BMC Cancer 2005, 5:102 http://www.biomedcentral.com/1471-2407/5/102 tumors studied here do not result from somatic mutations in their mtDNA. In contrast to previous reports of genetic heterogeneity in normal human brain mtDNA, [40,67] we found no evi- dence for genetic heterogeneity in mtDNA isolated from synaptic and nonsynaptic mitochondria in the C57BL/6J and VM mouse strains by direct sequencing. On the other hand, our findings that most mtDNA changes in the mouse brain tumors were in hypervariable regions of mononucleotide repeats are in general agreement with previous studies in human brain tumors (Table 4). We detected a single heteroplasmic mutation in the 16srRNA gene of the VM-M metastatic cell line, which was deter- mined by sequence analysis [68]. Since this heteroplasmic mutation was not detected in another independently derived cell line from this tumor, it is unlikely that the mutation is associated with either tumorigenesis or metas- tasis. The EPEN cell line contained a nucleotide variation compared to the B6 genotype at position 9461 in the ND3 gene. This nucleotide variation in the EPEN cell line also was found in the VM-NM and VM-M cell lines. This poly- morphism could be due to a base pair wobble at that loca- tion or could result from a nucleotide variation in the original C57BL/6J mouse from which the EPEN tumor arose [44]. The four mouse brain tumor cell lines we examined repre- sent both spontaneous and chemically induced tumors that differ in cell origin, biochemical composition, and angiogenesis. The selection of the cell lines provides a comparison for defining the contribution that mtDNA might play in carcinogenesis. Although we did not find any mtDNA pathogenic mutations in these brain tumors, we do not rule out the possibility that such mutations might occur in other spontaneous mouse brain tumors or in mouse brain tumors induced with other chemical car- cinogens. Our findings suggest that mtDNA mutations do not likely contribute to the development or progression of CT-2A or EPEN chemically induced tumors or the meta- static and nonmetastatic spontaneous VM brain tumors. With the exception of a single variation at nucleotide 11780 in the ND5 gene, our whole mitochondrial genome sequence for the B6 strain was in complete agree- ment with the recently republished sequence for this mouse strain [23]. This single nucleotide difference involved a T->A transversion that changed isoleucine to methionine. This difference was also found in the VM mouse strain as well as in all examined mouse brain tumor cell lines. It is not yet clear if this variation represents an error in the republished B6 mtDNA sequence or a population genetic variation among B6 mouse strains. Brain mitochondria were fractionated into different populations for genotyping to confirm that there was no mtDNA heterogeneity in hypervariable regions that might be associated with metabolic heterogeneity. Sequencing of all three brain populations also served as an internal control for the genotype of the mouse strains. In summary, our results show that the mitochondrial genome of these mouse brain tumor cell lines contain mutations in regions of mononucleotide repeats. These mtDNA mutations in the mouse brain tumors are similar to those found previously in hypervariable regions of the D-loop in human brain tumors. However, none of the mouse mutations were pathogenic suggesting that mtDNA changes do not contribute to the carcinogenic progression of these chemically induced and spontaneous mouse brain tumors. List of abbreviations The abbreviations used are: OL, origin of replication of the light strand; mtDNA, mitochondrial DNA; kb, kilobase; D-loop, displacement loop; B6, C57BL/6J Competing interests The author(s) declare that they have no competing interests. Authors' contributions MK carried out mitochondrial isolation, amplification and sequencing of brain and tumor mtDNA, sequence analysis and drafted the manuscript. TS participated in the study's design, coordination, and editing of the manuscript. Acknowledgements We would like to thank Laura Abate and Kristen Gorham for technical assistance. This work was supported from NIH grants HD39722 and CA102135, a grant from the American Institute of Cancer Research and the Boston College Research Expense Fund. References 1. Wolstenholme DR: Animal mitochondrial DNA: structure and evolution. Int Rev Cytol 1992, 141:173-216. 2. Barrell BG, Bankier AT, Drouin J: A different genetic code in human mitochondria. Nature 1979, 282:189-194. 3. 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