Effects of sodium fusidate in animal models of insulin-dependent diabetes mellitus and septic shock.

We have evaluated the effects of the novel immunosuppressant sodium fusidate (fusidin) in the non-obese diabetic (NOD) mouse and in D-galactosamine (D-Gal)-presensitized BALB/c mice challenged with the bacterial superantigen, Staphylococcus aureus enterotoxin B (SEB) or with the endotoxin, Escherichia coli lipopolysaccharide (LPS). The NOD mouse model has clinical and histoimmunological features similar to those of human insulin-dependent diabetes mellitus (IDDM). The SEB- and LPS-treated BALB/c mouse models exhibit pathogenic similarities with human septic shock conditions. In the NOD mouse, fusidin suppressed the spontaneous development of insulitis (mean inhibition 73%) and hyperglycaemia (IDDM incidence 25% versus 0%) when administered at 40 mg/kg five times weekly for 8 consecutive weeks from the fourth week of age; concurrently treated animals exhibited reduced percentages of splenic T lymphocytes. This anti-diabetogenic effect was confirmed in the accelerated model of diabetes induced in the NOD mouse with cyclophosphamide (CY) (IDDM incidence 55% versus 21-6% using dosages of fusidin from 40 to 80 mg/kg five times weekly); protection from IDDM development was achieved even when the drug (80 mg/kg/day) was first administered 7 days after CY challenge. In contrast, fusidin did not reverse hyperglycaemia when administered to CY-treated animals within 3 days of IDDM development. In the two models of septic shock, prophylactic treatment with fusidin, 80 mg/kg given three times for 2 days prior to D-Gal/SEB or D-Gal/LPS challenge, drastically reduced the lethality compared with D-Gal/buffer-treated mice. This effect may depend on the inhibitory action of fusidin on the secretion of cytokines such as interferon-gamma and tumour necrosis factor-alpha, the serum levels of which were greatly diminished in the fusidin-treated mice (mean inhibition 50-90%). These results demonstrate that fusidin may have a role in the treatment of cell-mediated autoimmune diseases and cytokine-mediated infectious diseases in humans.     

Histones interact with anionic phospholipids with high avidity; its relevance for the binding of histone-antihistone immune complexes.

Antibodies recognizing anionic phospholipids have been described in systemic lupus erythematosus (SLE) and other autoimmune diseases. Recent studies have shown that some of these antibodies may recognize a cardiolipin-binding protein (apolipoprotein H) rather than phospholipids. A similar possibility is conceivable for other cardiolipin-binding proteins that are targets of autoantibodies. In this study we have addressed whether this might be the case for histones, a set of highly cationic and widely distributed proteins that react in a well known autoantibody system. Our results indicate that: (i) histones bind to anionic phospholipids (cardiolipin and phosphatidylserine) with high avidity, but not to zwitterionic phospholipids (phosphatidylcholine); (ii) monoclonal and polyclonal antihistone antibodies recognize histones bound to cardiolipin; (iii) the addition of histones to serum samples containing antihistone antibodies often enhances their anticardiolipin reactivity. In addition, we have found that antihistone-producing hybridomas derived from MRL-lpr mice may show anticardiolipin activity due to the presence of histones in the cell culture supernatants with the resultant formation of immune complexes. Taken together, the results suggest a potential role for histones in the anti-cardiolipin activity detected in sera containing antihistone antibodies. These histone-phospholipid interactions should be taken into account when evaluating the pathogenic effects of antihistone antibodies or other autoantibodies reacting with nuclear components (e.g. nucleosomes) containing histones.     

Apoptotic DNA binds to HLA class II molecules inhibiting antigen presentation and participating in the development of anti-inflammatory functional behavior of phagocytic macrophages

Resident macrophages are mainly responsible for the clearance of apoptotic cells from tissue by phagocytosis. Phagocytosis of apoptotic cells is not accompanied by activation of inflammatory mechanisms, unlike what happens when necrotic phenomena occur. We analyzed the effect of phagocytosis of apoptotic bodies on macrophage cell functions. After phagocytosis of apoptotic cells macrophages were unable to present an exogenous antigen to autologous antigen-specific T-cell lines. The inhibition was mediated by different mechanisms including binding of apoptotic DNA to human leukocyte antigen (HLA) class II molecules of macrophages, decreased expression of co-stimulatory molecules and increased secretion of tumor growth factor beta (TGFbeta). When dendritic cells were cultured with macrophages phagocytosing apoptotic cells, or with their supernatant, impaired dendritic cell antigen presenting activity and reduced tumor necrosis factor alpha (TNFalpha) secretion were found. Our results suggest that: (1) the phagocytosis of apoptotic bodies inhibits macrophage antigen presentation; (2) such inhibition is mediated by the binding of apoptotic DNA to macrophage HLA class II molecules as well as by the activation of biological mechanisms that induce an anti-inflammatory functional behavior in macrophages; and (3) macrophages phagocytosing apoptotic cells inhibit antigen presentation of neighboring dendritic cells via TGFbeta secretion. These events are likely related to the preservation of healthy tissues from the onset of inflammation.     

Study of molecular mechanisms of pro-apoptotic activity of NCX 4040, a novel nitric oxide-releasing aspirin, in colon cancer cell lines

Background  

Despite numerous studies aimed at verifying the antitumor activity of nitric oxide-releasing nonsteroidal antiflammatory drugs (NO-NSAIDs), little is known about the molecular targets responsible for their antineoplastic properties. In the present study, we investigated the mechanisms underlying the cytotoxicity of NCX 4040, a novel NO-aspirin with promising antineoplastic action, in in vitro human colon cancer models.     

Mediastinal large-cell lymphoma with sclerosis

Summary The Southern blot hybridization technique has been applied to study the configuration of immunoglobulin and T-cell receptor genes in 6 cases of the so called mediastinal large cell lymphoma with sclerosis. This lymphoma has been recently recognized as a separate entity among non-Hodgkin lymphomas mainly affecting young adult patients. The B-cell origin of this neoplasm was suggested by means of immunohistochemical analysis. However, the immunophenotypical B-cell related markers used do not always exhibit lineage fidelity. The Southern blot analysis demonstrated the presence of unique heavy and k-light chain immunoglobulin gene rearrangements, establishing genotypically their B-cell origin.This work was supported by the Associazione Italiana per la Ricerca sul Cancro, Milano, Italy, and Progetto finalizzato Oncologia (contratto n^o 86.00461.44), CNR, Rome, Italy. Aldo Scarpa and Maurizio Lestani are supported by a Scholarship from the Associazione Italiana per la Ricerca sul Cancro, Milano, Italy     

Immune Response Induced in Vitro by CD16− and CD16+ Monocyte-Derived Dendritic Cells in Patients with Metastatic Renal Cell Carcinoma Treated with Dendritic Cell Vaccines

Monocyte-derived dendritic cells (mDC) are increasingly used as cancer vaccines. However, human monocytes are a heterogeneous cell population. We showed previously that DC derived from a monocyte subset expressing CD16 (16+mDC) stimulated allogeneic naïve T lymphocytes to secrete higher levels of IL-4 than DC derived from regular CD14^highCD16^? monocytes (16?mDC). Th1-type responses have been associated with effective antitumor responses, thus the use of mDC containing 16+mDC as cancer vaccines might be disadvantageous. Here, we evaluate the primary and memory immune response elicited in vitro by 16+mDC and 16?mDC in five patients with metastatic renal cell carcinoma vaccinated with autologous mDC pulsed with tumor lysates (TuLy) and keyhole limpet hemocyanin (KLH). After therapy, three of the five patients had stable disease. Surprisingly, patients with longer survival showed the highest amount of peripheral blood CD16+ monocytes. Analysis of KLH-specific antibodies revealed high titers of IgG2 in patients with longer survival. CD4+ T lymphocyte proliferation against KLH and TuLy increased after treatment, and some patients showed an augmented rate of CD4+ T lymphocyte proliferation against KLH (3/5) and TuLy (2/3) when 16+mDC were used as antigen presenting cells (APC). Before treatment, the IFN-γ/IL-4 ratio against TuLy and KLH was higher when using 16?mDC as APC, but after vaccination four of five patients had an increased ratio for TuLy with 16+mDC. These results suggest that the immune response elicited by 16?mDC and 16+mDC is modified when memory or naïve T cells are stimulated, and 16+mDC could favor a stronger and more beneficial antitumoral Th1 memory response in vivo.     

Muscle strength, volume and activation following 12-month resistance training in 70-year-old males

In elderly males muscle plantar flexor maximal voluntary contraction (MVC) torque normalised to muscle volume (MVC/VOL) is reduced compared to young males as a result of incomplete muscle activation in the elderly. The aim of the present study was to determine the influence of a 12-month resistance training programme on muscle volume, strength, MVC/VOL, agonist activation and antagonist coactivation of the plantarfexors in elderly males. Thirteen elderly males aged 70 years and over (range 70–82 years), completed a 12-month whole body resistance-training programme (TRN), training three times a week. Another eight males (range 18–30 years), who maintained their habitual physical activity for the same 12-month period as the TRN group acted as controls (CTRL). Isometric plantarflexor maximal voluntary contraction (MVC) torque increased in the TRN group by 20% (P<0.01), from 113.1±22.0 Nm to 141.5±19.2 Nm. Triceps surae volume (TS VOL) assessed using MRI, increased by 12%, from 796.3±78.9 cm³ to 916.8±144.4 cm³ . PF activation, measured using supramaximal double twitch interpolation, increased from 83.6±11.0% pre training, to 92.1±7.6% post training (P<0.05). Dorsiflexion MVC and antagonist coactivation (assessed using surface electromyography) did not change with training. Plantarflexor MVC torque normalized for triceps surae muscle volume (MVC/VOL) was 142.6±32.4 kN m^–2 before training and 157.0± 27.9 kN m^–2 after training (a non-significant increase of 8%). No significant change in any measurement was observed in the CTRL group. This study has shown that the gain in muscle strength in response to long-term (12-month) training in older men is mostly accounted for by an increased muscle volume and activation.     

CD10 is expressed in a subset of chromophobe renal cell carcinomas.

CD10 has been considered a useful marker in the diagnosis of renal carcinomas, because of its expression in clear cell and papillary renal cell carcinomas and its absence in chromophobe renal cell carcinomas. On the other hand, chromophobe renal cell carcinoma expresses parvalbumin, which is absent in clear cell and papillary renal cell carcinomas. To further address the relevance of these markers, we studied the expression of CD10 and parvalbumin in 42 samples of chromophobe renal cell carcinoma (seven of which had aggressive features, including invasion beyond the renal capsule, renal vein invasion, metastases, or sarcomatoid transformation), 75 clear cell renal cell carcinomas (eight metastatic) and 51 papillary renal cell carcinomas (two metastatic). CD10 was found in 100% of clear cell renal cell carcinomas, 63% of papillary renal cell carcinomas and in all metastatic cases of both types. At variance with previous studies, we found CD10 expression in from 30 to 90% of the neoplastic cells, in 11 of 42 (26%) chromophobe renal cell carcinomas. The CD10-positive cases included five of the seven (71%) chromophobe renal cell carcinoma with aggressive features. Statistical analysis showed significant association of CD10-positive tumors with clinicopathologic aggressiveness (P=0.003) and mitotic figures (P=0.04). Parvalbumin was strongly expressed in all primary and metastatic chromophobe renal cell carcinomas. Western blot analysis was utilized to confirm the expression of both CD10 and parvalbumin in chromophobe renal cell carcinomas.     

Vilse, a conserved Rac/Cdc42 GAP mediating Robo repulsion in tracheal cells and axons

Slit proteins steer the migration of many cell types through their binding to Robo receptors, but how Robo controls cell motility is not clear. We describe the functional analysis of vilse, a Drosophila gene required for Robo repulsion in epithelial cells and axons. Vilse defines a conserved family of RhoGAPs (Rho GTPase-activating proteins), with representatives in flies and vertebrates. The phenotypes of vilse mutants resemble the tracheal and axonal phenotypes of Slit and Robo mutants at the CNS midline. Dosage-sensitive genetic interactions between vilse, slit, and robo mutants suggest that vilse is a component of robo signaling. Moreover, overexpression of Vilse in the trachea of robo mutants ameliorates the phenotypes of robo, indicating that Vilse acts downstream of Robo to mediate midline repulsion. Vilse and its human homolog bind directly to the intracellular domains of the corresponding Robo receptors and promote the hydrolysis of RacGTP and, less efficiently, of Cdc42GTP. These results together with genetic interaction experiments with robo, vilse, and rac mutants suggest a mechanism whereby Robo repulsion is mediated by the localized inactivation of Rac through Vilse.