Ann. N.Y. Acad. Sci. ISSN 0077-8923 ANNALS OF THE NEW YORK ACADEMY OF SCIENCES Issue: Thymosins in Health and Disease Thymosin 4 expression reveals intriguing similarities between fetal and cancer cells Gavino Faa, 1 Sonia Nemolato, 1 Tiziana Cabras, 2 Daniela Fanni, 1 Clara Gerosa, 1 Mattia Fanari, 1 Annalisa Locci, 1 Vassilios Fanos, 3 Irene Messana, 2 and Massimo Castagnola 4 1 Department of Pathology, University Hospital San Giovanni di Dio, 2 Department of Life and Environmental Sciences, 3 NICU Center, University of Cagliari, Cagliari, Italy. 4 Institute of Biochemistry and Clinical Biochemistry, Faculty of Medicine, Catholic University, Rome, Italy Address for correspondence: Daniela Fanni, Department of Pathology, University Hospital San Giovanni di Dio, University of Cagliari, Via Ospedale 54, 09124 Cagliari, Italy. fandan73@yahoo.it Thymosin 4 (T4) is highly expressed in saliva of human newborns but not in adults. Here preliminary immuno- histochemical analyses on different human tissues are reported. Immunoreactivity for T4 in human salivary glands show high quantities of T4 before birth, followed by downregulation of expression in adulthood. In contrast, T4 is detected in tumors of salivary glands, suggesting that tumor cells might utilize fetal programs, including T4 syn- thesis. Immunohistochemical analyses in the gastrointestinal tract showed strong reactivity for T4 in enterocytes during development, but weak immunostaining in mature enterocytes. In colorectal cancer, the association of a high expression of T4 with epithelial–mesenchymal transition was observed. On the basis of these data, the process of epithelial–mesenchymal transition could represent the unifying process that explains the role of T4 during fetal development and in cancer progression. Keywords: thymosin beta 4; immunohistochemistry; development; cancer; embryogenesis Introduction Thymosin 4(T4) is a naturally occurring peptide, first isolated in 1966 by Goldstein et al . from the calf thymus among other lymphocytopoietic factors. 1 The complete amino acid sequence of T4 was de- scribed in 1981; it contains 43 amino acids, with a high proportion of lysyl and glutamyl residues. 2 The human T4 gene (hT4) is located on chromosome X and comprises three exons and two introns. 3 The primary translation product is modified by removal of the N-terminal methionine and acetylation. 4,5 Recent studies highlighted extracellular roles for T4 (for a recent review, see Ref. 6). However, because the peptide does not contain N-terminal leader sequences, the mechanism of release is com- pletely unknown. 7 T4 is considered the most abundant among -thymosin peptides in mammalian tissues: its activity has been mainly related to the regula- tion of actin polymerization in living cells. 8,6 T4 is also thought to be involved in many criti- cal biological activities, 9 including angiogenesis, 10 wound healing, 11 inflammatory response, 12 and cell migration. 13 Most mammalian cells express two -thymosin variants simultaneously; T4, associ- ated with T10, is the most abundant -thymosin in human cells. 14–16 Our interest in T4 takes its origin from the find- ing that this peptide is highly expressed in saliva of human newborns, but not in saliva of adults. 17 It was necessary to first determine in which tis- sue, and in which cells, T4 is synthesized and secreted into the oral cavity. To answer these ques- tions, we analyzed tissue samples from human sali- vary glands of neonates with different gestational ages and performed immunohistochemical anal- yses on these tissues looking for the presence of the peptide. T4 immunostaining was identified in acinar cells of the parotid, submandibular, and sublingual glands, as well as in minor salivary glands, clearly indicating these cells as the source of T4 in doi: 10.1111/j.1749-6632.2012.06679.x Ann. N.Y. Acad. Sci. 1269 (2012) 53–60 c  2012 New York Academy of Sciences. 53