title: Tissue kallikrein protects SH-SY5Y neuronal cells against oxygen and glucose deprivation-induced injury through bradykinin B2 receptor-dependent regulation of autophagy induction.
abstract: Previous studies have demonstrated that tissue kallikrein (TK) protects against cerebral ischemia injury mainly through inhibition of apoptosis via bradykinin B2 receptor (B2R). In this study, we proposed that autophagy induction contributed to the neuroprotective mechanism of TK. To validate this hypothesis, we investigated TK-induced autophagy and its signaling mechanisms in human SH-SY5Y cells exposed to oxygen and glucose deprivation (OGD). We found that TK treatment enhanced autophagy induction, reflected by augmented LC3 conversion and Beclin1 expression, decreased p62 levels and increased monomeric red fluorescent protein-LC3 puncta formation. Green fluorescent protein-monomeric red fluorescent protein-LC3 adenovirus assay indicated that TK maintained autophagic flux. Moreover, bafilomycin A1 (Baf.A1) caused obvious LC3-II accumulation either in the presence or absence of TK. Autophagy inhibition by Beclin1 knockdown or Baf.A1 treatment abrogated the neuroprotective effects of TK. Mitogen-activated protein kinase kinase 1/2 (MEK1/2)/extracellular signal-regulated kinase (ERK)1/2 and AMP-activated protein kinase (AMPK)/tuberous sclerosis complex 2 (TSC2)/mammalian target of rapamycin (mTOR) signaling were induced by OGD stress and enhanced by TK. MEK/ERK inhibitor U0126 alone elevated autophagy in OGD conditions, but impaired TK-induced autophagy. Blockade of AMPK/TSC2/mTOR signaling by AMPK inhibitor compound C and shRNA mediated the knockdown of AMPK α1 and TSC2 but abolished autophagy in SH-SY5Y cells exposed to OGD treated either with or without TK. Moreover, B2R expression was up-regulated by OGD exposure. B2R knockdown attenuated autophagy and suppressed MEK1/2/ERK1/2 and AMPK/TSC2/mTOR signaling in OGD conditions in either the presence or absence of TK. In sum, we revealed the significance of B2R-mediated MEK/ERK and AMPK signaling in autophagy induction under OGD stress, and proposed novel mechanisms involved in the neuropotective function of TK through B2R-dependent regulation of autophagy. We propose the depicted model for the neuroprotective mechanism of tissue kallikrein (TK) during OGD stress: TK enhances bradykinin B2 receptor (B2R)-mediated MEK1/2/ERK1/2 and AMPK/TSC2/mTOR signaling, thus inducing protective autophagy. The findings reported in this study should provide new evidence for the pro-survival role of B2R-mediated autophagy in cerebral ischemia.
authors.affiliation: Department of Neurology, Huashan Hospital, State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai, China.
authors.forename: Yanping
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Department of Neurology, Yueyang Hospital of Integrated Traditional Chinese and Western Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai, China.
authors.forename: Zhengyu
authors.initials: Z
authors.lastname: Lu
authors.affiliation: Department of Neurology, Huashan Hospital, State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai, China.
authors.forename: Mei
authors.initials: M
authors.lastname: Cui
authors.affiliation: Department of Neurology, Huashan Hospital, State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Yang
authors.affiliation: Department of Neurology, Huashan Hospital, State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai, China. tangyuping39@hotmail.com.
authors.forename: Yuping
authors.initials: Y
authors.lastname: Tang
authors.affiliation: Department of Neurology, Huashan Hospital, State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai, China. dong_qiang@fudan.edu.cn.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Dong
mesh_terms: D001343:Autophagy / Q000187:drug effects*; D000071186:Beclin-1 / Q000096:biosynthesis; D015687:Cell Hypoxia / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D005947:Glucose / Q000172:deficiency*; D006801:Humans; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008869:Microtubule-Associated Proteins / Q000096:biosynthesis; D009474:Neurons / Q000187:drug effects*; D018696:Neuroprotective Agents / Q000494:pharmacology*; D043782:Receptor, Bradykinin B2 / Q000187:drug effects*; D058570:TOR Serine-Threonine Kinases / Q000096:biosynthesis / Q000235:genetics; D020840:Tissue Kallikreins / Q000494:pharmacology*
chemical_list: C491997:BECN1 protein, human; D000071186:Beclin-1; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; D018696:Neuroprotective Agents; D043782:Receptor, Bradykinin B2; C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases; D020840:Tissue Kallikreins; D005947:Glucose
keywords: autophagy; bradykinin B2 receptor; oxygen and glucose deprivation; signaling; tissue kallikrein