Expression of macrophage markers by a population of T cells obtained from synovial fluid of a subgroup of patients with juvenile rheumatoid arthritis

OBJECTIVE: To characterize distinctive lymphoid cell populations in the synovial fluid (SF) of patients with juvenile rheumatoid arthritis (JRA) that have the specific ability to display monocytic markers when cultured in vitro. METHODS: Mononuclear cells obtained from SF of patients with JRA and depleted of adherent macrophages were cultured in vitro in RPMI 1640 medium supplemented with only fetal calf serum (FCS). Phenotypic evaluation of these cells was by flow cytometry and immunohistochemical analysis was by specific fluorochrome labeled antibodies. RESULTS: T cells from a JRA subgroup displayed some typical macrophage attributes, i.e., abundant cytoplasm, adherence to plastic, and phagocytosis of latex beads when cultured in vitro. These cells have the ability to survive in culture for several weeks in RPMI 1640 medium containing only 10% FCS. The macrophage-like T cells rosetted with sheep red blood cells and proliferated when stimulated with phytohemagglutinin or anti-CD3, indicating functional T cell responses. CONCLUSION: Our data indicate that a population of T cells obtained from the SF of a subgroup of patients with JRA exhibited characteristics of macrophages, yet retained their CD3 and T cell receptor expression. Whether this promiscuous behavior is caused by malignant transformation of lymphoid precursor cells or is induced by the concerted effect of a myriad of cytokines and growth factors present in the SF remains unknown. The presence of these cells in the SF of 2 patients with JRA with different onset types raises the question of their function and significance in an autoimmune disorder such as JRA.     

Supersites within superfolds. Binding site similarity in the absence of homology

A method is presented to assess the significance of binding site similarities within superimposed protein three-dimensional (3D) structures and applied to all similar structures in the Protein Data Bank. For similarities between 3D structures lacking significant sequence similarity, the important distinction was made between remote homology (an ancient common ancestor) and analogy (likely convergence to a folding motif) according to the structural classification of proteins (SCOP) database. Supersites were defined as structural locations on groups of analogous proteins (i.e. superfolds) showing a statistically significant tendency to bind substrates despite little evidence of a common ancestor for the proteins considered. We identify three potentially new superfolds containing supersites: ferredoxin-like folds, four-helical bundles and double-stranded beta helices. In addition, the method quantifies binding site similarities within homologous proteins and previously identified supersites such as that found in the beta/alpha (TIM) barrels. For the nine superfolds, the accuracy of predictions of binding site locations is assessed. Implications for protein evolution, and the prediction of protein function either through fold recognition or tertiary structure comparison, are discussed.     

Coronary artery spasm complicating anaphylaxis secondary to skin disinfectant

We report a patient in whom presumed vasospasm of an angiographically normal coronary artery led to severe transmural myocardial ischemia. To our knowledge, this is the first case in which an allergic reaction to locally applied chlorhexidine caused such a severe reaction.     

ILAE-defined epilepsy syndromes in children: correlation with quantitative MRI

PURPOSE: The role of quantitative magnetic resonance imaging (MRI) in evaluation of childhood epilepsy remains poorly defined, with minimal published data. Previous work from our center questioned the specificity of hippocampal asymmetry (HA) in an outpatient group whose epilepsy was defined by using clinical and interictal data only. By using childhood volunteer controls and defining epilepsy syndromes using video-EEG monitoring, we readdressed the utility of HA in differentiating mesial temporal lobe epilepsy (MTLE) from other partial and generalized epileptic syndromes in children. METHODS: Seventy children were enrolled; entry criteria were age younger than 18 years with predominant seizure type recorded on video-EEG telemetry with volumetric MRI in all cases. Thirty healthy child volunteers had volumetric MRI. Epilepsy syndrome classification was according to ILAE. RESULTS: Control data revealed symmetric hippocampi, mean smaller/larger ratio of 0.96 (0.95-0.97, 95% CI) with no gender or right/left predominance. Overall 23% of patients had significant HA. Mean hippocampal ratio for MTLE was 0.78 (95% CI, 0.70-0.86), significantly lower than controls and from all other epilepsy syndromes. HA was highly specific (85%) to the syndrome of MTLE. Other potential epileptogenic lesions were found in 27 (39%) patients, lowest yield in frontal and mesial temporal syndromes. Dual pathology was present in 10% of patients. There was no significant association between HA and risk factors. CONCLUSIONS: In this study, we found that HA in children with a well-defined epilepsy syndrome is highly sensitive and specific for MTLE. Whether this will correlate with surgical outcome, as in adults, is the subject of ongoing study.     

Conditioned fear-induced tachycardia in the rat: vagal involvement

The effects of conditioned fear on gross activity, heart rate, PQ interval, noradrenaline and adrenaline were studied in freely moving rats. Subcutaneous (s.c.) injections of atropine methyl nitrate (0.5 mg/kg) during rest resulted in a significant shortening of the PQ interval, indicating that the PQ interval can be used as a measure of vagal activity. Conditioned fear was induced by 10-min forced exposure to a cage in which the rat had previously experienced footshocks (5 x 0.5 mA x 3 s). In non-shocked controls, an increase in gross activity was found and a pronounced tachycardia, without changes in PQ interval. Conditioned fear rats showed immobility behaviour, associated with a less pronounced tachycardia and an increase in PQ interval. Noradrenaline was similarly increased in both groups, whereas adrenaline was increased in conditioned fear rats only. To further evaluate the role of the vagus, rats were exposed to conditioned fear after pre-treatment with atropine methyl nitrate (0.5 mg/kg, s.c.). Again, immobility was observed with a concomitant tachycardia, but without an increase in PQ interval. These results indicate that the autonomic nervous system is differentially involved in heart rate regulation in conditioned fear rats and in non-shocked controls: in non-shocked controls a predominant sympathetic nervous system activation results in an increase in heart rate, whereas in conditioned fear rats the tachycardiac response is attenuated by a simultaneous activation of sympathetic nervous system and parasympathetic nervous system.     

MyoD and the specification of muscle and non-muscle fates during postembryonic development of the C. elegans mesoderm

Basic-helix-loop helix factors of the myoD/myf5/ myogenin/MRF4 family have been implicated in acquisition and elaboration of muscle cell fates. Here we describe both myogenic and non-myogenic roles for the Caenorhabditis elegans member of this family (CeMyoD) in postembryonic mesodermal patterning. The postembryonic mesodermal lineage in C. elegans provides a paradigm for many of the issues in mesodermal fate specification: a single mesoblast ('M') divides to generate 14 striated muscles, 16 non-striated muscles, and two non-muscle cells. To study CeMyoD function in the M lineage, we needed to circumvent an embryonic requirement for the protein. Two approaches were used: (1) isolation of mutants that decrease CeMyoD levels while retaining viability, and (2) analysis of genetic mosaics that had lost CeMyoD in the M lineage. With either manipulation, we observed a series of cell-fate transformations affecting a subset of both striated muscles and non-muscle cells. In place of these normal fates, the affected lineages produced a number of myoblast-like cells that initially failed to differentiate, instead swelling to acquire a resemblance to sex myoblasts (M-lineage-derived precursors to non-striated uterine and vulval muscles). Like normal sex myoblasts, the ectopic myoblast-like cells were capable of migration and proliferation followed by differentiation of progeny cells into vulval and uterine muscle. Our results demonstrate a cell-intrinsic contribution of CeMyoD to specification of both non-muscle and muscle fates.     

Interleukin-10 gene transfer inhibits murine mammary tumors and elevates nitric oxide

Transfection of cDNA for IL-10 into line 66.1 murine mammary tumor cells results in marked suppression of tumor growth and metastasis. Others have reported that nitric oxide has potent antitumor activity and IL-10 is known to regulate the inducible isoform of nitric oxide synthase (iNOS) expressed in macrophages. We identified nitric oxide production in mammary tumors as indicated by electron paramagnetic resonance detection of nitric oxide-hemoglobin (NO-Hb). IL-10 expression resulted in elevated levels of NO-Hb in mammary tumors. Immunohistochemical examination of mammary tumors for iNOS protein revealed few positively staining cells in parental or control neo-transfected tumors but strong iNOS staining in all IL-10 transfected tumors, consistent with the NO-Hb data. To determine if mammary epithelial tumor cells themselves, express nitric oxide synthase activity, cultured tumor cells were treated with pro-inflammatory cytokines and nitrite accumulation was assessed in the conditioned medium. All IL-10 producing cell lines accumulated uM concentrations of nitrite in response to short term (24 hr) cytokine stimulation. Cells not expressing IL-10 (parental and neo-transfectants) accumulated no nitrite under similar culture conditions. After longer stimulation (48 hr), parental and 66-neo cells accumulated lower amounts of nitrite. IL-10 gene transfer is associated with increased iNOS protein expression and enzymatic activity detected both in vitro and in vivo. Our findings suggest that the antimetastatic and antitumor activity of IL-10 is related to enhanced production of nitric oxide.     

Novel phosphate mimetics for the design of non-peptidyl inhibitors of protein tyrosine phosphatases

Benzylic alpha,alpha-difluorosulfonates, alpha,alpha-difluorotetrazoles, and alpha,alpha-difluorocarboxylates of type 5 and 6 were synthesized and examined as potential phosphate biosteres for PTP1B inhibition. The alpha,alpha-difluorosulfonates and alpha,alpha-difluorotetrazoles were found to be more effective inhibitors than the analogous compounds bearing the fluoromalonyl group, a phosphate biostere currently being used for PTP inhibition.