Atra For Lack Of Sugar Antiapoptotic Effect And Mechanism Of Hypoxia Pc12 Cell Injury

Background Vitamin A, iron and iodine deficiencies are identified asthe world’s three major nutritional deficiency diseases by the WHO andUNICEF. VAD widely exists all over the world, and pregnant women andchildren are the high-risk population of VAD.The neonatal hypoxicischemicbrain injury (HIBD) is the highly frequent and critical illness during theneonatal period, which results in neonatal central nervous system disordersand the main etiology of influence neonatal mortality and disability. Lots ofstudies have shown that the main pathogenesis of HIBD involves in thechanges of cerebral hemodynamics, energy metabolism failure of braincells, ischemia-reperfusion injury and activation of nitric oxide and freeradical production, intracellular calcium accumulation, neurotoxic effectsof excitatory amino acids, delayed neuronal death and apoptosis. Theapoptosis plays the key role of the HIBD pathological process. Ourprevious studies have demonstrated that the levels of VA in the HIBD ofneonatal pups are closely associated with the restoration of their neural functions, however, its molecular mechanism is still unknown.Objective To investigate in vitro the protective effect of ATRA on theapoptosis of PC12cells following OGD injury.Methods Using the oxygen and glucose-deprivation (OGD) model, thePC12cells were given the four different concentration (0.5,2,4and20μmol/L) of ATRA treatment. Meanwhile, the OGD injured cells were thecontrol group. The apoptosis ratio and the changes of mitochondrialmembrane potential in the PC12cells after OGD injury were analyzed withflow cytometry technique and staining of fluorescence probe JC-1. Theexpression levels of apoptotic genes expression in the mitochondrialapoptosis pathway were detected by Real-time PCR and Western blot,respectively.Results The PC12cells following OGD treatment were showed thecell body shrinked and refractive index declined under the microscopy. Thepercentage of apoptosis of the OGD injured PC12cells were significantlyincreased, and the mitochondrial membrane potential was statisticallydecreased compared with normal PC12cells. The treatment of2and4μmol/L ATRA obviously reduced the apoptosis rate of the PC12cells andmaintained the stability of the mitochondrial membrane potential comparedwith OGD cells, especially the best efficiency of4μmol/L ATRA treatment.The expression level of Bax apoptotic gene was significantlydown-regulation and Bcl-2anti-apoptotic gene up-regulation following 4μmol/L ATRA treatment. The changes of protein expression in Bax andBcl-2were consistent with the levels of mRNA expression.Conclusions The4μmol/L ATRA reduces apoptosis of OGD injuredPC12cells. The molecular mechamism is that ATRA regulatesmitochondrial apoptotic pathway through inducing Bcl-2increase and Baxdecrease to inhibit apoptosis of the damaged PC12cells. Background Apoptosis of neural cells is the important form duringthe HIBD pathological process. The expressional imbalance of Bcl-2protein family in the mitochondria apoptotic signaling pathway is thekeypoint, which leads to the extensive damage cellular stucture anddysfunction. The previous study has identified that the nomal level of VAin vivo more effectively enhances the recovery of neural cells function inthe HIBD rat. VAD reduced expression of RAR and decreased neuronalCa2+excitability to result in failure of learning-memory function. The dataof the front study has demonstrited that4μmol/L ATRA can inhibiteapoptosis of OGD injured PC12cells via regulating mitochondrialapoptotic signaling pathway. Our next study will focus on whether RAregulates RAR to play the role of anti-apoptosis.Objective To investigate the effects of RAR in the RA signalpathway on ATRA against apoptosis of PC12cells following OGD injury. Methods We constructed Ad-siRAR recombinant adenovirus byourselves to infect PC12cells for36h, and then detected level of RARmRNA expression to identify inhibition effeciency of Ad-siRAR byRT-PCR. Meanwhile, the empty adenovirus plasmid was the control ofsiRAR. After staining of Annexin V-PI and fluorescence probe JC-1,apoptosis rate and the mitochondrial membrane potential were detected byflow cytometry technique. The changes of expression level in Bax andBcl-2were tested by Real-time PCR and Western blot.Results The expression level of RAR gene decreased in the PC12cells following OGD treatment. And Ad-siRAR infection effectivelyinhibited expression of RAR in the PC12cells. The apoptotisis rate andmitochondrial membrane potential of OGD injured PC12cells bothsignificantly induced following infected Ad-SiRAR adenovirus. Inaddition, we also found that siRAR up-regulated Bax expression anddownregulated the expression of Bcl-2.Conclusions Ad-siRAR recommendient adenovirus significantlyinhibites the anti-apoptosis effect of4μmol/L ATRA on the PC12cellsfollowing OGD injury. These results reveal that ATRA enhances the Baxdecrease and Bcl-2increase in mitochondrial apoptotic singaling pathwaythrough regulating RAR expression to play the protective role ofanti-apoptosis in the OGD injured PC12cells.