Tracking seasonal changes in diversity of pollen allergen exposure: Targeted metabarcoding of a subtropical aerobiome.

The importance of grass pollen to the global burden of allergic respiratory disease is well established but exposure to subtropical and temperate pollens is difficult to discern. Current monitoring of airborne pollen relies on light microscopy, limiting identification of taxa to family level. This informs seasonal fluctuations in pollen aerobiology but restricts analysis of aerobiological composition. We aimed to test the utility of DNA metabarcoding to identify specific taxa contributing to the aerobiome of environmental air samples, using routine pollen and spore monitoring equipment, as well as assess temporal variation of Poaceae pollen across an entire season. Airborne pollen concentrations were determined by light microscopy over two pollen seasons in the subtropical city of Brisbane (27°32'S, 153°00E), Australia. Thirty daily pollen samples were subjected to high throughput sequencing of the plastid rbcL amplicon. Amplicons corresponded to plants observed in the local biogeographical region with up to 3238 different operational taxonomic units (OTU) detected. The aerobiome sequencing data frequently identified pollen to genus levels with significant quantitative differences in aerobiome diversity between the months and seasons detected. Moreover, multiple peaks of Chloridoideae and Panicoideae pollen were evident over the collection period confirming these grasses as the dominant Poaceae pollen source across the season. Targeted high throughput sequencing of routinely collected airborne pollen samples appears to offer utility to track temporal changes in the aerobiome and shifts in pollen exposure. Precise identification of the composition and temporal distributions of airborne pollen is important for tracking biodiversity and for management of allergic respiratory disease.

Features of the angiographic evaluation of the INTACT study. International Nifedipine Trial on Antiatherosclerotic Therapy.

INTACT (International Nifedipine Trial on Antiatherosclerotic Therapy) is a prospective, placebo-controlled, randomized, double-blind, multicenter trial analyzing the influence of 80 mg nifedipine/day on the angiographic progression of early stage coronary atherosclerosis. Coronary angiograms were taken in identical projections before and after a treatment period of 3 years. Quantitative analysis of the angiograms was performed with the computer-assisted contour detection system CAAS. For definition purposes, the coronary artery system was subdivided into 25 different segments, including all anatomic variants. Measurement parameters of segments were mean and minimal diameter, and of stenoses minimal diameter, percentage diameter reduction (at least 20%), length, and plaque area. The variable extent of the changes of these parameters in the different projections analyzed per patient in the two study angiograms was considered by separate computation of the maximal, mean, and minimal changes over these projections; the comparison of the parameter changes between the two treatment groups was performed separately according to these three modes. For all parameters, this comparison was performed on the basis of the individual 25 segments, as well as after aggregation of individual segments to arteries (RCA, LAD, and LCX), to groups of large and small segments, and to the entire coronary artery system. Assessment of changes of the coronary (patho)morphology by quantitative analysis of coronary angiograms is associated with a number of methodical limitations, which may lead to a certain variability of the results. However, due to the double-blind feature of INTACT, this variability should be comparable in the two groups of this study, allowing for a conclusive comparison.

Retardation of angiographic progression of coronary artery disease by nifedipine. Results of the International Nifedipine Trial on Antiatherosclerotic Therapy (INTACT). INTACT Group Investigators.

425 patients showing mild coronary artery disease (CAD) on arteriography were enrolled in a multicentre trial and randomised to treatment with nifedipine (80 mg/day) or placebo. The two groups were well matched for age, sex, and risk factors. 348 patients (82%) underwent repeat arteriography 3 years later; 282 (134 nifedipine, 148 placebo) had received treatment throughout, but treatment had been stopped in 39 nifedipine-treated and 27 placebo-treated patients after average periods of 354 and 467 days. Computer-assisted measurements of arteriograms from all restudied patients (175 placebo, 173 nifedipine) showed no significant differences in the number or severity of lesions on initial arteriograms, or in the progression or regression of existing lesions over 3 years. In contrast, the number of new lesions per patient was significantly lower in the nifedipine group than in the placebo group (0.59 vs 0.82 lesions per patient, a 28% reduction). Thus in patients with mild CAD nifedipine substantially suppresses disease progression as shown by the appearance of new lesions detectable by quantitative coronary arteriography.

International nifedipine trial on anti-atherosclerotic therapy (INTACT)--methodologic implications and results of a coronary angiographic follow-up study using computer-assisted film analysis.

Animal experiments demonstrated a significant suppressive effect of various calcium channel blockers on the formation of atherosclerotic lesions. Therefore, a prospective, placebo-controlled, randomized, double blind multicenter study was performed to investigate the inhibitory influence of the calcium channel blocker nifedipine (80 mg/day) on the progression of coronary artery disease in man. Study endpoints were changes of coronary morphology documented by coronary angiography with particular respect to the formation of new coronary stenoses. In 348 out of 425 patients included in the study, coronary angiograms were repeated after three years. The angiograms were standardized by induction of a maximal coronary vasodilation with high doses of nitrates and by using absolutely identical angiographic projections. Quantitative analysis of coronary cineangiograms was performed with the computer-assisted contour detection system CAAS. Parameters were mean and minimal diameter of all segments and minimal stenosis diameter, percent diameter stenosis, length and plaque area of all stenoses. Continuous intake of study medication was registered in 282 patients, 134 on nifedipine and 148 patients on placebo. In these patients, a total of 3808 coronary segments with 893 stenoses (greater than or equal to 20% diameter reduction in at least one angiographic projection) were compared on the baseline and follow-up cineangiograms. The changes in all angiographic parameters analyzed averaged over all patients by considering all angiographic projections analyzed, indicated significant progression of the disease (p less than 0.006). The average changes in all parameters were even about three times more profound, when in the individual patients only the respective projections indicating the maximal changes were considered for the calculation (p less than 0.001). However, with neither of these two analysis modes, the differences in progression between the treatment groups were statistically significant. In the follow-up angiograms, a total of 196 new coronary lesions (185 stenoses, 11 occlusions) were found at previously normal arterial sites. In patients on nifedipine, an average of only 0.58 new lesions per patient were detected versus 0.80 lesions per patient on placebo (-27%; p = 0.031). INTACT is the first prospective angiographic trial on the progression of coronary artery disease using computer-assisted quantitative coronary angiography in such a high number of patients. All parameters analyzed indicated significant progression of coronary artery sclerosis. Nifedipine had no influence on the progression of preexisting coronary stenoses, but inhibited significantly the formation of new angiographically recognizable lesions. Further prospective coronary angiographic trials with calcium channel blockers using a comparably exact method are needed to confirm the results of this study.

[Computer-assisted geometric measuring technic in coronary angiography interval studies: results of initial angiograms of the International Nifedipine Trial of Anti-atherosclerotic Therapy (INTACT) study]. / Computergestützte geometrische Messtechnik in koronarangiographischen Intervallstudien: Ergebnisse bei Erstangiogrammen der INTACT-Studie.

In 396 of 423 (93.6%) patients with mild to moderate coronary artery disease participating in a coronary angiographic follow-up trial, diameters of the epicardial coronary arteries were measured with an automatic contour detection system (CAAS). The underlying INTACT study (International Nifedipine Trial on Antiatherosclerotic Therapy), a prospective, placebo-controlled, randomized, double-blind multicenter trial, investigates the influence of the calcium antagonist nifedipine (80 mg/day) on the progression of coronary atherosclerosis over a three-year interval. The study is based on coronary angiograms repeated in identical projections after premedication with 10 mg isosorbide dinitrate sublingually. For quantitative analysis the coronary artery system was, in consideration of all anatomical variations, subdivided into 25 segments. In the first angiograms of the 396 patients evaluated up to April 1, 1988, altogether 5,425 different coronary segments could be analyzed over their entire length in one or more angiographic projections--on the average 13.7 +/- 2.8 segments per patient. Analysis parameters were the mean diameters of the entire segments and of the individual subsegments (about 5 mm in length). The 10 major proximal segments could be evaluated in 76-94% of patients respectively in more than two different angiographic projections on the average; in 0-13% of patients the respective segments were occluded. In 1-4% of patients the evaluation of these segments was prevented by poor film quality and in 1-16% of patients prevented by anatomical abnormalities (e.g., segment too short or too small). 184 segments were found occluded (RCA 42%, LAD 30%, CX 28%) and 909 mostly low-grade to moderate stenoses (RCA 34%, CX 32%, LAD 30%, left main 4%) were analyzed with a special algorithm. The following obstruction parameters were derived: minimal diameter, percentage severity, length, and plaque area. The present data demonstrate that in an angiographical multicenter follow-up study such as INTACT a nearly complete quantitative morphometric analysis of the visualized coronary artery system can indeed be obtained in virtually all angiograms when a computer-assisted contour detection system is applied.

Clinical application of quantitative coronary angiography using the CAAS system: preliminary results of the INTACT study (International Nifedipine Trial on Antiatherosclerotic Therapy).

It is the objective of the INTACT-study to test in man, whether a significant retardation of the progression of coronary artery disease is attainable with the Ca-antagonist nifedipine; this may be possible on the basis of numerous animal experiments. INTACT is a prospective, randomized, double blind, placebo controlled investigation in 423 patients with preferably early stages of coronary sclerosis in whom a progression of the disease seems likely. A proper coronary angiogram led to inclusion of the patients in the study (October 1983-June 1985). Over the following 3-years-period patients received either nifedipine 80 mg/day or placebo. The study is concluded by a control coronary angiogram with angiographic projections which are identical to those of the first coronary angiography. The extent of coronary sclerosis is objectivated by computer-assisted quantitative measurement of the entire coronary arterial system with the CAAS-system (Rotterdam). For definition purposes the coronary artery system subdivided into 25 segments. Parameters for progression assessment will be mean segment diameter, minimal obstruction diameter, percentage severity of obstruction, length of obstruction and plaque area. So far 4826 coronary segments have been analyzed from the first angiograms of 383 patients. Per patient an average of 12.6 different segments could be evaluated in at least one angiographic projection. The major coronary segments could be measured in 72-93% of the patients in one or more angiographic projections (at the average about 2 different projections). Five hundred and forty-six coronary obstructions were analyzed; 131 of these were total occlusions. Only 9% of the length of the vessel contours detected by the computer algorithm required manual correction by the operators, suggesting a high reliability of the system. It can be concluded that quantitative measurement of the complete coronary artery system can indeed be obtained in a large angiographical multicenter study such as INTACT.

Comparison of mackerel-oil and lard-fat enriched diets on plasma lipids, cardiac membrane phospholipids, cardiovascular performance, and morphology in young pigs.

Purified mackerel-oil extract or lard fat (9.1% wt/wt) was added to a basal diet of young pigs for 8 wk. Effects on plasma lipids, glucose and insulin, cardiac membrane phospholipids, cardiovascular performance, and morphology were studied. A time-dependent reduction of plasma triglyceride (62%), total cholesterol (41%), and HDL cholesterol (47%) was found in the mackerel-oil-fed pigs. The postprandial glucose and insulin response may indicate a mackerel-oil-induced resistance of insulin receptors. Although the polyunsaturated fatty acid composition of cardiac sarcolemma widely differed between the two groups, all determined indices of heart function were equal. It is concluded that consumption of a fish-oil diet with a low content of monoenes and supplemented with antioxidants reduces plasma lipid levels without producing pathological side effects.

On a possible role for calcium antagonists in atherosclerosis. A personal view.

An overview is provided of current thinking regarding the origin(s) of atherosclerosis on the basis of clinical evidence, experimental models and pharmacological investigations with calcium antagonists. Because this group of drugs inhibits the influx of Ca2+ into the smooth muscle cell of the cardiovascular wall, it has been argued that when endothelial damage is anticipated or has occurred, the 'protective' action of calcium antagonists might avoid, delay or even help to repair the calcium overload seen with cell membrane damage. In the real world the situation must be far more complex as will become evident but as a concept the avoidance of Ca2+ overloading is enticing. In fact, based on circumstantial evidence, a major clinical trial is currently in progress in which the long term effect of nifedipine on coronary atherosclerosis is being evaluated in a double blind randomized fashion. The end-point of this study lies in the reduction of the severity and number of coronary artery lesions. Although not yet completed, the design and goals of that study are provided.

The influence of fish oil diet and norepinephrine treatment on fatty acid composition of rat heart phospholipids and the positional fatty acid distribution in phosphatidylethanolamine.

The effect of chronic norepinephrine (NE) administration with increasing dosage from 1-4 mg/kg over a period of 2 weeks was studied on cardiac phospholipids and their fatty acid distribution in rats. Animals were fed a control diet or a 10% cod liver oil (CLO)-enriched diet. The relative distribution of various polyunsaturated fatty acids esterified to the 1- and 2-position of the phosphatidylethanolamine fraction was estimated. NE stress during control feeding significantly reduced the total phospholipid content in rat heart. No differences in the phospholipid class distribution were found. However, CLO feeding as well as chronic NE administration resulted in a decrease of omega 6 fatty acids, mainly C 18:2 omega 6 and C 20:4 omega 6, which was compensated with an increase in omega 3 fatty acids, mainly C 20:5 omega 3 and C 22:6 omega 3. The changes in fatty acid composition qualitatively agree with those reported by Gudbjarnason et al. (23), except that the mortality in our NE-treated control or CLO-fed groups was considerably lower. It can probably be attributed to a different mode of NE administration. On the other hand, at the end of the CLO feeding period in rats treated with NE or not, comparing with control fed rats without NE treatment, the incidence rate of ST segment elevation in electrocardiogram (ECG) recorded under light diethylether-induced anesthesia was higher. Independent of whether the fatty acid composition of myocardial phospholipids was dietary or pharmacologically manipulated, most of the polyunsaturated fatty acids were found at the 2-position of the phosphatidylethanolamine molecules. The polyunsaturated fatty acids account for 45-50% of the fatty acyl residues and preferentially occupy the 2-position, where they can exchange for each other.

Effect of long-term oral nifedipine therapy on left ventricular regional wall function at rest and during supine bicycle exercise.

15 patients, 1 to 3 year after coronary bypass surgery, underwent symptom limited supine bicycle exercise tests without nifedipine and after acute and chronic (3 months) administration of the drug. Haemodynamic variables were monitored as was epicardial marker motion, using biplane cineradiography during exercise, the markers having been implanted at the time of surgery. We found significant (P less than 0.001) reductions in end-diastolic and end-systolic regional dimensions at maximal exercise after oral nifedipine, associated with a significant reduction in exertional angina, which persisted during long-term treatment. No adverse effects of the drug were observed.

Treatment of unstable angina with emphasis on calcium antagonists.

From the literature which has appeared over the last decade a selection was made of the three major calcium antagonists: nifedipine, verapamil and diltiazem. In the clinical situation, the net hemodynamic and electrophysiological effect of these drugs is the result of complex interactions between their peripheral and their central effects. The degree of baroceptor stimulation and reflex mediated beta-adrenergic activity, which counteracts and influences the intrinsic negative dromotopic, chronotopic and inotropic effects of calcium antagonists, are related to the degree of peripheral dilation. Nifedipine is the most potent arterial vasodilator and is consequently associated with the most intense reflex adrenergenic activity. Although their effects on vascular and cardiac muscle are similar but not identical, there exist major differences in their antiarrhythmic properties. All act as an antiarrhythmic agent when ischemia or reperfusion cause the arrhythmias, while verapamil selectively blocks the A-V node conduction. All three discussed calcium antagonists are effective in treating patients with coronary spasm, variant angina and unstable angina. In our personal experience with 73 patients with unstable angina with prolonged severe pain at rest with transient ST-segments and T-wavechanges without elevated enzyme levels, 21 became asymptomatic within 8 hours of treatment with conventional therapy, which included nitrates and betablockers. Of 52 who remained refractory to such therapy, the addition of 10 mg of nifedipine orally every two hours to a maximum of 60 mg rendered 42 of the 52 asymptomatic within 8 hours. Arguments why we believe that the timely administration of nifedipine to these and similar patients will reduce or delay the incidence of arrhythmias and myocardial infarction are given on the basis of experimental data. Nifedipine greatly dilates coronary arteries, an effect which persists even after the drug's general hemodynamic effects disappear. It is shown that antagonists have anti-ischemic properties which are primarily related to the persistent reduction of the basic coronary vascular tone which increases oxygen supply whilst reduced myocardial contractility leads to decreased oxygen consumption. Furthermore, preservation of cellular integrity is achieved via protection against intracellular Ca2+ excess, as demonstrated by preserved intracellular high energy phosphate. A randomized multicenter trial in the Netherlands is now in progress to clarify the definitive role of beta blockade versus calcium antagonists therapy of their combination in this syndrome.

Mechanism(s) of calcium antagonists and ischemia.

An overview is provided of the mechanismen of action of the three main calcium antagonists: nifedipine, verapamil and diltiazem, in ischemic heart disease. The thesis is proposed that in individual cases with angina pectoris and ischemic heart disease different and differing actions must be expected with this type of pharmaco-therapy.

Anti-anginal, electrophysiologic and hemodynamic effects of combined beta-blocker/calcium antagonist therapy.

Nitrates and beta-blockers have been the mainstay in the therapy of chronic stable angina pectoris for many years. Since an important number of patients remains symptomatic, new potent anti-ischemic agents like the calcium antagonists fulfil a great clinical need. Combined therapy with beta-blockers and calcium antagonists is attractive, since both classes of drugs have differing and eventually complementary modes of action. On the other hand, both have direct negative inotropic and chronotropic effects. We reviewed the anti-anginal, electrophysiologic and hemodynamic effects of combined treatment with a beta-blocker and verapamil or nifedipine. Combined therapy provides greater symptomatic relief than monotherapy with beta-blockers or slow channel blockers alone. While incidental adverse negative inotropic and chronotropic interactions have been reported, particularly when verapamil is involved, their hemodynamic interplay appears beneficial rather than detrimental in the majority of patients. Indeed, combined therapy is effective and safe, at least when a preserved or only moderately impaired left ventricular function is present. However, caution must be exercised in patients with more impaired left ventricular function, and combined therapy with verapamil must be avoided when conduction disturbances are likely to occur.

Influence of intracoronary nifedipine on left ventricular function, coronary vasomotility, and myocardial oxygen consumption.

The effect of intracoronary nifedipine on regional and global left ventricular performance, coronary vasomotility, and myocardial oxygen consumption is reported. Left ventricular pressures and volume indices of contractility and relaxation were simultaneously recorded in five patients without coronary artery disease. In these patients, nifedipine in the left main coronary artery not only delayed (+115 ms) anterior wall contraction but also slowed (3.5 vs 1.9 cm/s) and depressed it (-26%), resulting in a depression of global left ventricular ejection. This asynchrony and depression of regional contraction is considered to be responsible for the slowed isovolumic contraction and relaxation of the whole ventricle. In 10 other patients with coronary artery disease, coronary sinus blood flow and myocardial oxygen consumption were measured before and after intracoronary nifedipine. The observed decrease in myocardial oxygen consumption (-28%) depended primarily on a decrease in contractility and left ventricular performance. In a third study group of 12 patients with coronary artery disease, the effects of intracoronary nifedipine on the coronary vasomotility of 40 coronary segments (normal, prestenotic, stenotic, poststenotic) were quantitatively determined. Left ventricular haemodynamics and coronary sinus saturation were monitored while the cineangiograms were recorded before and after nifedipine. Nifedipine provoked vasodilatation of the normal (+10.3%), prestenotic, stenotic (+4 to 30%), and poststenotic (+16.4%) coronary segments, which persisted after the disappearance of its direct effects on the myocardium. This transient regional "cardioplegic" effect of nifedipine, associated with an increase in coronary blood flow, a reduction in myocardial oxygen consumption, and a vasodilatation of the epicardial vessels is likely to be beneficial during temporary coronary occlusion such as occurs in spasm or transluminal angioplasty.

Acute myocardial ischaemia: what information can be expected in the near future from ongoing clinical trials with nifedipine?

Nifedipine is a calcium antagonist with established anti-vasospastic action in man and, possibly cardioprotective effects during ischaemia. Although it is regularly used in the treatment of acute myocardial ischaemia, few double-blind randomized trials have been performed in this area. Recently, one trial was published which showed that nifedipine, when added to conventional treatment (beta-blockers, nitrates) of patients hospitalized for unstable angina pectoris (UAP) prevents recurrent ischaemia. Several further trials are known to be under way. The American nifedipine angina-myocardial infarction study (NAMIS) comprises sub-protocols both for patients diagnosed as UAP and patients diagnosed as acute myocardial infarction (AMI). In the former, a comparison is made between a nifedipine and a propranolol based regimen. Recurrence of ischaemia is the end point. In the NAMIS-protocol for AMI; nifedipine is compared with placebo. Infarct-size and complications are the end points. The Holland Interuniversity Nifedipine/metoprolol Trial (HINT) is similar in design to the NAMIS UAP-study. However, it has a placebo group for comparison and also deals with the combination of nifedipine and beta-blockade. NAMIS and HINT may resolve several important questions in reference to the treatment of acute myocardial ischaemia but they are of insufficient size to study effects on total mortality. Finally, in the area of secondary prevention after MI, a major trial (secondary prevention reinfarction nifedipine trial: SPRINT) is known to be under way in Israel.

What is preferable in unstable angina, beta-blockade or calcium-inhibition?

Nifedipine is a strong calcium antagonist; it blocks the excitation-contraction coupling, yet at therapeutic dosage levels it has few side effects. It is said that a single molecule of nifedipine can neutralize the effects of several thousand calcium ions in the excitation-process. Thus this agent reduces the ability of the cardiac cells to develop mechanical tension yet it does not affect electrical excitation. Calcium antagonists are also potent dilators, particularly of the arteries on the periphery of the human body and in the coronary arterial system. The basic mode of action is thought to be the same, i.e. reduction of the calcium transport, this time into the smooth muscle cells of the arterial wall, thus "forcing' the arteriole to dilate. The resultant action is a reduction in afterload, while there is an increase in perfusion of the coronary vascular bed. Thus there is a dual action, the one centrally, the other peripherally both of which will be particularly effective in patients with unstable angina (UA) in whom excess coronary vascular tone is suspected. Beta-blockers, particularly propranolol and metoprolol, with which we have the most experience, work via beta 2 adrenoceptor blockade in the heart. Cardiac frequency decreases, arrhythmias are suppressed, blood pressure may decrease, all reducing cardiac work. If in UA variations in coronary vascular tone and frank spasm do occur then is would be more logical to use Ca-antagonists instead of beta-blockade. Our clinical experience to date has not shown that patients with UA are worsened by propranolol or metoprolol but there are scores of clinical reports showing worsening of chest pain in Prinzmetal's syndrome, presumably because beta-blockade leads to excessive alpha-adrenergic vascular tone. On the other hand, in one year experience with 73 patients with UA, 52 did not experience relief from adequate beta-blockade until nifedipine was added. Within 2 hours, 42 of the 52 became asymptomatic--thus indicating that in UA the need for spasmolytic therapy prevails over the need for beta-blockade. In many cases of UA both drugs may have a place however, their exact relative value remains to be established.