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Thrombin Mediates Migration of Rat Brain Astrocytes via PLC, Ca2+, CaMKII, PKCα, and AP-1-Dependent Matrix Metalloproteinase-9 Expression

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Abstract

Matrix metalloproteinase-9 (MMP-9) plays a crucial role in pathological processes of brain inflammation, injury, and neurodegeneration. Thrombin has been known as a regulator of MMP-9 expression and cells migration. However, the mechanisms underlying thrombin-induced MMP-9 expression in rat brain astrocytes (RBA-1 cells) remain unclear. Here, we demonstrated that thrombin induced the expression of pro-form MMP-9 and migration of RBA-1 cells, which were inhibited by pretreatment with the inhibitor of Gq-coupled receptor (GPAnt2A), Gi/o-coupled receptor (GPAnt2), PC-PLC (D609), PI-PLC (U73122), Ca2+-ATPase (thapsigargin, TG), calmodulin (CaMI), CaMKII (KN62), PKC (Gö6976 or GF109203X), MEK1/2 (PD98059), p38 MAPK (SB202190), JNK1/2 (SP600125), or AP-1 (Tanshinone IIA) or the intracellular calcium chelator (BAPTA/AM) and transfection with siRNA of PKCα, Erk2, JNK1, p38 MAPK, c-Jun, or c-Fos. In addition, thrombin-induced elevation of intracellular Ca2+ concentration was attenuated by PPACK (a thrombin inhibitor). Thrombin further induced CaMKII phosphorylation and PKCα translocation, which were inhibited by U73122, D609, KN62, TG, or BAPTA/AM. Thrombin also induced PKCα-dependent p42/p44 MAPK and JNK1/2, but not p38 MAPK activation. Finally, we showed that thrombin enhanced c-Fos expression and c-Jun phosphorylation. c-Fos mRNA levels induced by thrombin were reduced by PD98059, SP600125, and Gö6976, but not SB202190. Thrombin stimulated in vivo binding of c-Fos to the MMP-9 promoter, which was reduced by pretreatment with SP600125 or PD98059, but not SB202190. These results concluded that thrombin activated a PLC/Ca2+/CaMKII/PKCα/p42/p44 MAPK and JNK1/2 pathway, which in turn triggered AP-1 activation and ultimately induced MMP-9 expression in RBA-1 cells.

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Acknowledgments

This work was supported by the Ministry of Education, Taiwan, grant numbers EMRPD1C0261 and EMRPD1C0271; National Science Council, Taiwan, grant numbers NSC101-2321-B-182-013, NSC101-2320-B-182-039-MY3, NSC99-2321-B182-003, and NSC98-2320-B-255-001-MY3; and Chang Gung Medical Research Foundation, grant numbers CMRPD180373, CMRPD1B0381, CMRPG391033, and CMRPG3B1091.

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Authors and Affiliations

  1. Department of Anesthetics, Chang Gung Memorial Hospital at Lin-Kou and College of Medicine, Chang Gung University, Kwei-San, Tao-Yuan, Taiwan

    Chih-Chung Lin & I-Ta Lee

  2. School of Medicine, Fu-Jen Catholic University, Xinzhuang, New Taipei City, Taiwan

    Wen-Bin Wu

  3. Department of Physiology and Pharmacology and Health Aging Research Center, College of Medicine, Chang Gung University, 259 Wen-Hwa 1st Road, Kwei-San, Tao-Yuan, Taiwan

    Chiung-Ju Liu, Li-Der Hsiao, Chien-Chung Yang & Chuen-Mao Yang

  4. Division of Basic Medical Sciences, Department of Nursing, Chang Gung University of Science and Technology, Tao-Yuan, Taiwan

    Hsi-Lung Hsieh

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  1. Chih-Chung Lin
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Correspondence to Chuen-Mao Yang.

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Lin, CC., Lee, IT., Wu, WB. et al. Thrombin Mediates Migration of Rat Brain Astrocytes via PLC, Ca2+, CaMKII, PKCα, and AP-1-Dependent Matrix Metalloproteinase-9 Expression. Mol Neurobiol 48, 616–630 (2013). https://doi.org/10.1007/s12035-013-8450-6

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  • DOI: https://doi.org/10.1007/s12035-013-8450-6

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