Research Paper Volume 13, Issue 16 pp 20569—20584

NPAS2 ameliorates myocardial ischaemia/reperfusion injury in rats via CX3CL1 pathways and regulating autophagy

NPAS2 transcriptionally promoted CX3CL1 expression in vitro. (A, B) Flow cytometry detected the changes of apoptosis in H9c2 cells and quantified. (C) The protein level of Cleaved-Caspase-3 (17kDa) and Caspase-3 (17kDa) in H9c2 cells was determined by Western Blot. (D) The protein level of CX3CL1 (100kDa) in H9c2 cells was determined by Western Blot. (E) The protein level of LC3B (14 and 16kDa) and p62 (62kDa) in H9c2 cells was determined by Western Blot. (F) The protein level of p-AKT (60kDa), t-AKT (60kDa), p-mTOR (289kDa) and t-mTOR (289kDa) in H9c2 cells was determined by Western Blot. (G) Typical images of immunofluorescence staining of mRFP-GFP-LC3 in H9c2 cells. Typical profiles of autophagosomes (RFP+GFP+dots) and autolysosomes (RFP+GFP-dots). (H) Autophagic vacuoles (autophagosomes) determined by transmission electron microscopy (TEM). Representative TEM images are shown, and typical autophagosomes are marked with black arrows. Data are expressed as mean ± SEM (n = 3).

Figure 7. NPAS2 transcriptionally promoted CX3CL1 expression in vitro. (A, B) Flow cytometry detected the changes of apoptosis in H9c2 cells and quantified. (C) The protein level of Cleaved-Caspase-3 (17kDa) and Caspase-3 (17kDa) in H9c2 cells was determined by Western Blot. (D) The protein level of CX3CL1 (100kDa) in H9c2 cells was determined by Western Blot. (E) The protein level of LC3B (14 and 16kDa) and p62 (62kDa) in H9c2 cells was determined by Western Blot. (F) The protein level of p-AKT (60kDa), t-AKT (60kDa), p-mTOR (289kDa) and t-mTOR (289kDa) in H9c2 cells was determined by Western Blot. (G) Typical images of immunofluorescence staining of mRFP-GFP-LC3 in H9c2 cells. Typical profiles of autophagosomes (RFP+GFP+dots) and autolysosomes (RFP+GFP-dots). (H) Autophagic vacuoles (autophagosomes) determined by transmission electron microscopy (TEM). Representative TEM images are shown, and typical autophagosomes are marked with black arrows. Data are expressed as mean ± SEM (n = 3).