Theileria annulata in CD5(+) macrophages and B1 B cells

Theileria parasites infect and transform bovine leukocytes. We have analyzed laboratory-established Theileria sp.-infected leukocyte lines and observed that transformed macrophages express CD5. Low-level expression of CD5 by macrophages was further confirmed on three independent Theileria annulata clinical isolates from Tunisia. Interestingly, the fourth CD5(+) clinical isolate (MB2) was morphologically different, expressed surface immunoglobulin M (IgM) and BoLA class II, and had rearranged Ig light-chain genes. To demonstrate that MB2 did indeed contain CD5(+) B cells, individual clonal lines were obtained by limiting dilution, and CD5 expression and Ig gene rearrangement were confirmed. This suggests that in natural infections T. annulata can invade and transform CD5(+) B cells.     

Characterization of T-cell-receptor gamma (TRG) gene rearrangements in alloreactive T-cell clones

Rearrangements of the T-cell Rearranging Gene (TRG) or T-cell-receptor gamma-chain genes were analyzed in 24 in vivo-sensitized alloreactive T-cell clones. This analysis represents the first complete assignment of TRG gene rearrangements to given variable and joining gene segments in nonleukemic T cells and provides some evidence for the hypothesis of sequential gamma genes rearrangements during T-lymphocyte differentiation. TRG gene rearrangements in our T-cell panel involved the known "active" V gamma genes, with a preferential use of V2 and V4 genes. In most clones, rearrangements occurred on both chromosomes and involved the J2 segment, but only 2 and 4 out of the 49 described rearrangements involved the additional J gamma segments JP1 and JP2, respectively. Two peculiar rearrangements were found. The first one was probably due to the creation of a new restriction enzyme site in the N-region at the V-J junction; the second can be explained by an aberrant rearrangement of a V gene to a sequence located between exons 2 and 3 of the TRGC1 gene.     

Detailed immunoautoradiographic mapping of enkephalinase (EC 3.4.24.11) in rat central nervous system: comparison with enkephalins and substance P

The metallopeptidase enkephalinase known to participate in the inactivation of endogenous enkephalins and, possibly, other neuropeptides such as tachykinins, was visualized by autoradiography using a [125I]iodinated monoclonal antibody. A detailed mapping of the enzyme in rat brain and spinal cord was established on 10-micron serial sections prepared in a frontal plane as well as a few sections in a sagittal plane. On adjacent sections, and for the purpose of comparison, substance P-like and enkephalin-like immunoreactivities were also visualized by autoradiography using a 125I-monoclonal antibody and a polyclonal antibody detected by a secondary 125I-anti-rabbit antibody respectively. Histological structures were identified on adjacent Nissl-stained sections. Using the highly sensitive 125I-probe, enkephalinase immunoreactivity was found to be distributed in a markedly heterogeneous manner in all areas of the central nervous system. Immunoreactivity was undetectable in white matter areas, for example the corpus callosum or fornix, and had a laminar pattern in, for example, the cerebral cortex or hippocampal formation. Hence, although immunodetection was not performed at the cellular level, a major neuronal localization of the peptidase is suggested. The latter is consistent with the detection of a strong immunoreactivity in a pathway linking the striatum to the globus pallidum, the entopeduncular nucleus and the substantia nigra, as well as with a series of biochemical and lesion data. The strong immunoreactivity also present in choroid plexuses and ependymal cells as well as in the intermediate lobe and in scattered cells of the anterior lobe of the pituitary suggests that populations of glial and endocrine cells also express the peptidase. The highest density of enkephalinase immunoreactivity was observed in basal ganglia and limbic areas (caudate putamen, globus pallidus, nucleus accumbens, olfactory tubercles) as well as in areas involved in pain control mechanisms (superficial layers of the spinal nucleus of the trigeminal nerve or of the dorsal horn of the spinal cord) which also display the highest immunoreactivities for both enkephalins and substance P (except in globus pallidus for the latter). These localizations account for the opioid-like analgesic and motor effects of enkephalinase inhibitors inasmuch as a selective or predominant participation of the peptidase in enkephalin inactivation is assumed. A number of other areas appear richly endowed in both enkephalinase and enkephalins whereas substance P is hardly detectable. This is particularly the case for the olfactory bulb, bed nucleus of the accessory olfactory tract, the cerebellum (where enkephalinase mainly occurs in the molecular layer) and the hippocampal formation (namely in the molecular layer of the dentate gyrus).(ABSTRACT TRUNCATED AT 400 WORDS)     

Evaluation of the inflammatory process in hepatic disorders: immunochemistry of C alpha-reactive protein and of alpha 1-acid glycoprotein

C-reactive protein (CRP) and alpha 1-acid glycoprotein (alpha 1-GPA) two acute phase reactants, have both been monitored in patients suffering from hepatocellular diseases, compared with healthy subjects. Immunochemical findings, in hepatic amebiasis, revealed a higher incidence of increased alpha 1-GPA levels (86% of patients, as compared to 46% for CRP), whereas during liver primitive cancer and cirrhosis inverse pattern occurs. In viral chronic hepatitis, lesser perturbations were observed. In contrast, a simultaneous increase of both proteins is strong supporting evidence for the severity of disease with an unfavourable prognostic.     

Trisomy 3 in cold agglutinin disease

Cold agglutinin disease (CAD) is a form of autoimmune hemolytic anemia generally considered to result from an antibody-producing clonal expansion of B lymphocytes. We studied 16 patients with CAD and demonstrated a chromosomally abnormal clone in the peripheral blood (PB) of six. Trisomy 3 was the only abnormality in two patients, trisomy 12 the only abnormality in one, and both trisomy 3 and trisomy 12 were present in three patients. A lymphoma was subsequently diagnosed in two of these six patients and in none of the patients without a karyotypic abnormality. The results suggest that trisomy 3 confers a slight growth advantage in the B-cell lineage, particularly at a relatively late stage of differentiation. The resultant clone may present clinically as CAD and, in some cases, progress to a clinical lymphoma.     

Lack of intermediate-affinity interleukin-2 receptor in mice leads to dependence on interleukin-2 receptor α, β and γ chain expression for T cell growth

An interleukin (IL)-4 dependant mouse T cell clone 8.2 derived from an IL-2-dependent T cell line was characterized. As measured by flow cytometric analysis and Northern blotting, it expresses IL-2 receptor β (IL-2Rβ) and γ (IL-2Rγ) chains, but has lost expression of IL-2 receptor α chain (IL-2Rα). To investigate the properties of the mouse IL-2Rβγ complex and the role of IL-2Rα gene expression, this clone was further studied. T cell clone 8.2 has lost the capacity to bind ¹²⁵I-labeled human IL-2 under experimental conditions able to detect intermediate-affinity IL-2R in human cells. Mouse IL-2 is unable to block the binding of mAb TMβ1 to 8.2 cells. Under the same experimental conditions, mouse IL-2 blocks the binding of TMβ1 to C30-1 cells expressing the IL-2αβγ complex. Since TMβ1 recognizes an epitope related to the IL-2 binding site of IL-2Rβ, these results can be taken as a demonstration that mouse IL-2Rβγ does not bind mouse IL-2. Furthermore, T cell clone 8.2 does not proliferate in response to recombinant mouse or human IL-2. On the other hand, T cell transfectant lines expressing heterospecific receptors made of the human IL-2Rβ and mouse IL-2Rγ chains bind ¹²⁵I-labeled human IL-2 and proliferate in response to IL-2. This establishes the difference between mouse and human IL-2Rβ chains. Transfection of T cell clone 8.2 with human IL-2Rα genes restores their capacity to proliferate in response to IL-2. In addition, all transfectants grown in IL-2 express the endogeneous mouse IL-2Rα chain. When grown in IL-4, the endogeneous mouse IL-2Rα gene remains silent in all these transfectants. These results show that, contrary to the human, the mouse does not express an intermediate-affinity IL-2R. Expression of the IL-2Rα gene is therefore required for the formation of the functional IL-2R in mice.     

Microarray-based comparative genomic analyses of the human malaria parasite Plasmodium falciparum using Affymetrix arrays

Microarray-based comparative genomic hybridization (CGH) provides a powerful tool for whole genome analyses and the rapid detection of genomic variation that underlies virulence and disease. In the field of Plasmodium research, many of the parasite genomes that one might wish to study in a high throughput manner are not laboratory clones, but clinical isolates. One of the key limitations to the use of clinical samples in CGH, however, is the miniscule amounts of genomic DNA available. Here we describe the successful application of multiple displacement amplification (MDA), a non-PCR-based amplification method that exhibits clear advantages over all other currently available methods. Using MDA, CGH was performed on a panel of NF54 and IT/FCR3 clones, identifying previously published deletions on chromosomes 2 and 9 as well as polymorphism in genes associated with disease pathology.