Pharmacological potentiation of monocyte-derived dendritic cell cancer immunotherapy.

Dendritic cells have been at the forefront of cancer-immunotherapy research for over 2 decades. They elicited that attention by having an unprecedented capacity to mount T cells responses against tumors. However, the clinical use of DC-based vaccination against established malignancies has resulted in limited clinical benefits. Several factors are responsible for limiting the efficacy of DC-based immunotherapy, such as the harmful influence of the tumor microenvironment on DCs activity. New insights into the inner process of DC-mediated T cell activation have supported the development of new strategies that potentiate DCs-based therapies. Herein, we identify signaling cascades that have recently been targeted by small molecules and biologicals to promote the activation of monocyte-derived DCs and decrease their susceptibility to becoming tolerogenic. While Statins can markedly enhance antigen presentation, protein kinase inhibitors can be used to increase the expression of co-receptors and adhesion molecules. STAT3 and IDO can be modulated to limit the production of regulatory factors that work against differentiation and activation. The targeting of multiple pathways simultaneously has also been found to produce synergism and drastically enhance DCs activity. Some of these strategies have recently yielded positive results in clinical settings against established malignancies such as non-small cell lung cancer. The emergence of these approaches opens the door for a new generation of potent dendritic cell-based therapeutics to fight cancer.

Self-Assembly of Rhamnolipid Bioamphiphiles: Understanding the Structure-Property Relationship Using Small-Angle X-ray Scattering.

The structure-property relationship of rhamnolipids, RLs, well-known microbial bioamphiphiles (biosurfactants), is explored in detail by coupling cryogenic transmission electron microscopy (cryo-TEM) and both ex situ and in situ small-angle X-ray scattering (SAXS). The self-assembly of three RLs with reasoned variation of their molecular structure (RhaC10, RhaC10C10, and RhaRhaC10C10) and a rhamnose-free C10C10 fatty acid is studied in water as a function of pH. It is found that RhaC10 and RhaRhaC10C10 form micelles in a broad pH range and RhaC10C10 undergoes a micelle-to-vesicle transition from basic to acid pH occurring at pH 6.5. Modeling coupled to fitting SAXS data allows a good estimation of the hydrophobic core radius (or length), the hydrophilic shell thickness, the aggregation number, and the surface area per RL. The essentially micellar morphology found for RhaC10 and RhaRhaC10C10 and the micelle-to-vesicle transition found for RhaC10C10 are reasonably well explained by employing the packing parameter (PP) model, provided a good estimation of the surface area per RL. On the contrary, the PP model fails to explain the lamellar phase found for the protonated RhaRhaC10C10 at acidic pH. The lamellar phase can only be explained by values of the surface area per RL being counterintuitively small for a di-rhamnose group and folding of the C10C10 chain. These structural features are only possible for a change in the conformation of the di-rhamnose group between the alkaline and acidic pH.

In Situ Stimulation of Self-Assembly Tunes the Elastic Properties of Interpenetrated Biosurfactant-Biopolymer Hydrogels.

Hydrogels are widespread soft materials, which can be used in a wide range of applications. The control over the viscoelastic properties of the gel is of paramount importance. Ongoing environmental issues have raised the consumer's concern toward the use of more sustainable materials, including hydrogels. However, are greener materials compatible with high functionality? In a safe-by-design approach, this work demonstrates that functional hydrogels with in situ responsivity of their elastic properties by external stimuli can be developed from entirely "sustainable" components, a biobased amphiphile and biopolymers (gelatin, chitosan, and alginate). The bioamphiphile is a stimuli-responsive glycolipid obtained by microbial fermentation, which can self-assemble into fibers, but also micelles or vesicles, in water under high dilution and by a rapid variation of the stimuli. The elastic properties of the bioamphiphile-/biopolymer-interpenetrated hydrogels can be modulated by selectively triggering the phase transition of the glycolipid and/or the biopolymer inside the gel by mean of temperature or pH.

Interpenetrated Biosurfactant-Biopolymer Orthogonal Hydrogels: The Biosurfactant's Phase Controls the Hydrogel's Mechanics.

Controlling the viscoelastic properties of hydrogels is a challenge for many applications. Low molecular weight gelators (LMWGs) like bile salts and glycolipids and biopolymers like chitosan and alginate are good candidates for developing fully biobased hybrid hydrogels that combine the advantages of both components. Biopolymers lead to enhanced mechanics, while LMWGs add functionality. In this work, hybrid hydrogels are composed of biopolymers (gelatin, chitosan, and alginate) and microbial glycolipid bioamphiphiles, known as biosurfactants. Besides their biocompatibility and natural origin, bioamphiphiles can present chameleonic behavior, as pH and ions control their phase diagram in water around neutrality under strongly diluted conditions (<5 wt%). The glycolipid used in this work behaves like a surfactant (micellar phase) at high pH or like a phospholipid (vesicle phase) at low pH. Moreover, at neutral-to-alkaline pH in the presence of calcium, it behaves like a gelator (fiber phase). The impact of each of these phases on the elastic properties of biopolymers is explored by means of oscillatory rheology, while the hybrid structure is studied by small angle X-ray scattering. The micellar and vesicular phases reduce the elastic properties of the hydrogels, while the fiber phase has the opposite effect; it enhances the hydrogel's strength by forming an interpenetrated biopolymer-LMWG network.

Shear recovery and temperature stability of Ca2+ and Ag+ glycolipid fibrillar metallogels with unusual ß-sheet-like domains.

Low-molecular weight gelators (LMWGs) are small molecules (Mw < ∼1 kDa), which form self-assembled fibrillar network (SAFiN) hydrogels in water. A great majority of SAFiN gels are described by an entangled network of self-assembled fibers, in analogy to a polymer in a good solvent. Here, fibrillation of a biobased glycolipid bolaamphiphile is triggered by Ca2+ or Ag+ ions which are added to its diluted micellar phase. The resulting SAFiN, which forms a hydrogel above 0.5 wt%, has a "nano-fishnet" structure, characterized by a fibrous network of both entangled fibers and ß-sheet-like rafts, generally observed for silk fibroin, actin hydrogels or mineral imogolite nanotubes, but generally not known for SAFiN. This work focuses on the strength of the SAFIN gels, their fast recovery after applying a mechanical stimulus (strain) and their unusual resistance to temperature, studied by coupling rheology to small angle X-ray scattering (rheo-SAXS) using synchrotron radiation. The Ca2+-based hydrogel maintains its properties up to 55 °C, while the Ag+-based gel shows a constant elastic modulus up to 70 °C, without the appearance of any gel-to-sol transition temperature. Furthermore, the glycolipid is obtained by fermentation from natural resources (glucose and rapeseed oil), thus showing that naturally engineered compounds can have unprecedented properties, when compared to the wide range of chemically derived amphiphiles.

Ca2+ and Ag+ orient low-molecular weight amphiphile self-assembly into "nano-fishnet" fibrillar hydrogels with unusual ß-sheet-like raft domains.

Low-molecular weight gelators (LMWGs) are small molecules (Mw < ∼1 kDa), which form self-assembled fibrillar network (SAFiN) hydrogels in water when triggered by an external stimulus. A great majority of SAFiN gels involve an entangled network of self-assembled fibers, in analogy to a polymer in a good solvent. In some rare cases, a combination of attractive van der Waals and repulsive electrostatic forces drives the formation of bundles with a suprafibrillar hexagonal order. In this work, an unexpected micelle-to-fiber transition is triggered by Ca2+ or Ag+ ions added to a micellar solution of a novel glycolipid surfactant, whereas salt-induced fibrillation is not common for surfactants. The resulting SAFiN, which forms a hydrogel above 0.5 wt%, has a "nano-fishnet" structure, characterized by a fibrous network of both entangled fibers and ß-sheet-like rafts, generally observed for silk fibroin, actin hydrogels or mineral imogolite nanotubes, but not known for SAFiNs. The ß-sheet-like raft domains are characterized by a combination of cryo-TEM and SAXS and seem to contribute to the stability of glycolipid gels. Furthermore, glycolipid is obtained by fermentation from natural resources (glucose, rapeseed oil), thus showing that naturally engineered compounds can have unprecedented properties, when compared to the wide range of chemically derived amphiphiles.

TAOK3 limits age-associated inflammation by negatively modulating macrophage differentiation and their production of TNFα.

BACKGROUND: Human aging is characterized by a state of chronic inflammation, termed inflammaging, for which the causes are incompletely understood. It is known, however, that macrophages play a driving role in establishing inflammaging by promoting pro-inflammatory rather than anti-inflammatory responses. Numerous genetic and environmental risk factors have been implicated with inflammaging, most of which are directly linked to pro-inflammatory mediators IL-6, IL1Ra, and TNFα. Genes involved in the signaling and production of those molecules have also been highlighted as essential contributors. TAOK3 is a serine/threonine kinase of the STE-20 kinase family that has been associated with an increased risk of developing auto-immune conditions in several genome-wide association studies (GWAS). Yet, the functional role of TAOK3 in inflammation has remained unexplored. RESULTS: We found that mice deficient in the serine/Threonine kinase Taok3 developed severe inflammatory disorders with age, which was more pronounced in female animals. Further analyses revealed a drastic shift from lymphoid to myeloid cells in the spleens of those aged mice. This shift was accompanied by hematopoietic progenitor cells skewing in Taok3-/- mice that favored myeloid lineage commitment. Finally, we identified that the kinase activity of the enzyme plays a vital role in limiting the establishment of proinflammatory responses in macrophages. CONCLUSIONS: Essentially, Taok3 deficiency promotes the accumulation of monocytes in the periphery and their adoption of a pro-inflammatory phenotype. These findings illustrate the role of Taok3 in age-related inflammation and highlight the importance of genetic risk factors in this condition.

Sunflower Proteins at Air-Water and Oil-Water Interfaces.

The adsorption of a sunflower protein extract at two air-water and oil-water interfaces is investigated using tensiometry, dilational viscoelasticity, and ellipsometry. For both interfaces, a three step mechanism was evidenced thanks to master curve representations of the data taken at different aging times and protein concentrations. At short times, a diffusion limited adsorption of proteins at interfaces is demonstrated. First, a two-dimensional protein film is formed with a partition of the polypeptide chains in the two phases that depends strongly on the nature of the hydrophobic phase: most of the film is in the aqueous phase at the air-water interface, while it is mostly in the organic phase at the oil-water interface. Then a three-dimensional saturated monolayer of proteins is formed. At short times, adsorption mechanisms are analogous to those found with typical globular proteins, while strong divergences are observed at longer adsorption times. Following the saturation step, a thick layer expands in the aqueous phase and appears associated with the release of large objects in the bulk. The kinetic evolution of this second layer is compatible with a diffusion limited adsorption of the minor population of polymeric complexes with hydrodynamic radius RH ∼ 80 nm, evidenced in equilibrium with hexameric globulins (RH ∼ 6 nm) in solution. These complexes could result from the presence of residual polyphenols in the extract and raise the question of the role of these compounds in the interfacial properties of plant protein extracts.

PTPRS is a novel marker for early Tau pathology and synaptic integrity in Alzheimer's disease.

We examined the role of protein tyrosine phosphatase receptor sigma (PTPRS) in the context of Alzheimer's disease and synaptic integrity. Publicly available datasets (BRAINEAC, ROSMAP, ADC1) and a cohort of asymptomatic but "at risk" individuals (PREVENT-AD) were used to explore the relationship between PTPRS and various Alzheimer's disease biomarkers. We identified that PTPRS rs10415488 variant C shows features of neuroprotection against early Tau pathology and synaptic degeneration in Alzheimer's disease. This single nucleotide polymorphism correlated with higher PTPRS transcript abundance and lower p(181)Tau and GAP-43 levels in the CSF. In the brain, PTPRS protein abundance was significantly correlated with the quantity of two markers of synaptic integrity: SNAP25 and SYT-1. We also found the presence of sexual dimorphism for PTPRS, with higher CSF concentrations in males than females. Male carriers for variant C were found to have a 10-month delay in the onset of AD. We thus conclude that PTPRS acts as a neuroprotective receptor in Alzheimer's disease. Its protective effect is most important in males, in whom it postpones the age of onset of the disease.

PTPRS is a novel marker for early tau pathology and synaptic integrity in Alzheimer's disease.

We examined the role of protein tyrosine phosphatase receptor sigma (PTPRS) in the context of Alzheimer's disease and synaptic integrity. Publicly available datasets (BRAINEAC, ROSMAP, ADC1) and a cohort of asymptomatic but "at risk" individuals (PREVENT-AD) were used to explore the relationship between PTPRS and various Alzheimer's disease biomarkers. We identified that PTPRS rs10415488 variant C shows features of neuroprotection against early tau pathology and synaptic degeneration in Alzheimer's disease. This single nucleotide polymorphism correlated with higher PTPRS transcript abundance and lower P-tau181 and GAP-43 levels in the CSF. In the brain, PTPRS protein abundance was significantly correlated with the quantity of two markers of synaptic integrity: SNAP25 and SYT-1. We also found the presence of sexual dimorphism for PTPRS, with higher CSF concentrations in males than females. Male carriers for variant C were found to have a 10-month delay in the onset of AD. We thus conclude that PTPRS acts as a neuroprotective receptor in Alzheimer's disease. Its protective effect is most important in males, in whom it postpones the age of onset of the disease.

The induction of SHP-1 degradation by TAOK3 ensures the responsiveness of T cells to TCR stimulation.

Thousand-and-one-amino acid kinase 3 (TAOK3) is a serine and threonine kinase that belongs to the STE-20 family of kinases. Its absence reduces T cell receptor (TCR) signaling and increases the interaction of the tyrosine phosphatase SHP-1, a major negative regulator of proximal TCR signaling, with the kinase LCK, a component of the core TCR signaling complex. Here, we used mouse models and human cell lines to investigate the mechanism by which TAOK3 limits the interaction of SHP-1 with LCK. The loss of TAOK3 decreased the survival of naïve CD4+ T cells by dampening the transmission of tonic and ligand-dependent TCR signaling. In mouse T cells, Taok3 promoted the secretion of interleukin-2 (IL-2) in response to TCR activation in a manner that depended on Taok3 gene dosage and on Taok3 kinase activity. TCR desensitization in Taok3-/- T cells was caused by an increased abundance of Shp-1, and pharmacological inhibition of Shp-1 rescued the activation potential of these T cells. TAOK3 phosphorylated threonine-394 in the phosphatase domain of SHP-1, which promoted its ubiquitylation and proteasomal degradation. The loss of TAOK3 had no effect on the abundance of SHP-2, which lacks a residue corresponding to SHP-1 threonine-394. Modulation of SHP-1 abundance by TAOK3 thus serves as a rheostat for TCR signaling and determines the activation threshold of T lymphocytes.

Chameleonic amphiphile: The unique multiple self-assembly properties of a natural glycolipid in excess of water.

Chameleons are stunning reptiles which change colour according to the surrounding environment. In astrophysics, chameleons are particles whose mass varies in the surrounding matter. Here, we show the chameleonic self-assembly behavior of a low molecular weight (LMW) amphiphile, a broad class of molecules widely studied for several decades. Their ability to self-assemble in water make them both fascinating and useful compounds for a number of applications. Under thermodynamic conditions, their thermotropic and lyotropic phase behavior is generally predicted in relation to their molecular shape, as seen for classical head-tail molecules like surfactants or phospholipids. However, many exceptions do exist, either when amphiphiles have unconventional shapes, e.g., bolaform or gemini, or when they contain functional groups which undergo specific interactions such as H-bonding or π-π stacking. In excess water, surfactants form micelles, phospholipids form vesicles or lamellar phases, and functional amphiphiles often form micelles or fibers. Here, we show the multiphase behavior, much richer and more unpredictable than what it is known for most amphiphiles, of a biobased glycolipid produced by the yeast S. bombicola ΔugtB1. In excess water and within a narrow pH range around neutrality, this compound assembles into micelles, uni- and multilamellar vesicles, lamellae and fibers, simply as a function of changing pH, temperature and counterions. This rich phase behavior is not only interesting in itself, it also generates a number of diverse biocompatible and biodegradable soft self-assembled materials like hydrogels, complex coacervates and drug carriers.

Cation-Induced Fibrillation of Microbial Glycolipid Biosurfactant Probed by Ion-Resolved In Situ SAXS.

Biological amphiphiles are molecules with a rich phase behavior. Micellar, vesicular, and even fibrillar phases can be found for the same molecule by applying a change in pH or by selecting the appropriate metal ion. The rich phase behavior paves the way toward a broad class of soft materials, from carriers to hydrogels. The present work contributes to understanding the fibrillation of a microbial glycolipid, glucolipid G-C18:1, produced by Starmerella bombicola ΔugtB1 and characterized by a micellar phase at alkaline pH and a vesicular phase at acidic pH. Fibrillation and prompt hydrogelation is triggered by adding either alkaline earth, Ca2+, or transition metal, Ag+, Fe2+, Al3+, ions to a G-C18:1 micellar solution. A specifically designed apparatus coupled to a synchrotron SAXS beamline allows the performing of simultaneous cation- and pH-resolved in situ monitoring of the morphological evolution from spheroidal micelles to crystalline fibers, when Ca2+ is employed, or to wormlike aggregates, when Fe2+ or Al3+ solutions are employed. The fast reactivity of Ag+ and the crystallinity of Ca2+-induced fibers suggest that fibrillation is driven by direct metal-ligand interactions, while the shape transition from spheroidal to elongated micelles with Fe2+ or Al3+ rather suggest charge screening between the lipid and the hydroxylated cation species.

pH-switchable pickering emulsions stabilized by polyelectrolyte-biosurfactant complex coacervate colloids.

HYPOTHESIS: Polyelectrolyte-surfactant complexes (PESCs) have long been employed as oil-in-water (o/w) emulsions stabilizers, but never in the structure of colloidal complex coacervates providing a Pickering effect. The complexed state of PESCs could make them unsuitable o/w Pickering emulsifiers, which instead require a balance between colloidal structure and stability, amphiphilicity and wettability. Here we hypothesize that PESCs coacervates are efficient Pickering stabilizers. Instead of classical surfactants, we employ sophorolipid (SL) biosurfactants, atypical anionic/neutral stimuli-responsive biosurfactants. Despite their tunable charge and mild amphiphilic character, they can be used in combination with cationic/neutral polyelectrolytes (chitosan, CHL, or poly-l-lysine, PLL) to form PESC coacervates for the development of biobased, but also pH-switchable, Pickering emulsions. EXPERIMENTS: Aqueous solutions of SL-CHL (or SL-PLL) complex coacervates are emulsified with dodecane. Confocal laser scanning microscopy (CLSM) and scanning electron microscopy under cryogenic conditions (cryo-SEM) demonstrate the Pickering effect, while optical microscopy and oscillatory rheology respectively assess the emulsion formation and relative viscoelastic properties. FINDINGS: Both SL-CHL and SL-PLL PESCs stabilize o/w emulsions up to Φoil of 0.7 only in the pH region of complex coacervation (6 < pH < 9): outside this range, phase separation occurs. Rheology shows a typical solid-like response and mechanical recovery upon applying large deformations. CLSM and cryo-SEM highlight a colloidal structure, associated to the complex coacervates, of the oil/water interface and suggest a Pickering effect. These findings demonstrate the Pickering effect from PESC coacervates and the possibility to use biobased and biocompatible components, with application potential in cosmetics, food science, or oil recovery.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) in Alzheimer's disease: A genetic and proteomic multi-cohort study.

BACKGROUND: Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a hepatic enzyme that regulates circulating low-density lipoprotein (LDL) cholesterol levels by binding to LDL receptors (LDLR) and promoting their degradation. Although PCSK9 inhibitors were shown to reduce the risk of cardiovascular disease, a warning was issued concerning their possible impact on cognitive functions. In Alzheimer's disease (AD), it is believed that cognitive impairment is associated with cholesterol metabolism alterations, which could involve PCSK9. The main objective of this study is to determine if PCSK9 plays a significant role in the pre-symptomatic phase of the disease when the pathophysiological markers of AD unfolds and, later, when cognitive symptoms emerge. METHODS AND FINDINGS: To test if PCSK9 is associated with AD pathology, we measured its expression levels in 65 autopsy confirmed AD brains and 45 age and gender matched controls. Messenger ribonucleic acid (mRNA) were quantified using real-time polymerase chain reaction (RT-PCR) and protein levels were quantified using enzyme-linked immunosorbent assay (ELISA). PCSK9 was elevated in frontal cortices of AD subjects compared to controls, both at the mRNA and protein levels. LDLR protein levels were unchanged in AD frontal cortices, despite and upregulation at the mRNA level. To verify if PCSK9 dysregulation was observable before the onset of AD, we measured its expression in the cerebrospinal fluid (CSF) of 104 "at-risk" subjects and contrasted it with known apolipoproteins levels and specific AD biomarkers using ELISAs. Positive correlations were found between CSF PCSK9 and apolipoprotein E (APOE), apolipoprotein J (APOJ or CLU), apolipoprotein B (APOB), phospho Tau (pTau) and total Tau. To investigate if PCSK9 levels were driven by genetic variants, we conducted an expression quantitative trait loci (eQTL) study using bioinformatic tools and found two polymorphisms in strong association. Further investigation of these variants in two independent cohorts showed a female specific association with AD risk and with CSF Tau levels in cognitively impaired individuals. CONCLUSIONS: PCSK9 levels differ between control and AD brains and its protein levels correlate with those of other lipoproteins and AD biomarkers even before the onset of the disease. PCSK9 regulation seems to be under tight genetic control in females only, with specific variants that could predispose to increased AD risk.

Multistep building of a soft plant protein film at the air-water interface.

Gliadins are edible wheat storage proteins well known for their surface active properties. In this paper, we present experimental results on the interfacial properties of acidic solutions of gliadin studied over 5 decades of concentrations, from 0.001 to 110 g/L. Dynamic pendant drop tensiometry reveals that the surface pressure Π of gliadin solutions builds up in a multistep process. The series of curves of the time evolution of Π collected at different bulk protein concentrations C can be merged onto a single master curve when Π is plotted as a function of αt where t is the time elapsed since the formation of the air/water interface and α is a shift parameter that varies with C as a power law with an exponent 2. The existence of such time-concentration superposition, which we evidence for the first time, indicates that the same mechanisms govern the surface tension evolution at all concentrations and are accelerated by an increase of the bulk concentration. The scaling of α with C is consistent with a kinetic of adsorption controlled by the diffusion of the proteins in the bulk. Moreover, we show that the proteins adsorption at the air/water interface is kinetically irreversible. Correlated evolutions of the optical and elastic properties of the interfaces, as probed by ellipsometry and surface dilatational rheology respectively, provide a consistent physical picture of the building up of the protein interfacial layer. A progressive coverage of the interface by the proteins occurs at low Π. This stage is followed, at higher Π, by conformational rearrangements of the protein film, which are identified by a strong increase of the dissipative viscoelastic properties of the film concomitantly with a peculiar evolution of its optical profile that we have rationalized. In the last stage, at even higher surface pressure, the adsorption is arrested; the optical profile is not modified while the elasticity of the interfacial layer dramatically increases with the surface pressure, presumably due to the film ageing.