Co-localization of galectin-1 with GM1 ganglioside in the course of its clathrin- and raft-dependent endocytosis.

Mammalian galectin-1 (Gal-1), a beta-galactoside-binding lectin has a prominent role in regulating cell adhesion, cell growth and immune responses. Downregulation of these biological functions may occur via internalization of Gal-1. In the present study we have investigated the mechanism and possible mediator(s) of Gal-1 endocytosis. We show that internalization occurs at a temperature higher than 22 degrees C in an energy dependent fashion. After one hour incubation Gal-1 localizes in the Golgi system within the cells, and then disappears without accumulation in degradation compartments, such as lysosomes. Based on their strong intracellular co-localization, two glycoconjugates, GM1 ganglioside and CD7 are implicated in the sorting of internalized Gal-1 into Golgi. Other known Gal-1 binding glycoproteins on T cells (CD2, CD3, CD43 and CD45) do not cointernalize with the lectin. Internalization of Gal-1 depends on its lectin activity and follows dual pathways involving clathrin-coated vesicles and raft-dependent endocytosis.

Optimization of the cellular import of functionally active SH2-domain-interacting phosphopeptides.

Phosphopeptides interacting with src homology 2 (SH2) domains can activate essential signaling enzymes in vitro. When delivered to cells, they may disrupt protein-protein interactions, thereby influencing intracellular signaling. We showed earlier that phosphopeptides corresponding to the inhibitory motif of Fcgamma receptor IIb and a motif of the Grb2-associated binder 1 adaptor protein activate SH2-containing tyrosine phosphatase 2 in vitro. To study the ex vivo effects of these peptides, we have now compared different methods for peptide delivery: (i) permeabilization of the target cells and (ii) the use of cell-permeable vectors, which are potentially able to transport biologically active compounds into B cells. We found octanoyl-Arg(8) to be an optimal carrier for the delivery of phosphopeptides to the cells. With this strategy, the function of cell-permeable SHP-2-binding phosphopeptides was analyzed. These peptides modulated the protein phosphorylation in B cells in a dose- and time-dependent manner.

Differential expression of GL7 activation antigen on bone marrow B cell subpopulations and peripheral B cells.

GL7 was originally described as a 35-kDa late activation antigen on mouse T and B cells. GL7 expression has also been demonstrated on thymocytes, germinal center B cells and some neuronal cell types. Flow-cytometry and immunohistochemistry were used to follow changes in the expression of GL7 during B cell development, amongst B cell subpopulations and various anatomical locations. GL7 is expressed as early as the pro-B cell stage and increases up to the pre-B-I stadium. Expression remains high on pre-B-II and on immature B cells, although slightly decreases during maturation. GL7 is almost completely downregulated when IgD appears on the cell surface. On the periphery only a few B cells are positive and these cells are almost exclusively found in the sIgD- germinal center areas of lymph nodes and spleen. The staining pattern of GL7 is very similar to that of PNA in the lymph nodes but in the bone marrow we have found both B220+PNA+GL7- and B220+PNA+GL7+ populations, showing that GL7 and the antigen recognized by PNA are different. After in vitro stimulation, the GL7(hi) B cell population has also been found to be IgD negative. Functional comparison between in vitro activated and MACS sorted GL7(hi) and GL7(lo/-) spleen B cells of immunized mice showed significantly higher specific and total antibody production as well as antigen presenting capacity in the GL7(hi) population.

Comparative study on the effect of phosphorylated TCR zeta chain ITAM sequences on early activation events in Jurkat T cells.

One of the main dilemma in T cell receptor (TCR) signal transduction is whether the presence of multiple Immunoreceptor Tyrosine-based Activation Motifs (ITAMs) within the TCR signaling module serves for signal amplification or signal distribution. To contribute to answer this question, we analyzed the effect of synthetic oligopeptides representing the three bi-phosphorylated zeta chain-ITAMs on the early signaling events in permeabilized leukemia T cells. Our main observations were as follows: 1/Stimulation of the cells with the bi-phosphorylated membrane proximal and central ITAMs (zeta (1)y(p)y(p) and zeta (2)y(p)y(p), respectively) resulted in a strong phosphorylation of proteins with a similar pattern. In contrast, the membrane distal ITAM, zeta (3)y(p)y(p) had a reduced ability to promote tyrosine phosphorylation and failed to induce the phosphorylation of a number of proteins. 2/ The phospho-peptide induced tyrosine phosphorylation events were at least partially mediated by p56(lck) and Syk/ZAP70 protein tyrosine kinases as it was shown in p56(lck) and Syk/ZAP70 deficient Jurkat variants. 3/The patterns of the association of the adaptor protein, Grb2 with tyrosine phosphorylated proteins following cell stimulation with the bi-phosphorylated membrane proximal or the central ITAMs were similar, while the membrane distal ITAM was unable to induce any of these associations. Our data provide additional evidence that the three zetaITAMs differ in their capacity to induce tyrosine phosphorylation of intracellular proteins in permeabilized T cells, depending to their primary sequence. The first and second ITAM sequences of the zeta chain may have similar but not totally overlapping functions. This conclusion results from their similar but not identical abilities to induce tyrosine phosphorylation and association of Grb-2 with intracellular phosphoproteins. In contrast, the third ITAM (zeta3) may have distinct functions since this peptide fails to induce tyrosine phosphorylation of a number of proteins compared to the other two ITAMs, and it is unable to induce either new association or the increase in the amount of Grb-2 associated phosphoproteins.

Emergency percutaneous nephrostomy in the septic kidney.

From 250 upper tract obstructive uropathy cases we have studied 64 patients hospitalized with toxico-septic shock. The constant symptom was arterial hypotension. Other 3 patients with long-standing urinary infection due to lithiasis developed this dreaded complication after PNL (staghorn stones-2, pyelic stone-1). In complicated obstructive uropathy cases associated with toxico-septic shock, percutaneous nephrostomy for high urinary derivation in emergency is usually made under local anaesthesia. Its aim is rapid and efficient clearance of kidney obstruction, with minimal damage for the patient; then it is followed by strong antibiotherapy associated with other reanimation and intensive care measures. There were 11 deaths. The stone generating obstructive uropathy was removed subsequently, after the improvement of biological constants and general state of the patient, under the protection of percutaneous nephrostomy.

The place of echography in percutaneous nephrostomy and nephrostolithotomy.

The advantages and limits of echography in the performance of percutaneous nephrostomy and nephrostolithotomy are described, based on our own cases.

Anterograde urethrotomy, a new method for internal urethrotomy.

The authors elaborated a new device permitting to perform internal urethrotomy for urethral stricture in an anterograde way, even in cases where optical retrograde urethrotomy could not be applied. The new device is similar to a bladder biopsy forceps with a diameter of Ch 5. Its cutting knife is movable. It can be used with Sachse's urethrotome, inserting it in its guide channel with closed knife and passing it through the stricture. By opening the knife, anterograde urethrotomy can be performed. Out of 145 urethrotomies, this method was utilized in only 3 cases.

Original transvesical prostatectomy.

Starting out from a series of theoretical considerations, and checked clinically, the authors describe an original technique for prostatic adenomectomy, being more efficient in limitation of surgical haemorrhage, urinary infection and postoperative hypogastric fistulae. The operative technique and its results in comparison with classic techniques are presented.

Improved suprapubic vesical drainage in transvesical prostatectomy.

An improved drainage is used in transvesical prostatectomy in order to avoid urinary retention after removal of the Novak-type suprapubic drainage with bladder neck cerclage haemostasis (described by some authors in 1975). The causes of urinary retention are analysed, the drainage technique as well as the mechanisms eliminating the above-mentioned complications are described.

[Technic of transvesical suprapubic adenomectomy for the control of hemorrhage and urinary infection]. / Technik der transvesikalen suprapubischen Adenomektomie zur Beherrschung der Blutung und Harninfektion

The advantages of our method: 1. By omission of the urethral catheter the infection is nearly completely avoided. 2. It guarantees a good haemostasis. 3. It reduces the intraoperative and later complications of adenectomy (infection and anaemia). 4. It facilitates the postoperative treatment. 5. One avoids urethritis and strictures caused by the urethral catheter. 6. It shortens the postoperative period of rest. 7. One economises medicaments (antibiotically, antianaemically). 8. The possible break of the catgut thread at its removal is of no danger, as it is resorbed.