Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: S107 is the most effective drug to prevent the aberrant Ca 2+ release in RyR2-H29D hiPSC-CMs. A Display of original line-scan images of Ca 2+ transients and corresponding tracings and pacing trail in RyR2-H29D hiPSC-CMs, isogenic control, RyR2-H29D hiPSC-CMs treated with propranolol (3 µM for 10 min), verapamil (10 nM for 10 min), flecainide (5 µM for 10 min) and S107 (5 µM for 10 min) under 1 Hz pacing (20 V and 5 ms duration). Additional and aberrant Ca 2+ release events are shown with the arrows. B Normalized Ca 2+ -transient amplitude in RyR2-H29D hiPSC-CMs (red bars and red dots plot), isogenic control (black bars and black dots plot), RyR2-H29D treated with propranolol (blue bars and blue dots plot), verapamil (green bars and green dots plot), flecainide (purple bars and purple dots plot) and S107 (gray bars and gray dots plot) under 1 Hz pacing. C Frequency of occurrence of aberrant Ca 2+ -transients in RyR2-H29D, isogenic control, RyR2-H29D treated with propranolol, verapamil, flecainide and S107. D Frequency of occurrence of diastolic leaky events in RyR2-H29D, isogenic control, RyR2-H29D treated with propranolol, verapamil, flecainide and S107. E Rate of RyR2 Ca 2+ release (dF/dt max in ΔF/s) in RyR2-H29D, isogenic control, RyR2-H29D treated with propranolol, verapamil, flecainide and S107. (F) Decay time in RyR2-H29D, isogenic control, RyR2-H29D treated with propranolol, verapamil, flecainide and S107. The number of experiments is based on 3 independent biological replicates. The number of experiments varies from 20 to 152 cells for each scatter plot. Significance was calculated by Kruskal–Wallis test. Data are presented as mean ± SEM. *, p < 0.05, **, p < 0.01, ***, p < 0.001

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: Summary and grading of the effects of propranolol, verapamil, flecainide and S107 on the SR Ca 2+ handling, contractile properties and RyR2 remodeling in RyR2-H29D (PMVT) hiPSC-CMs

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: Propranolol improves the contractile properties in RyR2-H29D hiPSC-CMs. A Representative traces of contractile parameters in RyR2-H29D and RyR2-H29D hiPSC-CMs treated with 3 µM propranolol for 20 min. Aberrant contraction oscillations are marked with the arrows. B Normalized beat rate in RyR2-H29D hiPSC-CMs (red bars and red dots plot) and RyR2-H29D hiPSC-CMs treated with propranolol (blue bars and blue dots plot). C Normalized contraction time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with propranolol. D Normalized relaxation time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with propranolol. E Normalized resting time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with propranolol. F Normalized homogeneity in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with propranolol. All parameters were normalized to the isogenic control (dotted line). The number of experiments is based on 3 independent biological replicates. The number of experiments varies from 39 to 40 videos for each scatter plot. Significance was calculated by Wilcoxon test. Data are presented as mean ± SEM. *, p < 0.05, **, p < 0.01

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: Verapamil does not prevent the aberrant contractile properties in RyR2-H29D hiPSC-CMs. A Representative traces of contractile parameters in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with 100 nM verapamil for 20 min. Aberrant contraction oscillations are marked with the arrows. B Normalized beat rate in RyR2-H29D hiPSC-CMs (red bars and red dots plot) and RyR2-H29D hiPSC-CMs treated with verapamil (green bars and green dots plot). C Normalized contraction time in RyR2-H29D and RyR2-H29D treated with verapamil. D Normalized relaxation time in RyR2-H29D and RyR2-H29D treated with verapamil. E Normalized resting time in RyR2-H29D and RyR2-H29D treated with verapamil. F Normalized homogeneity in RyR2-H29D and RyR2-H29D treated with verapamil. All parameters were normalized to the isogenic control (dotted line). The number of experiments is based on 3 independent biological replicates. The number of experiments varies from 28 to 30 videos for each scatter plot. Significance was calculated by Wilcoxon and paired t tests. Data are presented as mean ± SEM. **, p < 0.01

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: Flecainide is effective to prevent the aberrant contractile properties in RyR2-H29D hiPSC-CMs. A Representative traces of contractile parameters in RyR2-H29D and RyR2-H29D hiPSC-CMs treated with 5 µM flecainide for 20 min. Aberrant contraction oscillations are marked with the arrows. B Normalized beat rate in RyR2-H29D hiPSC-CMs (red bars and red dots plot) and RyR2-H29D hiPSC-CMs treated with flecainide (purple bars and purple dots plot). C Normalized contraction time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with flecainide. D Normalized relaxation time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with flecainide. E Normalized resting time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with flecainide. F Normalized homogeneity in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with flecainide. All parameters were normalized to the isogenic control (dotted line). The number of experiments is based on 3 independent biological replicates. The number of experiments varies from 39 to 40 videos for each scatter plot. Significance was calculated by Wilcoxon and paired t tests. Data are presented as mean ± SEM. *, p < 0.05, **, p < 0.01

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: S107 is effective to prevent the aberrant contractile properties in RyR2-H29D hiPSC-CMs. A Representative traces of contractile parameters in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with 5 µM S107 for 20 min. Aberrant contraction oscillations are marked with the arrows. B Normalized beat rate in RyR2-H29D hiPSC-CMs (red bars and red dots plot) and RyR2-H29D hiPSC-CMs treated with S107 (gray bars and gray dots plot). C Normalized contraction time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with S107. D Normalized relaxation time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with S107. E Normalized resting time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with S107. F Normalized homogeneity in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with S107. All parameters were normalized to the isogenic control (dotted line). The number of experiments is based on 3 independent biological replicates. The number of experiments varies from 26 to 30 videos for each scatter plot. Significance was calculated by Wilcoxon and paired t tests. Data are presented as mean ± SEM. **, p < 0.01

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: S107 is the most effective drug to prevent the aberrant Ca 2+ release in RyR2-H29D hiPSC-CMs. A Display of original line-scan images of Ca 2+ transients and corresponding tracings and pacing trail in RyR2-H29D hiPSC-CMs, isogenic control, RyR2-H29D hiPSC-CMs treated with propranolol (3 µM for 10 min), verapamil (10 nM for 10 min), flecainide (5 µM for 10 min) and S107 (5 µM for 10 min) under 1 Hz pacing (20 V and 5 ms duration). Additional and aberrant Ca 2+ release events are shown with the arrows. B Normalized Ca 2+ -transient amplitude in RyR2-H29D hiPSC-CMs (red bars and red dots plot), isogenic control (black bars and black dots plot), RyR2-H29D treated with propranolol (blue bars and blue dots plot), verapamil (green bars and green dots plot), flecainide (purple bars and purple dots plot) and S107 (gray bars and gray dots plot) under 1 Hz pacing. C Frequency of occurrence of aberrant Ca 2+ -transients in RyR2-H29D, isogenic control, RyR2-H29D treated with propranolol, verapamil, flecainide and S107. D Frequency of occurrence of diastolic leaky events in RyR2-H29D, isogenic control, RyR2-H29D treated with propranolol, verapamil, flecainide and S107. E Rate of RyR2 Ca 2+ release (dF/dt max in ΔF/s) in RyR2-H29D, isogenic control, RyR2-H29D treated with propranolol, verapamil, flecainide and S107. (F) Decay time in RyR2-H29D, isogenic control, RyR2-H29D treated with propranolol, verapamil, flecainide and S107. The number of experiments is based on 3 independent biological replicates. The number of experiments varies from 20 to 152 cells for each scatter plot. Significance was calculated by Kruskal–Wallis test. Data are presented as mean ± SEM. *, p < 0.05, **, p < 0.01, ***, p < 0.001

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: Summary and grading of the effects of propranolol, verapamil, flecainide and S107 on the SR Ca 2+ handling, contractile properties and RyR2 remodeling in RyR2-H29D (PMVT) hiPSC-CMs

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: Propranolol improves the contractile properties in RyR2-H29D hiPSC-CMs. A Representative traces of contractile parameters in RyR2-H29D and RyR2-H29D hiPSC-CMs treated with 3 µM propranolol for 20 min. Aberrant contraction oscillations are marked with the arrows. B Normalized beat rate in RyR2-H29D hiPSC-CMs (red bars and red dots plot) and RyR2-H29D hiPSC-CMs treated with propranolol (blue bars and blue dots plot). C Normalized contraction time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with propranolol. D Normalized relaxation time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with propranolol. E Normalized resting time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with propranolol. F Normalized homogeneity in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with propranolol. All parameters were normalized to the isogenic control (dotted line). The number of experiments is based on 3 independent biological replicates. The number of experiments varies from 39 to 40 videos for each scatter plot. Significance was calculated by Wilcoxon test. Data are presented as mean ± SEM. *, p < 0.05, **, p < 0.01

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: Verapamil does not prevent the aberrant contractile properties in RyR2-H29D hiPSC-CMs. A Representative traces of contractile parameters in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with 100 nM verapamil for 20 min. Aberrant contraction oscillations are marked with the arrows. B Normalized beat rate in RyR2-H29D hiPSC-CMs (red bars and red dots plot) and RyR2-H29D hiPSC-CMs treated with verapamil (green bars and green dots plot). C Normalized contraction time in RyR2-H29D and RyR2-H29D treated with verapamil. D Normalized relaxation time in RyR2-H29D and RyR2-H29D treated with verapamil. E Normalized resting time in RyR2-H29D and RyR2-H29D treated with verapamil. F Normalized homogeneity in RyR2-H29D and RyR2-H29D treated with verapamil. All parameters were normalized to the isogenic control (dotted line). The number of experiments is based on 3 independent biological replicates. The number of experiments varies from 28 to 30 videos for each scatter plot. Significance was calculated by Wilcoxon and paired t tests. Data are presented as mean ± SEM. **, p < 0.01

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: Flecainide is effective to prevent the aberrant contractile properties in RyR2-H29D hiPSC-CMs. A Representative traces of contractile parameters in RyR2-H29D and RyR2-H29D hiPSC-CMs treated with 5 µM flecainide for 20 min. Aberrant contraction oscillations are marked with the arrows. B Normalized beat rate in RyR2-H29D hiPSC-CMs (red bars and red dots plot) and RyR2-H29D hiPSC-CMs treated with flecainide (purple bars and purple dots plot). C Normalized contraction time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with flecainide. D Normalized relaxation time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with flecainide. E Normalized resting time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with flecainide. F Normalized homogeneity in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with flecainide. All parameters were normalized to the isogenic control (dotted line). The number of experiments is based on 3 independent biological replicates. The number of experiments varies from 39 to 40 videos for each scatter plot. Significance was calculated by Wilcoxon and paired t tests. Data are presented as mean ± SEM. *, p < 0.05, **, p < 0.01

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: Stem Cell Research & Therapy

Article Title: Personalized medicine in the dish to prevent calcium leak associated with short-coupled polymorphic ventricular tachycardia in patient-derived cardiomyocytes

doi: 10.1186/s13287-023-03502-5

Figure Lengend Snippet: S107 is effective to prevent the aberrant contractile properties in RyR2-H29D hiPSC-CMs. A Representative traces of contractile parameters in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with 5 µM S107 for 20 min. Aberrant contraction oscillations are marked with the arrows. B Normalized beat rate in RyR2-H29D hiPSC-CMs (red bars and red dots plot) and RyR2-H29D hiPSC-CMs treated with S107 (gray bars and gray dots plot). C Normalized contraction time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with S107. D Normalized relaxation time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with S107. E Normalized resting time in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with S107. F Normalized homogeneity in RyR2-H29D hiPSC-CMs and RyR2-H29D hiPSC-CMs treated with S107. All parameters were normalized to the isogenic control (dotted line). The number of experiments is based on 3 independent biological replicates. The number of experiments varies from 26 to 30 videos for each scatter plot. Significance was calculated by Wilcoxon and paired t tests. Data are presented as mean ± SEM. **, p < 0.01

Article Snippet: 4–20% SDS-PAGE gradient gel was used to separate the proteins which were incubated with the following primary antibodies: rabbit 5029 Y2 anti-RyR2 (1:5000), anti-phospho-RyR2-pSer2809 (homemade antibody: polyclonal rabbit CRTRRI- (pS)-QTSQ, 1:1000), anti-RyR2-pSer2815 (homemade antibody: polyclonal rabbit CSQTSQV-(pS)-VD), anti-DNP antibody (Millipore,1:2000), anti-Cys-NO (Sigma-Aldrich, 1:1000) and mouse anti-FKBP12.6 (Santa Cruz, ref: 376,135, 1:1000).

Techniques:

Journal: PLoS ONE

Article Title: 2-Deoxy-D-Glucose Treatment Induces Ketogenesis, Sustains Mitochondrial Function, and Reduces Pathology in Female Mouse Model of Alzheimer's Disease

doi: 10.1371/journal.pone.0021788

Figure Lengend Snippet: Hippocampal homogenate samples from both the Ctrl and 2-DG groups were analyzed for protein levels of APP, multiple forms of Aβ oligomer (56 kD, 27 kD, 16 kD), sAPPα fragment, ADAM10, BACE1, and PS-1 by western blot. Brain sections were stained for amyloid species (6E10) and microglia marker (Iba I). A, 2-DG induced significant decrease in APP, 56 kD, 27 kD and 16 kD Aβ oligomer protein level. In contrast, sAPPαlevel was significantly increased by 2-DG (*, P<0.05 compared to Ctrl, bars represent mean values ± SEM); B, immunofluorescent labeling of amyloid with 6E10 antibody in the hippocampal CA1 region. Image i–iv: lower magnification, scale bar: 100 µm; images v–viii, higher magnification, scale bar: 20 µm. Images i, ii, v, and vi: 6E10 only, Images iii, iv, vii, and viii: 6E10 with microglia marker IbaI labeling and DAPI; C, 2-DG treatment significantly increased expression of α secretase ADAM10 protein levels. In contrast, γ secretase PS-1 protein level was significantly decreased by 2-DG.β secretase was not changed by 2-DG (*, P<0.05 compared to Ctrl, bars represent mean value ± SEM).

Article Snippet: Equal amounts of proteins (20 µg/well) were loaded in each well of a 12% SDS-PAGE gel, electrophoresed with a Tris/glycine running buffer, and transferred to a 0.45 µm pore size polyvinylidene difluoride (PVDF) membrane and immunobloted with 6E10 antibody (1∶1000, Covance, Princeton, NJ), sAPPα antibody (1∶500, Covance, Princeton, NJ), PHF13/pTau antibody (1∶500, Covance, Princeton, NJ), PDH E1 alpha antibody (1∶1000, Mitosciences, Eugene, OR), ADAM10 antibody (1∶500, Millipore, Temecula, CA), PS-1 antibody (1∶1000, Millipore, Temecula, CA), BACE1 antibody (1∶500, Covance, Princeton, NJ), FGF-2 antibody (1∶200, Millipore, Temecula, CA), NGFβ antibody (1∶500, Millipore, Temecula, CA) and BDNF antibody (1∶200, Chemicon, Ramona, CA), Arc antibody (1∶500, Proteintech, Chicago, IL), OGDH antibody (1∶500, Proteintech, Chicago, IL), SCOT Antibody (1∶100, Santa Cruz, Santa Cruz, CA), ACAT1 antibody (1∶500, Proteintech, Chicago, IL), MnSOD antibody (1∶1000, BD Biosciences, San Diego, CA), PrdxV antibody (1∶200, BD Biosciences, San Diego, CA), Hsp60 antibody (1∶500, Millipore, Temecula, CA), β-actin antibody (1∶5000, Chemicon, Ramona, CA), and porin/VDAC antibody (1∶500, Cell Signaling, Danvers, MA).

Techniques: Western Blot, Staining, Marker, Labeling, Expressing

Journal: Neurobiology of Disease

Article Title: Analysis of homozygous and heterozygous Csf1r knockout in the rat as a model for understanding microglial function in brain development and the impacts of human CSF1R mutations

doi: 10.1016/j.nbd.2021.105268

Figure Lengend Snippet:

Article Snippet: Anti-PS-NCAM , Millipore , Cat# MAB5324, RRID: AB_95211.

Techniques: Software