Korean Circ J.  2007 Sep;37(9):399-407. 10.4070/kcj.2007.37.9.399.

Arrhythmogenic Gene Change and Nerve Sprouting after Acute Myocardial Infarction in Mice

• Oh YS
• Kim JH
• Choi YS
• Park HW
• Lee DH
• Shin WS
• Youn HJ
• Lee MY
• Chung WS
• Rho TH
• Kim JH
• Chen PS
• Choi KB

• KMID: 2093975
• DOI: http://doi.org/10.4070/kcj.2007.37.9.399

Abstract

BACKGROUND AND OBJECTIVES: Myocardial infarction (MI) elicits nerve sprouting. However, the time course and spatial distribution of this nerve sprouting and its relationship to the expression of neurotrophic factors is unclear. The aim of this study was to identify the association of nerve sprouting with the expression of neurotrophic factors.
MATERIALS AND METHODS
We induced MI in FVB mice by ligating the left coronary artery. The hearts were removed at 3 hours to 13 months after MI for growth associated protein 43 (GAP-43) immunostaining. The nerve density (micrometer2/mm2) was determined by ImagePro software. In another group of mice, their myocardial tissues were processed and analyzed with using an Affymetrix RG U74V2 array.
RESULTS
The density of the nerve fibers that were immunopositive for GAP-43 was the highest 3 hours after MI in both the peri-infarct areas and the remote areas. The outer loop of the ventricle had a higher nerve density than that in the inner loop of the ventricle. The differences were at a peak 3 hours after MI, but they persisted for 2 months afterwards. The expressions of nerve growth factor, insulin-like growth factor, leukemia inhibitory factor, transforming growth factor-beta3 and interleukin-1alpha were increased for up to 2 months after MI as compared to the normal control. qRT PCR analyses showed increased mRNA for tyrosine hydroxylase, synaptophysin, nerve growth factor and leukemia inhibiting factor in the peri-infarct areas for up to 2 months after MI, but this occurred only for roughly 3 days after MI in the remote areas.
CONCLUSION
We conclude that MI resulted in immediate upregulation of nerve growth factor, insulin-like growth factor, leukemia inhibitory factor, transforming growth factor-beta3 and interleukin-1alpha in the peri-infarct areas and this all occurred to a lesser extent in the remote areas. These changes persisted for at least 2 months, and they were associated with increased nerve sprouting activity, which was most active in the outer loop of the heart.

Keyword

Electrophysiology; Ventricular remodeling; Regeneration; DNA

MeSH Terms

Animals
Coronary Vessels
DNA
Electrophysiology
GAP-43 Protein
Heart
Interleukin-1alpha
Leukemia
Leukemia Inhibitory Factor
Mice*
Myocardial Infarction*
Nerve Fibers
Nerve Growth Factor
Nerve Growth Factors
Polymerase Chain Reaction
Regeneration
RNA, Messenger
Synaptophysin
Tyrosine 3-Monooxygenase
Up-Regulation
Ventricular Remodeling
DNA
GAP-43 Protein
Interleukin-1alpha
Leukemia Inhibitory Factor
Nerve Growth Factor
Nerve Growth Factors
RNA, Messenger
Synaptophysin
Tyrosine 3-Monooxygenase

Figure

• 

  Fig. 1 Mouse MI. A shows the surface ECG (lead II) at baseline and after left coronary ligation during the first surgery. The arrows point to the ST segment elevation after ligation, indicating myocardial ischemia. Also note the accelerated heart rate after coronary artery ligation. B shows a trichrome stained cross section of the left ventricle. The fibrous tissues stained blue and the myocardium stained red. A dashed line was drawn half way through the tissue to separate the peri-infarct region from the remote region. MI: myocardial infarction.

• 

  Fig. 2 Nerve sprouting within 3 hours after MI. A: trichrome staining showing the suture tracts (green arrows), and the dark red area that's compatible with acute myocardial necrosis. B and C show GAP-43 immunostaining in the peri-infarct region. D shows the same staining in the remote region. The brown twigs in B, C and D are immunopositive nerves. Note that the peri-infarct sites had a higher density of GAP-43 positive nerves than the remote site. The magnification of the objective is 40X. The horizontal bar at the right lower portion of each panel is 0.1 mm in length.

• 

  Fig. 3 Densities of the GAP-43 immunopositive nerves and the time after MI. Each column represents 1 mouse. The nerve count was the highest within 3 hours after MI, and then it progressively decreased to normal. A: peri-infarct area. B: remote area. NL: normal, hrs: hours, d: day, wk: week, mo: month.

• 

  Fig. 4 Nerve densities in the inner and outer loops in a normal mouse. A: Cross section of a normal mouse ventricle. The inner and outer loops of the left ventricle can be identified based on their location and fiber orientation. The fiber orientation in the inner loop is roughly perpendicular while to the outer loop is parallel to the circumference of the ventricles. The nerve densities were determined by analyzing 3 pairs of selected fields in the inner and outer loops as shown in Panel A. Panels B and C shows GAP-43 immunopositive nerves in the inner and outer loops, respectively. The arrows point to brown nerve twigs. The magnification of the objective is 20X. The horizontal bar is 1 mm in length.

• 

  Fig. 5 Nerve densities of the inner and outer loops in normal mice and MI mice. Each column represents 1 mouse. In the outer loop, the nerve count increased within the first 3 hours after MI. The nerve count then progressively decreased to the normal level. In the inner loop, however, there was only an insignificant rise of nerve densities throughout the post-MI period. A: peri-infarct area. B: remote area. NL: normal, hrs: hours. d: day, wk: week, mo: month, MI: myocardial infarction.

• 

  Fig. 6 The mRNA expression as a function of time. We used qRT PCR to determine the mRNA expression at baseline and during each time point after MI. We then divided the latter by the former, resulting in a ratio of the mRNA to that of the control (ordinate). A ratio of >1.5 is considered a significant increase and it is marked by an asterisk. A and B show the mRNA expression in the peri-infarct area and in the remote area, respectively. MI: myocardial infarction, TH: tyrosine hydroxylase, SYN: synaptophysin, NGF: nerve growth factor, LIF: leukemia inhibitory factor.

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