| Objectives:The aims of this study were to investigate the cytoprotection of glycine (Gly) on thecerebral ischemia/reperfusion injury and the possible mechanisms, the studies areuseful to further explore the clinical value of glycine and to provide theorecicalreasons and new ideas to prevent cerebral ischemia/reperfusion injury.Methods:(1) This study used cerebral ischemia/reperfusion injury by right MCAO, A5-0monofilament surgical nylon suture with a heat-blunted tip was introduced into theleft internal carotid artery through the stump of the external carotid to the base of theright to stop blood flow. After90min, the filament was withdrawn to allow bloodreperfusion. Animals were divided into four groups, control animals received normalsaline solutions by intraperitoneal injection and treated animals received glycine at250mg/kg or1000mg/kg of body weight, which dissolved in saline solution. Micewere examined for neurological deficit before giving anesthesia,24h, and72h afterMCAO and reperfusion, mice were killed by decapitation24h or72h after MCAO.Brains infarct size was determined by means of saline solution containing2%2,3,5-triphenyl tetrazolium chloride (TTC) staining technique and a digital imagingsystem, in addition, the apoptotic neurons were analyzed by terminaldeoxynucleotidyl transferase biotin-dUTP nick end labeling(TUNEL) assay.(2) The right ischemic cortex was harvested and cellular proteins and RNA wereisolated for Western blot and real-time PCR. Western blot detected P-JNK, caspase-3,bcl-2, bax, FasL and real-time PCR detected bcl-2/bax, FasL changes between different groups. The purpose of these studies were to clarify the possible downstreampathways of glycine protective effects.(3) We used oxygen and glucose deprivation (OGD) reoxygenation, an in vitro modelthat best mimics in vivo cerebral ischemia reperfusion in SH-SY5Y cells as well as inprimary cultured neurons, MTT assay and LDH activity were performed to measurecell injury. SiRNA transfection and antibody against glycine receptor α1were used toclarify whether the glycine receptor α1was essential for the glycine protectiveeffects.Results:Our results showed that glycine could attenuate ischemia/reperfusion (I/R) inducedcerebral infarction and improved neurological outcomes in mice. The protective effectof glycine was associated with reduction of terminal deoxynucleotidyl transferasebiotin-dUTP nick end labeling (TUNEL) positive cells, inhibition of caspase-3cleavage, down-regulation of FasL/Fas, and up-regulation of bcl-2and bcl-2/bax inthe mouse I/R penumbra. The beneficial effect of glycine against oxygen and glucosedeprivation induced injury was also confirmed in SH-SY5Y cells as well as inprimary cultured neurons, which was significantly dampened by knockdown ofglycine receptora1with siRNA transfection or by preventing glycine binding withglycine receptor using a specific antibody against glycine receptor.Conclusions:These results suggest that glycine antagonize cerebral I/R induced injury by inhibitingapoptosis in mice and these glycine protective effects maybe mediated by glycinereceptor. |
