Frequent polyclonal origin of micro-dissected thyroid follicles in normal but not in nodular thyroid tissue

X-chromosome inactivation has been frequently used to infer the clonal expansion of a tumour and decide between neoplasia or hyperplasia. Such studies using heterozygous polymorphisms in X-chromosome-linked markers demonstrate a predominant clonal origin of tumor tissues including the thyroid gland. However, recent results of clonal analysis after PCR amplification of X-linked markers from micro-dissected tumours need to be interpreted with caution because the sample could lie within a monoclonal patch of cells that arise from a single progenitor cell and share an identical pattern of X-chromosome inactivation. In that case X-chromosome based diagnosis would not be a reliable marker for neoplasia. Because reports concerning the exact thyroid patch size are rare and since the patch volume does not illustrate the patch architecture we intended to answer the question how often in normal thyroid parenchyma a single follicle (the smallest structural unit) originates from more than one progenitor cell. We micro-dissected single follicles from 12µm sections of fresh frozen normal and thyroid disease tissue. Using a PCR based HUMARA clonality assay we then studied the clonal origin of about 900 follicles. In the three normal thyroid tissue samples 18 of 99, 22 of 79 and 22 of 94 follicles were of polyclonal origin. This clearly suggests that single thyroid follicles from normal thyroid frequently belong to different patches which implies a rather small thyroid patch size. In contrast, all 43 follicles and 74 of 79 follicles of nodular tissue did not only present monoclonal but showed X-inactivation for the same allele. This strongly indicates that the follicles from the investigated nodular tissue originate from a single precursor cell. Graves' disease tissue presented similar to normal thyroid with 19% to 27% of follicles with polyclonal origin. Therefore, clonal analysis based on a number of single micro-dissected follicles (20–50) can differentiate normal from nodular thyroid tissue and could be helpful in the clonal characterisation of a tumour.