Sixteen hours after injection, the dye was obviously accumulated in the brains of the mutant mice, but not in WT mice (Fig. 7A, B), demonstrating impaired BBB permeability in the mutant mice. YAP/astrocytes thus showed hyperactivation of the JAKSTAT inflammatory pathway and reactive astrogliosis. Expression of SOCS3 in YAP/astrocytes prevented the hyperactivation of STAT3 and partially restored the astrocytic activation. Finally, reactive astrogliosis was associated with bloodbrain barrier dysfunction in YAP brain-selective knockout mice. Taken together, these results identify unrecognized functions of YAP in preventing reactive astrogliosis and reveal a pathway of YAP-SOCS for the negatively control of neuroinflammation. Keywords: bloodbrain barrier, reactive astrogliosis, SOCS3, STAT3, YAP == Introduction == Astrocytes, a major type of glial cells in the brain, are crucial for a wide variety of functions in the central nervous system (CNS), including regulating synaptic neurotransmission (Eroglu and Barres 2010) and modulating blood flow (Takano et al. 2006). Under physiological condition, astrocytes give rise to a dense network of finely branching processes, some of which envelop blood vessel, and form bloodbrain barrier JV15-2 (BBB) with endothelial cells and pericytes. BBB (neurovascular unit) is a physical and metabolic barrier between the CNS and the systemic circulation to maintain the microenvironment of the CNS (Abbott et al. 2010; Sofroniew and Vinters 2010; Liebner et al. 2011; Tajes et al. 2014). Recently, astrocytes are also considered to be a special type of immune neuroglia at the CNS (Farina et al. 2007; Ransohoff and Engelhardt 2012; Burda and Sofroniew 2014; Xie and Yang 2015). Under stress or pathological insults, astrocytes are activated (so-called reactive astrogliosis) (Burda and Sofroniew 2014). The reactive astrocytes produce and release diverse pro- or anti-inflammatory cytokines, chemokines, and neutrophins to cause tissue damage or repair (Shen et al. 2012; Yang et al. 2012; Burda and Sofroniew 2014; Choi et al. 2014; Xie and Yang 2015). Although some signaling pathways, such as STAT3, are identified to regulate astrocyte activation (Burda and Sofroniew 2014), the molecular mechanisms of reactive astrogliosis under different pathological insults remain unclear. The Hippo pathway is critical for the development of multiple ST271 organs, including liver, heart, kidney, and intestine (Pan 2010; Mo et al. 2014; Piccolo et al. 2014). Activation of this pathway leads to phosphorylation of Yes-associated protein (YAP), a transcriptional cofactor, and its subsequent proteosomal degradation or cytoplasmic retention. Dephosphorylated YAP enters nuclei and interacts with ST271 transcriptional enhancer factor domain family proteins to induce target gene expression. YAP is believed to control organ size by promoting cell proliferation, differentiation, and survival (Zhao et al. 2007; Pan 2010; Mo et al. 2014; Piccolo et al. 2014). A glaring gap in our understanding of YAP function is that little is known about its role in developing nervous system. Here, we provide evidence that in developing brains, ST271 YAP was selectively expressed in astrocytes and neural stem cells (NSCs). YAP deletion resulted in reactive astrogliosis, astrocyte-driven microglial activation, which was associated with reduced BBB function. At the molecular level, YAP in astrocytes was necessary for preventing hyperactivation of the JAK/STAT inflammatory pathway, which was likely due to YAP induction of suppressor of cytokine signaling (SOCS) family gene expression. Taken together, these results reveal a pathway of YAP-SOCS for the negatively control of STAT-mediated inflammatory response and reactive astrogliosis, a crucial event for maintaining adequate BBB function. == Materials and Methods == == Animals and Mouse Breeding == Yapnestin-CKO conditional knockout mice were generated by crossing floxed Yap allele (Yapf/f) with nestin-Cre transgenic mice. Both Yapf/fand nestin-Cre mice were maintained in C57BL/6 strain background. Yapf/fmice were generated as previously described (Zhang et al. 2010; Wang et al. 2014). Nestin-Cre transgenic mice were purchased from the Jackson Laboratory (donated by Dr Rudiger Klein). Briefly, we first generated nestin-Cre+/YAPf/wmouse, by crossing nestin-Cre+male mice with Yapf/ffemale mice. Nestin-Cre+/YAPf/wmale mice were then breed with Yapf/ffemale mice. This breeding strategy yielded 25% progeny with the homozygous mice (Yapnestin-CKO) and 25% Yapf/fas control littermates. Embryonic day (E) 0. 5 was defined as noon of the day when the vaginal plug was detected. Identification of these mice was performed as described inSupplementary Figure 1 . The use of experimental animals has been approved by the Institutional Animal Care and Use Committee (IACUC) at Georgia Regents University in accordance with the NIH guidelines. == Primary Cultures of Neural Stem Cells, Astrocytes, Microglia, and Neurons == For Yapnestin-CKO neural stem ST271 cell culture, Nestin-Cre+/YAPf/wmale mice were used to breed with YAPf/ffemale mice. The pregnant mice (E14. 5) were sacrificed, and embryonic mice were taken out. Genomic DNAs of each embryonic mouse were collected for genotyping, and littermates were used as controls..