An improved mouse model of sepsis based on intraperitoneal injections of the enriched culture of cecum slurry.

AIM: Sepsis is a life-threatening clinical syndrome comprising multiorgan dysfunctions caused by a disproportionate body immune response. There are several animal sepsis models which are based on cecum ligation, cecal puncture, and cecum slurry injection. The major limitation of all current sepsis models is the high variability owing to the variable degree of ligation, puncture and inconsistent microbial composition used for sepsis initiation. The primary objective of this work is to demonstrate the feasibility of a standardized method for sepsis development. MATERIALS AND METHODS: The cecal slurry bacterial culture was developed and preserved in glycerol stocks. Antibiotics aztreonam and vancomycin were used for generating several defined, enriched cecal slurry bacterial cultures. Mice survival was assessed until 48 hrs post injection, and the tissue samples were collected after 10 hrs from sepsis initiation. KEY FINDINGS: The results indicate that increasing polymicrobial load resulted in lower survival rates and was associated with the higher number of infiltrating immune cells and necrosis. H&E (haematoxylin & eosin) staining & serum markers revealed that septic mice exhibited increased inflammation and significant damage to the liver and kidneys. The defined Gram-negative and Gram-positive specific cecal slurry bacterial cultures were developed and their efficiency in inducing sepsis was characterized. SIGNIFICANCE: Enriched cecal slurry bacterial cultures can be stored in glycerol stocks at -80 °C. This has an ethical advantage of avoiding unnecessary animal euthanasia for each experiment and provides a standardization capability of sepsis development.

Adaptor molecules mediate negative regulation of macrophage inflammatory pathways: a closer look.

Macrophages play a central role in initiating, maintaining, and terminating inflammation. For that, macrophages respond to various external stimuli in changing environments through signaling pathways that are tightly regulated and interconnected. This process involves, among others, autoregulatory loops that activate and deactivate macrophages through various cytokines, stimulants, and other chemical mediators. Adaptor proteins play an indispensable role in facilitating various inflammatory signals. These proteins are dynamic and flexible modulators of immune cell signaling and act as molecular bridges between cell surface receptors and intracellular effector molecules. They are involved in regulating physiological inflammation and also contribute significantly to the development of chronic inflammatory processes. This is at least partly due to their involvement in the activation and deactivation of macrophages, leading to changes in the macrophages' activation/phenotype. This review provides a comprehensive overview of the 20 adaptor molecules and proteins that act as negative regulators of inflammation in macrophages and effectively suppress inflammatory signaling pathways. We emphasize the functional role of adaptors in signal transduction in macrophages and their influence on the phenotypic transition of macrophages from pro-inflammatory M1-like states to anti-inflammatory M2-like phenotypes. This endeavor mainly aims at highlighting and orchestrating the intricate dynamics of adaptor molecules by elucidating the associated key roles along with respective domains and opening avenues for therapeutic and investigative purposes in clinical practice.

Novel peptide inhibitors targeting CD40 and CD40L interaction: A potential for atherosclerosis therapy.

Atherosclerosis is a chronic inflammatory disease characterized by plaque build-up in the arteries, leading to the obstruction of blood flow. Macrophages are the primary immune cells found in the atherosclerotic lesions and are directly involved in atherosclerosis progression. Macrophages are derived from extravasating blood monocytes. The monocytic CD40 receptor is important for monocyte recruitment on the endothelium expressing the CD40 ligand (CD40L). Thus, targeting monocyte/macrophage interaction with the endothelium by inhibiting CD40-CD40L interaction may be a promising strategy for attenuating atherosclerosis. Monoclonal antibodies have been used against this target but shows various complications. We used an array of computer-aided drug discovery tools and molecular docking approaches to design a therapeutic inhibitory peptide that could efficiently bind to the critical residues (82Y, 84D, and 86N) on the CD40 receptor essential for the receptor's binding to CD40L. The initial screen identified a parent peptide with a high binding affinity to CD40, but the peptide exhibited a positive hepatotoxicity score. We then designed several novel peptidomimetic derivatives with higher binding affinities to CD40, good physicochemical properties, and negative hepatotoxicity as compared to the parent peptide. Furthermore, we conducted molecular dynamics simulations for both the apo and complexed forms of the receptor with ligand, and screened peptides to evaluate their stability. The designed peptidomimetic derivatives are promising therapeutics targeting the CD40-CD40L interaction and may potentially be used to attenuate atherosclerosis.

The indispensability of macrophage adaptor proteins in chronic inflammatory diseases.

Adaptor proteins represent key signalling molecules involved in regulating immune responses. The host's innate immune system recognizes pathogens via various surface and intracellular receptors. Adaptor molecules are centrally involved in different receptor-mediated signalling pathways, acting as bridges between the receptors and other molecules. The presence of adaptors in major signalling pathways involved in the pathogenesis of various chronic inflammatory diseases has drawn attention toward the role of these proteins in such diseases. In this review, we summarize the importance and roles of different adaptor molecules in macrophage-mediated signalling in various chronic disease states. We highlight the mechanistic roles of adaptors and how they are involved in protein-protein interactions (PPI) via different domains to carry out signalling. Hence, we also provide insights into how targeting these adaptor proteins can be a good therapeutic strategy against various chronic inflammatory diseases.

Nirmatrelvir exerts distinct antiviral potency against different human coronaviruses.

Nirmatrelvir is the main component of Paxlovid, an oral antiviral drug approved for the treatment of COVID-19 caused by SARS-COV-2 infection. Nirmatrelvir targets the main protease (Mpro), which is substantially conserved among different coronaviruses. Here, our molecular docking analysis indicates comparable affinity of nirmatrelvir binding to the Mpro enzymes of SARS-CoV-2 and three seasonal coronaviruses (OC43, 229E and NL63). However, in cell culture models, we found that nirmatrelvir potently inhibited SARS-CoV-2, OC43 and 229E, but not NL63. The insensitivity of NL63 to nirmatrelvir treatment was demonstrated at both viral replication and infectious titer levels. The antiviral activity of nirmatrelvir against OC43 and 229E was further confirmed in human airway organoids. The combination of nirmatrelvir and molnupiravir exerted differential patterns of antiviral response against OC43 and 229E. These results revealed disparities in the ability of nirmatrelvir to inhibit different coronaviruses, and caution against repurposing of nirmatrelvir as a pan-coronavirus treatment.