Reliability and validity of the COOP/WONCA health status measure in patients with chronic obstructive pulmonary disease.

The objective of the study was to assess the reliability and validity of the Dartmouth Primary Care Cooperative Information Project/World Organization of National Colleges, Academies, and Academic Associations of General Practice/Family Physicians (COOP/WONCA) questionnaire in outpatients with chronic obstructive pulmonary disease (COPD). The test-retest reliability of individual items of the COOP/WONCA questionnaire was assessed using a weighted kappa-statistic, and construct validity was assessed by correlating items of the COOP/WONCA with the EQ-5D health status measure. Discriminant validity was assessed by comparing scores for known groups, at the same time comparing the results with those of a lung-specific health status questionnaire. The individual items of the COOP/WONCA had test-retest reliabilities of 0.67-0.78 (weighted kappa). Spearman's rank correlations between COOP/WONCA single-item scores and corresponding EQ-5D ranged 0.45-0.72, which were generally higher than associations between non-corresponding items. Four of the five COOP/WONCA items did not discriminate between patient groups divided according to forced expiratory flow in 1 sec (FEV1) in percent of predicted and 6-min walking distance, while four of five items of the lung-specific questionnaire discriminated well between these groups. The COOP/WONCA chart system was reliable and showed properties supporting the construct validity of the measure. The items, however, did not discriminate well between known groups, indicating that this questionnaire is not very sensitive in patients with COPD. The reliability of the COOP/WONCA items was acceptable for use at group level, but lower than current recommendations for use in individual patients.

Lipid accumulation in the perfused rat heart after isoproterenol administration.

Isoproterenol is a potent lipolytic agent. It has also been shown to induce accumulation of lipid droplets in the myocardium in vivo. The isolated working rat heart model was applied to evaluate the effect in vitro. Twenty-four rats were randomly separated into four experimental groups of six rats, all receiving Krebs-Hensleit buffer with 11mM glucose. The control group had no supplement, the other groups received either 10(-5) M isoproterenol, 1.2 mM palmitate, or a combination of the two. Tissue levels of triglycerides and phospholipids, and the fractional volume of lipid droplets were determined at the end of the 30 min perfusion in the working heart mode. Ventricular function was monitored continuously and showed a marked increase in the isoproterenol treated groups. The level of tissue phospholipids remained unchanged in all groups. Whereas tissue triglycerides were significantly increased in the group receiving only palmitate as compared to the other groups. A corresponding increase was, however, not found to be significant for lipid droplet quantitation. The results are discussed in terms of possible mechanisms for isoproterenol-induced accumulation of neutral lipids in the myocardium.

Effects of verapamil on intracellular lipid accumulation in cat hearts with 3 h of regional-ischaemia.

In the regionally ischaemic heart lipid droplet accumulation is found in the ischaemic area and is most pronounced in the periphery. The purpose of the present study is to explore the potential effects of the calcium-channel-blocker verapamil on this accumulation. The drug is known to reduce the intensity of myocardial ischaemic injury. The myocardial ultrastructure was studied in anaesthetized open chest cats with 3 h of coronary artery occlusion. Biopsies were taken from the ischaemic, border and normally perfused myocardium defined in vivo injections of fluorescein and verified by blood flow measurements using radiolabelled microspheres. Arterial concentration of non esterified fatty acids (NEFA) was measured during the ischaemic period. A higher accumulation of lipid droplets was found in the central ischaemic myocardium of verapamil-treated cats than in control animals (p less than 0.05). The normally perfused and borderline areas were unaffected by verapamil as far as lipid accumulation was concerned and showed the same pattern as in the untreated group. The increased accumulation of lipid droplets in the ischaemic myocardium, after treatment with verapamil, may reflect a preserved metabolic activity in the ischaemic tissue or result from a higher supply of fatty acids due to increased perfusion of the central ischaemic tissue.

Oxfenicine-induced accumulation of lipid in the rat myocardium.

Oxfenicine inhibits myocardial metabolism of nonesterified fatty acids (NEFA). The purpose of the present study was to examine the effects of oxfenicine on triglyceride accumulation and the development of histologically visible lipid droplets. The beta-agonist isoproterenol was used to induce elevated arterial NEFA. Four groups of rats were used in the experiment (12 to 14 rats in each group), and each group received two subcutaneous injections, the second injection 25 min after the first, of oxfenicine-isoproterenol, oxfenicine-saline, saline-isoproterenol and saline twice, respectively. One hour after the second injection, the rats were anesthetized, and the hearts from six rats from each group were quickly removed and frozen for later analysis of triglyceride content. From the remaining rats blood samples were drawn for NEFA analysis, and biopsies were taken from the left ventricular wall before the hearts were frozen in liquid nitrogen and prepared for analysis of esters of carnitine and CoA. Quantitative morphometric techniques were used to determine the fractional volume of lipid droplets in myocardial biopsies. Our results show a marked increase in the triglyceride and lipid droplet content in all groups receiving oxfenicine or isoproterenol. The effect was most pronounced after treatment with both drugs. The close association between the increase in triglyceride and lipid droplet supports the notion that the lipid droplets are composed of triglycerides. Our finding that oxfenicine induces lipid droplet accumulation independent of NEFA increase supports the hypothesis that oxfenicine exerts its effect by inhibiting carnitine acyl transferase.

Effects of verapamil and timolol on cellular morphometric changes in cat hearts with regional ischaemia.

In twenty-one anaesthetized open chest cats the left anterior descending coronary artery (LAD) was occluded for three hours. Seven cats were pretreated with a bolus injection of Verapamil, followed by a continuous infusion of Verapamil during the ischaemic period. Seven cats were pretreated with a bolus injection of Timolol to a heart rate reduction of 20 beats/min or more and seven cats were given saline. In the latter two groups the cats received a continuous infusion of saline during the period of coronary occlusion. Biopsies were taken from the mid-myocardium of the normal, border and ischaemic zones, as defined by fluorescein staining, and verified by blood flow measurements with radiolabelled microspheres. Standard point counting techniques were used for calculations of fractional volumes of mitochondria, cytoplasm and myofibrils as well as of mitochondrial surface density and surface to volume ratio. We observed a cytoplasmic oedema in the border and ischaemic zones, that was not altered by medical treatment. In the border zone of the control cats there is greater mitochondrial swelling than in the ischaemic zone. This particular swelling is not seen in the treatment groups. However, in the normal and border zones of the verapamil group the mitochondria are smaller when compared with the respective zones in the two other groups, but increases relatively more in size in the border and ischaemic zones. Furthermore, we measured the water content, sarcomere length and per cent heavily damaged cells. These variables were not altered by Verapamil or Timolol in any zone when compared with the respective zones in the control group.

Myocardial cAMP and calcium levels in the calcium paradox.

In a graded model (minimal, subtotal, total) of the calcium paradox phenomenon induced by a progressive increase in the flow rate and volume (5 ml, 10 ml, 45 ml) of calcium-free perfusion (5 min) the severity of tissue injury on calcium repletion was related to a potential elevation of cAMP during calcium depletion. In the subtotal and total models of the calcium paradox a 50% increase was found for tissue cAMP after calcium-free perfusion, but no such rise could be associated with the minimal calcium paradox. In the study tissue injury, as assessed by whole tissue and mitochondrial accumulation of calcium and by myocardial leakage of creatine kinase, could only partly be related to cAMP changes. It is concluded that calcium-free coronary perfusion induces a complex series of events favouring excessive calcium entry, only one of which may be related to a change in cAMP.

Cellular lipid accumulation in different regions of myocardial infarcts in cats during beta adrenergic blockade with timolol.

The effects of non-selective beta adrenergic blockade on intracellular lipid accumulation in hearts with acute ischaemia were studied by electron microscopy of myocardial biopsy specimens using quantitative stereological techniques. Pentobarbital anaesthetised cats with coronary ligation were divided into eight controls and eight cats treated with timolol intravenously just before ligation. Biopsy specimens were collected from ischaemic, borderline, and normally perfused myocardium, defined by an in vivo injection of fluorescein and verified by regional myocardial blood flow measurements with 15 microns radiolabelled microspheres. During a 3 h occlusion period timolol treated cats had a lower heart rate, left ventricular dP/dt, and plasma free fatty acid concentration. In control cats the cytosolic volume fraction of lipids was 0.71 X 10(-3) in non-ischaemic myocardium, 2.63 X 10(-3) in central ischaemic tissue, and 6.53 X 10(-3) in borderline tissue. Timolol reduced the appreciable lipid accumulation in borderline tissue by 24% (to 4.97 X 10(-3)) compared with controls, whereas accumulation in central ischaemic tissue was not affected. Thus timolol diminished lipid accumulation in borderline myocardial tissue. The mechanism is most likely related to reduced ischaemic intensity and better preserved metabolic function.

Lipid accumulation in the myocardium during acute regional ischaemia in cats.

Accumulation of lipid material in the myocardium was studied in cat hearts with acute regional ischaemia of 3 h duration. The fractional volume of lipid droplets in cytosol was analysed by electron microscopy of myocardial biopsies using a quantitative stereologic technique. Ischaemic and normally perfused myocardium were identified by fluoresceine injection, and tissue blood flow measurements were performed with labelled microspheres. In normal myocardium only small amounts of lipid droplets were found. A marked accumulation of lipid droplets occurred in borderline tissue between the two types of myocardium, whereas lipid accumulation in ischaemic myocardium was less pronounced. The arterial concentration of nonesterified fatty acids was clearly increased during the 3 h coronary artery occlusion period. Increased triglyceride synthesis from arterial fatty acids, or redistribution of intracellular lipids, are suggested as possible explanations for lipid accumulation during acute myocardial ischaemia.

Propranolol induced lipid accumulation and mitochondrial granularity in myocardial cells.

The ultrastructure of the left ventricular wall was studied in mice given intraperitoneal injections of propranolol (5 mg or 20 mg.kg-1 b.w. twice a day for 48 h with an additional dose 3 h before death), and in saline injected controls. Injections of propranolol increase the number of lipid droplets in areas of the myocardium. Fractional volume of lipid droplets in myocardial cells determined using quantitative stereological techniques, showed an increase of 280% and 544% in the group receiving the lowest and highest doses respectively. Injections of propranolol were also followed by an increased granularity of the mitochondria.

Protective effects of verapamil against isoprenaline- induced mobilization of mitochondrial calcium and cellular lipid droplets in the myocardium.

Calcium level and lipid droplets accumulation is studied in the mouse myocardium of animals receiving a single injection of isoprenaline (ISP), of animals receiving ISP after 4-8 days of pretreatment with verapamil (Isoptin), and, of control animals. The animals are sacrificed 4h after the ISP-injection. Calcium is determined in isolated mitochondria and in whole tissue homogenates by atomic absorption spectrometry. Lipid droplets accumulation is analysed by quantitative stereological technique. Mitochondrial calcium is raised by 65% and calcium of whole tissue homogenates by 21% following ISP stimulation. At the same time, the myocardial fractional volume occupied by lipid droplets in the ISP treated animals increases to 14 times that of control. The ISP stimulated elevation of mitochondrial calcium is reduced by 73% and that of lipid fractional volume by 71% after pretreatment with verapamil. No similar reduction of calcium content is observed in the whole tissue homogenates. These results are discussed in terms of the role of Ca2+ in ISP induced myocardial necrosis.

Effect of isoproterenol on lipid accumulation in myocardial cells.

Plasma free fatty acid (FFA) levels and myocardial accumulation of lipid droplets were studied in mice receiving 85 mg isoproterenol (ISP)/kg b.wt. with and without pretreatment with nicotinic acid (NA) 30 min before ISP injection, and in saline-injected controls. The animals were killed 30 min after the ISP injection. Plasma FFA were determined by a radiochemical method. Lipid droplet accumulation was analyzed by quantitative stereologic techniques. Plasma FFA were raised from 505.8 +/- 15.7 to 1,808.8 +/- 87.8 mumol/l after ISP treatment and to 1,108.0 +/- 34.7 mumol/l in the group receiving NA pretreatment. However, NA pretreatment did not bring about a corresponding decrease in the fractional volume of lipid droplets, as both groups showed an increase of five times that of the controls. These results are discussed in terms of the pathogenesis of ISP-induced lipid accumulation.

Calcium and magnesium levels in isolated cardiac mitochondria from mice injected with isoproterenol.

Calcium (Ca) and Magnesium (Mg) are determined by atomic absorption flame spectrometry in isolated cardiac mitochondria from mice receiving subcutaneous injections of DL-isoproterenol HCl (ISO), and in mitochondria of untreated controls. In the controls, mitochondria were isolated in the presence or absence of ruthenium red. On the absence of ruthenium red in the isolation medium, mitochondrial Ca levels increase by about 300%, while levels of Mg remain unchanged. Focal myocardial necrosis following a single ISO-injection is shown by electron microscopy. Ca and Mg levels are largely unaffected by a single dose of ISO until 24 h after the injection. A slight increase in Ca occurs in the 48 h samples. When multiple injections of ISO are given every 12th hour of 48 h, 72 h and 96 h, respectively, endogenous Ca and Mg increase significantly. It is suggested that this increase might be associated with ISO-induced cardiac hypertrophy rather than with the pharmacological effects of ISO per se.

Accumulation of myocardial lipid droplets in dexamethasone-treated mice.

Conventionally fixed and plastic-embedded myocardial tissue from mice treated with 1 mg/kg body weight dexamethasone for 48 h was examined in the electron microscope. The dexamethasone-treated mice showed a marked increase in the accumulation of lipid droplets, as compared with control animals. The percent volume of lipid droplets, calculated by morphometric analysis, showed a significant increase in the dexamethasone-treated mice. No other ultrastructural difference between dexamethasone-treated mice and controls was observed.

Effects of isoproterenol on the dense core and perigranular membrane of atrial specific granules.

Following subcutaneous injections of isoproterenol hydrochloride (ISO), atrial cells present a large number of partly degranulated or completely clear "specific granules" enclosed by an intact membrane. Such profiles were never encountered in normal controls and might suggest ISO-induced release of a secretory product. Permeability of perigranular membrane was tested using the extracellular macromolecular tracer horseradish peroxidase (HRP). Reaction product was entirely absent within granules of atrial cells in which the sarcolemma was made permeable to HRP molecules by the ISO injections. This seemed to be the case even in heavily labelled cells in which the peroxidase had penetrated the mitochondrial membranes. In atrial cells impermeable to the tracer, the specific granules closely apposed to the sarcolemma were always HRP-negative. The release mechanism of a possible secretory substance from the specific granules is discussed.