Journal: Nature

Article Title: The extracellular matrix protein agrin promotes heart regeneration in mice

doi: 10.1038/nature22978

Figure Lengend Snippet: a, b, Representative fields of heart cultures stained with DAPI (blue), cTnT (green) and Ki67 (red). White arrows indicate Ki67+cTnT+ cells. CMs, cardiomyocytes. c, d, Percentage of proliferating cardiomyocytes from P1 (c) or P7 (d) hearts in response to P1 and P7 ECM particles. n = 2,069 cardiomyocytes from three samples (c); n = 2,221 cardiomyocytes from four samples (d). e, f, Percentage of proliferating cardiomyocytes (Ki67+cTnT+) in response to P1 and P7 ECM in P1 (e) or P7 (f) cultures, with or without the broad MMP inhibitor (Marimastat). n = 3,480 cardiomyocytes from three samples (e); n = 23,445 cardiomyocytes from four samples (f). g, h, Percentage of P1 (g) or P7 (h) proliferating cardiomyocytes in response to MMP9- or MMP12-cleaved ECM fragments. n = 11,820 cardiomyocytes from four samples (g); n = 15,509 cardiomyocytes from four samples (h). i, qPCR of Agrn mRNA in P1 and P7 hearts. n = 8 P1 and 3 P7 hearts. j, Quantification of western blots for agrin from P1, P7 and 12-week-old (12W) adult heart lysates. A.U., arbitrary units. n = 3 samples per group. k, Images of P1 and P7 heart sections stained for agrin (green) and DAPI (blue). n = 3 samples per group. l, qPCR analysis of cardiac populations (FB, fibroblasts; CM, cardiomyocytes; EC, endothelial cells). n = 4 cardiomyocyte, 4 non-cardiomyocyte, 4 fibroblast, 4 non-fibroblast, 7 endothelial cells and 7 non-endothelial cell samples. Scale bars, 40 μm (a) and 10 μm (k). Data are presented as mean ±s.e.m. *P < 0.05, **P< 0.01, ***P< 0.001; statistical significance was calculated using ANOVA followed by a Dunnett’s post hoc test relative to the control group (c–h) or a Tukey’s post hoc test (j), statistical significance was calculated using a one-tailed t-test (i, l).

Article Snippet: Separation to distinct cardiac cell populations was performed by using the Neonatal Cardiac Endothelial Cell Isolation kit (130-104-183, Miltenyi Biotec), Neonatal Cardiac Fibroblast Cell Isolation kit (130-101-372, Miltenyi Biotec) or by Neonatal Cardiomyocyte Isolation kit (130-100-825, Miltenyi Biotec), according to the manufacturer’s instructions.

Techniques: Staining, Western Blot, Control, One-tailed Test

Journal: Microbial cell factories

Article Title: Secretion of tumoricidal human tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) by recombinant Lactococcus lactis: optimization of in vitro synthesis conditions.

doi: 10.1186/s12934-018-1028-2

Figure Lengend Snippet: Fig. 9 Selectivity of hsTRAIL-induced death to cancer cells. HCT116 cells and human cardiac fibroblasts were incubated for 48 h with increasing concentration of hsTRAIL present in supernatant from broth culture of L. lactis (hsTRAIL+). As controls in experimental setup were used: corresponding volumes of supernatants from broth culture of L. lactis (hsTRAIL−)—negative control; corresponding concentrations of recombinant human TRAIL (Peprotech)—positive control. Viability of cancer cells and non-malignant, cardiac fibroblasts was assessed by MTS assay. Results are presented as % of viability of cells incubated in standard culture medium only. Secreted hsTRAIL remained non-cytotoxic to fibroblasts (Fig. 9a), while decreased viability of cancer cells in a dose-dependent manner (Fig. 9b). Table below—concentration of hsTRAIL [ng/ml] in specified volume of supernatant. The bars indicate the mean value ± SD of three independent experiments, each performed in triplicates. Statistical significance was calculated using two-way ANOVA with Tukey’s multiple comparison post-test. *p < 0.05, **p < 0.01, ***p < 0.001

Article Snippet: Human primary proliferating cardiac fibroblasts were obtained from Cell Applications, Inc. (San Diego, CA).

Techniques: Incubation, Concentration Assay, Negative Control, Recombinant, Positive Control, MTS Assay, Comparison

Journal: Scientific Reports

Article Title: TRPC3-GEF-H1 axis mediates pressure overload-induced cardiac fibrosis

doi: 10.1038/srep39383

Figure Lengend Snippet: ( a–c ) Effect of Pyr3 (1 μM) on induction of fibrosis-related mRNAs (CTGF and TGF-β1 or -β2) by MS in human iPSC-derived cardiomyocytes (n = 3). Cells were subjected to 20% static-MS for 3 h. GAPDH and 18 S ribosomal RNA were used as internal controls. ( d–g ) Effects of knockdown ( d ) or pharmacological inhibition ( e ) of TRPC3 on CTGF mRNA expression ( d,e ) and α-SMA protein expression ( f,g ) induced by TGF-β2 stimulation in human cardiac fibroblasts (n = 3). Error bars, s.e.m. *P < 0.05, **P < 0.01. ( h ) Schema for the mechanism of TRPC3-mediated cardiac fibrosis induced by pressure overload. TRPC3 mediates MS-induced expression of profibrotic factors (CTGF and TGF-β) in cardiomyocytes and TGF-β-induced fibrotic responses of cardiac fibroblasts, partially through Nox2-dependent GEF-H1 activation.

Article Snippet: For protein knockdown, cells were transfected with siRNAs (100 nM) using Lipofectamine 2000 for 72 h. Primary human cardiac fibroblasts were purchased from Lonza and cultured according to manufacturer’s instruction.

Techniques: Derivative Assay, Inhibition, Expressing, Activation Assay