Effects Of Simulated Microgravity On Cardiovascular And Mechanisms Research

Microgravity/simulated microgravity can induce cardiovascular deconditioning, the structural remodeling and functional changes of heart and arterial vascular smooth muscle are the important mechanisms responsible for the cardiovascular dysfunction. However, until now, there is not any research report about cardiovascular dysfunction after long-time simulated microgravity on human in China. Moreover, the mechanisms of structural remodeling and functional changes in heart and arterial vascular smooth muscle induced by microgravity/simulated microgravity aren't fully elucidated. Thus, the objective of this study is following:(1) Acquire the cardiovascular experimental data from Chinese volunteers by 60d HDT and evaluate the countermeasure effects of Vibration and Chinese medicine"TaiKong Yangxin Prescription".(2) Illustrate the molecular mechanisms of cardiac atrophy induced by simulated microgravity.(3) Illustrate the regulating mechanisms of changes in contractile response of hindlimb arterial vascualrs after simulated microgravity.(4) Study the effects of microgravity and simulated microgravity on VSMC proliferation and theβ-catenin signal pathway. The following methods will be used in our experiments.(1) 60d head down bedrest was used to simulate the physiological effects of microgravity. The echography was used to measure the structure and functions of heart and vascular. The strain gauge venous occlusion plethysmography was used to assess limb venous vascular characteristics. Moreover, the protectable effects of two kinds of countermeasures were evaluated.(2) Tai-suspension rats were used animal model of simulated microgravity, TaiKong Yangxin Prescription was used to countermeasure the cardiac dysfunction. The isolated rat heart functions were measured by Langendorff method. The cardiac muscle cells were stained by FITC-Lectin and the cross section area were calculated. Western blot was used to detect the protein expression of TnI, Bcl-2, Hsp20 and the phosphorylation level of Akt, GSK3-β, FAK, P38 MAPK and Hsp27.(3) The contractile responses of femoral and abdomen arteries to NE or PE were measured by powerlab recording system. By the special inhibitors, PD98059, ML-7, LY-294002, SB203580, Nifedipine and Cytochalasin D, respectively for ERK/CaD, MLCK, PI3K, P38 MAPK, L type calcium channel and actin polymerization, the contributing role of these special pathways to changes of contractile responses after simulated microgravity were evaluated.(4) In clinostat and"Shenzhou 7"spacecraft, the VSMCs were cultured for 48h, the proliferation was measured. Western blot and immunofluorescence were used to detect the protein expression ofβ-catenin,N-Cadherin,Cyclin D1,P21Cip1 and the phosphorylation level of Akt and GSK-3βafter simulated microgravity by clinostat. Immunofluorescence was used to detect changes of nucleus in VSMCs and the expression ofβ-catenin,N-Cadherin,Cyclin D1,P21Cip1. From the experiments, the following results were acquired. 1. HDT experiment(1) 58d HDT induced the cardiac atrophy and decreased the LDVD and SV. The decrease of cardiac mass and functions can be protected partly by countermeasures of vibration and TaiKong Yangxin Prescription. Moreover, TaiKong Yangxin Prescription had better countermeasure effects than vibration did.(2) 58d HDT had no effect on the diameters of carotid and femoral arteries, but can decrease the cross section area of portal vein in three groups. Especially, the decreased range in control group reached the statistical difference.(3) 58d HDT affected the contractile functions of vascular. The contractile speeds in three groups all decreased. Except carotid artery, the contractile speed of pulmonary artery significantly decreased in vibration and TaiKong Yangxin Prescription group. The contractile speed of femoral artery in three groups decreased significantly compared with those of before HDT. Moreover, there was a decreased trend of femoral arteries RI in control and vibration groups, but in TaiKong Yangxin Prescription group, femoral arteries RI were increased slightly. there was a increased trend of Tibial arteries RI in control and TaiKong Yangxin Prescription group, but in vibration group, there was a decreased trend compared with that of before HDT.(4) 38d HDT induced the significant decrease of leg vein compliance, but compared with control and vibration groups, the decrease range in TaiKong Yangxin Prescription group was obviously smaller, which showed that TaiKong Yangxin Prescription has better countermeasure effect on the decreased leg vein compliance induced by 38d HDT. After 52d HDT, the leg vein compliance in TaiKong Yangxin Prescription group and control group had a ongoing decrease, but the decreased range in vibration group reduced to the level of below 38d HDT, which showed that vibration countermeasure had a better effects on the decreased leg vein compliance after 52d HDT.(5) HDT increased the leg venous resistance. After 38d HDT, the increased range of leg venous resistance in control and vibration groups were higher than that of TaiKong Yangxin Prescription group, which showed that TaiKong Yangxin Prescription had a better countermeasure effect compared with vibration. However, after 52d HDT, the leg venous resistance of control and TaiKong Yangxin Prescription group had a increased trend, but the increased range in vibration reduced to the level below that of 38d HDT, which showed that vibration had better countermeasure on the increased leg venous resistance than that of TaiKong Yangxin Prescription after 52d HDT.2. Effects of simulated microgravity on cardiac structure and function and countermeasure study(1) Simulated microgravity induced by 7d tail-suspension decreased the rat cardiac functions which can be countermeasured by TaiKong Yangxin Prescription.(2) Simulated microgravity after 7d tail-suspension induced the rat cardiac atrophy, decreased the protein expression of TnI and inhibited the Akt/GSK-3βsignal pathway, which can be counter- measured by TaiKong Yangxin Prescription.(3) The protein expression of TnI was decreased ongoing after 7d, 14d and 21d tail-suspension. The Akt/GSK-3βsignal pathway significantly inhibited by 7d and 21d tail-suspension, but increased by 14d tail-suspension.(4) Simulated microgravity induced by 14d tail-suspension decreased significantly the signal of P38 MAPK/Hsp27 pathway in cardiac muscle.3. Effects of simulated microgravity on vascular function and mechanisms study(1) Simulated microgravity decreased the contractile response of femoral arteries and inhibited the signal of ERK/CaD pathway.(2) 7d tail-suspension had little effects on signal of MLCK/MLC20 pathway in femoral arteries, however, 14d tail-suspension inhibited significantly the signal of MLCK/MLC20 pathway.(3) The function of L type calcium channel decreased significantly after 7d and 14d tail-suspension.(4) 14d tail-suspension decreased the contractile response of abdomen arteries and inhibited the signal of P38 MAPK/Hsp27 pathway, but had little effects on the P38 MAPK/Hsp27 pathway in femoral arteries.(5) The signal of PI3K pathway decreased significantly after 7d and 14d tail-suspension.(6) Simulated microgravity induced by tail-suspension had no effect on the actin polymerization in femoral arteries.4. Effects of microgravity and simulated microgravity on VSMC proliferation andβ-catenin signal pathway(1) Microgravity and simulated microgravity decreased the VSMC proliferation, in whichβ-catenin signal pathway had important role.(2) Microgravity and simulated microgravity inhibited the expression ofβ-catenin and N-Cadherin. Simulated microgravity decreased the activity of Akt and GSK-3β.(3) Microgravity and simulated microgravity inhibited theβ-catenin signal pathway,but the direct reason which decreased the VSMC proliferation was the reduced expression of Cyclin D1, cell cycle retarded at G1 and different expression of P21Cip1. For the first time, we found that microgravity and simulated microgravity had different regulating mode on protein expression of P21 Cip1.This study got following conclusions.(1) For the first time, we found that 60d HDT can induce cardiac atrophy, functional inhibition, decreased arterial contractility, decreased leg vein compliance and increased leg venous flow resistance in Chinese volunteers. The countermeasures of TaiKong Yangxin Prescription and vibration has partly protectable effects on cardiac dysfunction induced by 60d HDT.(2) Simulated microgravity after tail-suspension can induce the cardiac atrophy by Akt/GSK-3βsignal pathway which has important role in regulating cardiac hypertrophy. TaiKong Yangxin Prescription can inhibit cardiac atrophy by activating the signal of Akt/GSK-3βpathway. It can also increase the protein expression of Hsp20 and promote its cardioprotection effects. Moreover, simulated microgravity can also induced cardiac atrophy by inhibiting the signal pathway of P38 MAPK/Hsp27.(3) Simulated microgravity after tail-suspension decreased the contractile responses of femoral and abdomen arteries. In femoral arteries, the functions of ERK/CaD, MLCK/MLC20, PI3K signal pathways and L type calcium channel decreased by tail-suspension. In abdomen arteries, tail-suspension inhibited the signal of P38 MAPK/Hsp27 pathway.(4) For the first time, this experiment found microgravity can decrease the VSMC proliferation. Microgravity and simulated microgravity inhibited the protein expression ofβ-catenin and N-Cadherin. Simulated microgravity can decreased the activity of Akt/GSK-3βsignal pathway. The direct reason which decreased the VSMC proliferation was the reduced expression of Cyclin D1, cell cycle retarded at G1 and different expression of P21Cip1. For the first time, we found that microgravity and simulated microgravity had different regulating mode on protein expression of P21 Cip1.