Interhemispheric asymmetry of c-Fos expression in glomeruli and the olfactory tubercle following repeated odor stimulation

Odor adaptation allows the olfactory system to regulate sensitivity to different stimulus intensities, which is essential for preventing saturation of the cell-transducing machinery and maintaining high sensitivity to persistent and repetitive odor stimuli. Although many studies have investigated the structure and mechanisms of the mammalian olfactory system that responds to chemical sensation, few studies have considered differences in neuronal activation that depend on the manner in which the olfactory system is exposed to odorants, or examined activity patterns of olfactory-related regions in the brain under different odor exposure conditions. To address these questions, we designed three different odor exposure conditions that mimicked diverse odor environments and analyzed c-Fos-expressing cells (c-Fos+ cells) in the odor columns of the olfactory bulb (OB). We then measured differences in the proportions of c-Fos-expressing cell types depending on the odor exposure condition. Surprisingly, under the specific odor condition in which the olfactory system was repeatedly exposed to the odorant for 1 min at 5-min intervals, one of the lateral odor columns and the ipsilateral hemisphere of the olfactory tubercle had more c-Fos+ cells than the other three odor columns and the contralateral hemisphere of the olfactory tubercle. However, this interhemispheric asymmetry of c-Fos expression was not observed in the anterior piriform cortex. To confirm whether the anterior olfactory nucleus pars externa (AONpE), which connects the left and right OB, contributes to this asymmetry, AONpE-lesioned mice were analyzed under the specific odor exposure condition. Asymmetric c-Fos expression was not observed in the OB or the olfactory tubercle. These data indicate that the c-Fos expression patterns of the olfactory-related regions in the brain are influenced by the odor exposure condition and that asymmetric c-Fos expression in these regions was observed under a specific odor exposure condition due to synaptic linkage via the AONpE. © 2020 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.1

Patent challenges and factors associated with successful patent challengers under the patent linkage system: recent evidence from South Korea after the Korea United States free trade agreement

Objectives The patent linkage system upgraded patent challenges to an important factor in granting timely market approval for generic drugs. We aim to understand patent challenges and identify the factors that are associated with successful patent challengers under the patent linkage system in South Korea. Methods We constructed a novel dataset that combined information on manufacturers with detailed data about their patent challenges after introduction of the patent linkage system. Based on the number of successful patent challenges, manufacturers were categorized into non-challengers, passive challengers, and aggressive challengers. Then, two types of logistic models were applied to identify the factors associated with successful and aggressive challengers. Findings Only 39 active ingredients were challenged by 77 manufacturers from March 2015 to December 2019. Of 171 manufacturers, 94 (55 %) were non-challengers, 58 (34 %) were passive challengers who had succeeded in fewer than 4 patent challenges, and 19 (11 %) were aggressive challengers who had succeeded in 4 or more patent challenges. Higher sales, more employees, and a greater number of reimbursed drugs were associated with being a patent challenger, while a greater number of reimbursed drugs was associated with being an aggressive challenger. Conclusion Some manufacturers utilize patent challenges to strengthen their product portfolios in the market. However, under the patent linkage system, the frequency of patent challenges is limited in South Korea compared to the United States. In particular, patent challenges against drugs in injection form and biologics are very rare

A pathogen-derived metabolite induces microglial activation via odorant receptors

Microglia (MG), the principal neuroimmune sentinels in the brain, continuously sense changes in their environment and respond to invading pathogens, toxins, and cellular debris, thereby affecting neuroinflammation. Microbial pathogens produce small metabolites that influence neuroinflammation, but the molecular mechanisms that determine whether pathogen-derived small metabolites affect microglial activation of neuroinflammation remain to be elucidated. We hypothesized that odorant receptors (ORs), the largest subfamily of G protein-coupled receptors, are involved in microglial activation by pathogen-derived small metabolites. We found that MG express high levels of two mouse ORs, Olfr110 and Olfr111, which recognize a pathogenic metabolite, 2-pentylfuran, secreted by Streptococcus pneumoniae. These interactions activate MG to engage in chemotaxis, cytokine production, phagocytosis, and reactive oxygen species generation. These effects were mediated through the G(alpha s)-cyclic adenosine monophosphate-protein kinase A-extracellular signal-regulated kinase and G(beta gamma)-phospholipase C-Ca2+ pathways. Taken together, our results reveal a novel interplay between the pathogen-derived metabolite and ORs, which has major implications for our understanding of microglial activation by pathogen recognition. Database Model data are available in the PMDB database under the accession number PM0082389.N

3D printable hyaluronic acid-based hydrogel for its potential application as a bioink in tissue engineering

Abstract Background After recognition of 3D printing and injectable hydrogel as a critical issue in tissue/organ engineering and regenerative medicine society, many hydrogels as bioinks have been developed worldwide by using polymeric biomaterials such as gelatin, alginate, hyaluronic acid and others. Even though some gels have shown good performances in 3D bioprinting, still their performances do not meet the requirements enough to be used as a bioink in tissue engineering. Method In this study, a hydrogel consisting of three biocompatible biomaterials such as hyaluronic acid (HA), hydroxyethyl acrylate (HEA) and gelatin-methacryloyl, i.e. HA-g-pHEA-gelatin gel, has been evaluated for its possibility as a bioprinting gel, a bioink. Hydrogel synthesis was obtained by graft polymerization of HEA to HA and then grafting of gelatin- methacryloyl via radical polymerization mechanism. Physical and biological properties of the HA-based hydrogels fabricated with different concentrations of methacrylic anhydride (6 and 8%) for gelatin-methacryloylation have been evaluated such as swelling, rheology, morphology, cell compatibility, and delivery of small molecular dimethyloxalylglycine. Printings of HA-g-pHEA-Gelatin gel and its bioink with bone cell loaded in lattice forms were also evaluated by using home-built multi-material (3D bio-) printing system. Conclusion The experimental results demonstrated that the HA-g-pHEA-gelatin hydrogel showed both stable rheology properties and excellent biocompatibility, and the gel showed printability in good shape. The bone cells in bioinks of the lattice-printed scaffolds were viable. This study showed HA-g-pHEA-Gelatin gel’s potential as a bioink or its tissue engineering applications in injectable and 3D bioprinting forms

Fatty Acid Increases cAMP-dependent Lactate and MAO-B-dependent GABA Production in Mouse Astrocytes by Activating a Gαs Protein-coupled Receptor

Medium-chain fatty acids (MCFAs) are mostly generated from dietary triglycerides and can penetrate the blood-brain barrier. Astrocytes in the brain use MCFAs as an alternative energy source. In addition, MCFAs have various regulatory and signaling functions in astrocytes. However, it is unclear how astrocytes sense and take up MCFAs. This study demonstrates that decanoic acid (DA; C10), a saturated MCFA and a ligand of G(alpha s) protein-coupled receptors (G(alpha s)-GPCRs), is a signaling molecule in energy metabolism in primary astrocytes. cAMP synthesis and lactate release were increased via a putative G(alpha s)-GPCR and transmembrane adenylyl cyclase upon short-term treatment with DA. By contrast, monoamine oxidase B-dependent gamma-aminobutyric acid (GABA) synthesis was increased in primary cortical and hypothalamic astrocytes upon long-term treatment with DA. Thus, astrocytes respond to DA by synthesizing cAMP and releasing lactate upon short-term treatment, and by synthesizing and releasing GABA upon long-term treatment, similar to reactive astrocytes. Our data suggest that astrocytes in the brain play crucial roles in lipid-sensing via GPCRs and modulate neuronal metabolism or activity by releasing lactate via astrocyte-neuron lactate shuttle or GABA to influence neighboring neurons.1

Small-chain fatty acid activates astrocytic odorant receptor Olfr920

Odorant receptors are the largest subfamily of G protein-coupled receptors and were recently suggested to play critical roles in nonolfactory tissues. However, the expression and function of odorant receptors in astrocytes, the most abundant cells in the brain, are not well known. We demonstrate that Olfr920 is highly expressed and propose that it functions as a short-chain fatty acid sensor in primary cortical astrocytes. The short-chain fatty acid isobutyric acid (IBA) was identified via a luciferase assay as an Olfr920 ligand. We show that IBA activates the Gs protein-adenylyl cyclase-cAMP pathway via Olfr920 in primary cortical astrocytes by using cAMP and knockdown analyses. In addition, IBA reduces lipopolysaccharide-induced glial fibrillary acidic protein expression in reactive astrocytes. These results suggest that astrocytic Olfr920 is a potential novel target for increased reactive astrocytes. © 20191

MRPrimer: a MapReduce-based method for the thorough design of valid and ranked primers for PCR

Primer design is a fundamental technique that is widely used for polymerase chain reaction (PCR). Although many methods have been proposed for primer design, they require a great deal of manual effort to generate feasible and valid primers, including homology tests on off-target sequences using BLAST-like tools. That approach is inconvenient for many target sequences of quantitative PCR (qPCR) due to considering the same stringent and allele-invariant constraints. To address this issue, we propose an entirely new method called MRPrimer that can design all feasible and valid primer pairs existing in a DNA database at once, while simultaneously checking a multitude of filtering constraints and validating primer specificity. Furthermore, MRPrimer suggests the best primer pair for each target sequence, based on a ranking method. Through qPCR analysis using 343 primer pairs and the corresponding sequencing and comparative analyses, we showed that the primer pairs designed by MRPrimer are very stable and effective for qPCR. In addition, MRPrimer is computationally efficient and scalable and therefore useful for quickly constructing an entire collection of feasible and valid primers for frequently updated databases like RefSeq. Furthermore, we suggest that MRPrimer can be utilized conveniently for experiments requiring primer design, especially real-time qPCR. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

The macrophage odorant receptor Olfr78 mediates the lactate-induced M2 phenotype of tumor-associated macrophages

Expression and function of odorant receptors (ORs), which account for more than 50% of G protein-coupled receptors, are being increasingly reported in nonolfactory sites. However, ORs that can be targeted by drugs to treat diseases remain poorly identified. Tumorderived lactate plays a crucial role in multiple signaling pathways leading to generation of tumor-associated macrophages (TAMs). In this study, we hypothesized that the macrophage OR Olfr78 functions as a lactate sensor and shapes the macrophage-tumor axis. Using Olfr78+/+ and Olfr78-/- bone marrow-derived macrophages with or without exogenous Olfr78 expression, we demonstrated that Olfr78 sensed tumor-derived lactate, whichwas themain factor in tumor-conditioned media responsible for generation of protumoral M2-TAMs. Olfr78 functioned together with Gpr132 to mediate lactate-induced generation of protumoral M2-TAMs. In addition, syngeneic Olfr78-deficient mice exhibited reduced tumor progression and metastasis together with an increased anti- versus protumoral immune cell population. We propose that the Olfr78-lactate interaction is a therapeutic target to reduce and prevent tumor progression and metastasis. © 2021 National Academy of Sciences. All rights reserved.1

Impact of sphingosine and acetylsphingosines on the aggregation and toxicity of metal-free and metal-treated amyloid-beta

Pathophysiological shifts in the cerebral levels of sphingolipids in Alzheimer's disease (AD) patients suggest a link between sphingolipid metabolism and the disease pathology. Sphingosine (SP), a structural backbone of sphingolipids, is an amphiphilic molecule that is able to undergo aggregation into micelles and micellar aggregates. Considering its structural properties and cellular localization, we hypothesized that SP potentially interacts with amyloid-beta (A beta) and metal ions that are found as pathological components in AD-affected brains, with manifesting its reactivity towards metal-free A beta and metal-bound A beta (metal-A beta). Herein, we report, for the first time, that SP is capable of interacting with both A beta and metal ions and consequently affects the aggregation of metal-free A beta and metal-A beta. Moreover, incubation of SP with A beta in the absence and presence of metal ions results in the aggravation of toxicity induced by metal-free A beta and metal-A beta in living cells. As the simplest acyl derivatives of SP, N-acetylsphingosine and 3-O-acetylsphingosine also influence metal-free A beta and metal-A beta aggregation to different degrees, compared to SP. Such slight structural modifications of SP neutralize its ability to exacerbate the cytotoxicity triggered by metal-free A beta and metal-A beta. Notably, the reactivity of SP and the acetylsphingosines towards metal-free A beta and metal-A beta is determined to be dependent on their formation of micelles and micellar aggregates. Our overall studies demonstrate that SP and its derivatives could directly interact with pathological factors in AD and modify their pathogenic properties at concentrations below and above critical aggregation concentrations

Mouse neutrophils express functional umami taste receptor T1R1/T1R3

Neutrophils play an important role in the initiation of innate immunity against infection and injury. Although many different types of G-protein coupled receptors are functionally expressed in neutrophils, no reports have demonstrated functional expression of umami taste receptor in these cells. We observed that mouse neutrophils express the umami taste receptor T1R1/T1R3 through RNA sequencing and quantitative RT-PCR analysis. Stimulation of mouse neutrophils with L-alanine or L-serine, which are ligands for the umami taste receptor, elicited not only ERK or p38 MAPK phosphorylation but also chemotactic migration. Moreover, addition of L-alanine or L-serine markedly reduced the production of several cytokines including TNF-α induced by lipopolysaccharide (LPS) through inhibition of NF-κB activity or STAT3 phosphorylation in neutrophils. Our findings demonstrate that neutrophils express the umami taste receptor, through which tastants stimulate neutrophils, resulting in chemotactic migration, and attenuation of LPS-induced inflammatory response. © 2014 by the The Korean Society for Biochemistry and Molecular Biology.1