Analysis of lymphocytes reactive to histocompatibility antigens. III. Detection of inclusion among allo-reactive lymphocyte populations by specific depletion of reactive cells.

We have used a quantitative titration assay for mixed lymphocyte interactions to determine the additivity of responses of rat lymphocyte populations to simultaneous stimulation with major histocompatibility complex alloantigens from pairs of apparently unrelated rat strains. Our inability to detect strict additive stimulation led us to investigate inclusion among allo-reactive lymphocyte populations. After specific depletion of reactivity to PVG by negative selection through an AO × PVG F₁ hybrid rat, AO thoracic duct lymphocytes showed an asymmetrical loss of reactivity to the unrelated strains DA and F344. We suggest that this differential loss of reactivity probably reflects alloantigen sharing among rat strains which had been previously undetected using other less sensitive techniques for the quantitation of lymphocyte reactivity after specific depletion of allo-reactive cells.     

The ubiquitin C‐terminal hydrolase UCH‐L1 promotes bacterial invasion by altering the dynamics of the actin cytoskeleton

Invasion of eukaryotic target cells by pathogenic bacteria requires extensive remodelling of the membrane and actin cytoskeleton. Here we show that the remodelling process is regulated by the ubiquitin C‐terminal hydrolase UCH‐L1 that promotes the invasion of epithelial cells by Listeria monocytogenes and Salmonella enterica. Knockdown of UCH‐L1 reduced the uptake of both bacteria, while expression of the catalytically active enzyme promoted efficient internalization in the UCH‐L1‐negative HeLa cell line. The entry of L. monocytogenes involves binding to the receptor tyrosine kinase Met, which leads to receptor phosphorylation and ubiquitination. UCH‐L1 controls the early membrane‐associated events of this triggering cascade since knockdown was associated with altered phosphorylation of the c‐cbl docking site on Tyr1003, reduced ubiquitination of the receptor and altered activation of downstream ERK1/2‐ and AKT‐dependent signalling in response to the natural ligand Hepatocyte Growth Factor (HGF). The regulation of cytoskeleton dynamics was further confirmed by the induction of actin stress fibres in HeLa expressing the active enzyme but not the catalytic mutant UCH‐L1_C90S. These findings highlight a previously unrecognized involvement of the ubiquitin cycle in bacterial entry. UCH‐L1 is highly expressed in malignant cells that may therefore be particularly susceptible to invasion by bacteria‐based drug delivery systems.     

Interferon-γ effect on growth regulation of cells with different levels of EGF receptor expression

Interferon gamma (IFNγ) is known to inhibit the proliferation of some transformed cell lines. Recently, we demonstrated the transactivation of the epidermal growth factor receptor (EGFR) in response to IFNγ (Burova et al., 2007) and provided direct evidence for the dependence of IFNγ-induced EGFR transactivation on the EGFR expression level in epithelial cells (Gonchar et al., 2008). This study examines an antiproliferative effect of IFNγ on human epithelial cell lines—A431 and HeLa that express high levels of EGFR, as well as HEK293 that expresses low levels of EGFR. To characterize the IFNγ-induced changes in these cells, we studied cell growth, the cell cycle, and induction of apoptosis. The response to IFNγ differed in the compared cell lines; cell growth was inhibited in both A431 and HeLa cells, but not in HEK293 cells, as was shown by the cell count and MTT. The cell-cycle phases analyzed by flow cytometry were disturbed in A431 and HeLa cells in response to IFNγ. On the contrary, in HEK293 cells, the IFNγ treatment did not alter distribution by cell cycle phases. Our results indicate that IFNγ produces an antiproliferative effect that depends on the increased expression of EGFR in A431 and HeLa cells. Furthermore, it was demonstrated that IFNγ induced the caspase 3 activation in A431 cells, which suggests the involvement of active caspase 3 in the IFNγ-induced apoptosis.     

Immune relations between mother and foetus in the viviparous poeciliid fish Xiphophorus hellerii Haeckel

Advanced embryos of the viviparous poeciliid fish Xiphophorus hellerii (swordtail) were removed from the site of gestation within the ovary and re-implanted into the peritoneal cavities of unrelated immunologically competent adults of the same species. Most of those transplanted with intact fertilization membranes were normal in appearance, and many were clearly still alive. Those transplanted without a fertilization membrane were all dead, with lymphocyte infiltration and other evidence of rejection as allografts. The integrity of the membrane in late pregnancy was indicated by swelling of follicles in hypoosmotic liquid. It was concluded that the fertilization membrane is capable of protecting an antigenically foreign embryo in an immunologically hostile environment, and that it probably serves this function during normal pregnancy.     

Preliminary clinical experience with the antiarrhythmic effect of PGF2a

A total dosage up to 1 mg PGF_2a as i.v. infusions of 10–40 μg/min. was investigated on patients with arrhythmias of several kinds. We found therapeutic effects in 5 of 6 patients with constant extrasystoles and in one patient with digitalis - induced partial AV-block respectively. In 3 of 4 patients with acute tachyarrhythmias the results were not convincing, probably due to a dosage not high enough. An increase of the diastolic stimulation threshold usually seen with other antiarrhythmics was not to be observed in 3 patients. The mechanism of action of PGF_2a has not yet been clarified.