Patterns of CD16 and CD56 expression in persistent expansions of CD3+NKa+ lymphocytes are predictive for clonal T-cell receptor gene rearrangements

Phenotypic characteristics, and correlations between the expression of membrane NK-associated (NKa) determinants (CD11b, CD16, CD56 and CD57) and T cell receptor (TCR) genotypic patterns, were examined in 25 patients with persistent (greater than 6 months) expansions of CD3+WT31+NKa+ (CD8+ and CD8dim+) lymphocytes. These studies showed that distinct NKa phenotypic profiles were restricted to cases with rearranged TCR configurations and that clonal CD3+NKa+ components could be predicted in most cases by assessing relationships between membrane CD16 and CD56 expression. For all normal NKa subpopulations, there was a high correlation (P less than 0.0001; n = 31) between the expression of these two membrane determinants. Markedly increased CD16 expression by CD3+NKa+ cells, in relation to CD56 (i.e. a high CD16:CD56 ratio), was found exclusively in cases with rearranged TCR (13/16 cases); 2/3 of the remaining cases showing significantly reduced CD16:CD56 ratios and high (greater than 2.0) CD3+CD56+ absolute numbers. In contrast, 7/9 of the germline TCR cases had a normal CD16:CD56 ratio and 2/9 a decreased ratio with low (less than 1.0) CD3+CD56+ absolute numbers. A high ratio of CD16:CD56 expression by CD3+NKa+ lymphocytes was therefore informative for 82% of TCR rearrangements in this series; and analysis of CD16 and CD56 expression was predictive for germline and rearranged TCR configurations in 24/25 persistent CD3+NKa+ expansions.     

Expression profiling of metalloproteinases and inhibitors in cartilage

Objective   To profile the expression of all known members of the matrix metalloproteinase ( MMP ), a disintegrin and metalloproteinase with thrombospondin motifs ( ADAMTS ), and tissue inhibitor of metalloproteinases ( TIMP s) gene families in normal cartilage and that from patients with osteoarthritis (OA).
Methods   Human cartilage was obtained from femoral heads at joint replacement for either osteoarthritis or following fracture to the neck of femur. Total RNA was purified and expression of genes assayed using quantitative real-time PCR.
Results   Several members of the above gene families were regulated in OA. Genes increasing in expression in OA were: at P  < 0.001, MMP-13 , MMP-28 , ADAMTS-16 ; at P  < 0.01, MMP-9 , MMP-16 , ADAMTS-2 , ADAMTS-14 and at P  < 0.05, MMP-2 , TIMP-3 , ADAMTS-12 . Genes decreasing in expression in OA were: at P  < 0.001, MMP-1 , MMP-3 , ADAMTS-1 ; at P  < 0.01, MMP-10 , TIMP-1 , ADAMTS-9 and at P  < 0.05, TIMP-4 , ADAMTS-5 , ADAMTS-15 . Correlation analysis revealed that groups of genes across the gene families are co-expressed in cartilage.
Conclusion   This is the first comprehensive expression profile of all known MMP , ADAMTS and TIMP genes in cartilage. Patterns of expression provide a foundation on which to understand mechanisms of gene regulation in OA and potentially for refining the specificity of anti-proteolytic therapies.     

Impaired Microvascular Function in Normal Children: Effects of Adiposity and Poor Glucose Handling

Clustering of cardiovascular risk factors is thought to occur early in life. The endothelium is an important regulator of microvascular function. We investigated the relationship between microvascular function and cardiovascular risk factors in 145 normal, healthy children aged 11-14 years. Skin microvascular responses, measured using laser Doppler imaging, to iontophoresis of acetylcholine (ACh) and sodium nitroprusside (SNP), were negatively correlated with percentage body fat ( r =−0.20, P < 0.05 and r =−0.18, P < 0.05, respectively). Subjects were stratified into quintiles based on 2-h, post-feeding glucose levels. Subjects in the upper glucose quintile (range 7.4-11.4 mmol l⁻¹) showed significantly lower vasodilatation to both ACh (   P < 0.005  ) and SNP (   P < 0.02  ) than those in the lower quintile (range 3.9-4.9 mmol l⁻¹). Waist-to-hip ratio and the fasting insulin resistance index were significantly greater in subjects in the upper quintile than those in the lower quintile ( P < 0.001 and P < 0.05, respectively). Additionally, in subjects in the upper glucose quintile, fasting triglyceride correlated with fasting insulin ( r = 0.59, P < 0.001) and with the fasting insulin resistance index ( r = 0.49, P < 0.009), and plasma levels of cholesterol and 2-h glucose were also correlated ( r = 0.40, P < 0.05). In a cross-section of normal children, microvascular function was negatively associated with adiposity. Additionally, in a subgroup of subjects, there was a clustering of high post-feeding glucose, impaired microvascular function, increased insulin resistance and higher central fat distribution. These findings suggest that risk factors for adult cardiovascular disease begin to cluster in normal children, which might have important consequences for development of atherosclerosis later in life.     

Incarceration and the acquired immunodeficiency syndrome: Autopsy results in Texas prison inmates

The Texas Department of Criminal Justice (TDCJ) houses many subjects with acquired immunodeficiency syndrome (AIDS) who receive medical care in a comprehensive AIDS treatment center. In this case-control autopsy survey, we compared pathological outcomes of TDCJ inmates treated at the center (n = 155) with nonincarcerated patients who died during the same period (n = 155). Using multiple regression analysis and a proportional hazards model, survival time in the prisoners was equivalent to that in the controls. With few exceptions, the prevalences of opportunistic viral, fungal, protozoal, and bacterial infections contributing to mortality were equivalent between groups. Mycobacterium tuberculosis was isolated more frequently in the inmates, and M avium intracellulare was isolated less frequently (P < .0001). The inmates had a higher prevalence of bacterial infection of the central nervous system (CNS) (9.1% v 1.4%; P < .006); half of all CNS bacterial infections were caused by M tuberculosis. Inmates had significantly lower prevalences of vacuolar myelopathy (P < .006) and severe wasting disease (P < .0009). We conclude that survival of prison inmates with AIDS treated in a comprehensive AIDS treatment center was equivalent to that of nonincarcerated subjects with AIDS. Prevalences of certain complications of AIDS differed in the inmates, showing that the prison environment influenced some of the underlying causes of AIDS morbidity and mortality.     

Electrogram prolongation and nifedipine-suppressible ventricular arrhythmias in mice following targeted disruption of KCNE1

Mutations in KCNE1 , the gene encoding the β subunit of the slowly activating delayed rectifier potassium current ( I _Ks) channel protein, may lead to the long QT syndrome (LQTS), a condition associated with enhanced arrhythmogenesis. Mice with homozygous deletion of the coding sequence of KCNE1 have inner ear defects strikingly similar to those seen in the corresponding human condition. The present study demonstrated and assessed the mechanism of ventricular arrhythmias in Langendorff-perfused whole heart preparations from homozygous KCNE1-/- mice compared to wild-type mice of the same age. The effects of programmed electrical stimulation with decremental pacing from the basal right ventricular epicardial surface upon electrogram waveforms recorded from the basal left ventricle were assessed and quantified using techniques of paced electrogram fractionation analysis for the first time in an experimental system. All KCNE1-/-( n = 10) but not wild-type ( n = 14) mouse hearts empirically demonstrated marked pacing-induced ventricular arrhythmogenicity. This correlated with significant increases in electrogram dispersion, consistent with a wider spread in conduction velocities, in parallel with clinical findings from LQTS patients with potassium channel mutations. In contrast, introduction of 100 nM isoprenaline induced arrhythmogenicity in both KCNE1-/- ( n = 7) and wild-type ( n = 6) hearts during pacing. Furthermore, pretreatment with 1 μM nifedipine exerted a strong anti-arrhythmic effect in the KCNE1-/- hearts ( n = 12) that persisted even in the presence of 100 nM isoprenaline ( n = 6). Our findings associate KCNE1-/- with an arrhythmogenic phenotype that shows an increased dispersion of conduction velocities, and whose initiation is prevented by nifedipine, a finding that in turn may have therapeutic applications in conditions such as LQTS.     

GABAB receptor transduction mechanisms, and cross-talk between protein kinases A and C, in GABAergic terminals synapsing onto neurons of the rat nucleus basalis of Meynert

The transduction mechanisms underlying presynaptic GABA_B receptor-mediated inhibition of transmitter release have been characterized for a variety of synapses in the central nervous system (CNS). These studies have suggested a range of transduction mechanisms, including a role for second messengers such as protein kinases A (PKA) and C (PKC). In the present study, we have examined the intracellular signalling pathways underlying baclofen-induced inhibition of GABA release from terminals synapsing onto rat basalis of Meynert neurons using patch-clamp recordings. Baclofen, a selective GABA_B receptor agonist, reversibly decreased both evoked and spontaneous, miniature, GABAergic inhibitory postsynaptic currents (eIPSCs and mIPSCs, respectively). Such baclofen actions were completely abolished by CGP55845A, a selective GABA_B receptor antagonist, and by staurosporine, a non-selective PKA and PKC inhibitor. The mIPSC frequency was still decreased by baclofen even in the presence of 4 AP, a K⁺ channel blocker, and Cd²⁺, a voltage-dependent calcium channel blocker. Pharmacological activation or inhibition of PKC activity affected basal GABA release and mildly affected the response to baclofen. Inhibition of the cAMP/PKA cascade also affected basal GABA release and, in a subset of neurons, occluded the effects of baclofen, suggesting that the GABA_B receptor-mediated inhibitory action on GABA release was mediated via decreases in PKA activity. In addition, PKA inhibition occluded the effects of PKC modulation on both basal GABA release and on the response to baclofen. Our results characterize the transduction pathway of baclofen at these nucleus basalis of Maynert (nBM) synapses and show, for the first time, some cross-talk between the cAMP/PKA and PKC pathways in mammalian presynaptic nerve terminals.     

Modulation and function of caspase pathways in B lymphocytes

Summary:  During their development, B-lineage cells are selected to mature, to die, to divide, or to survive and wait, ready to respond to external signals. The homeostatic balance between growth, death, and survival is mediated by signaling pathways through the B-cell antigen receptor (BCR) complex, cytokine and chemokine receptors or cell–cell coreceptor interactions. The BCR complex is a master regulator essential at key checkpoints during development. These checkpoints involve various processes, including negative selection (deletion), anergy, receptor editing, and positive selection. Without BCRs or downstream BCR-signaling components, B-lineage cells arrest during development. Removal of BCRs from mature B cells leads to their death. Here, we discuss signaling pathways in B cells that activate members of the caspase family of cysteine proteases. In some B-cell subsets, BCR signaling activates caspases, which in turn induce a program leading to cell death. However, in other contexts, caspases are involved in the proliferation of B cells. The outcome depends in part on the presence or absence of modifiers that affect signaling thresholds and on which caspases are activated. These mechanisms allow the coordinated regulation of proliferation and apoptosis that is essential for lymphoid homeostasis.