SummaryWe investigate the proliferative activity, prostatic specific antigen (PSA) secretion, morphology and androgen response of human prostate tumour epithelial cells co-cultured with stromal cells in a bicameral system. Stromal and epithelial cells were isolated from prostate adenocarcinoma by enzyme digestion and cultured in defined media. Immunocytochemistry for prostate carcinoma tumour antigen (PCTA-1) was performed for culture purity evaluation. Also, the morphology of the epithelial cells in co-culture was evaluated by electron microscopy. PSA was determined by microparticle enzyme immunoassay (MEIA) automatized protocol and the proliferation was evaluated by a commercial spectrophotometric kit, based on formazan salt formation. Both cell cultures showed more than 90% of purity. The epithelial cell co-cultures showed marked membrane processes and cell interdigitations. The proliferative activity of the epithelial cells was increased in presence of stromal cells. Also, PSA secretion was significantly increased and maintained for at least 14 days, whereas the androgen response for PSA secretion was evidenced only in co-culture condition. Primary co-cultures of epithelial and stromal cells from human prostate carcinoma are able to maintain, for a prolonged time, proliferative and secretory properties as well hormone response, and represent a valuable tool for cellular and molecular studies on prostate cancer.

Four benzodiazepines flunitrazepam, nitrazepam, clonazepam, and alprazolam, have been analyzed from biological matrices by dual column gas chromatography using a nitrogen-phosphorus detector (NPD) and a micro-electron capture detector (m-ECD). The recoveries of the four benzodiazepines spiked into human whole blood and plasma were 88.8 -97.9 and 90.7 -97.5%, respectively. The regression equations for the four benzodiazepines showed excellent linearity. The detection limits (LODs) were 0.14 -0.95 ng/mL for whole blood and 0.13 -0.93 ng/mL for plasma. The method is simple and sensitive for the determination of benzodiazepines in human whole blood and plasma, and may be useful in forensic science practice.

A simple and rapid analytical method is developed and validated for the determination of antiepileptic and antipsychotic drugs in human whole blood using a solid-phase extraction and quantification by gas chromatography-mass spectrometry (GC/MS). Prazepam was used as internal standard (IS). The specificity, linearity, intra-and inter-assay precision and accuracy, and extraction recovery were fully evaluated. The limits of detection and quantification were in the range of 0.52 -0.98 mg/mL and 1.56 -2.97 mg/mL for antiepileptic drugs (pentobarbital, phenobarbital and carbamazepine) and 1.31 -2.94 ng/mL and 3.98 -8.89 ng/mL for antipsychotic drugs (chlorpromazine and thioridazine), respectively. The relative standard deviation (RSD) was less than 7.07%, while the intra-day accuracy was < 3.21% and the inter-day was < 8.80%, referred to RSD. The developed methodology was applied in forensic cases of death caused by phenothiazines in the North of Chile during 2010-2012.

The consumption of antidepressant drugs has increased in these last years, leading to severe and lethal poisonings. In this work, analytical tools, namely GC/MSD and GC/NPD/μECD, were used to identify and quantify several antidepressant drugs including amitriptyline, imipramine, sertraline, fluoxetine, and citalopram in forensic cases in the North of Chile during 2008-2011. Drugs were analyzed in biological arrays like blood. A solid phase extraction by Bond Elut Certify columns was applied in all these processes. Fluoxetine and sertraline were derivatized with pentafluoropropionic anhydride. Prazepam was used as internal standard (IS). The limit of detection (LOD) in blood were 0.5-20.07 ng/mL. The average extraction rate was 89.39% in blood. The relative standard deviation (RSD) was less than 3.6%, while the intra-day accuracy was < 5.5% and the inter-day was < 2.4%, referred to RSD. The procedures we have developed allow the quantification of drugs even at low therapeutic doses, a very important issue taking into account the nature of the analyzed arrays.