Attitudes and behaviors on physician-assisted death: a study of Michigan oncologists.

PURPOSE: To ascertain the attitudes of oncologists toward physician-assisted death, ie, physician-assisted suicide and active euthanasia, as well as their experiences with these activities and their opinions toward their legalization. METHODS: A survey was mailed to all practicing 250 oncologists in the state of Michigan, with subsequent development of psychometric scales and their correlation with self-reported behaviors in physician-assisted death. RESULTS: Analysis revealed five distinct, meaningful factors regarding approval or disapproval of physician-assisted death. These factors reflected global attitudes toward physician-assisted death, passive euthanasia, philosophical prohibitions toward physician-assisted death, concerns of legal consequences with physician-assisted death, and attitudes that physician-assisted death could be avoided with better end-of-life care (alpha = .94, .74, .76, .87, and .84, respectively). High levels of therapy withdrawal were reported (81%), with significant reservations toward assisted suicide and active euthanasia, although reported participation in such actions was noteworthy (18% and 4%, respectively). The scales reflecting global and philosophical attitudes correlated with several attitudes and behaviors toward physician-assisted death (P < .001). Legislation that would allow physician-assisted death was favored by 20.8% of respondents. CONCLUSION: Although they have reservations about physician-assisted death, significant numbers of oncologists are willing to consider such actions should they become legal. Given the substantial number of physicians who report that they have already participated in physician-assisted death, these findings may help better understand the attitudes that motivate physician behaviors toward assisted death.

Studies on type II collagen and aggrecan production in human articular chondrocytes in vitro and effects of transforming growth factor-beta and interleukin-1beta.

Type II collagen and aggrecan are major components of the extracellular matrix of articular cartilage. Their biosynthesis and catabolism are regulated by chondrocytes. They may be used as markers of chondrocyte phenotype for cells cultured in vitro. Type II collagen gene expression was detected by amplification of type II collagen-specific sequences, using cDNA produced by reverse transcription of mRNA extracted from freshly isolated and cultured human articular chondrocytes by the polymerase chain reaction (PCR). The synthesis of gene product was confirmed by immunohistochemical localization of type II collagen in cartilage sections and in cultured chondrocytes. Aggrecan core protein was also immunolocalized in cartilage sections and in chondrocytes in culture. Expression of type II collagen or aggrecan was not detected immunohistochemically in skin or bone. These results demonstrate that human articular chondrocytes can be characterized in culture, by the combined application of PCR and immunohistochemistry. Interleukin-1beta (IL-1beta) may play an important role in the destruction of cartilage matrix in arthritis, whereas transforming growth factor-beta (TGFbeta) may have an opposing effect and their combined actions may modulate chondrocyte phenotype. The effect of rhIL-1beta and rhTGFbeta on the production of type II collagen by chondrocytes in culture was investigated. It was shown that TGFbeta enhanced the production of type II collagen, localized immunocytochemically, in cultured chondrocytes. IL-1beta inhibited expression of mRNA for type II collagen. The implications of this study, in terms of a better understanding of degenerative cartilage disease, are discussed.

The effect of interleukin-1 on cytokine gene expression in cultured human articular chondrocytes analyzed by messenger RNA phenotyping.

OBJECTIVE: To investigate the pattern of cytokine gene expression in human articular chondrocytes in culture in response to interleukin-1 beta (IL-1 beta). The effect of serum and variations in culture conditions was also studied. METHODS: Messenger RNA was extracted from cells, reverse-transcribed to complementary DNA, and amplified by the polymerase chain reaction (PCR), using specific oligonucleotide primers. The PCR products were validated by restriction analysis with specific enzymes and by Southern blot analysis. RESULTS: In cultured articular chondrocytes, IL-1 beta, IL-1 alpha, granulocyte colony-stimulating factor (CSF), and granulocyte-macrophage CSF cytokine genes were expressed only after induction by IL-1 beta. However, IL-6, IL-8, and macrophage CSF genes were expressed constitutively. The expression of IL-1 beta was dose and time dependent. CONCLUSION: Using PCR, it was possible to demonstrate gene expression for several cytokines in human articular chondrocytes in culture. It was evident that some cytokine genes were expressed constitutively and some were inducible by IL-1 beta.

Expression of bone morphogenetic proteins in human prostatic adenocarcinoma and benign prostatic hyperplasia.

There are important interactions between prostatic tumours and bone. This study was designed to examine whether prostatic tissue can express bone inductive factors, in particular, the Bone Morphogenetic Proteins (BMPs). The polymerase chain reaction (PCR) has been used to screen for the expression of BMPs one to six in the prostatic tissue of patients with benign prostatic hyperplasia (BPH), non-metastatic prostatic adenocarcinoma and metastatic prostatic adenocarcinoma. BMPs were expressed in both benign and malignant prostate tissue and in the prostate tumour cell lines, PC3 and DU145. BMPs were also expressed in ocular melanoma tissue, a tissue which rarely metastasizes to bone. BMP-6 expression was detected in the prostate tissue of over 50% of patients with clinically defined metastatic prostate adenocarcinoma, but was not detected in non-metastatic or benign prostate samples or in ocular melanoma tissue. These findings suggest that the BMPs may play a role in the osteoinductive activity of prostate metastases and that the pattern of expression of BMPs may be important in the pathogenesis of osteoblastic metastases associated with prostate adenocarcinoma.

Mechanisms of action of cyclosporine and effects on connective tissues.

Cyclosporine is a potent immunomodulatory agent with an increasing number of clinical applications. Its major mode of action is inhibition of the production of cytokines involved in the regulation of T-cell activation. In particular, cyclosporine inhibits the transcription of interleukin 2. Although cyclosporine's major actions are on T cells, there is some evidence that it produces direct effects on other cell types. Its immunosuppressive action is closely linked to its binding of cyclophilin, a member of a family of high-affinity cyclosporine-binding proteins widely distributed in different cell types and in different species. The cyclophilins have been shown to have peptidyl-prolyl cis-trans isomerase enzyme activity that is blocked by cyclosporine. Although this may be a factor in cyclosporine's selective inhibition of cytokine gene transcription, it is still unclear whether inhibition of this activity is the mechanism through which cyclosporine exerts its effects on target cells. The ubiquitous presence of cyclophilins raises the question of why cyclosporine has major effects on T cells. Perhaps the critical proteins affected are transcriptional regulators restricted in their tissue distribution. The effects of cyclosporine on T cells and, directly or indirectly, on connective tissue cells, all of which can produce a range of cytokines, are of interest in relation to the tissue changes that occur in such inflammatory conditions as rheumatoid arthritis.

Detection of mRNA for the transforming growth factor beta family in human articular chondrocytes by the polymerase chain reaction.

The Transforming Growth Factor-beta (TGF beta) family of polypeptides elicits diverse biological actions on a wide range of cell types. There are known to be several isoforms of TGF beta coded for by different genes, with possibly differential expression and potencies. We have demonstrated that there is constitutive expression of three forms of transforming growth factor beta in adult human articular chondrocytes. The presence of 10% fetal calf serum in the culture medium may influence expression. The addition of transforming growth factor beta or interleukin 1 beta to the culture medium does not appear to consistently influence the expression of TGF beta by the cells.

Detection of the H-RAS oncogene in human thyroid anaplastic carcinomas.

We have transfected high-molecular-weight DNA from human thyroid carcinomas into murine 3T3 cells. As a result we identified several foci of morphologically distinct transformed cells in each of the tumour DNA transfected cultures. After a total of three rounds of transfection, the transformed cells were shown to form tumours in nude mice. Southern blot analysis of DNA prepared from third-round transfectants demonstrated the presence of human Alu repetitive sequences and, after hybridization with probes for known oncogenes, indicated the presence of the human H-RAS oncogene in 3T3 cells transfected with three out of four anaplastic carcinoma DNA samples. It appears therefore that activation of RAS genes may be an important event in the development of the anaplastic thyroid tumours.

Abnormal expression of the MOS proto-oncogene in human thyroid medullary carcinoma.

We have been studying the expression of a range of proto-oncogenes in human thyroid tumour tissue by using Northern blot analysis. We have demonstrated the expression of a MOS mRNA of 1 kb in all thyroid samples. Furthermore, in a medullary carcinoma sample we also observed additional mRNA species of 1.7 and 2.2 kb. Southern blot analysis of DNA prepared from the same tumour sample did not reveal a rearrangement of the gene. These findings are the first report of MOS expression in any human tissue, and indicate that MOS oncogene activation might be important in the development of some thyroid tumours.

Restriction fragment length polymorphisms in the ETS-1 proto-oncogene. Comparison of Saudi and Western populations.

We have cloned part of the ETS 1 proto-oncogene and demonstrated the presence of two polymorphic Sst I restriction sites. A probe derived from one of our clones revealed the presence of 8.3 kb, 9.5 kb and/or 11.5 kb fragments on Southern blots of human DNA samples. The relative frequencies of these alleles appear to be significantly different between Saudi and Western populations, but there are no apparent differences in these frequencies between Saudi non-leukemic and leukemic individuals.

T cell-macrophage interactions in the immune response to herpes simplex virus: the significance of interferon-gamma.

The antiviral properties of a herpex simplex virus type 1-specific 'helper' T cell clone were investigated. The clone was found to be deficient in interleukin 2 production, although it produced interleukin 3 and interferon-gamma upon stimulation with the virus in vitro. Supernatants containing these lymphokines were observed to increase the virocidal activity of macrophages in vitro and furthermore induced these cells to mediate cytotoxic activity against virus-infected target cells. Macrophage activation was linked to the presence of interferon-gamma in the clone supernatant. The implications of these results for protection against this virus in vivo are discussed.

Calmodulin in human serum and the specific release of calmodulin from calmodulin-rich platelets.

Calmodulin (CaM)-like activity was detected in human serum and foetal calf serum, with an activity 10 times more than that detectable in plasma. Serum CaM was largely accounted for by release from human platelets as confirmed by both radioimmunoassay and sodium-dodecyl-sulphate/polyacrylamide-gel electrophoresis of CaM partially purified from platelet releasate obtained in response to thrombin. Lactate dehydrogenase release was unaffected by thrombin. Platelet CaM content was very variable (1.3 to 11.3 micrograms/mg protein; n = 15). It is suggested that intact platelets are rich in CaM and that release of CaM during preparation explains the variation in CaM content reported here and in the literature.

Calcium, calmodulin, and the production of prostacyclin by cultured vascular endothelial cells.

The production of prostacyclin (PGI2) by cultured porcine aortic endothelial cells, in response to serum and the calcium ionophore A23187, was inhibited by TMB-8, an antagonist of intracellular calcium mobilization. The calcium-channel blocker methoxyverapamil (D600) inhibited serum-induced PGI2 production in but had little effect on A23187-induced PGI2 production. Calmodulin activity was detected in endothelial-cell lysates and was inhibited by the calmodulin antagonist W7, which also inhibited PGI2 production in response to both agonists. Calcium and calmodulin appear to play an important role in mediating PGI2 production by the vascular endothelium.

The presence in normal plasma, serum and platelets of factors that stimulate the production of prostacyclin (PGI2) by cultured endothelial cells.

1. The effect of plasma and serum from normal subjects on the production of prostacyclin by cultured porcine endothelial cells was investigated. 2. Both plasma and serum from all subjects studied significantly stimulated the production of prostacyclin by cultured endothelial cells, measured by the radioimmunoassay of its stable metabolite 6-oxoprostaglandin F1 alpha. 3. Serum caused a consistently greater stimulation than plasma from the same individual. The stimulation was dose-dependent and inhibited by indomethacin. Heparin added to serum also inhibited this response. 4. Extracts from isolated washed platelets were tested for their ability to increase prostacyclin production. Extracts from platelets which had been induced to aggregate and release their granule contents in response to thrombin, caused stimulation. 5. These results indicated the invariable presence in plasma and serum of factors that stimulate the production of prostacyclin by endothelial cells in vitro. At least one of these factors is derived from platelets. These factors may be involved in the regulation of prostacyclin production by the vascular endothelium under normal conditions and in disease states.

Increased thymidine incorporation into fetal rat cartilage in vitro in the presence of human somatomedin, epidermal growth factor and other growth factors.

The incorporation of [3H]thymidine by rat costal cartilage in vitro was studied at different fetal and postnatal ages and the effect of partially purified human somatomedin, mouse epidermal growth factor, platelet secretion products, insulin and growth hormone on thymidine uptake by fetal cartilage was examined. Thymidine uptake in plasma-free medium was many times greater in late fetal life than after birth. The incorporation of [3H]thymidine into costal cartilage from 21-day fetuses was significantly (P less than 0.05) increased above control values in the presence of 10 micrograms somatomedin/1, and when cartilage was incubated in medium containing somatomedin and diluted human plasma there was a synergistic action. Epidermal growth factor at a concentration of 1 ng/l was a potent stimulator of thymidine uptake. Secretion products from human platelets after their aggregation by thrombin stimulated [3H]thymidine uptake at a concentration of 2% (v/v), but were inhibitory at high concentrations. High concentrations of platelet secretion products stimulated the incorporation of [35S]sulphate by cartilage. A pharmacological concentration of 10 mu. insulin/ml stimulated [3H]thymidine uptake, but not concentrations of 1 or 100 mu./ml. Growth hormone had no effect. The results showed that fetal cartilage had a greater endogenous mitogenic activity than postnatal cartilage. While somatomedins may be important in the regulation of fetal body growth, other protein growth factors also stimulate fetal skeletal tissues.

Stimulation of prostacyclin production from vascular endothelial cells by plasma, serum and platelets under normal and pathological conditions.

We have investigated the effects of plasma and serum from normal subjects on the production of prostacyclin (PGI2) by cultured porcine aortic vascular endothelial cells, measured by radioimmunoassay of its stable metabolite 6-keto prostaglandin F1 alpha (6-keto PGF1 alpha). Both plasma and serum caused significant stimulation of the production of 6-keto PGF1 alpha over basal levels. Serum caused consistently greater stimulation than plasma from the same individual. Washed platelet suspensions were induced to aggregate using thrombin and the supernatants stimulated the production of 6-keto PGF1 alpha by cultured endothelial cells. Preliminary studies also show that a stimulatory factor is released from cultured human leucocytes. Serum from patients with systemic lupus erythematosus (SLE) and severe systemic sclerosis (SS), two connective tissue diseases with autoimmune features and vascular complications, showed significantly reduced levels of stimulation when compared with a control group.