Themis controls T cell activation, effector functions, and metabolism of peripheral CD8 T cells.

Themis is important in regulating positive selection of thymocytes during T cell development, but its role in peripheral T cells is less understood. Here, we investigated T cell activation and its sequelae using a tamoxifen-mediated, acute Themis deletion mouse model. We find that proliferation, effector functions including anti-tumor killing, and up-regulation of energy metabolism are severely compromised.

Platelet TLR7 is essential for the formation of platelet-neutrophil complexes and low-density neutrophils in lupus nephritis.

Objectives: Platelets and low-density neutrophils (LDNs) are major players in the immunopathogenesis of SLE. Despite evidence showing the importance of platelet-neutrophil complexes (PNCs) in inflammation, little is known about the relationship between LDNs and platelets in SLE. We sought to characterize the role of LDNs and Toll-like receptor 7 (TLR7) in clinical disease.

Analytical technology development to monitor the stability of Polysaccharide-Protein conjugate vaccines.

The covalent attachment of a bacterial-derived capsular polysaccharide to protein is of critical importance in transforming the polysaccharide from an antigen with limited immunogenicity in infants and older adults to an antigen that can prevent potentially fatal disease. For a polysaccharide-protein conjugate vaccine (PCV) candidate to be successful, it must be sufficiently stable. Chemical breakage of carbohydrate bonds in the polysaccharide may result in the reduction of "conjugate dose" and could negatively impact immunogenicity and the ability of the vaccine to prime for memory responses.

Bayesian analysis of cytokines and chemokine identifies immune pathways of HBsAg loss during chronic hepatitis B treatment.

Our objective was to examine differences in cytokine/chemokine response in chronic hepatitis B(CHB) patients to understand the immune mechanism of HBsAg loss (functional cure) during antiviral therapy. We used an unbiased machine learning strategy to unravel the immune pathways in CHB nucleo(t)side analogue-treated patients who achieved HBsAg loss with peg-interferon-α(peg-IFN-α) add-on or switch treatment in a randomised clinical trial. Cytokines/chemokines from plasma were compared between those with/without HBsAg loss, at baseline, before and after HBsAg loss.

Immunogenicity generated by mRNA vaccine encoding VZV gE antigen is comparable to adjuvanted subunit vaccine and better than live attenuated vaccine in nonhuman primates.

Shingles is a painful, blistering rash caused by reactivation of latent varicella-zoster virus (VZV) and most frequently occurs in elderly and immunocompromised individuals. Currently, two approved vaccines for the prevention of shingles are on the market, a live attenuated virus vaccine ZOSTAVAX® (Merck & Co., Inc.

FTIR Spectroscopy Detects Intermolecular β-Sheet Formation Above the High Temperature T for Two Monoclonal Antibodies.

The temperature-dependent secondary structure of two monoclonal IgG antibodies, anti-IGF1R and anti-TSLP, were examined by transmission mode Fourier Transform Infrared (FTIR) spectroscopy. Anti-IGF1R and anti-TSLP are IgG monoclonal antibodies (mAbs) directed against human Insulin-like Growth Factor 1 Receptor for anti-tumor activity and Thymic Stromal Lymphopoietin cytokine for anti-asthma activity, respectively. Differential scanning calorimetry (DSC) clearly indicates both antibodies in their base formulations have a lower temperature protein conformational change near 70 °C (T) and a higher temperature protein conformational change near 85 °C (T).

Met-Flow, a strategy for single-cell metabolic analysis highlights dynamic changes in immune subpopulations.

A complex interaction of anabolic and catabolic metabolism underpins the ability of leukocytes to mount an immune response. Their capacity to respond to changing environments by metabolic reprogramming is crucial to effector function. However, current methods lack the ability to interrogate this network of metabolic pathways at single-cell level within a heterogeneous population.

High Throughput Differential Scanning Fluorimetry (DSF) Formulation Screening with Complementary Dyes to Assess Protein Unfolding and Aggregation in Presence of Surfactants.

Purpose: The purpose was to evaluate DSF for high throughput screening of protein thermal stability (unfolding/ aggregation) across a wide range of formulations. Particular focus was exploring PROTEOSTAT® - a commercially available fluorescent rotor dye - for detection of aggregation in surfactant containing formulations. Commonly used hydrophobic dyes (e.

Quantitative Analysis of Vaccine Antigen Adsorption to Aluminum Adjuvant Using an Automated High-Throughput Method.

Aluminum-containing adjuvants have been widely used in vaccine formulations to safely and effectively potentiate the immune response. The examination of the extent of antigen adsorption to aluminum adjuvant is always evaluated during the development of aluminum adjuvant containing vaccines. A rapid, automated, high-throughput assay was developed to measure antigen adsorption in a 96-well plate format using a TECAN Freedom EVO (TECAN).

Germline NLRP1 Mutations Cause Skin Inflammatory and Cancer Susceptibility Syndromes via Inflammasome Activation.

Inflammasome complexes function as key innate immune effectors that trigger inflammation in response to pathogen- and danger-associated signals. Here, we report that germline mutations in the inflammasome sensor NLRP1 cause two overlapping skin disorders: multiple self-healing palmoplantar carcinoma (MSPC) and familial keratosis lichenoides chronica (FKLC). We find that NLRP1 is the most prominent inflammasome sensor in human skin, and all pathogenic NLRP1 mutations are gain-of-function alleles that predispose to inflammasome activation.

Improving the immunogenicity of a trivalent Neisseria meningitidis native outer membrane vesicle vaccine by genetic modification.

Trivalent native outer membrane vesicles (nOMVs) derived from three genetically modified Neisseria meningitidis serogroup B strains have been previously evaluated immunologically in mice and rabbits. This nOMV vaccine elicited serum bactericidal activity (SBA) against multiple N. meningitidis serogroup B strains as well as strains from serogroups C, Y, W, and X.

Metabolism Is Central to Tolerogenic Dendritic Cell Function.

Immunological tolerance is a fundamental tenant of immune homeostasis and overall health. Self-tolerance is a critical component of the immune system that allows for the recognition of self, resulting in hyporeactivity instead of immunogenicity. Dendritic cells are central to the establishment of dominant immune tolerance through the secretion of immunosuppressive cytokines and regulatory polarization of T cells.

Efficacy of a nanocochleate-encapsulated 3,5-diaryl-s-triazole derivative in a murine model of graft-versus-host disease.

We have previously demonstrated that the compound 3-(2-ethylphenyl)-5-(3-methoxyphenyl)-1H-1,2,4-triazole exerts immunosuppressive effects in several experimental models of autoimmunity. These results were achieved by subcutaneously administering ST1959 after dissolution in an oily vehicle, because of its poor water solubility. To circumvent this problem, we sought to determine whether nanocochleate technology could be successfully exploited to deliver ST1959 and protect mice undergoing lethal acute graft-versus-host disease (GVHD).

Effect of alternative aluminum adjuvants on the absorption and immunogenicity of HPV16 L1 VLPs in mice.

Aluminum adjuvants are commonly used in prophylactic vaccines to enhance antigen immunogenicity through induction of high-titer antibody responses. Three major forms of aluminum adjuvants with substantially different physical and chemical properties have been described: aluminum phosphate (AlPO(4)), aluminum hydroxide (AlOH) and amorphous aluminum hydroxyphosphate sulfate (AAHS). Here we describe the effect of these different aluminum adjuvants on the formulation and subsequent immunogenicity in mice of virus-like particles (VLPs) consisting of the L1 protein of Human Papillomavirus (HPV) Type 16.

Interdigitation--fusion liposomes.

IF-liposomes are formed by a unique process that involves fusing small liposomes into interdigitated lipid sheets, using either ethanol or hydrostatic pressure. The interdigitation-fusion method requires liposome formulations with lipids that form the L beta I phase. Preparing ethanol-induced IF-liposomes is simple and quick.

Association of hydrophobically-modified poly(ethylene glycol) with fusogenic liposomes.

We present results on using cooperative interactions to shield liposomes by incorporating multiple hydrophobic anchoring sites on polyethylene glycol (PEG) polymers. The hydrophobically-modified PEGs (HMPEGs) are comb-graft polymers with strictly alternating monodisperse PEG blocks (M(w)=6, 12, or 35 kDa) bonded to C18 stearylamide hydrophobes. Cooperativity is varied by changing the degree of oligomerization at a constant ratio of PEG to stearylamide.

A novel N-acyl phosphatidylethanolamine-containing delivery vehicle for spermine-condensed plasmid DNA.

A unique method for formulation of plasmid DNA with phospholipids has been devised for the purpose of producing vehicles that can mediate gene delivery and transfection of living cells. The polycation, spermine, was used to condense plasmid DNA within a water-in-chloroform emulsion stabilized by phospholipids. After organic solvent removal, the particles formed could be extruded to a number average size of about 200 nm and retained DNA that was protected from nuclease digestion.

Elastase activated liposomal delivery to nucleated cells.

The specific activation of liposomes for delivery has been explored by enzyme mediated cleavage of a peptide substrate covalently conjugated to a fusogenic lipid. We have previously shown an elastase sensitive peptide conjugated to 1, 2-dioleoyl-sn-glycero-3-phosphoethanolamine [corrected] (DOPE) could be activated by enzymatic cleavage, triggering liposome-liposome lipid mixing and fusion with erythrocyte ghosts (Pak et al., Biochim.

Enhancement of the in vivo circulation lifetime of L-alpha-distearoylphosphatidylcholine liposomes: importance of liposomal aggregation versus complement opsonization.

Incorporation of N-(omega-carboxy)acylamido-phosphatidylethanolamines (-PEs) into large unilamellar vesicles (LUVs) of L-alpha-distearoylphosphatidylcholine (DSPC) was found to dramatically increase the in vivo liposomal circulation lifetime in rats, reaching a maximal effect at 10 mol.% of the total phospholipid. Neither pure DSPC liposomes nor those with the longest circulating derivative, N-glutaryl-dipalmitoylphosphatidylethanolamine (-DPPE), were found to significantly bind complement from serum.

Association and release of prostaglandin E1 from liposomes.

PGE1-lipid interactions were studied in several liposome systems. Data from both circular dichroic (CD) measurements and differential scanning calorimetry (DSC) indicated that PGE1 in the protonated form seeks the less polar environment of the lipid bilayer. CD measurements made on PGE1 in solution showed that the wavelength of maximum absorbance red shifted approximately 8 nm with decreasing solvent polarity.

Solute-induced shift of phase transition temperature in Di-saturated PC liposomes: adoption of ripple phase creates osmotic stress.

We have examined the calorimetric behavior of large liposomes consisting of symmetric saturated chain phosphatidylcholines. Most notably, for systems made in solutions containing solute (e.g.

Interdigitation-fusion: a new method for producing lipid vesicles of high internal volume.

Previously we demonstrated that fused phospholipid sheets can be formed from small unilamellar vesicles (SUVs) comprised of saturated symmetric chain lipids by exposing them to concentrations of ethanol sufficient to cause bilayer interdigitation (Boni et al. (1993) Biochim. Biophys.

The determination of liposome captured volume.

Manipulating the process by which lipids assemble to form bilayer membranes has produced a myriad of protocol-dependent liposome types. For each of these systems the arrangement of bilayers is characteristic and can be described by parameters such as aqueous entrapment per mole lipid or captured volume, vesicle size distribution, the average number of lamellae per vesicle and shape. For specific applications as model systems or drug delivery systems specific characteristics are desired.