Toll-like receptor 4 (TLR4), the signal-transducing molecule of the LPS receptor complex, plays a fundamental role in the sensing of LPS from Gram-negative bacteria. Activation of TLR4 signaling ...pathways by LPS is a critical upstream event in the pathogenesis of Gram-negative sepsis, making TLR4 an attractive target for novel antisepsis therapy. To validate the concept of TLR4-targeted treatment strategies in Gram-negative sepsis, we first showed that TLR4⁻/⁻ and myeloid differentiation primary response gene 88 (MyD88)⁻/⁻ mice were fully resistant to Escherichia coli-induced septic shock, whereas TLR2⁻/⁻ and wild-type mice rapidly died of fulminant sepsis. Neutralizing anti-TLR4 antibodies were then generated using a soluble chimeric fusion protein composed of the N-terminal domain of mouse TLR4 (amino acids 1-334) and the Fc portion of human IgG1. Anti-TLR4 antibodies inhibited intracellular signaling, markedly reduced cytokine production, and protected mice from lethal endotoxic shock and E. coli sepsis when administered in a prophylactic and therapeutic manner up to 13 h after the onset of bacterial sepsis. These experimental data provide strong support for the concept of TLR4-targeted therapy for Gram-negative sepsis.
To assess the diagnostic value of procalcitonin (PCT), interleukin (IL)-6, IL-8, and standard measurements in identifying critically ill patients with sepsis, we performed prospective measurements in ...78 consecutive patients admitted with acute systemic inflammatory response syndrome (SIRS) and suspected infection. We estimated the relevance of the different parameters by using multivariable regression modeling, likelihood-ratio tests, and area under the receiver operating characteristic curves (AUC). The final diagnosis was SIRS in 18 patients, sepsis in 14, severe sepsis in 21, and septic shock in 25. PCT yielded the highest discriminative value, with an AUC of 0.92 (CI, 0.85 to 1.0), followed by IL-6 (0.75; CI, 0.63 to 0.87), and IL-8 (0.71; CI, 0.59 to 0.83; p < 0.001). At a cutoff of 1.1 ng/ml, PCT yielded a sensitivity of 97% and a specificity of 78% to differentiate patients with SIRS from those with sepsis-related conditions. Median PCT concentrations on admission (ng/ ml, range) were 0.6 (0 to 5.3) for SIRS; 3.5 (0.4 to 6.7) for sepsis; 6.2 (2.2 to 85) for severe sepsis; and 21.3 (1.2 to 654) for septic shock (p < 0.001). The addition of PCT to a model based solely on standard indicators improved the predictive power of detecting sepsis (likelihood ratio test; p = 0.001) and increased the AUC value for the routine value-based model from 0.77 (CI, 0.64 to 0.89) to 0.94 (CI, 0.89 to 0.99; p = 0.002). In contrast, no additive effect was seen for IL-6 (p = 0.56) or IL-8 (p = 0.14). Elevated PCT concentrations appear to be a promising indicator of sepsis in newly admitted, critically ill patients capable of complementing clinical signs and routine laboratory parameters suggestive of severe infection.
To review the role of macrophage migration inhibitory factor (MIF) in host responses to infection and to explore the potential of MIF as a novel target for therapeutic intervention in patients with ...severe sepsis and septic shock.
Published articles on the role of MIF in innate immune responses against microbial pathogens.
MIF has emerged recently as an important effector molecule of the innate immune system. MIF is expressed constitutively by monocytes/macrophages, T cells, B cells, endocrine cells, and epithelial cells. Microbial toxins and cytokines are powerful inducers of MIF release by immune cells. MIF expression is up-regulated during the course of inflammatory and infectious diseases and was found to play an important role in the pathogenesis of sepsis and septic shock.
Given the role of MIF in innate immune responses against microbial pathogens and in the regulation of inflammatory responses, modulation of MIF production or neutralization of its activity may offer new therapeutic options for the management of patients with sepsis.
Macrophages are essential effector cells of innate immunity that play a pivotal role in the recognition and elimination of invasive microorganisms. Mediators released by activated macrophages ...orchestrate innate and adaptive immune host responses. The cytokine macrophage migration inhibitory factor (MIF) is an integral mediator of the innate immune system. Monocytes and macrophages constitutively express large amounts of MIF, which is rapidly released after exposure to bacterial toxins and cytokines. MIF exerts potent proinflammatory activities and is an important cytokine of septic shock. Recent investigations of the mechanisms by which MIF regulates innate immune responses to endotoxin and gram-negative bacteria indicate that MIF acts by modulating the expression of Toll-like receptor 4, the signal-transducing molecule of the lipopolysaccharide receptor complex. Given its role in innate immune responses to bacterial infections, MIF is a novel target for therapeutic intervention in patients with septic shock.
The cytokine macrophage migration inhibitory factor (MIF) has emerged recently as an important mediator of inflammation and innate immunity. MIF is rapidly released by macrophages after stimulation ...with microbial products and pro-inflammatory cytokines and, in turn, stimulates the production of pro-inflammatory mediators by immune cells. Immunoneutralization of MIF or deletion of the Mif gene was shown to protect animals from lethal endotoxemia, staphylococcal toxic shock and septic shock in experimental models of bacterial peritonitis. To investigate the function of MIF in innate immunity, we studied the response of macrophages expressing reduced levels of MIF to microbial products. These cells were generated by transduction of an antisense MIF adenovirus or by stable transfection with an antisense MIF plasmid or were obtained from MIF-knockout mice. MIF-deficient macrophages were shown to be hyporesponsive to stimulation with LPS and Gram-negative bacteria. The defect was associated with a down-regulation of Toll-like receptor 4 (TLR4), the signal transducing molecule of the LPS receptor complex. Immunoneutralization of extracellular MIF decreased TLR4 expression and responses of macrophages to LPS, indicating that MIF may exert autocrine effects. These findings identify an important role for MIF in innate immunity and provide a rationale for the development of anti-MIF strategy for the treatment of patients with Gram-negative septic shock.
The cytokine macrophage migration inhibitory factor (MIF) has emerged recently as an important mediator of inflammation and innate immunity. MIF is rapidly released by macrophages after stimulation ...with microbial products and pro-inflammatory cytokines and, in turn, stimulates the production of pro-inflammatory mediators by immune cells. Immunoneutralization of MIF or deletion of the Mif gene was shown to protect animals from lethal endotoxemia, staphylococcal toxic shock and septic shock in experimental models of bacterial peritonitis. To investigate the function of MIF in innate immunity, we studied the response of macrophages expressing reduced levels of MIF to microbial products. These cells were generated by transduction of an antisense MIF adenovirus or by stable transfection with an antisense MIF plasmid or were obtained from MIFknockout mice. MIF-deficient macrophages were shown to be hyporesponsive to stimulation with LPS and Gram-negative bacteria. The defect was associated with a down-regulation of Toll-like receptor 4 (TLR4), the signal transducing molecule of the LPS receptor complex. Immunoneutralization of extracellular MIF decreased TLR4 expression and responses of macrophages to LPS, indicating that MIF may exert autocrine effects. These findings identify an important role for MIF in innate immunity and provide a rationale for the development of anti-MIF strategy for the treatment of patients with Gram-negative septic shock.
Here, we highlight the potential translational benefits of delivering FLASH radiotherapy using ultra-high dose rates (>100 Gy·s−1). Compared with conventional dose-rate (CONV; 0.07–0.1 Gy·s−1) ...modalities, we showed that FLASH did not cause radiation-induced deficits in learning and memory in mice. Moreover, 6 months after exposure, CONV caused permanent alterations in neurocognitive end points, whereas FLASH did not induce behaviors characteristic of anxiety and depression and did not impair extinction memory. Mechanistic investigations showed that increasing the oxygen tension in the brain through carbogen breathing reversed the neuroprotective effects of FLASH, while radiochemical studies confirmed that FLASH produced lower levels of the toxic reactive oxygen species hydrogen peroxide. In addition, FLASH did not induce neuroinflammation, a process described as oxidative stress-dependent, and was also associated with a marked preservation of neuronal morphology and dendritic spine density. The remarkable normal tissue sparing afforded by FLASH may someday provide heretofore unrealized opportunities for dose escalation to the tumor bed, capabilities that promise to hasten the translation of this groundbreaking irradiation modality into clinical practice.
Abstract
Background
Sjögren’s syndrome is a systemic autoimmune disease characterized by immune cells predominantly infiltrating the exocrine glands and frequently forming ectopic lymphoid ...structures. These structures drive a local functional immune response culminating in autoantibody production and tissue damage, associated with severe dryness of mucosal surfaces and salivary gland hypofunction. Cenerimod, a potent, selective and orally active sphingosine-1-phosphate receptor 1 modulator, inhibits the egress of lymphocytes into the circulation. Based on the mechanism of action of cenerimod, its efficacy was evaluated in two mouse models of Sjögren’s syndrome.
Methods
Cenerimod was administered in two established models of Sjögren’s syndrome; firstly, in an inducible acute viral sialadenitis model in C57BL/6 mice, and, secondly, in the spontaneous chronic sialadenitis MRL/lpr mouse model. The effects of cenerimod treatment were then evaluated by flow cytometry, immunohistochemistry, histopathology and immunoassays. Comparisons between groups were made using a Mann-Whitney test.
Results
In the viral sialadenitis model, cenerimod treatment reduced salivary gland immune infiltrates, leading to the disaggregation of ectopic lymphoid structures, reduced salivary gland inflammation and preserved organ function. In the MRL/lpr mouse model, cenerimod treatment decreased salivary gland inflammation and reduced T cells and proliferating plasma cells within salivary gland ectopic lymphoid structures, resulting in diminished disease-relevant autoantibodies within the salivary glands.
Conclusions
Taken together, these results suggest that cenerimod can reduce the overall autoimmune response and improve clinical parameters in the salivary glands in models of Sjögren’s syndrome and consequently may reduce histological and clinical parameters associated with the disease in patients.
•An original strategy of enzyme immobilization by entrapment in plasma-polymerized thin films is proposed.•RPECVD is used to obtain the ppTMDSO thin films overcoating the adsorbed ...β-galactosidase.•The role of both enzyme solvent and adsorption support is investigated.•The enzyme leaching from the support and its resulting activity are analyzed as function of various coating thicknesses.•β-galactosidase activity is characterized by studying its stability, the diffusional limitations and the kinetic parameters.
Over the years, immobilization of biologically active species such as enzymes onto solid support gave rise to a wide range of analytical and industrial applications. The development of fast, simple and efficient immobilization strategies is becoming of great importance in specific Biological Micro-Electromechanical Systems (BioMEMS) manufacturing. Thus, the current work focuses on an original methodology and mild procedure for β-galactosidase immobilization. Using as support either silicon or a thin film obtained from polymerization of 1,1,3,3-tetramethyldisiloxane (ppTMDSO) deposited by Plasma Enhanced Chemical Vapor Deposition in afterglow mode, the strategy developed here consisted in adsorption of β-galactosidase followed by its overcoating by the same siloxane plasma polymer. After sample washing, the enzymes were characterized to be efficiently entrapped within the porous polymer matrix while allowing the penetration and hydrolysis of the synthetic substrate ortho-nitrophenyl-β-d-galactopyranoside (o-NPG) with stability over at least 8 assays. The entrapment procedure allowed obtaining bio-functionnal coatings where β-galactosidase was expected to be included in the plasma-polymerized films while preserving its native structure and its activity. This latter was modulated by mass transfer limitations of the substrate according to the thickness of the ppTMDSO coatings. The dry-process-based-preparation of such a thin bio-functional film (from ∼200nm to ∼650nm) is fast and compatible with biochip or microreactor fabrication processes while avoiding the use of lot of chemicals and multi-step treatments commonly encountered in enzyme immobilization procedures.
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