Mitochondria are cellular organelles bounded by two distinct membranes. They contain one-tenth of the cellular proteins. Their DNA replicates autonomously and is transmitted through the maternal germline independently of nuclear and chromosomal DNA [
6]. Each cell in the human body contains up to several hundred mitochondria. Each mitochondrion contains ≤10 copies of mtDNA. In addition to producing energy, mitochondria support and participate in several essential cellular functions, including: intermediary metabolism; ion homeostasis; synthesis of lipids, amino acids and nucleotides; active transport; and apoptosis [
7]. The aging process in humans has been shown to be associated with reduced levels of mtDNA transcripts and increased mtDNA content in the brain and lung [
8,
9]. There are undoubtedly tissue-specific effects of aging because the skeletal muscles and livers of rats showed an age-related decline in mtDNA copy number. Different from nuclear DNA, replication of mtDNA is error-prone and mammalian mtDNA contains no introns, has no protective histones, and is exposed to deleterious ROS generated by oxidative phosphorylation [
10]. These factors contribute to the accumulation of mutations in mtDNA at an approximately tenfold greater rate than in nuclear DNA [
11]. The mtDNA of tumor cells may not only suffer from the change of structure but also the change of number. Some studies have shown that copy number is increased in tumors. Wang et al. found that the average mtDNA copy number in pathological low-grade tumours was over two-fold higher than that in high-grade endometrial carcinomas, and the Change in mtDNA content was not related with patients’ age or tumour stages [
12]. The copy number of 1p36.33 and mtDNA peripheral blood mononuclear cells infected by the Epstein–Barr virus in patients with lymphocytic leukemia was tested in quantitative PCR by Jeon et al. They suggested that increased mitochondrial biogenesis is indicative of the progression of EBV-mediated B-cell transformation [
13]. However, some studies showed that the mtDNA copy number was decreased in tumors. Lee et al. found: a marked decrease in cellular mtDNA and ATP content, concomitant with a lack of mRNAs encoded by mtDNA. The mtDNA- depleted cells showed a decreased sensitivity and accumulation of anti-cancer drugs, suggesting that mtDNA depletion could develop multidrug resistance (MDR) phenotype in HCT-8 cells. The expression level of MDR1 mRNA and its translated product P-glycoprotein was increased in the mtDNA- depleted cells. The decline in mtDNA content may cause multidrug resistance, which may be raised by increasing the stability of MDR1 mRNA expression [
14]. Lee et al. examined three colon cancer cells and found that the decline in mtDNA copy number may be independent of tumor-cell homogeneity or heterogeneity of point mutations or large fragment deletion. In human tumors, the instability of the mitochondrial genome and the decline of mtDNA copy number may be independent factors [
15]. Decline in mtDNA copy number may be related to the reduction in the number of oxidative phosphorylation proteins. In various tumors, reduction of mtDNA copy number may be correlated with clinicopathological parameters and tumor invasiveness, For example, compared with normal liver tissues, the mtDNA copy number in hepatocellular carcinoma(HCC) was significantly reduced, and the results showed that it was related to tumor size and cirrhosis. It is evident that a low mtDNA copy number in HCC patients might help to identify a poor prognosis [
16]. Scientists could establish cell lines containing varying levels of mtDNA by treatment with low concentrations of ethidium bromide. Ethidium bromide can insert into mtDNA molecules and inhibit the activity of mtDNA polymerase gamma during mtDNA replication. Amuthan and colleagues demonstrated that an invasive phenotype is produced within the original cell line of the non-invasive mouse skeletal muscle C1C12 cell line and human lung cancer cell line A549 if mtDNA is absent, and that the overexpression of the tumor-specific markers cathepsin L and β-transforming growth factor can be detected. These studies suggest that the loss of mtDNA can promote the development and metastasis of tumors [
17]. The loss of mtDNA may affect the development of cancer and cancer metastasis by preventing apoptosis and promoting the generation of cancer-related proteins. Wu et al. suggest that somatic mtDNA mutations and mtDNA depletion occur in gastric cancer and that mtDNA depletion is involved in carcinogenesis and/or cancer progression of gastric carcinoma [
18]. Some evidences suggest that a loss in the integrity of the mitochondrial genome (sequence and/or copy number) can progress a cancer to an advanced phenotype by way of ROS dependent and independent events [
19].
Tumor growth requires an appreciable amount of energy, which is mainly from mitochondrial aerobic oxidation and glycolysis, which is required for excessive cell proliferation. Previously, it was believed that long-term hypoxia as well as glycolysis in the tumor played a leading part in the rapid growth of tumors. Recently, it was found that these two types of energy supply in tumor growth have equally important roles. Mitochondria are the only gene-containing structures outside the nucleus in eukaryotic cells. The rate of somatic mutation of mtDNA has been presumed to be 10–100-times greater than that of nuclear DNA. Reported sequence changes include point mutations, multiple deletions and microsatellite instability in coding and non-coding regions. However, little is known about rapid decreases in mtDNA, microsatellite instability and alteration of the copy number in human CRC.
We found that the mean copy number in CRC was significantly lower than the copy number in adjacent tissues. The copy numbers obtained from the present study were significantly lower than the copy number of myocardial and peripheral blood mononuclear cells reported from other studies [
5,
20]. We also found that the decrease in mtDNA copy number was significantly correlated with lymph-node metastasis but not with sex, age, pathological type, or TNM stage. We also found that the three-year tumor-free survival was lower among CRC patients with lower mtDNA copy numbers. This may be because the low copy numbers in tumors result in a stronger tolerance to hypoxia. Therefore, tumors can reduce the dependence of mitochondrial oxidative phosphorylation. Hence, the main source of energy for tumor progression is rendered to anaerobic metabolism in the glycolytic pathway. This is a favorable measure for tumor invasion, and which is a necessary condition for survival of the tumor under hypoxic conditions [
21]. Tumors with relatively high copy numbers of mtDNA have poor tolerance to hypoxia grow slowly, they are less invasive, and more sensitive to chemotherapy. Thus, patients with such tumors have a better prognosis [
22], Tumors that metastasize to lymph nodes are more likely to recur even after surgical removal. We found that the copy number of mtDNA was negatively correlated with lymph-node metastasis. Patients with a relatively high copy number of mtDNA had a higher three-year tumor-free survival than patients with a lower copy number of mtDNA. but the difference was not significant. The small sample size might limit the significance of our study. A future should focus on a functional analysis of the reduction of mtDNA in CRC.