The effectiveness of protective clothing in the reduction of potential DNA contamination of the scene of crime.

The use of ultra-sensitive low copy number (LCN) DNA typing allows the analysis of picogram amounts of DNA. Trace evidence accidentally left at a scene of crime (SOC) by the investigating team may be inadvertently collected and analysed, potentially leading to spurious evidence being introduced into the criminal investigation. A series of experiments were undertaken to determine the extent to which an investigator could contribute to any DNA contamination of a scene of crime under different simulated activities. Further, the degree to which any contamination was reduced by the use of commercially available protective clothing was demonstrated. Precautions that should routinely be taken at a scene of crime to reduce the risk of DNA contamination are recommended.

Acidosis correction with a new 25 mmol/l bicarbonate/15 mmol/l lactate peritoneal dialysis solution.

OBJECTIVE: The aim of this study was to evaluate the effects of a combined 25 mmol/L bicarbonate/15 mmol/L lactate-based solution (Bic/Lac), compared to a 35 mmol/L lactate solution (Lac)--the most commonly used solution for patients in southern Europe--on the venous plasma bicarbonate level in patients treated with continuous ambulatory peritoneal dialysis (CAPD). DESIGN: This was a randomized, parallel, controlled, open-label study, with patients studied for a period of 3 months preceded by a 1-month baseline and followed by a 1-month follow-up. Patients used the 35 mmol/L lactate solution during baseline and follow-up periods. SETTING: Four Spanish nephrology centers. PATIENTS: Thirty-one (20 Bic/Lac, 11 Lac) well-dialyzed (creatinine clearance > 55 L/week/1.73 m2 body surface area) CAPD patients. INTERVENTIONS: Blood samples were taken for biochemistry tests at all visits. A physical examination was completed at baseline and month 3, and a medical update was completed after 1, 2, and 3 months, and at the follow-up visit. Adverse-event monitoring and notation of prescription changes were carried out continuously. MAIN OUTCOME MEASURE: Effect on venous plasma bicarbonate level. RESULTS: Venous plasma bicarbonate rose by 3.1 mmol/L (confidence intervals 1.6-4.8),from a baseline level of 23.0 mmol/L during the treatment period in those patients treated with Bic/Lac (p < 0.05 vs Lac). The number of acidotic patients (venous plasma bicarbonate < 24 mmol/L) was statistically significantly reduced at every treatment period visit in the Bic/Lac group (p < 0.05). There were no adverse findings with respect to vital signs, physical examination, or clinical symptoms, apart from one death in the control group. CONCLUSIONS: The new Bic/Lac solution allowed better correction of acid-base status than the lactate solution.

Tryptophan depletion impairs stimulus-reward learning while methylphenidate disrupts attentional control in healthy young adults: implications for the monoaminergic basis of impulsive behaviour.

RATIONALE: Altered serotonergic and dopaminergic function have been widely implicated in behavioural disorders associated with impulsivity and risk-taking. However, little research has addressed the specific cognitive consequences of changed monoaminergic function that might contribute to the production of impulsive behaviour. OBJECTIVES AND METHODS: We compared the effects of rapid plasma tryptophan depletion, acute doses of the mixed indirect catecholamine agonist, methylphenidate (40 mg), and acute doses of the alpha(1)/alpha(2 )agonist, clonidine (1.5 microg/kg), on aspects of visual discrimination learning involving either acquisition of altered stimulus-reward associations (i.e. updating the affective valence of exteroceptive stimuli) or the control of attention towards relevant as opposed to irrelevant stimulus dimensions. RESULTS: Relative to subjects who received placebo, subjects with reduced tryptophan exhibited a deficit in the ability to learn changed stimulus-reward associations, but were still able to shift an acquired attentional set away from a now-irrelevant stimulus dimension towards a newly relevant dimension. By contrast, subjects who received methylphenidate were able to learn effectively about changing stimulus-reward associations, but showed an enhanced ability to shift an attentional bias, in combination with slowed response times. Subjects who received clonidine showed neither of these changes. CONCLUSIONS: These results suggest that reduction in central serotonin leads to altered neuromodulation of the cortical and subcortical regions (e.g. orbitofrontal cortex, striatum and anterior temporal structures) that mediate important aspects of associative learning whereby exteroceptive stimuli acquire altered incentive motivational value. On the other hand, facilitation of catecholamine neurotransmitters may disrupt the allocation of attention between relevant and irrelevant features of the environment, perhaps through altered modulation of the dorsolateral prefrontal cortex. The implications of these results for understanding the differential neuromodulation of cognitive functions are discussed.

An actin gene-related polymerase chain reaction (PCR) test for identification of chicken in meat mixtures.

Using the polymerase chain reaction (PCR) and DNA extracted from muscle, a single pair of oligonucleotide primers can yield amplification products from several members of the actin multigene family simultaneously. These multiple PCR products form species-specific "fingerprints" on gel electrophoresis which may be useful for meat authentication. However, for analysis of meat mixtures, the presence of a single band unique to a species would have many advantages over a multi-component fingerprint. A procedure is described in which primers amplify at a single actin gene locus, giving a positive band with DNA extracted from chicken and turkey, but no reaction with duck, pheasant, porcine, bovine, ovine or equine DNA. The chicken signal was clearly detectable with DNA from meat admixtures containing 1% chicken/99% lamb and from meat heat-treated at 120°C. For further discrimination, the chicken PCR product could be differentiated from turkey by restriction enzyme digestion.

A controlled trial of two bicarbonate-containing dialysis fluids for CAPD--final report.

BACKGROUND: The combination of a low pH and a high concentration of lactate which is present in most dialysis fluids is found to be cytotoxic in vitro. For these reasons it would seem logical to use a bicarbonate-containing solution and thus automatically provide a solution with a neutral pH. METHODS: A parallel, randomized, open-label, prospective 2-month trial with an optional 4 month extension was undertaken to compare two novel bicarbonate-based solutions; one containing 38 mmol/l of bicarbonate (B), and one containing a mixture of 25 mmol/l bicarbonate and 15 mmol/l of lactate (B/L), with a control solution (C) containing 40 mmol/l lactate. RESULTS: Three groups of 19 (C), 20 (B), and 20 (B/L) patients were recruited and data from approximately 55 patient months were accumulated in each group. The data show that both bicarbonate-based solutions maintain acid-base levels within the normal range, that there were no changes in any of the other blood biochemistry parameters measured in the peritoneal equilibration test or with regard to adequacy of dialysis, and that furthermore, both solutions were well tolerated. CONCLUSIONS: This study showed that either the bicarbonate or bicarbonate/lactate solutions could be utilized efficaciously in patients undergoing CAPD.

The actin multigene family and livestock speciation using the polymerase chain reaction.

Actins constitute a family of highly-conserved multifunctional intracellular proteins, best known as myofibrillar components in striated muscle fibres. Most vertebrate genomes contain numerous actin genes with high sequence homology in protein coding regions but considerable variability in intron number and sizes. This genetic diversity can be utilised for livestock speciation purposes. The high sequence conservation has enabled a single pair of oligonucleotides to be used to prime the polymerase chain reaction (PCR) with DNA extracted from all animals so far studied. Multiple amplification products were obtained which on gel electrophoresis constituted characteristic species-specific 'fingerprints'. The patterns were reproducible, did not vary between individuals of the same breed or between different breeds within a species, and could be generated even from heat-processed muscle held at 120 degrees C for one hour. Given the capacity of PCR to amplify relatively short sequences in highly-degraded DNA, this approach may be suitable for authentication of processed meat products.

Meat speciation by restriction fragment length polymorphism analysis using an α-actin cDNA probe.

Classical DNA fingerprinting is based on separation of DNA restriction fragments by electrophoresis and hybridisation to nucleic acid probes containing repetitive nucleotide sequences. The use of such mini- or micro-satellite probes tends to yield patterns specific to an individual rather than to a species, hence their value in forensic analysis but general unsuitability for meat speciation. In the present study, a cDNA probe based on conserved sequences contained in members of the actin multigene family has been evaluated for potential application in meat speciation. Genomic DNA was extracted from muscle and digested with BamHI before electrophoresis and hybridisation to a murine α-actin cDNA probe. Beef, pork, lamb, horse, chicken and fish DNA restriction fragments formed characteristic 'fingerprints' which were reproducible and varied sufficiently to allow discrimination even between closely-related species. However no major differences were seen between individuals of the same breed or between different breeds within a species. When DNA obtained from fresh tissue and also from meat heated at 120 °C was analysed, the gel patterns were essentially the same. An attractive feature of this approach is that it employs a single cross-reacting probe and set of conditions, and gives different patterns with all species so far studied. This simplicity suggests applications in meat speciation or related areas of biology.

DNA typing from human faeces.

A method has been developed for the forensic analysis of faeces by DNA amplification and direct sequencing of a polymorphic segment of mitochondrial DNA. Starting from as little as 10 mg wet weight of faeces, DNA was extracted by a variety of protocols and amplified using primers specific to hypervariable region 1 of the mitochondrial control region. The resulting amplification products were sequenced in solid phase using an automated DNA sequencer. In total, mtDNA sequences were generated from the faeces of nine Caucasians and compared with sequences generated from their respective blood samples. Sequences of faeces and blood samples from the same individual were identical in every case, but a range of 1-10 nucleotide differences was observed between individuals, with an average sequence variation of approximately 4.88 per 400 bp. Of the various extraction protocols assessed in this study, greatest success rates were achieved using magnetisable beads to bind and purify the DNA. STR analysis of DNA extracted from faeces was not routinely possible.

Adenosine 5'-triphosphate, adenosine and endothelium-derived relaxing factor in hypoxic vasodilatation of the heart.

The involvement of ATP in hypoxic vasodilatation was investigated using isolated perfused guinea-pig hearts (Langendorff). Reactive blue 2, a selective P2Y-purinoceptor antagonist, attenuated dilatations due to ATP and hypoxia. Hydroquinone, an agent which destroys endothelium-derived relaxing factor, substantially decreased dilatations due to 2-methylthioATP, a potent P2Y-purinoceptor agonist, and hypoxia, but not to adenosine. ATP may, therefore, have an important role to play in the initiation of hypoxic dilatation which is mediated by the release of endothelium-derived relaxing factor.

A randomised comparative trial of nicardipine versus amiloride and hydrochlorothiazide in mild to moderate hypertension. A report from the General Practitioner Hypertension Study Group.

In 1984 the General Practitioner Hypertension Study Group undertook a rescreening of their patient population, looking for patients who still had untreated mild to moderate essential hypertension. Suitable patients were entered into a clinical trial comparing the safety and efficacy of nicardipine (a calcium antagonist) and amiloride + hydrochlorothiazide (HCTZ) (moduretic). The study included one year of long-term follow-up. Both drugs significantly lowered BP in both the short and long term. Numbers and percentages of patients from each group reporting adverse experiences were similar in the short term, but in the long term the frequency of adverse event reporting was much lower with nicardipine treatment than with amiloride + HCTZ treatment (2/10 versus 9/17). Treatment with amiloride + HCTZ led to elevations in serum levels of cholesterol, uric acid and urea, which were maintained at one year, whilst no abnormalities in blood biochemistry were seen in patients treated with nicardipine. In conclusion we have found that nicardipine compares very favourably with amiloride + HCTZ in the treatment of mild to moderate hypertensive patients.

Ultrastructural localisation of substance P and choline acetyltransferase in endothelial cells of rat coronary artery and release of substance P and acetylcholine during hypoxia.

Substance P and choline acetyltransferase have been localised in a small proportion of endothelial cells of rat coronary arteries using electron microscopic immunocytochemistry. During a hypoxic period of 1 min, coronary vasodilatation was produced in the Langendorff heart preparation and increased levels of substance P and acetylcholine were released into the perfusate. The possibility that these substances are released from endothelial cells during hypoxia and contribute to the hyperaemic response is discussed.

GTP-independent stimulation of rabbit heart adenylate cyclase by isoproterenol at physiological ATP concentrations.

Isoproterenol increased the activity of the adenylate cyclase of rabbit heart sarcolemmal membranes in the absence of added GTP. ATP, the ATP-regenerating system, and the sarcolemmal membrane preparation were eliminated as possible sources of contaminating GTP. Isoproterenol-stimulation increased as ATP was raised. At 0.5 mM ATP, isoproterenol increased activity by 19% whereas at 5 mM ATP isoproterenol increased activity by 121%. There was no change in basal activity between 0.5 and 5 mM ATP. Stimulation by Gpp(NH)p and NaF increased slightly between 0.5 and 5 mM ATP; stimulation by KCl was unaffected. GTP does not activate cyclase d. GTP does not activate cyclase to the same extent as Gpp(NH)p even though the two act at the same site on Ns (the stimulatory guanine nucleotide-binding protein). GTP decreased cyclase activation by Gpp(NH)p in a concentration-dependent fashion when the two were added to the assay simultaneously. Increasing ATP from 0.5 to 5 mM did not reduce activation by Gpp(NH)p when both were added simultaneously to the assay. This suggests that ATP does not interact with the same site as Gpp(NH)p. ATP gamma S, an analogue of ATP which irreversibly thiophosphorylates proteins, did not irreversibly support activation by isoproterenol. The effect of ATP in supporting isoproterenol stimulation is not, therefore, thought to be due to phosphorylation of a protein.

Serotonin is localized in endothelial cells of coronary arteries and released during hypoxia: a possible new mechanism for hypoxia-induced vasodilatation of the rat heart.

In this report we demonstrate the immunocytochemical localization of serotonin in endothelial cells of rat coronary vessels and a significant increase in the release of serotonin into the perfusate of Langendorff rat heart preparations during hypoxia. It is suggested that serotonin, localized in endothelial cells, is released during hypoxia and could provide part of a pathophysiological mechanism for vasodilatation to protect the heart from damage due to hypoxia.

Pertussis toxin reduces the antiadrenergic effect of 2-chloroadenosine on papillary muscle and the direct negative inotropic effect of 2-chloroadenosine on atrium.

2-Chloroadenosine reduced the contractile tension of guinea-pig atria directly, and inhibited the increase in tension produced by beta-adrenergic stimulation of guinea-pig papillary muscle. Both effects were reduced by 8-phenyltheophylline, a competitive antagonist at extracellular P1-purinoceptors. Treatment of guinea-pigs with pertussis toxin reduced the sensitivity of both atria and ventricles to 2-chloroadenosine. Atria were significantly affected after treatment with 125 micrograms/kg toxin, but not 100 micrograms/kg. 60 micrograms/kg toxin had no effect on the sensitivity of the ventricles, but 100 and 125 micrograms/kg significantly decreased the antiadrenergic effect of 2-chloroadenosine. We conclude that both the direct and antiadrenergic effects are mediated by an inhibitory guanine nucleotide binding protein.

An antiadrenergic effect of adenosine on guinea-pig but not rabbit ventricles.

The antiadrenergic effect of adenosine was investigated using isolated guinea-pig heart and guinea-pig and rabbit papillary muscle. Adenosine, 15 microM, completely abolished the increased tension stimulated by 0.1-1.0 nM isoprenaline in Langendorff-perfused guinea-pig hearts. With guinea-pig papillary muscles, adenosine decreased by 40% the increased force stimulated by 1-10 nM isoprenaline. When 5 microM 2-chloroadenosine was used in conjunction with 1 unit ml-1 adenosine deaminase, a complete inhibition of the isoprenaline-stimulated tension was seen in guinea-pig papillary muscles. The antiadrenergic effect of 2-chloroadenosine was blocked by 8-phenyltheophylline. In rabbit, there was little effect of 2-chloroadenosine (plus deaminase) on isoprenaline-stimulated tension. (-)-N6 (R-phenylisopropyl)-adenosine (PIA) had no effect on basal or isoprenaline-stimulated adenylate cyclase activity of guinea-pig or rabbit sarcolemmal membranes. We conclude that the antiadrenergic effect of adenosine is mediated by A type receptors and is seen in guinea-pig but not rabbit. Production of adenosine by superfused papillary muscle may obscure the effect of added adenosine. We find no evidence that the antiadrenergic effect is mediated by inhibition of adenylate cyclase.

ATP mediates coronary vasoconstriction via P2x-purinoceptors and coronary vasodilatation via P2y-purinoceptors in the isolated perfused rat heart.

The effect of ATP and its analogues on the perfusion pressure of the rat coronary vasculature and the left ventricular pressure of the isolated Langendorff perfused rat heart was examined. The response to ATP was generally biphasic, causing an increase followed by a decrease in perfusion pressure. The rank order of potency of the analogues for eliciting the vasoconstriction component was alpha,beta-methyleneATP greater than 2-methylthioATP greater than ATP, which resembles the pattern previously observed for the P2x-purinoceptor. In causing vasodilation, the rank order of antagonist potency was 2-methylthioATP greater than ATP, with alpha,beta-methyleneATP being without effect; this is a characteristic of P2y-purinoceptors. 1,3-Dipropyl-8-(2-amino-4-chlorophenyl)xanthine (PACPX) antagonised some of the vasodilatory effects of ATP, showing that some of the effect is due to breakdown to adenosine. Reactive blue 2, a putative P2y-purinoceptor antagonist was found to be five times more effective at antagonising the vasodilatory responses to 2-methylthioATP than to adenosine or the vasoconstriction responses to alpha,beta-methyleneATP. We suggest that ATP exerts its biphasic effects in the coronary vasculature of the rat by interacting with P2x- and P2y-purinoceptors and partly via P1-purinoceptors after breakdown to adenosine. Reactive blue 2 was more effective at antagonising responses mediated via P2y-purinoceptors than by P2x- or P1-purinoceptors.