Journal: Science Advances

Article Title: Mapping cardiac remodeling in chronic kidney disease

doi: 10.1126/sciadv.adj4846

Figure Lengend Snippet: ( A ) Mice underwent sham surgery, STNx, or IRI and were sacrificed at 70 or 147 days after surgery. ( B ) Serum urea, serum creatinine (S-creatinine), heart weight, normalized to tibia length, and ejection fraction at the time point of sacrifice in sham (SH), STNx, and IRI mice. ( C ) Representative trichrome-stained images of kidney and heart. Scale bars, 50 μm. ( D ) UMAP embedding of nuclei isolated from left ventricle of all mice. n = 33,039 nuclei from n = 12 mice. ( E ) Expression of marker genes per cluster (cardiomyocytes: Ryr2 , Dmd , Myh6 , Fhl2 , and Ttn ; endothelial cells: Vwf , Cgnl1 , Smoc1 , Bmx , Pecam1 , Flt1 , Adgrl1 , and Prprb ; fibroblasts: Pdgra , Egfr , Dcn , Gsn , Col3a1 , and Col8a1 ; lymphatic endothelial cells: Flt4 , Prox1 , Lyve1 , and Pdpn ; lymphocytes: Btla , Cd2 , Bcl11b , and Sell ; macrophages: Mrc1 , Lyz2 , Cd86 , Cd68 , and Cd80 ; mesothelial cells: Muc16 , Msln , and Upk3b ; monocytes: Cd45 , Csfr1r; RBC: Lock2 , and Cd36 ; pericytes: Pdgfrb , Notch3 , Mcam , and Rgs5 ; and VSMC : Myh11 , Myl9 , and Lmod ). ( F ) PROGENGY pathway analysis in all cell types, STNx versus sham and IRI versus sham. ( G ) Predicted receptor-ligand interactions per group, sham, STNx, and IRI. * P < 0.05; ** P < 0.001. CM, cardiomyocytes; EC, endothelial cells; Fib, fibroblasts; Lym, lymphocytes; LEC, lymphatic endothelial cells; Mac, macrophages; Mes, mesothelial cells; Mon, monocytes; Per, pericytes; RBC, red blood cells; VSMC, vascular smooth muscle cells.

Article Snippet: Human AC16 cardiomyocytes (Merck Millipore, USA) were cultured in Dulbecco’s modified Eagle’s medium (DMEM)–F12 (D6434 Sigma-Aldrich, Germany) supplemented with 2 mM glutamine (TMS-002-C Millipore, USA), 12.5% fetal bovine serum (ES-009-B Millipore, USA), and 1% penicillin/streptomycin.

Techniques: Staining, Isolation, Expressing, Marker

Journal: Science Advances

Article Title: Mapping cardiac remodeling in chronic kidney disease

doi: 10.1126/sciadv.adj4846

Figure Lengend Snippet: ( A ) Representative confocal images of WGA-stained left ventricular myocardium from mice after sham surgery, STNx, or IRI. Scale bars, 20 μm. ( B ) Feret diameter of cardiomyocytes from sham, STNx, and IRI mice. ( C ) Hypertrophy gene set enrichment in cardiomyocytes 1 to 4 from sham, STNx, and IRI mice (GSEA). ( D ) Diffusion map (DM) embedding of cardiomyocytes 1 to 4 from sham, STNx, and IRI mice color-coded per condition, ( E ) See (D) but color-coded per cardiomyocyte cluster. ( F ) Differentiation trajectory toward hypertrophic cardiomyocytes within DM from (D) to (E). ( G ) Hypertrophy gene set enrichment in DM of cardiomyocytes from (D) to (E). ( H ) Genes sorted along the pseudotime trajectory of cardiomyocyte hypertrophy in (F); the color represents z score of the log-normalized gene expression across pseudotime (0 to 100) cells. ( I ) GO terms sorted along the pseudotime trajectory of cardiomyocyte hypertrophy in (F); the color represents the −log 10 transformed P value of GO terms derived from highly expressed genes across pseudotime (one to five) cells. ( J ) AC16 cardiomyocyte cell line, treated with 5 or 10% uremic serum, compared to healthy serum; planimetric analysis of cross-sectional area per cardiomyocyte from fluorescence microscopic images, TMRM staining. ( K ) Hypertrophic gene set, expressed in AC16 cells after treatment with 5% healthy or 5% uremic serum. ( L ) PROGENY pathway analysis in AC16 cells after treatment with 5% healthy or 5% uremic serum. ( M ) Planimetric analysis of cross-sectional area per cardiomyocyte from fluorescence microscopic images, TMRM staining, cells either treated with healthy serum, uremic serum + vehicle, uremic serum + ruxolitinib, or uremic serum + etanercept. * P < 0.05; # P < 0.001.

Article Snippet: Human AC16 cardiomyocytes (Merck Millipore, USA) were cultured in Dulbecco’s modified Eagle’s medium (DMEM)–F12 (D6434 Sigma-Aldrich, Germany) supplemented with 2 mM glutamine (TMS-002-C Millipore, USA), 12.5% fetal bovine serum (ES-009-B Millipore, USA), and 1% penicillin/streptomycin.

Techniques: Staining, Diffusion-based Assay, Gene Expression, Transformation Assay, Derivative Assay, Fluorescence

Journal: Science Advances

Article Title: Mapping cardiac remodeling in chronic kidney disease

doi: 10.1126/sciadv.adj4846

Figure Lengend Snippet: ( A ) Gene set enrichment of genes associated to UTs on all cell types; STNx versus sham (IRI versus sham is shown in fig. S13). ( B ) HUVEC tube formation assay, treatment with 5 or 10% uremic serum versus healthy control serum, quantification of isolated branches and bright-field microscopic images. ( C ) HUVEC tube formation assay, treatment with endothelin 1 (Edn1) or endothelin 1 neutralizing antibody, quantification of total mesh area and bright-field microscopic images. Scale bars, 500 μm. Sham: # P < 0.001. ( D ) Expression of endothelin converting enzyme 1 ( Ece1 ) in lymphatic endothelial cells and endothelial cells 1 to 3, per treatment group. ( E ) Size of AC16 cells after treatment with Edn1 or vehicle for 3 days. Scale bars, 25 μm.

Article Snippet: Human AC16 cardiomyocytes (Merck Millipore, USA) were cultured in Dulbecco’s modified Eagle’s medium (DMEM)–F12 (D6434 Sigma-Aldrich, Germany) supplemented with 2 mM glutamine (TMS-002-C Millipore, USA), 12.5% fetal bovine serum (ES-009-B Millipore, USA), and 1% penicillin/streptomycin.

Techniques: HUVEC Tube Formation Assay, Control, Isolation, Expressing

Journal: Journal of Translational Medicine

Article Title: CAV3 alleviates diabetic cardiomyopathy via inhibiting NDUFA10-mediated mitochondrial dysfunction

doi: 10.1186/s12967-024-05223-6

Figure Lengend Snippet: CAV3 was decreased in diabetic mice hearts and knock-down CAV3 induced mitochondria-dependent intrinsic apoptosis in cardiomyocytes. A Western blotting and associated quantitative analysis the CAV3 protein expression levels in the hearts of wt and db/db mice (n = 7 mice). B Representative immunohistochemical stains of CAV3 in the hearts of wt and db/db mice (n = 7 mice). C Representative Western blotting and quantitative analyses for CAV3, Bax, Cleaved caspase 3 and Bcl2 protein levels in cardiomyocytes treated with or without HGHF and with or without CAV3 downregulation (n = 4). D Representative Western blotting and quantitative analyses for the content of cytochrome C in cytoplasm treated with or without HGHF and with or without CAV3 downregulation (n = 3). E Flow cytometry and quantitative analysis of apoptosis cells by AnnexinV-FITC and propidium iodide (PI) staining in cardiomyocytes treated with or without HGHF and with or without CAV3 downregulation in vitro (n = 5). F Flow cytometry and quantification analysis of mitochondrial membrane potential by JC-1 in cardiomyocytes treated with or without HGHF and with or without CAV3 downregulation. High levels of green fluorescence (x-axis) represent reduced mitochondrial membrane potential (ΔΨm) and high levels of red fluorescence (y-axis) represents increased ΔΨm. A decrease in the red/green fluorescence is indicative of loss of ΔΨm (n = 6). G Normalized ATP production levels in cardiomyocytes treated with or without HGHF and with or without CAV3 downregulation in vitro (n = 6). H Representative confocal microscope of MitoSOX staining, immunofluorescence staining of 8-oxoG andquantitative analyses in cardiomyocytes treated with or without HGHF and with or without CAV3 downregulation (MitoSOX fluorescence, red; 8-oxoG fluorescence) (n = 5). I Immunofluorescence images of 8-oxoG and TOM20 in the AC16 cells treated with HGHF. AC16 cells were stained with 8-oxoG antibodies (Alexa Fluor 594), DAPI, and TOM20 (Alexa Fluor 488). Data are depicted as mean ± SD. *p < 0.05, **p < 0.01

Article Snippet: Human cardiomyocytes (AC16) and mouse cardiomyocytes (HL-1) were purchased from iCell Bioscience (Shanghai, China).

Techniques: Western Blot, Expressing, Immunohistochemical staining, Flow Cytometry, Staining, In Vitro, Membrane, Fluorescence, Microscopy, Immunofluorescence

Journal: Journal of Translational Medicine

Article Title: CAV3 alleviates diabetic cardiomyopathy via inhibiting NDUFA10-mediated mitochondrial dysfunction

doi: 10.1186/s12967-024-05223-6

Figure Lengend Snippet: Overexpression of CAV3 protected against cardiac dysfunction and cardiac remodeling in db/db mice. A Western blotting and associated quantitative analysis the CAV3 protein expression levels in the hearts of db/db mice with or without CAV3 overexpression in vivo (n = 7 mice). B – F The body weight, food intake and blood glucose levels of wt and db/db mice among each group were showed by a line chart (n = 7 mice). G Hemodynamic analysis (dp/dtmin) of wt mice and db/db mice with or without CAV3 overexpression. (n = 7 mice). H Representative images of echocardiograph (E/A) and relevant quantification analysis in wt mice and db/db mice with or without CAV3 overexpression (n = 7 mice). I The HW/TL ratio was quantitatively analyzed in wt mice and db/db mice with or without CAV3 overexpression (n = 7 mice). J Representative images of H&E staining and WGA staining to detect cardiomyocyte cross-sectional area, Sirius red staining to detect myocardial interstitial collagen and relevant quantification analysis in wt mice and db/db mice with or without CAV3 overexpression (n = 7 mice). Data are depicted as mean ± SD. *p < 0.05, **p < 0.01

Article Snippet: Human cardiomyocytes (AC16) and mouse cardiomyocytes (HL-1) were purchased from iCell Bioscience (Shanghai, China).

Techniques: Over Expression, Western Blot, Expressing, In Vivo, Staining

Journal: Journal of Translational Medicine

Article Title: CAV3 alleviates diabetic cardiomyopathy via inhibiting NDUFA10-mediated mitochondrial dysfunction

doi: 10.1186/s12967-024-05223-6

Figure Lengend Snippet: Overexpression of CAV3 prevented mitochondrial dysfunction in diabetic cardiomyocytes in vivo and in vitro models. A Representative transmission electron microscope images of cardiac mitochondria and quantification analysis of mitochondrial cristae number and the proportion of mitochondria with disorganized cristae in wt mice and db/db mice with or without CAV3 overexpression (n = 7 mice). B Normalized ATP production levels in cardiomyocytes of wt mice and db/db mice with or without CAV3 overexpression (n = 7 mice). C Flow cytometry analysis and quantification of mitochondrial membrane potential by JC-1 in cardiomyocytes treated with or without HGHF and with or without CAV3 overexpression. (n = 5). D Extracellular flux analysis of OCRs and respective quantitative analysis in cardiomyocytes treated with or without HGHF and with or without CAV3 overexpression (n = 3). E Representative images of DHE staining and quantitative analysis of ROS levels in the hearts of wt mice and db/db mice with or without CAV3 overexpression (n = 7 mice). F Representative images of immunofluorescence staining of 8-oxoG and quantitative analyses in cardiomyocytes treated with or without HGHF and with or without CAV3 overexpression (8-oxoG fluorescence, red) (n = 5). G Representative Western blotting and quantitative analyses for SOD2, Bcl2, Bax and Cleaved caspase 3 protein levels in cardiomyocytes of wt mice and db/db mice with or without CAV3 overexpression (n = 7 mice). H TUNEL assay by double staining with TUNEL (red) and DAPI (blue) detected apoptotic cells in mice hearts and quantitative analysis of wt mice and db/db mice with or without CAV3 overexpression (n = 7 mice). Data are depicted as mean ± SD. *p < 0.05, **p < 0.01

Article Snippet: Human cardiomyocytes (AC16) and mouse cardiomyocytes (HL-1) were purchased from iCell Bioscience (Shanghai, China).

Techniques: Over Expression, In Vivo, In Vitro, Transmission Assay, Microscopy, Flow Cytometry, Membrane, Staining, Immunofluorescence, Fluorescence, Western Blot, TUNEL Assay, Double Staining

Journal: Journal of Translational Medicine

Article Title: CAV3 alleviates diabetic cardiomyopathy via inhibiting NDUFA10-mediated mitochondrial dysfunction

doi: 10.1186/s12967-024-05223-6

Figure Lengend Snippet: CAV3 interacted with mitochondrial complex I by anchoring NDUFA10 and maintaining its stability. A IP assay was carried out using CAV3 antibody and Coomassie Brilliant blue was stained to indicate CAV3 bands. B Through bioinformatics methods, NDUFA10 and TARDBP were overlapped from the LC–MS/MS analysis, DCM-related Genes (DisGeNet & Gencards) and Mitochondrial dysfunction-related genes (GeneCards). C , D IP assay was carried out using CAV3 or NDUFA10 antibody in cardiomyocytes. E Representative Western blotting assessing the purity of the mitochondria isolated. F – G IP assay was carried out using CAV3 or NDUFA10 antibody in mitochondria isolated from AC16 cells. H Immunofluorescence images of cardiomyocytes were stained with mitochondria-targeting dye (Mito-Tracker, Deep Red, Alexa Fluor 647), DAPI, NDUFA10 antibodies (Alexa Fluor 594), and CAV3 antibodies (Alexa Fluor 488). I Western blotting and quantitative analyses protein levels in cardiomyocytes of wt mice and db/db mice with or without CAV3 overexpression (n = 7 mice). J The levels of NDUFA10 mRNA in cardiomyocytes treated with or without HGHF and with or without CAV3 overexpression (n = 6). K CAV3 plasmid (pcDNA3.1-CAV3) was transfected into cardiomyocytes using Lipofectamine 2000 and treated with HGHF. The transfected cells were treated with 100 µmol/L cycloheximide for 0, 2, 4, 6, 8, 10 h. The protein levels of NDUFA10 were shown by a line chart (n = 3). L Cardiomyocytes were treated with or without HGHF, CHX (100 µmol/L), and MG132 (10 µmol/L) or CQ (50 µmol/L) for 24 h. Quantitative analyses of NDUFA10 protein levels among each group (n = 3). M Immunofluorescence images of NDUFA10 and lysosomes-targeting dye (Lamp1) in the cardiomyocytes treated with or without CAV3 overexpression. Cardiomyocytes were stained with NDUFA10 antibodies (Alexa Fluor 594), DAPI, and Lamp1 (Alexa Fluor 488). Data are depicted as mean ± SD. *p < 0.05, **p < 0.01

Article Snippet: Human cardiomyocytes (AC16) and mouse cardiomyocytes (HL-1) were purchased from iCell Bioscience (Shanghai, China).

Techniques: Staining, Liquid Chromatography with Mass Spectroscopy, Western Blot, Isolation, Immunofluorescence, Over Expression, Plasmid Preparation, Transfection

Journal: Journal of Translational Medicine

Article Title: CAV3 alleviates diabetic cardiomyopathy via inhibiting NDUFA10-mediated mitochondrial dysfunction

doi: 10.1186/s12967-024-05223-6

Figure Lengend Snippet: CAV3 overexpression restored mitochondrial function partially relied on NDUFA10 in vivo and in vitro models. A Representative transmission electron microscope images of cardiac mitochondria and quantification analysis of mitochondrial cristae number and the proportion of mitochondria with disorganized cristae in different db/db mice groups (n = 7 mice). B Normalized ATP production levels in cardiomyocytes of db/db mice transfected with or without AAV-CAV3 or AAV-shNDUFA10 (n = 7 mice). C Flow cytometry and quantitative analysis of mitochondrial membrane potential by JC-1 in cardiomyocytes treated with HGHF and transfected with or without pc3.1-CAV3 or siNDUFA10 (n = 5). D Extracellular flux analysis of OCRs and respective quantitative analysis in cardiomyocytes treated with HGHF and transfected with or without pc3.1-CAV3 or siNDUFA10 (n = 3). E The activity of mitochondrial complex I in cardiomyocytes of db/db mice transfected with or without AAV-CAV3 or AAV-shNDUFA10 (n = 7 mice). F Representative images of DHE staining and quantitative analysis of ROS levels in the hearts of db/db mice transfected with or without AAV-CAV3 or AAV-shNDUFA10 (n = 7 mice). G Representative images of immunofluorescence staining of 8-oxoG and quantitative analyses in cardiomyocytes treated with HGHF and transfected with or without pc3.1-CAV3 or siNDUFA10 (8-oxoG fluorescence, red) (n = 5). H Representative Western blotting and quantitative analyses for SOD2, Bcl2, Bax and Cleaved caspase 3 protein levels in cardiomyocytes of db/db mice transfected with or without AAV-CAV3 or AAV-shNDUFA10 (n = 7 mice). I TUNEL assay by double staining with TUNEL (red) and DAPI (blue) detected apoptotic cells in the hearts and quantitative analysis of different db/db mice groups (n = 7 mice). Data are depicted as mean ± SD. *p < 0.05, **p < 0.01

Article Snippet: Human cardiomyocytes (AC16) and mouse cardiomyocytes (HL-1) were purchased from iCell Bioscience (Shanghai, China).

Techniques: Over Expression, In Vivo, In Vitro, Transmission Assay, Microscopy, Transfection, Flow Cytometry, Membrane, Activity Assay, Staining, Immunofluorescence, Fluorescence, Western Blot, TUNEL Assay, Double Staining

Journal: Journal of Translational Medicine

Article Title: CAV3 alleviates diabetic cardiomyopathy via inhibiting NDUFA10-mediated mitochondrial dysfunction

doi: 10.1186/s12967-024-05223-6

Figure Lengend Snippet: Cardiac-restricted NDUFA10 expression partially attenuated CAV3 benefits in diabetic cardiomyopathy. A Hemodynamic analysis (dp/dtmin) of db/db mice transfected with or without AAV-CAV3 or AAV-shNDUFA10 (n = 7 mice). B Representative images of echocardiograph (E/A) and relevant quantification analysis in db/db mice transfected with or without AAV-CAV3 or AAV-shNDUFA10 (n = 7 mice). C The HW/TL ratio was quantitatively analyzed in db/db mice transfected with or without AAV-CAV3 or AAV-shNDUFA10 (n = 7 mice). D Representative images of H&E staining and WGA staining to detect cardiomyocyte cross-sectional area, Sirius red staining to detect myocardial interstitial collagen and relevant quantification analysis in db/db mice transfected with or without AAV-CAV3 or AAV-shNDUFA10 (n = 7 mice). Data are depicted as mean ± SD. *p < 0.05, **p < 0.01

Article Snippet: Human cardiomyocytes (AC16) and mouse cardiomyocytes (HL-1) were purchased from iCell Bioscience (Shanghai, China).

Techniques: Expressing, Transfection, Staining

Journal: Journal of Translational Medicine

Article Title: CAV3 alleviates diabetic cardiomyopathy via inhibiting NDUFA10-mediated mitochondrial dysfunction

doi: 10.1186/s12967-024-05223-6

Figure Lengend Snippet: The schematic representation of the mechanisms by which CAV3 downregulation exacerbates diabetic cardiomyopathy via inducing NDUFA10-mediated mitochondrial dysfunction in diabetes. CAV3 expression is decreased in cardiomyocytes under diabetic conditions. Downregulation of CAV3 leads to the downregulation of NDUFA10, which is degraded via the lysosomal pathway. Consequently, the reduction of NDUA10 in mitochondria lead to a decrease in the mitochondrial complex I activity, resulting in mitochondrial dysfunction. As a result, ATP production decreases, mitochondrial ROS levels increase and cell apoptosis occurs, thereby leading to cardiac dysfunction

Article Snippet: Human cardiomyocytes (AC16) and mouse cardiomyocytes (HL-1) were purchased from iCell Bioscience (Shanghai, China).

Techniques: Expressing, Activity Assay