Scalable signaling mediated by T cell antigen receptor-CD3 ITAMs ensures effective negative selection and prevents autoimmunity.

The T cell antigen receptor (TCR)-CD3 complex is unique in having ten cytoplasmic immunoreceptor tyrosine-based activation motifs (ITAMs). The physiological importance of this high TCR ITAM number is unclear. Here we generated 25 groups of mice expressing various combinations of wild-type and mutant ITAMs in TCR-CD3 complexes. Mice with fewer than seven wild-type CD3 ITAMs developed a lethal, multiorgan autoimmune disease caused by a breakdown in central rather than peripheral tolerance. Although there was a linear correlation between the number of wild-type CD3 ITAMs and T cell proliferation, cytokine production was unaffected by ITAM number. Thus, high ITAM number provides scalable signaling that can modulate proliferation yet ensure effective negative selection and prevention of autoimmunity.

Impact of arsenic-polluted groundwater on soil and produce quality: a food chain study.

The experiment was conducted to assess the impact of arsenic (As)-contaminated groundwater irrigation on soil health and crop quality. Geo-referenced groundwater, soil, and crop produce samples were collected from the middle Gangetic plains of Maner block of Patna and were analyzed for As content. The result showed that long-term application of As-contaminated groundwater (0.017 to 0.677 mg L-1) buildup significant amount of As in the soil (0.41 to 8.66 mg kg-1). A significant correlation (r2 = 0.922) was also observed between As content in groundwater and the soil. The content of As in groundwater also affected crop quality and accumulated metal content in different crop parts. Total As content in crop samples ranged from 0.010 to 0.963 µg g-1 of dry weight. The average As content in crop followed order: oilseeds > cereals > vegetables > pulses. Therefore, produce quality should be monitored frequently for As uptake as there is a great chance of As accumulation in food crops. Hence, these approaches are useful for the formulation of policy guidelines for the management of As-containing groundwater and routine risk assessment of As-contaminated soils.

Sulfonated nitrogen and added-sulfur sources influence productivity, quality, and nutrient acquisition of soybean-wheat cropping system.

Soybean-wheat is the predominant cropping system covering >2.5 Mha area in India. The lower productivity of soybean-wheat cropping system (SWCS), remains a serious concern primarily due to inadequate nutrient management. Increasing sulfur (S) deficiency is widespread, especially under oilseed-based cropping system. Hence, to standardize the S requirement through customized fertilization, an experiment was conducted in completely randomized block design (RBD) comprised of 12 nutrient sources, replicated thrice. The study aims to evaluate the agronomic performance of sulfonated nitrogen (SN) in comparison to conventional S nutrient sources in SWCS. The maximum soybean productivity was recorded under NPK + S through 40-0-0-13 (SN1), although NPK + 50% S (15 kg/ha) as basal and 50% (15 kg/ha) as top dressing through SN2 10-0-0-75 produced maximum wheat grain yield. When compared with no nitrogen (control), the application of 30 kg S ha-1 to both crops increased the productivity of the soybean-wheat cropping system up to 39%. The maximum system (SWCS) productivity (8.45 tha-1) was obtained with the application of 50% S as basal and 50% as top dressing (SN2-based), remaining N through urea. The highest sustainable yield index of soybean (SYIS), i.e. 0.90 was under SN1+ remaining N through urea and likewise highest sustainable yield index of wheat (SYIW) was under S splitting. The application of SN also improved the nutrient acquisition and grain quality of soybean and wheat with a positive nutrient balance in the soil. The protein content and yield of soybean and wheat grains also improved. The higher gluten content in wheat grain was produced with 60 kg S ha-1 applied. The agronomic efficiency of N and S (AEN and AEs) were highest under SN1 and SN2, respectively (32.8 kg grain/kg N applied; 15 kg grain/kg S applied) in soybean, however in wheat, S splitting and urea application resulted in highest agronomic efficiency (AEN and AES) of N and S (17.1 kg grain/kg N applied; 22.3 kg grain/kg S applied respectively). Hence splitting of S doses of SN along with urea and recommend P, K was found efficient for the soybean-wheat cropping system.

Management of plant nutrient dynamics under alkaline soils through graded application of pressmud and gypsum.

An incubation experiment was conducted to monitor the effect of different organic matter inputs with the graded application of gypsum at different time intervals on soil pH, sodium (Na) content and available plant nutrients like nitrogen (N) and sulphur (S) in alkaline soil. The experiment was formulated with nine treatments, i.e. control (T1), recommended dose of fertilizer (RDF) (T2), RDF+Gyp1 (T3), RDF+FYM5+Gyp2 (T4), RDF+FYM10+Gyp1 (T5), RDF+PM5+Gyp2 (T6), RDF+PM10+Gyp1 (T7), RDF+FYM2.5+PM2.5+Gyp2 (T8), RDF+FYM5+PM5+Gyp1 (T9) with three replications. Periodical soil samples were taken at six and twelve months intervals. Results showed that the addition of organic matter reduced the pH and Na content in the soil. More reduction was observed at one year period as compared to six months. The addition of farmyard manure (FYM) and pressmud (PM) at 10 t/ha with gypsum (1 t/ha) improved available N and available S content as compared to organic inputs (5 t/ha) with gypsum (2 t/ha) in soil. Pressmud application with FYM showed better availability of plant nutrients and a reduction of soil pH (8.39 to 7.79) and Na content from 626 to 391 mEq/L in the soil during the incubation period. During the study, the application of treatment T9 (FYM and PM in equal ratio with 1 t/ha gypsum) showed a better availability of available N (175 to 235 kg/ha) and S (15.44 to 23.24 kg/ha) and reduced the active ion concentration of Na. This study is very useful for the management of sodium toxicity, improving soil health and the mineralization rate of organic matter through the application of organic inputs for sustainable crop production.

Cholinergic modulation of angiogenesis: role of the 7 nicotinic acetylcholine receptor.

Pathological angiogenesis contributes to tobacco-related diseases such as malignancy, atherosclerosis and age-related macular degeneration. Nicotine acts on endothelial nicotinic acetylcholine receptors (nAChRs) to activate endothelial cells and to augment pathological angiogenesis. In the current study, we studied nAChR subunits involved in these actions. We detected mRNA for all mammalian nAChR subunits except alpha(2), alpha(4), gamma, and delta in four different types of ECs. Using siRNA methodology, we found that the alpha(7) nAChR plays a dominant role in nicotine-induced cell signaling (assessed by intracellular calcium and NO imaging, and studies of protein expression and phosphorylation), as well as nicotine-activated EC functions (proliferation, survival, migration, and tube formation). The alpha(9) and alpha(7) nAChRs have opposing effects on nicotine-induced cell proliferation and survival. Our studies reveal a critical role for the alpha(7) nAChR in mediating the effects of nicotine on the endothelium. Other subunits play a modulatory role. These findings may have therapeutic implications for diseases characterized by pathological angiogenesis.

Understanding the motivation for conventional and complementary/alternative medicine use among men with prostate cancer.

HYPOTHESIS: The incidence of prostate cancer and the prevalence of complementary and alternative medicine (CAM) use among prostate cancer patients have been increasing. An understanding of the underlying motivations for men to turn to CAM is essential to achieve optimal outcomes. The authors hypothesized that CAM users and nonusers differ in their assessment of conventional and CAM treatments and explored perceptions, feelings, ideas, and experiences among prostate cancer patients who made a decision to use or not to use CAM. STUDY DESIGN: This qualitative study is based on in-person interviews with prostate cancer patients selected from a population-based survey. METHODS: The authors conducted semistructured interviews with 27 prostate cancer patients of Asian and Caucasian ethnicity, 18 of whom used CAM and 9 of whom did not. Using qualitative research methods, they examined perceptions of conventional medicine and CAM and contrasted viewpoints of CAM users and nonusers. Based on the patients' statements, the authors developed a model representing the viewpoints and thought patterns of CAM users as contrasted with those patients who did not use CAM. RESULTS: The interviews revealed notable differences in viewpoints between CAM users and nonusers in 4 areas. The following themes that were important to CAM users emerged from the analysis: a view of CAM as safe and holistic coupled with a view of conventional medicine as an aggressive and isolated treatment; concern about side effects, in particular, impotence and incontinence from conventional cancer therapy; a belief in the potential efficacy of CAM despite the lack of evidence; and a need to gain a sense of control. Although nonusers expressed similar concerns about side effects of conventional treatment and considered CAM harmless, they assigned different priorities to these issues in their decision making. CONCLUSIONS: In this study, no single theme was solely responsible for CAM use among prostate cancer patients. Instead, multiple ideas woven together led patients toward CAM use. An understanding of patients' thought processes may aid health care professionals in initiating a dialogue about decision making and potential side effects.

Broad-scale phosphoprotein profiling of beta adrenergic receptor (ß-AR) signaling reveals novel phosphorylation and dephosphorylation events.

ß-adrenergic receptors (ß-ARs) are model G-protein coupled receptors that mediate signal transduction in the sympathetic nervous system. Despite the widespread clinical use of agents that target ß-ARs, the signaling pathways that operate downstream of ß-AR stimulation have not yet been completely elucidated. Here, we utilized a lysate microarray approach to obtain a broad-scale perspective of phosphoprotein signaling downstream of ß-AR. We monitored the time course of phosphorylation states of 54 proteins after ß-AR activation mouse embryonic fibroblast (MEF) cells. In response to stimulation with the non-selective ß-AR agonist isoproterenol, we observed previously described phosphorylation events such as ERK1/2(T202/Y204) and CREB(S133), but also novel phosphorylation events such as Cdc2(Y15) and Pyk2(Y402). All of these events were mediated through cAMP and PKA as they were reproduced by stimulation with the adenylyl cyclase activator forskolin and were blocked by treatment with H89, a PKA inhibitor. In addition, we also observed a number of novel isoproterenol-induced protein dephosphorylation events in target substrates of the PI3K/AKT pathway: GSK3ß(S9), 4E-BP1(S65), and p70s6k(T389). These dephosphorylations were dependent on cAMP, but were independent of PKA and correlated with reduced PI3K/AKT activity. Isoproterenol stimulation also led to a cAMP-dependent dephosphorylation of PP1α(T320), a modification known to correlate with enhanced activity of this phosphatase. Dephosphorylation of PP1α coincided with the secondary decline in phosphorylation of some PKA-phosphorylated substrates, suggesting that PP1α may act in a feedback loop to return these phosphorylations to baseline. In summary, lysate microarrays are a powerful tool to profile phosphoprotein signaling and have provided a broad-scale perspective of how ß-AR signaling can regulate key pathways involved in cell growth and metabolism.

Reactivity profiles of broadly neutralizing anti-HIV-1 antibodies are distinct from those of pathogenic autoantibodies.

OBJECTIVE: Broadly neutralizing antibodies (bNt Abs) against HIV-1 are rarely produced during natural infection, and efforts to induce such Abs by vaccination have been unsuccessful. Thus, elucidating the nature and cellular origins of bNt Abs is a high priority for vaccine research. As the bNt monoclonal Abs (MAbs) 2F5, 4E10 and 2G12 have been reported to bind select autoantigens, we investigated whether these MAbs display a broader range of autoreactivity and how their autoreactivity compares with that of pathogenic autoAbs. METHODS: An autoantigen microarray comprising 106 connective tissue disease-related autoantigens and control antigens was developed and used, in combination with ELISAs, to compare the reactivity profiles of MAbs 4E10, 2F5 and 2G12 to those of four pathogenic autoAbs derived from patients with antiphospholipid-syndrome (APS), and to serum from a patient with systemic lupus erythematosus (SLE). RESULTS: The APS MAbs and SLE serum reacted strongly with multiple autoantigens on the microarray, whereas anti-HIV-1 MAb reactivity was limited mainly to HIV-1-related antigens. The APS autoAbs reacted strongly with CL, yet only 4E10 bound CL at high concentrations; both 2F5 and 4E10 bound their HIV-1 epitopes with a 2-3-log higher apparent affinity than CL. Moreover, the polyreactivity of 4E10, but not CL15, could be blocked with dried milk. CONCLUSION: The reactivity profiles of bNt anti-HIV-1 MAbs are fundamentally distinct from those of pathogenic autoAbs that arise from dysregulated tolerance mechanisms. This suggests that the limited polyreactivity observed for the bNt MAbs, and for HIV-1-Nt Abs in general, may arise through alternative mechanisms, such as extensive somatic mutation due to persistent antigen selection during chronic infection.

Comparing short-term outcomes of femoral-popliteal and iliofemoral deep venous thrombosis: early lysis and development of reflux.

This study compares the development of reflux, recanalization, and clinical outcomes of patients with femoral-popliteal and iliofemoral deep venous thrombosis (DVT). Emphasis is placed on the relationship between early lysis of clot through thrombolysis and the development of reflux and post-thrombotic syndrome (PTS) for iliofemoral patients. A retrospective chart review was conducted of 27 femoral-popliteal DVT limbs and 11 iliofemoral DVT limbs with average follow-up of 2.3 and 2.1 years, respectively. Rates of recanalization, development of reflux, and post-thrombotic syndrome were recorded through review of duplex scans and physical examinations. All femoral-popliteal patients received anticoagulant therapy. Nine of 11 iliofemoral patients (82%) received thrombolytic therapy in addition to anticoagulants. Statistical analysis included Kaplan-Meier estimation to take into account dropout in follow-up times, and chi-squared analysis to compare final outcomes. A significantly greater proportion of iliofemoral patients (73%) than femoral patients (31%) remained asymptomatic at the end of their follow-up (p < 0.025). Because of thrombolytic therapy, 82% of iliofemoral limbs showed partial or complete lysis 4 weeks after diagnosis of clot. As expected, only 22% of femoral-popliteal limbs developed some recanalization 4 weeks after diagnosis (p < 0.005). Interestingly, no significant difference in reflux development was observed between the two groups. After an average of 2.1 years, 60% of femoral-popliteal limbs developed reflux in the deep veins vs. 64% for iliofemoral limbs. The iliofemoral DVT patients showed improved clinical outcomes in the short term compared to that of femoral-popliteal patients in this pilot study. The improved clinical outcomes could be attributed to the early lysis of clot via thrombolytic therapy for the iliofemoral group. Although the extent of reflux development was similar in both groups, iliofemoral patients still showed fewer clinical symptoms after follow-up. This may suggest that the presence of both residual obstruction and reflux, rather than either one alone, significantly increases the chances for development of PTS. Since thrombolytics eliminates at least one of these factors, residual obstruction, it may aid in decreasing development of PTS in the short term. The data in this retrospective study warrant further long-term prospective analysis of thrombolysis and its relationship with PTS.