May 2003
Volume 44, Issue 13
ARVO Annual Meeting Abstract  |   May 2003
Activation of Downstream Pathways after Calpain-induced Proteolysis in the Hypoxic Rat Retina
Author Affiliations & Notes
  • Y. Tamada
    Senju Pharmaceutical Co Ltd, Kobe, Japan
  • E. Nakajima
    Senju Pharmaceutical Co Ltd, Kobe, Japan
  • M. Azuma
    Senju Pharmaceutical Co Ltd, Kobe, Japan
  • T.R. Shearer
    Oral Molecular Biology, Oregon Health & Science University, Portland, OR, United States
  • Footnotes
    Commercial Relationships  Y. Tamada, Senju Pharmaceutical Co., Ltd. E; E. Nakajima, Senju Pharmaceutical Co., Ltd. E; M. Azuma, Senju Pharmaceutical Co., Ltd. E; T.R. Shearer, Senju Pharmaceutical Co., Ltd. C.
  • Footnotes
    Support  NIH grant EY05786 to TRS
Investigative Ophthalmology & Visual Science May 2003, Vol.44, 3567. doi:
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      Y. Tamada, E. Nakajima, M. Azuma, T.R. Shearer; Activation of Downstream Pathways after Calpain-induced Proteolysis in the Hypoxic Rat Retina . Invest. Ophthalmol. Vis. Sci. 2003;44(13):3567.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract: : Purpose: In our previous studies with rats, calpain-induced proteolysis was an important mid-stage event in retinal cell death induced in vivo by ischemia-reperfusion injury or by hypoxic cell culture. The purpose of present study was to identify the downstream pathways activated after calpain-induced proteolysis in hypoxic rat retinas. Methods: To induce hypoxia, rat retinas were incubated in RPMI medium without glucose under 95 % N2/5 % CO2. Leakage of LDH into the medium measured retinal cell death. Activation of calpain and proteolysis of calpain substrates were analyzed by casein zymography and immunoblotting. Results: LDH leaked into the medium from retinas under hypoxic conditions. Caseinolytic activity of calpains decreased with hypoxia, suggesting calpain activation followed by autolytic degradation. Caspase-3, previously identified as a executor of apoptosis, was cleaved by calpain to a 29 kDa fragment. This initial cleavage of caspase-3 is thought to facilitate subsequent cleavage into active caspase. αB-crystallin, whose interaction with caspase-3 prevents apoptosis, was degraded in retina with hypoxia. Cytoskeletal proteins such as α-spectrin and tau, required for the maintenance of proper membrane order and integrity, were proteolyzed in hypoxic retina. Conclusions: These results suggested that activation of caspase-3, reduction of the anti-apoptotic effect of αB-crystallin, and proteolysis of α-spectrin and tau after calpain activation contributed to retinal cell death induced by hypoxia in vitro. Dr. Shearer has a significant financial interest (research contract and consulting fee) in Senju Pharmaceutical Co., Ltd.

Keywords: calcium • retinal degenerations: cell biology • proteolysis 

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