Reproductive toxicology最新文献

{"title":"Microcystin-LR disrupts ovarian granulosa cell glycolysis via GSK3β-Mediated HK2 mitochondrial dissociation: Evidence from integrated In Vivo and In Vitro models","authors":"Jintao Yuan ,&nbsp;Xinrui Li ,&nbsp;Songci Yan ,&nbsp;Chengyu Luo ,&nbsp;Sijia Xian ,&nbsp;Yuanyuan Li ,&nbsp;Jiang Wu","doi":"10.1016/j.reprotox.2025.109028","DOIUrl":"10.1016/j.reprotox.2025.109028","url":null,"abstract":"<div><div>Reproductive disorders, a significant global health challenge, impact roughly 10 % of couples of reproductive ages. Notably, among these couples, 60–70 % of reproductive disorders are associated with women. Current evidence highlights that glycolysis plays an essential role in female reproductive function and is critical to follicle development and maturation. Microcystins (MCs), monocyclic heptapeptide toxins produced by freshwater cyanobacteria, include various isomers. Among them, microcystin-leucine-arginine (MC-LR) is the most prevalent and toxic form and is commonly found in water and food sources. However, the precise effects of MC-LR on glycolysis and its underlying mechanisms remain poorly understood. Therefore, this study aims to explore whether MC-LR induces reproductive defects in female mammals by disrupting glycolysis in human ovarian granulosa cells and mouse ovarian tissue, and to clarify its underlying mechanism.</div></div>","PeriodicalId":21137,"journal":{"name":"Reproductive toxicology","volume":"137 ","pages":"Article 109028"},"PeriodicalIF":2.8,"publicationDate":"2025-08-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144822452","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Corrigendum to “Maternal application of topical antifungal medication is associated with reduced steroid hormone levels during minipuberty and shorter anogenital distance in offspring from 3 months to 9 years of age: Odense Child Cohort” [Reprod. Toxicol. (2025) 109007 137]","authors":"Sarah Munk Andreasen ,&nbsp;Anna-Patricia Iversen ,&nbsp;Lars Christian Lund ,&nbsp;Margit Bistrup Fischer ,&nbsp;Anna-Maria Andersson ,&nbsp;Naja Kamuk Rauer ,&nbsp;Gylli Mola ,&nbsp;Anders Juul ,&nbsp;Casper P. Hagen ,&nbsp;Tina Kold Jensen","doi":"10.1016/j.reprotox.2025.109019","DOIUrl":"10.1016/j.reprotox.2025.109019","url":null,"abstract":"","PeriodicalId":21137,"journal":{"name":"Reproductive toxicology","volume":"137 ","pages":"Article 109019"},"PeriodicalIF":2.8,"publicationDate":"2025-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144765341","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Removal and toxic intervention of polystyrene microplastics and nanoplastics by magnetic nano-Fe3O4 in spermatogonial GC-1 cells","authors":"Cheng Cheng ,&nbsp;Yanfan Cui ,&nbsp;Yujie Wang ,&nbsp;Jingfeng Huang ,&nbsp;Jiale Ma ,&nbsp;Tao Luo ,&nbsp;Wen Chen","doi":"10.1016/j.reprotox.2025.109020","DOIUrl":"10.1016/j.reprotox.2025.109020","url":null,"abstract":"<div><div>Microplastics and nanoplastics (MNPs) are widespread in the environment and have male reproductive toxicity. However, toxic interventions involving MNPs have not been extensively examined. In this investigation, we explored the elimination capacity of magnetic nano-Fe₃O₄ on polystyrene microplastics and nanoplastics (PS-MNPs) of different sizes. This study also investigated whether magnetic nano-Fe₃O₄ could alleviate the toxicity of PS-MNPs in spermatogonial GC-1 cells. After coprecipitation by magnetic nano-Fe₃O₄ in ddH₂O, the removal rates of polystyrene microplastics (PS-MPs, 4 and 10 μm) are much higher than those of PS-NPs (25 nm, 100 nm, and 500 nm). The removal rate of the PS-NPs dramatically enhanced in the salt ion solutions. In addition, 25-nm, 100-nm, 500-nm, and 4-µm PS-MNPs penetrated GC-1 cells. Nevertheless, exclusively 25-nm PS-NPs decreased cell viability, elevated reactive oxygen species, disrupted the mitochondrial membrane potential, and induced apoptosis and inflammation through the P38/MAPK and Nrf2/HO-1 signaling pathways in GC-1 cells. Interestingly, magnetic nano-Fe₃O₄ alleviated these harmful impacts of the 25-nm PS-NPs on the GC-1 cells. In conclusion, we demonstrated the toxicity of PS-NPs in GC-1 cells and provided a viable way to alleviate their toxicity.</div></div>","PeriodicalId":21137,"journal":{"name":"Reproductive toxicology","volume":"137 ","pages":"Article 109020"},"PeriodicalIF":2.8,"publicationDate":"2025-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144757756","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Is ferroptosis a cause for concern in male infertility?","authors":"Bhagyalaxmi Sahoo ,&nbsp;Diptimayee Guru ,&nbsp;Anwesha Pradhan ,&nbsp;Soumya Ranjan Jena ,&nbsp;Lisa Goutami ,&nbsp;Jasmine Nayak ,&nbsp;Ashutosh Sahu ,&nbsp;Luna Samanta","doi":"10.1016/j.reprotox.2025.109022","DOIUrl":"10.1016/j.reprotox.2025.109022","url":null,"abstract":"<div><div>Ferroptosis, a recently identified form of regulated cell death, has emerged as a key player in the pathophysiology of various disorders, including male reproductive dysfunction. This review explores the interplay of ferroptosis with male reproductive health, focusing on the molecular mechanisms involved. Global male reproductive health has been deteriorating due to a combination of genetic, environmental, and lifestyle factors, including oxidative stress, reactive oxygen species (ROS), epigenetic alterations, and posttranslational modifications (PTMs). Ferroptosis is characterized by iron overload and lipid peroxidation, leading to testicular damage and impaired spermatogenesis, thus contributing to male infertility. While iron plays an essential role in maintaining spermatogenesis and testosterone production, its overload induces oxidative stress, ROS accumulation, and testicular ferroptosis, which disrupts normal reproductive function. Factors such as failure of antioxidant defense systems, exposure to xenobiotics (e.g., arsenite, cadmium, phthalates, bisphenol A, PM2.5), and alterations in ferroptosis-related genes (FRGs) further exacerbate testicular dysfunction. This review examines the critical role of lipid, iron, and glutathione metabolic pathways in ferroptosis induction, the effects of environmental xenobiotics on testicular health, and the potential therapeutic benefits of ferroptosis inhibitors in treating male infertility.</div></div>","PeriodicalId":21137,"journal":{"name":"Reproductive toxicology","volume":"137 ","pages":"Article 109022"},"PeriodicalIF":2.8,"publicationDate":"2025-07-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144765342","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Assessing microplastics as a novel threat to maternal-fetal health: Placental barrier penetration and fetal developmental consequences","authors":"Amy S. Mathew ,&nbsp;Abhirami Harjith ,&nbsp;C. Guruvayoorappan","doi":"10.1016/j.reprotox.2025.109021","DOIUrl":"10.1016/j.reprotox.2025.109021","url":null,"abstract":"<div><div>Microplastics (MPs) are now ubiquitous environmental contaminants that pose major risks to the environment and ecosystem, and in the last decade, concern about their potential threats to the reproductive system has gained widespread recognition. MPs can readily pass through biological barriers such as the placental and blood-brain barriers due to their small size. The ability of MPs to cross the placental barrier causes utmost concern as they could pass to the next generation and affect the health of the offspring. This review addresses the placental biology and its barrier function, as well as the ability of the MPs to translocate across the placenta. It also draws emphasis to responses of the placenta to the exposure of MPs at the cellular level, <em>in vivo</em> animal models, and human studies. To give a thorough grasp of the impacts of MPs at the maternal-fetal interface, investigations on the effects of MPs on fetal growth and development are also extensively reviewed.</div></div>","PeriodicalId":21137,"journal":{"name":"Reproductive toxicology","volume":"137 ","pages":"Article 109021"},"PeriodicalIF":2.8,"publicationDate":"2025-07-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144765340","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Subchronic exposure to Voliam Targo® affects ovarian histology and reproductive performance in rabbits (Oryctolagus cuniculus).","authors":"Thiziri Tlili ,&nbsp;Hassina Khaldoun ,&nbsp;Nacira Zerrouki Daoudi ,&nbsp;Rebiha AROUN ,&nbsp;Chahrazed Makhlouf ,&nbsp;Amina Settar ,&nbsp;Liza Benamara ,&nbsp;Nacima Djennane ,&nbsp;Smail Krabi","doi":"10.1016/j.reprotox.2025.109015","DOIUrl":"10.1016/j.reprotox.2025.109015","url":null,"abstract":"<div><div>In the current study, we evaluated the subchronic toxic effects of the Voliam Targo® (VT) insecticide on the ovaries of rabbits (<em>Oryctolagus cuniculus</em>) as well as the potential reproductive performance effects. The experiment was conducted using thirty females and thirty males, which were divided into two treated groups: control (distilled water) and VT (15 mg/kg b.w., by gavage, daily for 85 days). After a treatment period of 17 days, male and female rabbits from the two groups were randomly assigned to four artificial insemination (AI) mating groups using heterospermy or homospermy. The regimen continued through the gestation periods until 35 days of lactation, during which the first-generation (F1) offspring were monitored. At the end of the study, histomorphometric and immunohistochemical analyses were used to assess ovarian damage. The results revealed that body, ovary, uterine horn, cervix, and vagina weights did not vary significantly in the VT group compared to the control. Concerning reproductive performance, paternal exposure to VT insecticide caused a significant (p &lt; 0.01) increase in the number of live fetuses and a decrease in the percentage of death in pups during the postnatal period. Also, VT treatment resulted in ovarian tissue structure disorganization, including follicular atresia, hemorrhagic follicles, and ovarian degeneration. Moreover, Ki67, P53, and Bcl-2 protein expression in the ovaries of the VT-treated group differed from the control group. This study suggests that subchronic exposure to Voliam Targo® may affect ovarian structure and reproductive performance by altering cell proliferation and apoptosis.</div></div>","PeriodicalId":21137,"journal":{"name":"Reproductive toxicology","volume":"137 ","pages":"Article 109015"},"PeriodicalIF":2.8,"publicationDate":"2025-07-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144721814","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Reprotoxic effect of bisphenol F and bisphenol S on female Drosophila melanogaster exposed during development","authors":"Elize Musachio ,&nbsp;Bianca Munieweg da Silva ,&nbsp;Graziela Moro Meira ,&nbsp;Barbara Osmarin Turra ,&nbsp;Cibele Ferreira Teixeira ,&nbsp;Fernanda Barbisan ,&nbsp;Luana Barreto Meichtry ,&nbsp;Eliana Jardim Fernandes ,&nbsp;Dieniffer Espinosa Janner ,&nbsp;Gustavo Petri Guerra ,&nbsp;Marina Prigol","doi":"10.1016/j.reprotox.2025.109017","DOIUrl":"10.1016/j.reprotox.2025.109017","url":null,"abstract":"<div><div>What is the effect of exposure to Bisphenol F (BPF) and Bisphenol S (BPS) during the embryonic and developmental period on the reproductive system of filial generation 1 (F1) female flies? To answer this question, <em>Drosophila melanogaster</em> were used, which remained throughout the development period, at different concentrations of BPF and BPS (0.25, 0.5, and 1 mM, separately). Upon hatching, evaluation of wing size and body weight, in addition to the expression of the ecdysone receptor (EcR) and enzymes catalase (CAT) and superoxide dismutase (SOD), quantification of reactive species (RS) and CYP 450 activity were performed in part of virgin females (F1). Another part of the females (F1) were mated with untreated males to assess reprotoxicity through tests such as fertility and fecundity, by counting the number of eggs laid and hatched, and evaluation of ovary morphology and viability of ovarian cells. Exposure to 1 mM BPF, 0.5 and 1 mM BPS reduced the ability of flies to lay eggs, changed the shape and size of the ovaries, reduced the viability of ovarian cells, increased body weight, and reduced wing size. EcR reduction was observed at all concentrations, accompanied by an increase of CAT and SOD expression and an increase in CYP450 activity. Even with increases in antioxidant enzymes, the reproductive system of females exposed to 1 mM BPF, 0.5 and 1 mM BPS may have succumbed to the rise in RS and reduction of EcR, which indicates endocrine dysregulation, thus manifesting a reprotoxic effect, highlighting BPS as more harmful.</div></div>","PeriodicalId":21137,"journal":{"name":"Reproductive toxicology","volume":"137 ","pages":"Article 109017"},"PeriodicalIF":2.8,"publicationDate":"2025-07-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144724894","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Folate deficiency mediates ovarian dysfunction through appetite and inflammatory pathways","authors":"Afridi Shaikh ,&nbsp;Bharti Choudhary ,&nbsp;Mukund Chhatpar ,&nbsp;Dewaunshi Panchakshari ,&nbsp;Dhaval Fefar ,&nbsp;Hetal Roy","doi":"10.1016/j.reprotox.2025.109016","DOIUrl":"10.1016/j.reprotox.2025.109016","url":null,"abstract":"<div><div>The long-established link between nutrition and reproduction is known to have critical consequences for reproductive function. However, the available experimental data on the effects of folate deficiency on ovarian health remains scarce. It is still unclear whether folate deficiency is directly responsible for causing ovarian-dysfunction and, if so, what are the underlying mechanisms. Therefore, our objective was to establish evidence for association between folate deficiency, hormone dynamics, and ovarian function using in vivo model. Folate-deprived female zebrafish were developed using intraperitoneal administration of methotrexate (MTX; DHFR inhibitor) and were used to study the possible implications of folate deprivation on ovarian health. Changes in the expression of transcripts regulating appetite and ovarian function was observed. We observed that folate deprivation resulted in impaired appetite behaviour and alteration in its regulatory gene expression. Due to folate deficiency, the neuroendocrine function of the brain was affected that resulted in altered reproductive hormone levels. Histology of ovary shows follicles arrested in primary oocyte stage and scarring of tissue is seen. Furthermore, elevated lipid peroxidation and catalase enzyme activity along with increased IL-6, indicates folate deficiency induced oxidative stress and inflammation in ovary as one of the possible mechanisms to aid- ovarian dysfunction. Our study provides experimental evidence, using an in vivo folate-deficient fish model, that underscores the essential role of folate in maintaining reproductive health. The intricate relationship between folate deficiency, appetite regulation, and its impact on the synthesis and release of female reproductive hormones calls for deeper investigation, particularly through studies involving mammalian models.</div></div>","PeriodicalId":21137,"journal":{"name":"Reproductive toxicology","volume":"137 ","pages":"Article 109016"},"PeriodicalIF":2.8,"publicationDate":"2025-07-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144724905","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The consumption of monosodium glutamate during the periconceptional period can impair preimplantation embryo development","authors":"Zuzana Šefčíková,&nbsp;Alexandra Špirková,&nbsp;Veronika Kovaříková,&nbsp;Laura Rušinová,&nbsp;Vladimír Baran,&nbsp;Jozef Pisko,&nbsp;Janka Babeľová,&nbsp;Dušan Fabian,&nbsp;Štefan Čikoš","doi":"10.1016/j.reprotox.2025.109014","DOIUrl":"10.1016/j.reprotox.2025.109014","url":null,"abstract":"<div><div>Monosodium glutamate (MSG) is one of the most commonly used food additives and is consumed worldwide as part of commercially processed foods. To study the possible reproductive risks of consuming monosodium glutamate during the periconceptional period (comprising oocyte maturation and the earliest stages of embryo development), we administered MSG (at doses of 500, 200 or 50 mg/kg body weight, by oral gavage) to female mice in this period. Preimplantation embryos were then isolated at D4 of gestation and analyzed. In blastocysts developed from female mice fed with MSG, we found a reduced proportion of blastocysts, a lower number of cells, and an increased proportion of dead cells. The expression of the proapoptotic gene Bak1 and active caspase-3 were significantly increased and telomeres were shorter in blastocysts isolated from MSG-fed females. The results of our previous in vitro study revealed that glutamate receptors are involved in the negative effects of glutamate on mouse blastocysts. In silico analysis of RNA-seq datasets containing data from human preimplantation embryos performed in this study revealed that NMDA receptors (formed from the GRIN2D or GRIN2B and GRIN3B subunits) and the GRM4 and GRM8 metabotropic receptors are expressed in human blastocysts. These results indicate that glutamate receptors could mediate the effects of MSG in human blastocyst cells. In summary, our results show that oral intake of relatively low doses of MSG during the periconception period can adversely affect early embryonic development and embryo quality and may pose a risk to successful conception and subsequent embryo development.</div></div>","PeriodicalId":21137,"journal":{"name":"Reproductive toxicology","volume":"137 ","pages":"Article 109014"},"PeriodicalIF":2.8,"publicationDate":"2025-07-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144725061","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Investigating the protective role of Astragalus polysaccharides against fluoride-induced testicular injury via network pharmacology, molecular docking, and in vivo experiments","authors":"Yue Pan ,&nbsp;Xingyan Du ,&nbsp;Yuanyuan Xiao ,&nbsp;Qiuhan Pi ,&nbsp;Yu Chen ,&nbsp;Jiajun Huang ,&nbsp;Didong Lou ,&nbsp;Wenchao Tang","doi":"10.1016/j.reprotox.2025.109012","DOIUrl":"10.1016/j.reprotox.2025.109012","url":null,"abstract":"<div><h3>Objective</h3><div>This study aimed to investigate the protective effects of Astragalus polysaccharides (APS) against sodium fluoride (NaF)-induced testicular damage and to provide a theoretical basis for developing natural strategies to mitigate fluoride-induced reproductive toxicity.</div></div><div><h3>Methods</h3><div>Network pharmacology was employed to identify potential targets and pathways of APS in treating fluoride-induced testicular injury. Molecular docking was used to assess the binding affinity between APS active compounds and core targets. An in vivo rat model was established to evaluate testicular index, fluoride accumulation, and histopathological changes. Oxidative stress markers including SOD, CAT, GSH, and MDA were measured. Immunohistochemistry was performed to assess blood–testis barrier integrity and the expression of key targets (AKT1, ESR1, CASP3).</div></div><div><h3>Results</h3><div>Network pharmacology identified 34 potential APS targets, enriched in apoptosis, PI3K-Akt, and IL-1B signaling pathways. Molecular docking revealed strong binding affinity of APS components to core targets. APS significantly improved NaF-induced reductions in testicular index (<em>p</em> &lt; 0.01) and fluoride accumulation (<em>p</em> &lt; 0.05), alleviated seminiferous tubule atrophy and mitochondrial swelling, enhanced SOD, CAT activities and GSH levels (<em>p</em> &lt; 0.01), and reduced MDA levels (<em>p</em> &lt; 0.01). Moreover, APS upregulated CLDN1, ZO-1, and Occludin expression to repair the blood–testis barrier. Abnormal expression of AKT1, ESR1, CASP3, and HSP90AA1 was reversed by APS (<em>p</em> &lt; 0.01).</div></div><div><h3>Conclusion</h3><div>APS mitigates fluoride-induced testicular toxicity via multi-target regulation of oxidative stress and blood–testis barrier pathways, offering a novel therapeutic approach for fluoride-related reproductive damage.</div></div>","PeriodicalId":21137,"journal":{"name":"Reproductive toxicology","volume":"137 ","pages":"Article 109012"},"PeriodicalIF":3.3,"publicationDate":"2025-07-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144704164","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}