Advance directives: the views of health care professionals.

OBJECTIVES: This study examined the views and experiences of 20 physicians and 20 nurses at a major Canadian teaching hospital regarding the use of advance directives in clinical care. DESIGN: The participants were purposively drawn from four clinical specialties: family and community medicine, oncology, intensive care and geriatrics. Detailed interviews were conducted in person. Content analysis was used to code the data, which were further analysed with both quantitative and qualitative techniques. MAIN RESULTS: Thirty-nine of the 40 participants favoured the use of advance directives in clinical care; physicians had somewhat less positive attitudes than nurses toward such directives. Advance directives were thought by participants to be helpful in resolving disagreements between patients and their families about treatment options; in making patients more comfortable, both physically and psychologically, during the process of dying; and in opening up communication and trust among patients, their families and health care professionals. Concerns about the use of advance directives focused on the lack of clarity in some patients'' instructions, the absence of legal status for directives, the possible interference with a practitioner''s clinical judgement, the adequacy and appropriateness of patients'' information about their circumstances, and the type of intervention (passive or active) requested by patients. CONCLUSIONS: New regulations and legislation are making the use of advance directives more widespread. Health care professionals should participate in the development and implementation of these directives. Continuing professional education is essential in this regard.     

Biochemical characterization of the tobacco 42-kD protein kinase activated by osmotic stress

In tobacco (Nicotiana tabacum), hyperosmotic stress induces rapid activation of a 42-kD protein kinase, referred to as Nicotiana tabacum osmotic stress-activated protein kinase (NtOSAK). cDNA encoding the kinase was cloned and, based on the predicted amino acid sequence, the enzyme was assigned to the SNF1-related protein kinase type 2 (SnRK2) family. The identity of the enzyme was confirmed by immunoprecipitation of the active kinase from tobacco cells subjected to osmotic stress using antibodies raised against a peptide corresponding to the C-terminal sequence of the kinase predicted from the cloned cDNA. A detailed biochemical characterization of NtOSAK purified from stressed tobacco cells was performed. Our results show that NtOSAK is a calcium-independent Ser/Thr protein kinase. The sequence of putative phosphorylation sites recognized by NtOSAK, predicted by the computer program PREDIKIN, resembled the substrate consensus sequence defined for animal and yeast (Saccharomyces cerevisiae) AMPK/SNF1 kinases. Our experimental data confirmed these results, as various targets for AMPK/SNF1 kinases were also efficiently phosphorylated by NtOSAK. A range of protein kinase inhibitors was tested as potential modulators of NtOSAK, but only staurosporine, a rather nonspecific protein kinase inhibitor, was found to abolish the enzyme activity. In phosphorylation reactions, NtOSAK exhibited a preference for Mg(2+) over Mn(2+) ions and an inability to use GTP instead of ATP as a phosphate donor. The enzyme activity was not modulated by 5'-AMP. To our knowledge, these results represent the first detailed biochemical characterization of a kinase of the SnRK2 family.     

The copper transport protein Atox1 promotes neuronal survival

Atox1, a copper transport protein, was recently identified as a copper-dependent suppressor of oxidative damage in yeast lacking superoxide dismutase. We have previously reported that Atox1 in the rat brain is primarily expressed in neurons, with the highest levels in distinct neuronal subtypes that are characterized by their high levels of metal, like copper, iron, and zinc. In this report, we have transfected the Atox1 gene into several neuronal cell lines to increase the endogenous level of Atox1 expression and have demonstrated that, under conditions of serum starvation and oxidative injury, the transfected neurons are significantly protected against this stress. This level of protection is comparable with the level of protection seen with copper/zinc superoxide dismutase and the anti-apoptotic gene bcl-2 that had been similarly transfected. Furthermore, neuronal cell lines transfected with a mutant Atox1 gene, where the copper binding domain has been modified to prevent metal binding, do not afford protection against serum starvation resulting in apoptosis. Therefore, Atox1 is a component of the cellular pathways used for protection against oxidative stress.