microarray analysis exosomes  (Exosome Diagnostics)

Journal: Signal Transduction and Targeted Therapy

Article Title: Natural killer cell-derived exosomal miR-1249-3p attenuates insulin resistance and inflammation in mouse models of type 2 diabetes

doi: 10.1038/s41392-021-00805-y

Figure Lengend Snippet: NK-derived exosomes from lean mice attenuate obesity-induced insulin resistance. a Fasting blood glucose of NCD and HFD mice after group feeding. b – d OGTT ( b ), ITT ( c ), and HOMA-IR index ( d ) of HFD mice and NCD mice ( n = 48) were recorded at 42 days after group feeding. HOMA-IR = Fasting blood glucose value × fasting serum insulin value/22.5. e Transmission electron micrographs of NK-derived exosomes. Scale bar, 100 nm. f NanoSight particle tracking analysis showing the particle size of NK-derived exosomes isolated from NCD and HFD mice. g Western blot assays of exosomal markers TSG101, HSP70, CD63, and CD9. h – k After blank liposomes, NCD-Exos, and HFD-Exos were separately injected into NCD or HFD mice via tail vein, fasting blood glucose ( h ), OGTT ( i ), ITT ( j ), and HOMA-IR ( k ) were assessed in each group. G1: HFD mice treated with blank liposomes ( n = 3); G2: HFD mice treated with HFD-Exos ( n = 3); G3: HFD mice treated with NCD-Exos ( n = 3); G4: NCD mice treated with blank liposomes ( n = 3); G5: NCD mice treated with HFD-Exos ( n = 3); and G6: NCD mice treated with NCD-Exos ( n = 3). l After NCD-Exos labeled with PKH26 were transferred into recipient mice, fluorescence images of the liver, islets, SATs, VATs, and skeletal muscles were observed by in vitro imaging system. Scale bar, 5 mm. m The weight of VATs from HFD mice treated with NCD-Exos, HFD-Exos, or blank liposomes. n Liver triglyceride content of HFD mice treated with NCD-Exos, HFD-Exos, or blank liposomes. o , p ELISA assays of IL-6, IL-1β, and TNF-α expression in VATs and livers of HFD mice treated with NCD-Exos, HFD-Exos, or blank liposomes. Experiments were performed at least in triplicate, and the results are shown as the mean ± s.d. Student’s t-test was used to analyze the data. (* P < 0.05; ** P < 0.01; *** P < 0.001)

Article Snippet: Exosomal miRNA microarray analysis was performed at CapitalBio Technology, Inc. (Beijing, China) using Agilent Mouse miRNA 8 × 60K V21.0 Microarray (Agilent Technologies, California, USA).

Techniques: Derivative Assay, Transmission Assay, Isolation, Western Blot, Liposomes, Injection, Labeling, Fluorescence, Muscles, In Vitro, Imaging, Enzyme-linked Immunosorbent Assay, Expressing

Journal: Signal Transduction and Targeted Therapy

Article Title: Natural killer cell-derived exosomal miR-1249-3p attenuates insulin resistance and inflammation in mouse models of type 2 diabetes

doi: 10.1038/s41392-021-00805-y

Figure Lengend Snippet: NK-derived exosomal miR-1249-3p mediates cellular insulin sensitivity and inflammation. a Microarray analysis of significantly expressed exosomal miRNAs between NCD-Exos and HFD-Exos was presented in a heatmap. b qRT-PCR assay of miR-1249-3p expression in splenic NK cells, NK-derived exosomes, and circulating exosomes from NCD or HFD mice. c NK cells transfected with a Cy3-labeled miR-1249-3p mimic were co-cultured with 3T3-L1 adipocytes or AML12 cells in a Transwell TM plate (membrane pore = 0.4 mm) plate. Scale bar, 100 μm. d – g After transfecting with miR-1249-3p mimic, miR-NC, inhibitor, and inh-NC, the glucose uptake content of 3T3-L1 adipocytes ( d ) and glucose production content of AML12 cells ( f ) were measured, and the concentrations of IL-6, TNF-α, and IL-1β secreted by 3T3-L1 adipocytes ( e ) and AML12 cells ( g ) were detected by ELISA. Experiments were performed at least in triplicate, and the results are shown as the mean ± s.d. Student’s t -test was used to analyze the data. (* P < 0.05; ** P < 0.01; *** P < 0.001)

Article Snippet: Exosomal miRNA microarray analysis was performed at CapitalBio Technology, Inc. (Beijing, China) using Agilent Mouse miRNA 8 × 60K V21.0 Microarray (Agilent Technologies, California, USA).

Techniques: Derivative Assay, Microarray, Quantitative RT-PCR, Expressing, Transfection, Labeling, Cell Culture, Membrane, Enzyme-linked Immunosorbent Assay

Journal: Signal Transduction and Targeted Therapy

Article Title: Natural killer cell-derived exosomal miR-1249-3p attenuates insulin resistance and inflammation in mouse models of type 2 diabetes

doi: 10.1038/s41392-021-00805-y

Figure Lengend Snippet: Exosomal miR-1249-3p directly targets SKOR1 to mediate insulin sensitivity. a The potential targets of miR-1249-3p were predicted by integrating the results of two databases (TargetScan and miRDB). b Western blot analysis of SKOR1 in 3T3-L1 adipocytes and AML12 cells with the indicated treatments. c The wild-type and a mutated type of binding site between miR-1249-3p and SKOR1. d Relative luciferase activity of AML12 cells in the presence of indicated treatments. e Western blot analysis of SKOR1 expression in the VATs and livers of HFD mice after NCD-Exos or HFD-Exos treatment. f – i The effect of sh-SKOR1 on glucose uptake capacity in 3T3-L1 adipocytes ( f ), glucose production capacity in AML12 cells ( g ), and expression levels of IL-6, TNF-α, and IL-1β ( h , i ). j – q Glucose uptake content of 3T3-L1 adipocytes ( j ) and glucose production content of AML12 cells ( n ) with the indicated treatments were measured, as well as the expression levels of IL-6, TNF-α, and IL-1β ( k – m and o – q ). Experiments were performed at least in triplicate, and the results are shown as the mean ± s.d. Student’s t -test was used to analyze the data. (* P < 0.05; ** P < 0.01; *** P < 0.001)

Article Snippet: Exosomal miRNA microarray analysis was performed at CapitalBio Technology, Inc. (Beijing, China) using Agilent Mouse miRNA 8 × 60K V21.0 Microarray (Agilent Technologies, California, USA).

Techniques: Western Blot, Binding Assay, Luciferase, Activity Assay, Expressing

Journal: Signal Transduction and Targeted Therapy

Article Title: Natural killer cell-derived exosomal miR-1249-3p attenuates insulin resistance and inflammation in mouse models of type 2 diabetes

doi: 10.1038/s41392-021-00805-y

Figure Lengend Snippet: MiR-1249-3p relieves insulin resistance and inflammation via the SKOR1-SMAD6-TLR4-NF-κB axis. a – c After transfection with a miR-1249-3p mimic, miR-NC, or specific siRNA to SKOR1 or SMAD6, qRT-PCR analysis of miR-1249-3p ( a ), SKOR1 ( b ) and SMAD6 ( c ) expression in 3T3-L1 adipocytes cells with the indicated treatments was performed. d , g The expression of p-p65 and p65 in 3T3-L1 adipocytes cells with the indicated treatments was performed by western blot. e , f , h IL-1β, IL-6, and TNF-α expression in 3T3-L1 adipocytes cells subjected to the indicated treatments was assessed by ELISA. i The signaling pathway through which NK-derived exosomal miR-1249-3p regulates insulin resistance in type 2 diabetes mice. Experiments were performed at least in triplicate, and the results are shown as the mean ± s.d. Student’s t -test was used to analyze the data. (* P < 0.05; ** P < 0.01; *** P < 0.001)

Article Snippet: Exosomal miRNA microarray analysis was performed at CapitalBio Technology, Inc. (Beijing, China) using Agilent Mouse miRNA 8 × 60K V21.0 Microarray (Agilent Technologies, California, USA).

Techniques: Transfection, Quantitative RT-PCR, Expressing, Western Blot, Enzyme-linked Immunosorbent Assay, Derivative Assay