Chronic noise exposure induces Alzheimer's disease-like neuropathology and cognitive impairment via ferroptosis in rat hippocampus.

BACKGROUND: Chronic noise exposure poses a remarkable public health concern, drawing attention to its impacts on the brain. Ferroptosis is involved in several brain-related diseases. However, the role of ferroptosis in the effects of chronic noise on the brain remains elusive. This study aimed to investigate the effects of chronic noise exposure on the brain and elucidate the underlying mechanisms. METHODS: A chronic noise-induced cognitive impairment model in rats was constructed and validated. The pathological state and ferroptosis level of the rat hippocampus were determined using Western blotting and immunohistochemistry. Bioinformatics was employed to investigate the interrelationship between chronic noise exposure and genes. Genetic relationships were analyzed using Mendelian randomization (MR) analysis. Cytoscape was employed for the prediction of upstream molecular and drug targets. RESULTS: In vivo experiments revealed that chronic noise exposure could induce Alzheimer's disease (AD)-like neuropathological changes in rat hippocampus and cognitive impairment. Moreover, protein markers indicative of ferroptosis and levels of lipid peroxidation were quantified to elucidate underlying mechanisms. Thereafter, oxidative stress- and ferroptosis-related differentially expressed genes (DEGs) underwent functional enrichment and PPI network analyses. Additionally, 8 genes with diagnostic significance were identified. In MR analysis, retinoic acid receptor responder 2 (Rarres2) gene exhibited a negative genetic relationship with AD. CONCLUSIONS: Chronic noise exposure could induce AD-like neuropathological changes and cognitive impairment via ferroptosis. The results of MR analysis indicated that Rarres2 gene may act as a protective factor in AD. This gene may be upstream of ferroptosis and serve as a target for the prevention and treatment of chronic noise-induced cognitive impairment.

Targeting copper death genotyping associated gene RARRES2 suppresses glioblastoma progression and macrophages infiltration.

BACKGROUND: Copper homeostasis is associated with malignant biological behavior in various tumors. The excessive accumulation of copper can induce tumor death, which is named cuproptosis, and it is also closely related to tumor progression and the formation of the immune microenvironment. However, the associations of cuproptosis with glioblastoma (GBM) prognosis and microenvironment construction are poorly understood. METHOD: First, TCGA and GEO (GSE83300, GSE74187) merged datasets were used to analyze the association of cuproptosis-related genes (CRGs) with GBM. Then, we performed cluster analysis of CRGs in GBM from the GEO (GSE83300, GSE74187) and TCGA merged datasets. Subsequently, the prognostic risk model was constructed by least absolute shrinkage and selection operator (LASSO) according to gene expression features in CRG clusters. Next, we performed a series of in-depth analyses, including tumor mutational burden (TMB) analysis, cluster analysis, and GBM IDH status prediction. Finally, RARRES2 was identified as a target gene for GBM treatment, especially IDH wild-type GBM. In addition, we further analyzed the correlation of CRG clusters and RARRES2 expression with the GBM immune microenvironment by ESTIMATE and CIBERSORT analyses. In vitro experiments were conducted to demonstrate that targeting RARRES2 inhibits glioblastoma progression and macrophage infiltration, particularly IDH wild-type GBM. RESULTS: In the present study, we demonstrated that the CRG cluster was closely related to GBM prognosis and immune cell infiltration. Moreover, the prognostic risk model constructed with the three genes (MMP19, G0S2, RARRES2) associated with the CRG clusters could well evaluate the prognosis and immune cell infiltration in GBM. Subsequently, after further analyzing the tumor mutational burden (TMB) in GBM, we confirmed that RARRES2 in the prognostic risk model could be used as a crucial gene signature to predict the prognosis, immune cell infiltration and IDH status of GBM patients. CONCLUSION: This study fully revealed the potential clinical impact of CRGs on GBM prognosis and the microenvironment, and determined the effect of the crucial gene (RARRES2) on the prognosis and tumor microenvironment construction of GBM, meanwhile, our study also revealed over-expressed RARRES2 is related to the IDH satus of GBM, which provides a novel strategy for the treatment of GBM, particularly IDH wild-type GBM.

Competitive Hybridization of a Microarray Identifies CMKLR1 as an Up-Regulated Gene in Human Bone Marrow-Derived Mesenchymal Stem Cells Compared to Human Embryonic Fibroblasts.

Mesenchymal stem cells (MSCs) have been widely applied to the regeneration of damaged tissue and the modulation of immune response. The purity of MSC preparation and the delivery of MSCs to a target region are critical factors for success in therapeutic application. In order to define the molecular identity of an MSC, the gene expression pattern of a human bone marrow-derived mesenchymal stem cell (hBMSC) was compared with that of a human embryonic fibroblast (hEF) by competitive hybridization of a microarray. A total of 270 and 173 genes were two-fold up- and down-regulated with FDR < 0.05 in the hBMSC compared to the hEF, respectively. The overexpressed genes in the hBMSC over the hEF, including transcription factors, were enriched for biological processes such as axial pattern formation, face morphogenesis and skeletal system development, which could be expected from the differentiation potential of MSCs. CD70 and CD339 were identified as additional CD markers that were up-regulated in the hBMSC over the hEF. The differential expression of CD70 and CD339 might be exploited to distinguish hEF and hBMSC. CMKLR1, a chemokine receptor, was up-regulated in the hBMSC compared to the hEF. RARRES2, a CMKLR1 ligand, stimulated specific migration of the hBMSC, but not of the hEF. RARRES2 manifested as ~two-fold less effective than SDF-1α in the directional migration of the hBMSC. The expression of CMKLR1 was decreased upon the osteoblastic differentiation of the hBMSC. However, the RARRES2-loaded 10% HA-silk scaffold did not recruit endogenous cells to the scaffold in vivo. The RARRES2−CMKLR1 axis could be employed in recruiting systemically delivered or endogenous MSCs to a specific target lesion.

Possible involvement of the RARRES2/CMKLR1-system in metabolic and reproductive parameters in Holstein dairy cows.

BACKGROUND: In dairy cows, the energy cost of milk yield results in a negative energy balance (EB) and body fat mobilization that impairs reproductive efficiency. Emerging evidence suggests that the novel adipokines, Retinoic acid receptor responder protein 2 (RARRES2), and its main receptor, Chemokine-like receptor 1 (CMKLR1) are involved in the regulation of metabolic and ovarian functions. So, we investigated in a first experiment the plasma RARRES2, and RARRES2 and CMKLR1 mRNA expression levels in subcutaneous adipose tissue (SAT) and granulosa cells (GC) at different times of body fat mobilization in dairy cows (4, 8, 20 and 44 weeks postpartum, wk. pp. for SAT and 8, 20 and 44 wk. pp. for GC). Then, in a second experiment we examined the effect of high (HE) and low energy (LE) diets on the RARRES2 system and its links with metabolic and reproductive parameters. METHODS: The first experiment included 9 animals fed with HE diet from 4 to 44 wk. pp. and the second one included animals fed either a HE diet (n = 8) or a LE diet (n = 8) from - 4 to 16 wk. peripartum. In both experiments, various metabolic and reproductive parameters were determined and associated with plasma RARRES2 as measured by bovine ELISA. RARRES2 and CMKLR1 mRNA expression levels were analyzed by RT-qPCR in SAT after biopsy and GC after aspiration of follicles. RESULTS: Plasma RARRES2 levels were higher at 4 wk. pp. as compared to 20 and 44 wk. pp. and they were positively correlated with body fat mobilization and milk yield. RARRES2 and CMKLR1 mRNA expression levels increased from 4 to 8 wk. pp. (fat mobilization, EB < 0) and remained unchanged at 20 and 44 wk. pp. (fat reconstitution, EB > 0) as compared to 4 wk. pp. in SAT. RARRES2 and CMKLR1 mRNA levels decreased from 8 to 44 wk. pp. in GC from small follicles. In the second experiment, plasma RARRES2 increased from - 4 to 8 wk. peripartum similarly in both LE and HE cows. In addition, the area under of plasma RARRES2 curve was highly negatively associated with the number of small follicles obtained in HE animals during the cycle before the first artificial insemination. In SAT of HE cows, RARRES2 mRNA expression decreased at 1 wk. pp. compared to - 4 and 16 wk. peripartum whereas opposite expression patterns were obtained for CMKLR1. Similar results were observed for CMKLR1 mRNA expression in LE cows while there was no variation in RARRES2 mRNA expression. Moreover, RARRES2 mRNA was higher expressed in LE than in HE cows at 1 wk. pp. CONCLUSIONS: The lactation-induced fat and energy mobilization influenced plasma RARRES2 profile and mRNA expression pattern of RARRES2 and CMKLR1 similarly in both SAT and GC. In addition, the energy content of the diet did not affect plasma RARRES2 but it altered RARRES2 mRNA expression in SAT and the area under the curve of plasma RARRES2 that was negatively associated to the number of small follicles in HE animals. Thus, RARRES2 could be a metabolic or ovarian signal involved in the interactions between metabolic and reproductive functions in dairy cows.

Circulating Chemerin Levels, but not the RARRES2 Polymorphisms, Predict the Long-Term Outcome of Angiographically Confirmed Coronary Artery Disease.

Chemerin, a novel adipokine, has been associated with metabolic, inflammatory, and atherosclerotic diseases. We aimed to determine the genetic basis of chemerin levels by conducting a genome-wide association study (GWAS) and to investigate the role of RARRES2 polymorphisms and circulating chemerin levels in the long-term outcome of coronary artery disease (CAD). A total of 2197 participants from the Taiwan Biobank (TWB) were recruited for the GWAS analysis, and 481 patients with angiographically confirmed CAD were enrolled for long-term outcome analysis. One locus of genome-wide significance with a single independent association signal was identified in the GWAS for chemerin levels with the peak association at the RARRES2 gene promoter region polymorphism rs3735167 (p = 2.35 × 10-21). In the CAD population, borderline significance was noted between RARRES2 polymorphisms and chemerin levels, whereas high chemerin levels were associated with obesity, female sex, diabetes mellitus, hypertension, current smoking, high platelet and leukocyte counts, anemia, impaired renal function, high C-reactive protein (CRP) levels, and multi-vessel disease. Kaplan⁻Meier survival curves indicated that the patients with high chemerin and CRP levels, but not those with RARRES2 polymorphisms, had a lower survival rate and higher combined cerebral and cardiovascular event rates. Combined chemerin and CRP levels further revealed a stepwise increase in poor clinical outcomes from low- to high-risk subgroups. In conclusion, rs3735167 is the lead RARRES2 polymorphism for chemerin levels in Taiwanese. Chemerin levels, but not the rs3735167 genotypes, predicted the long-term outcome of CAD, especially when combined with CRP levels.

Peptidase inhibitor 16 is a membrane-tethered regulator of chemerin processing in the myocardium.

A key response of the myocardium to stress is the secretion of factors with paracrine or endocrine function. Intriguing in this respect is peptidase inhibitor 16 (PI16), a member of the CAP family of proteins which we found to be highly upregulated in cardiac disease. Up to this point, the mechanism of action and physiological function of PI16 remained elusive. Here, we show that PI16 is predominantly expressed by cardiac fibroblasts, which expose PI16 to the interstitium via a glycophosphatidylinositol (-GPI) membrane anchor. Based on a reported genetic association of PI16 and plasma levels of the chemokine chemerin, we investigated whether PI16 regulates post-translational processing of its precursor pro-chemerin. PI16-deficient mice were engineered and found to generate higher levels of processed chemerin than wildtype mice. Purified recombinant PI16 efficiently inhibited cathepsin K, a chemerin-activating protease, in vitro. Moreover, we show that conditioned medium from PI16-overexpressing cells impaired the activation of pro-chemerin. Together, our data indicate that PI16 suppresses chemerin activation in the myocardium and suggest that this circuit may be part of the cardiac stress response.

High-resolution molecular validation of self-renewal and spontaneous differentiation in clinical-grade adipose-tissue derived human mesenchymal stem cells.

Improving the effectiveness of adipose-tissue derived human mesenchymal stromal/stem cells (AMSCs) for skeletal therapies requires a detailed characterization of mechanisms supporting cell proliferation and multi-potency. We investigated the molecular phenotype of AMSCs that were either actively proliferating in platelet lysate or in a basal non-proliferative state. Flow cytometry combined with high-throughput RNA sequencing (RNASeq) and RT-qPCR analyses validate that AMSCs express classic mesenchymal cell surface markers (e.g., CD44, CD73/NT5E, CD90/THY1, and CD105/ENG). Expression of CD90 is selectively elevated at confluence. Self-renewing AMSCs express a standard cell cycle program that successively mediates DNA replication, chromatin packaging, cyto-architectural enlargement, and mitotic division. Confluent AMSCs preferentially express genes involved in extracellular matrix (ECM) formation and cellular communication. For example, cell cycle-related biomarkers (e.g., cyclins E2 and B2, transcription factor E2F1) and histone-related genes (e.g., H4, HINFP, NPAT) are elevated in proliferating AMSCs, while ECM genes are strongly upregulated (>10-fold) in quiescent AMSCs. AMSCs also express pluripotency genes (e.g., POU5F1, NANOG, KLF4) and early mesenchymal markers (e.g., NES, ACTA2) consistent with their multipotent phenotype. Strikingly, AMSCs modulate expression of WNT signaling components and switch production of WNT ligands (from WNT5A/WNT5B/WNT7B to WNT2/WNT2B), while upregulating WNT-related genes (WISP2, SFRP2, and SFRP4). Furthermore, post-proliferative AMSCs spontaneously express fibroblastic, osteogenic, chondrogenic, and adipogenic biomarkers when maintained in confluent cultures. Our findings validate the biological properties of self-renewing and multi-potent AMSCs by providing high-resolution quality control data that support their clinical versatility.

The Dual Role of Chemerin in Lung Diseases.

Chemerin is an atypical chemokine first described as a chemoattractant agent for monocytes, natural killer cells, plasmacytoid and myeloid dendritic cells, through interaction with its main receptor, the G protein-coupled receptor chemokine-like receptor 1 (CMKLR1). Chemerin has been studied in various lung disease models, showing both pro- and anti-inflammatory properties. Given the incidence and burden of inflammatory lung diseases from diverse origins (infectious, autoimmune, age-related, etc.), chemerin has emerged as an interesting therapeutical target due to its immunomodulatory role. However, as highlighted by this review, further research efforts to elucidate the mechanisms governing chemerin's dual pro- and anti-inflammatory characteristics are urgently needed. Moreover, although a growing body of evidence suggests chemerin as a potential biomarker for the diagnosis and/or prognosis of inflammatory lung diseases, this review underscores the necessity for standardizing both sampling types and measurement techniques before drawing definitive conclusions.

RARRES2 regulates lipid metabolic reprogramming to mediate the development of brain metastasis in triple negative breast cancer.

BACKGROUND: Triple negative breast cancer (TNBC), the most aggressive subtype of breast cancer, is characterized by a high incidence of brain metastasis (BrM) and a poor prognosis. As the most lethal form of breast cancer, BrM remains a major clinical challenge due to its rising incidence and lack of effective treatment strategies. Recent evidence suggested a potential role of lipid metabolic reprogramming in breast cancer brain metastasis (BCBrM), but the underlying mechanisms are far from being fully elucidated. METHODS: Through analysis of BCBrM transcriptome data from mice and patients, and immunohistochemical validation on patient tissues, we identified and verified the specific down-regulation of retinoic acid receptor responder 2 (RARRES2), a multifunctional adipokine and chemokine, in BrM of TNBC. We investigated the effect of aberrant RARRES2 expression of BrM in both in vitro and in vivo studies. Key signaling pathway components were evaluated using multi-omics approaches. Lipidomics were performed to elucidate the regulation of lipid metabolic reprogramming of RARRES2. RESULTS: We found that down-regulation of RARRES2 is specifically associated with BCBrM, and that RARRES2 deficiency promoted BCBrM through lipid metabolic reprogramming. Mechanistically, reduced expression of RARRES2 in brain metastatic potential TNBC cells resulted in increased levels of glycerophospholipid and decreased levels of triacylglycerols by regulating phosphatase and tensin homologue (PTEN)-mammalian target of rapamycin (mTOR)-sterol regulatory element-binding protein 1 (SREBP1) signaling pathway to facilitate the survival of breast cancer cells in the unique brain microenvironment. CONCLUSIONS: Our work uncovers an essential role of RARRES2 in linking lipid metabolic reprogramming and the development of BrM. RARRES2-dependent metabolic functions may serve as potential biomarkers or therapeutic targets for BCBrM.

RARRES2 is Downregulated in Isocitrate Dehydrogenase 1 Mutant Glioma Patients and Served as an Unfavorable Prognostic Factor of Glioma.

BACKGROUND: Mutations in isocitrate dehydrogenase 1 (IDH1) induce extensive transcriptional alterations to promote glioma development. However, IDH1 mutation contributes the better clinical outcomes of glioma. Further understanding the transcriptional and DNA methylation changes mediated by IDH1 mutation will provide new therapeutic targets for glioma. METHODS: Public glioma cohorts were collected and processed using R software. The transcriptional changes mediated by IDH1 mutation were determined and presented using heatmap. The differentially expressed genes in IDH1 mutant glioma were overlapped using TBtools. The prognostic effects of IDH1 regulated genes were determined by Kaplan-Meier survival analysis. RESULTS: Retinoic acid receptor responder 2 (RARRES2) was upregulated in IDH1 wild type lower-grade glioma (LGG) patients, and higher expression levels of RARRES2 were associated with worse clinical outcomes of LGG. Moreover, IDH1 wild type LGG patients with higher expression levels of RARRES2 had even worse overall survival. Compared with LGG, RARRES2 was upregulated in grade IV glioma (glioblastoma multiforme, GBM). Also, RARRES2 represented an unfavorable prognostic factor of glioma. In GBM, RARRES2 was also associated with IDH1 mutation. In both LGG and GBM, IDH1 mutation induced extensive DNA hypermethylation, and more than half genes that were downregulated in IDH1 mutant glioma were contributed by DNA hypermethylation. RARRES2 was also hypermethylated in IDH1 mutant LGG or GBM patients. Furthermore, RARRES2 hypomethylation was an unfavorable prognostic factor in patients with LGG. CONCLUSIONS: RARRES2 was downregulated by IDH1 mutation and served as an unfavorable prognostic factor in glioma.

Association of serum levels of Visfatin, Intelectin-1, RARRES2 and their genetic variants with bone mineral density in postmenopausal females.

Background: Adipokines are engaged in bone physiology and regulate bone mineral density (BMD) by playing protective or cynical role in bone metabolism. The study is designed to measure and compare BMD, adipokines (retinoic acid receptor responder protein-2 RARRES2, visfatin and Intelectin-1) and their genetic variants in postmenopausal osteoporotic, osteopenic and non-osteoporotic females. Methods: This comparative study included postmenopausal non-osteoporotic (n=72), osteopenic (n=72) and osteoporotic (n=100) females with two years of amenorrhea and age between 50 to 70 years. Gold standard DXA was used to measure BMD. Hardy-Weinberg equilibrium was established. Kruskal-Wallis test for comparisons, logistic and multivariate regression analysis were used to rule out the predictors of BMD. Results: On comparing the three groups, significant differences were observed in serum RARRES2 (p <0.001) and serum visfatin (p=0.050). The significant positive predictor of BMD at lumbar spine and total hip was serum visfatin. BMD at right and left femoral neck was predicted negatively by serum chemerin while BMD at left femoral neck was also predicted positively by serum calcium levels. There was significant difference in BMD at right femoral neck (p = 0.033) between rs7806429 genotypes. The odds of having low BMD increases with increasing serum levels of chemerin and decreasing serum levels of visfatin and calcium. Conclusion: The adipokines RARRES2 and visfatin are associated with BMD. RARRES2 is an independent negative and visfatin is positive predictor of BMD in postmenopausal females. BMD at right femoral neck was significantly low in RARRES2 rs7806429 TC heterozygotes.

Common variants of RARRES2 and RETN contribute to susceptibility to hand osteoarthritis and related pain.

Aim: To examine the relationship of the RETN and RARRES2 genes with hand osteoarthritis (HOA) susceptibility risk, clinical severity and pain. Methods: A total of 3740 subjects comprising 1180 participants with HOA and 2560 controls were enrolled. Genetic association was evaluated at both single marker and haplotype levels using PLINK. Results: Two significant hits, single-nucleotide polymorphism (SNP) rs4721 from RARRES2 and SNP rs3745368 from RETN, were identified as being related to an increased risk of HOA. Significant associations were obtained for SNP rs3745368 with Kellgren-Lawrence grade in HOA patients and SNP rs4721 with pain analog scales of HOA patients. Conclusion: The authors' results indicate that RARRES2 and RETN affect HOA risk and are associated with clinical features and severity in patients with HOA.

Chemerin levels and its genetic variants are associated with Gestational diabetes mellitus: A hospital-based study in a Chinese cohort.

BACKGROUND: Gestational diabetes mellitus (GDM) is a glucose intolerance condition encounters for the first time in a fraction of pregnant women. The role of different host inflammatory molecules in GDM etiology has been deciphered. Chemerin is a chemoattractant protein primarily associated with the pathogenesis of type 2 diabetes, obesity, and metabolic syndrome. However, the association of chemerin and its genetic variants with the predisposition of GDM is not clear, and our present study is aimed to address the issue. MATERIALS AND METHODS: A total of 703 Chinese women comprising of GDM (n = 303), glucose tolerant pregnant women (n = 211), and non-pregnant glucose tolerant controls (n = 189) were recruited in the present investigation. GDM was diagnosed according to the World Health Organization recommendation for diagnosis of gestational diabetes during pregnancy. Plasma levels of chemerin were quantified by an Enzyme-linked Immunosorbent Assay (ELISA). Common variants in the chemerin gene (rs4721, rs17173617, rs7806429, and rs17173608) were genotyped by using TaqMan assay. RESULTS: Plasma chemerin level was found higher in subjects with GDM as compared to glucose tolerant pregnant and non-pregnant women. Further, a positive correlation between plasma chemerin and HOMA-IR index suggesting an essential role of chemerin in mediating insulin resistance. Variants of rs4721 and rs17173608 polymorphisms were associated with lower levels of plasma chemerin and low HOMA-IR index. Furthermore, mutants of rs4721 and rs17173608 polymorphisms were associated with protection against the development of GDM in the Chinese cohort. CONCLUSIONS: Plasma chemerin is elevated in GDM patients. Genetic variation in chemerin gene associated with lower plasma levels of chemerin, HOMA-IR index and protects against the development of GDM in Chinese.

Hypertension is associated with a variant in the RARRES2 gene in populations of Ouro Preto, Minas Gerais, Brazil: a cross-sectional study.

BACKGROUND: Arterial hypertension (AH) is implicated in vascular health and contributes significantly to cardiovascular morbidity and mortality. In addition to the contribution of usual risk factors for AH, elucidating the influence of genetic factors is a promising area of investigation. Therefore, we evaluated the association between AH and cardiovascular risk factors (CVRFs) and genetic polymorphisms in communities in Southeast Brazil. METHODS: A total of 515 adults aged 18-91 years, who were cross-sectionally assessed between 2015-2016, were included. Demographic, clinical, behavioral, anthropometric characteristics, and laboratory parameters and 12 single nucleotide polymorphisms in seven candidate genes involved in cardiovascular risk (RARRES2, AGT, NOS3, GNB3, APOE, APOB, APOC3, LDLR, and PPARG) were evaluated, with AH as the outcome. Sex, age, and laboratory parameters were considered the main confounding factors. RESULTS: There was a significant association between age >60 years (odds ratio [OR] =6.74), alcohol dependence (OR=3.84), smoking (OR=1.74), overweight (OR=1.74), high plasma triglyceride (TG) levels (OR=1.98) and low high-density lipoprotein (HDL-c) (OR=6.22), diabetes (OR=3.68), and insulin resistance (OR=2.40) and AH. A significant association was observed between rs4721 in RARRES2 and AH. The T allele in homozygosis was a potent chance modifier for AH. The highest chance gradients for AH were characterized by the presence of the TT genotype and DMT2 (OR=9.70), high TG (OR=6.26), low HDL-c (OR=8.20), and age more than 60 years (OR=9.96). CONCLUSION: The interaction of the T allele of the rs4721 polymorphism in RARRES2 with CVRFs may predispose carriers to a higher cardiovascular risk.

Expression of CCRL2 Inhibits Tumor Growth by Concentrating Chemerin and Inhibiting Neoangiogenesis.

CCRL2 belongs to the G protein-coupled receptor family and is one of the three chemerin receptors. It is considered as a non-signaling receptor, presenting chemerin to cells expressing the functional chemerin receptor ChemR23/CMKLR1 and possibly GPR1. In the present work, we investigate the role played by CCRL2 in mouse cancer models. Loss of function of Ccrl2 accelerated the development of papillomas in a chemical model of skin carcinogenesis (DMBA/TPA), whereas the growth of B16 and LLC tumor cell grafts was delayed. Delayed tumor growth was also observed when B16 and LLC cells overexpress CCRL2, while knockout of Ccrl2 in tumor cells reversed the consequences of Ccrl2 knockout in the host. The phenotypes associated with CCRL2 gain or loss of function were largely abrogated by knocking out the chemerin or Cmklr1 genes. Cells harboring CCRL2 could concentrate bioactive chemerin and promote the activation of CMKLR1-expressing cells. A reduction of neoangiogenesis was observed in tumor grafts expressing CCRL2, mimicking the phenotype of chemerin-expressing tumors. This study demonstrates that CCRL2 shares functional similarities with the family of atypical chemokine receptors (ACKRs). Its expression by tumor cells can significantly tune the effects of the chemerin/CMKLR1 system and act as a negative regulator of tumorigenesis.

The antitumoral effects of chemerin are independent from leukocyte recruitment and mediated by inhibition of neoangiogenesis.

Chemerin, a multifunctional protein acting through the receptor ChemR23/CMKLR1, is downregulated in various human tumors and was shown to display antitumoral properties in mouse models of cancer. In the present study, we report that bioactive chemerin expression by tumor cells delays the growth of B16 melanoma and Lewis lung carcinoma in vivo. A similar delay is observed when chemerin is not expressed by tumor cells but by keratinocytes of the host mice. The protective effect of chemerin is mediated by CMKLR1 and appears unrelated to the recruitment of leukocyte populations. Rather, tumors grown in the presence of chemerin display a much smaller number of blood vessels, hypoxic regions early in their development, and larger necrotic areas. These observations likely explain the slower growth of the tumors. The anti-angiogenic effects of chemerin were confirmed in a bead sprouting assay using human umbilical vein endothelial cells. These results suggest that CMKLR1 agonists might constitute therapeutic molecules inhibiting the neoangiogenesis process in solid tumors.

Single Nucleotide Polymorphism rs17173608 in the Chemerin Encoding Gene: Is It a Predictor of Insulin Resistance and Severity of Coronary Artery Disease in Non-Obese Type 2 Diabetes?

(1) Background: Chemerin, or the RARRES2 (Retinoic Acid Receptor Responder 2) gene, is found to be associated with an increased incidence of insulin resistance, endothelial dysfunction, type 2 diabetes (T2D), and coronary artery disease (CAD). This study investigates associations of RARRES2rs17173608 with insulin resistance and the severity of CAD in non-obese T2D patients in relation to the clinical and genetic factors. (2) Methods: A total of 300 patients with T2D and CAD were recruited in this study. The associations of insulin resistance and the severity of CAD with RARRES2rs17173608 and clinical factors were assessed. The genotyping procedures were performed using the TaqMan method. The significant associations (p ≤ 0.05) from preliminary tests were employed to carry out the secondary analysis. (3) Results: RARRES2rs17173608 (TT, TG, and GG polymorphisms in the preliminary analysis; TG and GG polymorphisms in a secondary analysis) was associated with insulin resistance and the severity of CAD in both the preliminary and secondary analysis (all p-values were < 0.05). Additionally, in the secondary analysis, FPG and ACEI were also associated with insulin resistance and the severity of CAD (all p-values were < 0.05). (4) Conclusion: From the preliminary findings, rs17173608 is a significant predictor of insulin resistance and the severity of CAD.

Variants in the RARRES2 gene are associated with serum chemerin and increase the risk of diabetic kidney disease in type 2 diabetes.

Genetic susceptibility plays an important role in the pathogenesis of diabetic kidney disease (DKD). Recent studies have suggested that chemerin (encoded by the RARRES2 gene) is a risk factor for the development of DKD. We investigate the relationship between RARRES2 single nucleotide polymorphisms (SNPs) and DKD and their correlation with serum chemerin levels in Chinese individuals with type 2 diabetes (T2D). A total of 256 individuals with T2D were enrolled in this case-control study and classified into normo-, micro- and macroalbuminuria groups according to their urinary albumin/creatinine ratio (UACR). All exons of the RARRES2 gene were sequenced by polymerase chain reaction-direct sequencing, and 7 SNPs were genotyped. We found that the minor alleles of rs1047207, rs1047575 and rs1047586 were significantly associated with macroalbuminuria. Carriers of the minor allele of rs1047575 and rs1047586 also had higher urinary albumin (U-Alb) and UACR levels under both homo- and heterozygotic conditions than carriers of the major allele under the homozygotic condition. The minor alleles of rs1047207, rs1047575 and rs1047586 were significantly associated with increased serum chemerin levels under the homozygotic condition. These SNPs in the RARRES2 gene probably affect chemerin expression and likely confer susceptibility to albuminuria in individuals with T2D.

Chemerin Suppresses Breast Cancer Growth by Recruiting Immune Effector Cells Into the Tumor Microenvironment.

Infiltration of immune cells into the tumor microenvironment (TME) can regulate growth and survival of neoplastic cells, impacting tumorigenesis and tumor progression. Correlations between the number of effector immune cells present in a tumor and clinical outcomes in many human tumors, including breast, have been widely described. Current immunotherapies utilizing checkpoint inhibitors or co-stimulatory molecule agonists aim to activate effector immune cells. However, tumors often lack adequate effector cell numbers within the TME, resulting in suboptimal responses to these agents. Chemerin (RARRES2) is a leukocyte chemoattractant widely expressed in many tissues and is known to recruit innate leukocytes. CMKLR1 is a chemotactic cellular receptor for chemerin and is expressed on subsets of dendritic cells, NK cells, and macrophages. We have previously shown that chemerin acts as a tumor suppressive cytokine in mouse melanoma models by recruiting innate immune defenses into the TME. Chemerin/RARRES2 is down-regulated in many tumors, including breast, compared to normal tissue counterparts. Here, using a syngeneic orthotopic EMT6 breast carcinoma model, we show that forced overexpression of chemerin by tumor cells results in significant recruitment of NK cells and T cells within the TME. While chemerin secretion by EMT6 cells did not alter their phenotypic behavior in vitro, it did significantly suppress tumor growth in vivo. To define the cellular effectors required for this anti-tumor phenotype, we depleted NK cells or CD8+ T cells and found that either cell type is required for chemerin-dependent suppression of EMT6 tumor growth. Finally, we show significantly reduced levels of RARRES2 mRNA in human breast cancer samples compared to matched normal tissues. Thus, for the first time we have shown that increasing chemerin expression within the breast carcinoma TME can suppress growth by recruitment of NK and T cells, thereby supporting this approach as a promising immunotherapeutic strategy.

Expression and Regulation of Retinoic Acid Receptor Responders in the Human Placenta.

INTRODUCTION: Retinoic acid (RA) signaling through its receptors (RARA, RARB, RARG, and the retinoic X receptor RXRA) is essential for healthy placental and fetal development. An important group of genes regulated by RA are the RA receptor responders (RARRES1, 2, and 3). We set out to analyze their expression and regulation in healthy and pathologically altered placentas of preeclampsia (PE) and intrauterine growth restriction (IUGR) as well as in trophoblast cell lines. METHODS: We performed immunohistochemical staining on placental sections and analyzed gene expression by real-time polymerase chain reaction. Additionally, we performed cell culture experiments and stimulated Swan71 and Jeg-3 cells with different RA derivates and 2'-deoxy-5-azacytidine (AZA) to induce DNA demethylation. RESULTS: RARRES1, 2, and 3 and RARA, RARB, RARG, and RXRA are expressed in the extravillous part of the placenta. RARRES1, RARA, RARG, and RXRA were additionally detected in villous cytotrophoblasts. RARRES gene expression was induced via activation of RARA, RARB, and RARG in trophoblast cells. RARRES1 was overexpressed in villous trophoblasts and the syncytiotrophoblast from PE placentas, but not in IUGR without PE. Promoter methylation was detectable for RARRES1 and RARB based on their sensitivity toward AZA treatment of trophoblast cell lines. DISCUSSION: RARRES1, 2 and 3 are expressed in the functional compartments of the human placenta and can be regulated by RA. We hypothesize that the epigenetic suppression of trophoblast RARRES1 and RARB expression and the upregulation of RARRES1 in PE trophoblast cells suggest an involvement of environmental factors (eg, maternal vitamin A intake) in the pathogenesis of this pregnancy complication.