Slit and Glypican-1 mRNAs Are Coexpressed in the Reactive Astrocytes of the Injured Adult Brain SEITA HAGINO, 1,2 * KEN ISEKI, 2,3 TETSUJI MORI, 2 YUXIANG ZHANG, 2 TSUYOSHI HIKAKE, 2 SACHIHIKO YOKOYA, 2 MAYUMI TAKEUCHI, 2 HIROMI HASIMOTO, 2 SHINICHI KIKUCHI, 1 AND AKIO WANAKA 4 1 Department of Orthopedic Surgery, Fukushima Medical University School of Medicine, Fukushima, Japan 2 Department of Cell Science, Institute of Biomedical Sciences, Fukushima Medical University School of Medicine, Fukushima, Japan 3 Department of Emergency Medicine, Fukushima Medical University School of Medicine, Fukushima, Japan 4 Department of Anatomy, Nara Medical University School of Medicine, Shijo-cho Kashihara-city, Nara, Japan KEY WORDS Slit; reactive astrocyte; gliosis; brain injury; glypican-1 ABSTRACT The slit family serves as a repellent for growing axons toward correct targets during neural development. A recent report describes slit mRNAs expressed in various brain regions in adult rats. However, their functions in the adult nervous system remain unknown. In the present study, we investigated whether slit mRNAs were expressed in the cryo-injured brain, using in situ hybridization. All slit family members were expressed at the lesion. Slit2 mRNA was the most intensely expressed in the cells surrounding the necrotic tissue. A double-labeling study showed that slit2 mRNA was expressed in the glial fibrillary acidic protein (GFAP)-positive reactive astrocytes. In addition, glypican-1, a heparan sulfate proteoglycan that serves as a high-affinity receptor for Slit protein, was coexpressed with slit2 mRNA in the reactive astrocytes. These findings suggested that slit2 might prevent regenerating axons from entering into the lesion in concert with glypican-1. GLIA 42:130 –138, 2003. © 2003 Wiley-Liss, Inc. INTRODUCTION Axons in the adult mammalian central nervous sys- tem (CNS) are unable to regenerate spontaneously af- ter injury. The failure in regeneration is attributed, in part, to inhibitory environment (Fawcett and Asher, 1999). Nogo-A is a myelin-associated neurite out- growth inhibitor, present in oligodendrocyte and mye- lin at the intact tissue adjacent to the CNS lesion (Huber et al., 2002). Neutralization of Nogo-A by mono- clonal antibody IN-1 promotes axon regeneration and functional recovery after spinal cord injury (Schnell and Schwab, 1990, 1993; Bregman et al., 1995; Bro- samle et al., 2000; Chen et al., 2000; Merkler et al., 2001). In addition to the nonpermissive oligodendro- cyte, astrocytes form a glial scar that protects the in- tact tissue from spreading damage and simultaneously produces inhibitory molecules for regenerating axons (Silver, 1994). Chondroitin sulfate proteoglycans (CS- PGs) expressed in reactive astrocytes are one of the representative inhibitory molecules (McKeon et al., 1991). Many CS-PGs have been shown to be upregu- lated at sites of CNS injury (Jaworski et al., 1999; McKeon et al., 1999; Asher et al., 2000, 2002; Jones et al., 2002). In contrast, heparan sulfate proteoglycans (HS-PGs) are growth-supportive molecules in the de- Grant sponsor: Ministry of Education, Culture, Sports, Science, and Technol- ogy, Tokyo, Japan. *Correspondence to: Seita Hagino, Department of Orthopedic Surgery, Fuku- shima Medical University School of Medicine, 1 Hikarigaoka, Fukushima City, Fukushima 960-1295, Japan. E-mail: shagino@fmu.ac.jp Received 19 August 2002; Accepted 16 December 2002 DOI 10.1002/glia.10207 GLIA 42:130 –138 (2003) © 2003 Wiley-Liss, Inc.