RESUMEN
Chromatin accessibility plays a critical role in the regulation of cell fate decisions. Although gene expression changes have been extensively profiled at the single-cell level during early embryogenesis, the dynamics of chromatin accessibility at cis-regulatory elements remain poorly studied. Here, we used a plate-based single-cell ATAC-seq method to profile the chromatin accessibility dynamics of over 10 000 nuclei from zebrafish embryos. We investigated several important time points immediately after zygotic genome activation (ZGA), covering key developmental stages up to dome. The results revealed key chromatin signatures in the first cell fate specifications when cells start to differentiate into enveloping layer (EVL) and yolk syncytial layer (YSL) cells. Finally, we uncovered many potential cell-type specific enhancers and transcription factor motifs that are important for the cell fate specifications.
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Cromatina , Desarrollo Embrionario , Pez Cebra , Animales , Cromatina/genética , Cromatina/metabolismo , Yema de Huevo/metabolismo , Embrión no Mamífero/embriología , Embrión no Mamífero/metabolismo , Desarrollo Embrionario/genética , Pez Cebra/embriología , Pez Cebra/genética , Análisis de la Célula Individual , Dominios y Motivos de Interacción de Proteínas/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Intronic polyadenylation (IpA) usually leads to changes in the coding region of an mRNA, and its implication in diseases has been recognized, although at its very beginning status. Conveniently and accurately identifying IpA is of great importance for further evaluating its biological significance. Here, we developed IPAFinder, a bioinformatic method for the de novo identification of intronic poly(A) sites and their dynamic changes from standard RNA-seq data. Applying IPAFinder to 256 pan-cancer tumor/normal pairs across six tumor types, we discovered 490 recurrent dynamically changed IpA events, some of which are novel and derived from cancer-associated genes such as TSC1, SPERD2, and CCND2 Furthermore, IPAFinder revealed that IpA could be regulated by factors related to splicing and m6A modification. In summary, IPAFinder enables the global discovery and characterization of biologically regulated IpA with standard RNA-seq data and should reveal the biological significance of IpA in various processes.
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Neoplasias , Poliadenilación , Humanos , Intrones/genética , Neoplasias/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , RNA-SeqRESUMEN
H6 avian influenza virus is widely prevalent in wild birds and poultry and has caused human infection in 2013 in Taiwan, China. During our active influenza surveillance program in wild waterfowl at Poyang Lake, Jiangxi Province, an H6N2 AIV was isolated and named A/bean goose/JiangXi/452-4/2013(H6N2). The isolate was characterized as a typical low pathogenic avian influenza virus (LPAIV) due to the presence of the amino acid sequence PQIETR↓GLFGAI at the cleavage site of the hemagglutinin (HA) protein. The genetic evolution analysis revealed that the NA gene of the isolate originated from North America and exhibited the highest nucleotide identity (99.29%) with a virus recovered from wild bird samples in North America, specifically A/bufflehead/California/4935/2012(H11N2). Additionally, while the HA and PB1 genes belonged to the Eurasian lineage, they displayed frequent genetic interactions with the North American lineage. The remaining genes showed close genetic relationships with Eurasian viruses. The H6N2 isolate possessed a complex genome, indicating it is a multi-gene recombinant virus with genetic material from both Eurasian and North American lineages.
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Animales Salvajes , Virus de la Influenza A , Gripe Aviar , Filogenia , Virus Reordenados , Animales , China , Virus Reordenados/genética , Virus Reordenados/aislamiento & purificación , Virus Reordenados/clasificación , Gripe Aviar/virología , Animales Salvajes/virología , Virus de la Influenza A/genética , Virus de la Influenza A/aislamiento & purificación , Virus de la Influenza A/clasificación , Glicoproteínas Hemaglutininas del Virus de la Influenza/genética , Aves/virología , Evolución Molecular , Genoma Viral/genética , Neuraminidasa/genética , Proteínas Virales/genéticaRESUMEN
High-yielding dairy cows in early lactation often encounter difficulties in meeting the energy requirements essential for maintaining milk production. This is primarily attributed to insufficient dry matter intake, which consequently leads to sustained lipolysis of adipose tissue. Fatty acids released by lipolysis can disrupt metabolic homeostasis. Autophagy, an adaptive response to intracellular environmental changes, is considered a crucial mechanism for regulating lipid metabolism and maintaining a proper cellular energy status. Despite its close relationship with aberrant lipid metabolism and cytolipotoxicity in animal models of metabolic disorders, the precise function of diacylglycerol o-acyltransferase 1 (DGAT1) in bovine adipose tissue during periods of negative energy balance is not fully understood, particularly regarding its involvement in lipolysis and autophagy. The objective of the present study was to assess the effect of DGAT1 on both lipolysis and autophagy in bovine adipose tissue and isolated adipocytes. Adipose tissue and blood samples were collected from cows diagnosed as clinically ketotic (n = 15) or healthy (n = 15) following a veterinary evaluation based on clinical symptoms and serum concentrations of BHB, which were 3.19 mM (interquartile range = 0.20) and 0.50 mM (interquartile range = 0.06), respectively. Protein abundance of DGAT1 and phosphorylation levels of unc-51-like kinase 1 (ULK1), were greater in adipose tissue from cows with ketosis, whereas phosphorylation levels of phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), and mammalian target of rapamycin (mTOR) were lower. Furthermore, when adipocytes isolated from the harvested adipose tissue of 15 healthy cows were transfected with DGAT1 overexpression adenovirus or DGAT1 small interfering RNA followed by exposure to epinephrine (EPI), it led to greater ratios and protein abundance of phosphorylated hormone-sensitive triglyceride lipase (LIPE) to total LIPE and adipose triglyceride lipase (ATGL), while inhibiting the protein phosphorylation levels of ULK1, PI3K, AKT, and mTOR. Overexpression of DGAT1 in EPI-treated adipocytes reduced lipolysis and autophagy, whereas silencing DGAT1 further exacerbated EPI-induced lipolysis and autophagy. Taken together, these findings indicate that upregulation of DGAT1 may function as an adaptive response to suppress adipocytes lipolysis, highlighting the significance of maintaining metabolic homeostasis in dairy cows during periods of negative energy balance.
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Tejido Adiposo , Autofagia , Diacilglicerol O-Acetiltransferasa , Lipólisis , Animales , Bovinos , Diacilglicerol O-Acetiltransferasa/metabolismo , Diacilglicerol O-Acetiltransferasa/genética , Femenino , Tejido Adiposo/metabolismo , Lactancia , Cetosis/veterinaria , Cetosis/metabolismo , Metabolismo de los Lípidos , Adipocitos/metabolismoRESUMEN
Excessive concentrations of free fatty acids (FFA) are the main factors causing immune dysfunction and inflammation in dairy cows with ketosis. Polarization of macrophages (the process of macrophages freely switching from one phenotype to another) into M1 or M2 phenotypes is an important event during inflammation induced by environmental stimuli. In nonruminants, mammalian target of rapamycin (mTOR)-mediated autophagy (a major waste degradation process) regulates macrophage polarization. Thus, our objective was to unravel the role of mTOR-mediated autophagy on macrophage polarization in ketotic dairy cows. We performed 4 experiments: (1) In vitro differentiated monocyte-derived macrophages from healthy dairy cows or dairy cows with clinical ketosis (CK) were treated for 24 h with 100 ng/mL LPS and 100 ng/mL IFN-γ or with 10 ng/mL IL4 and 10 ng/mL IL10; (2) Immortalized bovine macrophages were treated for 24 h with 0, 0.3, 0.6, or 1.2 mM FFA, LPS, and IFN-γ, or with IL4 and IL10; (3) Macrophages were pretreated with 2 µM 4,6-dimorpholino-N-(4-nitrophenyl)-1,3,5-triazin-2-amine (MHY1485) for 30 min before treatment with LPS and IFN-γ or IL4 and IL10; (4) Macrophages were pretreated with 100 nM rapamycin (RAPA) for 2 h before treatment with LPS and IFN-γ or IL4 and IL10. Compared with healthy cows, cows with CK had a greater mean fluorescence intensity (MFI) of CD86+, but lower MFI of CD206+ and lower number of autophagosomes and autolysosomes in macrophages. Exogenous FFA treatment upregulated protein abundance of inducible nitric oxide synthase (iNOS) and the MFI of CD86, whereas it downregulated the protein abundance of arginase 1 and the MFI of CD206. In addition, FFA increased the p-p65/p65 protein abundance and tumor necrosis factor α, IL1B, and IL6 mRNA abundance, but decreased LC3-phosphatidylethanolamine conjugate protein abundance and the number of autophagosomes and autolysosomes number. Pretreatment with MHY1485 promoted macrophage M1 polarization and inhibited macrophage M2 polarization via decreased mTOR-mediated autophagy. Activation of mTOR-mediated autophagy by pretreatment with RAPA attenuated the upregulation of inflammation in M1 macrophages that was induced by FFA. These data revealed that high concentrations of FFA promote macrophage M1 polarization in ketotic dairy cows by impairing mTOR-mediated autophagy.
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Autofagia , Macrófagos , Serina-Treonina Quinasas TOR , Animales , Bovinos , Macrófagos/efectos de los fármacos , Autofagia/efectos de los fármacos , Femenino , Serina-Treonina Quinasas TOR/metabolismo , Ácidos Grasos/farmacología , Ácidos Grasos/metabolismo , Cetosis/veterinaria , Lipopolisacáridos/farmacología , Línea CelularRESUMEN
Postpartum cows experience lipolysis in adipose tissue due to negative energy balance (NEB), and accumulation of free fatty acids (FFA) leads to metabolic stress in adipose tissue. Ferroptosis is a type of cell death triggered by excessive buildup of iron-dependent lipid peroxides, which is involved in the occurrence and development of various metabolic diseases in nonruminants. However, it is still unclear whether ferroptosis occurs in the adipose tissue of ketotic cows and the regulatory mechanisms behind ferroptosis. Despite multiple studies demonstrating the significant involvement of hypoxia-inducible-factor-1α (HIF-1α) in regulating cellular dysfunction, its specific function in adipose tissue of ketotic dairy cows remains uncertain, particularly its regulation of oxidative stress and ferroptosis. The study aimed to explore the impact of HIF-1α on oxidative stress and ferroptosis in bovine subcutaneous adipose tissue and isolated adipocytes. The adipose tissue of clinical ketosis cows (n = 15) with a serum BHB concentration of 3.13 mM (interquartile range = 0.14) and healthy cows (n = 15) with a serum BHB concentration of and 0.58 mM (interquartile range = 0.13) was collected. The results showed that the concentrations of lipid peroxidation malondialdehyde (MDA), reactive oxygen species (ROS), Fe2+ and total iron were increased in adipose tissue of cows with ketosis, while the contents of glutathione (GSH) were reduced. Furthermore, the protein levels of HIF-1α, heme oxygenase 1 (HMOX1), catalase (CAT), superoxide dismutase 1 (SOD1), acyl-CoA synthetase 4 (ACSL4), and nuclear factor erythroid-derived 2-like 2 (NFE2L2) exhibited higher abundance in adipose tissue obtained from cows with ketosis, whereas the protein abundance of solute carrier family 7 member 11 (SLC7A11), glutamate cysteine ligase catalytic subunit (GCLC), kelch-like ECH-associated protein 1 (KEAP1), glutamate cysteine ligase regulatory subunit (GCLM) and glutathione peroxidase 4 (GPX4) were lower. To simulate the ferroptosis state of adipose tissue in ketotic cows, primary bovine adipocytes were isolated from the adipose tissue of healthy cows and cultured with erastin to construct ferroptosis model. Adipocytes were cultured with either an adenovirus overexpressing HIF-1α or small interfering RNA targeting HIF-for 48 h, followed by exposure to erastin (1 µM) for 24 h. Treatment with erastin led to higher protein abundance of CAT, SOD1, NFE2L2 and HMOX1, while it inhibited the protein expression levels of GCLC, SLC7A11, GCLM, GPX4 and KEAP1. Furthermore, erastin treatment elevated the levels of ROS, MDA, Fe2+, total iron and reduced the content of GSH. The overexpression of HIF-1α reversed the erastin-induced decreases in the protein abundance of GPX4 and SLC7A11, as well as the levels of MDA, ROS, Fe2+ and total iron, while significantly increasing protein abundance and content of CAT, SOD1, NFE2L2, HMOX1, GCLC, GCLM, GPX4, SLC7A11 and GSH. Conversely, the silencing of HIF-1α further exacerbated the erastin-induced levels of MDA, ROS, Fe2+ and total iron, while inhibiting the upregulation of SOD1, CAT, NFE2L2 and HMOX1. Collectively, these findings suggest that activation of HIF-1α may function as an adaptive mechanism to mitigate ferroptosis and alleviate oxidative stress in adipose tissue.
RESUMEN
Somatic single nucleotide variants (SNVs) in cancer genome affect gene expression through various mechanisms depending on their genomic location. While somatic SNVs near canonical splice sites have been reported to cause abnormal splicing of cancer-related genes, whether these SNVs can affect gene expression through other mechanisms remains an open question. Here, we analyzed RNA sequencing and exome data from 4,998 cancer patients covering ten cancer types and identified 152 somatic SNVs near splice sites that were associated with abnormal intronic polyadenylation (IPA). IPA-associated somatic variants favored the localization near the donor splice sites compared to the acceptor splice sites. A proportion of SNV-associated IPA events overlapped with premature cleavage and polyadenylation events triggered by U1 small nuclear ribonucleoproteins (snRNP) inhibition. GC content, intron length and polyadenylation signal were three genomic features that differentiated between SNV-associated IPA and intron retention. Notably, IPA-associated SNVs were enriched in tumor suppressor genes (TSGs), including the well-known TSGs such as PTEN and CDH1 with recurrent SNV-associated IPA events. Minigene assay confirmed that SNVs from PTEN, CDH1, VEGFA, GRHL2, CUL3 and WWC2 could lead to IPA. This work reveals that IPA acts as a novel mechanism explaining the functional consequence of somatic SNVs in human cancer.
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Neoplasias/genética , Polimorfismo de Nucleótido Simple , ARN , Bases de Datos Genéticas , Humanos , Intrones , PoliadenilaciónRESUMEN
The transition from meiotic spermatocytes to postmeiotic haploid germ cells constitutes an essential step in spermatogenesis. The epigenomic regulatory mechanisms underlying this transition remain unclear. Here, we find a prominent transcriptomic switch from the late spermatocytes to the early round spermatids during the meiotic-to-postmeiotic transition, which is associated with robust histone acetylation changes across the genome. Among histone deacetylases (HDACs) and acetyltransferases, we find that HDAC3 is selectively expressed in the late meiotic and early haploid stages. Three independent mouse lines with the testis-specific knockout of HDAC3 show infertility and defects in meiotic exit with an arrest at the late stage of meiosis or early stage of round spermatids. Stage-specific RNA-seq and histone acetylation ChIP-seq analyses reveal that HDAC3 represses meiotic/spermatogonial genes and activates postmeiotic haploid gene programs during meiotic exit, with associated histone acetylation alterations. Unexpectedly, abolishing HDAC3 catalytic activity by missense mutations in the nuclear receptor corepressor (NCOR or SMRT) does not cause infertility, despite causing histone hyperacetylation as HDAC3 knockout, demonstrating that HDAC3 enzyme activity is not required for spermatogenesis. Motif analysis of the HDAC3 cistrome in the testes identified SOX30, which has a similar spatiotemporal expression pattern as HDAC3 during spermatogenesis. Depletion of SOX30 in the testes abolishes the genomic recruitment of the HDAC3 to the binding sites. Collectively, these results establish the SOX30/HDAC3 signaling as a key regulator of the transcriptional program in a deacetylase-independent manner during the meiotic-to-postmeiotic transition in spermatogenesis.
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Fertilidad/genética , Regulación de la Expresión Génica , Histona Desacetilasas/fisiología , Meiosis/genética , Espermatogénesis/genética , Activación Transcripcional , Acetilación , Animales , Reprogramación Celular/genética , Histona Desacetilasas/genética , Histona Desacetilasas/metabolismo , Histonas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Factores de Transcripción SOX/metabolismo , Espermátides/citología , Espermátides/metabolismo , Testículo/metabolismoRESUMEN
BACKGROUND: The impact of maternal pre-pregnancy bodyweight on gestational diabetes mellitus (GDM) following assisted reproductive technology (ART) treatment has been insufficiently investigated. The aim of this study was to investigate the association between maternal pre-pregnancy bodyweight and GDM following ART. METHODS: From January 2014 to March 2019, this population-based retrospective cohort study included pregnancies achieved by ART treatment in a pregnancy registration database in China. Multivariate regression analysis and restricted cubic splines were used to explore the association between bodyweight and GDM. RESULTS: A total of 6,598 pregnancies were included. The incidence of GDM was 26.0% (1715/6598). A total of 868 (13.2%) pregnant women were underweight, 665 (10.8%) were overweight, and 145 (2.20%) were obesity. We found a linear dose-response relation between maternal body mass index and GDM by restricted cubic splines, where one unit body mass index increase was associated with the 15% elevated risk of GDM (adjusted odds ratio [OR] 1.15, 95% CI 1.08-1.22). Compared to the normal weight group, maternal underweight was associated with lower risk of GDM (adjusted OR 0.68, 95% CI 0.57-0.82), while increased risk was found for overweight (adjusted OR 1.54 95% CI 1.29-1.84) and obesity (adjusted OR 1.74, 95% CI 1.23-2.47). CONCLUSIONS: Our study found a linear dose-effect relationship between pre-pregnancy bodyweight and GDM following ART treatment. The findings in this study support the clinical recommendation of advising women with overweight or obesity to lose weight prior to ART treatment.
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Diabetes Gestacional , Índice de Masa Corporal , Estudios de Cohortes , Diabetes Gestacional/epidemiología , Diabetes Gestacional/etiología , Femenino , Humanos , Obesidad/complicaciones , Sobrepeso/epidemiología , Embarazo , Técnicas Reproductivas Asistidas/efectos adversos , Estudios Retrospectivos , Factores de Riesgo , Delgadez/complicaciones , Delgadez/epidemiologíaRESUMEN
CircSERPINA3 has been shown to be upregulated in laryngeal squamous cell carcinoma (LSCC); however, whether it regulates the development of LSCC and the specific molecular mechanism remains unclear, which is to be explored in this study. Expressions of circSERPINA3, miR-885-5p, and Malic enzyme 1 (ME1) in LSCC tissues or cell lines were determined by quantitative reverse transcription polymerase chain reaction (qRT-PCR). The regulation of circSERPINA3 on the biological behavior of LSCC cells was confirmed by loss and gain experiments (cell counting kit-8, transwell, and colony formation assay). The correlation between circSERPINA3/ME1 and miR-885-5p was predicted and confirmed by bioinformatics analysis, dual-luciferase reporter assay, and qRT-PCR. The effect of circSERPINA3/miR-885-5p axis on the biological behavior of LSCC cells and expressions of epithelial-mesenchymal transition-related proteins was confirmed by rescue experiments. CircSERPINA3 and ME1 was upregulated in LSCC tissues, whereas miR-885-5p was downregulated. MiR-885-5p was the target gene of circSERPINA3, whereas ME1 was the target gene of miR-885-5p. Silent circSERPINA3 suppressed viability, invasion, migration, colony formation, and expression of ME1, claudin-4, snail, and vimentin but elevated expression of miR-885-5p and E-cadherin, whereas overexpressed circSERPINA3 was the opposite. However, miR-885-5p inhibitor or mimic reversed the effects of silent circSERPINA3 or overexpressed circSERPINA3. Collectively, circSERPINA3 promotes proliferation, migration, and invasion of LSCC cells by targeting miR-885-5p.
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Neoplasias de Cabeza y Cuello , MicroARNs , Cadherinas/genética , Cadherinas/metabolismo , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Claudina-4/genética , Claudina-4/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias de Cabeza y Cuello/genética , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Vimentina/metabolismoRESUMEN
Bronchopulmonary dysplasia (BPD) represents a multifactorial chronic pulmonary pathology and a major factor causing premature illness and death. The therapeutic role of exosomes in BPD has been feverishly investigated. Meanwhile, the potential roles of exosomal circRNAs, lncRNAs, and mRNAs in umbilical cord blood (UCB) serum have not been studied. This study aimed to detect the expression profiles of circRNAs, lncRNAs, and mRNAs in UCB-derived exosomes of infants with BPD. Microarray analysis was performed to compare the RNA profiles of UCB-derived exosomes of a preterm newborn with (BPD group) and without (non-BPD, NBPD group) BPD. Then, circRNA/lncRNA-miRNA-mRNA co-expression networks were built to determine their association with BPD. In addition, cell counting kit-8 (CCK-8) assay was used to evaluate the proliferation of lipopolysaccharide (LPS)-induced human bronchial epithelial cells (BEAS-2B cells) and human umbilical vein endothelial cells (HUVECs). The levels of tumor necrosis factor (TNF)-α and interleukin (IL)-1ß in LPS-induced BEAS-2B cells and HUVECs were assessed through Western blot analysis. Then, quantitative reverse transcription-polymerase chain reaction assay was used to evaluate the expression levels of four differentially expressed circRNAs (hsa_circ_0086913, hsa_circ_0049170, hsa_circ_0087059, and hsa_circ_0065188) and two lncRNAs (small nucleolar RNA host gene 20 (SNHG20) and LINC00582) detected in LPS-induced BEAS-2B cells or HUVECs. A total of 317 circRNAs, 104 lncRNAs, and 135 mRNAs showed significant differential expression in UCB-derived exosomes of preterm infants with BPD compared with those with NBPD. Gene Ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were conducted to examine differentially expressed exosomal circRNAs, lncRNAs, and mRNAs. The results showed that the GO terms and KEGG pathways mostly involving differentially expressed exosomal RNAs were closely associated with endothelial or epithelial cell development. In vitro, CCK-8 and Western blot assays revealed that LPS remarkably inhibited the viability and promoted inflammatory responses (TNF-α and IL-1ß) of BEAS-2B cells or HUVECs. The expression levels of circRNAs hsa_circ_0049170 and hsa_circ_0087059 were upregulated in LPS-induced BEAS-2B cells; the expression level of hsa_circ_0086913 was upregulated and that of hsa_circ_0065188 was downregulated in LPS-induced HUVECs. Moreover, the expression level of lncRNA SNHG20 was upregulated and that of LINC00582 was downregulated in LPS-induced BEAS-2B cells. Further, 455 circRNA/lncRNA-miRNA-mRNA interaction networks were predicted, including hsa_circ_0086913/hsa-miR-103a-3p/transmembrane 4 L six family member 1 (TM4SF1) and lncRNA-SNHG20/hsa-miR-6720-5p/spermine synthase (SMS) networks, which may take part in BPD. CONCLUSION: This study provided a systematic perspective on UCB-derived exosomal circRNAs and lncRNAs and laid an important foundation for further investigating the potential biological functions of exosomal circRNAs and lncRNAs in BPD. WHAT IS KNOWN: ⢠BPD represents a multifactorial chronic pulmonary pathology and a major factor causing premature illness and death. ⢠The therapeutic role of exosomes in BPD has been feverishly investigated, and exosomal RNAs were ignored. WHAT IS NEW: ⢠The profiles of UCB-derived exosomal circRNAs, lncRNAs, and mRNAs were performed. ⢠Several differentially expressed circRNAs and lncRNAs were identified in LPS-induced BEAS-2B cells and HUVECs.
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Displasia Broncopulmonar , Exosomas , MicroARNs , ARN Largo no Codificante , Displasia Broncopulmonar/genética , Células Endoteliales/metabolismo , Exosomas/genética , Exosomas/metabolismo , Sangre Fetal/metabolismo , Humanos , Recién Nacido , Recien Nacido Prematuro , Lipopolisacáridos , MicroARNs/genética , ARN Circular/genética , ARN Largo no Codificante/genética , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
Adipose tissue of ketotic dairy cows exhibits greater lipolytic rate and signs of inflammation, which further aggravate the metabolic disorder. In nonruminants, the endoplasmic reticulum (ER) is a key organelle coordinating metabolic adaptations and cellular functions; thus, disturbances known as ER stress lead to inflammation and contribute to metabolic disorders. Enhanced activity of diacylglycerol O-acyltransferase 1 (DGAT1) in murine adipocytes undergoing lipolysis alleviated ER stress and inflammation. The aim of the present study was to investigate the potential role of DGAT1 on ER stress and inflammatory response of bovine adipose tissue in vivo and in vitro. Adipose tissue and blood samples were collected from cows diagnosed as clinically ketotic (n = 15) or healthy (n = 15) following a veterinary evaluation based on clinical symptoms and serum concentrations of ß-hydroxybutyrate, which were 4.05 (interquartile range = 0.46) and 0.52 mM (interquartile range = 0.14), respectively. Protein abundance of DGAT1 was greater in adipose tissue of ketotic cows. Among ER stress proteins measured, ratios of phosphorylated PKR-like ER kinase (p-PERK) to PERK and phosphorylated inositol-requiring enzyme 1 (p-IRE1) to IRE1, and protein abundance of cleaved ATF6 protein were greater in adipose tissue of ketotic cows. Furthermore, ratios of phosphorylated RELA subunit of NF-κB (p-RELA) to RELA and phosphorylated c-jun N-terminal kinase (p-JNK) to JNK were greater, whereas protein abundance of NF-κB inhibitor α (NFKBIA) was lower in adipose tissue of ketotic cows. In addition, mRNA abundance of proinflammatory cytokines including TNF and IL-6 was greater in adipose tissue of ketotic cows. To better address mechanistic aspects of these responses, primary bovine adipocytes isolated from the harvested adipose tissue of healthy cows were subjected to lipolysis-stimulating conditions via incubation with 1 µM epinephrine (EPI) for 2 h. In another experiment, adipocytes were cultured with DGAT1 overexpression adenovirus and DGAT1 small interfering RNA for 48 h, respectively, followed by EPI (1 µM) exposure for 2 h. Treatment with EPI led to greater ratios of p-PERK to PERK, p-IRE1 to IRE1, p-RELA to RELA, p-JNK to JNK, and cleaved ATF6 protein, whereas EPI stimulation inhibited protein abundance of NFKBIA. Furthermore, treatment with EPI upregulated the secretion of proinflammatory cytokines into culture medium, including TNF-α and IL-6. Overexpression of DGAT1 in EPI-treated adipocytes attenuated ER stress, the activation of NF-κB and JNK signaling pathways, and the secretion of inflammatory cytokines. In contrast, silencing DGAT1 further aggravated EPI-induced ER stress and inflammatory responses. Overall, these data indicated that activation of DGAT1 may act as an adaptive mechanism to dampen metabolic dysregulation in adipose tissue. As such, it contributes to relief from ER stress and inflammatory responses.
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Cetosis , Enfermedades de los Roedores , Femenino , Bovinos , Animales , Ratones , Ácido 3-Hidroxibutírico , Diacilglicerol O-Acetiltransferasa/metabolismo , Estrés del Retículo Endoplásmico , Inhibidor NF-kappaB alfa/metabolismo , FN-kappa B/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Cetosas/metabolismo , Cetosas/farmacología , ARN Interferente Pequeño/metabolismo , Interleucina-6/metabolismo , Cetosis/veterinaria , Tejido Adiposo/metabolismo , Citocinas/metabolismo , Inflamación/veterinaria , Inflamación/metabolismo , Proteínas Serina-Treonina Quinasas , Proteínas de Choque Térmico/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Epinefrina/farmacología , ARN Mensajero/metabolismo , Inositol/metabolismo , Inositol/farmacología , Enfermedades de los Roedores/metabolismoRESUMEN
Long non-coding RNAs (lncRNAs) contribute to disease pathogenesis and drug treatment effects. Both emodin and dexamethasone (DEX) have been used for treating severe acute pancreatitis-associated acute lung injury (SAP-ALI). However, lncRNA regulation networks related to SAP-ALI pathogenesis and drug treatment are unreported. In this study, lncRNAs and mRNAs in the lung tissue of SAP-ALI and control rats, with or without drug treatment (emodin or DEX), were assessed by RNA sequencing. Results showed both emodin and DEX were therapeutic for SAP-ALI and that mRNA and lncRNA levels differed between untreated and treated SAP-ALI rats. Gene expression profile relationships for emodin-treated and control rats were higher than DEX-treated and -untreated animals. By comparison of control and SAP-ALI animals, more up-regulated than down-regulated mRNAs and lncRNAs were observed with emodin treatment. For DEX treatment, more down-regulated than up-regulated mRNAs and lncRNAs were observed. Functional analysis demonstrated both up-regulated mRNA and co-expressed genes with up-regulated lncRNAs were enriched in inflammatory and immune response pathways. Further, emodin-associated lncRNAs and mRNAs co-expressed modules were different from those associated with DEX. Quantitative polymerase chain reaction demonstrates selected lncRNA and mRNA co-expressed modules were different in the lung tissue of emodin- and DEX-treated rats. Also, emodin had different effects compared with DEX on co-expression network of lncRNAs Rn60_7_1164.1 and AABR07062477.2 for the blue lncRNA module and Nrp1 for the green mRNA module. In conclusion, this study provides evidence that emodin may be a suitable alternative or complementary medicine for treating SAP-ALI.
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Lesión Pulmonar Aguda/etiología , Emodina/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Redes Reguladoras de Genes/efectos de los fármacos , Pancreatitis/complicaciones , ARN Largo no Codificante/genética , ARN Mensajero/genética , Lesión Pulmonar Aguda/metabolismo , Lesión Pulmonar Aguda/patología , Animales , Biomarcadores , Biopsia , Biología Computacional/métodos , Citocinas/metabolismo , Modelos Animales de Enfermedad , Susceptibilidad a Enfermedades , Ontología de Genes , Mediadores de Inflamación/metabolismo , Masculino , RatasRESUMEN
The aim of this study was to investigate the impact of maternal hepatitis B virus (HBV) status on pregnancy complications and neonatal outcomes for women undergoing assisted reproductive technology (ART). A total of 7,011 pregnancies achieved by ART were included from a population-based database involving 523,111 pregnancies. Exposures of hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) among pregnant women were routinely tested at the first antenatal visit for all pregnancies. We collected pregnancy complications (e.g., gestational diabetes mellitus [GDM], intrahepatic cholestasis of pregnancy [ICP]), neonatal outcomes and confounding variables from the same database. Univariate and multivariate analyses by adjusting confounders were conducted to evaluate the impact of maternal HBV infection. Prevalence of HBsAg seropositivity (HBsAg+) was 11.34% (95% CI 10.6-12.1) and that of HBsAg and HBeAg co-seropositivity (HBsAg+HBeAg+) was 2.55% (2.1-3.0) among included population. Compared with HBsAg-group, ICP risk in the HBsAg+group was higher (4.03% vs. 1.79%; adjusted odds ratio [aOR] 2.49, 1.65-3.77). Similarly, ICP prevalence in the HBsAg+HBeAg+ group was higher than that in the HBsAg-HBeAg- group (6.47% vs. 1.61%; aOR 4.78, 2.28-9.98). No associations were found between maternal HBV infection (i.e., HBsAg+, HBsAg+HBeAg+, or HBsAg+HBeAg-) and other adverse outcomes for women undergoing ART (i.e., GDM, pre-eclampsia, placental previa, premature separation of placenta, premature rupture of membranes, preterm birth and low birthweight) in this study. In conclusion, maternal HBV infection (HBsAg+or HBsAg+HBeAg+) probably increase ICP risk, but may not associate with other pregnancy complications or neonatal outcomes for pregnant women who underwent ART.
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Hepatitis B , Complicaciones Infecciosas del Embarazo , Nacimiento Prematuro , Femenino , Hepatitis B/complicaciones , Hepatitis B/epidemiología , Antígenos de Superficie de la Hepatitis B , Antígenos e de la Hepatitis B , Virus de la Hepatitis B , Humanos , Recién Nacido , Transmisión Vertical de Enfermedad Infecciosa , Placenta , Embarazo , Complicaciones Infecciosas del Embarazo/epidemiología , Nacimiento Prematuro/epidemiología , Técnicas Reproductivas AsistidasRESUMEN
BACKGROUND: The prevalence of diabetes is increasing worldwide. Our study aimed to estimate the changing trends in the prevalence and incidence of diagnosed type 2 diabetes mellitus (T2DM) among Xiamen residents and the floating population using real-world data. METHOD: We used real-world data from the System of Xiamen Citizens Health Information from 2014 to 2019 to estimate the changing trends in the prevalence and incidence of diagnosed T2DM. The System included the diagnosis of diabetes and the prescription of hypoglycemic drugs. Prevalent cases of T2DM were individuals who were diagnosed with T2DM and/or using hypoglycemic drugs. Incident cases were individuals with diagnosed T2DM and/or using hypoglycemic drugs in 2014 or 2019 who had not been diagnosed and/or did not use hypoglycemic drugs in the past. RESULTS: In 2014 and 2019, the prevalence of T2DM in Xiamen was 4.04 and 4.84%, respectively. In 2014 and 2019, the incidence rate of T2DM in Xiamen was 14.1 per 1000 person-year and 15.0 per 1000 person-year, respectively. There was a significant increase in both the prevalence (Prevalence difference: 0.80, 95%CI 0.76-0.83%, P < 0.001) and the incidence of T2DM (Incidence difference: 0.9, 95%CI 0.7-1.1, P < 0.001). in Xiamen. The prevalence and incidence of T2DM in people aged 18-39 increased significantly (P < 0.001), while the prevalence and incidence of T2DM in people aged 40-69 reduced significantly (P < 0.001). CONCLUSIONS: There was a significant increase in the prevalence and incidence of T2DM in Xiamen from 2014 to 2019 especially among those with younger age.
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Diabetes Mellitus Tipo 2/epidemiología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , China/epidemiología , Femenino , Historia del Siglo XXI , Humanos , Incidencia , Masculino , Persona de Mediana Edad , Prevalencia , Factores de Riesgo , Adulto JovenRESUMEN
Adipose tissue concentration of reactive oxygen species (ROS) increases in dairy cows with ketosis, suggesting that the tissue experiences oxidative stress. Autophagy, an adaptive response to cellular stress, has been shown to promote survival and plays a critical role in antioxidant responses. Dysregulation of adenosine 5'-monophosphate-activated protein kinase (AMPK) is closely related to antioxidant responses and autophagy of adipocytes in animal models of metabolic disorders, but its role in bovine adipose tissue during periods of stress is unknown. We hypothesized that AMPK may play important roles in the regulation of oxidative stress in adipose tissue of ketotic cows. Specific objectives were to evaluate autophagy status and AMPK activity in adipose tissue of ketotic cows, and their link with oxidative stress in isolated bovine adipocytes. Selection of 15 healthy and 15 clinically ketotic Holstein cows at 17 (±4) d postpartum was performed after a thorough veterinary evaluation for clinical symptoms and also based on serum ß-hydroxybutyrate concentrations before collection of subcutaneous adipose tissue samples. Primary cultures of bovine adipocytes isolated from the harvested adipose tissue were stimulated with varying concentrations of H2O2 (0, 50, 100, 200, or 400 µM) for 2 h. In another experiment, adipocytes were cultured with the AMPK activator A769662 or adenovirus-containing small interfering RNA (ad-AMPKα-siRNA) for 3 or 48 h, respectively, followed by H2O2 exposure (200 µM) for 2 h. Compared with healthy cows, clinical ketosis led to increased abundance of AMPK and nuclear factor erythroid-derived 2-like 2 (NFE2L2), but lower abundance of Kelch-like ECH-associated protein 1 (KEAP1) in adipose tissue. Abundance of the key proautophagy proteins Beclin1, sequestosome 1 (SQSTM1), autophagy-related gene 7 (ATG7), ATG5, and ratio of microtubule-associated protein light chain 3 (LC3) II to LC3I were greater in adipose tissue of ketotic cows. In bovine adipocytes, treatment with H2O2 induced accumulation of ROS and malondialdehyde (MDA), whereas H2O2 stimulation inhibited activities of the antioxidant enzymes glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD). Addition of AMPK activator A769662 increased antioxidant response via activating NFE2L2 and its downstream targets heme oxygenase 1 (HMOX1), superoxide dismutase 1 (SOD1), catalase (CAT), and glutathione-S-transferase (GST) to improve H2O2-induced oxidative stress in adipocytes. Simultaneously, activation of AMPK increased abundance of Beclin1, SQSTM1, ATG7, ATG5, and ratio of LC3II to LC3I. In contrast, inhibition of AMPK downregulated abundance of NFE2L2, HMOX1, SOD1, CAT, Beclin1, SQSTM1, ATG7, ATG5, and ratio of LC3II to LC3I, and further aggravated H2O2-induced oxidative stress. Overall, these data indicate that activation of AMPK, as an adaptive mechanism for acute metabolic regulation of adipose tissue homeostasis, can induce antioxidant responses and autophagy, and further reduce oxidative stress in bovine adipocytes.
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Antioxidantes , Factor 2 Relacionado con NF-E2 , Adenosina , Adipocitos/metabolismo , Animales , Autofagia , Bovinos , Femenino , Peróxido de Hidrógeno , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Estrés Oxidativo , Proteínas Quinasas , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Preadipocyte proliferation and differentiation are critical for normal adipose tissue development, including achieving a mature phenotype, characterized by its ability to accumulate triacylglycerol and release fatty acids. In nonruminants, it is well known that all-trans retinoic acid (ATRA), the most-active form of vitamin A, helps regulate proliferation, differentiation, and apoptosis in several types of cells including adipocytes. The purpose of this study was to evaluate the effect of ATRA on proliferation, apoptosis, differentiation, and lipolysis of primary bovine adipocytes isolated from subcutaneous adipose tissue of 5 healthy Holstein cows at 17 (±4 standard deviations) d postpartum. Cells were stimulated with increasing concentrations of ATRA (0.2, 2, and 20 nM) at the preconfluent (2 d) and postconfluent (8 d) preadipocyte stage or at the mature adipocyte stage (2 d). All concentrations of ATRA inhibited preconfluent preadipocyte proliferation with decreased proportion of S-phase cells and reduced protein abundance of cyclins (CCND1, CCND2, CCND3, CCNE1) and cyclin-dependent kinases (CDK2, CDK4, CDK6). Compared with vehicle, ATRA treatment induced apoptosis in preconfluent preadipocytes. Additionally, ATRA (0.2, 2, and 20 nM) supplementation also inhibited differentiation of postconfluent preadipocytes through downregulation of protein abundance of PPARγ and C/EBPα. After induction of differentiation, basal lipolysis in mature adipocytes increased upon treatment with all concentrations of ATRA. However, data on phosphorylated hormone-sensitive lipase or PLIN1 indicated that ATRA had no effect on epinephrine-stimulated lipolysis in mature adipocytes. Overall, these results demonstrate that ATRA might inhibit lipid accumulation by suppressing preadipocyte proliferation and differentiation, subsequently leading to apoptosis in postconfluent preadipocytes and promoting basal lipolysis in mature adipocytes. Overall, these in vitro responses provide some insights into the potential for nutritional management to modulate adipose tissue lipolysis, particularly in overconditioned cows during the dry period, which are more susceptible to suffer metabolic disorders due to excessive fat mobilization postpartum.
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Adipocitos , Lipólisis , Animales , Bovinos , Diferenciación Celular , Proliferación Celular , Femenino , Tretinoina/farmacologíaRESUMEN
Ketosis is a serious metabolic disorder characterized by systemic and hepatic oxidative stress, inflammation, and apoptosis, as well as reduced milk yield. Because of the paucity of data on mammary responses during ketosis, the aim of this study was to evaluate alterations in oxidative stress, NF-κB signaling, NLRP3 inflammasome, and caspase apoptotic pathways in mammary gland of dairy cows with ketosis. Blood, mammary gland tissue, and milk samples were collected from healthy cows [Control, blood concentration of ß-hydroxybutyrate (BHB) <0.6 mM, n = 10] and cows with subclinical ketosis (SCK, blood concentration of BHB >1.2 mM and <3 mM, n = 10) or clinical ketosis (CK, blood concentration of BHB >3 mM, n = 10) at median 8 d in milk (range = 6-12). Compared with Control, serum concentration of glucose was lower (3.91 vs. 2.86 or 2.12 mM) in cows with SCK or CK, whereas concentrations of fatty acids (0.25 vs. 0.57 or 1.09 mM) and BHB (0.42 vs. 1.81 or 3.85 mM) were greater. Compared with Control, the percentage of milk fat was greater in cows with SCK or CK. In contrast, the percentage of milk protein was lower in cows with SCK or CK. We detected no differences in milk lactose content across groups. Compared with Control, activities of glutathione peroxidase, superoxide dismutase, and catalase were lower in mammary gland tissue of cows with SCK or CK. In contrast, concentrations of hydrogen peroxide and malondialdehyde were greater in cows with SCK or CK. Compared with Control, mRNA abundances of TNFA, IL6, and IL1B were greater in mammary tissues of cows with SCK or CK. In addition, activity of IKKß and the ratio of phosphorylated inhibitor of κBα to IκBα, and of phosphorylated NF-κB p65 to NF-κB p65, were also greater in mammary tissues of cows with SCK or CK. Subclinical or clinical ketosis also led to greater activity of caspase 1 and protein abundance of caspase 1, NLRP3, Bax, caspase 3, and caspase 9. In contrast, abundance of the antiapoptotic protein was lower in SCK or CK cows. The data indicate that the mammary gland of SKC or CK cows undergoes severe oxidative stress, inflammation, and cell death.
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Enfermedades de los Bovinos/metabolismo , Cetosis/veterinaria , Glándulas Mamarias Animales/metabolismo , FN-kappa B/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Estrés Oxidativo/fisiología , Ácido 3-Hidroxibutírico/sangre , Animales , Apoptosis/fisiología , Caspasas/metabolismo , Bovinos , Femenino , Inflamasomas/metabolismo , Inflamación/metabolismo , Inflamación/veterinaria , Cetosis/metabolismo , Cetosis/patología , Lactancia/fisiología , Glándulas Mamarias Animales/química , Glándulas Mamarias Animales/patología , Leche/química , Transducción de SeñalRESUMEN
Bovine mammary epithelial cells undergo an increase in metabolic rate, mitochondrial dysfunction, and oxidative stress after calving. Nuclear factor erythroid 2-related factor 2 (NFE2L2), a master regulator of cellular redox homeostasis, plays crucial roles in the regulation of mitochondrial function. The objective of this study was to investigate the role of NFE2L2 on mitochondrial function in bovine mammary epithelial cells under hyperlipidemic conditions. Three experiments were conducted as follows: (1) the immortalized bovine mammary epithelial cell line MAC-T was treated with various concentrations of free fatty acids (FFA; 0, 0.6, 1.2, or 2.4 mM) for 24 h to induce stress; (2) MAC-T cells were transfected with small interfering RNA targeting NFE2L2 (si-NFE2L2) and scrambled nontarget negative control (si-Control) for 48 h; and (3) MAC-T cells were pretreated with 10 µM sulforaphane (SFN), an activator of NFE2L2, for 24 h followed by treatment with 1.2 mM FFA for an additional 24 h. Results indicated that exogenous FFA challenge induced linear and quadratic increases in concentrations of mitochondrial reactive oxygen species (ROS). Compared with 0 mM FFA, mitochondrial membrane potential, mRNA abundance of oxidative phosphorylation complexes (CO I-V), protein abundance of nuclear respiratory factor 1 (NRF1), peroxisome proliferator-activated receptor γ coactivator 1 α (PGC-1α), mitochondrial transcription factor A (TFAM), and NFE2L2 along with the contents of ATP, mitochondrial DNA (mtDNA), and total mitochondria were greater in the MAC-T challenged with 0.6 mM FFA group, but lower in the 1.2 and 2.4 mM FFA cultures. Knockdown of NFE2L2 via small interfering RNA led to greater mitochondrial ROS content and lower mitochondrial membrane potential along with contents of ATP, mtDNA, and total mitochondria. The SFN pretreatment upregulated protein abundance of NFE2L2 and attenuated the downregulation of NFE2L2 induced by FFA. Pretreatment with SFN attenuated the downregulation induced by FFA of PGC-1α, NRF1, and TFAM protein abundance along with contents of mtDNA and total mitochondria. Furthermore, SFN pretreatment attenuated the upregulation of mitochondrial ROS content, the downregulation of mitochondrial membrane potential, and the decreases in ATP, mtDNA, and mitochondrial content induced by FFA. Overall, data indicated that FFA inhibit NFE2L2, resulting in mitochondrial dysfunction and ROS production in bovine mammary epithelial cells. Thus, NFE2L2 may be a promising therapeutic target against metabolic challenge-driven mitochondrial dysfunction and oxidative stress in bovine mammary epithelial cells.
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Ácidos Grasos no Esterificados , Factor 2 Relacionado con NF-E2 , Estrés Oxidativo , Animales , Bovinos , Células Epiteliales , Ácidos Grasos no Esterificados/metabolismo , Femenino , Glándulas Mamarias Animales/metabolismo , Mitocondrias/metabolismo , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Dairy cows with ketosis exhibit signs of pronounced adipose tissue lipolysis and systemic inflammation, both of which exacerbate this metabolic disorder. In nonruminants, CIDEC plays a pivotal role in the formation of large unilocular lipid droplets. The present study aimed to ascertain the role of CIDEC in the lipolytic and inflammatory response of white adipose tissue (WAT) in vivo and in vitro. Subcutaneous adipose tissue and blood samples were collected from 15 healthy cows (blood ß-hydroxybutyrate concentration < 1.2 mM) and 15 cows with clinical ketosis (blood ß-hydroxybutyrate concentration > 3.0 mM) that had a similar number of lactations (median = 3, range = 2-4) and days in milk (median = 6 d, range = 3-9). Adipocytes isolated from 5 healthy Holstein calves (1 d old, female, 30-40 kg) were used for in vitro studies. Isolated adipocytes were treated with 0, 0.1, 1, or 10 ng/mL TNF-α for 3 h, transfected with CIDEC small interfering RNA for 48 h, or transfected with CIDEC overexpression adenovirus for 48 h followed by treatment with TNF-α (0.1 ng/mL) for 3 h. Serum concentrations of fatty acids were greater, and dry matter intake, milk yield, and serum glucose concentrations lower in cows with clinical ketosis. Protein and mRNA abundance of CIDEC were lesser in subcutaneous WAT of clinically ketotic versus healthy cows. Furthermore, the ratio of phosphorylated hormone sensitive lipase (p-LIPE) to LIPE, phosphorylated RELA (p-RELA) to RELA, and protein abundance of PNPLA2 and phosphorylated inhibitor of κBα (p-NFKBIA) were greater in dairy cows with clinical ketosis. The mRNA abundance of proinflammatory cytokines TNFA and IL1B were greater, and the anti-inflammatory cytokine IL10 was lower in WAT of dairy cows with clinical ketosis. In calf adipocytes, exogenous TNF-α (0.1, 1, or 10 ng/mL) decreased protein and mRNA abundance of CIDEC. In addition, exogenous TNF-α or knockdown of CIDEC reduced the secretion of the anti-inflammatory cytokine IL-10, but increased the ratio of p-LIPE to LIPE, p-RELA to RELA, protein abundance of PNPLA2 and p-NFKBIA, glycerol content, and the secretion of IL-1ß in calf adipocytes. Overexpression of CIDEC in TNFα-treated adipocytes attenuated lipolysis and activation of the NF-κB signaling pathway. Overall, these data suggest that inhibition of lipid droplet-associated protein CIDEC by TNF-α contributes to the pronounced lipolysis and inflammation of calf adipocytes, and CIDEC is a relevant target in clinically ketotic cows.