ABSTRACT
BACKGROUND: Tachykinins and their cognate receptors, neurokinin receptors (NKs) including NK1, NK2, and NK3 play vital roles in regulating various physiological processes including neurotransmission, nociception, inflammation, smooth muscle contractility, and stimulation of endocrine and exocrine gland secretion. Their abnormal expression has been reported to be associated with neurological disorders, inflammation, and cancer. Even though NKs are expressed in the same cells with their expression being inversely correlated in some conditions, there is no direct evidence to prove their interaction. Understanding the functional crosstalk between NKs in mediated downstream signaling and cellular responses may elucidate the roles of each receptor in pathophysiology. RESULTS: In this study, we showed that NKs were co-expressed in some cells. However, different from NK3, which only forms homodimerization, we demonstrated a direct interaction between NK1 and NK2 at the protein level using co-immunoprecipitation and NanoBiT-based protein interaction analysis. Through heterodimerization, NK2 downregulated substance P-stimulated NK1 signals, such as intracellular Ca2+ mobilization and ERK phosphorylation, by enhancing ß-arrestin recruitment, even at the ligand concentration that could not activate NK2 itself or in the presence of NK1 specific antagonist, aprepitant. In A549 cells with receptors deleted and reconstituted, NK2 exerted a negative effect on substance P/NK1-mediated cell migration. CONCLUSION: Our study has provided the first direct evidence of an interaction between NK1 and NK2, which highlights the functional relevance of their heterodimerization in cellular responses. Our findings demonstrated that through dimerization, NK2 exerts negative effects on downstream signaling and cellular response mediated by NK1. Moreover, this study has significant implications for understanding the complexity of GPCR dimerization and its effect on downstream signaling and cellular responses. Given the important roles of tachykinins and NKs in pathophysiology, these insights may provide clues for developing NKs-targeting drugs.
ABSTRACT
The spexin-based GALR2 agonist (NS200) is a novel drug, which has shown antidepressant and anxiolytic action in a recent experimental study. In this study, we investigated the effects of NS200 on renal injury in an animal model of type 2 diabetes. Eight-week-old diabetic db/db mice were administered NS200 for 12 weeks. NS200 was intraperitoneally administered at a dose of 1.0â mg/kg/day. Metabolic parameters and structural and molecular changes in the kidneys were compared among the three groups: non-diabetic db/m control, db/db mice, and NS200-treated db/db mice. In db/db mice, NS200 administration did not impact the body weight, food and water intake, urinary volume, fasting blood glucose level, or HbA1c levels. Insulin and glucose tolerance were also unaffected by NS200 treatment. However, NS200 improved urinary albumin excretion and glomerulosclerosis in diabetic kidneys. Activation of TGFß1 and insulin signaling pathways, such as PI3â K /AKT/ERK, were inhibited by NS200. In conclusion, a spexin-based GALR2 agonist attenuated diabetic nephropathy by alleviating renal fibrosis in mice with type 2 diabetes. Spexin-based GALR2 agonists have considerable potential as novel treatment agents in diabetic nephropathy.
ABSTRACT
C-X-C motif chemokine ligand 12(CXCL12) is an essential chemokine for organ development and homeostasis in multiple tissues. Its receptor, C-X-C chemokine receptor type 4(CXCR4), is expressed on the surface of target cells. The chemokine and receptor are expressed almost ubiquitously in human tissues and cells throughout life, and abnormal expression of CXCL12 and CXCR4 is observed in pathological conditions, such as inflammation and cancer. CXCR4 is reportedly translated into five splicing variants of different lengths, which each have different amino acids in the N-terminus. As the N-terminus is the first recognition site for chemokines, CXCR4 variants may respond differently to CXCL12. Despite these differences, the molecular and functional properties of CXCR4 variants have not been thoroughly described or compared. Here, we explored the expression of CXCR4 variants in cell lines and analyzed their roles in cellular responses using biochemical approaches. RT-PCR revealed that most cell lines express more than one CXCR4 variant. When expressed in HEK293 cells, the CXCR4 variants differed in protein expression efficiency and cell surface localization. Although variant 2 demonstrated the strongest expression and cell surface localization, variants 1, 3, and 5 also mediated chemokine signaling and induced cellular responses. Our results demonstrate that the N-terminal sequences of each CXCR4 variant determine the expression of the receptor and affect ligand recognition. Functional analyses revealed that CXCR4 variants may also affect each other or interact during CXCL12-stimulated cellular responses. Altogether, our results suggest that CXCR4 variants may have distinct functional roles that warrant additional investigation and could contribute to future development of novel drug interventions.
Subject(s)
Chemokine CXCL12 , Receptors, CXCR4 , Humans , HEK293 Cells , Ligands , Receptors, CXCR4/metabolism , Chemokine CXCL12/genetics , Chemokine CXCL12/metabolism , Signal Transduction , Protein Processing, Post-TranslationalABSTRACT
CXCR3 regulates leukocyte trafficking, maturation, and various pathophysiological conditions. Alternative splicing generates three CXCR3 isoforms in humans. Previous studies investigated the roles of CXCR3 isoforms, and some biochemical data are not correlated with biological relevance analyses. RT-PCR analyses indicate that most cells express all three splicing variants, suggesting that they may mutually affect the chemokine binding and cellular responses of other splicing variants. Here, we performed an integrative analysis of the functional relations among CXCR3 splicing variants and their chemokine-dependent signaling using NanoBiT live cell protein interaction assays. The results indicated that the CXCR3 N-terminal region affected cell surface expression levels and ligand-dependent activation. CXCR3A was efficiently expressed in the plasma membrane and responded to I-TAC, IP-10, and MIG chemokines. By contrast, CXCR3B had low plasma membrane expression and mediated I-TAC-stimulated cellular responses. CXCR3Alt was rarely expressed on the cell surface and did not mediate any cell responses to the tested chemokines; however, CXCR3Alt negatively affected the plasma membrane expression of CXCR3A and CXCR3B and their chemokine-stimulated cellular responses. Jurkat cells express endogenous CXCR3, and exogenous CXCR3A expression enhanced chemotactic activity in response to I-TAC, IP-10, and MIG. By contrast, exogenous expression of CXCR3B and CXCR3Alt eliminated or reduced the CXCR3A-induced chemotactic activity. The PF-4 chemokine did not activate any CXCR3-mediated cellular responses. NanoBiT technology are useful to integrative studies of CXCR3-mediated cell signaling, and expand our knowledge of the cellular responses mediated by molecular interactions among the splicing variants, including cell surface expression, ligand-dependent receptor activation, and chemotaxis.
Subject(s)
Chemokine CXCL10 , Signal Transduction , Humans , Chemokine CXCL10/genetics , Chemokine CXCL10/metabolism , Ligands , Alternative Splicing , Protein Isoforms/genetics , Protein Isoforms/metabolism , Receptors, CXCR3/genetics , Receptors, CXCR3/metabolismABSTRACT
Despite the various medications used in clinics, the efforts to develop more effective treatments for depression continue to increase in the past decades mainly because of the treatment-resistant population, and the testing of several hypotheses- and target-based treatments. Undesirable side effects and unresponsiveness to current medications fuel the drive to solve this top global health problem. In this study, we focused on neuroinflammatory response-mediated depression which represents a cluster of depression etiology both in animal models and humans. Several meta-analyses reported that proinflammatory cytokines such as interleukin 6 (IL-6) and tumor necrosis factor-α (TNF-α) were increased in major depressive disorder patients. Inflammatory mediators implicated in depression include type-I interferon and inflammasome pathways. To elucidate the molecular mechanisms of neuroinflammatory cascades underlying the pathophysiology of depression, we introduced hycanthone, an antischistosomal drug, to check whether it can counteract depressive-like behaviors in vivo and normalize the inflammation-induced changes in vitro. Lipopolysaccharide (LPS) treatment increased proinflammatory cytokine expression in the murine microglial cells as well as the stimulation of type I interferon-related pathways that are directly or indirectly regulated by Janus kinase-signal transducer and activator of transcription (JAK-STAT) activation. Hycanthone treatment attenuated those changes possibly by inhibiting the JAK-STAT pathway and inflammasome activation. Hycanthone also ameliorated depressive-like behaviors by LPS. Taken together, we suggest that the inhibitory action of hycanthone against the interferon pathway leading to attenuation of depressive-like behaviors can be a novel therapeutic mechanism for treating depression.
ABSTRACT
BACKGROUND: C-C motif chemokine receptor 2 (CCR2), the main receptor for monocyte chemoattractant protein-1 (MCP-1), is expressed on immune cells, including monocytes, macrophages, and activated T cells, and mediates cell migration toward MCP-1 in inflammation-related diseases. The CCR2 gene encodes two isoforms: CCR2A and CCR2B. The CCR2B open reading frame is localized in a single exon, similar to other chemokine receptors, and CCR2A and CCR2B feature different amino acid sequences in their C-terminal intracellular loops due to alternative splicing. Most biochemical studies on CCR2-related cellular responses in the immune system have focused on CCR2B, with few reports focused on CCR2A. Understanding the functional properties of CCR2A in cellular responses may elucidate the roles played by MCP-1 and CCR2 in pathophysiological responses. RESULTS: CCR2 gene expression analysis in several cell types revealed that most adherent cells only expressed CCR2A, whereas CCR2B expression was dominant in monocytic cells. The C-terminal Helix 8 region of CCR2A contains few basic amino acids, which may be unfavorable for cell surface localization, as confirmed with the HiBiT assay. CCR2B contains many C-terminal Ser/Thr residues, similar to other chemokine receptors, which may be phosphorylated by G protein-coupled receptor kinases (GRKs) to promote ß-arrestin recruitment and subsequent endocytosis. By contrast, CCR2A contains few C-terminal Ser/Thr residues, which are unlikely to be phosphorylated by GRKs. CCR2A localized on the cell surface is resistant to internalization, despite the interaction between Gß and GRKs induced by ligand binding with CCR2A. CCR2A induced cellular responses at a relatively higher degree than CCR2B, although both receptors mediated signaling events through Gαq and Gαi. HeLa cells lacking CCR2A showed slowed growth compared with parent cells, regardless of MCP-1 stimulation, and their chemotactic activity toward MCP-1, in addition to basal motility, was significantly impaired. CONCLUSION: MCP-1 and CCR2 may play pivotal roles in cancer progression by recruiting macrophages into cancer tissue. This study demonstrates that CCR2A but not CCR2B is expressed in solid cancer-derived cells. CCR2A is resistant to internalization by ß-arrestin due to a distinct C-terminal region from CCR2B, which enhances MCP-1-stimulated responses, indicating that CCR2A may play essential roles in solid cancer progression.
ABSTRACT
Internalization of membrane proteins plays a key role in many physiological functions; however, highly sensitive and versatile technologies are lacking to study such processes in real-time living systems. Here we describe an assay based on bioluminescence able to quantify membrane receptor trafficking for a wide variety of internalization mechanisms such as GPCR internalization/recycling, antibody-mediated internalization, and SARS-CoV2 viral infection. This study represents an alternative drug discovery tool to accelerate the drug development for a wide range of physiological processes, such as cancer, neurological, cardiopulmonary, metabolic, and infectious diseases including COVID-19.
Subject(s)
Drug Discovery/methods , Membrane Proteins , Protein Transport/physiology , Spectrometry, Fluorescence/methods , COVID-19 , Drug Development/methods , HEK293 Cells , Humans , Luciferases/genetics , Luciferases/metabolism , Membrane Proteins/chemistry , Membrane Proteins/metabolism , Microscopy, Fluorescence , Nanotechnology , Receptors, G-Protein-Coupled , SARS-CoV-2/chemistry , SARS-CoV-2/metabolism , Virus InternalizationABSTRACT
Spexin (SPX) is a recently identified neuropeptide that is believed to play an important role in the regulation of energy homeostasis. Here, we describe a mediating function of SPX in hypothalamic leptin action. Intracerebroventricular (icv) SPX administration induced a decrease in food intake and body weight gain. SPX was found to be expressed in cells expressing leptin receptor ObRb in the mouse hypothalamus. In line with this finding, icv leptin injection increased SPX mRNA in the ObRb-positive cells of the hypothalamus, which was blocked by treatment with a STAT3 inhibitor. Leptin also increased STAT3 binding to the SPX promoter, as measured by chromatin immunoprecipitation assays. In vivo blockade of hypothalamic SPX biosynthesis with an antisense oligodeoxynucleotide (AS ODN) resulted in a diminished leptin effect on food intake and body weight. AS ODN reversed leptin's effect on the proopiomelanocortin (POMC) mRNA expression and, moreover, decreased leptin-induced STAT3 binding to the POMC promoter sequence. These results suggest that SPX is involved in leptin's action on POMC gene expression in the hypothalamus and impacts the anorexigenic effects of leptin.
Subject(s)
Leptin , Neuropeptides , Animals , Feeding Behavior , Hypothalamus/metabolism , Leptin/metabolism , Leptin/pharmacology , Mice , Neuropeptides/metabolism , Pro-Opiomelanocortin/genetics , Pro-Opiomelanocortin/metabolism , Pro-Opiomelanocortin/pharmacologyABSTRACT
AIM: To develop more effective and long-lasting antiobesity and antidiabetic therapeutics by employing novel chemical modifications of glucagon-like peptide-1 receptor (GLP-1R) agonists. METHODS: We constructed novel unimolecular dual agonists of GLP-1R and glucagon receptor prepared by linking sEx-4 and native glucagon (GCG) via lysine or triazole [sEx4-GCG(K) and sEx4-GCG(T), respectively] and evaluated their antiobesity and antidiabetic efficacy in the diabetic and obese mouse model. RESULTS: Both sEx4-GCG(K) and sEx4-GCG(T) showed the beneficial metabolic effects of GLP-1 and glucagon: they promoted weight loss and ameliorated insulin resistance and hepatic steatosis. They also increased thermogenesis in brown adipose tissue, and lipolysis and ß-oxidation in white adipose tissue, with concomitant suppression of lipogenesis. Furthermore, both dual agonists activated the 5'-AMP-activated protein kinase signalling pathway and prevented palmitate-induced oxidative stress in skeletal muscle cells. CONCLUSION: Through their complementary dual agonism, sEx4-GCG(T) and sEx4-GCG(K) induce more marked weight loss and metabolic improvements than conventional agonists, and could be developed as novel therapeutic agents for the treatment of obesity and associated metabolic disorders in humans.
Subject(s)
Glucagon-Like Peptide-1 Receptor , Glucagon , Animals , Glucagon/physiology , Glucagon-Like Peptide 1/metabolism , Glucagon-Like Peptide-1 Receptor/agonists , Humans , Mice , Receptors, Glucagon/metabolism , ThermogenesisABSTRACT
Neurogenesis and functional brain activity require complex associations of inherently programmed secretory elements that are regulated precisely and temporally. Family with sequence similarity 19 A1 (FAM19A1) is a secreted protein primarily expressed in subsets of terminally differentiated neuronal precursor cells and fully mature neurons in specific brain substructures. Several recent studies have demonstrated the importance of FAM19A1 in brain physiology; however, additional information is needed to support its role in neuronal maturation and function. In this study, dendritic spine morphology in Fam19a1-ablated mice and neurite development during in vitro neurogenesis were examined to understand the putative role of FAM19A1 in neural integrity. Adult Fam19a1-deficient mice showed low dendritic spine density and maturity with reduced dendrite complexity compared to wild-type (WT) littermates. To further explore the effect of FAM19A1 on neuronal maturation, the neurite outgrowth pattern in primary neurons was analyzed in vitro with and without FAM19A1. In response to FAM19A1, WT primary neurons showed reduced neurite complexity, whereas Fam19a1-decifient primary neurons exhibited increased neurite arborization, which was reversed by supplementation with recombinant FAM19A1. Together, these findings suggest that FAM19A1 participates in dendritic spine development and neurite arborization.
Subject(s)
Brain/metabolism , Chemokines/metabolism , Dendritic Spines/metabolism , Neurites/metabolism , Neuronal Outgrowth , Age Factors , Animals , Brain/pathology , Cells, Cultured , Chemokines/genetics , Dendritic Spines/pathology , Female , Gestational Age , Male , Mice, Inbred C57BL , Mice, Knockout , Neurites/pathology , Pregnancy , Signal TransductionABSTRACT
Family with sequence similarity 19 (chemokine (C-C motif)-like) member A5 (FAM19A5) is a chemokine-like secretory protein recently identified as involved in the regulation of osteoclast formation, post-injury neointima formation, and depression. Although roles for FAM19A5 have been described in nervous system development and psychiatric disorders, its role in the nervous system remains poorly understood. Here, we analyzed the evolutionary history of FAM19A genes in vertebrates and identified FAM19A5l, a paralogous zebrafish gene originating from a common ancestral FAM19A5 gene. Further, zebrafish FAM19A5l is expressed in trigeminal and dorsal root ganglion neurons as well as distinct neuronal subsets of the central nervous system. Interestingly, FAM19A5l+ trigeminal neurons are nociceptive neurons that localized with TRPA1b and TRPV1 and respond to mustard oil treatment. Behavioral analysis further revealed that the nociceptive response to mustard oil decreases in FAM19A5l-knockout zebrafish larvae. In addition, TRPA1b and NGFa mRNA levels are down- and upregulated in FAM19A5l-knockout and -overexpressing transgenic zebrafish, respectively. Together, our data suggest that FAM19A5l plays a role in nociceptive responses to mustard oil by regulating TRPA1b and NGFa expression in zebrafish.
Subject(s)
Cytokines/metabolism , Neurons/drug effects , Nociception/drug effects , Nociceptors/drug effects , Plant Oils/pharmacology , Animals , Animals, Genetically Modified , Cytokines/genetics , Mustard Plant , Neurons/metabolism , Nociception/physiology , Nociceptors/metabolism , ZebrafishABSTRACT
BACKGROUND: Some chemokine receptors referred to as atypical chemokine receptors (ACKRs) are thought to non-signaling decoys because of their inability to activate typical G-protein signaling pathways. CXCR7, also known as ACKR3, binds to only two chemokines, SDF-1α and I-TAC, and recruits ß-arrestins. SDF-1α also binds to its own conventional receptor, CXCR4, involving in homeostatic modulation such as development and immune surveillance as well as pathological conditions such as inflammation, ischemia, and cancers. Recently, CXCR7 is suggested as a key therapeutic target together with CXCR4 in such conditions. However, the molecular mechanisms underlying cellular responses and functional relation with CXCR7 and CXCR4 have not been elucidated, despite massive studies. Therefore, we aimed to reveal the molecular networks of CXCR7 and CXCR4 and compare their effects on cell migration. METHODS: Base on structural complementation assay using NanoBiT technology, we characterized the distinct mechanisms underlying ß-arrestin2 recruitment by both CXCR4 and CXCR7. Crosslinking and immunoprecipitation were conducted to analyze complex formation of the receptors. Gene deletion using CRISPR and reconstitution of the receptors were applied to analysis of ligand-dependent ERK phosphorylation and cell migration. All experiments were performed in triplicate and repeated more than three times. Unpaired Student's t-tests or ANOVA using PRISM5 software were employed for statistical analyses. RESULTS: Ligand binding to CXCR7 does not result in activation of typical signaling pathways via Gα subunits but activation of GRK2 via ßγ subunits and receptor phosphorylation with subsequent ß-arrestin2 recruitment. In contrast, CXCR4 induced Gαi activation and recruited ß-arrestin2 through C-terminal phosphorylation by both GRK2 and GRK5. SDF-1α-stimulated ERK phosphorylation was facilitated by CXCR4, but not CXCR7. Heterodimerization of CXCR4 and CXCR7 was not confirmed in this study, while homodimerization of them was verified by crosslinking experiment and NanoBiT assay. Regarding chemotaxis, SDF-1α-stimulated cell migration was mediated by both CXCR4 and CXCR7. CONCLUSION: This study demonstrates that SDF-1α-stimulated CXCR7 mediates ß-arrestin2 recruitment via different molecular networking from that of CXCR4. CXCR7 may be neither a simple scavenger nor auxiliary receptor but plays an essential role in cell migration through cooperation with CXCR4.
ABSTRACT
Peripheral nociceptors and their synaptic partners utilize neuropeptides for signal transmission. Such communication tunes the excitatory and inhibitory function of nociceptor-based circuits, eventually contributing to pain modulation. Corticotropin-releasing hormone (CRH) is the initiator hormone for the conventional hypothalamic-pituitary-adrenal axis, preparing our body for stress insults. Although knowledge of the expression and functional profiles of CRH and its receptors and the outcomes of their interactions has been actively accumulating for many brain regions, those for nociceptors are still under gradual investigation. Currently, based on the evidence of their expressions in nociceptors and their neighboring components, several hypotheses for possible pain modulations are emerging. Here we overview the historical attention to CRH and its receptors on the peripheral nociception and the recent increases in information regarding their roles in tuning pain signals. We also briefly contemplate the possibility that the stress-response paradigm can be locally intrapolated into intercellular communication that is driven by nociceptor neurons. Such endeavors may contribute to a more precise view of local peptidergic mechanisms of peripheral pain modulation.
ABSTRACT
Cytosolic Ca2+ levels ([Ca2+]c) change dynamically in response to inducers, repressors, and physiological conditions, and aberrant [Ca2+]c concentration regulation is associated with cancer, heart failure, and diabetes. Therefore, [Ca2+]c is considered as a good indicator of physiological and pathological cellular responses, and is a crucial biomarker for drug discovery. A genetically encoded calcium indicator (GECI) was recently developed to measure [Ca2+]c in single cells and animal models. GECI have some advantages over chemically synthesized indicators, although they also have some drawbacks such as poor signal-to-noise ratio (SNR), low positive signal, delayed response, artifactual responses due to protein overexpression, and expensive detection equipment. Here, we developed an indicator based on interactions between Ca2+-loaded calmodulin and target proteins, and generated an innovative GECI sensor using split nano-luciferase (Nluc) fragments to detect changes in [Ca2+]c. Stimulation-dependent luciferase activities were optimized by combining large and small subunits of Nluc binary technology (NanoBiT, LgBiT:SmBiT) fusion proteins and regulating the receptor expression levels. We constructed the binary [Ca2+]c sensors using a multicistronic expression system in a single vector linked via the internal ribosome entry site (IRES), and examined the detection efficiencies. Promoter optimization studies indicated that promoter-dependent protein expression levels were crucial to optimize SNR and sensitivity. This novel [Ca2+]c assay has high SNR and sensitivity, is easy to use, suitable for high-throughput assays, and may be useful to detect [Ca2+]c in single cells and animal models.
Subject(s)
Calmodulin/metabolism , Cytosol/metabolism , Proteins/metabolism , HEK293 Cells , HumansABSTRACT
Galanin receptors (GALRs) belong to the superfamily of G-protein coupled receptors. The three GALR subtypes (GALR1, GALR2, and GALR3) are activated by their endogenous ligands: spexin (SPX) and galanin (GAL). The synthetic SPX-based GALR2-specific agonist, SG2A, plays a dual role in the regulation of appetite and depression-like behaviors. Little is known, however, about the molecular interaction between GALR2 and SG2A. Using site-directed mutagenesis and domain swapping between GALR2 and GALR3, we identified residues in GALR2 that promote interaction with SG2A and residues in GALR3 that inhibit interaction with SG2A. In particular, Phe103, Phe106, and His110 in the transmembrane helix 3 (TM3) domain; Val193, Phe194, and Ser195 in the TM5 domain; and Leu273 in the extracellular loop 3 (ECL3) domain of GALR2 provide favorable interactions with the Asn5, Ala7, Phe11, and Pro13 residues of SG2A. Our results explain how SG2A achieves selective interaction with GALR2 and inhibits interaction with GALR3. The results described here can be used broadly for in silico virtual screening of small molecules for the development of GALR subtype-specific agonists and/or antagonists.
Subject(s)
Receptor, Galanin, Type 2/chemistry , Receptor, Galanin, Type 2/metabolism , Receptor, Galanin, Type 3/chemistry , Receptor, Galanin, Type 3/metabolism , Amino Acid Sequence , Animals , HEK293 Cells , Humans , Ligands , Mice , Mutation , Protein Domains , Receptor, Galanin, Type 3/genetics , Substrate SpecificityABSTRACT
Chronic low-grade inflammation contributes to the pathophysiology of major depressive disorder (MDD). This study aimed to examine the association between serum levels of FAM19A5, a novel chemokine-like peptide that reflects reactive astrogliosis and inflammatory activation in the brain, and the neurodegenerative changes of MDD by investigating the correlation between serum FAM19A5 levels and cortical thickness changes in patients with MDD. We included 52 drug-naïve patients with MDD and 60 healthy controls (HCs). Serum FAM19A5 levels were determined in peripheral venous blood samples using a sandwich enzyme-linked immunosorbent assay. All participants underwent T1-weighted structural magnetic resonance imaging. Serum FAM19A5 levels were greater in patients with MDD than in HCs. In the MDD group, there were significant inverse correlations between serum FAM19A5 levels and cortical thickness in the prefrontal regions (i.e., the left inferior and right medial superior frontal gyri), left posterior cingulate gyrus, right cuneus, and both precunei, which showed significantly reduced thickness in patients with MDD compared to HCs. However, no correlation between serum FAM19A5 level and cortical thickness was observed in the HC group. The results of our study indicate that serum FAM19A5 levels may reflect reactive astrogliosis and related neuroinflammation in MDD. Our findings also suggest that serum FAM19A5 may be a potential biomarker for the neurodegenerative changes of MDD.
Subject(s)
Depressive Disorder, Major , Biomarkers , Brain , Cerebral Cortex , Humans , Magnetic Resonance ImagingABSTRACT
Neurodevelopment and mature brain function are spatiotemporally regulated by various cytokines and chemokines. The chemokine-like neuropeptide FAM19A1 is a member of family with sequence similarity 19 (FAM19), which is predominantly expressed in the brain. Its highly conserved amino acid sequence among vertebrates suggests that FAM19A1 may play important physiological roles in neurodevelopment and brain function. Here we used a LacZ reporter gene system to map the expression pattern of the FAM19A1 gene in the mouse brain. The FAM19A1 expression was observed in several brain regions starting during embryonic brain development. As the brain matured, the FAM19A1 expression was detected in the pyramidal cells of cortical layers 2/3 and 5 and in several limbic areas, including the hippocampus and the amygdala. FAM19A1-deficient mice were used to evaluate the physiological contribution of FAM19A1 to various brain functions. In behavior analysis, FAM19A1-deficient mice exhibited several abnormal behaviors, including hyperactive locomotor behavior, long-term memory deficits and fear acquisition failure. These findings provide insight into the potential contributions of FAM19A1 to neurodevelopment and mature brain function.
Subject(s)
Behavior, Animal , Brain/metabolism , Chemokines/physiology , Conditioning, Psychological , Fear/physiology , Hyperkinesis/physiopathology , Memory, Long-Term/physiology , Amygdala/metabolism , Animals , Female , Hippocampus/metabolism , Male , Mice , Mice, KnockoutABSTRACT
The cytokine-like protein FAM19A5 is highly expressed in the brain, but little is known about its functions there. Here, we found that FAM19A5 was expressed in mouse hypothalamic cells expressing proopiomelanocortin (POMC) and neuropeptide Y (NPY)/agouti-related peptide (AgRP), and in the microglia. Tumor necrosis factor-α (TNF-α), which induces inflammatory sickness responses, greatly increased hypothalamic expression of FAM19A5. Knockdown of FAM19A5 expression resulted in decreased TNF-α-induced anorexia, body weight loss and TNF-α-induced expression of inflammatory factors. In contrast, intracerebroventricular administration of FAM19A5 induced anorexia, body weight loss and hyperthermia, together with increased expression of inflammatory factors. FAM19A5 injection also induced increases in c-fos activation and POMC mRNA level in hypothalamic POMC neurons. Together, these results suggest that FAM19A5 plays an important role in hypothalamic inflammatory responses.
Subject(s)
Cytokines/metabolism , Hypothalamus/metabolism , Hypothalamus/pathology , Inflammation/metabolism , Animals , Cytokines/administration & dosage , Cytokines/pharmacology , Humans , Hypothalamus/drug effects , Male , Mice, Inbred C57BL , Neurons/drug effects , Neurons/metabolism , Organ Specificity/drug effects , Pro-Opiomelanocortin/metabolism , Proto-Oncogene Proteins c-fos/metabolism , Tumor Necrosis Factor-alpha/administration & dosageABSTRACT
BACKGROUND: Neuromyelitis optica spectrum disorder (NMOSD) targets astrocytes and elevates the levels of astrocyte-injury markers during attacks. FAM19A5, involved in reactive gliosis, is secreted by reactive astrocytes following central nervous system (CNS) damage. OBJECTIVE: To investigate the significance of serum FAM19A5 in patients with NMOSD. METHODS: We collected clinical data and sera of 199 patients from 11 hospitals over 21 months. FAM19A5 levels were compared among three groups: NMOSD with positive anti-aquaporin-4 antibody (NMOSD-AQP4), other CNS demyelinating disease, and healthy controls. RESULTS: The median serum FAM19A5 level was higher in the NMOSD-AQP4 (4.90 ng/mL (3.95, 5.79)) than in the other CNS demyelinating (2.35 ng/mL (1.83, 4.07), p < 0.001) or healthy control (1.02 ng/mL (0.92, 1.14), p < 0.001) groups. There were significant differences in the median serum FAM19A5 levels between the attack and remission periods (5.89 ng/mL (5.18, 6.98); 4.40 ng/mL (2.72, 5.13), p < 0.001) in the NMOSD-AQP4 group. Sampling during an attack (p < 0.001) and number of past attacks (p = 0.010) were independently associated with increased serum FAM19A5. CONCLUSION: Serum FAM19A5 was higher in patients with NMOSD-AQP4 and correlated with clinical characteristics. Thus, serum FAM19A5 may be a novel clinical biomarker for NMOSD-AQP4.
Subject(s)
Neuromyelitis Optica , Aquaporin 4 , Autoantibodies , Biomarkers , Humans , Myelin-Oligodendrocyte Glycoprotein , Neuromyelitis Optica/diagnosisABSTRACT
Spexin (SPX) is an evolutionarily conserved neuropeptide that is expressed in the mammalian brain and peripheral tissue. Two orthologs are present in the teleost, SPX1 and SPX2. SPX1 is involved in reproduction and food intake. Recently, SPX1 neurons have been found to be located in the specific nuclei of dorsal habenula (dHb) and to project into the interpeduncular nucleus (IPN), in which galanin receptor 2a/2b (GALR2a/2b) expression was also observed. This indicates that habenula SPX1 neurons may interact with GALR2a/2b in the IPN; however, the function of SPX1 in the dHb-IPN neuronal circuit remains unknown. To determine the role of SPX1 in the dHb-IPN neural circuit, we generated transgenic zebrafish overexpressing SPX1 specifically in the dHb. We found that transgenic zebrafish overexpressing SPX1 in the dHb had anxiolytic behaviors compared with their wildtype siblings. Furthermore, quantitative PCR revealed that mRNA expression of galr2a and galr2b in the IPN and serotonin-related genes in the raphe was upregulated in the brains of transgenic zebrafish. Taken together, our data suggest that SPX1 function in the dHb-IPN neural circuits is implicated in the regulation of anxiety behaviors via modulation of the serotoninergic system in zebrafish.