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Attenuation of choroidal neovascularization by histone deacetylase inhibitor.

Chan N, He S, Spee CK, Ishikawa K, Hinton DR - PLoS ONE (2015)

Bottom Line: Most strikingly, TSA markedly down-regulates the expression of VEGF receptor-2 in human vascular endothelial cells and, thus, can knock down pro-angiogenic cell signaling.Additionally, TSA suppresses CNV-associated wound healing response and RPE epithelial-mesenchymal transdifferentiation.This report suggests that TSA, and possibly HDACi's in general, should be further evaluated for their therapeutic potential for the treatment of CNV.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Keck School of Medicine of the University of Southern California, Los Angeles, CA, United States of America; Doheny Eye Institute, Los Angeles, CA, United States of America.

ABSTRACT
Choroidal neovascularization (CNV) is a blinding complication of age-related macular degeneration that manifests as the growth of immature choroidal blood vessels through Bruch's membrane, where they can leak fluid or hemorrhage under the retina. Here, we demonstrate that the histone deacetylase inhibitor (HDACi) trichostatin A (TSA) can down-regulate the pro-angiogenic hypoxia-inducible factor-1α and vascular endothelial growth factor (VEGF), and up-regulate the anti-angiogenic and neuro-protective pigment epithelium derived factor in human retinal pigment epithelial (RPE) cells. Most strikingly, TSA markedly down-regulates the expression of VEGF receptor-2 in human vascular endothelial cells and, thus, can knock down pro-angiogenic cell signaling. Additionally, TSA suppresses CNV-associated wound healing response and RPE epithelial-mesenchymal transdifferentiation. In the laser-induced model of CNV using C57Bl/6 mice, systemic administration of TSA significantly reduces fluorescein leakage and the size of CNV lesions at post-laser days 7 and 14 as well as the immunohistochemical expression of VEGF, VEGFR2, and smooth muscle actin in CNV lesions at post-laser day 7. This report suggests that TSA, and possibly HDACi's in general, should be further evaluated for their therapeutic potential for the treatment of CNV.

No MeSH data available.


Related in: MedlinePlus

CNV volume measurements after TSA treatment.Mouse eyes were fixed with 10% formalin and eyecups were obtained by removing the anterior poles and neurosensory retina. Eyecups containing the RPE-choroid-sclera complex were blocked with PBS containing 1% BSA and 0.5% triton X-100 and then incubated with 10 μg/mL of FITC-isolectin B4 overnight at 4°C. Fluorescent images captured using the 20× objective of a scanning confocal microscope were analyzed. On both days 7 and 14, the sizes of CNV volumes were much smaller in TSA-treated mice than in mice that received PBS only. (*: t test p<0.05, **: t test p<0.005; n = 10/group) (bar = 50 μm).
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pone.0120587.g013: CNV volume measurements after TSA treatment.Mouse eyes were fixed with 10% formalin and eyecups were obtained by removing the anterior poles and neurosensory retina. Eyecups containing the RPE-choroid-sclera complex were blocked with PBS containing 1% BSA and 0.5% triton X-100 and then incubated with 10 μg/mL of FITC-isolectin B4 overnight at 4°C. Fluorescent images captured using the 20× objective of a scanning confocal microscope were analyzed. On both days 7 and 14, the sizes of CNV volumes were much smaller in TSA-treated mice than in mice that received PBS only. (*: t test p<0.05, **: t test p<0.005; n = 10/group) (bar = 50 μm).

Mentions: To study the effect of TSA on the development of laser-induced CNV, the formation of CNV was evaluated by three methods: fluorescein angiography (FA), histology and measurement of CNV volume. 5 drug-naïve mice per group were administered intraperitoneal injection of 20 mg/kg of TSA or the same volume of sterile PBS immediately after laser photocoagulation and every 48 h thereafter for 7 or 14 days. Late-phase FAs of both eyes in each mouse from the control and TSA-treated groups were evaluated according to the grading system described in the Materials and Methods section [53]. Ten animals, 20 eyes and 34 lesions were examined in the control group, and 10 animals, 20 eyes and 35 lesions were examined in the TSA group. Representative angiographic images in control and TSA-treated mice on days 7 and 14 are shown in Fig. 11A. TSA-treated mice exhibited attenuated CNV formation and leakage compared with PBS-treated mice. Mice treated with TSA demonstrated a significant reduction of FA score compared to the control animals on days 7 and 14 after laser photocoagulation (Fig. 11B; p<0.05). For histologic analysis, 8 animals, 8 eyes and 8 lesions were examined in the control group, and 9 animals, 9 eyes and 9 lesions were examined in the TSA group. TSA treatment resulted in statistically significant reductions in lesion area on days 7 and 14 after laser photocoagulation (Fig. 12A-B; p<0.05). Choroidal flatmount with fluorescein-conjugated isolectin B4 staining was used to assess the volume of the CNV lesions on days 7 and 14 after laser photocoagulation. Ten animals, 10 eyes and 20 lesions were examined in the control group, and 10 animals, 10 eyes and 28 lesions were examined in the TSA group. Mice treated with TSA demonstrated a significantly smaller lesion size compared to those treated with PBS (Fig. 13A). Quantitative measurement of the volumes of the CNV lesions showed that TSA treatment resulted in an approximately 65.5% and 65.9% reduction, respectively, in choroidal vascular volume, compared with that of control mice, 1 week and 2 weeks after laser photocoagulation (Fig. 13B; p<0.05).


Attenuation of choroidal neovascularization by histone deacetylase inhibitor.

Chan N, He S, Spee CK, Ishikawa K, Hinton DR - PLoS ONE (2015)

CNV volume measurements after TSA treatment.Mouse eyes were fixed with 10% formalin and eyecups were obtained by removing the anterior poles and neurosensory retina. Eyecups containing the RPE-choroid-sclera complex were blocked with PBS containing 1% BSA and 0.5% triton X-100 and then incubated with 10 μg/mL of FITC-isolectin B4 overnight at 4°C. Fluorescent images captured using the 20× objective of a scanning confocal microscope were analyzed. On both days 7 and 14, the sizes of CNV volumes were much smaller in TSA-treated mice than in mice that received PBS only. (*: t test p<0.05, **: t test p<0.005; n = 10/group) (bar = 50 μm).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4373846&req=5

pone.0120587.g013: CNV volume measurements after TSA treatment.Mouse eyes were fixed with 10% formalin and eyecups were obtained by removing the anterior poles and neurosensory retina. Eyecups containing the RPE-choroid-sclera complex were blocked with PBS containing 1% BSA and 0.5% triton X-100 and then incubated with 10 μg/mL of FITC-isolectin B4 overnight at 4°C. Fluorescent images captured using the 20× objective of a scanning confocal microscope were analyzed. On both days 7 and 14, the sizes of CNV volumes were much smaller in TSA-treated mice than in mice that received PBS only. (*: t test p<0.05, **: t test p<0.005; n = 10/group) (bar = 50 μm).
Mentions: To study the effect of TSA on the development of laser-induced CNV, the formation of CNV was evaluated by three methods: fluorescein angiography (FA), histology and measurement of CNV volume. 5 drug-naïve mice per group were administered intraperitoneal injection of 20 mg/kg of TSA or the same volume of sterile PBS immediately after laser photocoagulation and every 48 h thereafter for 7 or 14 days. Late-phase FAs of both eyes in each mouse from the control and TSA-treated groups were evaluated according to the grading system described in the Materials and Methods section [53]. Ten animals, 20 eyes and 34 lesions were examined in the control group, and 10 animals, 20 eyes and 35 lesions were examined in the TSA group. Representative angiographic images in control and TSA-treated mice on days 7 and 14 are shown in Fig. 11A. TSA-treated mice exhibited attenuated CNV formation and leakage compared with PBS-treated mice. Mice treated with TSA demonstrated a significant reduction of FA score compared to the control animals on days 7 and 14 after laser photocoagulation (Fig. 11B; p<0.05). For histologic analysis, 8 animals, 8 eyes and 8 lesions were examined in the control group, and 9 animals, 9 eyes and 9 lesions were examined in the TSA group. TSA treatment resulted in statistically significant reductions in lesion area on days 7 and 14 after laser photocoagulation (Fig. 12A-B; p<0.05). Choroidal flatmount with fluorescein-conjugated isolectin B4 staining was used to assess the volume of the CNV lesions on days 7 and 14 after laser photocoagulation. Ten animals, 10 eyes and 20 lesions were examined in the control group, and 10 animals, 10 eyes and 28 lesions were examined in the TSA group. Mice treated with TSA demonstrated a significantly smaller lesion size compared to those treated with PBS (Fig. 13A). Quantitative measurement of the volumes of the CNV lesions showed that TSA treatment resulted in an approximately 65.5% and 65.9% reduction, respectively, in choroidal vascular volume, compared with that of control mice, 1 week and 2 weeks after laser photocoagulation (Fig. 13B; p<0.05).

Bottom Line: Most strikingly, TSA markedly down-regulates the expression of VEGF receptor-2 in human vascular endothelial cells and, thus, can knock down pro-angiogenic cell signaling.Additionally, TSA suppresses CNV-associated wound healing response and RPE epithelial-mesenchymal transdifferentiation.This report suggests that TSA, and possibly HDACi's in general, should be further evaluated for their therapeutic potential for the treatment of CNV.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Keck School of Medicine of the University of Southern California, Los Angeles, CA, United States of America; Doheny Eye Institute, Los Angeles, CA, United States of America.

ABSTRACT
Choroidal neovascularization (CNV) is a blinding complication of age-related macular degeneration that manifests as the growth of immature choroidal blood vessels through Bruch's membrane, where they can leak fluid or hemorrhage under the retina. Here, we demonstrate that the histone deacetylase inhibitor (HDACi) trichostatin A (TSA) can down-regulate the pro-angiogenic hypoxia-inducible factor-1α and vascular endothelial growth factor (VEGF), and up-regulate the anti-angiogenic and neuro-protective pigment epithelium derived factor in human retinal pigment epithelial (RPE) cells. Most strikingly, TSA markedly down-regulates the expression of VEGF receptor-2 in human vascular endothelial cells and, thus, can knock down pro-angiogenic cell signaling. Additionally, TSA suppresses CNV-associated wound healing response and RPE epithelial-mesenchymal transdifferentiation. In the laser-induced model of CNV using C57Bl/6 mice, systemic administration of TSA significantly reduces fluorescein leakage and the size of CNV lesions at post-laser days 7 and 14 as well as the immunohistochemical expression of VEGF, VEGFR2, and smooth muscle actin in CNV lesions at post-laser day 7. This report suggests that TSA, and possibly HDACi's in general, should be further evaluated for their therapeutic potential for the treatment of CNV.

No MeSH data available.


Related in: MedlinePlus