alterations in these genes are involved in the mechanisms of tumorigenesis. In prostate cancer, additionally several chromosomal loci that should harbor mutated genes have been proposed. Some genes have been found altered in prostate cancer, such as PTEN, TP53, AR, RNASEL (HPC1), ELAC2 (HPC2), CDKN2A and MSR1 and those can be natural targets for new strategies of treatment. Besides, gene therapy has been suggested to be suitable for prostate cancer treatment. This approach includesex vivo corrective therapy, suicide, and antisense therapy.     

Comparing reconstruction with ileocecal graft to jejunal interposition pouch after total gastrectomy in rats.

After total gastrectomy, the ileocecal graft may act as a reservoir and protect against reflux but give rise to transposition of the ileum and cause possible changes in bile acid metabolism and nutrition. This study compared the ileocecal graft and jejunal pouch. Male Wistar rats weighing 265 +/- 22 g were submitted to sham operation (S), ileocecal interposition graft (IIG), and jejunal pouch interposition graft (JP) after total gastrectomy. Eight weeks later, the esophagus was examined for evidence of esophagitis. Nutritional biochemistry and weight profile were documented preoperatively and 8 weeks after surgery. The oral glucose tolerance test was performed. Thirty-three rats were operated on and 30 survived for 8 weeks. Esophagitis occurred in seven JP rats. Body weight was significantly higher in IIG than in JP rats (p < .05). Normal glucose tolerance to intragastric glucose load was observed in sham and operated rats. JP rats had a significant decrease in serum albumin, glucose, transferrin, hemoglobin, iron, folate, and calcium, compared to sham (p < .05). Cobalamine was significantly lower in IIG rats than in JP rats (p < .05). In the IIG and JP groups, serum/hepatic total bile acid did not differ significantly from preoperative and sham values. In conclusion, the IIG interposition graft in rats prevented esophagitis, preserved nutrition, and did not interfere with enterohepatic total bile acid circulation.     

Grafts of porcine intestinal submucosa for repair of cervical and abdominal esophageal defects in the rat.

Porcine small intestinal submucosa (SIS) is a cell-free collagen matrix that has demonstrated its ability as scaffold material for constructive remodeling of damaged or missing tissue. The purpose of this study was to evaluate the morphology and function of esophagoplasty in rat using a porcine SIS scaffold for the repair of a semi-circumferential defect in the cervical or in the abdominal esophagus. Sixty-seven rats underwent surgical excision of the anterior wall either of the cervical or of the abdominal esophagus and subsequent repair of the defect with an SIS patch graft. Outcomes of weight gain, signs of dysphagia, hematological and serum chemistry parameters, and barium swallow studies were used to assess the progress of healing and function over a 150-day time period. The grafts were studied for gross changes and histology at predetermined time points. Ninety-four percent of the SIS-treated rats survived, showing no significant differences in survival rate between groups. The grafted animals did well, without signs of dysphagia, and gaining weight. Barium swallow studies showed no evidence of fistula, significant stenosis, or diverticula. No hematological or serum biochemistry abnormalities were found. By 150 days, the SIS graft was replaced with esophageal-derived tissues. Specimens were completely lined by keratinized stratified squamous epithelium and showed complete regeneration of muscle fibers and scarce immunoreactivity for nerve. In the rat model, a patch graft technique using porcine SIS appears to induce esophageal regrowth either in cervical and abdominal esophagus. The repair mechanism occurred through a regenerative healing process.     

Multidimensional approach to the differences between muscular and articular temporomandibular patients: coping, distress, and pain characteristics.

OBJECTIVE: This study analyzes the differences in psychological variables and symptomatology between temporomandibular disorder diagnosis subgroups. STUDY DESIGN: The sample included 114 temporomandibular disorder patients that were evaluated in coping, distress, and temporomandibular characteristics. Diagnostic muscular (n = 58) and articular (n = 56) subgroups were compared in these variables through a MANCOVA. RESULTS: Muscular patients show a higher level of general distress, specifically in the anxiety and somatization subscales and a more active coping style, with a tendency of a minor use of humor and a higher number of parafunctional habits, specifically, biting nails, hangnails, and lips. CONCLUSION: We discuss the differences found in order to design the targets of the psychological intervention of temporomandibular patients.