title: Isoquercetin ameliorates tunicamycin-induced apoptosis in rat dorsal root ganglion neurons via suppressing ROS-dependent endoplasmic reticulum stress.
abstract: BACKGROUND
  Recent studies showed that Isoquercetin (Iso), a novel extracts of plants and fruits could protect neuronal cells from neurotoxicity and neuro-inflammation. However, its role in acute spinal cord injury (ASCI) have not been elucidated. In the present study, we investigated whether Iso could prevent Tunicamycin (TUN)-induced rat dorsal root ganglion (DRG) neurons from apoptosis and endoplasmic reticulum (ER) stress.
  
  
  METHODS
  DRG neurons were pre-treated with different doses of Iso for 24h (h) and then were stimulated with TUN (0.75μg/ml) for 24h. The cytotoxic effects and apoptosis were detected by MTT assay and TUNEL staining. The protein and mRNA expression levels were detected by Real Time PCR and Western blot, respectively. The localization of cleaved caspase-12 was evaluated by immunofluorescent staining and Western blot. The activation of caspase were measured by colorimetric assays and Western blot. Lactate Dehydrogenase (LDH) and Malondialdehyde (MDA) leakage were detected by the LDH or MDA Detection Kit PLUS.
  
  
  RESULTS
  Iso protected TUN-associated DRG neurons from being damaged by cytotoxicity and apoptosis in a dose-dependent manner. Increased LDH and MDA leakage and proportion of TUNEL-positive cells, activation of caspase-3 and -9, increased Bcl-2 Assaciated X protein (Bax)/B cell lymphoma/lewkmia-2 (Bcl-2) ratio and mRNA levels of p53 Upregulated Modulator Of Apoptosis (PUMA) and DP5, and mitochondrial Cytochrome c release. Additionally, Iso down-regulated mRNA levels of ER stress genes, such as glucose-regulated protein 78 (GRP78), GRP94, C/EBP homologous protein (CHOP), and cleaved caspase-12 in TUN-induced DRG neurons. Moreover, Iso blocked the activation of three key branches of unfolded protein response in DRG neurons, including phosphorylation of pancreatic ER stress kinase (PERK), eukaryotic initiation factor 2 alpha (eIF2α), inositol-requiring enzyme 1α (IRE1α), and transcription factor 6 (ATF6).
  
  
  CONCLUSIONS
  Collectively, Iso prevented TUN-induced DRG neurons apoptosis by inhibiting mitochondrial and ER stress-associated apoptotic signaling cascades, suggesting that Iso possessed neuro-protective ability which could be beneficial in the prevention of ASCI.
authors.affiliation: Department of Orthopeadic Surgery, the First Affiliated Hospital of Xinxiang Medical University, Xinxiang 453100, Henan Province, China. Electronic address: lutanhn@163.com.
authors.forename: Tan
authors.initials: T
authors.lastname: Lu
authors.affiliation: Department of Orthopeadic Surgery, the First Affiliated Hospital of Xinxiang Medical University, Xinxiang 453100, Henan Province, China.
authors.forename: Chao
authors.initials: C
authors.lastname: Zhang
authors.affiliation: Department of Orthopeadic Surgery, the First Affiliated Hospital of Xinxiang Medical University, Xinxiang 453100, Henan Province, China.
authors.forename: Mingxiang
authors.initials: M
authors.lastname: Chai
authors.affiliation: Department of Orthopeadic Surgery, the First Affiliated Hospital of Xinxiang Medical University, Xinxiang 453100, Henan Province, China.
authors.forename: Yongbo
authors.initials: Y
authors.lastname: An
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics; D001549:Benzamides / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology / Q000627:therapeutic use*; D020169:Caspases / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects* / Q000235:genetics; D004789:Enzyme Activation / Q000187:drug effects; D005727:Ganglia, Spinal / Q000166:cytology*; D005786:Gene Expression Regulation / Q000187:drug effects; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D009474:Neurons / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D011794:Quercetin / Q000494:pharmacology*; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013830:Thiadiazoles / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology / Q000627:therapeutic use*; D014415:Tunicamycin / Q000633:toxicity*; D056811:Unfolded Protein Response / Q000187:drug effects / Q000235:genetics
chemical_list: D001549:Benzamides; D017382:Reactive Oxygen Species; D013830:Thiadiazoles; C000613727:YLT256; D014415:Tunicamycin; D045304:Cytochromes c; D011794:Quercetin; D007770:L-Lactate Dehydrogenase; D020169:Caspases
keywords: Apoptosis; DRG neurons; ER stress; Isoquercetin; Mitochondrial dysfunction