Administration of Blood By Registered Nurses—Joint Statement by California Medical Association,California Hospital Association and California Nurses' Association

After several years of favorable experience with registered nurses giving intravenous injections of fluids under the criteria set out in a joint statement by the California Medical Association, the California Hospital Association and the California Nurses'' Association,^* it was proposed that it would be appropriate for registered nurses to administer blood. Careful study of current experience in various places with registered nurses giving blood transfusions convinced a joint committee that it recommend the adoption of the following statement which was approved by the associations indicated.     

Influence of euthanasia method on blood and gill variables in normoxic and hypoxic Gulf killifish Fundulus grandis

In many experiments, euthanasia, or humane killing, of animals is necessary. Some methods of euthanasia cause death through cessation of respiratory or cardiovascular systems, causing oxygen levels of blood and tissues to drop. For experiments where the goal is to measure the effects of environmental low oxygen (hypoxia), the choice of euthanasia technique, therefore, may confound the results. This study examined the effects of four euthanasia methods commonly used in fish biology (overdose of MS‐222, overdose of clove oil, rapid cooling and blunt trauma to the head) on variables known to be altered during hypoxia (haematocrit, plasma cortisol, blood lactate and blood glucose) or reflecting gill damage (trypan blue exclusion) and energetic status (ATP, ADP and ATP:ADP) in Gulf killifish Fundulus grandis after 24 h exposure to well‐aerated conditions (normoxia, 7·93 mg O₂ l^?1, c. 150 mm Hg or c. 20 kPa) or reduced oxygen levels (0·86 mg O₂ l^?1, c. 17 mm Hg or c. 2·2 kPa). Regardless of oxygen treatment, fish euthanized by an overdose of MS‐222 had higher haematocrit and lower gill ATP:ADP than fish euthanized by other methods. The effects of 24 h hypoxic exposure on these and other variables, however, were equivalent among methods of euthanasia (i.e. there were no significant interactions between euthanasia method and oxygen treatment). The choice of an appropriate euthanasia method, therefore, will depend upon the magnitude of the treatment effects (e.g. hypoxia) relative to potential artefacts caused by euthanasia on the variables of interest.     

Euthanasia in Belgium: trends in reported cases between 2003 and 2013

Background:In 2002, the Belgian Act on Euthanasia came into effect, regulating the intentional ending of life by a physician at the patient’s explicit request. We undertook this study to describe trends in officially reported euthanasia cases in Belgium with regard to patients’ sociodemographic and clinical profiles, as well as decision-making and performance characteristics.Methods:We used the database of all euthanasia cases reported to the Federal Control and Evaluation Committee on Euthanasia in Belgium between Jan. 1, 2003, and Dec. 31, 2013 (n = 8752). The committee collected these data with a standardized registration form. We analyzed trends in patient, decision-making and performance characteristics using a χ² technique. We also compared and analyzed trends for cases reported in Dutch and in French.Results:The number of reported euthanasia cases increased every year, from 235 (0.2% of all deaths) in 2003 to 1807 (1.7% of all deaths) in 2013. The rate of euthanasia increased significantly among those aged 80 years or older, those who died in a nursing home, those with a disease other than cancer and those not expected to die in the near future (p < 0.001 for all increases). Reported cases in 2013 most often concerned those with cancer (68.7%) and those under 80 years (65.0%). Palliative care teams were increasingly often consulted about euthanasia requests, beyond the legal requirements to do so (p < 0.001). Among cases reported in Dutch, the proportion in which the person was expected to die in the foreseeable future decreased from 93.9% in 2003 to 84.1% in 2013, and palliative care teams were increasingly consulted about the euthanasia request (from 34.0% in 2003 to 42.6% in 2013). These trends were not significant for cases reported in French.Interpretation:Since legalization of euthanasia in Belgium, the number of reported cases has increased each year. Most of those receiving euthanasia were younger than 80 years and were dying of cancer. Given the increases observed among non–terminally ill and older patients, this analysis shows the importance of detailed monitoring of developments in euthanasia practice.In 2002, Belgium legalized euthanasia, defined as the intentional ending of life by a physician at the patient’s explicit request.^1,2 For a patient to be eligible for euthanasia, certain formal criteria for due care must be met.¹ These include a voluntary, well-considered, repeated and written request, expressed by a person with full mental capacity who is fully informed about his or her medical condition and the remaining therapeutic possibilities.¹ The person must be in a state of constant and unbearable physical or mental suffering that cannot be alleviated. Due care criteria for the procedure include an a priori consultation with a second independent physician, consultation with a third physician in cases where death is not expected in the foreseeable future and a posteriori reporting of the case for evaluation purposes.¹To safeguard due process and legal compliance and to enable societal control and evaluation, a mandatory notification procedure was built into the legislation.³ Physicians are required to report each case of euthanasia to the multidisciplinary Belgian Federal Control and Evaluation Committee on Euthanasia by completing and submitting a registration form within 4 working days after a death by euthanasia.^1,3 The evaluation committee reviews the form and determines whether euthanasia was performed in accordance with the legal requirements. Initially, only anonymous information is reviewed; where there is doubt about legality, the committee can revoke anonymity by majority decision and can ask the reporting physician for additional information. If the committee is of the opinion, based on a two-thirds majority, that the legal requirements were not fulfilled, the case is sent to the public prosecutor.^1,3 Although not mentioned in the Belgian law, physician-assisted suicide is treated as a form of euthanasia by the committee.⁴To facilitate societal control, the Federal Control and Evaluation Committee on Euthanasia is legally required to issue biennial reports of all reported cases,^1,3–9 providing basic statistics, an evaluation of the law and further recommendations. However, these statistics do not provide an overview of long-term trends. A more complete and thorough evaluation of case characteristics and analysis of trends is needed. In this way, adherence to the legal criteria can be evaluated, and developments in euthanasia practice that might raise concerns can be identified and addressed.Belgium has 2 main language communities: those who speak Dutch (roughly 60% of the population), who mainly live in Flanders, and those who speak French (about 40%), who mainly live in Wallonia. The Brussels-Capital Region is officially bilingual, but predominantly French-speaking. Several empirical studies have found differences in end-of-life practices, knowledge and attitudes between the regions and language communities, showing that Dutch-speaking physicians more often receive and grant euthanasia requests and are more inclined to adhere to legal safeguards.^10–14 The reports issued by the Federal Control and Evaluation Committee on Euthanasia show a striking disparity in euthanasia reporting between the 2 language communities.^4–9 Trends in the characteristics of reported cases and differences among them have not yet been studied.The committee’s reports have shown a continuing increase in the number of euthanasia cases.^4–9 The primary objective of this study was to examine changes in the number and incidence of euthanasia cases and the proportion of euthanasia cases relative to all deaths in Belgium up to and including 2013. The secondary objectives were to determine and report the sociodemographic and clinical characteristics of patients, the decision-making and performance characteristics of reported cases and the differences in trends in characteristics between cases reported in Dutch and cases reported in French.     

Legal developments concerning active euthanasia on request in the Netherlands

The author describes social and legal developments in the debate over active euthanasia in the Netherlands. There, as in all of Western Europe, euthanasia is a crime. Although the medical profession in several European countries has rejected efforts to change the situation, the Dutch Medical Association in 1984 issued a statement indicating an increased willingness to accept euthanasia under stringent guidelines. Some court decisions have been lenient in applying penal law to doctors acting at the wish of their patients. The Netherlands State Commission on Euthanasia published a 1985 report advocating modification of the Penal Code. [An English summary of the report appears in the same issue of Bioethics]. An emotional debate is underway in medical and legal journals and in the mass media, but the Council of State has advised against any legislative change for now. Meanwhile, euthanasia continues to occur in the Netherlands at an estimated rate of 5,000 to 8,000 cases a year.     

The role of nurses in physician-assisted deaths in Belgium

Background

Belgium’s law on euthanasia allows only physicians to perform the act. We investigated the involvement of nurses in the decision-making and in the preparation and administration of life-ending drugs with a patient’s explicit request (euthanasia) or without an explicit request. We also examined factors associated with these deaths.

Methods

In 2007, we surveyed 1678 nurses who, in an earlier survey, had reported caring for one or more patients who received a potential life-ending decision within the year before the survey. Eligible nurses were surveyed about their most recent case.

Results

The response rate was 76%. Overall, 128 nurses reported having cared for a patient who received euthanasia and 120 for a patient who received life-ending drugs without his or her explicit request. Respectively, 64% (75/117) and 69% (81/118) of these nurses were involved in the physician’s decision-making process. More often this entailed an exchange of information on the patient’s condition or the patient’s or relatives’ wishes (45% [34/117] and 51% [41/118]) than sharing in the decision-making (24% [18/117] and 31% [25/118]). The life-ending drugs were administered by the nurse in 12% of the cases of euthanasia, as compared with 45% of the cases of assisted death without an explicit request. In both types of assisted death, the nurses acted on the physician’s orders but mostly in the physician’s absence. Factors significantly associated with a nurse administering the life-ending drugs included being a male nurse working in a hospital (odds ratio [OR] 40.07, 95% confidence interval [CI] 7.37–217.79) and the patient being over 80 years old (OR 5.57, 95% CI 1.98–15.70).

Interpretation

By administering the life-ending drugs in some of the cases of euthanasia, and in almost half of the cases without an explicit request from the patient, the nurses in our study operated beyond the legal margins of their profession.Medical end-of-life decisions with a possible or certain life-shortening effect occur often in end-of-life care.^1–5 The most controversial and ethically debated medical practice is that in which drugs are administered with the intention of ending the patient’s life, whether at the patient’s explicit request (euthanasia) or not. The debate focuses mainly on the role and responsibilities of the physician.⁶ However, physicians worldwide have reported that nurses are also involved in these medical practices, mostly in the decision-making and sometimes in the administration of the life-ending drugs.^1–3,7–9 Critical care,¹⁰ oncology¹¹ and palliative care nurses^12,13 have confirmed this by reporting their own involvement, particularly in cases of euthanasia.^14,15In Belgium, the law permits physicians to perform euthanasia under strict requirements of due care, one of which is that they must discuss the request with the nurses involved.¹⁶ There are no further explicit stipulations determining the role of nurses in euthanasia. Physician-assisted death is legally regulated in some other countries as well (e.g., the Netherlands, Luxemburg and the US states of Oregon and Washington State), without specifying the role of nurses. Reports from nurses in these jurisdictions are scarce, apart from some that are limited to particular settings, or lack details about their involvement.^13,14We conducted this study to investigate the involvement of nurses in Flanders, Belgium, in the decision-making and in the preparation and administration of life-ending drugs with, or without, a patient’s explicit request. We also examined patient- and nurse-related factors associated with the involvement of nurses in these deaths. In a related research article, Chambaere and colleagues describe the findings from a survey of physicians in Flanders about the practices of euthanasia and assisted suicide, and the use of life-ending drugs without an explicit request from the patient.¹⁷     

Physician-assisted deaths under the euthanasia law in Belgium: a population-based survey

Background

Legalization of euthanasia and physician-assisted suicide has been heavily debated in many countries. To help inform this debate, we describe the practices of euthanasia and assisted suicide, and the use of life-ending drugs without an explicit request from the patient, in Flanders, Belgium, where euthanasia is legal.

Methods

We mailed a questionnaire regarding the use of life-ending drugs with or without explicit patient request to physicians who certified a representative sample (n = 6927) of death certificates of patients who died in Flanders between June and November 2007.

Results

The response rate was 58.4%. Overall, 208 deaths involving the use of life-ending drugs were reported: 142 (weighted prevalence 2.0%) were with an explicit patient request (euthanasia or assisted suicide) and 66 (weighted prevalence 1.8%) were without an explicit request. Euthanasia and assisted suicide mostly involved patients less than 80 years of age, those with cancer and those dying at home. Use of life-ending drugs without an explicit request mostly involved patients 80 years of older, those with a disease other than cancer and those in hospital. Of the deaths without an explicit request, the decision was not discussed with the patient in 77.9% of cases. Compared with assisted deaths with the patient’s explicit request, those without an explicit request were more likely to have a shorter length of treatment of the terminal illness, to have cure as a goal of treatment in the last week, to have a shorter estimated time by which life was shortened and to involve the administration of opioids.

Interpretation

Physician-assisted deaths with an explicit patient request (euthanasia and assisted suicide) and without an explicit request occurred in different patient groups and under different circumstances. Cases without an explicit request often involved patients whose diseases had unpredictable end-of-life trajectories. Although opioids were used in most of these cases, misconceptions seem to persist about their actual life-shortening effects.Euthanasia and physician-assisted suicide are heavily debated issues in medical practice. In recent years, three European countries (Belgium and the Netherlands in 2002, and Luxemburg in 2009) and two US states (Oregon in 1997 and Washington State in 2009) decriminalized euthanasia and physician-assisted suicide under formal conditions.^1–5 Canada is among a number of countries where the debate over legalization has flared up, with a proposed bill reaching Parliament and a pro-euthanasia proposal by the Quebec College of Physicians.⁶Understandably, the issue of euthanasia triggers much emotion and can be fraught with speculative arguments. Opponents of euthanasia often argue that legalizing the procedure will lead to a rise in the use of life-ending drugs without a patient’s explicit request, especially in vulnerable patient groups.^7–10 Thus far, however, no indications of this have been found in studies of physician-assisted deaths before and after legalization in Belgium and the Netherlands.^9,11,12 In Belgium, the percentage of deaths in which life-ending drugs were used remained stable, and the proportion without an explicit request from the patient decreased.¹² Other studies have shown that euthanasia, physician-assisted suicide and the use of life-ending drugs without explicit patient request are not confined to countries where physician-assisted death is legal.^13–16In addition to knowing the overall occurrence of physician-assisted death, it is equally important for an adequately informed, empirically based debate to know its performance in vulnerable patient groups and the care put into the decision and performance. In light of legalization and its alleged effects on the use of life-ending drugs without patient request, it is also important to map similarities and differences between euthanasia and the use of life-ending drugs without explicit patient request. In this article, we report our investigation of demographic and clinical characteristics associated with physician-assisted deaths in Flanders, Belgium; the involvement of the patient, relatives and other caregivers in the decision-making process; reasons for the decisions; aspects of the treatment trajectory; and details of the performance in terms of drug use and the people administering the life-ending drugs.     

Special Reviews

Fair Isle and its Birds, Kenneth Williamson. Oliver & Boyd, Edinburgh and London, 1965. 30/-. xvi + 311 pages, 17 figures and maps, 27 photographs.

Waterfowl Tomorrow, edited by Joseph P. Linduska. U.S. Dept. of the Interior, Bureau of Sport Fisheries and Wildlife, Washington D.C., 1964. 4 dollars. 770 pages, 187 photographs, I (folded) map, many drawings by Bob Hines. For sale by the Superintendent of Documents, Washington D.C., 20402.

The Giant Canada Goose, Harold C. Hanson. Southern Illinois University Press (U.K. Agent, W. S. Hall, London), 1965. $ 9.75. 226 pages, 74 photos, text-figures and tables.

The Harvest that Kills, John Coleman-Cooke. Odhams, 1965. 25/-. 207 pages.

Gardening without Poisons, Beatrice Trum Hunter. Hamish Hamilton, London, 1 965. 25/-. 294 pages, line drawings.

The Life of the Robin, David Lack. H. F. & G. Witherby Ltd., London, 1965. 30/-. 240 pages, text-figures.

Vertebrate Fauna of the Halifax Parish, compiled by I. Morley. Halifax Scientific Society, 1965. 103 pages, 3 photographic plates.

The Bird Table Book, Tony Soper. David & Charles, Dawlish, and Macdonald, London, 1965. 21/-. 140 pages, 20 photos, numerous line drawings.

Song and Garden Birds of North America; Water, Prey and Game Birds of North America, Alexander Wetmore, with chapters by Dean Amadon, E. T. Gilliard, A. H. Miller, R. C. Murphy, R. T. Peterson, and others. $11.95 each (post paid), from Nat. Geog. Soc., Dept. 280, Washington, D.C., 20036. 400 and 464 pages respectively, numerous illustrations, nearly all in colour, 6 long-playing records in cover-flap of each.

The Birds of Shakespeare, J. E. Harting. Argonaut Inc. Publishers, Chicago, 1965. $7.50. 321 + xxxviii pages.

Serengeti Shall Not Die, B. & M. Grzimek. Collins, London, 1965. 25/-. 256 pages, 44 photos, text-figures.

Wings of Light, Garth Christian. Newnes, London, 1965. 35/-. 178 pages, 31 photos.

Antarctic Biology (Proceedings of the First Symposium on Antarctic Biology held at Paris in 1962), ed. R. Carrick, M. Holdgate and J. Prévost. Editions Hermann (115 Boulevard St-Germain, Paris 6), 1965. 54 N.F. 652 pages, 11 plates, numerous text-figures.

Birds of Town and Village, W. D. Campbell; paintings by Basil Ede.Country Life, London, 1965. 126/-. 156 pages, 36 coloured plates.

The Game Research Association, 5th Annual Report, 1965.

The Wildfowlers' Association of Great Britain and Ireland, Report and Year Book 1964–65.     

Shall Organized Medicine be Unified, or Separate?

At present, physicians in California who choose to join organized medicine do so through their county medical societies, and membership in the California Medical Association and the American Medical Association is then automatic. At the March meeting of the CMA House of Delegates, question was raised whether membership in CMA, and the AMA, or both should remain automatic.The House requested an ad hoc committee to cause a "poll and its attendant statements to be developed by May 21 for copy distribution to component medical societies and printing in the CMA membership news media-with mailing of the official questionnaires to the society members on September 1, 1971." Members will be asked to express their opinions by ballot in September.The Speaker of the House appointed an ad hoc committee of the House to conduct this informed opinion poll of the membership. The committee has met to set ground rules, prepare accurate pro and con statements and write the poll questions-in accord with the directions of the House action.The Informed Membership Opinion Poll Committee, with the advice of Decision Making Information, Inc., an independent consultant, prepared statements regarding unified and separate membership in CMA and AMA from comments which were solicited from every county medical society. A statement by legal counsel for the California Medical Association on the structural relationship of AMA, CMA and component societies, and the statements on unified or separate membership prepared by the committee appear on the following two pages.     

Mouse embryo yield and viability after euthanasia by CO2 inhalation or cervical dislocation

Efficient production of transgenic mice requires high yields of viable, healthy embryos. Cervical dislocation (without prior anesthesia) rather than CO2 inhalation as a means of euthanasia has been justified on the basis of the increased yield of viable ova, but controlled studies have not directly supported this contention. The American Veterinary Medical Association (AVMA) and Canadian Council on Animal Care (CCAC) Guides, and respective Institutional Animal Care and Use Committees (IACUC) have supported the use of CO2 as a preferred, humane method. The study reported here was undertaken to determine the relative yields of viable embryos from mice euthanized either by inhalation of 100% CO2 or by cervical dislocation. Inbred and hybrid mouse strains, representative of common strains used in genetic engineering experimentation included C57BL/6, FVB/N, and B6SJLF1. There was no difference in the embryo yields in comparisons using the two methods of euthanasia (P = 0.534). Decisions regarding the method of euthanasia can be made on the basis of criteria other than those associated with embryo yield and viability.     

WORKMEN'S COMPENSATION ACT—Some Aspects of Interest to California Physicians

This article was prepared by Mr. William M. Whelan, Director of Special Services, California Medical Association, under the supervision of Dr. Francis J. Cox, Chairman of the Medical Services Commission of the Association, and Mr. Howard Hassard, the Association''s Legal Counsel. It is intended as a brief synopsis of the California Workmen''s Compensation Law as it applies to the physician in private practice. It is not an exhaustive treatment of the subject. A physician who desires to acquaint himself in detail with California industrial practice should consult the article entitled “The Physician''s Role in Workmen''s Compensation,” California Medicine, 82:352-362, April, 1955. Inquiries regarding industrial medicine should be addressed to Mr. William M. Whelan, California Medical Association, 450 Sutter St., San Francisco 8.     

Attitudes of cancer patients, family caregivers, oncologists and members of the general public toward critical interventions at the end of life of terminally ill patients

Background:

Whereas most studies have focused on euthanasia and physician-assisted suicide, few have dealt comprehensively with other critical interventions administered at the end of life. We surveyed cancer patients, family caregivers, oncologists and members of the general public to determine their attitudes toward such interventions.

Methods:

We administered a questionnaire to four groups about their attitudes toward five end-of-life interventions — withdrawal of futile life-sustaining treatment, active pain control, withholding of life-sustaining measures, active euthanasia and physician-assisted suicide. We performed multivariable analyses to compare attitudes and to identify sociodemographic characteristics associated with the attitudes.

Results:

A total of 3840 individuals — 1242 cancer patients, 1289 family caregivers and 303 oncologists from 17 hospitals, as well as 1006 members of the general Korean population — participated in the survey. A large majority in each of the groups supported withdrawal of futile life-sustaining treatment (87.1%–94.0%) and use of active pain control (89.0%–98.4%). A smaller majority (60.8%–76.0%) supported withholding of life-sustaining treatment. About 50% of those in the patient and general population groups supported active euthanasia or physician-assisted suicide, as compared with less than 40% of the family caregivers and less than 10% of the oncologists. Higher income was significantly associated with approval of the withdrawal of futile life-sustaining treatment and the practice of active pain control. Older age, male sex and having no religion were significantly associated with approval of withholding of life-sustaining measures. Older age, male sex, having no religion and lower education level were significantly associated with approval of active euthanasia and physician-assisted suicide.

Interpretation:

Although the various participant groups shared the same attitude toward futile and ameliorative end-of-life care (the withdrawal of futile life-sustaining treatment and the use of active pain control), oncologists had a more negative attitude than those in the other groups toward the active ending of life (euthanasia and physician-assisted suicide).As more attention turns to when and how the lives of terminally ill patients end in the clinical setting, debate about the issues of euthanasia and physician-assisted suicide grows.^1–5 Euthanasia has been discussed in Europe and the United States for more than a century, and the public has become more accepting of it.^4–10 Announcing its first-ever ruling in favour of an unconscious patient’s right to die with dignity, the Korean Supreme Court recently ruled that doctors of an elderly woman in a persistent vegetative state remove the artificial respirator from her on the basis of her presumed wishes.¹¹ A public debate aimed at legalizing withdrawal of futile life-sustaining treatment, exposure to stories of dying patients in the mass media, and the court’s decision may have led to a greater awareness of, and sensibility toward, the rights of terminally ill patients. In 2000, only 16.5% of 535 Korean oncologists surveyed said that they would prescribe morphine for severe cancer pain, and more than half of 655 patients who had pain said they had inadequate pain management.¹²Although much has been written about attitudes toward how the general public would choose to die in the clinical setting,^4,13 most studies have focused on only euthanasia and physician-assisted suicide.^14–18 We conducted a large survey to examine attitudes among cancer patients, family caregivers, oncologists and members of the general public toward critical interventions at the end of life of terminally ill patients.     

Continuing Medical Education: Planning and Goals Conference in Continuing Medical Education—Workshop Recommendations

Following are recommendations drawn up by workshops held as a part of the Planning and Goals Conference in Continuing Medical Education, sponsored by the California Medical Association and directed by its Committee on Continuing Medical Education, San Diego, 11 to 12 March 1967. The general subject was divided among four workshops and the reports of two of them—No. 1, (a) and (b), and No. 2—were printed in the August issue of California Medicine. The Reports herewith are those of the other two workshops.The conference was supported in part by Contract No. PH 108-67-158, Bureau of Health Manpower, Public Health Service, Department of Health, Education and Welfare.     

An Italian Survey of Compliance With Major Guidelines for L-Thyroxine of Primary Hypothyroidism

Objective: The adherence by endocrinologists to guideline regarding levothyroxine (LT4) therapy and the compliance of patients may impact the management of hypothyroidism. The aim of this study was to compare the adherence of Italian endocrinologists to the ATA/AACE and ETA guidelines on the management of newly diagnosed primary hypothyroidism and to validate the Italian version of the Morisky-Green Medical Adherence Scale-8 (MMAS-8) questionnaire as applied to the evaluation of the adherence of patients with hypothyroidism to LT4 treatment.Methods: This was an observational, longitudinal, multicenter, cohort study, involving 12 Italian Units of Endocrinology.Results: The study enrolled 1,039 consecutive outpatients (mean age 48 years; 855 women, 184 men). The concordance of Italian endocrinologists with American Association of Clinical Endocrinologists/American Thyroid Association (AACE/ATA) and European Thyroid Association (ETA) recommendations was comparable (77.1% and 71.7%) and increased (86.7 and 88.6%) after the recommendations on LT4 dose were excluded, considering only the remaining recommendations on diagnosis, therapy, and follow-up. The MMAS-8 was filled out by 293 patients. The mean score was 6.71 with 23.9% low (score <6), 38.6% medium (6 to <8), 37.5% highly (= 8) adherers; the internal validation coefficient was 0.613. Highly adherent patients were not more likely to have good control of hypothyroidism compared with either medium (69% versus 72%, P = .878) or low (69% versus 43%, P = .861) adherers.Conclusion: Clinical management of hypothyroidism in Italy demonstrated an observance of international guidelines by Italian endocrinologists. Validation of the Italian version of the MMAS-8 questionnaire provides clinicians with a reliable and simple tool for assessing the adherence of patients to LT4 treatment.Abbreviations: AACE = American Association of Clinical Endocrinologists; ATA = American Thyroid Association; EDIPO = Endotrial SIE: DIagnosis and clinical management of Primitive hypothyrOidism in Italy; eCRF = electronic case report form; ETA = European Thyroid Association; fT3 = free triiodothyronine; fT4 = free thyroxine; LT4 = levothyroxine; MMAS-8 = Morisky-Green Medical Adherence Scale-8; PH = primary hypothyroidism; T3 = triiodothyronine; T4 = thyroxine; TSH = thyroid-stimulating hormone; US = ultrasonography     

Biogenesis of plasma lipoproteins in rat hepatoma McA-RH7777: Importance of diffusion-mediated events during cell growth

Summary Cultured McA-RH7777 rat hepatoma cells actively synthesize and secrete plasma lipoproteins. However, synthesis of [¹⁴C]triglyceride declines monotonically throughout the early growth period and remains low in postconfluent cultures; and net secretion of [¹⁴C]triglyceride is 10-fold more efficient in logarithmically growing cultures than in postconfluent cultures. Secretion of apolipoproteins associated with very low density and low density lipoproteins is selectively reduced in postconfluent cultures. The temporal reductions in [¹⁴C]triglyceride production are related more strongly to increasing cell concentration (cells/cm³ medium) than to increasing cell density (cells/cm² growth surface). We have allowed cells to grow either retained within small circular corrals or unrestricted in culture dishes. When seeded at equal density (10⁴ cells/cm²) but at one-fifth the cell concentration, corralled cells synthesize twice as much [¹⁴C]triglyceride per cell after 2 and 4 d, and are 10 times as efficient in [¹⁴C]triglyceride secretion by 6 d of growth, as noncorralled cells. When seeded at equal cell concentration (10⁵ cells/dish) but at 5 times the cell density, corralled cells are only 20% less efficient at [¹⁴C]triglyceride synthesis and secretion than noncorralled cells. Conditioned medium depresses synthesis and secretion efficiency of [¹⁴C]triglyceride. Orotic acid exposure also inhibits synthesis of [¹⁴C]triglyceride and secretion of certain [³⁵S]apolipoproteins in early cultures, but it has no significant effect on late cultures. We conclude that diffusion-mediated events are important regulators of triglyceride and apolipoprotein production in growing rat hepatoma cells, but that events associated with formation of cell-to-cell contacts play a minor role in regulation of plasma lipoprotein biogenesis. This research was supported by grants to R. H. from the American Heart Association (85-805), from NHLBI (15062, Specialized Center of Research in Atherosclerosis), and from the Louis Block Fund. S. T. was supported as an American Heart Association Medical Student Research Fellow. S. T. and H. S. are recipients of predoctoral training grants from the National Institutes of Health, Bethesda, MD (HL 723711, HD 07009). This work has been presented elsewhere in preliminary form (14, 48, 49).     

Viruses with fungal vectors

Book Review

Viruses with fungal vectorsJ.I. Cooper and M.J.C. Asher (Eds.), (AAB Developments in Applied Biology 2). Warwick: Association of Applied Biologists, 1988. vi+335 pages. £30.00. ISBN 0269-0713     

Role of microtubules in the intracellular transport of growth hormone

Summary Pulse-chase experiments utilising(³H)leucine have been used to study the effects of colchicine and vinblastine on intracellular transport and secretion of newly synthesised growth hormone from rat anterior pituitary fragments. Growth hormone was isolated from medium and fragments by polyacrylamide gel electrophoresis. When colchicine or vinblastine, which disrupt microtubules, were added immediately after pulse labelling, inhibition of the subsequent secretion of newly synthesised growth hormone was detected throughout the succeeding 5 h. Similar inhibition was seen if the drugs were added after a 1 h delay. However, if colchicine or vinblastine were added only after a 2 h chase incubation, then no significant effect on subsequent release of labelled growth hormone was seen. The results suggest that these agents may inhibit the transport of newly formed growth hormone storage granules from the Golgi complex to the cytoplasmic pool. Microtubules do not appear to be involved in the mechanism of the final secretion of newly synthesised hormone by exocytosis.These studies were supported by grants from the Medical Research Council and British Diabetic Association     

Comparison of glucokinase in C3H/He and C58 mice that differ in their hepatic activity

Partially purified preparations of the hepatic glucokinase from C3H/He and C58 inbred mice have been used to explore the molecular basis for the observed twofold difference in activity between the strains. The single codominant gene that appears to regulate activity, the alleles of which are designated Gk^a and Gk^b, respectively, for the two strains, could represent a structural gene change. This now seems unlikely because the mouse enzyme, although showing small differences from rat glucokinase, appeared to be identical in the two strains with respect to thermal stability, electrophoretic mobility in agarose gels, and kinetic properties such as the apparent K _m values for MgATP^2– and glucose and the unique cooperative interaction with the latter substrate. The enzymes also reacted identically in a range of immunological tests (double-diffusion, immunoelectrophoresis, immune precipitation and immune inhibition assays) and ELISA immune inhibition assays indicated that the twofold difference in activity was due to a similar difference in antigenically active enzyme. Genetic control over the physiologically significant regulation of enzyme amount is therefore probable.This work has been supported in part by a grant from the British Diabetic Association and a Training Studentship to PAJ from the Medical Research Council (U.K.).     

Multiple effects of protein kinase C activators on Na+ currents in mouse neuroblastoma cells

The effects of externally applied different protein kinase C (PKC) activators on Na⁺ currents in mouse neuroblastoma cells were studied using the perforated-patch (nystatin-based) whole cell voltage clamp technique. Two diacylglycerol-like compounds, OAG (1-oleoyl-2-acetyl-sn-glycerol), and DOG (1-2-dioctanoyl-rac-glycerol) attenuated Na⁺ currents without affecting the time course of activation or inactivation. The reduction in Na⁺ current amplitude caused by OAG or DOG was dependent on membrane potential, being more intense at positive voltages. The steady-state activation curve was also unaffected by these substances. However, both OAG and DOG shifted the steady-state inactivation curve of Na⁺ currents to more hyperpolarized voltages. Surprisingly, phorbol esters did not affect Na⁺ currents. Cis-unsaturated fatty acids (linoleic, linolenic, and arachidonic) attenuated Na⁺ currents without modifying the steady-state activation. As with DOG and OAG, cis-unsaturated fatty acids also shifted the steady-state inactivation curve to more negative voltages. Interestingly, inward currents were more effectively attenuated by cis-fatty acids than outward currents. Oleic acid, also a cis-unsaturated fatty acid, enhanced Na⁺ currents. This enhancement was not accompanied by changes in kinetic or steady-state properties of currents. Enhancement of Na⁺ currents caused by oleate was voltage dependent, being stronger at negative voltages. The inhibitory or stimulatory effects caused by all PKC activators on Na⁺ currents were completely prevented by pretreating cells with PKC inhibitors (calphostin C, H7, staurosporine or polymyxin B). By themselves, PKC inhibitors did not affect membrane currents. Trans-unsaturated or saturated fatty acids, which do not activate PKC's, did not modify Na⁺ currents. Taken together, the experimental results suggest that PKC activation modulates the behavior of Na⁺ channels by at least three distinct mechanisms. Because qualitatively different results were obtained with different PKC activators, it is not clear how Na⁺ currents would respond to activation of PKC under physiological conditions.This work was supported in part by a grant-in-aid from the American Heart Association (National Center), and by Loyola University Medical Center. Dr. Godoy is a recipient of a fellowship from Conselho Nacional de Pesquisas e Desenvolvimento (Brazil).     

Mitochondrial malic enzyme in mosaic skeletal muscle of mouse chimeras

The question was investigated whether mitochondria in the mammalian skeletal muscle fiber syncytium incorporate gene products encoded by one or many nuclei. Mouse chimeras were produced from strains which differ in their electrophoretic variants of the nuclear-coded mitochondrial protein, malic enzyme (MOD-2, E.C. 1.1.1.40, l-malate NADP⁺ oxidoreductase decarboxylating). The MOD-2 phenotypes of skeletal muscles of these chimeras were characterized in a starch gel electrophoretic system. The results indicate that individual mitochondria can contain products encoded by multiple nuclei and therefore that, for skeletal muscle mitochondria, the cell is not subdivided into nuclear territories. Possible mechanisms of gene product distribution in skeletal muscle fibers are discussed.This work was supported by Grants MT-1940 (K. B. F.) and MA-6411 (A. C. P.) from the Medical Research Council of Canada, and by the Muscular Dystrophy Association of Canada (A. C. P. and P. M. F.).