Experimental blunt chest trauma-induced myocardial inflammation and alteration of gap-junction protein connexin 43

Abstract

Objective: Severe blunt chest trauma in humans is associated with high mortality rates. Whereas lung tissue damage and lung inflammation after blunt chest trauma have extensively been investigated, the traumatic and posttraumatic effects on the heart remain poorly understood. Therefore, the purpose of this study was to define cardiac injury patterns in an experimental blunt chest trauma model in rats.

Methods: Experimental blunt chest trauma was induced by a blast wave in rats, with subsequent analysis of its effects on the heart. The animals were subjected either to a sham or trauma procedure. Systemic markers for cardiac injury were determined after 24 h and 5 days. Postmortem analysis of heart tissue addressed structural injury and inflammation 24 h and 5 days after trauma.

Results: Plasma levels of extracellular histones were elevated 24 h and 5 days after blunt chest trauma compared to sham-treated animals. In the heart, up-regulation of interleukin-1β 24 h after trauma and increased myeloperoxidase activity 24 h and 5 days after trauma were accompanied by reduced complement C5a receptor-1 expression 24 h after trauma. Histological analysis revealed extravasation of erythrocytes and immunohistochemical analysis alteration of the pattern of the gap-junction protein connexin 43. Furthermore, a slight reduction of α-actinin and desmin expression in cardiac tissue was found after trauma together with a minor increase in sarcoplasmatic/endoplasmatic reticlulum calcium-ATPase (SERCA) expression.

Conclusions: The clinically highly relevant rat model of blast wave-induced blunt chest trauma is associated with cardiac inflammation and structural alterations in cardiac tissue.

Fig 1. Systemic and local effects of blunt chest trauma.

Apparence of extracellular histones in plasma 24 h and 5 d after blunt chest trauma or sham, * differences to sham procedure were significant, p<0.05; n = 4 for each bar. B. Representative H.E staining of left ventricles 24 h or 5 d after blunt chest trauma or sham procedure as indicated.

Fig 2. Local inflammation in cardiac injury after blunt chest trauma.

A. Increased myeloperoxidase (MOP) activity in left ventricular cardiac tissue 24 h and 5 d after blunt chest trauma compared to sham procedure. B. Elevation of proinflammatory cardiodepressive cytokine IL-1β expression in left ventricular homogenates 24 h after blunt chest trauma compared to sham procedure as assessed by qRT-PCR. C. Representative western blot for C5aR1 of left ventricular tissue homogenates. Densitometry revealed diminished C5aR1 protein expression in left ventricular homogenates 24 h after blunt chest trauma compared to sham procedure. D. C5aR expression in left ventricular homogenates 24 h and 5 d after blunt chest trauma and after sham procedure as assessed by qRT-PCR. p<0.05; *differences were significant to sham procedure, For all frames n = 8 for each bar.

Fig 3. Structural alterations in the heart after blunt chest trauma.

Alteration of gap junctional protein connexin 43 (Cx43) after blunt chest trauma in the heart. A. Representative distribution of Cx43 in cardiac tissue after sham procedure or 24 h or 5d after blunt chest trauma as indicated. B. Representative western blot for Cx43 of left ventricular tissue homogenates. Densitometry revealed no significant increase in protein expression in left ventricular homogenates 24 h and 5 d after blunt chest trauma compared to sham procedure. For all frames n = 4 for each bar.

Fig 4. Apoptosis in the heart after blunt chest trauma.

A. Representative caspase 3 staining in cardiac tissue after sham procedure or 24 h and 5d after blunt chest trauma as indicated. B. Changes in pixel density of caspase 3 staining in left ventricular tissue 24h or 5d after blunt chest trauma compared to sham procedure as assessed by immunohistochemistry. C. Trauma induced cardiomyocyte apoptosis was determined by TUNEL (TdT-mediated dUTP-biotin nick end labeling) staining in left ventricular tissue 24h and 5d after blunt chest trauma compared to sham procedure as assessed by fluorescence mircoscopy. The total number of nuclei in cardiomyocytes were determined by DAPI staining and apoptotic nuclei were determined by positive TUNEL staining. The apoptotic index was calculated (number of positively stained/total number of nuclei). For all frames n = 4 for each bar. * p<0.05.