Pharmacological prevention of cardiac risk in claudicants with ischaemic heart disease.

BACKGROUND: Claudicants rarely progress to critical limb ischaemia but have a threefold increase in mortality, mainly due to cardiac disease. Antithrombotic therapy, beta blockers, ACE inhibitors and statins have been shown to reduce mortality and cardiovascular morbidity in patients with ischaemic heart disease. AIM: To investigate secondary pharmacological prevention of ischaemic heart disease in claudicants. MATERIALS AND METHODS: We prospectively recorded comorbidity and drug treatment in 89 patients (67 men and 22 women) with a history of ischaemic heart disease recruited in a supervised exercise and lifestyle modification programme to improve claudication distance and prognosis. RESULTS: Of the 89 cases, 40 had a history of angina only and 49 of myocardial infarction. Sixteen (18%) had diabetes, 47 (53%) had hypercholesterolaemia and 52 (58%) were hypertensive. Antithrombotic therapy was prescribed to 61 patients (68.5%), 64 (72%) with a history of myocardial infarction and 27 (67.5%) with angina only (p = 1). Beta-blockers were prescribed to 12 (13.5%) patients only, seven (15%) with a history of myocardial infarction and five (12.5%) with angina only (p = 1). Of the 47 patients with hypercholesterolaemia, 29 (62%) were on a statin. CONCLUSION: Secondary pharmacological prevention of ischaemic heart disease in claudicants remains suboptimal, with only two thirds of patients receiving antithrombotic therapy and a small minority receiving beta blockers. Pharmacological prevention in claudicants should improve to reduce cardiac morbidity and mortality.

The contribution of the external carotid artery to cerebral perfusion in carotid disease.

PURPOSE: In the presence of carotid occlusion, the external carotid artery (ECA) becomes an important source of cerebral blood flow, especially if the circle of Willis is incomplete. The contribution of the ECA to hemispheric blood flow in patients with severe ipsilateral carotid stenosis has never been previously investigated. METHODS: One hundred eight patients were monitored during sequential cross-clamping of the external (ECA) and then ipsilateral internal carotid artery (ICA) during carotid endarterectomy using transcranial Doppler sonography (TCD) (Neuroguard CDS, Los Angeles, Calif), to measure middle cerebral artery blood flow velocity, and near-infrared spectroscopy, to measure regional cerebral oxygen saturation (CsO(2)) (Invos 3100A; Somanetics, Troy, Mich). RESULTS: On the ipsilateral ECA cross-clamp, the median fall in CsO(2) was 3% (interquartile range, 1%-4%; P <.0001). On addition of the ICA cross-clamp there was a further fall of 3% and a total fall of 6% (3%-9%; P <.0001). The median percentage fall in middle cerebral artery blood flow velocity on ECA clamping was 12% (4%-24%; P <.0001); on ICA clamping it was 48% (25%-74%; P <.0001). Falls in TCD on ECA clamping were greater with increasing severity of ipsilateral ICA stenosis. The correlation between CsO(2) and TCD on external clamping, although less strong than that on internal clamping, was statistically significant r = 0.32; P =.01; Spearman rank correlation). CONCLUSIONS: The falls in TCD and CsO(2) were of a similar order of magnitude and must therefore reflect a fall in cerebral perfusion. The ipsilateral ECA contributes significantly to intracranial blood flow and oxygen saturation in severe carotid stenosis.

Should heparin be reversed after carotid endarterectomy? A randomised prospective trial.

OBJECTIVE: To investigate whether heparin reversal after carotid endarterectomy reduces the incidence of haemorrhagic complications. DESIGN: A randomised prospective trial. METHODS: Sixty-four patients randomised to reversal of heparin or no reversal, of whom 31 received protamine titrated to the residual circulating heparin at closure of arteriotomy. Measurements included serial activated clotting times (ACTs), wound drainage, neck swelling using duplex Doppler imaging to measure the depth from skin to carotid bifurcation, and the recording of all complications. RESULTS: Wound drainage volumes were significantly reduced by protamine reversal (68.5 ml compared to 35 ml, p < 0.001), but neck swelling was not (72 mm compared to 70 mm, p = 0.77). Two patients who were not reversed developed neck haematomas requiring evacuation. More importantly, two patients receiving protamine, thrombosed the operated internal carotid artery (ICA) postoperatively and died despite urgent thrombectomy. A further patient who was not randomised in this study but who received protamine also developed ICA thrombosis within the same 3 month period. CONCLUSIONS: Reversing heparin with protamine reduces postoperative wound drainage after carotid surgery but may predispose to ICA thrombosis and stroke. This is in keeping with a previous retrospective study published during our trial.

Leg ulcer odour detection identifies beta-haemolytic streptococcal infection.

beta-Haemolytic streptococci were identified in bacteriological cultures from 14 of 24 chronic venous leg ulcers in 21 patients. Multi-element odour detection (MEOD) analysis demonstrated a significant difference in odour in those ulcers from which beta-haemolytic streptococci were isolated (p < 0.01). MEOD has potential to detect pathogenic organisms instantaneously in the clinical setting.

The Effects of Heavy Metals and Root Immersion on Isoflavonoid Metabolism in Alfalfa (Medicago sativa L.).

Modest increases in the concentration of medicarpin, 6-fold in leaves and 4-fold in roots, were observed in alfalfa (Medicago sativa L.) seedlings treated with 1 mM metal salts for 72 h. However, medicarpin-3-O-glucoside-6"-O-malonate (MGM) and formononetin-7-O-glucoside-6"-O-malonate (FGM) levels were up to 50-fold lower in metal-treated compared to control roots. Approximately 10% of the "missing" conjugates could be accounted for in the root treatment solution, where FGM and MGM transiently accumulated prior to their hydrolysis. Time-course studies revealed that total isoflavonoid content (roots plus solution) increased slightly after CuCl2 treatment, whereas the levels of FGM and MGM increased rapidly in alfalfa roots immersed in water. This increase was reduced by aeration. The phenylalanine ammonia-lyase inhibitor L-[alpha]-aminooxy-[beta]-phenylpropionic acid was used to show that immersion of the roots reduced conjugate rates of degradation, which explains their accumulation. In contrast, conjugate rates of degradation were elevated in CuCl2-treated roots, with 50% of the increase being due to hydrolysis. Up to 90% of formononetin and medicarpin produced in response to CuCl2 treatment arose via conjugate hydrolysis. Our results demonstrate that both immersion/anaerobiosis and abiotic elicitation modify isoflavonoid metabolism in alfalfa, and that metal-stimulated accumulation of phytoalexins may arise through the release from preformed stores rather than de novo synthesis.

The management of cardiac trauma by general surgeons in non-cardiothoracic units.

Cardiac trauma requiring immediate surgical intervention is usually thought of as a rare event. However, our recent experience at Leicester Royal Infirmary suggests that this clinical situation is on the increase, and has highlighted potential problems in the management of these patients. It is essential that all units that may have to deal with these patients have the necessary facilities and equipment to perform an emergency thoracotomy in the Accident and Emergency department, and that the medical and nursing staff involved have been properly trained to deal with these life-threatening emergencies.

Abscisic acid biosynthesis in roots : I. The identification of potential abscisic acid precursors, and other carotenoids.

The pathway of water-stress-induced abscisic acid (ABA) biosynthesis in etiolated and light-grown leaves has been elucidated (see A.D. Parry and R. Horgan, 1991, Physiol. Plant. 82, 320-326). Roots also have the ability to synthesise ABA in response to stress and it was therefore of interest to examine root extracts for the presence of carotenoids, including those known to be ABA precursors in leaves. All-trans- and 9'-cis-neoxanthin, all-trans- and 9-cis-violaxanthin, antheraxanthin (all potential ABA precursors), lutein and ß-carotene were identified on the basis of absorbance spectra, reactions with dilute acid, retention times upon high-performance liquid chromatography and by comparison with leaf carotenoids that had been analysed by mass spectrometry. The source of the extracted carotenoids was proved to be root tissue, and not contaminating compost or leaf material. The levels of total carotenoids in roots varied between 0.03-0.07% of the levels in light-grown leaves (Arabidopsis thaliana (L.) Heynh, Nicotiana plumbaginifolia Viv., Phaseolus vulgaris L. and Pisum sativum L.) up to 0.27% (Lycopersicon esculentum Mill.). The relative carotenoid composition was very different from that found in leaves, and varied much more between species. All-trans-neoxanthin and violaxanthin were the major carotenoids present (64-91 % of the total), but while Lycopersicon contained 67-80% all trans-neoxanthin, Phaseolus, Pisum and Zea mays L. contained 61-79% all-trans-violaxanthin. Carotenoid metabolism also varied between species, with most of the carotenoids in older roots of Phaseolus being esterified. Roots and leaves of the ABA-deficient aba mutant of Arabidopsis had reduced epoxy-xanthophyll levels compared to the wild-type.

Abscisic acid biosynthesis in roots : II. The effects of water-stress in wild-type and abscisic-acid-deficient mutant (notabilis) plants of Lycopersicon esculentum Mill.

The ubiquity of the apo-carotenoid abscisic acid (ABA) biosynthetic pathway elucidated in water-stressed, etiolated leaves of Phaseolus vulgaris (see A.D. Parry and R. Horgan, 1991, Physiol. Plant. 82, 320-326), has been difficult to establish. Light-grown leaves contain very high carotenoid: ABA ratios, preventing correlative studies, and no etiolated leaves so far studied, other than those of Phaseolus, have been found capable of synthesising significant amounts of ABA in response to stress. Roots are known to synthesise ABA and contain low carotenoid levels; therefore ABA biosynthesis was investigated in soil- and hydroponically grown roots of Lycopersicon esculentum Mill. Hydroponically grown roots were stressed by immersion in 100 mM mannitol and soil-grown roots by withholding water. In both cases stress led to an increase in ABA levels and a decrease in the levels of specific xanthophylls, namely all-trans- and 9'-cis-neoxanthin and all-trans-violaxanthin. In hydroponically grown roots, and soil-grown roots stressed after removal of the shoot, ratios of xanthophyll cleaved:ABA synthesised of approx. 1∶1 were obtained. These findings are consistent with the operation of an apo-carotenoid pathway in roots, involving the conversion of all-trans-violaxanthin via all-trans-neoxanthin, to 9'-cis-neoxanthin, and the specific cleavage of 9'-cis-neoxanthin to yield the C15 ABA precursor xanthosin. Similar experiments with roots of the "leaky", ABA-deficient mutant of Lycopersicon, notabilis, indicate that the mutation does not affect the perception or transduction of stress, or the ability of the plant to cleave carotenoids. Rather, it appears that notabilis possesses an enzyme with reduced substrate specificity which cleaves more all-trans-than 9'-cis-neoxanthin.

Abscisic-acid metabolism in a wilty mutant of Nicotiana plumbaginifolia.

A mutant of Nicotiana plumbaginifolia, CKR1, isolated on the basis of its enhanced resistance to cytokinins was found to have a greater tendency to wilt than the wild type (Blonstein et al., 1991, Planta 183, 244-250). Further characterisation has shown that the wiltiness in the mutant is not caused by an insensitivity to abscisic acid (ABA) because the external application of ABA leads to stomatal closure and phenotypic reversion. The basal ABA level in the mutant is < 20% of that in the wild type. Following stress, the ABA level in wild-type leaves increases by approx 9-to 10-fold while the mutant shows only a slight increase. This deficiency in ABA is unlikely to be the consequence of accelerated catabolism as the levels of two major metabolites of ABA, phaseic and dihydrophaseic acid, are also much reduced in the mutant. The qualitative and quantitative distributions of carotenoids, the presumed presursors of ABA, are the same for the leaves of both wild type and mutant. Biosynthesis of ABA at the C15 level was investigated by feeding xanthoxin (Xan) to detached leaves. Wild-type leaves convert between 9-19% of applied Xan to ABA while the mutant converts less than 1%. The basal level of trans-ABA-alcohol (t-ABA-alc) is 3-to 10-fold greater in the mutant and increases by a further 2.5-to 6.0-fold after stress. This indicates that the lesion in the wilty mutant of N. plumbaginifolia affects the conversion of ABA-aldehyde to ABA, as in the flacca and sitiens mutants of tomato and the droopy mutant of potato (Taylor et al., 1988, Plant Cell Environ. 11, 739-745; Duckham et al., 1989, J. Exp. Bot. 217, 901-905). Wild-type tomato and N. plumbaginifolia leaves can convert trans-Xan into t-ABA-alc, and Xan into ABA, while those of flacca and the wilty N. plumbaginifolia mutant convert both Xan and t-Xan to t-ABA-alc.

A cytokinin-resistant mutant of Nicotiana plumbaginifolia is wilty.

Selection for cytokinin resistance by incubating M2 seed of Nicotiana plumbaginifolia, after ethylmethanesulphonate mutagenesis, on 20 µM 6-benzylami-nopurine resulted in the isolation of a monogenic, recessive mutant, CKR1. Germination of the mutant is less sensitive to cytokinin inhibition than the wild type, and leaf development of the mutant occurs at cytokinin concentrations inhibitory to the wild type. Germination of the mutant is also resistant to auxin but not to abscisic acid. Three other traits jointly inherited with cytokinin resistance in the F2 are lack of root branching, precocious germination and wiltiness. The wilty phenotype is the consequence of the failure of stomatal closure during water stress.

The role of cis-carotenoids in abscisic acid biosynthesis.

Evidence has been obtained which is consistent with 9'-cis-neoxanthin being a major precursor of abscisic acid (ABA) in higher plants. A mild, rapid procedure was developed for the extraction and analysis of carotenoids from a range of tissues. Once purified the carotenoids were identified from their light-absorbance properties, reactions with dilute acid, high-performance liquid chromatography Rts, mass spectra and the quasiequilibria resulting from iodine-catalysed or chlorophyllsensitised photoisomerisation. Two possible ABA precursors, 9'-cis-neoxanthin and 9-cis-violaxanthin, were identified in extracts of light-grown and etiolated leaves (of Lycopersicon esculentum, Phaseolus vulgaris, Vicia faba, Pisum sativum, Cicer arietinum, Zea mays, Nicotiana plumbaginifolia, Plantago lanceolata and Digitalis purpurea), and roots of light-grown and etiolated plants (Lycopersicon, Phaseolus and Zea). The 9,9'-di-cisisomer of violaxanthin was synthesised but its presence was not detected in any extracts. Levels of 9'-cis-neoxanthin and all-trans-violaxanthin were between 20- to 100-fold greater than those of ABA in light-grown leaves. The levels of 9-cis-violaxanthin were similar to those of ABA but unaffected by water stress. Etiolated Phaseolus leaves contained reduced amounts of carotenoids (15-20% compared with light-grown leaves) but retained the ability to synthesise large amounts of ABA. The amounts of ABA synthesised, measured as increases in ABA and its metabolites phaseic acid and dihydrophaseic acid, were closely matched by decreases in the levels of 9'-cis-neoxanthin and all-trans-violaxanthin. In etiolated seedlings grown on 50% D2O, deuterium incorporation into ABA was similar to that into the xanthophylls. Relative levels of carotenoids in roots and light-grown and etiolated leaves of the ABA-deficient mutants, notabilis, flacca and sitiens were the same as those found in wild-type tomato tissues.

Xanthoxin levels and metabolism in the wild-type and wilty mutants of tomato.

Using (13)C-labelled internal standards and gas chromatography-mass spectrometry/multiple-ion monitoring the levels of xanthoxin (Xan) and 2-trans-xanthoxin (t-Xan) have been determined in stressed and non-stressed leaves of wildtype tomato (Lycopersicon esculentum Mill cv. Ailsa Craig), and the wilty mutants, notabilis (not), flacca (flc) and sitiens (sit). Levels of Xan were very low in all tissues. Ratios of t-Xan: Xan ranged from 10:1 to <500:1. In the wild-type and flc, t-Xan levels increased following stress. The results from feeding experiments using [(13)C]Xan and t-Xan demonstrated that whilst wild-type and not plants readily converted Xan into abscisic acid (ABA), flc and sit plants converted only a small amount of applied Xan into ABA. In all plants t-Xan was not converted into ABA. These results indicate that the flc and sit mutants are impaired in ABA biosynthesis because they are unable to convert Xan into ABA, whereas the not mutant is blocked at a metabolic step prior to Xan. Another possible ABA precursor, ABA-1',4'-trans-diol (ABA-t-diol) was found to occur in wild-type and mutant tissue. All four tissues could convert [(2)H]ABA-t-diol to ABA. Incubation of stressed leaves in the presence of (18)O2 provided evidence consistent with Xan and ABA originating via oxidative cleavage of a xanthophyll such as violaxanthin.

The carotenoid and abscisic acid content of viviparous kernels and seedlings ofZea mays L.

Carotenoid and abscisic acid (ABA) levels were determined in endosperm, embryos and seedlings of wild-type and viviparous (vp) mutants ofZea mays L. Carotenoid concentrations were determined by absorption spectrometry following purification by high-performance liquid chromatography and ABA concentrations by combined gas chromatography-mass spectrometry. Lutein and zeaxanthin were the terminal carotenoids in wild-type tissue. The carotenoid profiles ofvp-1 andvp-8 tissue were similar to that of the wild type; invp-2, vp-5, vp-7 andvp-9 carotenogenesis was blocked at early stages so that xanthophylls were absent. Except forvp-1, where the ABA content was similar to the wild type, the ABA content ofvp embryos was substantially reduced, to 6-16% of the corresponding wild type. Thus, the absence of xanthophylls was associated with reduced ABA content, which was in turn correlated with vivipary. Kernels ofvp-8 had a reduced ABA content although xanthophylls were present. Seedlings of carotenoid-deficient mutants rescued from viviparous kernels contained less ABA than did wild-type seedlings grown in the same way. Furthermore, the ABA concentration of such seedlings did not increase in response to water deficit. Conversely,vp-1 seedlings contained normal levels of carotenoids and ABA. Carotenoid-deficient seedlings did not contain appreciable amounts of chlorophyll so that chloroplast development was not normal. Thus ABA-deficiency could be associated with abnormal plastid development rather than the absence of carotenoids per se.