Temporal pattern of Toll-like receptor 9 upregulation in neurons and glial cells following cerebral ischemia reperfusion in mice

Abstract

Purpose: The family of Toll-like receptors (TLRs) has recently been reported to play a role in ischemic injury, but the time course and cell types of the post-stroke TLR9 upregulation remain unclear. In this study, we investigated the dynamic changes of TLR9 expression and the expression of TLR9 in neurons and glial cells after cerebral ischemia reperfusion in mice. Methods: Focal cerebral ischemia was induced by middle cerebral artery occlusion for 90 min in male C57BL/6 mice. The TLR9 expression levels in the tissue surrounding the infarct were detected by Western Blot at 6 h, 3 d, 7 d, 14 d, 21 d, and 28 d after reperfusion. The expression of TLR9 in neurons and glial cells was observed by immunofluorescence staining. Results: The expression of TLR9 protein first increased and then decreased, with the peak observed at 14 d–21 d. Only small punctate intracellular TLR9 was occasionally observed in the neurons at each time point, and the TLR9-positive rate showed no difference at different time points. By contrast, the activated microglia gathered at the margin of the infarct, and the intracellular TLR9 changed from scattered small punctate to coarse and lumpy. The TLR9-positive rate of microglia was first increased and then decreased with time, with the peak observed at 3 d. No positive TLR9 staining was found in the astrocytes and oligodendrocytes. Conclusions: TLR9 expression showed dynamic changes for a long period of time and microglias were the main brain cells to express TLR9 after cerebral ischemia and reperfusion.

KEYWORDS:

• cerebral ischemia
• reperfusion
• Toll-like receptor 9
• microglia
• neuron

Acknowledgements

This study was supported by National Natural Science Foundation of China (No. 81171103) and Guangdong Natural Science Foundation (No.9151008901000028, and S2011010006043).

Declaration of Interest

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper.