Putative role of protein kinase C in neurotoxic inflammation mediated by extracellular heat shock protein 70 after ischemia-reperfusion

Journal of Neuroinflammation

Dmitry Ivanov²  Xenia Dvoriantchikova¹  Ali M Saeed³  Andrea Rachelle C Santos¹  Galina Dvoriantchikova¹ 

[1] Bascom Palmer Eye Institute, Department of Ophthalmology, University of Miami Miller School of Medicine, 1638 NW 10th Ave, Miami, FL 33136, USA;Department of Microbiology and Immunology, University of Miami Miller School of Medicine, 1600 NW 10th Ave, Miami, FL 33136, USA;Sheila and David Fuente Program in Cancer Biology, University of Miami Miller School of Medicine, 1550 NW 10th Ave, Miami, FL 33136, USA

关键词: Neuronal death;    Protein kinase C (PKC);    Polymyxin B;    Endotoxins;    Toll-like receptor 4 (Tlr4);    Extracellular heat shock protein 70 (Hsp70);    Damage-associated molecular patterns (DAMPs);    Sterile inflammation;    Ischemia-reperfusion (IR) injury;   

Others  :  804463
DOI  :  10.1186/1742-2094-11-81

 received in 2014-01-07, accepted in 2014-04-04,  发布年份 2014

【 摘 要 】

Background

Sterile inflammation occurs in the absence of live pathogens and is an unavoidable consequence of ischemia-reperfusion (IR) injury in the central nervous system (CNS). It is known that toll-like receptor 4 (Tlr4) contributes to damage and sterile inflammation in the CNS mediated by IR. However, the mechanism of Tlr4 activation under sterile conditions in ischemic tissue is poorly understood. We performed this study to clarify the mechanism. To this end, we focused on the extracellular heat shock protein 70 (Hsp70), the prototypic Tlr4 ligand.

Methods

Tlr4-, Myd88- and Trif-knockout animals, as well as C57BL/6 mice, were used for the wild type control. For the in vivo study, we used a mouse model of retinal IR injury. To test the role of protein kinase C (PKC) in IR injury, IR retinas were treated with the PKC inhibitors (polymyxin B and Gö6976) and retinal damage was evaluated by directly counting neurons in the ganglion cell layer of flat-mounted retinas seven days after IR. Primary retinal neurons (retinal ganglion cells) and glial cells were used for in vitro experiments. Quantitative RT-PCR, ELISA and western blot analysis were used to study the production of pro-inflammatory factors in IR retinas and in primary cell cultures.

Results

We found significant accumulation of extracellular Hsp70 in a model of retinal IR injury. We noted that PKC was involved in Tlr4 signaling, and found that PKC inhibitors promoted neuroprotection by reducing pro-inflammatory activity in ischemic tissue. To put all of the pieces in the signaling cascade together, we performed an in vitro study. We found that PKC was critical to mediate the Hsp70-dependent pro-inflammatory response. At the same time, the contamination of Hsp70 preparations with low-dose endotoxin was not critical to mediate the production of pro-inflammatory factors. We found that extracellular Hsp70 can promote neuronal death at least, by mediating production of cytotoxic levels of tumor necrosis factor alpha, predominantly due to the Tlr4/Myd88 signaling cascade.

Conclusions

Our findings suggest that PKC acts as a switch to amplify the pro-inflammatory activity of Hsp70/Tlr4 signaling, which is sufficient to mediate neuronal death.

【 授权许可】

   
2014 Dvoriantchikova et al.; licensee BioMed Central Ltd.

【 图 表 】

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