MyD88-dependent Toll-like receptor 2 signaling modulates macrophage activation on lysate-adsorbed Teflon™ AF surfaces in an biomaterial host response model.

The adsorbed protein layer on an implanted biomaterial surface is known to mediate downstream cell-material interactions that drive the host response. While the adsorption of plasma-derived proteins has been studied extensively, the adsorption of damage-associated molecular patterns (DAMPs) derived from damaged cells and matrix surrounding the implant remains poorly understood. Previously, our group developed a DAMP-adsorption model in which 3T3 fibroblast lysates were used as a complex source of cell-derived DAMPs and we demonstrated that biomaterials with adsorbed lysate potently activated RAW-Blue macrophages via Toll-like receptor 2 (TLR2).

The extended effect of adsorbed damage-associated molecular patterns and Toll-like receptor 2 signaling on macrophage-material interactions.

Implanted biomaterials elicit an immune-mediated foreign body reaction (FBR) that results in the fibrous encapsulation of the implant and can critically impact the performance of some implants. Consequently, understanding the molecular mechanisms that underpin cell-materials interactions that initiate biomaterial-induced inflammation and fibrosis is critical to improving the performance of biomaterial implants negatively impacted by the FBR. Damage-associated molecular patterns (DAMPs) are endogenous mediators of inflammation that are released upon tissue injury and induce sterile inflammation Toll-like receptors (TLRs).

Arginase-1 deficiency in neural cells does not contribute to neurodevelopment or functional outcomes after sciatic nerve injury.

Arginase-1 (Arg1) is an enzyme controlling the final step of the urea cycle, with highest expression in the liver and lower expression in the lungs, pancreas, kidney, and some blood cells. Arg1 deficiency is an inherited urea cycle disorder presenting with neurological dysfunction including spastic diplegia, intellectual and growth retardation, and encephalopathy. The contribution of Arg1 expression in the central and peripheral nervous system to the development of neurological phenotypes remains largely unknown.

Perivascular adipose tissue-derived extracellular vesicle miR-221-3p mediates vascular remodeling.

Adipose tissue-secreted extracellular vesicles (EVs) containing microRNAs (miRNAs) convey intercellular message signaling. The biogenesis of EV-miRNAs from perivascular adipose tissue (PVAT) and their roles in intercellular communication in response to obesity-associated inflammation have not yet been fully explored. By feeding mice a high-fat diet for 16 wk, we established obesity-associated, chronic low-grade inflammation in PVAT, characterized as hypertrophy of perivascular adipocytes, decreased adipogenesis, and proinflammatory macrophage infiltration.

Isoform-Specific Compensation of Cyclooxygenase (Ptgs) Genes during Implantation and Late-Stage Pregnancy.

The participation of cyclooxygenase (COX) in embryo implantation and parturition has been studied extensively. However, the distinct role of the two COX isoforms in these processes still remains unclear. Using three characterized mouse lines where the Ptgs1 and Ptgs2 genes substitute for one another, this study focused on the reproductive significance of their distinct roles and potential biological substitution.

Differential compensation of two cyclooxygenases in renal homeostasis is independent of prostaglandin-synthetic capacity under basal conditions.

The distinct functions of each cyclooxygenase (COX) isoform in renal homeostasis have been the subject of intense investigation for many years. We took the novel approach of using 3 characterized mouse lines, where the prostaglandin (PG)-endoperoxide synthase genes 1 and 2 ( Ptgs1 and Ptgs2) substitute for one another to delineate distinct roles and the potential for COX isoform substitution. Flipped Ptgs genes generate a reversed COX-expression pattern in the kidney, where the knockin COX-2 is highly expressed.

Transplantation of Gene-Edited Hepatocyte-like Cells Modestly Improves Survival of Arginase-1-Deficient Mice.

Progress in gene editing research has been accelerated by utilizing engineered nucleases in combination with induced pluripotent stem cell (iPSC) technology. Here, we report transcription activator-like effector nuclease (TALEN)-mediated reincorporation of Arg1 exons 7 and 8 in iPSCs derived from arginase-1-deficient mice possessing Arg1 alleles lacking these terminal exons. The edited cells could be induced to differentiate into hepatocyte-like cells (iHLCs) in vitro and were subsequently used for transplantation into our previously described (Sin et al.

Flipping the cyclooxygenase () genes reveals isoform-specific compensatory functions.

Two prostaglandin (PG) H synthases encoded by genes, colloquially known as cyclooxygenase (COX)-1 and COX-2, catalyze the formation of PG endoperoxide H, the precursor of the major prostanoids. To address the functional interchangeability of these two isoforms and their distinct roles, we have generated COX-2>COX-1 mice whereby is knocked in to the locus. We then "flipped" genes to successfully create the Reversa mouse strain, where knock-in COX-2 is expressed constitutively and knock-in COX-1 is lipopolysaccharide (LPS) inducible.

Genomic and lipidomic analyses differentiate the compensatory roles of two COX isoforms during systemic inflammation in mice.

Both cyclooxygenase (COX)-1 and COX-2, encoded by and , function coordinately during inflammation. But the relative contributions and compensations of COX-1 and COX-2 to inflammatory responses remain unanswered. We used three engineered mouse lines where the and genes substitute for one another to discriminate the distinct roles and interchangeability of COX isoforms during systemic inflammation.

Proof-of-Concept Gene Editing for the Murine Model of Inducible Arginase-1 Deficiency.

Arginase-1 deficiency in humans is a rare genetic disorder of metabolism resulting from a loss of arginase-1, leading to impaired ureagenesis, hyperargininemia and neurological deficits. Previously, we generated a tamoxifen-inducible arginase-1 deficient mouse model harboring a deletion of Arg1 exons 7 and 8 that leads to similar biochemical defects, along with a wasting phenotype and death within two weeks. Here, we report a strategy utilizing the Clustered, Regularly Interspaced, Short Palindromic Repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system in conjunction with piggyBac technology to target and reincorporate exons 7 and 8 at the specific Arg1 locus in attempts to restore the function of arginase-1 in induced pluripotent stem cell (iPSC)-derived hepatocyte-like cells (iHLCs) and macrophages in vitro.

Liver-specific knockout of arginase-1 leads to a profound phenotype similar to inducible whole body arginase-1 deficiency.

Arginase-1 (Arg1) converts arginine to urea and ornithine in the distal step of the urea cycle in liver. We previously generated a tamoxifen-inducible Arg1 deficient mouse model (Arg1-Cre) that disrupts Arg1 expression throughout the whole body and leads to lethality ≈ 2 weeks after gene disruption. Here, we evaluate if liver-selective Arg1 loss is sufficient to recapitulate the phenotype observed in global Arg1 knockout mice, as well as to gauge the effectiveness of gene delivery or hepatocyte transplantation to rescue the phenotype.

Strategies to rescue the consequences of inducible arginase-1 deficiency in mice.

Arginase-1 catalyzes the conversion of arginine to ornithine and urea, which is the final step of the urea cycle used to remove excess ammonia from the body. Arginase-1 deficiency leads to hyperargininemia in mice and man with severe lethal consequences in the former and progressive neurological impairment to varying degrees in the latter. In a tamoxifen-induced arginase-1 deficient mouse model, mice succumb to the enzyme deficiency within 2 weeks after inducing the knockout and retain <2 % enzyme in the liver.

Endogenously generated omega-3 fatty acids attenuate vascular inflammation and neointimal hyperplasia by interaction with free fatty acid receptor 4 in mice.

Background: Omega-3 polyunsaturated fatty acids (ω3 PUFAs) suppress inflammation through activation of free fatty acid receptor 4 (FFAR4), but this pathway has not been explored in the context of cardiovascular disease. We aimed to elucidate the involvement of FFAR4 activation by ω3 PUFAs in the process of vascular inflammation and neointimal hyperplasia in mice.

Methods And Results: We used mice with disruption of FFAR4 (Ffar4(-/-)), along with a strain that synthesizes high levels of ω3 PUFAs (fat-1) and a group of crossed mice (Ffar4(-/-)/fat-1), to elucidate the role of FFAR4 in vascular dysfunction using acute and chronic thrombosis/vascular remodeling models.

Multiple-site activation of the cysteinyl leukotriene receptor 2 is required for exacerbation of ischemia/reperfusion injury.

Objective: Transgenic overexpression of the human cysteinyl leukotriene receptor 2 (CysLT2R) in murine endothelium exacerbates vascular permeability and ischemia/reperfusion injury. Here, we explore the underlying mechanisms of CysLT2R activation-mediated inflammation and delineate the relative contributions of endogenous murine CysLT2R and the transgene-derived receptor.

Approach And Results: We created a novel mouse with only endothelial-expressed CysLT2R (endothelium-targeted overexpression mice [EC]/CysLT2R-knockout mice [KO]) by crossing EC with KO to dissect the role of endothelial CysLT2R in tissue injury.

Inducible arginase 1 deficiency in mice leads to hyperargininemia and altered amino acid metabolism.

Arginase deficiency is a rare autosomal recessive disorder resulting from a loss of the liver arginase isoform, arginase 1 (ARG1), which is the final step in the urea cycle for detoxifying ammonia. ARG1 deficiency leads to hyperargininemia, characterized by progressive neurological impairment, persistent growth retardation and infrequent episodes of hyperammonemia. Using the Cre/loxP-directed conditional gene knockout system, we generated an inducible Arg1-deficient mouse model by crossing "floxed" Arg1 mice with CreER(T2) mice.

A selective cysteinyl leukotriene receptor 2 antagonist blocks myocardial ischemia/reperfusion injury and vascular permeability in mice.

Cysteinyl leukotrienes (CysLTs) are potent inflammatory mediators that predominantly exert their effects by binding to cysteinyl leukotriene receptors of the G protein-coupled receptor family. CysLT receptor 2 (CysLT(2)R), expressed in endothelial cells of some vascular beds, has been implicated in a variety of cardiovascular functions. Endothelium-specific overexpression of human CysLT(2)R in transgenic mice (hEC-CysLT(2)R) greatly increases myocardial infarction damage.