{"title":"Myocardial infarction elevates endoplasmic reticulum stress and protein aggregation in heart as well as brain.","authors":"Nirjal Mainali, Xiao Li, Xianwei Wang, Meenakshisundaram Balasubramaniam, Akshatha Ganne, Rajshekhar Kore, Robert J Shmookler Reis, Jawahar L Mehta, Srinivas Ayyadevara","doi":"10.1007/s11010-023-04856-3","DOIUrl":"10.1007/s11010-023-04856-3","url":null,"abstract":"<p><p>Cardiovascular diseases, including myocardial infarction (MI), constitute the leading cause of morbidity and mortality worldwide. Protein-aggregate deposition is a hallmark of aging and neurodegeneration. Our previous study reported that aggregation is strikingly elevated in hearts of hypertensive and aged mice; however, no prior study has addressed MI effects on aggregation in heart or brain. Here, we present novel data on heart and brain aggregation in mice following experimental MI, induced by left coronary artery (LCA) ligation. Infarcted and peri-infarcted heart tissue, and whole cerebra, were isolated from mice at sacrifice, 7 days following LCA ligation. Sham-MI mice (identical surgery without ligation) served as controls. We purified detergent-insoluble aggregates from these tissues, and quantified key protein constituents by high-resolution mass spectrometry (LC-MS/MS). Infarct heart tissue had 2.5- to 10-fold more aggregates than non-infarct or sham-MI heart tissue (each P = 0.001). Protein constituents from MI cerebral aggregates overlapped substantially with those from human Alzheimer's disease brain. Prior injection of mice with mesenchymal stem cell (MSC) exosomes, shown to limit infarct size after LCA ligation, reduced cardiac aggregation ~ 60%, and attenuated markers of endoplasmic reticulum (ER) stress in heart and brain (GRP78, ATF6, P-PERK) by 50-75%. MI also elevated aggregate constituents enriched in Alzheimer's disease (AD) aggregates, such as proteasomal subunits, heat-shock proteins, complement C3, clusterin/ApoJ, and other apolipoproteins. These data provide novel evidence that aggregation is elevated in mouse hearts and brains after myocardial ischemia, leading to cognitive impairment resembling AD, but can be attenuated by exosomes or drug (CDN1163) interventions that oppose ER stress.</p>","PeriodicalId":18724,"journal":{"name":"Molecular and Cellular Biochemistry","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11455681/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"71434402","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Ling Gao, Li Peng, Hong Tang, Chuang Wang, Qingsong Wang, Yujie Luo, Weiming Chen, Ying Xia
{"title":"Screening and identification of differential-expressed RNAs in thrombin-induced in vitro model of intracerebral hemorrhage.","authors":"Ling Gao, Li Peng, Hong Tang, Chuang Wang, Qingsong Wang, Yujie Luo, Weiming Chen, Ying Xia","doi":"10.1007/s11010-023-04879-w","DOIUrl":"10.1007/s11010-023-04879-w","url":null,"abstract":"<p><p>Survival of olfactory mucosal mesenchymal stem cells (OM-MSCs) remains the low level in the cerebral microenvironment during intracerebral hemorrhage (ICH). This article aims to reveal the differential expression profile of ICH-stimulated OM-MSCs based on whole transcriptome sequence analysis. OM-MSCs were isolated from 6-week C57BL/6 mice. Morphology and surface markers of OM-MSCs were investigated by light microscope and flow cytometry, respectively. OM-MSCs were incubated with 20 U/mL thrombin for 24 h to mimic ICH-induced injury in vitro. Total RNA was extracted for whole transcriptome sequencing and qPCR. OM-MSCs were characterized by negative for CD45 and CD34, and positive for CD44, CD90 and CD29. Thrombin led to decrease in cell viability and increase in senescence and apoptosis in OM-MSCs. In total, 736 lncRNAs (upregulated: 393; downregulated: 343), 21 miRNAs (upregulated: 7; downregulated: 14) and 807 mRNAs (upregulated: 422; downregulated: 385) were identified. GO and KEGG pathways were enriched in protein heterodimerization activity, trans-synaptic signaling, membrane pathway, alcohol metabolic process, organic hydroxy compound biosynthesis process, secondary alcohol metabolic process, alcoholism, neutrophil extracellular trap formation, systemic lupus erythematosus, metabolic process, steroid biosynthesis and drug metabolism-cytochrome P450. 200 lncRNA-miRNA-mRNA were predicted in thrombin-induced OM-MSCs. Based on qPCR, we validated COMMD1B, MOAP1, lncRNA CAPN15, lncRNA ALDH1L2, miR-3473b and miR-1964-3p were upregulated in thrombin-stimulated OM-MSCs, and GM20431, lncRNA GAPDH and miR-122b-3p were downregulated. Our findings provide novel understanding for thrombin-induced injury in OM-MSCs. Differently-expressed RNAs can be the targets of improving therapeutic application of OM-MSCs.</p>","PeriodicalId":18724,"journal":{"name":"Molecular and Cellular Biochemistry","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"71522104","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Naaleum Song, Jeong Eun Yu, Eunhye Ji, Kyoung-Hee Choi, Sahmin Lee
{"title":"Hydrogen sulfide inhibits gene expression associated with aortic valve degeneration by inducing NRF2-related pro-autophagy effect in human aortic valve interstitial cells.","authors":"Naaleum Song, Jeong Eun Yu, Eunhye Ji, Kyoung-Hee Choi, Sahmin Lee","doi":"10.1007/s11010-023-04881-2","DOIUrl":"10.1007/s11010-023-04881-2","url":null,"abstract":"<p><p>Aortic valve stenosis (AS) is the most common valvular heart disease but there are currently no effective medical treatments that can delay disease progression due to a lack of knowledge of the precise pathophysiology. The expression of sulfide: quinone oxidoreductase (SQOR) and nuclear factor erythroid 2-related factor 2 (NRF2) was decreased in the aortic valve of AS patients. However, the role of SQOR and NRF2 in the pathophysiology of AS has not been found. We investigated the effects of hydrogen sulfide (H<sub>2</sub>S)-releasing compounds on diseased aortic valve interstitial cells (AVICs) to explain the cellular mechanism of SQOR and elucidate the medical value of H<sub>2</sub>S for AS treatment. Sodium hydrosulfide (NaHS) treatment increased the expression of SQOR and NRF2 gene and consequently induced the NRF2 target genes, such as NAD(P)H quinone dehydrogenase 1 and cystathionine γ-lyase. In addition, NaHS dose-dependently decreased the expression level of fibrosis and inflammation-related genes (MMP9, TNF-α, IL6) and calcification-related genes (ALP, osteocalcin, RUNX2, COL1A1) in human AVICs. Furthermore, NaHS activated the AMPK-mTOR pathway and inhibited the PI3K-AKT pathway, resulting in a pro-autophagy effect in human AVICs. An NRF2 inhibitor, brusatol, attenuated NaHS-induced AMPK activation and decreased the autophagy markers Beclin-1 and LC3AB, suggesting that the mechanism of action of H<sub>2</sub>S is related to NRF2. In conclusion, H<sub>2</sub>S decreased gene expression levels related to aortic valve degeneration and activated AMPK-mTOR-mediated pro-autophagy function associated with NRF2 in human AVICs. Therefore, H<sub>2</sub>S could be a potential therapeutic target for the development of AS treatment.</p>","PeriodicalId":18724,"journal":{"name":"Molecular and Cellular Biochemistry","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49679715","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Fernanda Huf, Jessié Martins Gutierres, Gabrielle N da Silva, Adriana M Zago, Luiz Felipe C Koenig, Marilda C Fernandes
{"title":"Neuroprotection elicited by taurine in sporadic Alzheimer-like disease: benefits on memory and control of neuroinflammation in the hippocampus of rats.","authors":"Fernanda Huf, Jessié Martins Gutierres, Gabrielle N da Silva, Adriana M Zago, Luiz Felipe C Koenig, Marilda C Fernandes","doi":"10.1007/s11010-023-04872-3","DOIUrl":"10.1007/s11010-023-04872-3","url":null,"abstract":"<p><p>This study aimed to analyze whether taurine has a nootropic effect on short-term and long-term memory in a model of sporadic dementia of the Alzheimer's type (SDAT). Moreover, we evaluated the immunoreactivity and insulin receptor (IR) distribution and markers for neurons and glial cells in the hippocampus of rats with SDAT and treated with taurine. For this, Male Wistar rats received STZ (ICV, 3 mg/kg, bilateral, 5ul per site, aCFS vehicle) and were treated with taurine (100 mg/kg orally, 1 time per day, saline vehicle) for 25 days. The animals were divided into 4 groups: vehicle (VE), taurine (TAU), ICV-STZ (STZ) and ICV-STZ plus taurine (STZ + TAU). At the end of taurine treatment, short- and long-term memory were assessed by performance on object recognition and Y-maze tasks. Insulin receptor (IR) was evaluated by immunoperoxidase while mature neurons (NeuN), astrocytes (GFAP, S100B, SOX9), and microglia (Iba-1) were evaluated by immunofluorescence. STZ induced worse spatial and recognition memory (INDEX) in YM and ORT tasks. Taurine protected against STZ-induced memory impairment. SDAT reduced the population of mature neurons as well as increased astrocytic and microglial reactivity, and taurine protected against these STZ-induced effects, mainly in the CA1 region of the hippocampus. Taurine increases IR expression in the hippocampus, and protects against the reduction in the density of this receptor in CA1 induced by STZ. In conclusion, these findings demonstrate that taurine is able to enhance memory, up-regulates IR in the hippocampus, protects the neuron population, and reduces the astrogliosis found in SDAT.</p>","PeriodicalId":18724,"journal":{"name":"Molecular and Cellular Biochemistry","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"49691464","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Therapeutic applications and challenges in myostatin inhibition for enhanced skeletal muscle mass and functions.","authors":"Brock Wetzlich, Benard B Nyakundi, Jinzeng Yang","doi":"10.1007/s11010-024-05120-y","DOIUrl":"https://doi.org/10.1007/s11010-024-05120-y","url":null,"abstract":"<p><p>Myostatin, a potent negative regulator of skeletal muscle mass, has garnered significant attention as a therapeutic target for muscle dystrophies. Despite extensive research and promising preclinical results, clinical trials targeting myostatin inhibition in muscle dystrophies have failed to yield substantial improvements in muscle function or fitness in patients. This review details the mechanisms behind myostatin's function and the various inhibitors that have been tested preclinically and clinically. It also examines the challenges encountered in clinical translation, including issues with drug specificity, differences in serum myostatin concentrations between animal models and humans, and the necessity of neural input for functional improvements. Additionally, we explore promising avenues of research beyond muscle dystrophies, particularly in the treatment of metabolic syndromes and orthopedic disorders. Insights from these alternative applications suggest that myostatin inhibition may hold the potential for addressing a broader range of pathologies, providing new directions for therapeutic development.</p>","PeriodicalId":18724,"journal":{"name":"Molecular and Cellular Biochemistry","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-09-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142350255","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Xin Yan, Feng Li, Igor Dozmorov, Mark Barton Frank, Ming Dao, Michael Centola, Wei Cao, Dan Hu
{"title":"Retraction Note: External Qi of Yan Xin Qigong induces cell death and gene expression alterations promoting apoptosis and inhibiting proliferation, migration and glucose metabolism in small-cell lung cancer cells.","authors":"Xin Yan, Feng Li, Igor Dozmorov, Mark Barton Frank, Ming Dao, Michael Centola, Wei Cao, Dan Hu","doi":"10.1007/s11010-024-05107-9","DOIUrl":"https://doi.org/10.1007/s11010-024-05107-9","url":null,"abstract":"","PeriodicalId":18724,"journal":{"name":"Molecular and Cellular Biochemistry","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-09-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142350254","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Interconnections of screen time with neuroinflammation.","authors":"Ashish Verma, Anmol Kumar, Satendra Chauhan, Nisha Sharma, Anuradha Kalani, Prakash Chandra Gupta","doi":"10.1007/s11010-024-05123-9","DOIUrl":"https://doi.org/10.1007/s11010-024-05123-9","url":null,"abstract":"<p><p>The increasing prevalence of screen time among modern citizens has raised concerns regarding its potential impact on neuroinflammation and overall brain health. This review examines the complex interconnections between screen time and neuroinflammatory processes, particularly in children and adolescents. We analyze existing literature that explores how excessive digital media use can lead to alterations in neurobiological pathways, potentially exacerbating inflammatory responses in the brain. Key findings suggest that prolonged exposure to screens may contribute to neuroinflammation through mechanisms such as disrupted sleep patterns, diminished cognitive engagement, and increased stress levels. Similarly, we discuss the implications of these findings for mental health and cognitive development, emphasizing the need for a balanced approach to screen time. This review highlights the necessity for further research to elucidate the causal relationships and underlying mechanisms linking screen time and neuroinflammation, thereby informing guidelines for healthy media consumption.</p>","PeriodicalId":18724,"journal":{"name":"Molecular and Cellular Biochemistry","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-09-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142308057","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Rania A Radwan, Walaa A Elsalakawy, Doaa M Abdelaziz, Doaa M Abdelrazek, Sara M Radwan
{"title":"BsmI, ApaI and FokI variants of vitamin D receptor gene polymorphism as predictors of response to treatment in immune thrombocytopenia patients.","authors":"Rania A Radwan, Walaa A Elsalakawy, Doaa M Abdelaziz, Doaa M Abdelrazek, Sara M Radwan","doi":"10.1007/s11010-024-05100-2","DOIUrl":"https://doi.org/10.1007/s11010-024-05100-2","url":null,"abstract":"<p><p>Vitamin D receptor (VDR) polymorphisms are linked with the incidence and severity of several autoimmune diseases. The current work aimed at evaluating if VDR rs1544410 (BsmI), rs7975232 (ApaI) and rs2228570 (FokI) gene polymorphisms could be predictors of response to steroid treatment in patients with immune thrombocytopenia (ITP). The study involved 75 steroid treatment responders and 75 resistant ITP patients. All participants were subjected to VDR BsmI, ApaI and FokI gene polymorphisms analysis through genotyping by RT-PCR. Carrying the FokI F allele was significantly associated with low vitamin D level and increased risk of developing steroid resistance. Interestingly, the tri-allelic haplotypes BAF and BaF were significantly only present in steroid resistant ITP patients. Thus, the present study suggests that VDR FokI F allele may contribute to ITP pathogenesis and resistance to steroid treatment. Knowing the genetic background of patients helps to individualize treatment to obtain a better outcome.</p>","PeriodicalId":18724,"journal":{"name":"Molecular and Cellular Biochemistry","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-09-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142291458","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Somayeh Ghiasi Hafezi, Rana Kolahi Ahari, Maryam Saberi-Karimian, Zahra Eslami Giski, Amin Mansoori, Gordon A Ferns, Mahmoud Ebrahimi, Alireza Heidari-Bakavoli, Mohsen Moohebati, Sara Yousefian, Farnaz Farrokhzadeh, Habibollah Esmaily, Majid Ghayour-Mobarhan
{"title":"Association of high-sensitivity C-reactive protein and hematologic-inflammatory indices with risk of cardiovascular diseases: a population-based study with partial least squares structural equation modeling approach.","authors":"Somayeh Ghiasi Hafezi, Rana Kolahi Ahari, Maryam Saberi-Karimian, Zahra Eslami Giski, Amin Mansoori, Gordon A Ferns, Mahmoud Ebrahimi, Alireza Heidari-Bakavoli, Mohsen Moohebati, Sara Yousefian, Farnaz Farrokhzadeh, Habibollah Esmaily, Majid Ghayour-Mobarhan","doi":"10.1007/s11010-024-05122-w","DOIUrl":"https://doi.org/10.1007/s11010-024-05122-w","url":null,"abstract":"<p><p>Partial least squares structural equation modeling is a simple approach that may be used to determine the factors associated with diseases. In the current study, we aimed to explore the most associated high-sensitivity C-reactive protein (hs-CRP) as well as hematologic-inflammatory indices for the risk of cardiovascular disease (CVD). A total of 7362 healthy (non-CVD) participants aged 35-65 years old from baseline investigation were evaluated in the Phase 2 follow-up. Of these, 1022 individuals were found to have CVDs in the second phase (10-year follow-up) of the Mashhad Stroke and Heart Atherosclerotic Disorder (MASHAD) cohort study. We used partial least squares structural equation modeling to develop a prediction model for association of CVD risk factors and hs-CRP as well as hematologic-inflammatory indices in the study population. According to the study, age had the most significant impact on the presence of CVD. Increasing in age by one unit raises the risk of CVD by 0.166. Also, serum hs-CRP was found to have the second-highest impact on CVD; increasing in age by one unit raises the risk of CVD by 0.042. The study also discovered a strong and significant correlation between red cell distribution width (RDW) and CVD. Moreover, the study found that several factors such as hemoglobin (HGB), neutrophil (NEUT), neutrophil-to-lymphocyte ratio (NLR), systemic immune-inflammation index (SII), and platelet-to-lymphocyte ratio (PLR) have indirect effects on CVD that are mediated by hs-CRP while controlling for age, sex and social-economic factors. Generally, the results showed that age, hs-CRP, and RDW were the most important risk factors on CVD.</p>","PeriodicalId":18724,"journal":{"name":"Molecular and Cellular Biochemistry","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-09-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142291457","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Stress-induced extracellular vesicles: insight into their altered proteomic composition and probable physiological role in cancer.","authors":"Vaidehi Bhavsar, Ashish Sahu, Ravindra Taware","doi":"10.1007/s11010-024-05121-x","DOIUrl":"https://doi.org/10.1007/s11010-024-05121-x","url":null,"abstract":"<p><p>EVs (extracellular vesicles) are phospholipid bilayer vesicles that can be released by both prokaryotic and eukaryotic cells in normal as well as altered physiological conditions. These vesicles also termed as signalosomes, possess a distinctive cargo comprising nucleic acids, proteins, lipids, and metabolites, enabling them to play a pivotal role in both local and long-distance intercellular communication. The composition, origin, and release of EVs can be influenced by different physiological conditions and a variety of stress factors, consequently affecting the contents carried within these vesicles. Therefore, identifying the modified contents of EVs can provide valuable insights into their functional role in stress-triggered communication. Particularly, this is important when EVs released from tumor microenvironment are investigated for their role in the development and dissemination of cancer. This review article emphasizes the importance of differential EV shedding and altered proteomic content in response to reduced oxygen concentration, altered levels of glucose and glutamine, pH variations, oxidative stress and Ca<sup>2+</sup> ion concertation and it is subsequent effects on the behavior of recipient cells.</p>","PeriodicalId":18724,"journal":{"name":"Molecular and Cellular Biochemistry","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-09-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142291460","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}