The significance of low bcl-2 expression by CD45RO T cells in normal individuals and patients with acute viral infections. The role of apoptosis in T cell memory.

The bcl-2 gene product has been shown to prevent apoptotic cell death. We have now investigated the bcl-2 protein expression by resting and activated mature T cell populations. Freshly isolated CD45RO+ T cells within CD4+ and CD8+ subsets expressed significantly less bcl-2 than CD45RO- (CD45RA+) T cells (p < 0.001). When CD45RA+ T cells within both CD4+ and CD8+ subsets were activated in vitro, the transition to CD45RO phenotype was associated with a decrease in bcl-2 expression. Patients with acute viral infections such as infectious mononucleosis caused by Epstein-Barr virus infections or chickenpox, resulting from varicella zoster virus infection, had circulating populations of activated CD45RO+ T cells which also showed low bcl-2 expression. In these patients, a significant correlation was seen between low bcl-2 expression by activated T cells and their apoptosis in culture (r = 0.94, p < 0.001). These results suggest that the primary activation of T cells leads to the expansion of a population that is destined to perish unless rescued by some extrinsic event. Thus the suicide of CD45RO+ T cells could be prevented by the addition of interleukin 2 to the culture medium which resulted in a concomitant increase in the bcl-2 expression of these cells. Alternatively, apoptosis was also prevented by coculturing the activated T lymphocytes with fibroblasts, which maintained the viability of lymphoid cells in a restinglike state but with low bcl-2 expression. The paradox that the CD45RO+ population contains the primed/memory T cell pool yet expresses low bcl-2 and is susceptible to apoptosis can be reconciled by the observations that maintenance of T cell memory may be dependent on the continuous restimulation of T cells, which increases their bcl-2 expression. Furthermore, the propensity of CD45RO+ T cells to extravasate may facilitate encounter with fibroblast-like cells in tissue stroma and thus be an important additional factor which promotes the survival of selected primed/memory T cells in vivo.

Genetic diversity in farm animals--a review.

Domestication of livestock species and a long history of migrations, selection and adaptation have created an enormous variety of breeds. Conservation of these genetic resources relies on demographic characterization, recording of production environments and effective data management. In addition, molecular genetic studies allow a comparison of genetic diversity within and across breeds and a reconstruction of the history of breeds and ancestral populations. This has been summarized for cattle, yak, water buffalo, sheep, goats, camelids, pigs, horses, and chickens. Further progress is expected to benefit from advances in molecular technology.

Inhibition of T cell apoptosis in the rheumatoid synovium.

Synovial T cells in rheumatoid arthritis are highly differentiated and express a phenotype suggesting susceptibility to apoptosis (CD45RB dull, CD45RO bright, Bcl-2 low, Bax high, Fas high). However, no evidence of T cell apoptosis was found in synovial fluid from any of 28 patients studied. In contrast, synovial fluid from 10 patients with crystal arthritis showed substantial levels of T cell apoptosis. The failre of apoptosis was not an intrinsic property of rheumatoid synovial T cells, as they showed rapid spontaneous apoptosis on removal from the joint. Synovial T cells from rheumatoid arthritis and gout patients could be rescued from spontaneous apoptosis in vitro either by IL-2R gamma chain signaling cytokines (which upregulate Bcl-2 and Bcl-XL) or by interaction with synovial fibroblasts (which upregulates Bcl-xL but not Bcl-2). The phenotype of rheumatoid synovial T cells ex vivo (Bcl-2 low, Bcl-xL high) suggested a fibroblast-mediated mechanism in vivo. This was confirmed by in vitro culture of synovial T cells with fibroblasts which maintained the Bcl-xL high Bcl-2 low phenotype. Synovial T cells from gout patients were Bcl-2 low Bcl-xL low and showed clear evidence of apoptosis in vivo. Inhibition experiments suggested that an integrin-ligand interaction incorporating the Arg-Gly-Asp motif is involved in fibroblast-mediated synovial T cell survival. We propose that environmental blockade of cell death resulting from interaction with stromal cells is a major factor in the persistent T cell infiltration of chronically inflamed rheumatoid synovium.

The kinetics of interaction between lymphocytes and magnetic polymer particles.

Magnetic polymer-coated particles linked to antibodies are considered to be an efficient rosetting matrix for immunoselection. We have shown that a 20:1 bead:target cell ratio and a 90 min incubation period are the optimal conditions for specific binding of monoclonal antibody-labelled cells to goat anti-mouse IgG-coated beads. Higher ratios or longer incubation periods resulted in considerable non-specific binding. Characterisation of the optimal conditions for specific depletion of lymphocyte subpopulations showed that (a) a range of bead:target cell ratios and incubation periods can be used, with resulting high efficiency and specificity; (b) multiple monoclonal antibodies can be used simultaneously for the depletion of diverse lymphocyte subpopulations; (c) non-specific bead-to-cell binding does not affect the specificity and efficiency of magnetic depletion; (d) specific binding of one bead only was adequate for effective magnetic separation. These findings define the most economical, specific and efficient conditions of use of beads for negative immunoselection but preclude the use of beads as an analytical rosetting medium.

Quantitative flow cytometry for the analysis of T cell receptor Vbeta chain expression.

Detailed characterisation of the T cell receptor (TCR) repertoire expressed by peripheral blood lymphocytes has been used to study specific T cell responses in disease conditions. The methods have mostly involved molecular biology analysis of transcribed gene products isolated from T cell subsets or individual clones. Extensive characterisation of the TCR Vbeta chain repertoire by flow cytometry is now possible due to the recently increased availability of specific monoclonal antibodies. However, there are major logistical problems inherent in this analysis relating to the number of cells required to obtain accurate results and the vast amounts of data generated. To reduce these factors to a practical level, we have performed a detailed study to define the limits of precision of cell subset analysis by flow cytometry. Maximal achievable precision was obtained by analysing 10(4) lymphocytes; no significant improvement was obtained by analysing greater numbers of cells up to 10(5) cells, even for cell subsets present at frequencies as low as 0.5%. Careful application of these precision profiles will also permit more effective use of clinical research samples for flow cytometry when the availability of cells is limited.

Purification of human blood eosinophils by negative selection using immunomagnetic beads.

A simple method for isolating highly purified eosinophils from human blood is described. Buffy coats from normal individuals (eosinophil counts less than 0.4 x 10(9)/litre) were centrifuged through a two layer Percoll density gradient, to produce a granulocyte fraction containing neutrophils and eosinophils. Neutrophils were extracted from this fraction using a monoclonal antibody (CLB FcR gran 1) against CD 16 (Fc gamma R III) in a direct or indirect selection procedure using immunomagnetic beads (Dynabeads). This negative immunoselection produced eosinophils of greater purity and with a superior capacity to mount a respiratory burst than eosinophils isolated by a method employing metrizamide.

Growth impairment in very low birthweight children at 12 years: correlation with perinatal and outcome variables.

AIM: To compare the growth of very low birthweight (VLBW) children in early adolescence with that of their normal birthweight peers; to examine the role of factors contributing to growth-parental height, perinatal variables, bone maturity and sexual maturation; to examine the correlation between head growth and cognitive and educational outcome. METHODS: Standing and sitting heights, weight, occipito-frontal circumference (OFC), skinfold thicknesses and pubertal staging were assessed in 137 VLBW children and 160 controls at 11-13.5 years of age. Ninety six (70%) of the VLBW children had their bone age assessed using the TW2 method. Reported parental heights were obtained by questionnaire. All children had standardised tests of cognitive and educational ability. Perinatal data had been collected prospectively as part of a longitudinal study. RESULTS: VLBW children had lower heights, weight, and OFC. Skinfold thicknesses were no different. The children's short stature was not accounted for by difference in parental height, degree of pubertal development, or by retarded bone age. Indeed, the TW2 RUS score was significantly advanced in the VLBW children. Using the bone ages to predict final adult height, 17% have a predicted height below the third centile and 33% below the tenth. Weight was appropriate for height, but there was a residual deficiency in OFC measurements after taking height into account. In the VLBW group smaller head size was associated with lower IQ and mathematics and reading scores. CONCLUSIONS: Growth problems persist in VLBW children and final heights may be even more abnormal than present heights suggest. VLBW children have smaller OFCs than expected from their short stature alone and this may be associated with poorer educational and cognitive outcomes.

Early antenatal ultrasound diagnosis of fetal intracranial teratoma.

The commonest fetal intracranial tumour is a teratoma. The prognosis is poor with death usually occurring shortly after birth. Modern high resolution ultrasound scanners facilitate examination of the cranial contents, allowing earlier diagnosis. We report a case where an intracranial teratoma was identified at 21 weeks gestation, the earliest gestational age that this has been reported. The ultrasound appearances are discussed.

The radiology of Meconium Ileus Equivalent.

Meconium Ileus Equivalent is a condition which occurs in patients with cystic fibrosis where the ileum becomes obstructed by bowel content of abnormal consistency. Intercurrent chest infection or absence of pancreatic extract supplements are often the exacerbating cause. Plain films of the abdomen show small bowel obstruction and in some a nodular pattern in the small bowel, the nodules being larger than those seen in quiescent cystic fibrosis. Gastrografin enemas and/or enemas with acetyl cysteine can be used in the treatment of the condition, the Gastrografin having the advantage of confirming the diagnosis radiologically. The condition is important as it can be the presenting feature of cystic fibrosis in childhood and even in early adult life, and the operative mortality and morbidity are high. Conservative treatment should be pursued vigorously and operation only undertaken if unavoidable.

The relationship of pulmonary interstitial emphysema to subsequent type of chronic lung disease.

Chronic lung disease of prematurity (CLD) can be classified using the chest radiograph taken on day 28 into type 1 (homogeneous opacification) or type 2 (the presence of cystic change) and it has been suggested that the presence of pulmonary interstitial emphysema (PIE) leads to the development of type 2 CLD. To further study this relationship we examined the chest radiographs taken on days 1-7, 14, 21 and 28, of 202 infants treated at Liverpool Maternity Hospital between January 1980 and December 1989 who developed CLD. 39% (54/137) of infants who developed type 1 CLD had suffered from PIE compared with 78% (51/65) with type 2 CLD (p < 0.001). There was no difference in the incidence of PIE in infants who developed CLD and subsequently died compared with those who survived. In the infants who survived there was no significant difference in length of oxygen dependency between those who suffered from PIE (median 99 days oxygen dependent) and those who had no PIE (median 105 days). We conclude that the presence of PIE is associated with subsequent development of type 2 CLD but is not a prerequisite for this radiographic type of chronic lung disease. Of those infants who survive to develop CLD, the presence of PIE does not alter prognosis in terms of duration of oxygen dependency or mortality.

The incidence and significance of testicular microlithiasis in a subfertile population.

The objective of this study was to assess the frequency of testicular microlithiasis (TM) in infertile men who underwent testicular ultrasound and to determine any causative or associated factors. The case notes of 159 consecutive patients who were referred for testicular ultrasound in the investigation of male factor infertility were reviewed. Microcalcification was found in 10 cases (6.2%). This was unilateral in all cases and six patients had clinical evidence of a varicocele. Five cases had minimal calcification and five had marked TM. On patient had a past history of testicular maldescent and another of testicular torsion. Sperm function (as assessed by sperm count, motility and the sperm migration test) was variable within the 10 patients and there was no correlation with hormone profiles or testicular size. There was a statistical difference between a number of investigations in those patients with minimal degrees of calcification and those with TM (sperm migration test (SMT), namely sperm migration and sperm motility (p < 0.05, Mann-Whitney U test)). The results showed a higher than expected incidence of TM. Patients with marked TM seemed to have poorer sperm function than those with minimal calcification. There was a high incidence of co-existing pathology, for instance scrotal varicocele and cryptorchidism, although the numbers in this study were small and further studies need to be carried out to determine the exact nature and significance of this condition.

The clinical significance of fetal right heart enlargement with a structurally normal heart.

Isolated right heart enlargement of the fetus without cardiac structural abnormalities appears to be associated with growth retardation or abnormal perinatal outcome. We report the outcome of eight fetuses with subjective enlargement of the right heart. All cases were diagnosed antenatally from a standard four-chamber view on an ultrasound scan. All fetuses had otherwise normal cardiac anatomy. Seven out of the eight fetuses ultimately had outcomes which deviated from the norm. An isolated finding of fetal right heart enlargement warrants close antenatal fetal surveillance.

Current imaging of childhood urinary infections: prospective survey.

OBJECTIVES: To assess whether ultrasonography alone is adequate for routine screening of childhood urinary infection, whether clinical features determine the need for further investigations, and which investigations are most appropriate. DESIGN: Prospective survey of children with proved urinary infection and a preinvestigation record of clinical features. Ultrasonography and intravenous urography were routine, with choice of further studies determined by ultrasonographic findings. SETTING: A children's hospital and two district general hospitals in Mersey region. MAIN OUTCOME MEASURES: Sensitivity and specificity of ultrasonography both generally and in relation to clinical features. Accuracy of intravenous urography compared with radioisotope examinations. RESULTS: Specificity of ultrasonography was good (99% (95% confidence interval 96% to 100%)) but sensitivity modest (43% (32% to 55%)), principally with respect to detecting vesicoureteric reflux and renal scarring. Among older children (aged 2-10 years) with positive ultrasound results and fever or vomiting the sensitivity in detecting reflux (with and without renal scarring) was 78% (62% to 89%) and the specificity 69% (60% to 78%); in detecting renal scarring (with and without reflux) the sensitivity was 100% (80% to 100%) and specificity 65% (56% to 74%). Renal scarring and obstructive uropathies were better assessed by radioisotope examinations than by intravenous urography. CONCLUSIONS: Ultrasonography alone is inadequate for routine screening of childhood urinary infection. Though further investigations remain advisable in infants, in older children they can be restricted to a minority who have positive ultrasound examinations or have had fever or vomiting. Radioisotope examinations largely eliminate the need for intravenous urography.

Interpretation of the neonatal chest X-ray.

Most neonatal X-rays are seen initially by a paediatrician without formal training in interpretation of chest X-rays. This article aims to help improve the information obtained from these X-rays which are often complex. Many factors affect accurate interpretation of the neonatal chest X-ray, including good quality radiographs, appropriate viewing conditions and thorough education.

The Skills and Employment Opportunities Project: report of an evaluation.

OBJECTIVES: the research was designed to evaluate the success of a Project set up to find employment or training in ordinary firms or organisations for people with disabilities. METHODS: three main sources of data were used: the Project's data-base, which included information on client 'outcomes'; postal questionnaires and interviews with Project clients; postal questionnaire and interviews with referrers to the Project. RESULTS: the rate of referral to the Project vastly exceeded expectations. According to the Project's 'outcome' categories, 66% of the clients were successfully placed, and clients' accounts verified this. The expectations of referrers of people with learning disabilities for their clients exceeded 'outcomes' but referrers of people with physical disabilities under-estimated them. Clients' views were generally favourable but they would have liked more choice and aspired to improving their work placements. CONCLUSIONS: The evidence indicated that the Project had a role in the area not filled by other agencies.