HDAC6 protein was immunoprecipitated from HCE cells using specific antibody against HDAC6 (Santa Cruz Biotechnology) and was incubated with HDAC colorimetric substrate that comprises an acetylated lysine side chain to determine HDAC6 activity. For immunoprecipitation, cells (5 × 107) were incubated in 1 mL lysis buffer (20 mM Tris pH 7.5, 137 mM NaCl, 1.5 mM, 2 mM EDTA, 10% glycerol, 1% Triton X-100, 1 mM phenylmethylsulfonyl fluoride, 10 μg/mL leupeptin, and 10 μg/mL aprotinin) on ice for 10 minutes. Cell lysates were sonicated and centrifuged at 13,000g for 10 minutes at 4°C. Supernatants were incubated with antibodies against HDAC6 at 4°C overnight. Immunocomplexes were recovered by incubation with 40 μL 25% protein A/G-Sepharose (Santa Cruz Biotechnology). Immunocomplex beads were washed twice with lysis buffer, dissolved in 1% SDS lysis buffer, and subjected to HDAC6 enzymatic activity assay using a kit purchased from (BioVision, Mountain View, CA). According to the manual, HDAC6 protein precipitated from HCE cells was incubated with HDAC colorimetric substrate at 37°C for 3 hours to deacetylate and sensitize the substrates. In the second step, the assay was stopped by the addition of 10 μL lysine developer and was incubated at 37°C for 30 minutes to produces a chromophore. Samples were read using an ELISA plate reader (LabSystem Multiskan MCC/340; Fisher Scientific, Pittsburgh, PA) at a wavelength of 405 nm.