Upregulation of RyR2 in hypoxic/reperfusion injury

Varun Kesherwani, Sandeep K. Agrawal

Research output: Contribution to journalArticle

9 Scopus citations

Abstract

Calcium influx into cells is responsible for initiating the cell death in neuronal tissue after hypoxic injury. Changes in intracellular calcium with subsequent increased expression of ryanodine receptor 2 (RyR2) are hypothesized to cause cell death after hypoxic injury. In the present study we have examined the time-dependent changes of RyR2 expression in hypoxic/reperfusion injury of spinal cord dorsal column. In this study we used western blotting, real time PCR (RT-PCR) and immunohistochemistry to examine changes in protein and gene expression of RyR2 after spinal cord injury (SCI) in the rat. Quantitative immunoblotting showed increase in the expression of RyR2 at 4h during hypoxic/reperfusion injury of dorsal column. Moreover, RT-PCR showed 36-fold increases in mRNA of RyR2 after 4h of hypoxic injury of white matter. By double immunofluorescence staining, RyR2 was localized on axons and astrocytes in the white matter of the spinal cord. After treatment with KN-62; (inhibitor of CaMKII) and SP600125 (inhibitor of JNK), there is a significant reduction in the expression of RyR2, indicating the role of these molecules in RyR2 regulation. Further removal of extracellular calcium does not have significant effect on RyR2 expression and phosphorylation of CaMKII, which was further confirmed by treatment with intracellular Ca ++ chelator BAPTA-AM. Finally, bioassay with quantitative analysis showed that treatment with inhibitor significantly reduced the cellular oxidative stress suggesting RyR2 is responsible for increased cellular oxidative load. In summary, we provide evidence that RyR2 gene and protein expression in astrocyte and axons is markedly increased after hypoxic injury. Further CaMKII/JNK pathway upregulates RyR2 expression after hypoxic injury. Therefore we propose that inhibitors of CaMKII/JNK pathway would reduce the cellular oxidative load and thereby have a neuroprotective role.

Original languageEnglish (US)
Pages (from-to)1255-1265
Number of pages11
JournalJournal of Neurotrauma
Volume29
Issue number6
DOIs
Publication statusPublished - Apr 10 2012

    Fingerprint

Keywords

  • Ca
  • CaMKII
  • JNK
  • RyR2
  • SCI
  • hypoxia/reperfusion

ASJC Scopus subject areas

  • Clinical Neurology

Cite this