J F Fiallo Diez, A P Tegeler, C G Flesher, T C Michelotti, H Ford, M N Hoque, B Bhattarai, O J Benitez, G F Christopher, C Strieder-Barboza
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Tissue samples were utilized for transcriptome analysis, targeted RT-qPCR for adipogenic markers, adipocyte sizing, assessment of viscoelastic properties and collagen accumulation, and then decellularized for native ECM isolation. For in vitro analyses, SAT and VAT samples were digested via collagenase, and ASPCs cultured for metabolic analysis. Adipogenic capacity was assessed by adipocyte size, quantification of ASPCs in stromal vascular fraction (SVF) via flow cytometry, and gene expression of adipogenic markers. In addition, functional assays including lipolysis and glucose uptake were performed to further characterize SAT and VAT adipocyte metabolic function. Data were analyzed using SAS (version 9.4; SAS institute Inc., Cary, NC) and GraphPad Prism 9. Subcutaneous AT adipogenic capacity was greater than VAT's, as indicated by increased ASPCs abundance, increased magnitude of adipocyte ADIPOQ and FASN expression during differentiation, and higher adipocyte lipid accumulation as shown by an increased proportion of larger adipocytes and abundance of lipid droplets. Rheologic analysis revealed that VAT is stiffer than SAT, which led us to hypothesize that differences between SAT and VAT adipogenic capacity were partly mediated by depot-specific ECM microenvironment. Thus, we studied depot-specific ECM-adipocyte crosstalk using a 3D model with native ECM (decellularized AT). Subcutaneous AT and VAT ASPCs were cultured and differentiated into adipocytes within depot-matched and mis-matched ECM for 14d, followed by ADIPOQ expression analysis. Visceral AT ECM impaired ADIPOQ expression in SAT cells. Our results demonstrate that SAT is more adipogenic than VAT and suggest that divergences between SAT and VAT adipogenesis are partially mediated by the depot-specific ECM microenvironment.</p>","PeriodicalId":354,"journal":{"name":"Journal of Dairy Science","volume":null,"pages":null},"PeriodicalIF":3.7000,"publicationDate":"2024-07-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Extracellular matrix modulates depot-specific adipogenic capacity in adipose tissue of dairy cattle.\",\"authors\":\"J F Fiallo Diez, A P Tegeler, C G Flesher, T C Michelotti, H Ford, M N Hoque, B Bhattarai, O J Benitez, G F Christopher, C Strieder-Barboza\",\"doi\":\"10.3168/jds.2024-25040\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>Adipose tissue (AT) expands through both hyperplasia and hypertrophy. During adipogenesis, adipose stromal and progenitor cells (ASPCs) proliferate and then accumulate lipids, influenced by the local AT microenvironment. Increased adipogenic capacity is desirable as it relates to metabolic health, especially in transition dairy cows where excess free fatty acids in circulation can compromise metabolic and immune health. Our aim was to elucidate the depot-specific adipogenic capacity and ECM properties of subcutaneous (SAT) and visceral (VAT) AT of dairy cows and define how the ECM affects adipogenesis. Flank SAT and omental VAT samples were collected from dairy cows in a local abattoir. Tissue samples were utilized for transcriptome analysis, targeted RT-qPCR for adipogenic markers, adipocyte sizing, assessment of viscoelastic properties and collagen accumulation, and then decellularized for native ECM isolation. For in vitro analyses, SAT and VAT samples were digested via collagenase, and ASPCs cultured for metabolic analysis. Adipogenic capacity was assessed by adipocyte size, quantification of ASPCs in stromal vascular fraction (SVF) via flow cytometry, and gene expression of adipogenic markers. In addition, functional assays including lipolysis and glucose uptake were performed to further characterize SAT and VAT adipocyte metabolic function. Data were analyzed using SAS (version 9.4; SAS institute Inc., Cary, NC) and GraphPad Prism 9. Subcutaneous AT adipogenic capacity was greater than VAT's, as indicated by increased ASPCs abundance, increased magnitude of adipocyte ADIPOQ and FASN expression during differentiation, and higher adipocyte lipid accumulation as shown by an increased proportion of larger adipocytes and abundance of lipid droplets. Rheologic analysis revealed that VAT is stiffer than SAT, which led us to hypothesize that differences between SAT and VAT adipogenic capacity were partly mediated by depot-specific ECM microenvironment. Thus, we studied depot-specific ECM-adipocyte crosstalk using a 3D model with native ECM (decellularized AT). Subcutaneous AT and VAT ASPCs were cultured and differentiated into adipocytes within depot-matched and mis-matched ECM for 14d, followed by ADIPOQ expression analysis. Visceral AT ECM impaired ADIPOQ expression in SAT cells. 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引用次数: 0
摘要
脂肪组织(AT)通过增生和肥大两种方式扩张。在脂肪生成过程中,脂肪基质细胞和祖细胞(ASPCs)会增殖,然后受当地脂肪组织微环境的影响积聚脂质。脂肪生成能力的提高与新陈代谢健康有关,因此是可取的,尤其是在转型期奶牛中,循环中过量的游离脂肪酸会损害新陈代谢和免疫健康。我们的目的是阐明奶牛皮下(SAT)和内脏(VAT)AT 的特异性脂肪生成能力和 ECM 特性,并确定 ECM 如何影响脂肪生成。从当地屠宰场采集了奶牛腹侧 SAT 和网膜 VAT 样本。组织样本用于转录组分析、脂肪生成标记物的靶向 RT-qPCR、脂肪细胞大小、粘弹性能和胶原积累评估,然后进行脱细胞以分离原生 ECM。在体外分析中,通过胶原酶消化 SAT 和 VAT 样品,培养 ASPC 进行代谢分析。通过脂肪细胞的大小、流式细胞术对基质血管部分(SVF)中 ASPC 的定量以及脂肪生成标志物的基因表达来评估脂肪生成能力。此外,还进行了脂肪分解和葡萄糖摄取等功能测试,以进一步确定 SAT 和 VAT 脂肪细胞代谢功能的特征。数据使用 SAS(9.4 版;SAS Institute Inc.皮下 AT 的成脂能力高于 VAT,表现在 ASPCs 数量增加、分化过程中脂肪细胞 ADIPOQ 和 FASN 表达量增加,以及脂肪细胞脂质积累增加,表现在较大脂肪细胞比例增加和脂滴数量增加。流变学分析表明,VAT 比 SAT 硬,因此我们假设 SAT 和 VAT 成脂能力的差异部分是由脂肪库特异性 ECM 微环境介导的。因此,我们使用具有原生 ECM(脱细胞 AT)的三维模型研究了特定于脂肪库的 ECM-脂肪细胞串联。皮下 AT 和 VAT ASPC 在与内脏匹配和不匹配的 ECM 中培养和分化成脂肪细胞 14 天,然后进行 ADIPOQ 表达分析。内脏 AT ECM 影响了 SAT 细胞中 ADIPOQ 的表达。我们的研究结果表明,SAT 比 VAT 的脂肪生成能力更强,并表明 SAT 和 VAT 脂肪生成之间的差异部分是由内脏特异性 ECM 微环境介导的。
Extracellular matrix modulates depot-specific adipogenic capacity in adipose tissue of dairy cattle.
Adipose tissue (AT) expands through both hyperplasia and hypertrophy. During adipogenesis, adipose stromal and progenitor cells (ASPCs) proliferate and then accumulate lipids, influenced by the local AT microenvironment. Increased adipogenic capacity is desirable as it relates to metabolic health, especially in transition dairy cows where excess free fatty acids in circulation can compromise metabolic and immune health. Our aim was to elucidate the depot-specific adipogenic capacity and ECM properties of subcutaneous (SAT) and visceral (VAT) AT of dairy cows and define how the ECM affects adipogenesis. Flank SAT and omental VAT samples were collected from dairy cows in a local abattoir. Tissue samples were utilized for transcriptome analysis, targeted RT-qPCR for adipogenic markers, adipocyte sizing, assessment of viscoelastic properties and collagen accumulation, and then decellularized for native ECM isolation. For in vitro analyses, SAT and VAT samples were digested via collagenase, and ASPCs cultured for metabolic analysis. Adipogenic capacity was assessed by adipocyte size, quantification of ASPCs in stromal vascular fraction (SVF) via flow cytometry, and gene expression of adipogenic markers. In addition, functional assays including lipolysis and glucose uptake were performed to further characterize SAT and VAT adipocyte metabolic function. Data were analyzed using SAS (version 9.4; SAS institute Inc., Cary, NC) and GraphPad Prism 9. Subcutaneous AT adipogenic capacity was greater than VAT's, as indicated by increased ASPCs abundance, increased magnitude of adipocyte ADIPOQ and FASN expression during differentiation, and higher adipocyte lipid accumulation as shown by an increased proportion of larger adipocytes and abundance of lipid droplets. Rheologic analysis revealed that VAT is stiffer than SAT, which led us to hypothesize that differences between SAT and VAT adipogenic capacity were partly mediated by depot-specific ECM microenvironment. Thus, we studied depot-specific ECM-adipocyte crosstalk using a 3D model with native ECM (decellularized AT). Subcutaneous AT and VAT ASPCs were cultured and differentiated into adipocytes within depot-matched and mis-matched ECM for 14d, followed by ADIPOQ expression analysis. Visceral AT ECM impaired ADIPOQ expression in SAT cells. Our results demonstrate that SAT is more adipogenic than VAT and suggest that divergences between SAT and VAT adipogenesis are partially mediated by the depot-specific ECM microenvironment.
期刊介绍:
The official journal of the American Dairy Science Association®, Journal of Dairy Science® (JDS) is the leading peer-reviewed general dairy research journal in the world. JDS readers represent education, industry, and government agencies in more than 70 countries with interests in biochemistry, breeding, economics, engineering, environment, food science, genetics, microbiology, nutrition, pathology, physiology, processing, public health, quality assurance, and sanitation.