Trading mtDNA uncovers its role in metastasis

It has been controversial for many years of whether mtDNA mutations are involved in phenotypes related to cancer due to the difficulty in excluding possible involvement of nuclear DNA mutations in these phenotypes. We addressed this issue by complete trading of mtDNAs between tumor cells expressing different metastatic phenotypes. Resultant trans-mitochondrial cybrids share the same nuclear background, but possess mtDNA from tumor cells expressing different metastatic phenotypes, and thus can be used to uncover the role of mtDNA in these phenotypes. The results showed that mtDNA controls development of metastasis in tumor cells, while tumor development is controlled by nuclear genome.Key words: pathogenic mtDNA mutations, respiration defects, enhanced glycolysis, ROS overproduction, rho-zero cells, mtDNA transfer technology, metastasisHuman mtDNAs with pathogenic mutations inducing significant respiration defects have been shown to be closely associated with mitochondrial diseases.^1,2 Although mitochondrial respiratory function is controlled by both nuclear and mitochondrial genomes, the pathogenicity of these mtDNA mutations has been proven by co-transmission of the mutant mtDNAs and mitochondrial respiration defects to mtDNA-less (ρ⁰) human cells: the resultant trans-mitochondrial cybrids sharing the same nuclear backgrouond showed respiration defects, only when they accumulated the mutated mtDNA from the patients.^3–6 Moreover, we generated transmitochondrial mito-mice sharing the same nuclear background, but carrying various proportions of mtDNA with a pathogenic mutation, and provided model systems for studying exactly how mtDNAs with pathogenic mutations are transmitted and distributed in tissues resulting in the pathogenesis of mitochondrial diseases that show various clinical phenotypes.^7–9With respect to the involvement of mtDNA in tumor phenotypes, it has been proposed that most chemical carcinogens bind preferentially to mtDNA rather than to nuclear DNA in mammalian cells,^10–12 and thus, mtDNA should be the major cellular target of chemical carcinogens, and resultant creation of mutations in mtDNAs is responsible for expression of tumor phenotypes.¹²Although, there has been no direct evidence for creation of mtDNA mutations by chemical carcinogens, and for their contribution to tumor development in mammalian cells, recent studies showed that somatic mtDNA mutations accumulated in human colorectal tumors¹³ and in various tumor types¹⁴ rather than in normal cells of the same subjects, probably by the clonal expansion of the mutated mtDNAs along with the repeated division of tumor cells. Many subsequent studies supported preferential accumulation of mutated mtDNAs in tumor cells,^15–18 suggesting that mutated mtDNAs in tumor cells have acquired replication advantages to be homoplasmic. However, these studies did not address the fundamental question of whether the mutated mtDNAs are involved in tumor development.Our previous studies directly addressed this issue using transmitochondrial cybrids obtained by mtDNA trading between normal and tumor cells, and provided convincing evidence that mutations in nuclear DNA, but not in mtDNA were involved in tumor development in the mouse^19,20 and in human cultured cells.^21,22 The possibility that these observations may represent some specific tumor cases can be excluded since there has been no statistical evidence for association of tumor development and pathogenic mtDNA mutations in the patients with mitochondrial diseases expressing respiration defects caused by pathogenic mutations in mtDNA. The possibility that some polymorphic mtDNA mutations that do not induce respiration defects, but somehow contribute to tumor development also can be excluded, because there has been no statistical evidence for the presence of maternal inheritance of tumor development in spite of the strictly maternal inheritance of mammalian mtDNA.^23,24Nonetheless, it was still possible that mtDNA mutations are involved in other processes than oncogenic transformation of normal cells to develop tumors, such as in malignant progression of tumor cells to develop a metastatic potential. Recent studies demonstrated that mitochondrial respiration defects in TCA-cycle enzymes caused by nuclear DNA mutations controls tumor phenotypes as a consequence of induction of a pseudo-hypoxic pathway under normoxia.^25–27 Thus, some mtDNA mutations also induce the pseudo-hypoxic pathway under normoxia by inducing mitochondrial respiration defects. However, there has been no direct evidence for involvement of mtDNA mutations in malignant progression or in the regulation of the pseudo-hypoxic pathway under normoxia, because of the difficulty in excluding possible contribution of nuclear DNA mutations in these processes.²⁸Recently, we addressed this issue using trans-mitochondrial cybrids²⁹ obtained by complete trading of mtDNAs between highly and poorly metastatic mouse lung carcinoma cells (Fig. 1). By this approach, we could provide convincing evidence for the control of malignant progression of tumor cells to develop metastatic potential by mtDNA:²⁹ all the trans-mitochondrial cybrids with mtDNA from highly metastatic tumor cells expressed high metastatic potential, while those with mtDNA from poorly metastatic tumor cells expressed low metastatic potential, irrespective of whether their nuclear genome was derived from highly or poorly metastatic tumor cells. The findings in our study²⁹ can be summarized as follows: (1) A missense G13997A mutation in the ND6 gene of mtDNA from highly metastatic lung tumor cells induces a complex I defect, and reversibly controls malignant progression of tumor cells to develop metastatic potential, but does not control oncogenic transformation of normal cells to develop tumors; (2) The complex I defect simultaneously induces enhanced glycolysis and ROS overproduction, but induction of metastasis is due to ROS overproduction; (3) ROS overproduction induces metastasis not by acceleration of genetic instability as usually proposed, but by reversible upregulation of nuclear-coded genes related to metastasis, such as Mcl-1; (4) ROS scavengers are therapeutically effective in suppressing mtDNA-mediated metastasis.Open in a separate windowFigure 1Scheme for the isolation of the trans-mitochondrial cybrids with completely exchanged mtDNA between parental cells expressing different metastatic phenotypes. Trading mtDNA shown in this scheme uncovered a role of mtDNA in metastasis. For trading mtDNA, parental P29 and A11 cells were treated with ditercalinium, an antitumor bis-intercalating agent, to isolate ρ⁰P29 and ρ⁰A11 cells (*), which have no mtDNA. Complete depletion of mtDNA was confirmed by PCR analysis. Enucleated cells of the mtDNA donors were prepared by their pretreatment with cytochalasin B and centrifugation. Resultant cytoplasts were fused with ρ⁰ cells by polyethylene glycol to obtain trans-mitochondrial cybrids. High metastatic potential is transferred to the P29mtA11 cybrids with the transfer of mtDNA from the A11 cells, and poor metastatic potential is transferred to the A11mtP29 cybrids with the transfer of mtDNA from the P29 cells. Involvement of cytoplasmic factors other than mtDNA from the A11 cells in expression of the high metastatic potential in the P29mtA11 cybrids can be ruled out by the observations that the A11mtP29 cybrids lost their high metastatic potential, even though they always contain cytoplasmic factors transcribed by the nuclear genes derived from the A11 cells.Thus, our study partly resolves the controversial issue on the relevance or irrelevance of mtDNA mutations in tumor development and/or tumor phenotypes by showing that mutations in mtDNA control development of metastasis in tumor cells.²⁹ Considering that complex I defects simultaneously induce enhanced glycolysis under normoxia (the Warburg effect) and ROS overproduction,²⁹ it remains possible that the Warburg effect alone can control metastasis independently from ROS overproduction. More recently, we examined this possibility by generating trans-mitochondrial cybrids with the deletion mutant mtDNA, which can be expected to induce overall respiration defects, and express enhanced glycolysis under normoxia, but not express ROS overproduction. The results showed that the Warburg effect alone did not control metastasis.     

Reversible regulation of metastasis by ROS-generating mtDNA mutations

It has been controversial whether mtDNA mutations are responsible for oncogenic transformation (normal cells to develop tumors), and for malignant progression (tumor cells to develop metastases). To clarify this issue, we created trans-mitochondrial cybrids with mtDNA exchanged between mouse tumor cells that express different metastatic phenotypes. The G13997A mutation in the ND6 gene of mtDNA from high metastatic tumor cells reversibly controlled development of metastases by overproduction of reactive oxygen species (ROS), but did not control development of tumors. The mtDNA-mediated reversible control of metastasis reveals a novel function of mtDNA, and suggests that ROS scavengers may be therapeutically effective in suppressing metastasis.     

Mitochondrial DNA mutations regulate metastasis of human breast cancer cells

Mutations in mitochondrial DNA (mtDNA) might contribute to expression of the tumor phenotypes, such as metastatic potential, as well as to aging phenotypes and to clinical phenotypes of mitochondrial diseases by induction of mitochondrial respiration defects and the resultant overproduction of reactive oxygen species (ROS). To test whether mtDNA mutations mediate metastatic pathways in highly metastatic human tumor cells, we used human breast carcinoma MDA-MB-231 cells, which simultaneously expressed a highly metastatic potential, mitochondrial respiration defects, and ROS overproduction. Since mitochondrial respiratory function is controlled by both mtDNA and nuclear DNA, it is possible that nuclear DNA mutations contribute to the mitochondrial respiration defects and the highly metastatic potential found in MDA-MB-231 cells. To examine this possibility, we carried out mtDNA replacement of MDA-MB-231 cells by normal human mtDNA. For the complete mtDNA replacement, first we isolated mtDNA-less (ρ(0)) MDA-MB-231 cells, and then introduced normal human mtDNA into the ρ(0) MDA-MB-231 cells, and isolated trans-mitochondrial cells (cybrids) carrying nuclear DNA from MDA-MB-231 cells and mtDNA from a normal subject. The normal mtDNA transfer simultaneously induced restoration of mitochondrial respiratory function and suppression of the highly metastatic potential expressed in MDA-MB-231 cells, but did not suppress ROS overproduction. These observations suggest that mitochondrial respiration defects observed in MDA-MB-231 cells are caused by mutations in mtDNA but not in nuclear DNA, and are responsible for expression of the high metastatic potential without using ROS-mediated pathways. Thus, human tumor cells possess an mtDNA-mediated metastatic pathway that is required for expression of the highly metastatic potential in the absence of ROS production.     

"ROS-generating mitochondrial DNA mutations can regulate tumor cell metastasis"--a critical commentary

In a recent publication (K. Ishikawa et al., 2008, Science320, 661-664), the authors described how replacing the endogenous mitochondrial DNA (mtDNA) in a weakly metastatic mouse tumor cell line with mtDNA from a highly metastatic cell line enhanced tumor progression through enhanced production of reactive oxygen species (ROS). The authors attributed the transformation from a low-metastatic cell line to a high-metastatic phenotype to overproduction of ROS (hydrogen peroxide and superoxide) caused by a dysfunction in mitochondrial complex I protein encoded by mtDNA transferred from the highly metastatic tumor cell line. In this critical evaluation, using the paper by Ishikawa et al. as an example, we bring to the attention of researchers in the free radical field how the failure to appreciate the complexities of dye chemistry could potentially lead to pitfalls, misinterpretations, and erroneous conclusions concerning ROS involvement. Herein we make a case that the authors have failed to show evidence for formation of superoxide and hydrogen peroxide, presumed to be generated from complex I deficiency associated with mtDNA mutations in metastatic cells.     

Mitochondrial ND5 gene variation associated with encephalomyopathy and mitochondrial ATP consumption

Mitochondrial encephalomyopathy and lactic acidosis with strokelike episodes (MELAS) is a severe young onset stroke disorder without effective treatment. We have identified a MELAS patient harboring a 13528A-->G mitochondrial DNA (mtDNA) mutation in the Complex I ND5 gene. This mutation was homoplasmic in mtDNA from patient muscle and nearly homoplasmic (99.9%) in blood. Fibroblasts from the patient exhibited decreased mitochondrial membrane potential (Deltapsim) and increased lactate production, consistent with impaired mitochondrial function. Transfer of patient mtDNA to a new nuclear background using transmitochondrial cybrid fusions confirmed the pathogenicity of the 13528A-->G mutation; Complex I-linked respiration and Deltapsim were both significantly reduced in patient mtDNA cybrids compared with controls. Inhibition of the adenine nucleotide translocase or the F1F0-ATPase with bongkrekic acid or oligomycin caused a loss of potential in patient mtDNA cybrid mitochondria, indicating a requirement for glycolytically generated ATP to maintain Deltapsim. This was confirmed by inhibition of glycolysis with 2-deoxy-D-glucose, which caused depletion of ATP and mitochondrial depolarization in patient mtDNA cybrids. These data suggest that in response to impaired respiration due to the mtDNA mutation, mitochondria consume ATP to maintain Deltapsim, representing a potential pathophysiological mechanism in human mitochondrial disease.     

Plasma membrane oxidoreductase activity in cultured cells in relation to mitochondrial function and oxidative stress

Dichlorophenol indophenol (DCIP) reduction by intracellualr pyridine nucleotides was investigated in two different lines of cultured cells characterized by enhanced production of reacive oxygen species (ROS) with respect to suitable controls. The first line denominated XTC-UC1 was derived from a metastasis of an oxyphilic thyroid tumor characterized by mitochondrial hyperplasia and compared with a line (B-CPAP) derived from a papillary thyroid carcinoma with normal mitochondrial mass. The second line (170 MN) was a cybrid line derived from rho0 cells from an osteosarcoma line (143B) fused with platelets from a patient with a nucleotide 9957 mutation in mitochondrial DNA (encoding for cytochrome c oxidase subunit III) in comparison with the parent 143B line. The experimental lines had no major decreases of electron transfer activities with respect to the controls; both of them, however, exhibited an increased peroxide production. The XTC-UC1 cell line exhibited enhanced activity with respect to control of dicoumarol-sensitive DCIP reduction, identified with membrane bound DT-diaphorase, whereas dicoumarol insensitive DCIP reduction was not significantly changed. On the other hand the mtDNA mutated cybrids exhibited a strong increase of both dicoumarol sensitive and insensitive DCIP reduction. The results suggest that enhanced oxidative stress and not deficient respiratory activity per se is the stimulus triggering over-expression of plasma membrane oxidative enzymes.     

Administration of an Antioxidant Prevents Lymphoma Development in Transmitochondrial Mice Overproducing Reactive Oxygen Species

Because of the difficulty to exclude possible involvement of nuclear DNA mutations, it has been a controversial issue whether pathogenic mutations in mitochondrial DNA (mtDNA) and the resultant respiration defects are involved in tumor development. To address this issue, our previous study generated transmitochondrial mice (mito-mice-ND6¹³⁹⁹⁷), which possess the nuclear and mtDNA backgrounds derived from C57BL/6J (B6) strain mice except that they carry B6 mtDNA with a G13997A mutation in the mt-Nd6 gene. Because aged mito-mice-ND6¹³⁹⁹⁷ simultaneously showed overproduction of reactive oxygen species (ROS) in bone marrow cells and high frequency of lymphoma development, current study examined the effects of administrating a ROS scavenger on the frequency of lymphoma development. We used N-acetylcysteine (NAC) as a ROS scavenger, and showed that NAC administration prevented lymphoma development. Moreover, its administration induced longevity in mito-mice-ND6¹³⁹⁹⁷. The gene expression profiles in bone marrow cells indicated the upregulation of the Fasl gene, which can be suppressed by NAC administration. Given that natural-killer (NK) cells mediate the apoptosis of various tumor cells via enhanced expression of genes encoding apoptotic ligands including Fasl gene, its overexpression would reflect the frequent lymphoma development in bone marrow cells. These observations suggest that continuous administration of an antioxidant would be an effective therapeutics to prevent lymphoma development enhanced by ROS overproduction.     

Involvement of highly sulfated chondroitin sulfate in the metastasis of the Lewis lung carcinoma cells

The altered expression of cell surface chondroitin sulfate (CS) and dermatan sulfate (DS) in cancer cells has been demonstrated to play a key role in malignant transformation and tumor metastasis. However, the functional highly sulfated structures in CS/DS chains and their involvement in the process have not been well documented. In the present study, a structural analysis of CS/DS from two mouse Lewis lung carcinoma (3LL)-derived cell lines with different metastatic potentials revealed a higher proportion of Delta(4,5)HexUA-GalNAc(4,6-O-disulfate) generated from E-units (GlcUA-GalNAc(4, 6-O-disulfate)) in highly metastatic LM66-H11 cells than in low metastatic P29 cells, although much less CS/DS is expressed by LM66-H11 than P29 cells. This key finding prompted us to study the role of CS-E-like structures in experimental lung metastasis. The metastasis of LM66-H11 cells to lungs was effectively inhibited by enzymatic removal of tumor cell surface CS or by preadministration of CS-E rich in E-units in a dose-dependent manner. In addition, immunocytochemical analysis showed that LM66-H11 rather than P29 cells expressed more strongly the CS-E epitope, which was specifically recognized by the phage display antibody GD3G7. More importantly, this antibody and a CS-E decasaccharide fraction, the minimal structure recognized by GD3G7, strongly inhibited the metastasis of LM66-H11 cells probably by modifying the proliferative and invading behavior of the metastatic tumor cells. These results suggest that the E-unit-containing epitopes are involved in the metastatic process and a potential target for the diagnosis and treatment of malignant tumors.     

Mitochondria dysfunction of Alzheimer's disease cybrids enhances Abeta toxicity

Alzheimer's disease (AD) brain reveals high rates of oxygen consumption and oxidative stress, altered antioxidant defences, increased oxidized polyunsaturated fatty acids, and elevated transition metal ions. Mitochondrial dysfunction in AD is perhaps relevant to these observations, as such may contribute to neurodegenerative cell death through the formation of reactive oxygen species (ROS) and the release of molecules that initiate programmed cell death pathways. In this study, we analyzed the effects of beta-amyloid peptide (Abeta) on human teratocarcinoma (NT2) cells expressing endogenous mitochondrial DNA (mtDNA), mtDNA from AD subjects (AD cybrids), and mtDNA from age-matched control subjects (control cybrids). In addition to finding reduced cytochrome oxidase activity, elevated ROS, and reduced ATP levels in the AD cybrids, when these cell lines were exposed to Abeta 1-40 we observed excessive mitochondrial membrane potential depolarization, increased cytoplasmic cytochrome c, and elevated caspase-3 activity. When exposed to Abeta, events associated with programmed cell death are activated in AD NT2 cybrids to a greater extent than they are in control cybrids or the native NT2 cell line, suggesting a role for mtDNA-derived mitochondrial dysfunction in AD degeneration.     

The protective roles of phosphorylated heat shock protein 27 in human cells harboring myoclonus epilepsy with ragged-red fibers A8344G mtDNA mutation

Mitochondrial DNA (mtDNA) mutations are associated with a large number of neuromuscular diseases. Myoclonus epilepsy with ragged-red fibers (MERRF) syndrome is a mitochondrial disease inherited through the maternal lineage. The most common mutation in MERRF syndrome, the A8344G mutation of mtDNA, is associated with severe defects in mitochondrial protein synthesis, which impair the assembly and function of the respiratory chain. We have previously shown that there is a decreased level of heat shock protein 27 (HSP27) in lymphoblastoid cells derived from a MERRF patient and in cytoplasmic hybrids (cybrids) harboring the A8344G mutation of mtDNA. In the present study, we found a dramatic decrease in the level of phosphorylated HSP27 (p-HSP27) in the mutant cybrids. Even though the steady-state level of p-HSP27 was reduced in the mutant cybrids, normal phosphorylation and dephosphorylation were observed upon exposure to stress, indicating normal kinase and phosphatase activities. To explore the roles that p-HSP27 may play, transfection experiments with HSP27 mutants, in which three specific serines were replaced with alanine or aspartic acid, showed that the phosphomimicking HSP27 desensitized mutant cybrids to apoptotic stress induced by staurosporine (STS). After heat shock stress, p-HSP27 was found to enter the nucleus immediately, and with a prolonged interval of recovery, p-HSP27 returned to the cytoplasm in wild-type cybrids but not in mutant cybrids. The translocation of p-HSP27 was correlated with cell viability, as shown by the increased number of apoptotic cells after p-HSP27 returned to the cytoplasm. In summary, our results demonstrate that p-HSP27 provides significant protection when cells are exposed to different stresses in the cell model of MERRF syndrome. Therapeutic agents targeting anomalous HSP27 phosphorylation might represent a potential treatment for mitochondrial diseases.     

Effect of selenite on basic mitochondrial function in human osteosarcoma cells with chronic mitochondrial stress

Mitochondrial chronic stress that originates from defective mitochondria is implicated in a growing list of human diseases. To enhance understanding of pathophysiology of chronic mitochondrial dysfunction we investigated human osteosarcoma cells with 2 types of chronic stress: corresponding to the mutation in ATP synthase subunit 6 encoded by mtDNA (NARP syndrome-mild stress) and to a total lack of mtDNA (Rho0 cells-heavy stress). We previously found that selenium influenced mitochondrial stress response and lowered ROS production. Therefore, in this study effect of selenite on other mitochondrial parameters was investigated. We showed that presence of selenium improved survival of starved cells, modified organization of mitochondrial network in NARP cybrids and decreased cytosolic calcium level in NARP and Rho0 cells. Selenium did not affect mitochondrial membrane potential, ATP level, activity of ATP synthase and activity of complex II of the respiratory chain.     

Free radicals-mediated damage in transmitochondrial cells harboring the T14487C mutation in the ND6 gene of mtDNA

We have studied the production of reactive oxygen species (ROS) in transmitochondrial cells, harboring homoplasmic levels of the T14487C mtDNA mutation in the ND6 gene of mitochondrial DNA (mtDNA). Previous work has shown that this mutation causes complex I deficiency. Here, we show that this mutation causes an overproduction of ROS leading to an increase in the oxidation of lipids and mtDNA without modification of antioxidant enzyme activities. We suggest that mutations in mtDNA affecting complex I activity may result in oxidative cellular damage, and reinforce the possible role of ROS-mediated mechanisms participating in some mtDNA-related disorders.     

A novel mutation in the mitochondrial 16S rRNA gene in a patient with MELAS syndrome, diabetes mellitus, hyperthyroidism and cardiomyopathy

Using RNase protection analysis, we found a novel C to G mutation at nucleotide position 3093 of mitochondrial DNA (mtDNA) in a previously reported 35-year-old woman exhibiting clinical features of mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS) syndrome together with diabetes mellitus, hyperthyroidism and cardiomyopathy. The patient also had an A3243G mutation in the tRNA(Leu(UUR)) gene and a 260-base pair duplication in the D-loop of mtDNA. The fibroblasts of the patient were cultured and used for the construction of cybrids using cytoplasmic transfer of the patient's mtDNA to the mtDNA-less rho(0) cells. RNA isolated from the cybrids was subjected to RNase protection analysis, and a C3093G transversion at the 16S rRNA gene and a MELAS-associated A3243G mutation of mtDNA were detected. The novel C3093G mutation together with the A3243G transition were found in muscle biopsies, hair follicles and blood cells of this patient and also in her skin fibroblasts and cybrids. The proportion of the C3093G mutant mtDNA in muscle biopsies of the patient was 51%. In contrast, the mutation was not detected in three sons of the proband. To characterize the impact of the mtDNA mutation-associated defects on mitochondrial function, we determined the respiratory enzyme activities of the primary culture of fibroblasts established from the proband, her mother and her three sons. The proportions of mtDNA with the C3093G transversion and the A3243G transition in the fibroblasts of the proband were 45 and 58%, respectively. However, the fibroblasts of the proband's mother and children harbored lower levels of mtDNA with the A3243G mutation but did not contain the C3093G mutation. The complex I activity in the proband's fibroblasts was decreased to 47% of the control but those of the fibroblasts of the mother and three sons of the proband were not significantly changed. These findings suggest that the C3093G transversion together with the A3243G transition of mtDNA impaired the respiratory function of mitochondria and caused the atypical MELAS syndrome associated with diabetes mellitus, hyperthyroidism and cardiomyopathy in this patient.     

Impaired Cellular Bioenergetics Causes Mitochondrial Calcium Handling Defects in MT-ND5 Mutant Cybrids

Mutations in mitochondrial DNA (mtDNA) can cause mitochondrial disease, a group of metabolic disorders that affect both children and adults. Interestingly, individual mtDNA mutations can cause very different clinical symptoms, however the factors that determine these phenotypes remain obscure. Defects in mitochondrial oxidative phosphorylation can disrupt cell signaling pathways, which may shape these disease phenotypes. In particular, mitochondria participate closely in cellular calcium signaling, with profound impact on cell function. Here, we examined the effects of a homoplasmic m.13565C>T mutation in MT-ND5 on cellular calcium handling using transmitochondrial cybrids (ND5 mutant cybrids). We found that the oxidation of NADH and mitochondrial membrane potential (Δψ_m) were significantly reduced in ND5 mutant cybrids. These metabolic defects were associated with a significant decrease in calcium uptake by ND5 mutant mitochondria in response to a calcium transient. Inhibition of glycolysis with 2-deoxy-D-glucose did not affect cytosolic calcium levels in control cybrids, but caused an increase in cytosolic calcium in ND5 mutant cybrids. This suggests that glycolytically-generated ATP is required not only to maintain Δψ_m in ND5 mutant mitochondria but is also critical for regulating cellular calcium homeostasis. We conclude that the m.13565C>T mutation in MT-ND5 causes defects in both mitochondrial oxidative metabolism and mitochondrial calcium sequestration. This disruption of mitochondrial calcium handling, which leads to defects in cellular calcium homeostasis, may be an important contributor to mitochondrial disease pathogenesis.     

Mitochondrial dysfunction and cancer metastasis

Mitochondria have an essential role in powering cells by generating ATP following the metabolism of pyruvate derived from glycolysis. They are also the major source of generating reactive oxygen species (ROS), which have regulatory roles in cell death and proliferation. Mutations in mitochondrial DNA (mtDNA) and dysregulation of mitochondrial metabolism have been frequently described in human tumors. Although the role of oxidative stress as the consequence of mtDNA mutations and/or altered mitochondrial functions has been demonstrated in carciongenesis, a causative role of mitochondria in tumor progression has only been demonstrated recently. Specifically, the subject of this mini-review focuses on the role of mitochondria in promoting cancer metastasis. Cancer relapse and the subsequent spreading of cancer cells to distal sites are leading causes of morbidity and mortality in cancer patients. Despite its clinical importance, the underlying mechanisms of metastasis remain to be elucidated. Recently, it was demonstrated that mitochondrial oxidative stress could actively promote tumor progression and increase the metastatic potential of cancer cells. The purpose of this mini-review is to summarize current investigations of the roles of mitochondria in cancer metastasis. Future development of diagnostic and therapeutic strategies for patients with advanced cancer will benefit from the new knowledge of mitochondrial metabolism in epithelial cancer cells and the tumor stroma.     

Bioenergetics of mitochondrial diseases associated with mtDNA mutations

This mini-review summarizes our present view of the biochemical alterations associated with mitochondrial DNA (mtDNA) point mutations. Mitochondrial cytopathies caused by mutations of mtDNA are well-known genetic and clinical entities, but the biochemical pathogenic mechanisms are often obscure. Leber's hereditary optic neuropathy (LHON) is due to three main mutations in genes for complex I subunits. Even if the catalytic activity of complex I is maintained except in cells carrying the 3460/ND1 mutation, in all cases there is a change in sensitivity to complex I inhibitors and an impairment of mitochondrial respiration, eliciting the possibility of generation of reactive oxygen species (ROS) by the complex. Neurogenic muscle weakness, Ataxia and Retinitis Pigmentosa (NARP), is due to a mutation in the ATPase-6 gene. In NARP patients ATP synthesis is strongly depressed to an extent proportional to the mutation load; nevertheless, ATP hydrolysis and ATP-driven proton translocation are not affected. It is suggested that the NARP mutation affects the ability of the enzyme to couple proton transport to ATP synthesis. A point mutation in subunit III of cytochrome c oxidase is accompanied by a syndrome resembling MELAS: however, no major biochemical defect is found, if we except an enhanced production of ROS. The mechanism of such enhancement is at present unknown. In this review, we draw attention to a few examples in which the overproduction of ROS might represent a common step in the induction of clinical phenotypes and/or in the progression of several human pathologies associated with mtDNA point mutations.     

细胞糖代谢方式改变与肿瘤转移的相关性

肿瘤转移是引起肿瘤相关死亡的主要原因,肿瘤细胞的代谢异常在肿瘤转移中扮演重要角色。肿瘤的糖代谢以“Warburg效应”为显著特征,即细胞在有氧条件下也以糖酵解为主要糖代谢途径提供能量。而这种现象在转移性肿瘤细胞中更为突出,表现为葡萄糖的大量摄取、高糖酵解速率和核酸合成速率等,这为肿瘤细胞的快速生长和增殖提供了重要的能量和物质基础。对于肿瘤转移过程中相关代谢改变的研究,将为最终揭示肿瘤转移的机制打下基础。本文综述肿瘤细胞糖代谢中糖酵解、线粒体有氧代谢及磷酸戊糖途径中的变化与肿瘤转移发生的相关性,其结果为进一步从调控肿瘤代谢角度发现新的肿瘤转移控制手段提供了启示。