Ab11805

Antibodies used in this study are as follows. Goat anti-FSTL1 (ab11805; Abcam), rabbit anti-FSTL1 (Proteintech), rabbit anti-DIAPH1 (5486; Cell Signaling Technology), rabbit anti-KSRP (A302-22A; Bethyl Laboratories), mouse anti-KRT14 (clone LLO01), mouse anti-EGF (clone 10825; R&D Systems), mouse anti-TGFβ1 (Novacastra), rabbit anti-GFP (Covance), rabbit anti-cMyc (Sigma-Aldrich), and rabbit anti-Wnt7a (ab100792; Abcam). Streptavidin Alexa Fluor 555, chicken anti-goat Alexa Fluor 488, and donkey anti-rabbit/mouse Alexa Fluor 488/555 were from Molecular Probes. Rabbit anti-Arp3 (4738), phospho (9101), and total ERK antibodies (9102) were from Cell Signaling Technology. Rabbit anti-Arpin (ABT251) was purchased from Merck Millipore. Recombinant human FSTL1 (His-tagged) expressed and purified from mammalian cells was purchased from Thermo Fisher Scientific.

Recombinant human FSTL1 (≥97% purity, as determined by HPLC) was obtained from PeproTech Co. (New Jersey, USA). Primary antibodies against FSTL1 (#ab11805, 1 : 1000), GAPDH (#ab181602, 1 : 1000), Nrf2 (#ab62352, 1 : 1000), and cleaved caspase-3 (#ab2302, 1 : 1000) were obtained from Abcam (Cambridge, UK). A mouse N-terminal brain natriuretic peptide precursor (NT-proBNP) assay kit was obtained from CUSABIO (#CSB-E05153m, Wuhan, China). DOX (#D1515) was provided by Sigma-Aldrich (MO, USA). A mouse cardiac troponin I (cTnI) ELISA kit was provided by CUSABIO (#CSB-E08421m). A mouse creatine kinase (CK) ELISA kit (#CSB-E14404m) and lactate dehydrogenase (LDH) ELISA kit (#CSB-E17733m) were also provided by CUSABIO.

Cells were harvested by scraper and lysed with RIPA buffer supplemented with 1× complete protease inhibitor cocktail (Roche) following the manufacturer’s protocol. Protein concentrations were determined using a Pierce BCA protein assay kit (Thermo Scientific). Lysates were separated on SDS-PAGE gels and transferred. Membranes were immunoblotted with the following antibodies: YAP (ab52771, EP1674Y, abcam; 1:1000), TRAM2 (ab109176, EPR2658, abcam; 1:5000), FSTL-1 (ab11805, abcam, 1:1000), Vimentin (550,513, BD Biosciences, 1:2000), HSP90 (610,418, BD Biosciences, 1:5000), and GAPDH (sc-32,233, Santa Cruz, 1:2000). Protein bands were visualized using corresponding secondary antibodies (Dako) and ECL reagent (GE Healthcare).