Structural and biochemical evidence of mitochondrial depletion in pigs with hypertrophic cardiomyopathy

https://doi.org/10.1016/S0034-5288(02)00189-3Get rights and content

Abstract

Pig hearts with naturally occurring hypertrophic cardiomyopathy (HCM) were isolated to investigate the effects of mitochondrial deficiency at biochemical and molecular levels. Enzyme activities of mitochondrial-encoded cytochrome c oxidase and NADH dehydrogenase in the HCM hearts (n=12) were lower than that in the controls (n=12) by 41±29% (P<0.01) and 43±21% (P<0.001), respectively. Additionally, Southern blot analysis was conducted to quantify the relative amount of mitochondrial DNA (mtDNA) from the HCM and controls. The relative amount of mtDNA in the HCM hearts was significantly 57±19% (P<0.001) lower than that in the controls. Both mitochondrial enzyme deficiency and mtDNA depletion were significantly correlated with the degree of cardiac hypertrophy judged based on the ratio of heart/body weight. In conclusion, our results reveal that a secondary effect of tissue-specific mtDNA depletion and mitochondrial dysfunction is in response to the HCM.

Introduction

Mitochondria are the major source of cellular ATP providing the energy for cardiac function by mitochondrial oxidative phosphorylation (OXPHOS). The cellular organelle contributes to myocyte injury via loss of physiologic function. Therefore, the physiologic significance of damage to mitochondrial DNA (mtDNA), proteins, or lipids should be established at the level of the mitochondrion, myocyte, and whole heart (Lesnefsky et al., 2001). Morphologic abnormalities of mitochondria and mitochondrial dysfunction occur in association with heart failure (Sharov et al., 2000), as well as in cardiomyocytes of ischemic, dilated, and hypertrophied hearts (Jarreta et al., 2000; Marin-Garcia et al., 1997, Marin-Garcia et al., 1998).

Hypertrophic cardiomyopathy (HCM) is a cardiac disease in which the most characteristic morphologic features are hypertrophied, nondilated ventricles, and myofibrillar disarray with increased amounts of matrix (Maron, 1997). This disease affects approximately 1 in every 500 of the general population (Spirito et al., 1997). Notably, over 50% of all reported cases of HCM are inherited, presenting disease-causing mutations in several genes encoding sarcomeric protein (Burch and Blair, 1999; McKenna et al., 1998) or in mtDNA (Marin-Garcia and Goldenthal, 1997). Although in some cases a correlation between genetic defects and decreased specific mitochondrial enzymatic activities has been noted, few investigations have focused on the characterization of respiratory chain function and the findings have often been contradictory (Marin-Garcia et al., 1997, Marin-Garcia et al., 1998; Ozawa et al., 1990). The lack of work on the area could be due to the limitations of a lack of available heart specimens in humans. Furthermore, the question of whether alterations in genetically mtDNA molecule and mitochondrially enzymatic defects play a primary or secondary role in HCM remains essentially unanswered.

Recently, the authors developed a pig model for naturally occurring HCM (Huang et al., 1996; Liu et al., 1994). The morphologic and pathologic features of pig HCM hearts comprise hypertrophic ventricles with cellular disorientation, fibrosis, and occluded intramural coronary arteries (Chiu et al., 1999; Dai et al., 1997; Liu et al., 1994). These characteristics all resemble those found in humans (Maron, 1997). The novel pig model also found mitochondrial damage in HCM hearts, which we hypothesized that oxidative stress caused the damage (Lin et al., 1997). Clinically, mitochondrial dysfunction associated with human cardiac disease has been evaluated at the biochemical and molecular levels (Anan et al., 1995; Marin-Garcia and Goldenthal, 1997). Mitochondrial regulation and deficiencies associated with HCM disease therefore can be profitably explored.

This study reports on pigs with HCM and abnormal activity levels of mitochondrial respiratory enzymes in the cardiomyocytes. The data also revealed a significant reduction in myocardial mtDNA compared with the controls. Therefore, mtDNA depletion may play a role in the pathogenesis of HCM.

Section snippets

Animal source, care, feeding, and management

Landrace pigs raised in the Nuclear Breeding Center of Northern Taiwan were bred specifically for the study of HCM (Huang et al., 1996). The pigs were fed 17.5% crude protein and 3050 kcal of metabolizable energy per kg. Feed and water were administered ad libitum. All pigs were reared in a natural lighting environment. The animals were treated according to guidelines established by the National Science Council of the Republic of China (National Science Council, 1993). The HCM pigs were

Characterization of HCM

Table 1 lists the physical characteristics of the 12 HCM pigs and the 12 unaffected pigs (normal controls). The control and HCM groups displayed no significant differences in either age or body weight, but the heart weight and heart/body weight ratio of the HCM pigs significantly exceeded those of the control group (P<0.001). Additionally, the thickness of the ventricular free walls in the HCM pigs significantly surpassed those of the control pigs (P<0.01). The physical characteristics of the

Discussion

Mitochondrial abnormalities at the biochemical and molecular level are increasingly recognized as playing a role in heart disease (Anan et al., 1995; Marin-Garcia and Goldenthal, 1997). Respiratory enzyme deficiencies and defects in mtDNA, including point mutations (Merante et al., 1994; Santorelli et al., 1996), deletions (Ozawa et al., 1990), and depletion (Marin-Garcia et al., 1997, Marin-Garcia et al., 1998), have been frequently associated with human HCM. However, the correlation between

Acknowledgements

We thank Dr. K.S. Dai, Y.T. Chiu, W.C. Lee, and S.Y. Huang at the Animal Technology Institute Taiwan for their collaboration. Our special thanks to Dr. Si-Kwang Liu (Animal Medical Center, New York, USA) for his encouragement and critical comments. This work was supported by Grants NSC89-2321-B-059-008-A20 and NSC89-2321-B-059-023-A20 from the National Science Council, Taiwan, Republic of China.

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