Reduced red cell 2,3-diphosphoglycerate concentrations in critical illness without decreased in vivo P50.

We investigated whether red cell 2,3-diphosphoglycerate (2,3-DPG) concentrations are reduced in critical illness, whether acidaemia, hypophosphataemia or anaemia influence 2,3-DPG, and whether there is any net effect on in vivo P50. Twenty healthy, non-smoking, male volunteers were compared with 20 male intensive care patients with APACHE 2 scores >20 on the preceding day. Those transfused in this time were excluded. Venous red cell 2,3-DPG concentrations were measured in both groups. In the patient group, routine multichannel biochemical profile and arterial blood gas analysis were also performed and in vivo P50 calculated. The mean 2,3-DPG concentration was significantly lower in the patient group than in the controls (4.2+/-1.3 mmol/l vs 4.9+/-0.5 mmol/l, P=0.016). The patients were well oxygenated (lowest arterial PO2=75 mm Hg) and showed a tendency to acidaemia (median pH 7.37, range 7.06 to 7.48) and anaemia (median haemoglobin concentration 113 g/l, range 89 to 154 g/l). By linear regression of patient data, pH had a significant effect on 2,3-DPG concentrations (r=0.6, P=0.011). Haemoglobin and phosphate concentrations did not, but there were few abnormal phosphate values. There was no correlation between 2,3-DPG concentrations and in vivo P50 (r2 < or = 0.08). We conclude that 2,3-DPG concentrations were reduced in a broad group of critically ill patients. Although this would normally reduce the P50, the reduction was primarily linked with acidaemia, which increases the P50. Overall, there was no net effect on the P50 and thus no affinity-related decrease in tissue oxygenation.

Cardiac troponin I predicts myocardial dysfunction in aneurysmal subarachnoid hemorrhage.

OBJECTIVES: We studied the incidence of myocardial injury in aneurysmal subarachnoid hemorrhage (SAH) using the more sensitive cardiac troponin I (cTnI) assay, correlated changes in cTnI with creatine kinase, MB fraction (CK-MB), myoglobin, and catecholamine metabolite assays, and examined the predictive value of changes in cTnI for myocardial dysfunction. BACKGROUND: Myocardial injury in aneurysmal SAH as evidenced by elevated CK-MB fraction has been reported. Little published data exist on the value of cTnI measurements in aneurysmal SAH. METHODS: Thirty-nine patients were studied for seven days. Clinical cardiovascular assessment, electrocardiographic (ECG), echocardiography, cTnI, CK, CK-MB and CK-MB index, myoglobin and 24-h urinary catecholamine assays were performed in all patients. The ECG abnormalities were defined by the presence of ST-T changes, prolonged QT intervals, and arrhythmias. An abnormal echocardiogram was defined by the presence of wall-motion abnormalities and a reduced ejection fraction. The severity of SAH was graded clinically and radiologically. RESULTS: Eight patients demonstrated elevations in cTnI (upper limit of normal is 0.1 microg/liter with the immunoenzymatic assay and 0.4 microg/liter with the sandwich immunoassay), while five had abnormal CK-MB levels (upper limit of normal is 8 microg/liter). Patients with more severe grades of SAH were more likely to develop a cTnI leak (p < 0.05). Patients with cTnI elevations were more likely to demonstrate ECG abnormalities (p < 0.01) and manifest clinical myocardial dysfunction (p < 0.01) as evidenced by the presence of a gallop rhythm on auscultation and clinical or radiological evidence of pulmonary edema as compared to those with CK-MB elevations. The sensitivity and specificity of cTnI to predict myocardial dysfunction were 100% and 91%, respectively, whereas the corresponding figures for CK-MB were 60% and 94%, respectively. Elevations in myoglobin levels (upper limit of normal <70 microg/liter) and urinary catecholamine metabolites (urinary vanilmandelate/creatinine ratio upper limit of normal, 2.6) are a nonspecific finding. CONCLUSIONS: Measurements of cTnI reveal a higher incidence of myocardial injury than predicted by CK-MB in aneurysmal SAH, and elevations of cTnI are associated with a higher incidence of myocardial dysfunction. Thus, cTnI is a highly sensitive and specific indicator of myocardial dysfunction in aneurysmal SAH.

Are metalloproteins and acute phase reactants associated with cardiovascular disease in end-stage renal failure?

End-stage renal failure (ESRF) is associated with a higher risk of cardiovascular disease (CVD) than predicted by the major risk factors. We investigate the hypothesis that metalloproteins such as transferrin and ceruloplasmin and the inflammatory response are associated with CVD risk in this population. In this cross-sectional study of 81 subjects stable on haemodialysis (HD), 43 with CVD and/or peripheral vascular disease (PAD) were compared to 38 subjects without clinical evidence of CVD/PAD. Serum concentrations of metalloproteins and acute phase reactants were compared by univariate analysis and logistic regression modelling. Body mass index, gender ratios, prevalence of diabetes, iron status, and homocysteine concentrations did not differ significantly between the groups. Those with CVD were older (P< 0.001) and had been on dialysis for longer (P = 0.004). CVD subjects had significantly higher concentrations of ceruloplasmin (325 vs 284 mg/L, P = 0.011), copper (18.2 vs 15.7 micromol/L, P = 0.002), and C-reactive protein (CRP) (median 9.0 vs 3.8 mg/L, P = 0.002). Transferrin iron binding capacity tended to be higher in the CVD group (P = 0.088). CVD risk for subjects with serum concentrations in the upper tertile was increased 9.4-fold (CI 2.8-31.0) for copper, 4.2-fold (CI 1.5-12.2) for ceruloplasmin, 3.9-fold (CI 1.3-12.1) for transferrin iron binding capacity, and 2.3-fold (CI 0.9-6.1) for CRP. In multivariate logistic regression models, age (P = 0.001) and time on dialysis (P = 0.002) were the strongest risk factors for CVD. After adjustment for age and time on dialysis, transferrin iron binding capacity (P = 0.013) and copper (P = 0.019) continued to be associated with CVD risk but ceruloplasmin (P = 0.065) and CRP (P = 0.634) were not. Total cholesterol was associated with a lower risk of CVD (ie protective), presumably due to cholesterol-lowering therapy in high-risk patients. In conclusion, copper and transferrin iron binding capacity may be associated with CVD risk in HD subjects.

Randomized, placebo-controlled trial of the angiotensin-converting enzyme inhibitor, ramipril, in patients with coronary or other occlusive arterial disease. PART-2 Collaborative Research Group. Prevention of Atherosclerosis with Ramipril.

OBJECTIVES: The primary objective of this study was to investigate the effects of the angiotensin-converting enzyme (ACE) inhibitor, ramipril, on carotid atherosclerosis in patients with coronary, cerebrovascular or peripheral vascular disease. BACKGROUND: Angiotensin-converting enzyme inhibitors have been shown to reduce the risk of coronary events in various patient groups and to prevent the development of atherosclerosis in animal models. It has been hypothesized that the clinical benefits of ACE inhibitors may, therefore, be mediated by effects on atherosclerosis. METHODS: Six hundred seventeen patients were randomized in equal proportions to ramipril (5-10 mg daily) or placebo. At baseline, two years and four years, carotid atherosclerosis was assessed by B-mode ultrasound, and left ventricular mass was assessed by M-mode echocardiography. RESULTS: Blood pressure (BP) was reduced by a mean of 6 mm Hg systolic and 4 mm Hg diastolic in the ramipril group compared with the placebo group (p<0.001). There was no difference between groups in the changes in common carotid artery wall thickness (p = 0.58) or in carotid plaque (p = 0.93). Left ventricular mass index decreased by 3.8 g/m2 (4%) in the ramipril group compared with the placebo group (2p = 0.04). CONCLUSIONS: The results provide no support for the hypothesis that reduced atherosclerosis is responsible for the beneficial effects of ACE inhibitors on major coronary events. It is more likely that the benefits are due to lower BP, reduced left ventricular mass or other factors such as reversal of endothelial dysfunction.

Prenatal diagnosis of dominant dystrophic epidermolysis bullosa, by COL7A1 molecular analysis.

We report the first direct molecular prenatal diagnosis, undertaken for the autosomal dominant form of dystrophic epidermolysis bullosa (DDEB). The proband had a moderately severe form of DDEB, with episodic blistering of skin and mucosal involvement. Diagnostic histopathological examination, using electron microscopy to evaluate skin from a fresh blister, demonstrated a zone of cleavage beneath the epidermal-dermal junction, thereby assigning the EB as dystrophic. DNA analysis of COL7A1, the gene encoding type VII collagen, identified a heterozygous transversion (G to A) in the triple helix domain (G2043R). For any subsequent pregnancy, the affected mother and the unaffected father of the proband requested prenatal prediction, which was thereafter carried out in DNA extracted from a chorionic villus sample obtained at 11 weeks of gestation. Restriction enzyme analysis of COL7A1 exons 73 and 74 amplified by PCR, demonstrated the presence of the G2043R mutation, and the pregnancy was subsequently terminated. Molecular analysis of DNA extracted from fetal tissues confirmed the prenatal prediction.

Artifactual elevation of measured plasma L-lactate concentration in the presence of glycolate.

OBJECTIVES: To determine whether glycolate, a toxic metabolite of ethylene glycol that is chemically similar to lactate, can cause artifactual elevation of measured L-lactate concentrations. DESIGN: Prospective in vitro study. SETTING: Intensive care unit and chemical pathology laboratory in a university-affiliated hospital. SUBJECTS: Heparinized normal human blood and four commercially available L-lactate analyzers. INTERVENTIONS: Four analyzers were tested, three of which used L-lactate oxidase and one of which used L-lactate dehydrogenase. Glycolic acid (10 g/L) in saline was added to blood in a series of aliquots. Corresponding plasma L-lactate concentrations and blood pH, PCO2, and hemoglobin concentrations were measured and base excess was calculated initially and after the addition of each aliquot. One of the two L-lactate oxidase-type analyzers, which was found to show interference, was then used to measure plasma L-lactate and glucose concentrations in blood with glycolic, oxalic, or formic acid added until the base excess was reduced by >15 mmol/L. MEASUREMENTS AND MAIN RESULTS: Artifactual plasma L-lactate elevations were observed in two analyzers, both of the L-lactate oxidase type. Small concentrations of glycolic acid (causing reductions of base excess of 2-5 mmol/L) were accompanied by artifactual plasma L-lactate elevations of 4-8 mmol/L. Artifactual plasma L-lactate elevations increased with further glycolic acid-induced reductions in base excess. Oxalate and formate did not interfere with plasma L-lactate measurements, and measured plasma glucose concentrations were unaffected by all three acids. CONCLUSIONS: Glycolate causes large artifactual elevations in plasma L-lactate measurements by two analyzers in common use, with potential for misdiagnosis of lactic acidosis in ethylene glycol poisoning. A possible cause of the interference is incomplete specificity of the analytical reagent L-lactate oxidase, allowing cross-reaction with glycolate.

Prevalence of lysosomal storage disorders.

CONTEXT: Lysosomal storage disorders represent a group of at least 41 genetically distinct, biochemically related, inherited diseases. Individually, these disorders are considered rare, although high prevalence values have been reported in some populations. These disorders are devastating for individuals and their families and result in considerable use of resources from health care systems; however, the magnitude of the problem is not well defined. To date, no comprehensive study has been performed on the prevalence of these disorders as a group. OBJECTIVE: To determine the prevalence of lysosomal storage disorders individually and as a group in the Australian population. DESIGN: Retrospective case studies. SETTING: Australia, from January 1, 1980, through December 31, 1996. MAIN OUTCOME MEASURE: Enzymatic diagnosis of a lysosomal storage disorder. RESULTS: Twenty-seven different lysosomal storage disorders were diagnosed in 545 individuals. The prevalence ranged from 1 per 57000 live births for Gaucher disease to 1 per 4.2 million live births for sialidosis. Eighteen of 27 disorders had more than 10 diagnosed cases. As a group of disorders, the combined prevalence was 1 per 7700 live births. There was no significant increase in the rate of either clinical diagnoses or prenatal diagnoses of lysosomal storage disorders during the study period. CONCLUSIONS: Individually, lysosomal storage disorders are rare genetic diseases. However, as a group, they are relatively common and represent an important health problem in Australia.

Validation of air as an equilibration medium in gastric tonometry: an in vitro evaluation of two techniques for measuring air PCO2.

This laboratory-based bench study was undertaken to evaluate the accuracy and equilibration characteristics of air and saline respectively as CO2 equilibrating media in the silicone balloon of a gastric tonometer and to compare two methods of measuring air PCO2. Two gastric tonometers were suspended in a bath containing 0.9% saline maintained at 37 degrees C. Certified calibration gases at three different CO2 concentrations were bubbled into the bath. When the bath PCO2 measurement was stable the tonometers were primed with 5 ml of air and 2.5 ml normal saline respectively and allowed to equilibrate for 30 and 90 minutes. Following equilibration, samples were aspirated and analysed in duplicate in a blood gas analyser. Bias and precision were calculated from the measured and expected PCO2 values. A consistent negative bias (21-23%) was seen with air at all three CO2 concentrations at 30 and 90 minutes with a coefficient of variation between 2.7 and 3.3%. Imprecise data were obtained with saline at different levels of CO2. A similar experimental set-up was used to compare air PCO2 measurement by a blood gas analyser and an infra-red analyser (Tonocap). Similar bias was obtained with the blood gas analyser with respect to air PCO2 measurement as in experiment 1. The infra-red analyser measurement was highly precise with negligible bias. Air appears to be a better CO2 equilibration medium during bench testing of tonometry producing a systematic negative offset and requiring a uniform correction factor of 1.25. This correction factor is independent of equilibration time and equilibrating CO2 concentration. The use of the infra-red analyser eliminates any bias in the measurement of air PCO2 and obviates the need for a correction factor.

Chronic copper toxicosis presenting as liver failure in an Australian child.

A 20 month old Caucasian male child, after a five week illness, developed liver failure which was successfully treated by liver transplantation. The explanted liver had a histology identical to that seen in Indian childhood cirrhosis and its copper content was increased tenfold. Water used to prepare the child's milk feeds came from a bore via copper conduits and at times contained 120 mumol/l of copper, eight times the recommended maximum for human consumption. Because non-Indian cases of Indian childhood cirrhosis associated with excess copper ingestion are increasingly being recognised, and as early treatment can restore normal liver morphology, we support the use of the previously suggested alternative term for this condition, ie; 'copper-associated liver disease in childhood'. Measurement of hepatic copper concentrations in all children less than six years of age who develop hepatic failure of unknown cause will increase its recognition. On diagnosis sources of increased dietary copper should be investigated to ensure that younger siblings are not similarly exposed.

Abnormal prostatic cells in ejaculates from men with prostatic cancer--a preliminary report.

OBJECTIVE: To relate findings from a novel approach, ejaculate cytology, to the established reference, histopathology from transrectal ultrasonography (TRUS)-guided prostatic biopsies, in patients at risk of having prostatic cancer on the basis of an abnormal digital rectal examination (DRE) and/or an elevated serum prostate specific antigen (PSA). PATIENTS SUBJECTS AND METHODS: Thirty-seven men suspected of having prostatic carcinoma provided ejaculate specimens which were collected in Hanks solution. The specimens were centrifuged to form a pellet from which smears were made for cytological examination. Immunohistochemical staining for PSA and prostatic acid phosphatase (PAP) were performed on embedded blocks of these cells. TRUS-guided sextant biopsies were performed for histological specimens using standard clinical procedures. A control group of 32 men < 30 years of age, with no family history of prostatic cancer, also produced specimens of ejaculate which were processed similarly. RESULTS: Frankly malignant and atypical prostatic cells were identified in ejaculate specimens from 14 of the 37 patients. Of 12 patients with TRUS biopsies positive for malignancy, nine (75%) had abnormal cells in their ejaculates. Furthermore, five of 25 patients with negative biopsies for adenocarcinoma also had abnormal ejaculate cytology; two of these five patients had high-grade prostatic intra-epithelial neoplasia (PIN). In the control group, no PSA- or PAP-positive prostatic epithelial cells were identified. Normal prostatic cells were not seen in any of the ejaculate specimens examined. CONCLUSIONS: These results indicate that ejaculate cytology, which is a non-invasive and easily repeated investigation, may prove to be a useful approach in the early detection of cancer of the prostate. However, its value in this role, together with the clinical significance of cytological findings, needs to be established, especially in relation to PSA and TRUS biopsy.

Prostate-specific antigen (PSA) and cancer-associated serum antigen (CASA) in distinguishing benign and malignant prostate disease.

The Prostate-Specific Antigen (PSA) and the Cancer-Associated Serum Antigen (CASA) assay for the MUC1 mucin were compared in the serum of 303 patients with malignant or benign prostatic disease. Using cutpoints of 4, 10, and 20 micrograms/l, PSA was elevated in 93%, 81%, and 64% of patients with prostate cancer (n = 113), with corresponding specificities of 55%, 84%, and 96% in benign prostate disease (prostatic hyperplasia or prostatitis, n = 190). Using the recommended cutpoint of 4 Units/ml, CASA was elevated in 38% of patients with prostate cancer, with a specificity of 91% in benign disease. PSA and CASA showed a poor correlation in prostate cancer (r = 0.367) and benign disease (r = 0.158), and CASA was elevated in some PSA negative samples. Used together, PSA > or = 20 micrograms/l and CASA > or = 4 kU/l gave perfect specificity in benign disease, with a corresponding sensitivity of 29% (positive and negative predictive values of 100% and 70%, respectively). However, this combination gave no improvement over the use of PSA alone, with sensitivity 47% when the cutpoint was raised to give perfect specificity. These data suggest that CASA is of little use as an adjunct to PSA in the differentiation of benign and malignant prostate disease.

Paraquat and diquat interference in the analysis of creatinine by the Jaffé reaction.

Paraquat and diquat were shown to interfere significantly in the measurement of plasma creatinine by the alkaline picrate (Jaffé) reaction in a young man who ingested a massive dose of a mixture of the 2 herbicides. It is likely that these bipyridylium compounds react in a manner similar but at different rates compared with creatinine in the Jaffé reaction.

A simple quality control protocol for haemoximetry using standard p50.

We have designed a quality control tool for haemoximetry using the principle that standard p50, calculated from blood gas and haemoximetry measurements using the Siggaard-Andersen algorithm, remains unchanged when a specimen of venous blood is oxygenated progressively to 97% saturation of haemoglobin. If the haemoximeter is inaccurate, progressively larger deviations of standard p50 from the constant value occur above 80% saturation. Deviations too small to be detected by analysis of quality control materials comprised of non-haemoglobin dyes and yet sufficient to cause significant errors in derived extractivity parameters can be detected by this method. The suggested protocol is easy to perform, and is recommended as a quality control tool for blood gas analyser/haemoximeters used to determine arterial extractivity parameters.

Comparative in vitro and in vivo studies of enteric-coated pancrelipase preparations for pancreatic insufficiency.

We evaluated three acid-resistant pancreatic enzyme preparations by in vitro assays, and by comparing degree of steatorrhea, creatorrhea, fecal wet weight, and stool energy losses in a randomized crossover study of patients with pancreatic insufficient cystic fibrosis. Aims of the study were to assess (a) the most practicable and reliable indicator of malabsorption; (b) the variation in enzyme batch potency; (c) the decline in enzyme batch potency with prolonged shelf life; and (d) the relative bioefficacy of the different preparations. In the in vivo study, absorption of energy, nitrogen, and fat did not differ when comparing the three preparations at roughly pharmaceutically equivalent doses, but when expressed per capsule of pancreatic supplement ingested, absorption reflected relative enzyme content, favoring the higher potency preparations. Although steatorrhea was reasonably controlled by these preparations, stool energy losses varied from 800 to 1,100 kJ per day, suggesting greater attention be paid to overall energy absorption rather than absorption of individual nutrients. In addition, fecal energy loss correlated more closely with fecal wet weight (r = 0.81; p < 0.05) than with steatorrhea (r = 0.40; ns), such that 1 g wet feces = 8.37 kJ (+/- 0.14). In vitro enzyme potency varied markedly between batches of the same brand, and also a decline of up to 20% in amylase, lipase, and trypsin activity was noted over an 8-month period for each batch. Both observations have clinical implications at times of represcription. Finally, the higher potency preparations were more effective per capsule and reduced capsule dosage is therefore attainable.

The calculation of reference intervals for oxygen availability parameters from measurements on peripheral venous blood.

An approach to the calculation of reference intervals for oxygen availability parameters using measurements on peripheral blood is described. The technique involves generating the haemoglobin oxygen dissociation curve from a single point using the technique of Siggaard-Andersen et al. and making the appropriate calculation from the curve.

Routine use of hair root or buccal swab specimens for PCR analysis: advantages over using blood.

We report the use of hair roots and buccal cells as specimens of choice for DNA analysis of genetic diseases in a service laboratory. Our protocols using these specimen types show superiority to those using blood specimens in the areas of collection, transport, storage and overall cost. Our experience using these specimen types for 319 cystic fibrosis delta F508 mutation tests and 62 Leber's hereditary optic neuroretinopathy mutation tests leads us to recommend that hair roots and buccal cells should be evaluated as specimens of first choice when developing PCR DNA analysis.