Increased retinal expression of the pro-angiogenic receptor GPR91 via BMP6 in a mouse model of juvenile hemochromatosis

Pachiappan Arjunan, Jaya P. Gnanaprakasam, Sudha Ananth, Michelle A. Romej, Veeranan Karmegam Rajalakshmi, Puttur D. Prasad, Pamela M. Martin, Mariappan Gurusamy, Muthusamy Thangaraju, Yangzom D. Bhutia, Vadivel Ganapathy

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

PURPOSE. Hemochromatosis, an iron-overload disease, occurs as adult and juvenile types. Mutations in hemojuvelin (HJV), an iron-regulatory protein and a bone orphogenetic protein (BMP) coreceptor, underlie most of the juvenile type. Hjv-/-mice accumulate excess iron in retina and exhibit aberrant vascularization and angiomas. A succinate receptor, GPR91, is pro-angiogenic in retina. We hypothesized that Hjv-/- retinas have increased BMP signaling and increased GPR91 expression as the basis of angiomas. METHODS. Expression of GPR91 was examined by qPCR, immunofluorescence, and Western blot in wild-type and Hjv-/- mouse retinas and pRPE cells. Influence of excess iron and BMP6 on GPR91 expression was investigated in ARPE-19 cells, and wild-type and Hjv-/- pRPE cells. Succinate was used to activate GPR91 and determine the effects of GPR91 signaling on VEGF expression. Signaling of BMP6 was studied by the expression of Smad1/5/8 and pSmad4, and the BMP-target gene Id1. The interaction of pSmad4 with GPR91 promoter was studied by ChIP. RESULTS. Expression of GPR91 was higher in Hjv-/- retinas and RPE than in wild-type counterparts. Unexpectedly, BMP signaling was increased, not decreased, in Hjv-/- retinas and RPE. Bone morphogenetic protein 6 induced GPR91 in RPE, suggesting that increased BMP signaling in Hjv-/- retinas was likely responsible for GPR91 upregulation. Exposure of RPE to excess iron and succinate as well as BMP6 and succinate increased VEGF expression. Bone morphogenetic protein 6 promoted the interaction of pSmad4 with GPR91 promoter in RPE. CONCLUSIONS. G-protein-coupled receptor 91 is a BMP6 target and Hjv deletion enhances BMP signaling in retina, thus underscoring a role for excess iron and hemochromatosis in abnormal retinal vascularization.

Original languageEnglish (US)
Pages (from-to)1612-1619
Number of pages8
JournalInvestigative Ophthalmology and Visual Science
Volume57
Issue number4
DOIs
StatePublished - Apr 1 2016

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Retina
Succinic Acid
Bone Morphogenetic Protein 6
Iron
Hemochromatosis
Hemangioma
Vascular Endothelial Growth Factor A
Iron-Regulatory Proteins
Iron Overload
Type 2 Hemochromatosis
G-Protein-Coupled Receptors
Fluorescent Antibody Technique
Up-Regulation
Western Blotting
Bone and Bones
Mutation
Genes
Proteins

Keywords

  • Age-related macular degeneration
  • BMP6 signaling
  • Juvenile hemochromatosis
  • Retinal pigment epithelium
  • Succinate receptor-GPR91

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

Cite this

Increased retinal expression of the pro-angiogenic receptor GPR91 via BMP6 in a mouse model of juvenile hemochromatosis. / Arjunan, Pachiappan; Gnanaprakasam, Jaya P.; Ananth, Sudha; Romej, Michelle A.; Rajalakshmi, Veeranan Karmegam; Prasad, Puttur D.; Martin, Pamela M.; Gurusamy, Mariappan; Thangaraju, Muthusamy; Bhutia, Yangzom D.; Ganapathy, Vadivel.

In: Investigative Ophthalmology and Visual Science, Vol. 57, No. 4, 01.04.2016, p. 1612-1619.

Research output: Contribution to journalArticle

Arjunan, Pachiappan ; Gnanaprakasam, Jaya P. ; Ananth, Sudha ; Romej, Michelle A. ; Rajalakshmi, Veeranan Karmegam ; Prasad, Puttur D. ; Martin, Pamela M. ; Gurusamy, Mariappan ; Thangaraju, Muthusamy ; Bhutia, Yangzom D. ; Ganapathy, Vadivel. / Increased retinal expression of the pro-angiogenic receptor GPR91 via BMP6 in a mouse model of juvenile hemochromatosis. In: Investigative Ophthalmology and Visual Science. 2016 ; Vol. 57, No. 4. pp. 1612-1619.
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abstract = "PURPOSE. Hemochromatosis, an iron-overload disease, occurs as adult and juvenile types. Mutations in hemojuvelin (HJV), an iron-regulatory protein and a bone orphogenetic protein (BMP) coreceptor, underlie most of the juvenile type. Hjv-/-mice accumulate excess iron in retina and exhibit aberrant vascularization and angiomas. A succinate receptor, GPR91, is pro-angiogenic in retina. We hypothesized that Hjv-/- retinas have increased BMP signaling and increased GPR91 expression as the basis of angiomas. METHODS. Expression of GPR91 was examined by qPCR, immunofluorescence, and Western blot in wild-type and Hjv-/- mouse retinas and pRPE cells. Influence of excess iron and BMP6 on GPR91 expression was investigated in ARPE-19 cells, and wild-type and Hjv-/- pRPE cells. Succinate was used to activate GPR91 and determine the effects of GPR91 signaling on VEGF expression. Signaling of BMP6 was studied by the expression of Smad1/5/8 and pSmad4, and the BMP-target gene Id1. The interaction of pSmad4 with GPR91 promoter was studied by ChIP. RESULTS. Expression of GPR91 was higher in Hjv-/- retinas and RPE than in wild-type counterparts. Unexpectedly, BMP signaling was increased, not decreased, in Hjv-/- retinas and RPE. Bone morphogenetic protein 6 induced GPR91 in RPE, suggesting that increased BMP signaling in Hjv-/- retinas was likely responsible for GPR91 upregulation. Exposure of RPE to excess iron and succinate as well as BMP6 and succinate increased VEGF expression. Bone morphogenetic protein 6 promoted the interaction of pSmad4 with GPR91 promoter in RPE. CONCLUSIONS. G-protein-coupled receptor 91 is a BMP6 target and Hjv deletion enhances BMP signaling in retina, thus underscoring a role for excess iron and hemochromatosis in abnormal retinal vascularization.",
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T1 - Increased retinal expression of the pro-angiogenic receptor GPR91 via BMP6 in a mouse model of juvenile hemochromatosis

AU - Arjunan, Pachiappan

AU - Gnanaprakasam, Jaya P.

AU - Ananth, Sudha

AU - Romej, Michelle A.

AU - Rajalakshmi, Veeranan Karmegam

AU - Prasad, Puttur D.

AU - Martin, Pamela M.

AU - Gurusamy, Mariappan

AU - Thangaraju, Muthusamy

AU - Bhutia, Yangzom D.

AU - Ganapathy, Vadivel

PY - 2016/4/1

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N2 - PURPOSE. Hemochromatosis, an iron-overload disease, occurs as adult and juvenile types. Mutations in hemojuvelin (HJV), an iron-regulatory protein and a bone orphogenetic protein (BMP) coreceptor, underlie most of the juvenile type. Hjv-/-mice accumulate excess iron in retina and exhibit aberrant vascularization and angiomas. A succinate receptor, GPR91, is pro-angiogenic in retina. We hypothesized that Hjv-/- retinas have increased BMP signaling and increased GPR91 expression as the basis of angiomas. METHODS. Expression of GPR91 was examined by qPCR, immunofluorescence, and Western blot in wild-type and Hjv-/- mouse retinas and pRPE cells. Influence of excess iron and BMP6 on GPR91 expression was investigated in ARPE-19 cells, and wild-type and Hjv-/- pRPE cells. Succinate was used to activate GPR91 and determine the effects of GPR91 signaling on VEGF expression. Signaling of BMP6 was studied by the expression of Smad1/5/8 and pSmad4, and the BMP-target gene Id1. The interaction of pSmad4 with GPR91 promoter was studied by ChIP. RESULTS. Expression of GPR91 was higher in Hjv-/- retinas and RPE than in wild-type counterparts. Unexpectedly, BMP signaling was increased, not decreased, in Hjv-/- retinas and RPE. Bone morphogenetic protein 6 induced GPR91 in RPE, suggesting that increased BMP signaling in Hjv-/- retinas was likely responsible for GPR91 upregulation. Exposure of RPE to excess iron and succinate as well as BMP6 and succinate increased VEGF expression. Bone morphogenetic protein 6 promoted the interaction of pSmad4 with GPR91 promoter in RPE. CONCLUSIONS. G-protein-coupled receptor 91 is a BMP6 target and Hjv deletion enhances BMP signaling in retina, thus underscoring a role for excess iron and hemochromatosis in abnormal retinal vascularization.

AB - PURPOSE. Hemochromatosis, an iron-overload disease, occurs as adult and juvenile types. Mutations in hemojuvelin (HJV), an iron-regulatory protein and a bone orphogenetic protein (BMP) coreceptor, underlie most of the juvenile type. Hjv-/-mice accumulate excess iron in retina and exhibit aberrant vascularization and angiomas. A succinate receptor, GPR91, is pro-angiogenic in retina. We hypothesized that Hjv-/- retinas have increased BMP signaling and increased GPR91 expression as the basis of angiomas. METHODS. Expression of GPR91 was examined by qPCR, immunofluorescence, and Western blot in wild-type and Hjv-/- mouse retinas and pRPE cells. Influence of excess iron and BMP6 on GPR91 expression was investigated in ARPE-19 cells, and wild-type and Hjv-/- pRPE cells. Succinate was used to activate GPR91 and determine the effects of GPR91 signaling on VEGF expression. Signaling of BMP6 was studied by the expression of Smad1/5/8 and pSmad4, and the BMP-target gene Id1. The interaction of pSmad4 with GPR91 promoter was studied by ChIP. RESULTS. Expression of GPR91 was higher in Hjv-/- retinas and RPE than in wild-type counterparts. Unexpectedly, BMP signaling was increased, not decreased, in Hjv-/- retinas and RPE. Bone morphogenetic protein 6 induced GPR91 in RPE, suggesting that increased BMP signaling in Hjv-/- retinas was likely responsible for GPR91 upregulation. Exposure of RPE to excess iron and succinate as well as BMP6 and succinate increased VEGF expression. Bone morphogenetic protein 6 promoted the interaction of pSmad4 with GPR91 promoter in RPE. CONCLUSIONS. G-protein-coupled receptor 91 is a BMP6 target and Hjv deletion enhances BMP signaling in retina, thus underscoring a role for excess iron and hemochromatosis in abnormal retinal vascularization.

KW - Age-related macular degeneration

KW - BMP6 signaling

KW - Juvenile hemochromatosis

KW - Retinal pigment epithelium

KW - Succinate receptor-GPR91

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