Single cell analysis of cancer cells using an improved RT-MLPA method has potential for cancer diagnosis and monitoring.

Single cell analysis techniques have great potential in the cancer genomics field. The detection and characterization of circulating tumour cells are important for identifying metastatic disease at an early stage and monitoring it. This protocol is based on transcript profiling using Reverse Transcriptase Multiplex Ligation-dependent Probe Amplification (RT-MLPA), which is a specific method for simultaneous detection of multiple mRNA transcripts. Because of the small amount of (circulating) tumour cells, a pre-amplification reaction is performed after reverse transcription to generate a sufficient number of target molecules for the MLPA reaction. We designed a highly sensitive method for detecting and quantifying a panel of seven genes whose expression patterns are associated with breast cancer, and optimized the method for single cell analysis. For detection we used a fluorescence-dependent semi-quantitative method involving hybridization of unique barcodes to an array. We evaluated the method using three human breast cancer cell lines and identified specific gene expression profiles for each line. Furthermore, we applied the method to single cells and confirmed the heterogeneity of a cell population. Successful gene detection from cancer cells in human blood from metastatic breast cancer patients supports the use of RT-MLPA as a diagnostic tool for cancer genomics.

Development of physical examination skills in a third-year surgical clerkship.

BACKGROUND: Previous studies have revealed deficiencies in physical examination (PE) skills of third-year medical students. The purpose of this study was to determine if formal teaching of PE skills and more supervised experiences with clinical faculty would increase the competency of these skills. METHODS: Forty-nine third-year medical students were assigned to one of two groups: group A was given formal instruction on PE skills at the beginning of the clerkship, and group B received the same instruction at the midpoint of the clerkship. The two groups received an assessment of the PE skills before, in the middle, and at the end of clerkship, utilizing a physician-developed checklist. RESULTS: Group A had a pretest score of 57.7%. Group B scored 59.8% on the pretest. Independent groups t tests were used to compare the two groups. At the midterm examination, group A increased their score to 71.0%, while group B decreased their score to 55.6%. From beginning to final, both groups had significant improvements in all areas, with group A scoring an overall mean of 75.1% (P = 0.0001), and group B scoring 75.5% (P = 0.0001). CONCLUSION: These findings show the effectiveness of structured learning situations for improving third-year surgery students' PE skills. There is also an indication that usual clerkship activities may not enhance learning of PE skills.

An evaluation of study habits of third-year medical students in a surgical clerkship.

BACKGROUND: This study was developed to assess study habits of medical students in a third-year surgical clerkship and to determine the relationship of these study habits to performance outcomes. METHODS: A questionnaire designed to assess medical student study habits was administered at the end of five consecutive 10-week multidisciplinary surgical clerkships. The results of questionnaires from 81 students were analyzed in respect to results on the National Board of Medical Education (NBME) surgical subtest and the multiple stations clinical examination (MSCE) given at the end of each clerkship. RESULTS: Although only 18 of the total 81 students reported studying in formal but self-directed groups, students who reported studying in a group on average scored 4 points higher on the MSCE than those who did not study in a group (P = 0.001). However, no significant differences or correlations were discovered between any of the study habits and the individual results on the NBME. CONCLUSION: Students may benefit from collaborative studying when it comes to clinical experience as demonstrated by improved performance on the MSCE.

Metabolism of garlic constituents in the isolated perfused rat liver.

The metabolic and kinetic behaviour of different garlic (Allium sativum L., Alliaceae) constituents were investigated in the isolated perfused rat liver, using aqueous extracts of garlic powder as well as isolated allicin, the main product of the enzymatic degradation of alliin. Allicin (allyl thiosulfinate) showed a remarkable first pass effect and passed the liver unmetabolized only at high concentrations which caused considerable cell injuries. Diallyl disulfide and allyl mercaptan were identified as metabolites of allicin, whereby diallyl disulfide probably is the metabolic precursor of allyl mercaptan as shown by perfusion with diallyl disulfide alone. The metabolites diallyl disulfide and allyl mercaptan could be determined in the perfusion medium as well as in the bile and the liver tissue. Other degradation products of garlic were also investigated in this model. Ajoenes and vinyldithiins were detected in perfusion medium after liver passage but no metabolites of them could be identified up to now.

Pharmacokinetics of vinyldithiins, transformation products of allicin.

The pharmacokinetic behaviour of vinyldithiins, the main constituents of oily preparations of garlic (Allium sativum L.), was investigated after oral administration of 27 mg 2-vinyl-4H-1,3-dithiin and 9 mg 3-vinyl-4H-1,2-dithiin to rats. In serum, kidney, and fat tissue, both vinyldithiins could be detected by GC-MS over a period of 24 h, whereas in liver only 1,3-vinyldithiin was found. Pharmacokinetic parameters (t1/2, ke, Cltot, AUC, and Vd) were determined using compartment models, elucidating the different pharmacokinetic behaviour of both vinyldithiins. 1,3-Vinyldithiin seems to be less lipophilic and is rapidly eliminated from serum, kidney, and fat tissue, whereas 1,2-vinyldithiin is more lipophilic and shows a tendency to accumulate in fat tissue. Experiments with liver homogenate confirmed the in vivo findings on the different degradation rates of both vinyldithiins. Allicin, the precursor of the vinyldithiins, is metabolized more rapidly in liver homogenate than the vinyldithiins.