Response to: Comment on ‘Human adult neurogenesis across the ages: An immunohistochemical study’

Dear Editor, It is with great interest that we read the comment by Marucci [1] referring to our publication ‘Human adult neurogenesis across the ages: An immunohistochemical study’ [2]. Since the seminal paper of Eriksson et al. in 1998, human adult neurogenesis has become a major area of research in neuroscience [3]. Although an agerelated decline in human adult neurogenesis is not disputed, opinions differ on the functional significance of the residual neuroblasts. The majority view, as shared by Marucci, is that adult neurogenesis, at least in the subgranular zone, continues to be functionally important throughout life. The implication being that this process can be potentially manipulated in its extent and direction to treat neurological disorders. Marucci suggests that our conclusions that ‘proliferative cells in these niches are not only sparse but also destined to be microglia’ [2], are ‘questionable’ and are ‘severely affecting the increasingly relevant efforts to exploit restorative neurobiology in some neurological disorders’ [1], implying that the techniques and markers we employed would have underestimated the true extent of neurogenesis. Here, we take the opportunity to address the issues raised by Marucci, and where appropriate, respectfully point out where our findings appear to have been misinterpreted. First, we agree with Marucci that SOX2 would have been a useful marker in our study. A small proportion of SOX2 cells (Type 2a) would have been expected to colocalize with our proliferation markers and doublecortin (DCX) [4]. However, in the mouse, at least, SOX2 is mainly expressed in non-proliferating neural stem cells [5] and would not necessarily have been more informative in determining cell fate than the markers we employed. Indeed, EGFR appears to be a specific marker of a subpopulation of activatable quiescent precursor cells [6]. Using DCX as a marker (largely) of a neuronal-specific fate [4], we were also able to compare our results with those of other studies in the field. Knoth and colleagues used DCX to quantify SGZ neurogenesis across the human lifespan in combination with PCNA [7] and demonstrated colocalization of SOX2 and DCX, as well as of SOX2 with PCNA or Ki67 (Knoth et al. Fig 7 [7]). Sanai and colleagues used DCX and Ki67 in their quantification of SVZ neurogenesis across the human lifespan [8]. We were not aware of the role of ALK5 in adult neurogenesis but note the reported overlap with DCX expression [9]. Secondly, Marucci reports that, compared to our finding, Weissleder et al. described a greater number of Ki67 cells in the human subventricular zone (SVZ [also referred to the subependymal zone] (Weissleder et al., Fig 1D [10])). However, it is difficult to directly compare these studies as Weissleder et al. counted Ki67 cells per section (each 50 microns thick) and normalized to the length of the SVZ, whereas we performed stereological quantification on 12 micron thick sections and normalized to the area rather than the length of the SVZ. The width of the SVZ varies considerably along its rostrocaudal length, being wider rostrally where it extends into the rostral migratory stream (Dennis et al. Fig. 7D, E and FigS5 [2]). Similar to Weissleder et al., we also found Ki67 cells in ‘doublets’ in the adult SVZ (Dennis et al. Fig. 1G [2]) although these were not specifically described in our original article. However, in the absence of colocalization data, it is not possible to determine the lineage of their cells. Based on the age-related increase in Iba1 mRNA reported by Weissleder et al. and our finding that the only Ki67 cells in adults were also Iba1 positive, it would seem reasonable to assume, that in the absence of further data, that the pairs and occasional groups of Ki67 nuclei identified by Weissleder et al. are microglial. Thirdly, Marucci argues that by using post-mortem tissue from individuals without known neurological abnormalities our findings are ‘likely influenced by strong biases’ and our ‘conclusions could be misleading’. He then describes the use of biopsy samples from patients undergoing surgery for medically intractable epilepsy as an alternative source of material in which to study neurogenesis and describes an ex vivo study where such samples were used to generate mature neurons [11]. However, experiments in animal models of induced