ABSTRACT
STUDY DESIGN: In vitro studies of the role of 17ß-estradiol (E2) and its possible targets in intervertebral disc degeneration (IDD). OBJECTIVE: To define the regulatory role of E2 in IDD and the potential mechanisms. SUMMARY OF BACKGROUND DATA: IDD has intricate etiology that is influenced by multiple risk factors. However, the underlying molecular mechanisms of occurrence and progression of IDD are not well elucidated. The degradation of extracellular matrix (ECM) has been extensively observed in IDD. E2 was found to inhibit ECM degradation in human nuleus pulposus cells (HNPCs), but the molecular mechanism remained to be determined. METHODS: Western blot and qPCR was performed to quantify the expression of target proteins in HNPCs. Luciferase reporter gene assay was applied to detect the effects of E2 and forkhead box O-3 (FOXO3) on matrix metalloproteinases (MMP)-3 promoter activity. Chromatin immunoprecipitation assay analyzed the binding of FOXO3 to MMP-3 and the effect of E2 on this process. RESULTS: We identified the upregulation of collagen II and aggrecan by E2 independent of time and concentration. And E2 downregulated MMP-3 expression in human nucleus pulposus cells. The phosphorylation of FOXO3 led to the reduction of MMP-3 promoter activity. Furthermore, 17ß-estradiol-induced the activation of PI3K/Akt pathway is required for FOXO3 phosphorylated. CONCLUSION: E2 prevents the degradation of ECM by upregulating collagen II and aggrecan expression via reducing MMP-3 expression in HNPCs, and PI3K/Akt/FOXO3 pathway is dispensable for MMP-3 downregulated. Therefore, E2 protects against IDD by preventing ECM degradation. LEVEL OF EVIDENCE: 3.
Subject(s)
Estradiol/pharmacology , Extracellular Matrix/drug effects , Extracellular Matrix/metabolism , Forkhead Box Protein O3/metabolism , Matrix Metalloproteinase 3/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Aggrecans/genetics , Down-Regulation/drug effects , Female , Humans , Intervertebral Disc Degeneration/drug therapy , Intervertebral Disc Degeneration/metabolism , Nucleus Pulposus/metabolism , Phosphorylation , Protective Agents/pharmacology , Up-Regulation/drug effectsABSTRACT
OBJECTIVE: Cervical spondylotic myelopathy (CSM) is the most common cause of spinal cord dysfunction. Our study aims to explore the correlation of osteoprotegerin (OPG) gene polymorphisms and the risk factors and severity of CSM. PATIENTS AND METHODS: The peripheral blood samples from 494 CSM patients and 515 healthy individuals were collected for detecting the 950T/C, 1181G/C and 163A/G genotypes and genetic equilibrium of OPG in the CSM and control groups and analyzing the genotype distribution and allele frequency. The severity of CSM and the impaired segments were evaluated by the Japanese Orthopedic Association (JOA) scoring combined with cervical magnetic resonance imaging (MRI), in order to investigate the relations between the three genotypes of OPG promoter gene loci (950T/C, 163A/G and 1181G/C) and occurrence as well as severity of CSM. RESULTS: The risk rate of TC genotype carrier suffered from CSM was 0.46, of TT genotype carrier was 0.27. The risk rate of T allele carrier suffered from CSM was 0.37. In 950T/C single nucleotide polymorphism (SNP), patients with TC, TT and T genotypes had lower risk to suffer from CSM. CONCLUSION: Taken together, OPG 950T/C SNP protects against CSM, and it is correlated with the severity of CSM, providing a new idea for the prevention and treatment of CSM.
Subject(s)
Asian People/genetics , Genetic Predisposition to Disease/genetics , Osteoprotegerin/genetics , Polymorphism, Single Nucleotide/genetics , Spinal Cord Diseases/genetics , Spondylosis/genetics , Adult , Cervical Vertebrae/diagnostic imaging , Female , Genetic Predisposition to Disease/epidemiology , Humans , Male , Middle Aged , Population Surveillance/methods , Spinal Cord Diseases/diagnostic imaging , Spinal Cord Diseases/epidemiology , Spondylosis/diagnostic imaging , Spondylosis/epidemiology , Young AdultABSTRACT
AIM: To determine how the relative amino acid contents and metabolic pathways regulate the pharmacological phenotypes in rats with cerebral ischemia after treatment with varying doses of DanHong injection (DHI). METHODS: Adult male rats underwent middle cerebral artery occlusion (MCAO), and were injected with DHI (DH-1: 1 mL/kg; DH-2: 2.5 mL/kg; DH-3: 5 mL/kg, and DH-4: 10 mL/kg, iv) daily for 3 d. The neurological deficit score, body weights and infarct volume were assessed. Serum levels of 20 free amino acids were determined using HPLC, and the values were transformed through the quantitative analysis of the amino acids in the serum metabolic spectrum. Multivariate statistical analysis methods (PCA and PLS-DA) and web-based metabolomics tools (MetPa and MetaboAnalyst) were used to analyze the biological data sets for the amino acids. RESULTS: Administration of DHI dose-dependently decreased cerebral infarct volume, and ameliorated neurological deficits. A total of 5, 6, 7 and 7 non-overlapping metabolites were identified in the DH-1, DH-2, DH-3, and DH-4 groups, respectively. Eight metabolites were shared between the DHI groups and the vehicle group. In addition, the serum levels of glutamic acid, aspartic acid and serine increased with increasing DHI dose. A total of 3, 2, 2 and 5 non-overlapping metabolic pathways were identified in the DH-1, DH-2, DH-3 and DH-4 groups, respectively, and glycine, serine, threonine and histidine metabolism were identified as overlapping pathways among the 4 dose groups. CONCLUSION: Overlapping and non-overlapping amino acid metabolites and metabolic pathways are associated with the dose-dependent neuroprotective effect of DHI.
