A porcine anterior segment perfusion and transduction model with direct visualization of the trabecular meshwork

Ralitsa T. Loewen, Pritha Roy, Daniel B. Park, Adrianna Jensen, Gordon Scott, Devora Cohen-Karni, Michael P Fautsch, Joel S. Schuman, Nils A. Loewen

Research output: Contribution to journalArticle

15 Citations (Scopus)

Abstract

PURPOSE. To establish a consistent and affordable, high quality porcine anterior segment perfusion and transduction model that allows direct visualization of the trabecular meshwork. METHODS. Porcine anterior segments were cultured within 2 hours of death by removing lens and uvea and securing in a specially designed petri dish with a thin bottom to allow direct visualization of the trabecular meshwork with minimal distortion. Twenty-two control eyes (CO) with a constant flow rate were compared to eight gravity perfused eyes (COgr, 15 mm Hg). We established gene delivery to the TM using eGFP expressing feline immunodeficiency virus (FIV) vector GINSIN at 108 transducing units (TU) per eye (GINSIN_8, n = 8) and 107 TU (GINSIN_7, n = 8). Expression was assessed for 14 days before histology was obtained. RESULTS. Pig eyes were a reliable source for consistent and high quality anterior segment cultures with a low failure rate of 12%. Control eyes had an intraocular pressure (IOP) of 15.8 ± 1.9 mm Hg at fixed pump perfusion with 3 µL/min compared to gravity perfused COgr with imputed 3.7 ± 1.6 µL/min. Vector GINSIN_8 eyes experienced a transient posttransduction IOP increase of 44% that resolved at 48 hours; this was not observed in GINSIN_7 eyes. Expression was higher in GINSIN_8 than in GINSIN_7 eyes. Trabecular meshwork architecture was well preserved. CONCLUSIONS. Compared with previously used human donor eyes, this inexpensive porcine anterior segment perfusion model is of sufficient, repeatable high quality to develop strategies of TM bioengineering. Trabecular meshwork could be observed directly. Despite significant anatomic differences, effects of transduction replicate the main aspects of previously explored human, feline and rodent models.

Original languageEnglish (US)
Pages (from-to)1338-1344
Number of pages7
JournalInvestigative Ophthalmology and Visual Science
Volume57
Issue number3
DOIs
StatePublished - Mar 1 2016

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Trabecular Meshwork
Swine
Perfusion
Gravitation
Intraocular Pressure
Uvea
Feline Immunodeficiency Virus
Infusion Pumps
Bioengineering
Felidae
Lenses
Rodentia
Histology

Keywords

  • EGFP
  • Glaucoma
  • Lentiviral transduction
  • Porcine eyes
  • Trabecular meshwork

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

Cite this

A porcine anterior segment perfusion and transduction model with direct visualization of the trabecular meshwork. / Loewen, Ralitsa T.; Roy, Pritha; Park, Daniel B.; Jensen, Adrianna; Scott, Gordon; Cohen-Karni, Devora; Fautsch, Michael P; Schuman, Joel S.; Loewen, Nils A.

In: Investigative Ophthalmology and Visual Science, Vol. 57, No. 3, 01.03.2016, p. 1338-1344.

Research output: Contribution to journalArticle

Loewen, Ralitsa T. ; Roy, Pritha ; Park, Daniel B. ; Jensen, Adrianna ; Scott, Gordon ; Cohen-Karni, Devora ; Fautsch, Michael P ; Schuman, Joel S. ; Loewen, Nils A. / A porcine anterior segment perfusion and transduction model with direct visualization of the trabecular meshwork. In: Investigative Ophthalmology and Visual Science. 2016 ; Vol. 57, No. 3. pp. 1338-1344.
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abstract = "PURPOSE. To establish a consistent and affordable, high quality porcine anterior segment perfusion and transduction model that allows direct visualization of the trabecular meshwork. METHODS. Porcine anterior segments were cultured within 2 hours of death by removing lens and uvea and securing in a specially designed petri dish with a thin bottom to allow direct visualization of the trabecular meshwork with minimal distortion. Twenty-two control eyes (CO) with a constant flow rate were compared to eight gravity perfused eyes (COgr, 15 mm Hg). We established gene delivery to the TM using eGFP expressing feline immunodeficiency virus (FIV) vector GINSIN at 108 transducing units (TU) per eye (GINSIN_8, n = 8) and 107 TU (GINSIN_7, n = 8). Expression was assessed for 14 days before histology was obtained. RESULTS. Pig eyes were a reliable source for consistent and high quality anterior segment cultures with a low failure rate of 12{\%}. Control eyes had an intraocular pressure (IOP) of 15.8 ± 1.9 mm Hg at fixed pump perfusion with 3 µL/min compared to gravity perfused COgr with imputed 3.7 ± 1.6 µL/min. Vector GINSIN_8 eyes experienced a transient posttransduction IOP increase of 44{\%} that resolved at 48 hours; this was not observed in GINSIN_7 eyes. Expression was higher in GINSIN_8 than in GINSIN_7 eyes. Trabecular meshwork architecture was well preserved. CONCLUSIONS. Compared with previously used human donor eyes, this inexpensive porcine anterior segment perfusion model is of sufficient, repeatable high quality to develop strategies of TM bioengineering. Trabecular meshwork could be observed directly. Despite significant anatomic differences, effects of transduction replicate the main aspects of previously explored human, feline and rodent models.",
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T1 - A porcine anterior segment perfusion and transduction model with direct visualization of the trabecular meshwork

AU - Loewen, Ralitsa T.

AU - Roy, Pritha

AU - Park, Daniel B.

AU - Jensen, Adrianna

AU - Scott, Gordon

AU - Cohen-Karni, Devora

AU - Fautsch, Michael P

AU - Schuman, Joel S.

AU - Loewen, Nils A.

PY - 2016/3/1

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N2 - PURPOSE. To establish a consistent and affordable, high quality porcine anterior segment perfusion and transduction model that allows direct visualization of the trabecular meshwork. METHODS. Porcine anterior segments were cultured within 2 hours of death by removing lens and uvea and securing in a specially designed petri dish with a thin bottom to allow direct visualization of the trabecular meshwork with minimal distortion. Twenty-two control eyes (CO) with a constant flow rate were compared to eight gravity perfused eyes (COgr, 15 mm Hg). We established gene delivery to the TM using eGFP expressing feline immunodeficiency virus (FIV) vector GINSIN at 108 transducing units (TU) per eye (GINSIN_8, n = 8) and 107 TU (GINSIN_7, n = 8). Expression was assessed for 14 days before histology was obtained. RESULTS. Pig eyes were a reliable source for consistent and high quality anterior segment cultures with a low failure rate of 12%. Control eyes had an intraocular pressure (IOP) of 15.8 ± 1.9 mm Hg at fixed pump perfusion with 3 µL/min compared to gravity perfused COgr with imputed 3.7 ± 1.6 µL/min. Vector GINSIN_8 eyes experienced a transient posttransduction IOP increase of 44% that resolved at 48 hours; this was not observed in GINSIN_7 eyes. Expression was higher in GINSIN_8 than in GINSIN_7 eyes. Trabecular meshwork architecture was well preserved. CONCLUSIONS. Compared with previously used human donor eyes, this inexpensive porcine anterior segment perfusion model is of sufficient, repeatable high quality to develop strategies of TM bioengineering. Trabecular meshwork could be observed directly. Despite significant anatomic differences, effects of transduction replicate the main aspects of previously explored human, feline and rodent models.

AB - PURPOSE. To establish a consistent and affordable, high quality porcine anterior segment perfusion and transduction model that allows direct visualization of the trabecular meshwork. METHODS. Porcine anterior segments were cultured within 2 hours of death by removing lens and uvea and securing in a specially designed petri dish with a thin bottom to allow direct visualization of the trabecular meshwork with minimal distortion. Twenty-two control eyes (CO) with a constant flow rate were compared to eight gravity perfused eyes (COgr, 15 mm Hg). We established gene delivery to the TM using eGFP expressing feline immunodeficiency virus (FIV) vector GINSIN at 108 transducing units (TU) per eye (GINSIN_8, n = 8) and 107 TU (GINSIN_7, n = 8). Expression was assessed for 14 days before histology was obtained. RESULTS. Pig eyes were a reliable source for consistent and high quality anterior segment cultures with a low failure rate of 12%. Control eyes had an intraocular pressure (IOP) of 15.8 ± 1.9 mm Hg at fixed pump perfusion with 3 µL/min compared to gravity perfused COgr with imputed 3.7 ± 1.6 µL/min. Vector GINSIN_8 eyes experienced a transient posttransduction IOP increase of 44% that resolved at 48 hours; this was not observed in GINSIN_7 eyes. Expression was higher in GINSIN_8 than in GINSIN_7 eyes. Trabecular meshwork architecture was well preserved. CONCLUSIONS. Compared with previously used human donor eyes, this inexpensive porcine anterior segment perfusion model is of sufficient, repeatable high quality to develop strategies of TM bioengineering. Trabecular meshwork could be observed directly. Despite significant anatomic differences, effects of transduction replicate the main aspects of previously explored human, feline and rodent models.

KW - EGFP

KW - Glaucoma

KW - Lentiviral transduction

KW - Porcine eyes

KW - Trabecular meshwork

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