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Complement gene expression by rabbit heart: upregulation by ischemia and reperfusion
K Yasojima1, K S Kilgore, R A Washington
1Kinsmen Laboratory of Neurological Research, University of British Columbia, Vancouver, Canada.
Insights
The heart produces complement proteins C3 and C9, challenging the liver-centric view. Ischemia and reperfusion significantly increase these complement gene expressions in the heart.
Area of Science:
- Cardiovascular Biology
- Immunology
- Molecular Biology
Background:
- The complement system plays a role in myocardial ischemia/reperfusion injury.
- The liver is traditionally considered the primary source of complement components.
Purpose of the Study:
- To investigate the expression of complement proteins C3 and C9 in the rabbit heart.
- To determine if ischemia and reperfusion affect complement gene expression in the heart.
Main Methods:
- Reverse-transcriptase polymerase chain reaction (RT-PCR) to detect mRNA for C3 and C9.
- Western blotting to confirm protein expression.
- Langendorff-perfused isolated rabbit hearts subjected to ischemia and reperfusion.
Main Results:
- Heart tissue expresses mRNAs for complement proteins C3 and C9, which are translated into proteins.
- Ischemia and reperfusion significantly upregulate C3 and C9 mRNA levels in the heart.
- Upregulation of C3 and C9 mRNA in the heart under stress exceeds levels found in normal liver.
Conclusions:
- The heart is capable of expressing complement system genes and proteins.
- Local complement production in the heart may significantly contribute to ischemic injury.
- Reperfusion augments complement expression, exacerbating myocardial damage.
Abstract:
Activation of the complement system has been implicated in the pathogenesis of myocardial ischemia/reperfusion injury. It has always been assumed that liver is the primary source of complement components. In the present study, we used the reverse-transcriptase polymerase chain reaction technique to establish that the mRNAs for complement proteins C3 and C9 are expressed in rabbit heart. Rabbit liver, brain, spleen, and kidney were also shown to express C3 and C9 mRNAs. We used Western blotting to establish that these mRNAs in heart are translated into the corresponding proteins. We further established that dramatic upregulation of the mRNAs occurred in Langendorff-perfused isolated hearts subjected to ischemia and reperfusion. C3 mRNA was always expressed at higher levels than was C9 mRNA, but C9 mRNA showed greater upregulation under stress. Compared with levels in control hearts subjected to 5 minutes of normoxic perfusion, hearts subjected to 0.5 hours of ischemia followed by 1 hour of reperfusion had a 4.72-fold increase in C3 mRNA and a 19.5-fold increase in C9 mRNA. By contrast, C3 mRNA in hearts subjected to 3.5 hours of normoxic perfusion showed no change, and those subjected to 3.5 hours of ischemia showed only a 1.72-fold increase, whereas C9 mRNA levels increased by 5.17-fold after 3.5 hours of normoxic perfusion and 12.5-fold after 3.5 hours of ischemia. The results of this study demonstrate for the first time that heart tissue is capable of expressing genes and proteins of the complement system, although it is not yet known which cell types are responsible. They further demonstrate that ischemia and reperfusion of the heart promotes a rapid upregulation of the mRNAs encoding the complement proteins C3 and C9 and that these abnormal levels considerably exceed those of normal liver. These observations are consistent with the hypothesis that local production of complement proteins may contribute significantly to the degree of ischemic injury to the myocardium and that complement expression is augmented by reperfusion.