Creatine kinase in human retinal pigment epithelium

Brian Kennedy, B. E. Haley, R. K. Geb, K. U. Loeffler, N. J. Mangini

Research output: Contribution to journalArticle

Abstract

Purpose. RPE transport activity, and consequent ATP consumption, vary dramatically as a function of photoreceptor light exposure. The present study was designed to assess ATP buffering capacity in the RPE. Specifically, to determine if the RPE possesses creatine kinase (CK) activity which could serve to maintain ATP levels during periods of high metabolic demand. Method. Multiple CK Isofprms (both cytosolic and mitochondrial) have been described in various tissues. Isoform specific antibodies were used to identify those isoforms present in the RPE. Immunohistochemistry was performed on paraffin-embedded sections of native human RPE (HRPE). Additionally, SDS-PAGE followed by Western blotting was carried out in HRPE maintained in cell culture. Finally, an enzyme-coupled spectrophotometric assay was used to quantitate CK activity in lysates from cultured HRPE. Result. By Western blotting, the cytosolic CK B monomer as well as the mitochondrial sMt-CK isoform were identified in cultured HRPE. Both isoforms were also found to be present, by positive immunostaining, in the native tissue sections. CK activity was measured in cell suspensions from four different donor cultures. Activity was 0.062± 0.010 nmol/mg protein/min. The predominant isoform by activity measurement was CK B. Conclusion. CK has been identified by Western blot, immunohistochemistry and activity measurements in HRPE.

Original languageEnglish (US)
JournalInvestigative Ophthalmology and Visual Science
Volume38
Issue number4
StatePublished - 1997

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Retinal Pigment Epithelium
Creatine Kinase
Protein Isoforms
Adenosine Triphosphate
Western Blotting
Mitochondrial Form Creatine Kinase
Immunohistochemistry
Population Groups
Paraffin
Polyacrylamide Gel Electrophoresis
Suspensions
Cell Culture Techniques
Tissue Donors
Light
Antibodies
Enzymes

ASJC Scopus subject areas

  • Ophthalmology

Cite this

Kennedy, B., Haley, B. E., Geb, R. K., Loeffler, K. U., & Mangini, N. J. (1997). Creatine kinase in human retinal pigment epithelium. Investigative Ophthalmology and Visual Science, 38(4).

Creatine kinase in human retinal pigment epithelium. / Kennedy, Brian; Haley, B. E.; Geb, R. K.; Loeffler, K. U.; Mangini, N. J.

In: Investigative Ophthalmology and Visual Science, Vol. 38, No. 4, 1997.

Research output: Contribution to journalArticle

Kennedy, B, Haley, BE, Geb, RK, Loeffler, KU & Mangini, NJ 1997, 'Creatine kinase in human retinal pigment epithelium', Investigative Ophthalmology and Visual Science, vol. 38, no. 4.
Kennedy, Brian ; Haley, B. E. ; Geb, R. K. ; Loeffler, K. U. ; Mangini, N. J. / Creatine kinase in human retinal pigment epithelium. In: Investigative Ophthalmology and Visual Science. 1997 ; Vol. 38, No. 4.
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