ABSTRACT
Bromodomain-containing proteins are vital for controlling the expression of many pro-inflammatory genes. Consequently, compounds capable of inhibiting specific bromodomain-facilitated protein-protein interactions would be predicted to alleviate inflammation, making them valuable agents in the treatment of diseases caused by dysregulated inflammation, such as age-related macular degeneration. Here, we assessed the ability of known inhibitors JQ-1, PFI-1, and IBET-151 to protect from the inflammation and cell death caused by etoposide exposure in the human retinal pigment epithelial cell line, ARPE-19. The potential anti-inflammatory effects of the bromodomain inhibitors were assessed by ELISA (enzyme-linked immunosorbent assay) profiling. The involvement of NF-κB and SIRT1 in inflammatory signaling was monitored by ELISA and western blotting. Furthermore, SIRT1 was knocked down using a specific siRNA or inhibited by EX-527 to elucidate its role in the inflammatory reaction. The bromodomain inhibitors effectively decreased etoposide-induced release of IL-6 and IL-8. This anti-inflammatory effect was not related to SIRT1 activity, although all bromodomain inhibitors decreased the extent of acetylation of p53 at the SIRT1 deacetylation site. Overall, since bromodomain inhibitors display anti-inflammatory properties in human retinal pigment epithelial cells, these compounds may represent a new way of alleviating the inflammation underlying the onset of age-related macular degeneration.
Subject(s)
Anti-Inflammatory Agents/pharmacology , Azabicyclo Compounds/pharmacology , Azepines/pharmacology , Benzodiazepines/pharmacology , Epithelial Cells/metabolism , Etoposide/antagonists & inhibitors , Pyridines/pharmacology , Transcription Factors/antagonists & inhibitors , Triazoles/pharmacology , Carbazoles/pharmacology , Cell Line , Cell Survival/drug effects , Epithelial Cells/cytology , Epithelial Cells/drug effects , Etoposide/pharmacology , Gene Expression Regulation , Humans , Inflammation , Interleukin-6/antagonists & inhibitors , Interleukin-6/genetics , Interleukin-6/metabolism , Interleukin-8/antagonists & inhibitors , Interleukin-8/genetics , Interleukin-8/metabolism , Models, Biological , NF-kappa B/genetics , NF-kappa B/metabolism , Protein Domains , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Retinal Pigment Epithelium/cytology , Retinal Pigment Epithelium/drug effects , Retinal Pigment Epithelium/metabolism , Signal Transduction , Sirtuin 1/antagonists & inhibitors , Sirtuin 1/genetics , Sirtuin 1/metabolism , Topoisomerase II Inhibitors/pharmacology , Transcription Factors/genetics , Transcription Factors/metabolism , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/metabolismABSTRACT
Acetylation of starch considerably decreases its swelling and enzymatic degradation. Thus, starch-acetate (SA) based delivery systems may be suitable for controlled drug delivery. The aim of the present study was to evaluate drug release from the SA microparticles (SA mps) and SA films. The average degree of acetyl substitution (DS) per glucose residue in the starch was either 1.9 (SA DS 1.9) or 2.6 (SA DS 2.6). Timolol (mw 332), calcein (mw 623) and bovine serum albumin (BSA, mw 68,000) were used as model drugs. A continuous timolol release from the both SA mps was observed in phosphate buffer solution (PBS) pH 7.4 (50-days incubation). The release of timolol was faster from the SA DS 1.9 mps than from the SA DS 2.6 mps. Calcein release from both SA mps was continuous in PBS pH 7.4 (5-days incubation). But, calcein release profile from the SA DS 2.6 film in PBS pH 7.4 showed discontinuities. However, the release of calcein from both SA films was continuous in human serum in vitro during the 7-day incubation, i.e. enzymes enhanced calcein release. Thus, alpha-amylase was incorporated into the SA films in order to enhance drug release from the films. However, the effects of incorporation of alpha-amylase on the model macromolecule (BSA) release from the SA films were modest. In conclusion, this study demonstrates the achievement of slow release of different molecular weight model drugs from the SA mps and films as compared to fast release from the native starch preparations. DS of SA, physicochemical properties of a drug and the presence of enzymes can all affect drug release profiles from SA based preparations.
