Modulating furin activity with designed mini-PDX peptides: synthesis and in vitro kinetic evaluation

A peptide was designed from reactive site loop structure of alpha1 Antitrypsin Portland known as alpha1 PDX as a novel mini-PDX inhibitor of furin. The sequence was derived from (367-394) that contains the crucial furin cleavage motif RIPR382. A P3 mutant replacing Ile380 by Leu was prepared as a first model peptide. A Cys residue was inserted at each terminal of the peptide for purpose of cyclisation which was accomplished by air or iodine-induced oxidation. This mini-PDX peptide both cyclic and acyclic form inhibited in vitro furin activity (IC50 in nM) when measured against either substrates Boc-RVRRdown double arrow MCA or QVEGF-C [Abz-QVHSIIRRdown double arrow SLP-Y(NO2)-A-CONH2, Abz=2-amino benzoic acid and Y(NO2)=3-nitro tyrosine], latter being derived from vascular endothelial growth factor-C (VEGF-C) processing site. The geometrically constrained structure mimicking PDX reactive loop is crucial for enzyme inhibition. Our study further revealed that both mini-PDX peptides inactivate furin in a slow tight binding manner, with disulfide-bridged cyclic form being slightly more potent. Unlike PDX, these peptides inhibit furin via a different mechanistic pathway. The study provides an alternate strategy for development of efficient peptide-based inhibitors of Proprotein Convertases including furin.     

Relationship of stimulated whole saliva cortisol level with the severity of a feeling of dry mouth in menopausal women

doi: 10.1111/j.1741‐2358.2010.00403.x Relationship of stimulated whole saliva cortisol level with the severity of a feeling of dry mouth in menopausal women Objectives: To evaluate the relationship of stimulated whole saliva cortisol level with the severity of a feeling of dry mouth (DM) in menopausal women. Background: A feel of DM is a major complaint for many elderly individuals and strongly associated with the menopause. The exact mechanisms that mediate sensation of DM in menopausal women have not been firmly established. Methods: A case–control study was carried out on 104 selected menopausal women with/without a feeling of DM, conducted at the Clinic of Oral Medicine, Tehran University of Medical Sciences. Xerostomia Inventory (XI) score was used as an index of DM severity. Stimulated whole saliva cortisol concentration (stimulated by chewing standard‐sized paraffin for 60 s) was measured by ELISA. Statistical analysis by Student’s t‐test and Spearman correlation was used. Results: The mean cortisol concentration of saliva, but not saliva cortisol output, was significantly higher in the cases than in the controls. There was significant positive correlation between XI score and concentration (r = 0.357, p = 0.000) or output (r = 0.223, p = 0.017) of stimulated whole saliva cortisol. Conclusions: It appears that stimulated whole saliva cortisol is high in menopausal women with a feeling of DM.     

Bio-effects and safety of low-intensity, low-frequency ultrasonic exposure

Low-frequency (LF) ultrasound (20-100 kHz) has a diverse set of industrial and medical applications. In fact, high power industrial applications of ultrasound mainly occupy this frequency range. This range is also used for various therapeutic medical applications including sonophoresis (ultrasonic transdermal drug delivery), dentistry, eye surgery, body contouring, the breaking of kidney stones and eliminating blood clots. While emerging LF applications such as ultrasonic drug delivery continue to be developed and undergo translation for human use, significant gaps exist in the coverage of safety standards for this frequency range. Accordingly, the need to understand the biological effects of LF ultrasound is becoming more important. This paper presents a broad overview of bio-effects and safety of LF ultrasound as an aid to minimize and control the risk of these effects. Its particular focus is at low intensities where bio-effects are initially observed. To generate a clear perspective of hazards in LF exposure, the mechanisms of bio-effects and the main differences in action at low and high frequencies are investigated and a survey of harmful effects of LF ultrasound at low intensities is presented. Mechanical and thermal indices are widely used in high frequency diagnostic applications as a means of indicating safety of ultrasonic exposure. The direct application of these indices at low frequencies needs careful investigation. In this work, using numerical simulations based on the mathematical and physical rationale behind the indices at high frequencies, it is observed that while thermal index (TI) can be used directly in the LF range, mechanical index (MI) seems to become less reliable at lower frequencies. Accordingly, an improved formulation for the MI is proposed for frequencies below 500 kHz.     

Combined triggering of dendritic cell receptors results in synergistic activation and potent cytotoxic immunity

Elimination of malignant cells and intracellular infections involves collaboration between CTLs and Th1 inflammation. Dendritic cells drive this response via costimulation and cytokines. We have defined key signals required for the exponential expansion of specific CD8(+) T cells in vivo in mice. Immunization with two or more TLR agonists, anti-CD40, IFN-gamma, and surfactant were sufficient to drive unprecedented levels of CD8 response to peptide or protein Ag and highly polarized Th1 CD4 responses. CD40 signaling was required for CD8 expansion but could be provided by a concomitant CD4 Th response in place of anti-CD40. Triggering of these pathways activated migration and activation of myeloid and plasmacytoid dendritic cells and secretion of IL-12. Cross-presentation can thus be exploited to induce potent cytotoxic responses and long-term memory to peptide/protein Ags. When combined with a tumor-associated peptide from tyrosinase-related protein 2, our combined adjuvant approach effectively halted tumor growth in an in vivo melanoma model and was more effective than anti-CD40 and a single TLR agonist. Antitumor immunity was associated with long-lived effector memory CD8 cells specific for the naturally processed and presented tumor Ag, and tumor protection was partially but not entirely dependent on CD8 T cells. This flexible strategy is more effective than existing adjuvants and provides a technological platform for rapid vaccine development.     

WFDC1/ps20 is a novel innate immunomodulatory signature protein of human immunodeficiency virus (HIV)-permissive CD4+ CD45RO+ memory T cells that promotes infection by upregulating CD54 integrin expression and is elevated in HIV type 1 infection

Understanding why human immunodeficiency virus (HIV) preferentially infects some CD4⁺ CD45RO⁺ memory T cells has implications for antiviral immunity and pathogenesis. We report that differential expression of a novel secreted factor, ps20, previously implicated in tissue remodeling, may underlie why some CD4 T cells are preferentially targeted. We show that (i) there is a significant positive correlation between endogenous ps20 mRNA in diverse CD4 T-cell populations and in vitro infection, (ii) a ps20⁺ permissive cell can be made less permissive by antibody blockade- or small-interference RNA-mediated knockdown of endogenous ps20, and (iii) conversely, a ps20^low cell can be more permissive by adding ps20 exogenously or engineering stable ps20 expression by retroviral transduction. ps20 expression is normally detectable in CD4 T cells after in vitro activation and interleukin-2 expansion, and such oligoclonal populations comprise ps20^positive and ps20^low/negative isogenic clones at an early differentiation stage (CD45RO⁺/CD25⁺/CD28⁺/CD57⁻). This pattern is altered in chronic HIV infection, where ex vivo CD4⁺ CD45RO⁺ T cells express elevated ps20. ps20 promoted HIV entry via fusion and augmented CD54 integrin expression; both of these effects were reversed by anti-ps20 antibody. We therefore propose ps20 to be a novel signature of HIV-permissive CD4 T cells that promotes infection in an autocrine and paracrine manner and that HIV has coopted a fundamental role of ps20 in promoting cell adhesion for its benefit. Disrupting the ps20 pathway may therefore provide a novel anti-HIV strategy.     

Optimization of Influencing Parameters on Phenanthrene Removal Efficiency in Soil Washing Process by Using Response Surface Methodology

Response surface methodology (RSM) under Box–Behnken design (BBD) was applied to evaluate the effect of the influencing parameters including surfactant concentration, liquid/soil ratio, Humic Acid concentration, and washing time on phenanthrene removal efficiency in soil washing process by using the nonionic surfactant Tween 80 and find an optimal operational conditions to achieve the highest removal efficiency. A polynomial quadratic model was used to correlate phenanthrene removal efficiency and four independent variables (R² = 0.9719). Based on the obtained results the most influential parameter on phenanthrene removal efficiency was surfactant concentration with an impact value of 69.519%. Liquid/soil ratio was also another factor that significantly influenced on removal efficiency with an impact value of 25.014%. The interaction between surfactant concentration and liquid/soil ratio was also shown to have a positive significant effect on removal efficiency (p_value = 0.0027). However, the other independent variables Humic Acid concentration and time were not significant in the ranges selected in this study. Based on the optimization results maximum removal efficiency of 70.692 ± 3.647% was achieved under the conditions of surfactant concentration 5000 mg L^?1, liquid/soil ratio 30 v/w, HA concentration 9.88 mg L^?1, and washing time 2 h, which was in good agreement with predicted value (66.643%).