Effects of potential neurotoxic pesticides on hearing loss: a review.

Several pesticides are supposed to be neurotoxic for humans, consequently, they may also affect the auditory system. This review analyzes human and experimental animal studies testing the hypothesis that exposure to pesticides is associated with hearing loss. The literature on this topic is still sparse and methodological limitations of some papers evaluated are identified. As a whole, available data indicate a possible ototoxic action of pesticides, but alternative hypotheses could not be ruled out, also considering some confounders, such as the co-exposure to noise. Therefore, further studies are necessary in order to clarify the association between pesticides exposure and hearing loss. While awaiting more evidence, for precautionary action we recommend considering pesticides as possible ototoxic agents, in particular for vulnerable targets, such as pregnant women and children during early development.

Phase III study in stage IV non-small-cell lung cancer patients treated with two courses of cisplatin/gemcitabine followed by a randomization to three additional courses of the same combination or gemcitabine alone.

BACKGROUND: This randomised phase III study investigated if in responsive and stable disease (SD) stage IV patients after two courses of cisplatin and gemcitabine, single-agent gemcitabine (experimental arm) was not inferior in terms of overall survival (OS) to cisplatin-gemcitabine (standard arm). PATIENTS AND METHODS: Noninferiority was defined as an increase in the hazard of death (HR) < or = 1.33 in the experimental arm. From January 2001 to February 2004, 340 patients were registered and 250 were randomised. Cisplatin was administered on day 1 at 75 mg/m2 and Gemcitabine on days 1 and 8 at 1250 mg/m2 every 3 weeks. RESULTS: Response rate after two courses was 29%. The 1-year progression-free survival was 13% in both arms. One-year survival was 52% in the standard and 42% in the experimental arm for an HR of 1.21 [90% confidence interval (CI) 0.97-1.51]. Postprogression survival was in favour of the standard arm (HR 1.30, 95% CI 0.99-1.70, P = 0.051). Grades 3-4 toxicity favoured in the experimental arm. CONCLUSION: In responsive and SD patients with stage IV non-small-cell lung cancer it was not possible to demonstrate that three courses of gemcitabine alone are not inferior, in terms of OS, to the standard approach of three courses of cisplatin-gemcitabine.

T cell apoptosis by tryptophan catabolism.

Indoleamine 2,3-dioxygenase (IDO) is a tryptophan-catabolizing enzyme that, expressed by different cell types, has regulatory effects on T cells resulting from tryptophan depletion in specific local tissue microenvironments. Different mechanisms, however, might contribute to IDO-dependent immune regulation. We show here that tryptophan metabolites in the kynurenine pathway, such as 3-hydroxyanthranilic and quinolinic acids, will induce the selective apoptosis in vitro of murine thymocytes and of Th1 but not Th2 cells. T cell apoptosis was observed at relatively low concentrations of kynurenines, did not require Fas/Fas ligand interactions, and was associated with the activation of caspase-8 and the release of cytochrome c from mitochondria. When administered in vivo, the two kynurenines caused depletion of specific thymocyte subsets in a fashion qualitatively similar to dexamethasone. These data suggest that the selective deletion of T lymphocytes may be a major mechanism whereby tryptophan metabolism affects immunity under physiopathologic conditions.

IL-6 inhibits the tolerogenic function of CD8 alpha+ dendritic cells expressing indoleamine 2,3-dioxygenase.

The outcome of dendritic cell (DC) presentation of tumor and/or self peptides, including P815AB (a tumor peptide of murine mastocytoma cells) and NRP-A7 (a synthetic peptide mimotope recognized by diabetogenic T cells), may depend on a balance between the activities of immunogenic (CD8alpha(-)) and tolerogenic (CD8alpha(+)) DC. By virtue of their respective actions on CD8(-) and CD8(+) DC, IL-12 and IFN-gamma have functionally opposing effects on peptide presentation by the CD8(-) DC subset, and IFN-gamma-activated CD8(+) DC mediate tolerogenic effects that prevail over the adjuvant activity of IL-12 on CD8(-) DC. We have previously shown that CD40 ligation abrogates the tolerogenic potential of CD8(+) DC, an effect associated with an impaired capacity of the CD40-modulated and IFN-gamma-treated DC to degrade tryptophan and initiate T cell apoptosis in vitro. We report here that IL-6 may both replace (upon administration of the recombinant cytokine) and mediate (as assessed by the use of neutralizing Abs) the effect of CD40 ligation in ablating the tolerogenic activity of CD8(+) DC. The activity of IL-6 includes down-regulation of IFN-gammaR expression in the CD8(+) DC subset and correlates to a reduced ability of these cells to metabolize tryptophan and initiate T cell apoptosis in vitro.

Positive regulatory role of IL-12 in macrophages and modulation by IFN-gamma.

Similar to myeloid dendritic cells, murine macrophages and macrophage cell lines were found to express a surface receptor for IL-12. As a result, peritoneal macrophages could be primed by IL-12 to present an otherwise poorly immunogenic tumor peptide in vivo. Using binding analysis and RNase protection assay, we detected a single class of high affinity IL-12 binding sites (K(d) of approximately 35 pM) whose number per cell was increased by IFN-gamma via up-regulation of receptor subunit expression. Autocrine production of IL-12 was suggested to be a major effect of IL-12 on macrophages when the cytokine was tested alone or after priming with IFN-gamma in vitro. In vivo, combined treatment of macrophages with IFN-gamma and IL-12 resulted in synergistic effects on tumor peptide presentation. Therefore, our findings suggest a general and critical role of IL-12 in potentiating the accessory function of myeloid APC.

Quality evaluation of health care offered by an Internal Medicine Department.

BACKGROUND: The aim of this work is to evaluate the quality and patients satisfaction for given services in an Internal Medicine Department during three months. METHODS: A questionnaire was given to all the patients admitted to our Medicine Department to evaluate our strength and to correct weakness. RESULTS: Our patients assessed doctors and nursing staff for skill and dedication. They gave suggestions about hotel management: bathroom cleaning and number of beds in the same room. They also asked for a pharmacy and a post office inside the hospital. CONCLUSIONS: It appears that our ward gives a satisfactory health care situation. Some of our patients suggestions can be put into practice in a short time, while others require longer, depending on public resources and not on private, such as happens, on the contrary, in the United States.

CD40 ligation ablates the tolerogenic potential of lymphoid dendritic cells.

The outcome of dendritic cell (DC) presentation of P815AB, a tolerogenic tumor/self peptide, depends on a balance between the respective immunogenic and tolerogenic properties of myeloid (CD8 alpha(-)) and lymphoid (CD8 alpha(+)) DC. We have previously shown that CD8(-) DC can be primed by IL-12 to overcome inhibition by the CD8(+) subset and initiate immunogenic presentation in vivo when the two types of peptide-pulsed DC are cotransferred into recipient hosts. IFN-gamma enhances the inhibitory activity of CD8(+) DC on Ag presentation by the other subset, blocking the ability of IL-12-treated CD8(-) DC to overcome suppression. We report here that CD40 ligation on lymphoid DC ablated their inhibitory function on Ag presentation as well as IFN-gamma potentiation of the effect. CD40 modulation of IFN-gamma action on lymphoid DC involved a reduction in IFN-gamma R expression and tryptophan-degrading ability. This effect was accompanied in vitro by an impaired capacity of the CD40-modulated and IFN-gamma-treated DC to initiate T cell apoptosis. In vivo, not only did CD40 triggering on lymphoid DC abrogate their tolerogenic activity, but it also induced the potential for immunogenic presentation of P815AB. Importantly, a pattern similar to P815AB as well as CD40 modulation of lymphoid DC function were observed on testing reactivity to NRP, a synthetic peptide mimotope recognized by diabetogenic CD8(+) T cells in nonobese diabetic mice.

Th1 and Th2 cell clones to a poorly immunogenic tumor antigen initiate CD8+ T cell-dependent tumor eradication in vivo.

Although CD8(+) T cells play a central role as immune effectors, CD4(+) T cells act to control the activation and persistence of the CD8(+) T cell response in autoimmune disease, antiviral immunity, and experimental systems with immunogenic model tumor Ag. However, little information is available on the effects of CD4(+) T cells on the function of endogenous CD8(+) T lymphocytes recognizing authentic tumor rejection Ag with limited immunogenicity. We report here that the prophylactic or postchallenge administration of T helper Th1-type and Th2-type CD4(+) clones specific for an unmutated rejection Ag (murine P815AB, resembling tumor-specific shared Ag in humans) leads to the induction of P815AB-specific reactivity in vivo and concomitant tumor destruction, with quantitative rather than qualitative differences characterizing the antitumor activity of Th1 vs Th2 cells. Because the transferred CD4(+) cells lacked direct antitumor activity in vitro and required the de novo generation of P815AB-specific CD8(+) T cells in vivo, these findings suggest that CD4(+) lymphocytes can enhance the ability of host APC to initiate an endogenous CD8(+) T cell response to authentic, poorly immunogenic tumor rejection Ag.

IFN-gamma inhibits presentation of a tumor/self peptide by CD8 alpha- dendritic cells via potentiation of the CD8 alpha+ subset.

Using an in vivo model of tumor/self peptide presentation for induction of class I-restricted skin test reactivity, we have previously shown that a minority population of CD8+ dendritic cells (DC) negatively regulates the induction of T cell reactivity by peptide-loaded CD8- DC in DBA/2 mice. However, the CD8- fraction can be primed by IL-12 to overcome inhibition by the CD8+ subset when the two types of DC are cotransferred into recipient hosts. We report here that exposure of CD8+ DC to IFN-gamma greatly enhances their inhibitory activity on Ag presentation by the other subset, blocking the ability of IL-12-treated CD8- DC to overcome suppression. In contrast, IFN-gamma has no direct effects on the APC function of the latter cells and does not interfere with IL-12 signaling. The negative regulatory effect triggered by IFN-gamma in CD8+ DC appears to involve interference with tryptophan metabolism in vivo. Through tryptophan depletion affecting T cell responses, IFN-gamma acting on CD8+ DC may thus contribute to regulation of immunity to tumor/self peptides presented by the CD8- subset.

Improved fibrinolytic capacity after withdrawal of steroid immunosuppression in renal transplant recipients.

BACKGROUND: Long-term steroid immunosuppression has been associated with the prothrombotic state observed in renal transplant (RT) patients, in whom both hypercoagulability due to an increase of von Willebrand factor/factor VIII complex, and impaired fibrinolysis due to PAI-1 excess have been demonstrated. Our aim was to investigate the effect of steroid withdrawal on fibrinolytic capacity in a group of RT patients. METHODS: The fibrinolytic study was performed in 28 RT patients under stable immunosuppression therapy with cyclosporine, azathioprine, and methylprednisolone; only 12 of these patients could repeat the study 6 months after steroid withdrawal. Euglobulin lysis time (ELT), tissue plasminogen activator activity (t-PA:act) and antigen (t-PA:Ag), PAI-1 activity (PAI-1:act), and antigen (PAI-1:Ag) were assayed on blood samples drawn before and 20 min after the venous occlusion test (VO). RESULTS: An hypofibrinolytic state due to a significant increase in PAI-1 levels was confirmed in RT patients receiving triple immunosuppression therapy. RT patient who stayed off steroids showed a significant shortening of ELT both before (P=0.01) and 20' after VO (P=0.005) at the 6-month control. Moreover, after steroid withdrawal, PAI-1:Ag levels decreased significantly (P=0.002) and normalized; in a similar manner PAI-1:act levels also showed a significant decrease both before (P=0.001), and after VO (P=0.0001). The prevalence of RT patients with impaired fibrinolytic capacity was as high as 83.3% during steroid treatment, and dropped to 16.7% after steroid withdrawal. CONCLUSIONS: Our findings confirm that steroid withdrawal may normalize impaired fibrinolytic capacity in RT patients; this improvement may further contribute to reduce the thrombotic risk associated with renal transplantation.

IL-12 induces SDS-stable class II alphabeta dimers in murine dendritic cells.

We assessed the effect of rIL-12 on the expression of class II molecules and on the ratio between SDS-stable and unstable alphabeta dimers in dendritic cells. We found that in vitro exposure of the cells to IL-12 increased their surface expression of mature class II molecules, despite a marked decline in class II biosynthesis. This effect was accompanied by a striking increase in the overall proportion of SDS-stable heterodimers.

Dual effect of IL-4 on resistance to systemic gram-negative infection and production of TNF-alpha.

To determine the effect of interleukin 4 (IL-4) administration in a live sepsis model characterised by high-level production of tumour necrosis factor a (TNF-alpha), mice infected systemically with lethal or sublethal inocula of Pseudomonas aeruginosa were given the recombinant cytokine at different times before infection. Improved survival and decreased TNF-alpha production were observed in lethally infected mice treated with the cytokine 1 day before challenge. In contrast, increased mortality and overproduction of TNF-alpha were observed in sublethally infected mice given IL-4 at the time of infection.

IL-9 protects mice from Gram-negative bacterial shock: suppression of TNF-alpha, IL-12, and IFN-gamma, and induction of IL-10.

IL-9 is a T cell-derived cytokine that, similar to the Th2 cytokines IL-4 and IL-10, has been implicated in the response to parasitic infections, allergy, and inflammatory processes. Because both IL-4 and IL-10 can confer protection to mice from septic shock, we investigated whether IL-9 may also be capable of conferring resistance on recipients of an otherwise lethal challenge with Pseudomonas aeruginosa. Prophylactic injections of rIL-9 appeared to be most effective in preventing the onset of a lethal shock, according to a pattern that was both dose dependent and time dependent. The protective effect of IL-9 was correlated with marked decreases in the production of the inflammatory mediators TNF-alpha, IL-12, and IFN-gamma, as well as the induction of the anti-inflammatory cytokine IL-10. Sustained levels of IL-9-specific transcripts could be detected in the spleens of mice recovering from sublethal P. aeruginosa infection. Therefore, IL-9 may be protective in septic shock via a rather unique mechanism involving a complex modulation of inflammatory and anti-inflammatory mediators.

IL-12 acts selectively on CD8 alpha- dendritic cells to enhance presentation of a tumor peptide in vivo.

Previous work has shown that a significant proportion of murine splenic dendritic cells (DC) express a high affinity receptor for IL-12, thus accounting for the adjuvanticity of the cytokine when DBA/2 mice are transferred with syngeneic DC exposed in vitro to rIL-12 and an otherwise poorly immunogenic tumor peptide. In DBA/2 mice, splenic DC consist of 90-95% CD8- and 5-10% CD8+ cells. To detect any difference in IL-12 responsiveness among phenotypically distinct DC subtypes, enriched CD8- (>99% pure) and CD8+ ( approximately 80% pure) populations of DC from DBA/2 spleens were assayed for APC function in vivo following exposure to rIL-12 and tumor peptide in vitro. Unlike unfractionated DC, the CD8- fraction was capable of effective presentation of the peptide even when the cells had not been pretreated with IL-12 before peptide pulsing. The addition of as few as 3% CD8+ cells during pulsing blocked in vivo priming by the CD8- fraction. However, pretreatment of CD8- DC with IL-12 before cell mixing and peptide pulsing ablated the inhibitory effect of the CD8+ fraction. CD8-, but not CD8+, DC showed significant message expression for the beta 1 and beta 2 subunits of the IL-12 receptor. These data suggest that a minority population of CD8+ DC, which appeared to secrete IL-10 in vitro, negatively regulates the induction of T cell reactivity by peptide-loaded CD8- DC in DBA/2 mice. However, the CD8- fraction can be primed by IL-12 to overcome the inhibitory effect of the CD8+ subtype.

Immunogenicity of tumor peptides: importance of peptide length and stability of peptide/MHC class II complex.

Nonameric P815AB, a cytotoxic-T-lymphocyte-defined minimal core peptide encoded by the murine mastocytoma gene P1A, fails to initiate CD4+ cell-dependent reactivity in vivo to class-I-restricted epitopes when mice are administered peptide-pulsed dendritic cells. Effective immunization requires T helper effects, such as those mediated by coimmunization with class-II-restricted (helper) peptides or by the use of recombinant interleukin-12 (rIL-12). Although P815AB does possess class-II-restricted epitopes, they are likely suboptimal, resulting in poor affinity and/or stability of MHC/P815AB complexes and inadequate activation of the antigen-presenting cell function of dendritic cells. The present study has examined a series of longer, P815AB-centered peptides (11-14 amino acids in length, all P1A-encoded) for their ability to initiate CD4+ and CD8+ cell-mediated responses to the nonamer in vivo, their ability to bind class II MHC in vitro, and their ability to assemble class II molecules stably. By means of a class-I-restricted skin test assay in mice receiving peptide-pulsed dendritic cells, we found that a 12-mer and a 13-mer effectively immunized against the core P815AB peptide, and that this correlated with IL-2 production in vitro by CD4+ cells in response to the nonamer. In vitro studies, involving affinity-purified class II molecules, showed that the capacity to assemble class II molecules stably, more than the affinity for class II MHC, correlated with the ability of the different P815AB peptides to prime the host to the core peptide seen by the T cells.

Autocrine IL-12 is involved in dendritic cell modulation via CD40 ligation.

Ligation of CD40 on dendritic cells (DC) triggers production of IL-12. Using an adoptive transfer model we have previously shown that rIL-12 acts directly on DC to enhance presentation of an otherwise poorly immunogenic tumor peptide. Using the same experimental model, we now describe a similar adjuvanticity of CD40 ligation on peptide presentation by DC. We also explore the possibility that the IL-12 resulting from CD40 ligation directly affects the APC function of DC, mediating or contributing to the adjuvant effect of CD40 ligation. CD40 engagement in vitro and rIL-12 at concentrations in the range induced by CD40 ligation were equally effective in priming DC for presentation of the tumor peptide in vivo. Remarkably, the copresence in vitro of neutralizing Ab to IL-12, but not to TNF-alpha, IL-1beta, or IFN-gamma, ablated the enhancing effect of CD40 engagement on the APC function of DC. These data suggest a major role for autocrine IL-12 in DC modulation via CD40 ligation.

IL-12 acts directly on DC to promote nuclear localization of NF-kappaB and primes DC for IL-12 production.

We analyzed the expression of an IL-12 receptor by fresh dendritic cells (DC) and a DC line. Using RT-PCR, RNAse protection, and electrophoretic mobility shift assay analysis, we found that DC possess an IL-12 receptor with beta1 subunit (downstream box 1)-related differences from that on T cells. IL-12 signaling through this receptor involved members of the NF-KB but not STAT family. The unique properties of the IL-12 receptor on DC, characterized by a single class of binding sites with a Kd of about 325 pM, may underlie rather unique effects, such as IFNgamma-independent augmentation of class II antigen expression and priming for LPS-induced production of IL-12.