Eugenol: A Promising Building Block for Synthesis of Radically Polymerizable Monomers

Eugenol, a natural phenol currently mainly obtained from clove oil, is an interesting aromatic building block for the synthesis of novel biobased monomers. It can also be obtained from lignin depolymerization, becoming a promising building block due to lignin availability as biomass feedstock. The synthesis of eight monomers derived from eugenol containing polymerizable functional groups is achieved. The (meth)acrylation of eugenol, isoeugenol, and dihydroeugenol is performed and the solution homopolymerization of these biobased monomers is studied. Moreover, aiming to prepare functional polymers, the introduction of epoxy and cyclic carbonate groups is executed via modification of the allylic double bond present in eugenol derived methacrylate. Thus, a novel platform of versatile biobased monomers derived from eugenol is presented, opening the opportunity to use them in a wide range of polymerization processes and applications

Photoinduced polymerization of eugenol-derived methacrylates

Biobased monomers have been used to replace their petroleum counterparts in the synthesis of polymers that are aimed at different applications. However, environmentally friendly polymerization processes are also essential to guarantee greener materials. Thus, photoinduced polymerization, which is low-energy consuming and solvent-free, rises as a suitable option. In this work, eugenol-, isoeugenol-, and dihydroeugenol-derived methacrylates are employed in radical photopolymerization to produce biobased polymers. The polymerization is monitored in the absence and presence of a photoinitiator and under air or protected from air, using Real-Time Fourier Transform Infrared Spectroscopy. The polymerization rate of the methacrylate double bonds was affected by the presence and reactivity of the allyl and propenyl groups in the eugenol- and isoeugenol-derived methacrylates, respectively. These groups are involved in radical addition, degradative chain transfer, and termination reactions, yielding crosslinked polymers. The materials, in the form of films, are characterized by differential scanning calorimetry, thermogravimetric, and contact angle analyses

Molecular basis for bacterial peptidoglycan recognition by LysM domains.

Carbohydrate recognition is essential for growth, cell adhesion and signalling in all living organisms. A highly conserved carbohydrate binding module, LysM, is found in proteins from viruses, bacteria, fungi, plants and mammals. LysM modules recognize polysaccharides containing N-acetylglucosamine (GlcNAc) residues including peptidoglycan, an essential component of the bacterial cell wall. However, the molecular mechanism underpinning LysM-peptidoglycan interactions remains unclear. Here we describe the molecular basis for peptidoglycan recognition by a multimodular LysM domain from AtlA, an autolysin involved in cell division in the opportunistic bacterial pathogen Enterococcus faecalis. We explore the contribution of individual modules to the binding, identify the peptidoglycan motif recognized, determine the structures of free and bound modules and reveal the residues involved in binding. Our results suggest that peptide stems modulate LysM binding to peptidoglycan. Using these results, we reveal how the LysM module recognizes the GlcNAc-X-GlcNAc motif present in polysaccharides across kingdoms

Human anti-60 kD heat shock protein autoantibodies are characterized by basic features of natural autoantibodies

Anti-human Hsp60 autoantibodies--known risk factor of atherosclerosis--were investigated in a mouse model and in samples of healthy subjects: polyreactivity, presence in cord blood samples of healthy newborns and life-long stability were tested. In IgM hybridoma panel from mouse spleens, polyreactivity of anti-Hsp60 autoantibodies was studied. In healthy pregnant women, umbilical vein and maternal blood samples were collected after childbirth, anti-Hsp-60 and -65 IgM and IgG levels were measured. Life-long stability of anti-Hsp-60 levels was studied on healthy patients during 5 years. ELISA was used in all studies. Polyreactivity of IgM clones of newborn mice and lifelong stability of these autoantibodies in healthy adults were established. IgM anti-Hsp60 autoantibodies in cord blood of healthy human infants were present, however, there was no correlation between maternal and cord blood IgM anti-Hsp60 concentrations. It is proposed that presence of anti-Hsp60 autoantibodies--as part of the natural autoantibody repertoire--may be an inherited trait. Level of anti-Hsp60 autoantibodies may be an independent, innate risk factor of atherosclerosis for the adulthood

Regulatory T cell epitopes (Tregitopes) in IgG induce tolerance in vivo and lack immunogenicity per se

Tregitopes are a set of epitopes, derived from IgG, that bind to MHCII, activate nTregs, and promote tolerance. We have now confirmed that coadministration of Tregitopes with a range of proteins (autoantigens and nominal antigens, such as OVA) in vitro and in vivo leads to suppression of T cell and antibody responses to the test antigens. In this study, we demonstrate that Tregitopes are not immunogenic in vivo even when emulsified with strong adjuvants, such as IFA or CFA. Moreover, in vivo administration of Tregitopes with IFA or CFA does not induce Th1 or Th2 cytokine expression under restimulation conditions in vitro. We investigated tolerance induction by codelivering Tregitopes with OVA using B cells. When B cells were pulsed with OVA plus Tregitopes and transferred into naïve mice, we found that cellular and humoral immune responses to the OVA were suppressed. As a result of their ability to induce Tregs and the absence of immunogenicity in the context of strong adjuvants, Tregitopes might be considered a novel immunomodulatory approach for the suppression of immune responses to protein therapeutics (such as FVIII and mAb), as well as for treatment of autoimmune diseases

Network Theory Analysis of Antibody-Antigen Reactivity Data: The Immune Trees at Birth and Adulthood

Motivation: New antigen microarray technology enables parallel recording of antibody reactivities with hundreds of antigens. Such data affords system level analysis of the immune system’s organization using methods and approaches from network theory. Here we measured the reactivity of 290 antigens (for both the IgG and IgM isotypes) of 10 healthy mothers and their term newborns. We constructed antigen correlation networks (or immune networks) whose nodes are the antigens and the edges are the antigen-antigen reactivity correlations, and we also computed their corresponding minimum spanning trees (MST) – maximal information reduced sub-graphs. We quantify the network organization (topology) in terms of the network theory divergence rate measure and rank the antigen importance in the full antigen correlation networks by the eigen-value centrality measure. This analysis makes possible the characterization and comparison of the IgG and IgM immune networks at birth (newborns) and adulthood (mothers) in terms of topology and node importance. Results: Comparison of the immune network topology at birth and adulthood revealed partial conservation of the IgG immune network topology, and significant reorganization of the IgM immune networks. Inspection of the antigen importance revealed some dominant (in terms of high centrality) antigens in the IgG and IgM networks at birth, which retain their importance at adulthood

Functional immunomics: Microarray analysis of IgG autoantibody repertoires predicts the future response of NOD mice to an inducer of accelerated diabetes

One's present repertoire of antibodies encodes the history of one's past immunological experience. Can the present autoantibody repertoire be consulted to predict resistance or susceptibility to the future development of an autoimmune disease? Here we developed an antigen microarray chip and used bioinformatic analysis to study a model of type 1 diabetes developing in non-obese diabetic (NOD) male mice in which the disease was accelerated and synchronized by exposing the mice to cyclophosphamide at 4 weeks of age. We obtained sera from 19 individual mice, treated the mice to induce cyclophosphamide-accelerated diabetes (CAD), and found, as expected, that 9 mice became severely diabetic while 10 mice permanently resisted diabetes. We again obtained serum from each mouse afterCAD induction. We then analyzed the patterns of antibodies in the individualmice to 266 different antigens spotted on the antigen chip. We identified a select panel of 27 different antigens (10% of the array) that revealed a pattern of IgG antibody reactivity in the pre-CAD serathat discriminated between the mice resistant or susceptible to CAD with 100% sensitivity and 82% specificity (p=0.017). Surprisingly, the set of IgG antibodies that was informative before CAD induction did not separate the resistant and susceptible groups after the onset of CAD; new antigens became criticalfor post-CAD repertoire discrimination. Thus, at least for a model disease, present antibody repertoires can predict future disease; predictive and diagnostic repertoires can differ; and decisive information about immune system behavior can be mined by bioinformatic technology. Repertoires matter.Comment: See Advanced Publication on the PNAS website for final versio

Role of the factor VIII-binding capacity of endogenous von Willebrand factor on the development of factor VIII inhibitors in patients with severe hemophilia A

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Effect of HLA DR epitope de-immunization of Factor VIII \u3ci\u3ein vitro\u3c/i\u3e and \u3ci\u3ein vivo\u3c/i\u3e

T cell-dependent development of anti-Factor VIII (FVIII) antibodies that neutralize FVIII activity is a major obstacle to replacement therapy in hemophilia A. To create a less immunogenic therapeutic protein, recombinant FVIII can be modified to reduce HLA binding of epitopes based on predicted anchoring residues. Here, we used immunoinformatic tools to identify C2 domain HLA DR epitopes and predict site-specific mutations that reduce immunogenicity. Epitope peptides corresponding to original and modified sequences were validated in HLA binding assays and in immunizations of hemophilic E16 mice, DR3 and DR4 mice and DR3 × E16 mice. Consistent with immunoinformatic predictions, original epitopes are immunogenic. Immunization with selected modified sequences lowered immunogenicity for particular peptides and revealed residual immunogenicity of incompletely de-immunized modified peptides. The stepwise approach to reduce protein immunogenicity by epitope modification illustrated here is being used to design and produce a functional full-length modified FVIII for clinical use