Journal: Journal of Cardiovascular Translational Research

Article Title: MicroRNA-122-5p Aggravates Angiotensin II-Mediated Myocardial Fibrosis and Dysfunction in Hypertensive Rats by Regulating the Elabela/Apelin-APJ and ACE2-GDF15-Porimin Signaling

doi: 10.1007/s12265-022-10214-3

Figure Lengend Snippet: MiR-122 inhibitor significantly rescued Ang II-mediated increases in oncosis, cellular proliferation, and oxidative stress in cultured CFs via activating the ELA/apelin signaling. A , B The reverse transcription RT-PCR analysis revealed that miR-122 inhibitor rescued Ang II-meditated downregulation of apelin ( A ) and ELA ( B ) levels in Ang II-treated CFs, which were blocked by cotreatment with apelin siRNA ( A ) and ELA siRNA ( B ), respectively. C – H MiR-122 inhibitor suppressed Ang II-induced ROS generation ( C , D ) in cultured rat CFs, as well as expression of porimin ( E , G ) and cellular migration ( F , H ), which were abolished by apelin siRNA and ELA siRNA transfection, respectively. I – J Western blot analysis showed that silencing of apelin ( I ) and ELA ( J ) by siRNA abrogated miR-122 inhibitor-mediated upregulation of p-YAP ( I ) and GDF15 ( J ). CF, cardiac fibroblast; Ang II, angiotensin II; NC, negative control; ELA, elabela; ROS, reactive oxygen species; GDF15, growth differentiation factor 15; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; YAP, yes-associated protein; A.U., arbitrary unit; R.E., relative expression. n = 5–6 for each group except for I – J where n = 3-4. * P < 0.05, ** P < 0.01, *** P < 0.001

Article Snippet: Each membrane was pre-incubated in tris-buffered saline with Tween20 (TBST) containing 5% nonfat milk for 1 h at room temperature and then incubated with specific primary antibodies against GDF15 (Santa Cruz, USA), porimin (Santa Cruz, USA), phosphorylated YAP (p-YAP; ABclonal, China), and YAP (Proteintech, USA) overnight at 4℃.

Techniques: Cell Culture, Reverse Transcription Polymerase Chain Reaction, Expressing, Migration, Transfection, Western Blot, Negative Control

Journal: Journal of Cardiovascular Translational Research

Article Title: MicroRNA-122-5p Aggravates Angiotensin II-Mediated Myocardial Fibrosis and Dysfunction in Hypertensive Rats by Regulating the Elabela/Apelin-APJ and ACE2-GDF15-Porimin Signaling

doi: 10.1007/s12265-022-10214-3

Figure Lengend Snippet: Treatment with apelin and ELA prevented Ang II-induced cellular oncosis and oxidative stress in CFs associated with upregulated levels of p-YAP and GDF15. A – D DHE staining and immunofluorescence revealed that pretreatemnet of apelin, ELA, and NAC prevented against Ang II-induced increases in ROS generation ( A , C ), and porimin expression ( B , D ). E The levels of LDH in each group. F Transmission electron microscopy showed that apelin and ELA attenuated Ang II-induced cellular oncosis-specific ultrastructural changes in CFs, characterized by swelling of organelles (yellow arrow) and nucleus (red arrow), and vacuolar degeneration of mitochondrial (black arrow). G – J Reverse transcription PCR ( G ) and western blot analysis ( H – J ) indicated that the ELA and apelin upregulated ACE2 mRNA expression ( G ), as well as p-YAP ( H , I ) and protein expression of GDF15 ( H , J ) in Ang II-treated CFs. CF, cardiac fibroblast; Ang II, angiotensin II; ELA, elabela; ROS, reactive oxygen species; LDH, lactate dehydrogenase; NAC, N-acetylcysteine; YAP, Yes-associated protein; GDF15, growth differentiation factor; GAPDH, glyceraldehyde-3-phosphate dehydrogenase. A.U., arbitrary unit; R.E., relative expression. n = 5–6 for each group except for H – J where n = 3. *** P < 0.001

Article Snippet: Each membrane was pre-incubated in tris-buffered saline with Tween20 (TBST) containing 5% nonfat milk for 1 h at room temperature and then incubated with specific primary antibodies against GDF15 (Santa Cruz, USA), porimin (Santa Cruz, USA), phosphorylated YAP (p-YAP; ABclonal, China), and YAP (Proteintech, USA) overnight at 4℃.

Techniques: Staining, Immunofluorescence, Expressing, Transmission Assay, Electron Microscopy, Western Blot

Journal: Journal of Cardiovascular Translational Research

Article Title: MicroRNA-122-5p Aggravates Angiotensin II-Mediated Myocardial Fibrosis and Dysfunction in Hypertensive Rats by Regulating the Elabela/Apelin-APJ and ACE2-GDF15-Porimin Signaling

doi: 10.1007/s12265-022-10214-3

Figure Lengend Snippet: ACE2 antagonized Ang II-induced augmentation of oxidative stress, migration, and proliferation in cultured CFs via activating the YAP-GDF15 signaling. A – E Flow cytometry assay ( A , B ), wound healing assay ( C , D ), and CCK8 ( E ) assay demonstrated that treatment with ACE2 alleviated Ang II-induced promotion of ROS production ( A , B ), cellular migration ( C , D ), and proliferation ( E ) in cultured rat CFs. F Treatment with recombinant ACE2 rescued Ang II-mediated reduction in NO production in rat CFs. G – I The levels of p-YAP ( G , H ), YAP ( G , H ), GDF15 ( H , I ) were detected by Western blotting, and the results showed that expression of GDF15 ( H , I ) and the ratio of p-YAP/YAP ( G , H ) were upregulated by ACE2 in the Ang II-treated CFs. CF, cardiac fibroblast; ROS, reactive oxygen species; ACE2, angiotensin-converting enzyme; GDF15, growth differentiation factor; YAP, Yes-associated protein; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; NO, nitric oxide; Ang II, angiotensin II; ELA, elabela; A.U., arbitrary unit; n = 5–6 for each group except for G – I where n = 4. * P < 0.05, ** P < 0.01, *** P < 0.001

Article Snippet: Each membrane was pre-incubated in tris-buffered saline with Tween20 (TBST) containing 5% nonfat milk for 1 h at room temperature and then incubated with specific primary antibodies against GDF15 (Santa Cruz, USA), porimin (Santa Cruz, USA), phosphorylated YAP (p-YAP; ABclonal, China), and YAP (Proteintech, USA) overnight at 4℃.

Techniques: Migration, Cell Culture, Flow Cytometry, Wound Healing Assay, Recombinant, Western Blot, Expressing