The Study On Expression Of Phosphodiesterase4D And Its Related Genes In Ischemic Stroke

Stroke, is also named as cerebral apoplexy, which is a common and frequent nerve system disease. Its disability rate and mortality are very high. It is one of the three major causes of death in human disease today, nearly accounts about10%of human diseases. There are two main types of stroke:cerebral hemorrhage and ischemia, ischemic stroke accounts for nearly85%. Stroke pathogenesis may be associated with heredity, calcium overload, oxidative stress, inflammation and any other factors. However, its mechanism is still unknown.PDE4(Phosphodiesterase4) is widely expressed in the body, PDE can degrade cAMP specifically. The second messenger cAMP can relax vascular smooth muscle, dilate blood vessels, promote nerve regeneration, transform abnormal cells and so on. It is also involved in enhancing the activity of intracellular respiratory chain oxidase, and improving myocardial ischemia. Then decrease of cAMP can induce the proliferation and migration of vascular smooth muscle cells and aggravate the local inflammatory reaction of damaged blood vessels. Beside that it may contribute to atherosclerosis and increase atheroma plaque. In normal cells PDE4D accounts approximately60%to70%of total activity.When the inflammation occurs, PDE4D activity is more than about80%. The study found that PDE4D SNP is associated with ischemic stroke.During the period of acute cerebral ischemia, the overload of intracellular Ca2+is one of most important factors that results in the death of neurons. Transient receptor potential channel1(TRPC1) is nonsecletive cation channel located on membrane.TRPC1is highly permeable to Ca+. It is also associated with central nervous system.TRPCl is a nonselective cation channels located on the cell membrane. Ischemic stroke can cause neuron necrosis involved in reactive oxygen species and inflammation. It can activate peripheral white blood cell and microglial cell, induce the cells to secrete amount of NO, reactive oxygen species and cytokines, such as hypoxia inducible factor la(HIF-la), tumor necrosis factor-a(TNF-a), and cyclooxygenase-2(COX-2) result in brain edema, cerebral hemorrhage and ultimately lead to neuronal death,Thioredoxin-1(Trx-1) is a low molecular weight protein of12kDa with highly conserved active sequence,-Cys-Gly-Pro-Cys-, regulates the balance of intracellular redox state, scavenges ROS, regulates activity of the transcription factors, inhibits apoptosis and provides the neurotrophic function. Thioredoxin-binding protein2(TBP-2) is a negative regulator of TRX and involved in redox regulation, the proliferation of cells and glucose and lipid metabolism. Forkheat protein3A (FOX03A) is a transcription factor regulating genes expressions related oxidative stress.We constructed a model of middle cerebral artery occlusion(MCAO) in mice and ischemia2h, collected peripheral blood and cortex tissues in mice. We checked the mRNA expressions of PDE4D、Trx、TBP-2、CREB、FOX03A、TRPC1、HIF-1α、COX-2、TNF-α by using the method of real-time quantitative PCR. We further compared these molecules expressions in peripheral blood between mice stroke patients and normal control. We explored the relationship between these factors and the pathogenesis of cerebral ischemia.The main results of this study are follows:(1) After ischemia for2h, PDE4D was increased in mice cortex, while the expression of PDE4D did not change in the blood. Neither in the blood nor in the cortex CREB changed. It is clear that during the period of acute cerebral ischemia PDE4D is increased in the cortex and reduced cAMP activity, induced the proliferation and migration of vascular smooth muscle cells and aggravated local inflammatory reaction of damaged blood vessels, contributed to atherosclerosis and increased atheroma plaque, finally aggravated ischemia damage.The expression of PDE4D was decreased, while CREB was increased in the peripheral blood of patients with ischemic stroke. The decreased PDE4D made cAMP increase, then increased the anti-inflammatory activity and reduced proliferation of vascular smooth muscle. This suggests the decreased or inhibited PDE4D has a protective role in ischemic stroke patients. The change of PDE4D is different between MCAO in mice model and patients, this needs to be further studied.(2) The mRNA of TRPC1was increased in the peripheral blood, cortex and in the peripheral blood of patients with ischemic stroke. It indicates that ischemia induce the influx of Ca2+, overload of Ca2+and neuronal death. This process, May be mediated by increase of TRPC1receptor expression.(3)The mRNA level of Trx was increased in the peripheral blood and cortex, while mRNA levels of TBP-2and FOX03A did not change. The mRNA levels of Trx-1and FOXO3A were increased in the peripheral blood of patients with ischemic stroke, while TBP-2was decreased. Trx increase suggests that the oxidative stress happenes in the body after ischemia. The change of Trx is same at the MACO mice model and ptients, thus Trx is a good target for studying ischemia. (4) The mRNA level of HIF-1α was increased in the peripheral blood and cortex. TNF-α was increased in the blood but it did not change in the cortex. The mRNA level of COX-2was increased in the cortex and but it did not change in the peripheral blood. The mRNA levels of HIF-1α and COX-2were increased while in the peripheral blood in patients with ischemic stroke, but TNF-α did not change. It suggests that hypoxia and a series of inflammatory cascades happen in the body. Although the expressions of HIF-α, COX-2mRNA andTNF-α are different, HIF-1α and COX-2are also taken as molecules for evaluating ischemia.Conclusion:This study compared difference of the mRNA levels of PDE4D, Trx, TBP-2, CREB, FOX03A, TRPC1, HIF-1α, COX-2and TNF-α in the peripheral blood and cortex in MACO mice model and peripheral blood of patients with ischemic stroke. This study provides basic theory with diagnose and treatment on ischemic stroke.