Fig. 2: Effects of Nap1l1 Knockdown on DNA Damage, Inflammatory Signaling, and Cellular Function in NRCMs and Cardiac Fibroblasts.

A Immunofluorescence staining showing the levels of γ-H2ax protein in NRCMs with siCon or siNap1l1; B Representative immunofluorescence staining showing the levels of DAPI, dsDNA and cGAS in NRCMs with siCon or siNap1l1; C qPCR analysis of Mito and B2m in NRCMs with siCon or siNap1l1. N = 6 for each group, and two-tailed unpaired Student’s t-test was applied; D, E Western blot and quantification showing the levels of Sting, Tbk1, Irf3, Ifna, Ifnb, and phosphorylation of Sting, Tbk1, and Irf3 in NRCMs with siCon or siNap1l1. N = 3 for each group, and two-tailed unpaired Student’s t-test was applied; F qPCR analysis of Ifna, Ifnb, Il-6, and Il-1b in NRCMs with siCon or siNap1l1. N = 6 for each group, and two-tailed unpaired Student’s t-test was applied; G Phalloidin staining and TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling) staining and quantification in NRCMs with siCon or siNap1l1. Relative cell surface: siCON, n = 15; siNap1l1, n = 10. TUNEL+ cell: N = 5 for each group, and two-tailed unpaired Student’s t-test was applied; H Measurement of heart rate and diastolic function over time after adding siRNA. N = 4 for each group and two-way ANOVA with Tukey tests were used for correction of multiple comparisons; I qPCR analysis of Ifit1, Ifit2, Ifi3, Zbp1, Isg15, Stat1, and Il6 in influenced cardiomyocytes and Ifit1, Ifit2, Ifi3, Isg15, Irf7, Il6 and Il1b in influenced cardiac fibroblasts. N = 6 for each group, and two-tailed unpaired Student’s t-test was applied; J Representative plot of KI67 staining and quantification. N = 5 for each group, and two-tailed unpaired Student’s t-test was applied; K qPCR analysis of Irf7, Il6, Stat1, Isg15, Ifit1, and Ifit2 in influenced cardiac fibroblasts. N = 6 for each group and a two-tailed unpaired Student’s t-test was applied. Data are presented as mean values +/− SEM.