Histone 4

SNOH-3, with purity >98%, was synthesized in Medicine Chemistry Laboratory at Shenyang Pharmaceutical University (see Figure 3a and Supplementary data). SAHA were obtained from Sigma (St. Louis, MO, USA). These agents were dissolved in DMSO to 100 mM and stored at −20 °C. Before treatment, the stock solution is diluted to different concentrations. The final concentration of DMSO in cultures is 0.1% (v/v) or less. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) and propidium idodide (PI) were purchased from Sigma. The primary antibodies against HDAC1, HDAC3, HDAC6, HDAC8, Histone 3, Histone 4, p21^WAF1/CIP1, PARP, Caspase-3, Survivin, XIAP, KLF4, VEGF, and β-actin were got from Cell Signaling Technology. The primary antibodies against CD31 were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA). The primary antibodies against Ac-H3 and Ac-H4 were purchased from Millipore (Boston, MA, USA). The HDAC1 Silencer Select Validated siRNA was got from Life Technologies (Waltham, MA, USA).

We employed a Hitachi 7650 analytical transmission electron microscope to visualize the isolated exosomes and nanoparticle tracking analysis (NanoSight NS300) to analyze particle size and concentration, as described earlier [14 (link),69 (link)].
We also used immunoblotting to identify exosomal markers from the isolated fraction to characterize the BALF- and lung-tissue-derived exosomes. In brief, 20 µg of exosomal lysate was resolved on a 10% sodium dodecyl sulfate (SDS)-polyacrylamide gel and electroblotted onto nitrocellulose membranes. Membranes were blocked using a 5% blocking buffer for 1 h and thereafter probed overnight with antibodies for exosomal surface markers. The antibodies included Histone 4(Cat# 2592) (Cell Signaling Technologies, Danvers, MA, USA), CD9 (Cat# ab92726), CD63 (Cat# ab134045) (Abcam, Waltham, MA, USA), and CD81 (Cat# EXOAB-CD81A-1) (SBI Biosciences, Palo Alto, CA, USA). In the following days, the blots were washed and probed with appropriate secondary antibodies. Chemiluminescence was detected using the Bio-Rad ChemiDoc MP Imaging System using the SuperSignal West Femto Maximum Sensitivity Substrate (Cat# 34096, Thermo Scientific, Waltham, MA, USA).

Total protein was isolated from the bone marrow samples of the AML patients or AML cells using RIPA lysis solution, supplemented with protease and phosphatase inhibitors. The protein concentration was established using a BCA kit. Equivalent quantities of protein samples were then subjected to 10–12% SDS-polyacrylamide gel electrophoresis and transferred to nitrocellulose membranes (Millipore). The membranes were then stained with primary antibodies against HDAC1–11 (Abcam), acetyle-histone 3, histone 3, acetyle-histone 4, histone 4, Apaf-1 and MRP1 (Cell Signaling Technology), followed by immune-probed using a horseradish peroxidase-conjugated secondary antibody. β-actin (Abcam) was used as an internal reference gene. The target protein was visualized using an ECL Western blotting substrate kit (Biovision).