The clinical importance of variation within the HLA-Bw4 complex.

BACKGROUND: Antibody screening of a patient with a failed renal transplant showed positive reactions with most, but not all HLA-Bw4-associated B-locus antigens. However, the patient's serological HLA class I type suggested the presence of HLA-Bw4. METHODS: Standard molecular techniques were used to re-type the patient and donor. ELISA antibody screening helped determine the patient's antibody specificity. RESULTS: The patient's type was HLA-B*1402,4703;Bw6 and the donor HLA-B*4703,51011;Bw4,6. Analysis of ELISA results identified three amino acids (positions 77,80,81) as the most likely epitope recognised by the patient's serum. These corresponded to HLA-B*51011 amino acid mismatches, explaining the lymphocytotoxic reactivity pattern. This epitope is located on a subgroup of the HLA-Bw4 antigen suggesting anti-Bw4 was not a sufficient description of this antibody. CONCLUSIONS: This report identifies an antibody to a sub-group of the Bw4 public specificity and also confirms the need for sequence-level analysis in the tissue-typing laboratory to determine future unacceptable mismatches.

Right ventricular dilatation in the fetus: a study of associated features and outcome.

Right ventricular dilatation is an infrequent finding at fetal echocardiography. Previous studies have documented an association with aortic coarctation. However, there are associations with other congenital abnormalities. We reviewed our experience of fetal right heart dilatation in order to recognize concurrent anomalies and to assess the outcome of the affected fetuses. We studied all fetuses with right ventricular dilatation over a 5-year period. We documented associated cardiac and noncardiac lesions, and outcome data were assessed in terms of the number of fetuses that were born live and the number surviving to 2 months of age. Forty-three fetuses with right heart dilatation were seen. Fifteen had associated cardiac abnormalities: most commonly coarctation (n = 4) and VSD (n = 4). Seven had associated noncardiac abnormalities. There were seven fetuses who also had chromosomal abnormalities. In total, there were three terminations of pregnancy, four intrauterine deaths, one stillbirth and 35 live-births. Twenty-eight were alive at 2 months of age (70% of the nonterminated pregnancies). Fetal right heart dilatation is frequently associated with both cardiac and noncardiac congenital abnormalities. Our outcome figures suggest a guarded prognosis be given during counseling of parents of fetuses with right heart dilatation.

Fetal diagnosis of lethal dysfunction of the right heart in three siblings.

A woman, having already delivered one child, underwent fetal echocardiography during three subsequent pregnancies. All three showed enlargement and poor function of the right-sided chambers. The first was still-born, the second died as a neonate, while the third pregnancy was terminated. Pathological examination revealed the same findings in each fetus, possibly representing a variation of Uhl's anomaly, or alternatively a hitherto unrecognised cardiomyopathic process.

Prenatal diagnosis of congenital heart disease in the northern region of England: benefits of a training programme for obstetric ultrasonographers.

OBJECTIVE: To examine the results of fetal cardiac scanning and audit the changes in performance resulting from the introduction of a training programme for obstetric ultrasonographers. METHODS: Using the database of the Northern Regional Congenital Abnormality Survey (NORCAS), fetuses with complex or significant congenital heart disease (CCHD) diagnosed prenatally in 1994 were identified. A simple programme of centralised and local training was instituted in 1995 by the department of paediatric cardiology to teach obstetric ultrasonographers in district general hospital maternity departments to identify congenital heart malformations. The results of the training programme were assessed by comparing the 1994 identification rate of CCHD with the rates for 1996 and 1997. RESULTS: Birth rate fell during the study from 35,026 in 1994 to 32,874 in 1997. Registration of CCHD also fell, from 115 in 1994 to 87 in 1997. Prenatal recognition of CCHD rose from 17% in 1994 to 30% in 1995 and 36% in 1996. In 1997 it fell slightly to 26.9%. The total number of scans did not change much year on year, but the number of parents choosing termination increased significantly (from 22.7% to 57%). CONCLUSIONS: A simple training programme for obstetric ultrasonographers increased their ability to detect serious congenital heart disease at a routine 18-20 week anomaly scan. With a termination rate of more than 50%, the incidence of CCHD in the population fell from 3.3/1000 to 2.6/1000 live births. This audit, conducted within a stable population using ascertainment by a well established fetal malformation registry, suggests that prenatal diagnosis may have a significant effect on the incidence of complex or serious congenital cardiac malformations.

Detection of HLA class I- and class II-specific antibodies by flow cytometry and PRA-STAT screening in renal transplant recipients.

BACKGROUND: Screening for HLA-specific antibodies has been performed by complement-dependent lymphocytotoxicity for many years. In recent years, methods involving the use of flow cytometry or ELISA have been developed. METHODS: This study has compared a flow cytometric screening technique for the detection of HLA class I- and class II-specific antibodies with a commercially available ELISA technique, PRA-STAT. RESULTS: A significant correlation was found between the two methods for the detection of antibodies in patients after transplantation (P<0.001). Specificity analysis confirmed that the PRA-STAT technique detected both HLA class I- and class II-specific antibodies. Screening of serum samples from patients who experienced graft loss by cytotoxic, flow cytometric, and PRA-STAT techniques showed that there was a significant correlation between all three methods for the detection of antibody, but that the best correlation for the panel-reactive antibody level was that between the flow cytometric and PRA-STAT techniques (r=0.86). This was principally due to the detection of both HLA class I- and class II-specific antibodies by these methods, whereas cytotoxic screening detected only class I-specific antibodies. CONCLUSIONS: These results suggest that PRA-STAT is a useful technique for the detection of both HLA class I- and class II-specific antibodies, rather than only class I-specific antibodies as previously described.

Estimation of pulmonary arterial pressure in the newborn: study of the repeatability of four Doppler echocardiographic techniques.

Despite the increasing use of Doppler echocardiographic (DE) techniques to determine pulmonary arterial pressure in the neonate undergoing intensive care, there have been no studies comparing their repeatability in this population. Our objective was to compare the repeatability of four such techniques in neonates. The study was conducted in two regional neonatal units serving the North East of England. Group A (repeatability between observers): Two experienced observers performed detailed DE examinations, one directly after the other. Group B (within observer repeatability/temporal variability): One observer performed two examinations 1 hour apart. Group A comprised 15 preterm babies (26-36 weeks' gestation, 975-2915 g), most with mild respiratory failure; 4 healthy term babies; and 7 with congenital heart disease, in whom tricuspid regurgitation (TR) only was measured. Their ages were 18 hours to 12 days. Group B comprised 11 babies aged 12-64 hours with moderate to severe respiratory failure; 10 were preterm (26-36 weeks, 785-2800 g). We recorded four measurements: (1) Peak velocity of TR in m/s; (2) peak left-to-right ductal flow velocity (PDAmax in m/s); (3) TPV/RVET ratio; and (4) PEP/RVET ratio, where TPV = time to peak velocity at the pulmonary valve, PEP = right ventricular preejection period, and RVET = right ventricular ejection time. The Bland-Altman analysis was used to produce the coefficient of repeatability (CR: 95% confidence limits of repeatability), also expressed as a repeatability index (CR/mean value) and as a number of "confidence steps"-a measure of sensitivity of the technique to hemodynamic change (range of values within the population/CR). Between-observer and within-observer repeatabilities were similar. Within-observer CR and index (%) results were for TR +/- 0.26 m/s (9%); for PDAmax, +/- 0.48 m/s (39%); TPV/RVET 0.1:1.0 (34%), PEP/RVET 0.12:1.00 (36%). TR and PDAmax had the largest number of confidence steps in the expected range of values (TR 8.5; PDA max 6.5; TPV/RVET 3.2; PEP/RVET 3.2). The most repeatable technique was TR, but PDAmax would also be useful for a serial study owing to the potential for large change. Systolic time interval ratios were less repeatable and likely to be less sensitive indicators of hemodynamic change.

IgG donor-specific crossmatches are not associated with graft rejection or poor graft survival after liver transplantation. An assessment by cytotoxicity and flow cytometry.

Early studies in liver transplantation suggested that there was no association between graft outcome or rejection and the presence of alloantibodies before transplantation. More recent reports have suggested lower graft survival rates and a higher incidence of chronic rejection in patients with IgG warm-T crossmatches. In the present study, panel reactive antibody, direct crossmatch testing, and flow cytometry were used to detect preformed antibodies in sera from 158 consecutive adult recipients of first hepatic grafts. The relationship between preformed antidonor antibodies and liver allograft survival and rejection was determined. Twenty-six (17%) patients were panel reactive antibody (PRA)-positive before transplantation, 22 (15%) had positive donor-specific crossmatches, and 14 (9%) were positive by IgG-specific flow cytometry. Cumulative survival distribution and multivariate analysis failed to reveal any significant associations between overall graft survival and antibody status. Graft survival in patients with PRA-positive sera was 81% compared with 77% for those with PRA-negative sera, 68% for those with positive donor-specific crossmatches compared with 80% for those who were donor-specific crossmatch negative, and 79% for those who were antibody positive by flow cytometric analysis compared with 78% for those who were antibody negative. Subgroup analysis also failed to reveal any significant associations. In addition, Cox proportional hazards regression analysis failed to reveal a relationship between acute or chronic graft rejection with the presence or absence of preformed antibodies, irrespective of immunoglobulin class, cell type (T or non-T), specificity, or technique used for antibody detection. In conclusion, there appears to be no association between either donor-specific or "third-party" alloreactive IgG or IgM antibodies and liver transplant survival or rejection. These data do not indicate a need for prospective crossmatching of liver transplant recipients.

In vitro cytokine profiles and their relevance to rejection following renal transplantation.

Graft rejection remains an important cause of renal allograft failure, despite improvements in immunosuppression and HLA typing. Although HLA matching is beneficial, ensuring an exact match it is often impractical. Thus, a reliable in vitro method for quantitating and qualitating alloreactivity is an important goal. In this study, we measured in vitro the cytokine secretion profiles of mononuclear cells from patients prior to renal transplantation by stimulating with anti-CD3 monoclonal antibody and suppressing with cyclosporine. Mononuclear cells from patients who subsequently developed acute cellular rejection secreted higher mean levels of interleukin (IL)-2 and gamma-interferon (IFN-gamma) than those from patients who had no rejection episodes. IFN-gamma secretion was significantly associated with rejection (P = 0.002), whereas IL-2 secretion did not quite reach statistical significance. There was no significant correlation between IL-4 levels and rejection. Although cyclosporine suppressed the secretion of both IL-2 and IFN-gamma, there was no difference in sensitivity to suppression between rejectors and nonrejectors. These results further emphasize the importance of the TH1 lymphocyte subset in renal allograft rejection. The IFN-gamma secretory capacity of alloreactive T cells may influence the outcome of a renal allograft by (1) activating graft infiltrating macrophages and/or (2) up-regulating HLA molecules on the graft.

Microbubble image enhancement and pericardiocentesis.

When performing pericardiocentesis, if blood-stained fluid is aspirated, one must ensure that the aspirating needle is not within one of the cardiac chambers. To clarify the position of the needle some contrast material may be injected using X-ray screening and for this reason pericardiocentesis is often performed in the catheter laboratory, particularly if the effusion is small, loculated or the heart is in an unusual position. Ultrasound immaging is now commonly used to guide the aspirating needle and if blood-stained fluid is withdrawn we reinject a few millilitres and this outlines the space from which the fluid is withdrawn with microbubbles. This modification of ultrasound immaging adds to the safety of the procedure and allows difficult procedures to be carried out in the ward or intensive care unit, where X-rays screening is not easily available.

Right heart pressure determination by Doppler in infants with tricuspid regurgitation.

Doppler and direct measurements of right ventricle to right atrial pressure drop were made during cardiac catheterisation on 28 occasions in 26 infants with congenital heart disease. Age was 10 days to 12 months (median 4.5 months), and weight was 3.1 to 9.0 kg (median 4.7 kg). We measured peak velocity of tricuspid regurgitation by continuous wave Doppler, and the pressure drop was calculated using the modified Bernoulli equation (delta p = 4v2). There was a high correlation (r = 0.95) between direct and Doppler measurements. Doppler values tended to underestimate the right ventricle to right atrial pressure drop, but this was not of clinical significance (mean 2 mm Hg). The 95% confidence interval for the Doppler velocity was -0.41 to +0.26 m/sec, and was consistent across the range of pressures studied. Variability between observers was tested, by two observers performing sequential paired examinations on 16 newborn babies with tricuspid regurgitation. The coefficient of repeatability was 6.3 mm Hg (95% confidence interval 4.7 to 9.5 mm Hg) or 0.26 m/sec (0.18 to 0.50 m/sec). This method of right ventricular pressure estimation, validated previously only in older children and adults, is a reproducible and accurate technique in infants with tricuspid regurgitation.

Reproducibility of ultrasonic measurement of fetal cardiac haemodynamics.

Two observers made independent estimates of volume flow across all four heart valves in 27 fetuses using a combination of cross-sectional and Doppler echocardiography. There were considerable discrepancies between observers in volume flow estimation arising from inconsistency both in cross-sectional and in Doppler measurements. One observer also produced a second estimate by making new measurements from the original recordings. There was considerable within-observer variability which was mainly due to errors in cross-sectional measurements. The problems of applying this technique to fetal cardiac haemodynamics are discussed.

Dipyridamole echocardiography: the bedside stress test for coronary artery disease.

Identification of dipyridamole-induced regional wall motion abnormalities by echocardiography has recently been proposed as an alternative diagnostic stress test for coronary artery disease. This study evaluates this new technique by comparing the results obtained (overall, regionally and by abnormality type) with those of thallium-201 myocardial imaging after dipyridamole stress in 25 patients. Acceptable echocardiograms were obtained in 20 patients (80%). Concordance of echocardiographic abnormalities for both overall and regional thallium abnormalities was 85%. Sensitivity, specificity and predictive value of dipyridamole echocardiography for overall and regional thallium defects were 92%, 71% and 85%, and 91%, 81% and 85% respectively. However, concordance between the two for abnormality type (i.e. ischaemia versus infarction) was only 66% and the sensitivity, specificity and predictive value of dipyridamole echocardiography for identifying ischaemia as opposed to infarction were only 43%, 82% and 63%, respectively. There was substantial agreement between thallium and echocardiographic imaging after dipyridamole infusion in the diagnosis of coronary artery disease. Echocardiography appears less well able to distinguish infarction from active ischaemia. Dipyridamole echocardiography provides a highly versatile, noninvasive bedside stress test for the detection and localization of coronary artery disease.

Valvar stenosis: a comparison of clinical assessment, echocardiography, Doppler ultrasound and catheterisation.

The relative merits of noninvasive techniques in the assessment of valve stenosis were examined by comparing the results of clinical assessment by two independent clinicians, the cross-sectional echocardiogram and Doppler ultrasound using the results of cardiac catheterisation as reference in 58 patients with a total of 60 stenotic valve lesions. Doppler ultrasound was the most reliable technique; it was correct in 57 (95%) of the 60 lesions. Clinical assessment and cross sectional echocardiography were correct in 48 (80%), and 46 (77%) of the 60 lesions, respectively. In 7 instances 2 noninvasive assessments were wrong in the same patient but on no occasion were all 3 techniques misleading in the same patient. In 17 patients with severe mitral stenosis, clinical assessment Doppler ultrasound and cross-sectional echocardiography were correct in 14 (82%), 16 (94%) and 17 (100%) patients, respectively, whilst in the 4 patients with moderate mitral stenosis the corresponding figures were 3 (75%), 4 (100%) and 2 (50%). In mild mitral stenosis (3 patients), the clinical assessment was correct in 2 (67%) patients, Doppler ultrasound in 3 (100%) patients and cross-sectional echocardiography in 2 (67%) patients. In 22 patients with severe aortic stenosis, the clinical assessment and Doppler ultrasound were correct in every patient (100%), whilst the cross-sectional echocardiogram was correct in 18 (82%) patients. In 11 patients with moderate aortic stenosis, the clinical assessment was correct in only 5 (45%) patients, the cross-sectional echocardiogram in 5 (45%) patients and Doppler assessment in 9 (82%) patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Isometric exercise in the denervated heart: a Doppler echocardiographic study.

The haemodynamic responses to isometric exercise of eight recipients of orthotopic heart transplants and eight healthy controls were studied. Each performed sustained exercise at 30% of maximal voluntary contraction for three minutes on a handgrip dynamometer. Cardiac output was measured by combined Doppler and cross sectional echocardiography before exercise and every 30 seconds during and after exercise. In the controls cardiac output and blood pressure increased significantly owing to an increase in heart rate with no change in stroke volume. In the transplant group cardiac output, heart rate, and stroke volume remained unchanged throughout exercise. In contrast with its response to dynamic exercise the denervated human heart is unable to increase cardiac output during isometric exercise. The pressor response that occurs is mediated via an increase in peripheral vascular resistance.

C-reactive protein, antiproteases and complement factors as objective markers of severity in acute pancreatitis.

In a series of patients with acute pancreatitis we have studied complement factors, antiproteases (alpha 2-macroglobulin and alpha 1-antiprotease) and C-reactive protein to determine the value of their sequential measurement in the prediction of outcome relative to clinical assessment and current multiple factor scoring systems. Complement factors were unhelpful in predicting the severity of an attack. alpha 2-Macroglobulin levels were significantly lower in complicated attacks during days 3-8 and alpha 1-antiprotease levels were significantly higher during days 4-8. C-reactive protein concentrations showed the best discrimination between mild and complicated attacks, levels rising higher and persisting for longer in complicated attacks; these differences were highly significant from day 2 (the morning after admission) to day 8. The concentrations providing the best discrimination were found to be greater than or equal to 210 mg/l for the peak C-reactive protein (on the second, third or fourth day) and greater than or equal to 120 mg/l for the C-reactive protein at the end of the first week. Analysis demonstrated both the peak or seventh-day C-reactive protein concentration to be of similar accuracy to either the Ranson or Glasgow multiple factor scoring systems and slightly better for attacks associated with gallstones. The C-reactive protein assay is simple, quick to perform, provides useful clinical information and is more likely to be of value and to be adopted into routine clinical practice than multiple factor scoring systems.

Reproducibility of cardiac output measurement by cross sectional and Doppler echocardiography.

The variability of Doppler echocardiographic estimation of cardiac output at the aortic orifice was investigated in eight healthy subjects. Cross sectional echocardiograms of the aortic orifice and aortic Doppler velocities were recorded and measured by four echocardiographers. Between subject variability was significantly larger than within subject variability for all variables. Variability owing to different echocardiographers and different measurement times was small compared with total variability. Coefficients of variation for aortic annular diameter, aortic velocity integral, and heart rate were 4.1%, 6.4%, and 5.0% respectively. The coefficient of variation for cardiac output was 8.8% and the 95% confidence interval for measurement of cardiac output by the Doppler method was 4.45 to 6.35 l/min. One echocardiographer reanalysed all the recordings and the results showed that recording the echocardiograms introduces a significantly larger source of error than measuring them. Thus serial measurements of cardiac output by the Doppler method can be performed with acceptable reproducibility; this indicates that the technique can be accurately applied in clinical practice.