Article, Emergency Medicine

Electrocardiographic changes after injury in a rat model of combined crush injury

a b s t r a c t

Background: Crush injury from debris, combined with hypoxia and water and food deprivation (combined crush injury), is common in industrial accidents and events such as earthquakes and terrorist attacks. Whether electrocardiographic changes are associated with combined crush injury is unclear.

Methods: Thirty-six rats underwent electrocardiography at baseline then were randomly assigned to 6 groups of 6. Bilateral hind limbs of all rats were compressed with custom-made clips (pressure 4.5 +- 0.3 kg), and the rats were put into a hypoxic compartment (oxygen concentration 10% +- 0.1%) for 72 hours without food or water. After 72 hours, the rats were moved to a normoxic environment, where the clips were removed (decompression) and food and water were freely accessible. Electrocardiography was performed in a different group at each of days 0, 1, 3, 7, 14, and 28 after decompression.

Results: One rat died at 0.6 days. Among the remaining 35 rats, 28 (80%) had abnormal electrocardiographic changes: ST-segment depression (n = 25), tall-peaked T waves (n = 16), arrhythmias (n = 4), abnormal Q waves (n = 2), wide QRS complexes (n = 2) and QT prolongation (n = 1). The abnormality rates among assessed rats were 100% on days 0, 1, and 3; 83% on day 7; and 50% on days 14 and 28.

Conclusions: The findings suggest that abnormal electrocardiographic changes were seen in rats after simulated combined crush injury and decompression and were slow to resolve.

(C) 2013

  1. Introduction

Crush injury due to burial underneath debris is a major cause of death and disability after bombings in the wars and terrorism and natural disasters such as earthquakes [1-4]. Individuals might remain buried for a long time in small spaces with reduced oxygen content. At the same time, water and food supplies might be depleted or nonexistent.

Previous studies were limited to simply crush injury. The effects of combined crush injury (crush injury plus hypoxia, water and food deprivation) on the heart have not been reported.

We used a rat model of combined crush injury to study electrocardiographic and microscopic cardiac changes after de- compression. Our study will offer laboratory data for combined crush injury.

? Disclosures: None.

?? Contributorship statement: Dewen Wang designed the experiment. Xiaoming Guo was responsible for Animal experiments, collecting electrocardiography, writing the article, and the statistics. Zhirui Liu was responsible for animal experiments and collection of specimens.

* Corresponding author.

E-mail addresses: [email protected] (X. Guo), [email protected] (D. Wang), [email protected] (Z. Liu).

  1. Materials and methods
    1. Animals

We obtained 36 healthy male Wister rats that weighed 220 to 260 g from the Animal Center of Academy of Military Medical Sciences, Beijing, China. All animal experiments were conducted according to the guidelines for animal use of the Academy of Military Medical Sciences.

Model of combined crush injury

After baseline electrocardiography, the rats were randomly assigned to 6 groups of 6, which were kept in six separate cages. Combined crush injury was simulated in all rats by compression of bilateral hind limbs with custom-made clips which were made by our laboratory (pressure 4.5 +- 0.3 kg) for 72 hours, during which they were housed in a hypoxic compartment (oxygen concentration 10% +- 0.1%) with no food or water. After 72 hours, the rats were moved to a normoxic environment, where the clips were removed. The rats were then kept under standard conditions with free access to food and water.

Electrocardiography

A different group of rats was assessed with electrocardiography at each of the following time-points: days 0, 1, 3, 7, 14, and 28 after

0735-6757/$ – see front matter (C) 2013 http://dx.doi.org/10.1016/j.ajem.2013.08.054

Fig. 1. Normal 6-lead electrocardiography at baseline.

Fig. 2. Tall-peaked T waves with narrow bases were observed immediately after decompression.

decompression. Before electrocardiography, all rats were anesthe- tized by intraperitoneal injection of pentobarbital sodium (50 mg/kg body weight). Four limb electrodes were placed for leads I, II, III, aVR, aVL, and aVF, and the Electrocardiographic features such as ST segment, Corrected QT interval, QRS duration, and T-wave changes were assessed.

Myocardial tissue light microscopy and electron microscopy

In each experimental group, myocardial tissues were subjected to light microscopy. For light microscopy, myocardial tissues were fixed in 10% formalin and embedded in paraffin wax. Sections were cut and stained with hematoxylin and eosin and were then carefully examined microscopically.

For electron microscopy, myocardial tissues of days 0 and 7 were fixed in 3% glutaraldehyde; ultrathin sections were cut and stained with uranyl acetate solution and lead citrate solution; transmission electron microscopy of myocardial ultrastructure was done, acceler- ating voltage to 80 kV (magnification x6000-20000).

  1. Results
    1. Death data

One rat died 0.6 days after decompression. Autopsy showed no macroscopic signs of infection of internal organs, bleeding, or other fatal diseases. The heart was soft and enlarged and the apex was blunt. The right and left ventricles were apparently expanded.

Electrocardiographic features

Electrocardiographic features at baseline were normal (Fig. 1). Overall, 28 (80%) of 35 rats had obvious electrocardiographic changes after decompression: ST-segment depression (n = 25), tall-peaked T waves with narrow bases (n = 16), arrhythmias (n = 4), abnormal Q waves (n = 2), QT prolongation (n = 2) and wide QRS complexes

(n = 1; Table, Figs. 2, 3, 4, and 5). In the early period after

decompression (days 0, 1, and 3), abnormal electrocardiographic changes were seen in 100% of rats assessed; on day 7, the rate was 83.3% (ST-segment depression and T-wave changes only), and on days

14 and 28, it was 50% (ST-segment depression only; Table). ST- segment depression was seen mostly in leads II, III, and aVF.

Myocardial tissue microscopy

Disorder and fracture of myocardial fibers was observed (Fig. 6A and B). Myocardial mitochondria structure was greatly altered. Mitochondria were swollen, even ruptured; cristae were disorganized or had disappeared (Fig. 6C and D). Perinuclear cisterna was widened (Fig. 6E).

  1. Discussion

Crush injury is typically severe and has a poor prognosis. Large numbers of people suffered from Crush injuries after massive earthquakes in Wenchuan and Yushu counties, China [1,3]. Some of the wounded were alert and gave appropriate verbal responses before being rescued but, in a short time (minutes to hours) after being removed from the debris, died suddenly. Some scholars thought that the sudden death should be due to acute renal failure caused by crush injury [2,4,5], but acute renal failure usually has a process of development; it is difficult to accept this cause as the only explanation. Thus, some sudden deaths might have had cardiogenic causes.

In this experiment, we found that the simulated combined crush injury was associated with multiple obvious electrocardiographic abnormalities. The most frequent abnormality was multi-lead ST- segment depression (71.4%) and indicates myocardial ischemia, anoxia, and damage. ST-segment depression appeared mostly in leads II, III, and aVF and, secondly, in leads I and aVL, therefore, myocardial ischemia and anoxia occurred mainly in the inferior and lateral walls of the heart.

Crush injury can cause muscle intracellular products such as potassium (K+) enters the circulation, resulting in hyperkalemia [6].

Table

Electrocardiographic features before and after crush injury and decompression

Observed samples

Abnormal samples

ST-segment depression

T-wave changes

Arrhythmia

Abnormal Q waves

QT prolongation

Wide QRS complexes

Before crush injury After decompression 0 h

36

6

6/6

5

5

2

2

2

1

1 d

6

5/5

4

4

1

3 d

6

6/6

5

5

1

7 d

6

5/6

5

2

14 d

6

3/6

3

28 d

6

3/6

3

Total

36

28/35

25

16

4

2

2

1

Fig. 3. ventricular premature beats were observed immediately after decompression.

Tall-peaked T waves with narrow bases were seen in 45.7% of rats and were consistent with hyperkalemia electrocardiographic features.

Arrhythmias affected more than 10% of rats. Second degree atrioventricular block indicated that the crush injury simulated deep buried condition that can damage the heart Conduction system. Wide QRS complexes [7] and QT prolongation are important predictors of sudden cardiac death [8-21]. The types of changes were multiple and diverse and recovery was slow, as half of rats were still affected at day 28. This study suggests that for deep buried wounds suffered from crush injury, treatment should not be only for Renal damage; heart damage should also be seriously concerned. For the evidence of heart

damage to be present, the Symptomatic treatment should be given.

Some mechanisms might explain the electrocardiographic changes caused by combined crush injury. Firstly, continuous prolonged external pressure applied to the body results in massive Skeletal muscle tissue destruction or traumatic rhabdomyolysis. Then muscle cell calcium overload, Ischemia-reperfusion injury, massive release of oxygen radicals, lipid peroxidation and an enhanced PMN-endothelium interaction will happen, which can influence or even hurt the heart. Secondly, muscle intracellular products such as potassium (K+) enter the circulation [22,23]. Hyperkalemia can cause malignant ventricular arrhythmia and sudden cardiac death. Thirdly, under the deep buried condition, sustained Hypoxic conditions, and no food and water will induce heart anoxia and ischemia. Fear and hypovolemia can activate

Fig. 4. Second-degree atrioventricular block was observed immediately after decompression.

Fig. 5. Atrial premature beats were observed 1 day after decompression.

the sympathetic nervous system, antidiuretic hormone, and the renin- angiotensin system which will hurt cardiovascular system.

Strengths and limitations

This study is unique. To our knowledge, it is the first study of electrocardiographic changes after injury in a rat model of combined crush injury.

The study is likely to have a number of limitations. If we use big animals who have much richer leg muscles, the experimental results may be even perfect. We just recorded a short time (about 1 minute)

of electrocardiographic changes. If we observe for a longer time, more abnormal electrocardiographic changes may be found.

  1. Conclusions

This study demonstrates that the heart is a sensitive target organ in cases of combined crush injury. More attention should be paid to possible Cardiac damage in addition to renal injury. Further study is necessary for electrocardiographic abnormalities in the recovery stage.

A

B

C

D

E

Fig. 6. Myocardial tissue instantly after depression. A, Magnification x200; hematoxylin-eosin staining. B, Magnification x400; hematoxylin-eosin staining. C, Magnification x8000; electron microscope. (D) Magnification x12000; electron microscope. E, Magnification x12000; electron microscope.

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