Diagnostic accuracy of DXA compared to conventional spine radiographs for the detection of vertebral fractures in children.

OBJECTIVES: In children, radiography is performed to diagnose vertebral fractures and dual energy x-ray absorptiometry (DXA) to assess bone density. In adults, DXA assesses both. We aimed to establish whether DXA can replace spine radiographs in assessment of paediatric vertebral fractures. METHODS: Prospectively, lateral spine radiographs and lateral spine DXA of 250 children performed on the same day were independently scored by three radiologists using the simplified algorithm-based qualitative technique and blinded to results of the other modality. Consensus radiograph read and second read of 100 random images were performed. Diagnostic accuracy, inter/intraobserver and intermodality agreements, patient/carer experience and radiation dose were assessed. RESULTS: Average sensitivity and specificity (95 % confidence interval) in diagnosing one or more vertebral fractures requiring treatment was 70 % (58-82 %) and 97 % (94-100 %) respectively for DXA and 74 % (55-93 %) and 96 % (95-98 %) for radiographs. Fleiss' kappa for interobserver and average kappa for intraobserver reliability were 0.371 and 0.631 respectively for DXA and 0.418 and 0.621 for radiographs. Average effective dose was 41.9 µSv for DXA and 232.7 µSv for radiographs. Image quality was similar. CONCLUSION: Given comparable image quality and non-inferior diagnostic accuracy, lateral spine DXA should replace conventional radiographs for assessment of vertebral fractures in children. KEY POINTS: • Vertebral fracture diagnostic accuracy of lateral spine DXA is non-inferior to radiographs. • The rate of unreadable vertebrae for DXA is lower than for radiographs. • Effective dose of DXA is significantly lower than radiographs. • Children prefer DXA to radiographs. • Given the above, DXA should replace radiographs for paediatric vertebral fracture assessment.

Editor's Choice - Inequalities in Abdominal Aortic Aneurysm Screening in England: Effects of Social Deprivation and Ethnicity.

OBJECTIVE: Population screening for abdominal aortic aneurysm (AAA) in men is currently ongoing in several countries. The aim was to examine the effects of deprivation and ethnicity on uptake of screening for abdominal aortic aneurysm (AAA) and prevalence of AAA. METHODS: This was a review of outcomes from a population screening programme using data collected contemporaneously on a bespoke national database. Men aged 65 in two annual cohorts (2013/14 and 2014/15) were invited for AAA screening. Attendance and prevalence of AAA (aortic diameter >2.9 cm) were recorded. Results were compared according to measures of social deprivation and recorded ethnicity. RESULTS: Some 593,032 men were invited and 461,898 attended for ultrasound screening; uptake 77.9%. Uptake was related to social deprivation: 65.1% in the most deprived decile, 84.1% in the least deprived: OR for least deprived 2.84, 95% CI 2.76-2.92, p<.0001. Men in deprived areas were more likely to actively decline screening: 6% versus 3.8% in the least deprived decile. AAA were twice as common in the most deprived compared with the least deprived decile: OR 2.1, 95% CI 1.77-2.27, p<.0001. AAA were more common in white British men than in black (OR 0.46, 95% CI 0.31-0.71) or Asian (OR 0.18, 95% CI 0.13-0.26) men. There was considerable local variation in all findings. CONCLUSIONS: Social deprivation affects uptake of AAA screening in 65 year old men. Local factors are the most important determinants of uptake, so solutions to improve uptake must be designed at local, not national level.

Impact of the first 5 years of a national abdominal aortic aneurysm screening programme.

BACKGROUND: The Abdominal Aortic Aneurysm (AAA) Screening Programme was introduced by the National Health Service (NHS) in England to reduce the rate of death from ruptured AAA in men. The programme commenced in 2009 and was implemented completely across the country in April 2013. The aim was to review the first 5 years of the programme, looking specifically at compliance and early outcome. METHODS: Men aged 65 years were invited for a single abdominal ultrasound scan. Data were entered into a bespoke database (AAA SMaRT). This was a planned analysis after the first 5 years of the programme. RESULTS: The summation analysis involved the first 700 000 men screened, and the first 1000 men with a large AAA referred for possible treatment. The prevalence of AAA (aortic diameter larger than 2·9 cm) in 65-year-old men was 1·34 per cent. Mean uptake was 78·1 per cent, but varied from 61·7 to 85·8 per cent across the country. Based on the Index of Multiple Deprivation, uptake was 65·1 per cent in the most deprived versus 84·1 per cent in the least deprived areas. Of the first 1000 men referred for possible treatment of a large AAA (greater than 5·4 cm), the false-positive rate was 3·2 per cent. Some 870 men underwent a planned AAA intervention (non-intervention rate 9·2 per cent), with seven deaths (perioperative mortality rate 0·8 per cent). CONCLUSION: The processes in the NHS AAA Screening Programme are effective in detecting and treating men with AAA.

Diagnosis of vertebral fractures in children: is a simplified algorithm-based qualitative technique reliable?

BACKGROUND: Identification of osteoporotic vertebral fractures allows treatment opportunity reducing future risk. There is no agreed standardised method for diagnosing paediatric vertebral fractures. OBJECTIVE: To evaluate the precision of a modified adult algorithm-based qualitative (ABQ) technique, applicable to children with primary or secondary osteoporosis. MATERIALS AND METHODS: Three radiologists independently assessed lateral spine radiographs of 50 children with suspected reduction in bone mineral density using a modified ABQ scoring system and following simplification to include only clinically relevant parameters, a simplified ABQ score. A final consensus of all observers using simplified ABQ was performed as a reference standard for fracture characterisation. Kappa was calculated for interobserver agreement of the components of both scoring systems and intraobserver agreement of simplified ABQ based on a second read of 29 randomly selected images. RESULTS: Interobserver Kappa for modified ABQ scoring for fracture detection, severity and shape ranged from 0.34 to 0.49 Kappa for abnormal endplate and position assessment was 0.27 to 0.38. Inter- and intraobserver Kappa for simplified ABQ scoring for fracture detection and grade ranged from 0.37 to 0.46 and 0.45 to 0.56, respectively. Inter- and intraobserver Kappa for affected endplate ranged from 0.31 to 0.41 and 0.45 to 0.51, respectively. Subjectively, observers' felt simplified ABQ was easier and less time-consuming. CONCLUSION: Observer reliability of modified and simplified ABQ was similar, with slight to moderate agreement for fracture detection and grade/severity. Due to subjective preference for simplified ABQ, we suggest its use as a semi-objective measure of diagnosing paediatric vertebral fractures.

Bayesian hierarchical modeling for a non-randomized, longitudinal fall prevention trial with spatially correlated observations.

Because randomization of participants is often not feasible in community-based health interventions, non-randomized designs are commonly employed. Non-randomized designs may have experimental units that are spatial in nature, such as zip codes that are characterized by aggregate statistics from sources like the U.S. census and the Centers for Medicare and Medicaid Services. A perennial concern with non-randomized designs is that even after careful balancing of influential covariates, bias may arise from unmeasured factors. In addition to facilitating the analysis of interventional designs based on spatial units, Bayesian hierarchical modeling can quantify unmeasured variability with spatially correlated residual terms. Graphical analysis of these spatial residuals demonstrates whether variability from unmeasured covariates is likely to bias the estimates of interventional effect. The Connecticut Collaboration for Fall Prevention is the first large-scale longitudinal trial of a community-wide healthcare intervention designed to prevent injurious falls in older adults. Over a two-year evaluation phase, this trial demonstrated a rate of fall-related utilization at hospitals and emergency departments by persons 70 years and older in the intervention area that was 11 per cent less than that of the usual care area, and a 9 per cent lower rate of utilization from serious injuries. We describe the Bayesian hierarchical analysis of this non-randomized intervention with emphasis on its spatial and longitudinal characteristics. We also compare several models, using posterior predictive simulations and maps of spatial residuals.

Contribution of nonneutralizing vaccine-elicited antibody activities to improved protective efficacy in rhesus macaques immunized with Tat/Env compared with multigenic vaccines.

Previously, chronic-phase protection against SHIV(89.6P) challenge was significantly greater in macaques primed with replicating adenovirus type 5 host range mutant (Ad5hr) recombinants encoding HIVtat and env and boosted with Tat and Env protein compared with macaques primed with multigenic adenovirus recombinants (HIVtat, HIVenv, SIVgag, SIVnef) and boosted with Tat, Env, and Nef proteins. The greater protection was correlated with Tat- and Env-binding Abs. Because the macaques lacked SHIV(89.6P)-neutralizing activity prechallenge, we investigated whether Ab-dependent cellular cytotoxicity (ADCC) and Ab-dependent cell-mediated viral inhibition (ADCVI) might exert a protective effect. We clearly show that Tat can serve as an ADCC target, although the Tat-specific activity elicited did not correlate with better protection. However, Env-specific ADCC activity was consistently higher in the Tat/Env group, with sustained cell killing postchallenge exhibited at higher levels (p < 0.00001) for a longer duration (p = 0.0002) compared with the multigenic group. ADCVI was similarly higher in the Tat/Env group and significantly correlated with reduced acute-phase viremia at wk 2 and 4 postchallenge (p = 0.046 and 0.011, respectively). Viral-specific IgG and IgA Abs in mucosal secretions were elicited but did not influence the outcome of the i.v. SHIV(89.6P) challenge. The higher ADCC and ADCVI activities seen in the Tat/Env group provide a plausible mechanism responsible for the greater chronic-phase protection. Because Tat is known to enhance cell-mediated immunity to coadministered Ags, further studies should explore its impact on Ab induction so that it may be optimally incorporated into HIV vaccine regimens.

Correlation of vaccine-elicited systemic and mucosal nonneutralizing antibody activities with reduced acute viremia following intrarectal simian immunodeficiency virus SIVmac251 challenge of rhesus macaques.

Cell-mediated immunity and neutralizing antibodies contribute to control of human immunodeficiency virus/simian immunodeficiency virus (HIV/SIV) infection, but the role of nonneutralizing antibodies is not defined. Previously, we reported that sequential oral/oral or intranasal/oral (I/O) priming with replication-competent adenovirus type 5 host range mutant (Ad5hr)-SIV recombinants, followed by intramuscular envelope protein boosting, elicited systemic and mucosal cellular immunity and exhibited equivalent, significant reductions of chronic viremia after rectal SIV(mac251) challenge. However, I/O priming gave significantly better control of acute viremia. Here, systemic and mucosal humoral immunity were investigated for potential correlates with the acute challenge outcome. Strong serum binding but nonneutralizing antibody responses against SIV(mac251) were induced in both groups. Antibody responses appeared earlier and overall were higher in the I/O group. Reduced acute viremia was significantly correlated with higher serum binding titer, stronger antibody-dependent cellular cytotoxicity activity, and peak prechallenge and 2-week-postchallenge antibody-dependent cell-mediated viral inhibition (ADCVI). The I/O group consistently displayed greater anti-envelope immunoglobulin A (IgA) antibody responses in bronchoalveolar lavage and a stronger rectal anti-envelope IgA anamnestic response 2 weeks postchallenge. Pre- and postchallenge rectal secretions inhibited SIV transcytosis across epithelial cells. The inhibition was significantly higher in the I/O group, although a significant correlation with reduced acute viremia was not reached. Overall, the replicating Ad5hr-SIV priming/envelope boosting approach elicited strong systemic and mucosal antibodies with multiple functional activities. The pattern of elevated immune responses in the I/O group is consistent with its better control of acute viremia mediated, at least in part, by ADCVI activity and transcytosis inhibition.

Cohort Profile: The Precipitating Events Project (PEP Study).

The Precipitating Events Project (PEP Study) is an ongoing longitudinal study of 754 nondisabled community-living persons age 70 years or older who were members of a large health plan in greater New Haven, Connecticut, USA. The study was established to rigorously evaluate the epidemiology of disability in older persons and to elucidate the role of intervening illnesses and injuries on the disabling process. Of the eligible members, 75.2% agreed to participate and were enrolled between March 1998 and October 1999. Participants have completed comprehensive home-based assessments at 18-month intervals and have been interviewed monthly over the phone with a completion rate of 99%. Detailed participant-level data on health care utilization are obtained annually through linkages with Medicare claims. Through June 2019, 702 (93.1%) participants have died after a median of 109 months, while 43 (5.7%) have dropped out of the study after a median of 27 months. Death certificates are available for all decedents. To date, 117 original reports have been published using data from the PEP Study, including many focusing on other high priority areas such as end of life, frailty, depressive symptoms, aging stereotypes, pain, sleep, and methodologic research. The PEP Study welcomes proposals to access data for meritorious analyses from qualified investigators.

U.s. Competitiveness: beyond the trade deficit.

Large trade deficits and the corresponding increase in U.S. international indebtedness have raised concerns about the long-run competitiveness of the United States. But being competitive requires more than balance in our foreign trade; it requires an improving standard of living. The long-term U.S. competitive problem is largely caused by low saving rates, high costs of capital, and the resulting inadequate level of both visible and invisible investment. As long as the U.S. national saving rate remains far below that of all our major competitor nations, there is little chance for restoring America's international economic position.

Crystal structure of bovine beta-trypsin at 1.5 A resolution in a crystal form with low molecular packing density. Active site geometry, ion pairs and solvent structure.

The crystal structure of bovine pancreatic beta-trypsin (BPT) has been determined from a novel orthorhombic crystal form which contains substantially more solvent (filling 57% of the volume of the unit cell) than previously determined orthorhombic (44%) and trigonal (37%) BPT structures. The native and benzamidine-inhibited crystal structures of BPT in ammonium sulphate at pH 5.3 have been determined for the new form by molecular replacement techniques. The structures have been refined at 1.5 A resolution with final R-values of 16.7% and 16.9%, respectively. Comparison with the previously refined old orthorhombic forms shows that the overall conformation of the protein backbone is highly conserved. A great number of previously undefined side-chains have been located in density. At the C terminus an extra ion pair involving lysines 87 and 107 has been revealed. A far more detailed picture of the ordered solvent structure has been derived. Thirty water clusters have been identified. A large water network extends from the calcium binding site to the activation area and the autolysis loop. There is evidence for a water channel reaching from the depth of the specificity pocket to the nearby protein surface which might be involved in the displacement of water molecules upon substrate binding. A sulphate anion which forms hydrogen bonds to the active site residues His57, Ser195 and Gly193 was for the first time positioned in clearly defined electron density. Interaction with the sulphate ion may explain the increase in the pKa value of His57 at high sulphate concentrations which was observed by nuclear magnetic resonance studies of a bacterial serine protease both in crystalline form and in solution. Thus, a His-Ser hydrogen bond will not exist in solvents containing sulphate at low pH (up to at least 6.8) where the imidazole of His57 is protonated. The new crystal form is of considerable interest for substrate binding studies. Wide solvent channels should allow diffusion of large substrates (comparable in size to, e.g. pancreatic trypsin inhibitor) into the enzyme crystal. The active site is accessible; intermolecular contact areas are further remote from the active site than in the old orthorhombic form.

X-ray analysis of the eye lens protein gamma-II crystallin at 1.9 A resolution.

We report the X-ray structure analysis and refinement at 1.9 A resolution of calf gamma-II crystallin, a lens-specific protein. The sequence of Croft (1972) has been modified to give a polypeptide chain of 174 residues (cf. 165). The protein has a symmetrical, hierarchical structure of two globular domains each comprising two similar "Greek key" motifs, consecutive along the polypeptide chain, and related by a pseudo 2-fold axis. The two domains pack together with a single connection and are related by a further pseudo 2-fold axis which bisects the angle between the intra-domain dyads. Forty-two pairs of C alpha positions for the two most similar motifs have root-mean-square separation at best fit of 0.69 A. The N and C-terminal domains gave root-mean-square separation of 0.89 A for 82 pairs of C alpha atoms at best fit. In each domain the two Greek key motifs form a pair of four-stranded antiparallel beta-pleated sheets, each sheet composed of three stands from one motif and one from the other. The sheets pack together in a wedge shape, closed at the top by the loops connecting the third and fourth strands of each motif. The first two strands of each motif form an extended beta-hairpin which is folded on to the beta-sheet. The packing of each motif into the globular domains involves a staggered bilayer of side-chains between each pair of beta-sheets which does not preserve the pseudo 2-fold axes observed in the C alpha position topology. In the core of each domain there are interactions between polarizable aromatic groups and sulphur-containing residues which may contribute to stability and may also serve to protect aromatic side-chains from ultraviolet light damage in the lens. At the surface of the molecule over half the ionic side-chains are closely paired, which probably stabilizes the tertiary fold and may reduce the water bound. Crystal lattice interactions are described which may be similar to those occurring in vivo in the lens between crystallins. Seven cysteine residues have been identified in the structure and these may have a role in the thermodynamic stability of the molecule, its intermolecular interactions under the normal reducing conditions of the lens, and also in the aggregation and cross-linking which occur in some forms of cataract. Three of these residues, Cys18, Cys23 and Cys74, form a cluster in the N-terminal domain.(ABSTRACT TRUNCATED AT 400 WORDS)

Crystals of the NC1 domain of human type IV collagen.

Crystals of the non-collagenous C-terminal region (NC1) of type IV collagen have been obtained from human placenta. These crystals diffract to 2.0 A, and belong to space group P22(1)2(1), with cell dimensions a = 81 A, b = 158 A, c = 138 A, alpha = beta = gamma = 90 degrees. The crystals contain one hexamer in the asymmetric unit; they are very stable with respect to X-rays.

Cognitive and other adverse effects of diphenhydramine use in hospitalized older patients.

BACKGROUND: Diphenhydramine hydrochloride is a commonly prescribed medicine in hospitalized patients, but its adverse effects on older patients remain unclear. METHODS: We enrolled 426 hospitalized medical patients aged 70 years or older in a prospective cohort study in a university hospital. Measurements included baseline and daily assessments including Mini-Mental State Examination scores, Confusion Assessment Method ratings, direct observations for medical devices (urinary catheter or physical restraints), and blinded medical record extractions for diphenhydramine use. RESULTS: Of the 426 patients, 114 (27%) received diphenhydramine during hospitalization and shared similar baseline characteristics including age, sex, delirium risk, and Mini-Mental State Examination scores compared with nonexposed patients. The diphenhydramine-exposed group was at an increased risk for any delirium symptoms (relative risk [RR], 1.7; 95% confidence interval [CI], 1.3-2.3) and for individual delirium symptoms, including inattention (RR, 3.0; 95% CI, 1.5-5.9), disorganized speech (RR, 5.5; 95% CI, 1.0-29.8), and altered consciousness (RR, 3.1; 95% CI, 1.6-6.1). Exposed patients also had increased risk for urinary catheter placement (RR, 2.5; 95% CI, 1.0-6.0) and longer median length of stay (7 vs 6 days; P =.009). A dose-response relationship was demonstrated for most adverse outcomes. Overall, 24% of diphenhydramine doses were administered inappropriately. CONCLUSIONS: Diphenhydramine administration in older hospitalized patients is associated with an increased risk of cognitive decline and other adverse effects with a dose-response relationship. Careful review of its use is necessary in this vulnerable population.

Multivariate flow cytometric characterization and enrichment of murine megakaryocyte progenitors (CFU-Meg).

Multivariate flow cytometric analysis and sorting of mouse bone marrow cells viably stained with Hoechst 33342 (HO) was utilized to obtain a subpopulation enriched at least 30- to 60-fold for clonogenic megakaryocyte precursors (CFU-Meg). HO fluorescence intensity, forward light scatter, and perpendicular light scatter were measured on each HO-stained bone marrow cell as it passed through a dual beam flow cytometer. Cells were sorted on the basis of their light scattering and HO fluorescence intensity properties. The sorting region which included cells with high forward light scatter, low perpendicular light scatter, and low (0.1-0.65 times the modal channel) HO fluorescence intensity contained approximately 0.6% CFU-Meg. Of all CFU-Meg, 75% were contained in the light scatter gate, and 56% of all CFU-Meg were in the low HO-sorting region. This technique offers a simple, one-step procedure to obtain preparations enriched in murine clonogenic megakaryocyte precursors.

Enrichment of murine hemopoietic clonogenic cells by multivariate analyses and sorting.

Multivariate analyses, dual beam flow cytometry and sorting, and list mode data processing were used to distinguish and enrich committed and pluripotent stem cells in mouse bone marrow. These cells were discriminated on the basis of their forward angle and perpendicular light scatter characteristics and their Hoechst 33342 fluorescence intensity. Myeloid committed progenitors (CFU-GM) and spleen colony-forming units (CFU-S) (day 9 and day 13) were enriched 100-fold by sorting on the basis of high forward angle light scatter, intermediate perpendicular light scatter, and very low HO fluorescence intensity. Approximately 10% of the sorted cells formed colonies in the CFU-GM assay and 2% formed CFU-S colonies. Morphologic analysis of the sorted subpopulation revealed 92% blast immature cell types. The DNA distribution of the sorted subpopulation, assessed by propidium iodide staining, indicated that 98% of the progenitor-enriched subpopulations contained 2N DNA content. This separation procedure offers a simple method to obtain preparations highly enriched in clonogenic cells in one pass through the cell sorter.

The relationship between onsets of depression and sudden drops in solar irradiation.

In a retrospective single-case study of a patient (L.S., the first author), who suffers from recurrent depression, we compared the onsets of his depression with meteorological data on light intensity, independently obtained, for the years 1975-1988. During those years there were 24 onsets or worsening of an existing depressive episode. Eleven of these occurred in the months September-December. Seven of these 11 autumn/winter onsets and 12 of the remaining 13 non-autumn/winter onsets coincided with a period of unusually low natural light intensity. Thus, 19 of 24 onsets (79%) were associated with falling light intensity, regardless of season. Some episodes that were not related to falling light intensity were associated with life events. For the years prior to the patient's use of light therapy, there was a statistically significant relationship between falling light and onset of depression. This supports the suggestion that depressive episodes (in certain individuals) may be triggered by unusually dull weather, regardless of season.

A computer graphics model of frog gamma-crystallin based on the three-dimensional structure of calf gamma-II crystallin.

Molecular models for Rana gamma-1 and gamma-2 crystallins have been constructed using computer graphics on the basis of the protein primary structure derived from the complementary DNA sequence and the three-dimensional structure of calf gamma-II crystallin that has been defined at high resolution by X-ray analysis. The models show that the cores of the two domains are conserved as hydrophobic, with the polypeptide chain arranged as a four Greek-key motif structure. Although many lysines replace arginines at equivalent positions in mammalian proteins, the Rana crystallins also have an extensive series of ion pairs on their surface; these are strongly implicated in their function as stable structural molecules, which are highly conserved in the evolution of the vertebrate eye lens.

Uptake of individual fatty acids into adipose tissue in relation to their presence in the diet.

BACKGROUND: The fatty acid composition of adipose tissue triacylglycerol reflects, but is not identical to, the fatty acid composition of the habitual diet. OBJECTIVE: We investigated whether the fatty acid composition of adipose tissue is explained by differences between fatty acids in early storage in adipose tissue after a meal. DESIGN: Nine healthy men ate a meal containing several fatty acids. Blood samples were taken for 6 h after the meal from an arterialized hand vein and a vein draining the anterior abdominal subcutaneous adipose tissue. RESULTS: Net storage of fatty acids in adipose tissue occurred between 1 and 4 h after the meal. In relation to the amount fed, storage of fatty acids differed (P < 0. 01) between classes (n-3 polyunsaturated < saturated < n-6 polyunsaturated < monounsaturated); oleic acid was stored in the greatest amounts. These differences agreed closely with published data, except for n-3 polyunsaturated fatty acids. The only individual metabolic step at which significant differences between fatty acids was shown was incorporation of fatty acids into chylomicron triacylglycerol. Differences between fatty acids in rate of extraction from chylomicron triacylglycerol and net uptake into adipose tissue in the postprandial period were significant (P < 0. 01), but not when expressed in relation to proportions in chylomicron triacylglycerol. CONCLUSIONS: The characteristic fatty acid pattern of adipose tissue may predominantly reflect the early metabolic handling of different fatty acids. Adipose tissue uptake of n-3 polyunsaturated fatty acids is slow in relation to that of other fatty acids.

Impaired postprandial tissue regulation of blood flow in insulin resistance: a determinant of cardiovascular risk?

The insulin resistant state is a major risk factor for coronary artery disease. This increased risk is likely to be due to associated lipid and coagulation abnormalities rather than just abnormalities in glucose metabolism or hyperinsulinaemia alone. Exaggerated postprandial lipaemia is a well-recognised associate of insulin resistance and postprandial hypertriglyceridaemia is particularly important in the development of coronary atheroma. It seems likely that insulin is one of the hormonal regulators of adipose tissue and skeletal muscle blood flow. The reduced blood flow and blunting of the postprandial rise of peripheral blood flow in insulin resistance may decrease chylomicron-triglyceride delivery to muscle in subjects with insulin resistance. This, in turn, will lead to increased production of atherogenic particles. We propose that impaired postprandial vasodilation, already recognised as a key feature of glucose intolerance, is also the cause of impaired lipid metabolism in insulin resistant subjects and predisposes them to cardiovascular disease.

Quantitative flow cytometric and clonogenic evaluation of glass bead affinity fractionation of antibody-labeled murine bone marrow.

Monoclonal antibodies and a glass bead affinity fractionation technique were used to selectively deplete subpopulations of murine bone marrow cells. Flow cytometric analyses permitted quantitative measurement of subpopulation depletion and characterization (light scatter and fluorescence intensity) of both the eluted and bound cell subpopulations. Mouse bone marrow cells were labeled with selected monoclonal rat anti-mouse antibodies directed against cell surface antigens and were eluted through a glass bead column coated with goat anti-rat immunoglobulin. Unlabeled cells passed through the column, whereas cells labeled with the antibody were selectively retained. Column operating conditions for optimal depletion of labeled cells were determined. With specific column conditions, 97% of the antibody positive cells were retained on the column. In addition, clonogenic assays on cells sorted from unfractionated and column fractionated preparations provided estimates of the fraction of granulocyte macrophage progenitors (CFU-GM) in the different cell subpopulations. The enrichment of CFU-GM achieved in the eluted cell populations was dependent upon the antibody used for cell labeling and ranged from four- to six-fold. Since large numbers of cells can be processed rapidly, this technique, in combination with antibodies specific for non-clonogenic cells, is particularly suitable when preparations enriched in colony-forming progenitors are required.