June 2023
Volume 64, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2023
HDAC1/2 inhibition induces cell-type dependent effects on viability and histone H3 and H4 acetylation
Author Affiliations & Notes
  • Kyung Sik Jung
    Surgery, East Tennessee State University James H Quillen College of Medicine, Johnson City, Tennessee, United States
  • Rashmi Dubey
    Surgery, East Tennessee State University James H Quillen College of Medicine, Johnson City, Tennessee, United States
  • Sushil Kumar Dubey
    Surgery, East Tennessee State University James H Quillen College of Medicine, Johnson City, Tennessee, United States
  • Nathaniel F Ashley
    East Tennessee State University James H Quillen College of Medicine, Johnson City, Tennessee, United States
  • Wenjian (Vincent) Tian
    East Tennessee State University James H Quillen College of Medicine, Johnson City, Tennessee, United States
  • Mark Ellsworth Kleinman
    Surgery, East Tennessee State University James H Quillen College of Medicine, Johnson City, Tennessee, United States
  • Footnotes
    Commercial Relationships   Kyung Sik Jung None; Rashmi Dubey None; Sushil Dubey None; Nathaniel Ashley None; Wenjian (Vincent) Tian None; Mark Kleinman None
  • Footnotes
    Support  NIH Grant R01EY028206
Investigative Ophthalmology & Visual Science June 2023, Vol.64, 2108. doi:
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      Kyung Sik Jung, Rashmi Dubey, Sushil Kumar Dubey, Nathaniel F Ashley, Wenjian (Vincent) Tian, Mark Ellsworth Kleinman; HDAC1/2 inhibition induces cell-type dependent effects on viability and histone H3 and H4 acetylation. Invest. Ophthalmol. Vis. Sci. 2023;64(8):2108.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Previous data from our lab revealed histone deacetylase 1 and 2 (HDAC1/2) expression was significantly reduced in human retinal pigment epithelium (RPE) with advanced dry AMD. Cellular response to HDAC inhibition is complex and the pharmacological HDAC1/2 inhibitor, romidepsin, induces widespread cellular changes including cell death, cell cycle arrest, autophagy, and altered protein acetylation. In this study, we have evaluated the effects of HDAC1/2 inhibition on cell viability and histone H3 and H4 acetylation (H3ac/H4ac) in RPE cell lines as compared to numerous other cell types.

Methods : ARPE-19, primary human RPE (hRPE), HMVEC (endothelial cells), HMC-3 (microglial cells), and THP-1 (monocyte) cell cultures (Lonza, ATCC) were treated with 0.5, 1.0, and 5.0 ng/ml romidepsin (Selleckchem, S3020, 24/48/72h). Cell viability was measured (MTS, Promega, G3580) at 0, 24, 48, and 72h, and statistical analyses was performed (Kruskal-Wallis). Histone acetylation (H3Ac/H4Ac) after romidepsin treatment was evaluated by Western blot.

Results : Romidepsin treatment significantly reduced RPE cell (ARPE19/hRPE) viability, and increased H3/H4 hyperacetylation. At 72h, romidepsin treated ARPE-19 cells (1.0 ng/ml) exhibited significantly decreased cell viability (69.04±1.97%). Similarly decreased cell viability was observed in hRPE (35.7±8.9%), HMVEC (46.6±1.09%) and HMC3(59.2±4.49%) cells. THP-1, a monocytic cell line, showed the least effect with romidepsin with enhanced cell viability at 24h post-treatment. THP-1 cells were resistant to romidepsin treatment and at 72h the loss in cell viability was only 6.56±2.22%. Densitometry analyses revealed romidepsin (5ng/mL) induced the highest levels of H3ac (22-fold) and H4ac (4-fold) in ARPE19, followed by hRPE (H3ac,14-fold; H4ac, 13.3-fold). In contrast, romidepsin treatment of THP-1 cells induced significantly less H3ac (8-fold) and H4ac (2.8-fold).

Conclusions : The cellular changes associated with HDAC1/2 inhibition are cell-type dependent. We observed variable loss of cell viability and global acetylation in numerous different cell types. The RPE is exquisitely sensitive to HDAC1/2 inhibition with robust loss of cell viability and significantly increased H3ac/H4a. Thus, loss of HDAC1/2 function could play an important role in the RPE degeneration in patient with atrophic dry AMD.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

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