Cytocidal and apoptotic effects of the ClyA protein from Escherichia coli on primary and cultured monocytes and macrophages.

Cytolysin A (ClyA) is a newly discovered cytolytic protein of Escherichia coli K-12 that mediates a hemolytic phenotype. We show here that highly purified ClyA and ClyA-expressing E. coli were cytotoxic and apoptogenic to fresh as well as cultured human and murine monocytes/macrophages.

Yersinia invasin, a bacterial beta1-integrin ligand, is a potent inducer of lymphocyte motility and migration to collagen type IV and fibronectin.

The Yersinia pseudotuberculosis invasin protein was found to be a potent inducer of pseudopodia formation and chemotactic and haptotactic migration in human T lymphocytes. Checkerboard analysis confirmed that migration was directional. The Yersinia invasin triggered migration of otherwise poorly migratory normal T cells on fibronectin and in particular on collagen type IV, and augmented the migration of leukemic T cell lines on these components. Invasin-induced lymphocyte migration was inhibited by staurosporin that selectively prevented pseudopodia formation but, noteworthy, augmented adhesion. The motogenic and attractant properties of invasin (Inv) were mediated via beta1-integrins, as shown by lack of effect of Inv on the motility of a beta1-integrin-negative lymphoid cell line and inhibition of invasin-induced lymphocyte motility by anti-beta1 Abs. Inv was markedly more effective than the extracellular matrix components fibronectin, collagen type IV, and laminin, which also interact with lymphocyte beta1-integrins, with respect to induction of pseudopodia, chemotaxis, and haptotaxis. Thus, Yersinia invasin is a model ligand for induction of lymphocyte motility via beta1-integrins. The extraordinary capacity of Inv to trigger and guide T lymphocyte motility and potentiate lymphocyte migration to extracellular matrix components may be of pathogenetic significance for the movement of lymphocytes to extraintestinal sites secondary to Yersinia infection.

Fibronectin and lymphocytes in inflammatory tissue. Studies of blood and synovial fluid lymphocytes from patients with rheumatoid arthritis and other inflammatory arthritides.

Lymphocytes infiltrating tissues under chronic inflammatory conditions are often surrounded by deposits of fibronectin. We have studied the possibility that this reflects capacity of lymphocytes to synthesize fibronectin and compared lymphocytes from blood and synovial fluid with respect to fibronectin interactions. In vitro activated blood lymphocytes exhibited synthesis of a fibronectin-like molecule. Synovial fluid cells appeared to synthesize the same high molecular weight component spontaneously. Activated blood lymphocytes have cell surface fibronectin and surface components of lower molecular weight which could be immunoprecipitated with anti-fibronectin antibodies as well as by insolubilized collagen. Synovial fluid cells showed cell surface fibronectin as revealed by immunocytochemical detection but seemed to lack or have relatively small amounts of the low-molecular weight fibronectin-like surface components. Synovial fluid T cells from arthritis patients showed adhesion to fibronectin. Immunocytochemistry demonstrated presence of alpha 4 and alpha 5 beta 1 integrins at the surface of the synovial fluid T cells and RGD and LDV peptides inhibited adhesion of the cells to fibronectin. Noteworthy, a portion of synovial fluid cells with lymphocyte markers also bound to plastic. Blood lymphocytes from the same arthritis patients displayed relatively poor or negligible adhesion to fibronectin unless activated to blast transformation and did not attach to plastic. Taken together these results suggest that activated lymphocytes from blood and synovial fluid may use fibronectin of exogenous or endogenous origin when interacting with tissues during inflammatory processes. Furthermore, the presence at the lymphocyte surface of components of different molecular weight precipitated by anti-fibronectin antibodies suggests that fibronectin or its fragments can bind to the lymphocyte surface.

Characterization of collagen binding lymphocyte membrane molecules.

Blood lymphocytes can adhere to two-dimensional collagen substrata. The lymphocyte plasma membrane contains 130 and 55 kd components with collagen-bindings capacity as well as high molecular weight collagen binding component (greater than 200 kd) pointing to the possibility that these molecules mediate lymphocyte adhesion to collagen. The 55 kd and the high molecular weight component also react with anti-fibronectin antibodies. Both the 130 kd and the 55 kd collagen binding components are present in lymphoid cell lines of T cell origin but the intensity of their expression varies. Under reducing conditions the 130 kd collagen binding components unchanged whereas the 55 kd component is increased to 66 kd.

Collagen receptor on T lymphocytes and the control of lymphocyte motility.

Human lymphocytes, freshly isolated from blood, were allowed to settle on surfaces coated by collagen type 1, fibronectin, laminin, IgG or albumin at different concentrations. In separate cultures the lymphocytes were also exposed to these proteins in soluble form. The lymphocytes, predominantly T cells, attached to two-dimensional collagen substrata both in the presence and absence of serum but did not adhere or adhered poorly to substrata coated with fibronectin, laminin, IgG and albumin. In contrast, T blasts induced in a mixed lymphocyte culture adhered to fibronectin-coated substratum. During contact with substratum-bound collagen for a 24-h period, 47 +/- 15% of the freshly purified lymphocytes from separate individuals developed motile behavior whereas 16 +/- 4% of the cells became motile on fibronectin. Gelatin (denatured collagen) also mediated attachment of lymphocytes to surfaces but only at comparatively high concentrations (40 mg/ml). Collagen and gelatin in solution also caused agglutination and motility of the vast majority of freshly isolated T lymphocytes whereas fibronectin and other proteins, when presented in soluble form, did not. Cell agglutination was maximal at moderate (10 or 20 mg/ml) and cell motility at low gelatin concentrations (1 to 10 mg/ml). High gelatin concentrations (20 and 40 mg/ml) did not induce motile behavior. Cytochalasin B augmented the proportion of adherent cells on gelatin-coated substrata. In the presence of cytochalasin B gelatin mediated substrate-adhesion at concentrations below those which normally induced adhesion indicating that motile behavior counteracted persistent lymphocyte adhesion to the substratum. Noteworthy, gelatin/collagen is unique among ligands (e.g. plasma fibronectin and other serum proteins) in its capacity to induce motility in the vast majority of resting lymphocytes freshly isolated from blood within a relatively short period. Taken together these results indicate that circulating lymphocytes have a collagen/gelatin-binding plasma membrane component. Cross-linking of this component is a likely explanation for the selective inducing effect of gelatin and collagen on lymphocyte motility. The present results showed that the lymphocyte plasma membrane contains collagen-binding components with a relative molecular mass of 130 and 55 kDa. The 55-kDa component also reacted with an anti-fibronectin antibody. Thus, interactions with the extracellular matrix may control lymphocyte locomotor capacity.

Tratamiento de la hepatitis crónica activa tipo B con interferón leucocitario humano

Se administró interferon leucocitario humano a 62 pacientes con hepatitis crónica activa tipo B, durante un período de seis meses a una dosis total de 400 millones U.I. Se siguieron mensualmente durante un periodo de un año, repitiéndose la biopsia hepática. Las cifras de alanino-amino-transferasa se normalizaron en el 67.7 por ciento de los pacientes y se logró la seroconversión antígeno-anticuerpo o en el 59 por ciento. El antígeno no fue detectable al final del tratamiento en solo cuatro pacientes. La histología hepática mostro una normalización en 17 pacientes y una mejoría en casi la mitad de los mismos. Solo 3 pacientes evolucionaron hacia una cirrosis hepática. Los mejores resultados se apreciaron en sujetos jóvenes y en aquellos que no habían recibido previamente terapia inmunosupresora

Serum androgen levels after streptozotocin administration in the male rat.

Serum levels of testosterone and dihydrotestosterone were measured in control and diabetic animals 5, 10 and 15 days after streptozotocin administration. The diabetic state produced a marked reduction in serum androgen levels 10 and 15 days after streptozotocin administration. Insulin treatment partially restored the circulating androgen levels when administered to diabetic rats.

[Long-term study of patients with active chronic hepatitis treated with leukocyte interferon]. / Estudio a largo plazo de pacientes con hepatitis crónica activa tratados con interferón leucocitario.

Seven patients with active chronic hepatitis who received Interferon Alfa showed a marked humoral and histological improvement one year after the treatment was concluded, in 3 patients the hepatic histology was almost normal, 2 evolved to a persistent chronic hepatitis and only one showed deterioration. These results differed from those obtained immediately after the treatment (p 0.05). Interferon Alfa proves to be of the most usefulness in this disease, studies should be continued up to one year after the treatment has ended.

Estudio a largo plazo de pacientes con hepatitis crónica activa tratados con interferón leucocitario. / [Long-term study of patients with active chronic hepatitis treated with leukocyte interferon]

Seven patients with active chronic hepatitis who received Interferon Alfa showed a marked humoral and histological improvement one year after the treatment was concluded, in 3 patients the hepatic histology was almost normal, 2 evolved to a persistent chronic hepatitis and only one showed deterioration. These results differed from those obtained immediately after the treatment (p 0.05). Interferon Alfa proves to be of the most usefulness in this disease, studies should be continued up to one year after the treatment has ended.

Induction of motility and alteration of surface membrane polypeptides in lymphocytes by contact with autologous and allogeneic fibroblasts.

Contact with cultured fibroblasts induced and maintained motile behavior in autologous and allogeneic human lymphocytes. After 3 h of contact with fibroblasts, 50 +/- 19% of the autologous lymphocytes were motile and after 24 h the corresponding figure was 49 +/- 18%. On a plastic surface the number of motile lymphocytes in the same individuals generally persisted below 15%. SDS-PAGE of iodine-labeled lymphocytes indicated that contact with fibroblasts but not with plastic for a 3-h period caused the appearance of a 300-kda band and the disappearance of several bands of lower molecular weight. During the course of T-lymphocyte activation by concanavalin A or allogeneic cells on a plastic surface, the number of motile forms did not reach a maximum (30 to 50% in separate individuals) until after 2 to 4 days in culture. Thus, in terms of both rate of development and number of motile forms, the fibroblast-dependent motility mechanism was more effective than conventional lymphocyte activation to blast transformation. Conditioned medium from fibroblasts did not induce motile behavior in the lymphocytes and did not provoke alteration of surface membrane polypeptides as revealed by iodination. Fibroblasts also triggered lymphocyte locomotion in serum-free medium, but their triggering effect was enhanced markedly by serum. The development and maintenance of lymphocyte motility required protein synthesis. These data suggest that during contact with fibroblasts lymphocytes acquire locomotor capacity by a mechanism different from activation to blast transformation.