Yolène Thomas

Cadmium induced apoptosis in a human T cell line

Abstract Cadmium, a potent toxic metal, poses a serious environmental threat but the mechanisms of its toxicity remain unclear. In the present study, we investigated the nature of cadmium-induced cell death in the human T cell line CEM-C12. Cadmium was time- and dose-dependently toxic for CEM-C12 cells, cell death being preceded by chromatin condensation and DNA fragmentation. Quantification of the latter indicated an increase above 4 μM cadmium, with maximal fragmentation at 8 to 10 μM. By contrast, when CEM-C12 cells were exposed to higher cadmium concentrations (50 μM), cell death increased without concomitant chromatin condensation or DNA fragmentation. Thus, cadmium at low and high concentration kills CEM-C12 cells by apoptosis and necrosis, respectively. Addition of cycloheximide reduced the apoptotic effect of cadmium, suggesting that cadmium-induced apoptosis is an process depending on protein synthesis. Verapamil, a calcium/potassium channel blocker, markedly increased the viability of CEM-C12 cells treated by low cadmium concentrations and prevented DNA fragmentation. The apoptotic effect of cadmium suggests a possible mechanism for lymphocyte damage occuring after in vivo exposure to cadmium.

Cystatin C Levels in Sera of Patients with Human Immunodeficiency Virus Infection. A New Avidin-Biotin ELISA Assay for Its Measurement

A solid-phase enzyme-linked immunosorbent assay (ELISA) for determining human serum cystatin C is described. In 50 normal samples, cystatin C concentration was 1247 +/- 224 micrograms/L (mean +/- SD) which is in agreement with previously reported levels. Serum levels of cystatin C and beta 2-microglobulin (beta 2-M) were investigated in a time-course study during the development of human immunodeficiency virus (HIV) infection. We found a persistent and uniform increase in the beta 2-M concentration (2762 +/- 1239 micrograms/L). In contrast to beta 2-M, on the basis of cystatin C levels, we found two distinct populations, one of which demonstrated an increased concentration (1620 +/- 618 micrograms/L). Interestingly a second group (21% of patients) exhibited an initial significant decrease in cystatin C concentration with a mean value of 377 (range 55-850) micrograms/L, followed by an increase. The biphasic pattern of cystatin C serum, a major cysteine proteinase inhibitor, during the course of HIV infection suggests a possible role for these proteinases (or proteinase inhibitors) in the development of this syndrome.

Paf-acether production by Escherichia coli

Paf is a potent mediator of inflammatory diseases and septic shock. In previous studies we showed that paf can be released by prokaryotic cells such as E. coli. In this report we define the production and release of paf by E. coli cultured under different experimental conditions. When cultures were supplemented with lyso paf, a dramatic increase in paf production was observed. Most of the paf synthesized by bacteria was released in the supernatant. Of interest C16 lyso paf was 4-fold more efficient than its C18 counterpart. Using normal and reverse phase HPLC bacterial paf exhibited physico-chemical characteristics identical to those of synthetic paf. These results may indicate that the putative E. coli acetyltransferase recognizes differently C16 and C18 lyso paf. They also could be of importance considering the pathogenetic role of enterobacteria.

Synthesis of paf-acether from exogenous precursors by the prokaryote Escherichia coli

Paf‐acether (paf) is a potent mediator of inflammatory diseases and septic shock. Using normal‐phase HPLC, a paf‐like activity was found in culture supernatants from E. coli. Prokaryotic paf exhibited the same biological and physico‐chemical properties as eukaryotic cells and synthetic paf. Further, reverse‐phase HPLC indicates that paf generated by bacteria is predominantly of the hexadecyl and octadecyl species. When cultures were supplemented with lyso‐paf, a dramatic increase in paf production was observed. The purity and molecular structure of bacterial paf were further characterized by mass spectral analysis. These results could be of importance considering the pathogenetic role of enterobacteria. Further, it appears that the competence to form and release paf is an early phylogenetic development.

Immunoregulatory functions of paf-acether. VI. Dual effect on human B cell proliferation

The role of paf-acether (paf), a phospholipid cytokine, in the modulation of human B cell function was investigated. Paf, from 1×10−5 M to 10−6 M, decreased B cell proliferation induced by both phorbol myristate acetate (PMA) and anti-IgM antibodies (anti-IgM Ab). By contrast, 1×10−7 M to 1×10−9 M paf enhanced PMA triggered, but not anti-IgM triggered B cell proliferation. B cell proliferation was modulated between 24 and 72 hr of culture indicating that the effect of paf did not merely reflect a shift in proliferation kinetics. Interestingly, paf also enhanced the spontaneous proliferation of a Burkitt lymphoma-derived B cell line, Raji, which suggests that paf can directly act on B cells. The modulatory effect of paf on peripheral blood B cells was independent of PMA concentration, yet the effect on Raji cells was dependent upon cell density. The data suggest that paf is a potent modulator of B cell function, and may be involved in the control of humoral immune response.

Presence of paf-acether in human thymus

Platelet‐activating factor‐acether; Thymus

Regulation of platelet-activating factor receptor expression in human B cells and B cell lines

We extended our previous data regarding the modulation of human platelet-activating factor receptor (hPAF-R) expression on human B cell lines as well as normal B cells. First, we showed that hPAF-R, but was absent from cell lines devoid of hPAF-R. Second, enhanced hPAF-R membrane expression induced in IM9 line by IL9 line by IL4 was preceeded by hPAF-R mRNA accumulation that was detectable by 8 h and which peaked at 24 h. Similar results were observed for 10 nM platelet-activating factor treatment, which increased hPAF-R mRNA content up to 120% at 48 h, whereas hPAF-R membrane expression was up-regulated by 130%. Third, our data indicate that functional hPAF-R are expressed on resting, as well as on activated, B cells and that B cell activation is required for maintaining hPAF-R membrane and mRNA expression. Thus, in normal B cells, as well as in B cell lines, transcriptional regulation and/or messager stability control hPAF-R expression.

Modulation of stress proteins by Cd2+ in a human T cell line☆

We previously showed in a human T cell line (CEM-C12 cells) that Cd2+ induced gene expression of stress proteins, metallothionein-IIA and heat shock protein 70 in a time- and dose-dependent manner. In the present study, CEM-C12 cells were pretreated for 24 h with 1 microM Cd2+ and then challenged with toxic concentrations of this metal. We found that maximal expression of the metallothionein-IIA and heat shock protein 70 genes was increased and this maximal level occurred at higher Cd2+ toxic concentrations. Actinomycin D chase experiments indicated that Cd2+ pretreatment did not modify metallothionein-IIA mRNA stability. The modulatory effect of Cd2+ pretreatment was dose-dependent from 100 pM to 1 microM. Such pretreatment also enhanced resistance to Cd2+ toxicity. Finally, verapamil, a calcium/potassium channel blocker displaced the dose-response curve for Cd2+ toxicity as well as metallothionein-IIA and heat shock protein 70 gene expression to higher Cd2+ concentrations.

Paf-acether in human skin

Paf is a phospholipid mediator present in human skin which induces inflammatory events such as neutrophil infiltration and increased vascular permeability. Recent data suggest that cutaneous cells, such as fibroblasts and keratinocytes, produce paf and that paf is released during allergic cutaneous reactions. It is tempting to speculate that paf may contribute to the development of various skin disorders with acute and chronic skin inflammation. Paf antagonists may help in bringing answers to this hypothesis and may offer new prospects for the treatment of cutaneous inflammatory diseases.