The increased GFAP expression, hypertrophy and elevated proliferation of mature astrocytes implied the induction of spontaneous astrogliosis in Mlc1-ErbB2V664E mice after Dox withdrawal. Consistent with the ectopic expression of ErbB2V664E, various brain regions in Mlc1-ErbB2V664E mice exhibited immunoreactivity with the ErbB2 antibody (Supplementary Figure S7). This immunoreactivity localized well in GFAP+ cells, confirming cell-targeting specificity in Mlc1-ErbB2V664E mice (Figure 4e). There was neither scarring nor aggregation of reactive astrocytes in the cerebral cortices, consistent with the idea that astrocytes in Mlc1-ErbB2V664E mice reacted to intrinsic factors instead of extrinsic ones released from injury sites or degenerative plaques. To further test this idea, primary astrocytes purified from Mlc1-ErbB2V664E and control mice were examined in vitro. Cultured cortical astrocytes from either control or Mlc1-ErbB2V664E mice exhibited reactive features such as the GFAP expression (Supplementary Figure S8a). However, normal cultured astrocytes appeared to be flat and spreading, adhering well to the bottom of the cell culture dish, with only a few of them expressing the radial glial cell marker RC2, whereas astrocytes from Mlc1-ErbB2V664E mice exhibited small but plump shapes and high levels of RC2, as well as accelerated proliferation rates (Supplementary Figure S8a and b). These results suggest that cell-autonomous ErbB activation promoted astrocyte proliferation. The observation of high RC2 expression in Mlc1-ErbB2V664E astrocytes was in line with the previous report that TGFα stimulates cultured astrocytes to dedifferentiate into NSCs .