SummaryWe examined structure and expression of the p53 and Rb genes in a C3HOS transplantable mouse model of osteosarcoma. The results were compared to analogous studies conducted with five human osteosarcoma cell lines. The p53 gene was found rearranged in the mouse tumour. The rearrangement mapped to the first intron region of the p53 gene and as a result, no p53 expression could be detected in C3HOS tumours. Using p53 genomic probes, we have detected the same rearrangement in the original radiation-induced tumour and the various clones that were isolated from it. Deletion and rearrangement of the p53 gene were also found in three out of five of the human osteosarcoma cell lines . No p53 expression could be detected in these three cell lines. In the affected human osteosarcoma cell lines, the rearrangement involved the first intron region. In addition, the mouse tumour was analysed for structural and expression changes in the Rb and the c-myc genes. Normal expression of both genes were detected in the murine tumour. Only one (Saos-2) human osteosarcoma cell line exhibited gross structural alteration in the retinoblastoma gene. The results suggest that the inactivation of p53 may be an important step in the development of osteosarcomas, and that a rearrangement affecting the first intron is common in osteosarcomas.

Objective. To measure changes in pharmacy and medical students' physician-pharmacist collaboration scores resulting from a workshop designed to promote understanding of the others' roles in health care. Methods. More than 88% of first-year pharmacy (n 5 215) and medical (n 5 205) students completed the Scale of Attitudes Toward Physician-Pharmacist Collaboration on 3 occasions in order to establish a baseline of median scores and to determine whether the scores were influenced by an interprofessional workshop. Results. Participation in the interprofessional workshop increased pharmacy students' collaboration scores above baseline (p50.02) and raised the scores of medical students on the education component of the collaboration survey instrument (p50.015). The collaboration scores of pharmacy students greatly exceeded those of medical students (p,0.0001). Conclusion. A workshop designed to foster interprofessional understanding between pharmacy and medical students raised the physician-pharmacist collaboration scores of both. Crucial practical goals for the future include raising the collaboration scores of medical students to those of pharmacy students.

A group-set of male Fischer 344 rats was kept on a choline-devoid (CD) diet for 3, 6, 9, 12 or 16 months. A second set was fed the same diet for 3, 6 and 9 months, followed by a control, choline-supplemented diet for 3 months. A third set was fed the CD diet for 0, 3, 6, 9 and 12 months and the control diet for the duration (16 months) of the experiment. [3H]Thymidine was injected into some of the animals to assess the extent of liver cell proliferation. In the latter animals, liver triacylglycerols were also determined as an index of the hepatonecrogenic action of a CD diet. Foci of enzyme-altered hepatocytes were detected histochemically and the presence of tumors was established histologically. Cell proliferation and triacylglycerols were both high after 3 months and declined steadily as the length of CD diet feeding increased. Upon subsequent feeding with the control diet, triacylglycerols promptly cleared from the liver, while cell proliferation remained at reduced but still relatively high levels for at least 3 months. Increasing but small numbers of foci of enzyme-altered hepatocytes were detected in some of the rats under experimentation for 6 months or longer. The incidence of hepatocellular carcinomas was 13, 27, 33 and 73% respectively in rats fed the CD diet for 3, 6, 9 or 12 months and the control diet for the duration of the experiment. On the other hand, only 26% of the rats fed exclusively the CD diet for 16 months developed carcinomas. The results were taken as evidence that: (i) liver cell proliferation persists beyond discontinuation of a CD diet; (ii) the liver contains initiated cells--capable of full evolution to cancer even in the absence of active promotion--after a relatively short 3-month exposure to a CD diet; and (iii) occurrence of a late event(s) is critical for the genesis of the tumors. Whether a CD diet is a complete carcinogen able to initiate de novo liver cells or acts merely as a promoter of the evolution of endogenous initiated cells is briefly discussed.

Objective. To determine the impact of performing critical-thinking and reflection assignments within interdisciplinary learning teams in a biochemistry course on pharmacy students' and prospective health professions students' collaboration scores. Design. Pharmacy students and prospective medical, dental, and other health professions students enrolled in a sequence of 2 required biochemistry courses. They were randomly assigned to interdisciplinary learning teams in which they were required to complete case assignments, thinking and reflection exercises, and a team service-learning project. Assessment. Students were asked to complete the Scale of Attitudes Toward Physician-Pharmacist Collaboration prior to the first course, following the first course, and following the second course. The physician-pharmacist collaboration scores of prospective health professions students increased significantly ( p,0.001). Conclusions. Having prospective health professions students work in teams with pharmacy students to think and reflect in and outside the classroom improves their attitudes toward physician-pharmacist collaboration.

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The p53 gene undergoes rearrangement in a high percentage of osteosarcomas, resulting in loss of its expression. A p53-null murine osteosarcoma cell line F6 was transfected with either a wild-type or a mutant p53 gene. Stably transfected cell lines were obtained, and their differentiation capabilities were compared in vitro with the parental cell line. Alkaline phosphatase and osteocalcin expression were measured as early and late differentiation markers, respectively. Induction of alkaline phosphatase expression was not affected by the presence of either p53 gene, whereas osteocalcin expression was seen in cells containing the wild-type p53 gene but not in the parental p53-null or mutant-expressing cell lines. That the induction of osteocalcin was intrinsically dependent on the presence of wild-type p53 was also indicated by the use of a temperature-sensitive Val 135 p53 mutant at 32 degrees C; predominant expression of p53 in the wild-type conformation resulted in osteocalcin expression. While the wild-type p53 gene could suppress tumor formation in vivo, the tumors expressing the mutant p53 gene grew two to three times as large as the tumors that did not express p53. Therefore, the absence of end-point differentiation in bone due to p53 rearrangements may contribute to the maintenance of the tumorigenic phenotype in osteosarcomas.

Liver tumors arise in rats fed a choline-devoid diet without added carcinogens. We found amplification of the c-myc gene in 13/13 of these tumors. The amplification ranged from 2-to 70-fold and was accompanied by an increase in c-myc gene expression. Amplification of c-myc was larger in tumors of rats fed a choline-devoid diet followed by a choline-supplemented diet than in tumors from animals fed a choline-devoid diet exclusively. In the former animals, low levels of c-myc gene amplification were also detected in nontumorous regions of tumor-bearing livers. The choline-devoid diet provides an in vivo experimental model for the induction of gene amplification in the rat liver. In this setting, amplification of the c-myc gene may be an early and critical event in carcinogenesis.There is evidence that the c-myc gene is involved in the control of normal cell division and in the process of cell transformation (1). In the liver, increased c-myc expression is seen during liver regeneration (2) and in hepatocellular carcinomas (3-8).Rats fed a choline-devoid (CD) diet, without exposure to chemical carcinogens, develop hepatocellular carcinomas after 14-16 months (9-11). The mechanism of this carcinogenesis is unknown, but major consequences of the diet include cell damage and a marked increase in hepatocyte turnover (12). DNA adducts were not detected in the livers of these animals, ruling out the possibility that contamination of the diet or the rats' environment with chemical carcinogens was responsible for the liver pathology and tumor induction (13). The CD diet also causes a general reduction in hepatocyte DNA methylation (14,15). During a study of ras genes, we noticed changes in endogenous viral sequences in tumor DNA and surmised that the changes may be characteristic of CD diet hepatocarcinogenesis (16). As liver cell death and regeneration persist during CD diet feeding, we undertook studies of c-myc expression and gene structure.In an earlier study, groups of rats were fed a continuous diet, either CD or choline-supplemented (CS), for 14-16 months, with tumor incidences of 26% and 0%, respectively (11,17). In later studies, groups were fed sequential diets CD diet for 3, 6, 9, and 12 months followed by CS diet for a combined total of 16 months; respective tumor incidences were 13%, 27%, 33%, and 73% (17). All tumors were well to moderately well differentiated hepatocellular carcinomas of trabecular, adenomatous, or mixed type. In this paper we report our analysis of c-myc gene structure and expression in these livers and tumors. MATERIALS AND METHODSExperimental Animals, Diets, and Tumors. All studies were performed on male Fisher 344 rats weighing 90-100 g when fed CD or CS diets, as described (11,16,17 Nucleic Acid Purification and Hybridization Methods. DNA and total RNA were purified from tumors and livers as described (16). Southern blotting and hybridization were also carried out as described (20). DNA was labeled by the random primer method (21) to a specific activity of 4 X 10' dpm/pxg and hy...

We previously demonstrated a correlation between wild-type p53 expression and appearance of osteoblastic-specific differentiation characteristics, as evidenced by basal osteocalcin gene expression in a mouse osteosarcoma tumor. The study reported here further explored the possibility of p53's having a distinct transcription-activating role in bone differentiation, in addition to its proposed role in G1 arrest and apoptosis. ROS17/2.3 osteoblastic osteosarcoma cells were stably transfected with a plasmid containing wild-type p53 binding sequences fused to the chloramphenicol acetyltransferase reporter gene. These cells were used to determine the transactivating role of p53 in regulation of osteocalcin gene expression. We chose two conditions under which osteocalcin expression is known to be upregulated: exposure of osteoblastic cells to differentiation-promoting medium and to vitamin D3. Exposure of the transfected cells to differentiation-promoting medium produced an increase in p53 transactivating activity correlating with the appearance of osteocalcin expression after about 1 wk. Vitamin D3 treatment resulted in upregulation of osteocalcin activity without a corresponding change in p53 transactivation activity or expression. In separate experiments, we tested whether changes in osteocalcin expression accompanied changes in p53 activity under conditions of downregulation of cell proliferation mediated by inhibition of DNA synthesis. Hydroxyurea treatment was used to inhibit DNA synthesis and produce growth arrest in osteoblastic cells. Inhibition of osteoblast cell proliferation was associated with a fourfold increase in p53 transactivating activity and a transient increase in osteocalcin steady-state expression. These results demonstrated a close relationship between p53 and osteocalcin and suggested a regulatory role for wild-type p53 in the control of basal osteocalcin gene expression in osteoblasts.