What Happens in “Chromospheres”?

The recent paper by Santana et al. titled “Isolation, Characterization, and Differentiation of Progenitor Cells From Adult Human Adrenal Medulla” [1] is an interesting contribution to human chromaffin cell biology. However, I am concerned about omissions of important information in the literature that could lead to alternate interpretations of the authors' data. The paper cites two publications from my laboratory showing that chromaffin cells proliferate throughout life and can acquire a neuron-like phenotype. However, the two cited papers are about chromaffin cells from rats, whereas a substantial literature on adult human chromaffin cells is overlooked. Two particularly important points are missed. The first is that human chromaffin cells acquire a neuron-like phenotype in monolayer cultures without any need for a “chromosphere” stage. This apparent transdifferentiation occurs spontaneously and is increased by nerve growth factor [2–4] (which the authors mention only as a hypothetical possibility for increasing neuronal differentiation in future experiments). Morphological changes include outgrowth of axon-like processes, and electron micrographs show apparent migration of numerous large secretory vesicles characteristic of chromaffin cells from the perikarya into processes as they begin to form. Biochemical changes include decreased epinephrine production, consistent with decreased phenylethanolamine N-methyltransferase expression as shown by Santana et al. [1], and increased synthesis of regulatory peptide neurotransmitters [4]. The second important overlooked point is that, in contrast to rat chromaffin cells, human chromaffin cells undergo these changes without any preceding cell proliferation. A sensitive and specific test for proliferation that employs double immunohistochemical staining for bromodeoxyuridine incorporation and tyrosine hydroxylase expression [5] shows no 5-bromo-2′-deoxyuridine (BrdU) labeling of any tyrosine hydroxylase-positive cells, with or without processes, in human adrenal medullary cell cultures plated directly as monolayers without first having been maintained as chromospheres [6]. It would be of interest to use the same test with differentiated cells derived from chromospheres. The absence of BrdU incorporation would suggest that the chromosphere cultures are upregulating genes required for transdifferentiation, perhaps in a manner analogous to “priming” of PC12 pheochromocytoma cells [7], rather than proliferatively expanding a progenitor cell population. If so, it would be important to determine whether the same transcriptional changes occur in direct monolayer cultures without chromospheres. Profiling the molecular changes that precede neuronal differentiation in those cultures would also be useful in its own right because, to my knowledge, the model was last studied almost 14 years ago [6] and has not been further characterized. If derivation of neurons from chromosphere cultures of adult human adrenal medullary cells results from reprogramming of differentiated chromaffin cells rather than proliferative expansion of neural progenitors, this type of culture would be an interesting developmental model but probably not a ready potential source of neurons for clinical purposes.