Establishment of human retinal microvascular endothelial cells with extended life-span

Meghana V. Kashyap, Amalendu Prakash Ranjan, Jwalitha Shankardas, Jamboor K. Vishwanatha

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

Aim: To generate and characterize a telomerase-immortalized human retinal microvascular endothelial cell (HREC) line. This cell line may be utilized as an in vitro model to study the molecular basis of several diseases of the human retina. Materials and Methods: Primary retinal neuronal cells were isolated and transfected with plasmid encoding full-length human telomerase reverse transcriptase (hTERT). Transfected cells were selected and characterized to determine telomerase activity, karyotype, proliferative capacity and functionality. Results: HREC-hTERT cells appear morphologically similar to primary endothelial cells and have an extended in vitro lifespan. HREC-hTERT cells express the progenitor/stem cell marker nestin. They have active telomerase and a high proliferative capacity. These cells also maintain a diploid karyotype. The HREC-hTERT cells showed high colony-formation capacity and plating efficiency compared to the primary cells. These cells are capable of differentiation into neuronal and glial cell phenotypes and the differentiated cells express the astrocyte marker glial fibrillary acidic protein (GFAP) and the neuronal marker microtubule-associated protein-2 (MAP2), respectively. Conclusion: The in vitro lifespan of human retinal neuronal endothelial cells can be extended by ectopic expression of hTERT without altering the genetic stability and functionality of these cells. These cells will be a valuable tool to further our understanding on the role of HRECs in the human blood-retinal-barrier and in angiogenesis and neovascularization.

Original languageEnglish
Pages (from-to)685-694
Number of pages10
JournalIn Vivo
Volume27
Issue number6
StatePublished - 1 Nov 2013

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Endothelial cells
Endothelial Cells
Telomerase
Nestin
Karyotype
Microtubule-Associated Proteins
Glial Fibrillary Acidic Protein
Stem cells
Plating
Stem Cells
Blood-Retinal Barrier
Plasmids
Blood
Cell Line
Cells
human TERT protein
Diploidy
Neuroglia
Astrocytes
Retina

Keywords

  • Extended life-span
  • Neuronal endothelial cells
  • Retina
  • Telomerase

Cite this

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title = "Establishment of human retinal microvascular endothelial cells with extended life-span",
abstract = "Aim: To generate and characterize a telomerase-immortalized human retinal microvascular endothelial cell (HREC) line. This cell line may be utilized as an in vitro model to study the molecular basis of several diseases of the human retina. Materials and Methods: Primary retinal neuronal cells were isolated and transfected with plasmid encoding full-length human telomerase reverse transcriptase (hTERT). Transfected cells were selected and characterized to determine telomerase activity, karyotype, proliferative capacity and functionality. Results: HREC-hTERT cells appear morphologically similar to primary endothelial cells and have an extended in vitro lifespan. HREC-hTERT cells express the progenitor/stem cell marker nestin. They have active telomerase and a high proliferative capacity. These cells also maintain a diploid karyotype. The HREC-hTERT cells showed high colony-formation capacity and plating efficiency compared to the primary cells. These cells are capable of differentiation into neuronal and glial cell phenotypes and the differentiated cells express the astrocyte marker glial fibrillary acidic protein (GFAP) and the neuronal marker microtubule-associated protein-2 (MAP2), respectively. Conclusion: The in vitro lifespan of human retinal neuronal endothelial cells can be extended by ectopic expression of hTERT without altering the genetic stability and functionality of these cells. These cells will be a valuable tool to further our understanding on the role of HRECs in the human blood-retinal-barrier and in angiogenesis and neovascularization.",
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author = "Kashyap, {Meghana V.} and Ranjan, {Amalendu Prakash} and Jwalitha Shankardas and Vishwanatha, {Jamboor K.}",
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Establishment of human retinal microvascular endothelial cells with extended life-span. / Kashyap, Meghana V.; Ranjan, Amalendu Prakash; Shankardas, Jwalitha; Vishwanatha, Jamboor K.

In: In Vivo, Vol. 27, No. 6, 01.11.2013, p. 685-694.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Establishment of human retinal microvascular endothelial cells with extended life-span

AU - Kashyap, Meghana V.

AU - Ranjan, Amalendu Prakash

AU - Shankardas, Jwalitha

AU - Vishwanatha, Jamboor K.

PY - 2013/11/1

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N2 - Aim: To generate and characterize a telomerase-immortalized human retinal microvascular endothelial cell (HREC) line. This cell line may be utilized as an in vitro model to study the molecular basis of several diseases of the human retina. Materials and Methods: Primary retinal neuronal cells were isolated and transfected with plasmid encoding full-length human telomerase reverse transcriptase (hTERT). Transfected cells were selected and characterized to determine telomerase activity, karyotype, proliferative capacity and functionality. Results: HREC-hTERT cells appear morphologically similar to primary endothelial cells and have an extended in vitro lifespan. HREC-hTERT cells express the progenitor/stem cell marker nestin. They have active telomerase and a high proliferative capacity. These cells also maintain a diploid karyotype. The HREC-hTERT cells showed high colony-formation capacity and plating efficiency compared to the primary cells. These cells are capable of differentiation into neuronal and glial cell phenotypes and the differentiated cells express the astrocyte marker glial fibrillary acidic protein (GFAP) and the neuronal marker microtubule-associated protein-2 (MAP2), respectively. Conclusion: The in vitro lifespan of human retinal neuronal endothelial cells can be extended by ectopic expression of hTERT without altering the genetic stability and functionality of these cells. These cells will be a valuable tool to further our understanding on the role of HRECs in the human blood-retinal-barrier and in angiogenesis and neovascularization.

AB - Aim: To generate and characterize a telomerase-immortalized human retinal microvascular endothelial cell (HREC) line. This cell line may be utilized as an in vitro model to study the molecular basis of several diseases of the human retina. Materials and Methods: Primary retinal neuronal cells were isolated and transfected with plasmid encoding full-length human telomerase reverse transcriptase (hTERT). Transfected cells were selected and characterized to determine telomerase activity, karyotype, proliferative capacity and functionality. Results: HREC-hTERT cells appear morphologically similar to primary endothelial cells and have an extended in vitro lifespan. HREC-hTERT cells express the progenitor/stem cell marker nestin. They have active telomerase and a high proliferative capacity. These cells also maintain a diploid karyotype. The HREC-hTERT cells showed high colony-formation capacity and plating efficiency compared to the primary cells. These cells are capable of differentiation into neuronal and glial cell phenotypes and the differentiated cells express the astrocyte marker glial fibrillary acidic protein (GFAP) and the neuronal marker microtubule-associated protein-2 (MAP2), respectively. Conclusion: The in vitro lifespan of human retinal neuronal endothelial cells can be extended by ectopic expression of hTERT without altering the genetic stability and functionality of these cells. These cells will be a valuable tool to further our understanding on the role of HRECs in the human blood-retinal-barrier and in angiogenesis and neovascularization.

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