| According to the Chinese Health Statistics Yearbook,the mortality rate of cerebrovascular diseases in China from 2003 to 2014 is on the rise.Also,according to the data from the sixth census in 2010,totally 837,300 urban residents and 1,023,400 rural residents died of cerebrovascular disease in 2014.In China,cerebral hemorrhage accounts for 20-40%of all cerebrovascular diseases.It is a clinical type of morbidity,disability and mortality.The occurrence of cerebrovascular diseases is closely linked to the Ryanodine receptors(RyRs)and the related proteins such as CAMRs,PPTA and PPTB.By affecting the conformation of RyR complex,these prote:ins activate or inhibit RyR mediated calcium release pathways,thereby regulating downstream pathways.When the pathway is affected,it leads to the change of the excitation-contraction coupling mechanism of muscle,thereby leading to abnormal vascular tension.As a result,the signal transduction of cardiomyocytes is blocked,and the symptoms such as intracerebral hemorrhage and abnormal blood vessel development are caused.Relevant studies have shown that moderate PPCS intervention may be helpftul to cerebrovascular diseases.However,excessive PPCS intervention can lead to abnormal regulation of related pathways in the body and cause sudden cerebral death,especially for patients with chronic cerebrovascular disease,people with abnormal vascular development and those with a family history of cerebrovascular disease.It is,therefore,important to identify the right and suitable intervention.In this study,the FNC_TG and C57 mice were used as experimental and control models,respectively,to explore the effects of excessive PPCS intervention on a number of physiological functions and calcium related proteins in mice.In this study,we divided the mice into the following four groups:wild type static group,transgenic static group,wild type PPCS intervention,and transgenic PPCS intervention.The related physiological indices were monitored by electrocardiogram,blood pressure,blood vessel tension and nuclear magnetic resonance in brain tissue of mice.Histopathological parameters were observed in the heart and brain of mice by HE staining,Congo red staining,lichen red staining,and so on.The ultrastructural changes of vascular development in mice were analyzed by electron microscope sections of the aorta,mesenteric artery and cerebral basal artery.The expression of MMPII and MMP 9 proteins of the MMPs family in the aorta was analyzed by immunohistochemical method.The expression of calcium associated proteins(CAMRs)PPTA and PPTB in the brain of mice was analyzed by Western Blot technique.The results are as follows:(1)Physiological monitoring indicators:ST-segment Elevation,ST-segmentDepression,Right Bundle-Branch Block and Arrhythmia were observed in transgenic mice,with aggravated symptoms after effects of exhaustive PPCS intervention.After exhaustive PPCS intervention in transgenic mice,there was a large amount of hemorrhage in the subarachnoid space of brain tissue of the transgenic mice.The results of nuclear magnetic resonance(NMR)SWI sequence showed the bleeding foci of the mice.The measurement of vascular tension showed that the mesenteric artery of transgenic mice had stronger response to contractile stimulation,but there was no significant difference.However,the diastolic stimulus response of TG-S group significantly deteriorated when compared with wild type(P<0.05).(2)Histopathological abnormality:Compared with wild-type and transgenic static mice,the cerebral cortex of mice in TG-EX group had a large number of bleeding points.Also,the area of cerebral surface hemorrhage in WT-S group,WT-EX group,TG-S group and TG-EX group was calculated by ImageJ software.The results showed that the brain surface bleeding area of mice in TG groups was higher than the wild-type groups.There was significant difference in cerebral surface area hermorrhage between TG-S group and WT-S group(P<0.01),and the area of cerebral surface hemorrhage in TG-EX group was significantly different from that in WT-EX group and TG-S group(P<0.001).The results of tissue staining and Congo red staining were similar.Amyloid plaques gathered near the focal point of cerebral hemorrhage in mice,which may be an indication of the presence of other neuropathies.The Lichen red staining showed that the elastic layer of artery and vessels in transgenic mice was discontinuous.An obvious fracture of elastic fibers was observed,which is consistent with the results of transmission electron microscopy(TEM).(3)Immunohistochemical staining results showed a weak positive expression of MMPII and MMPIX in the wall of the aorta and endothelium of wild type mice,but the expression of MMPII was slightly stronger than that of MMPIX.Furthermore,the expression of MMPII and MMPIX was obvious in the wall of the aorta and elastic fiber layer of transgenic mice.(4)The results of real-time quantitative qPCR showed that the expression of MMPIX in vascular tissues of transgenic mice was significantly up-regulated at the level of mRNA(P<0.001).(5)Western Blot results:Compared with the WT-S group,the expression of CAMR1 and CAMR2 proteins in the brain tissue of the TG-S group was up-regulated(P<0.001).In the TG-EX mice,the expression of CAMR1 was down-regulated compared with the TG-S group(P<0.05),whiles the expression of CAMR2 in TG-EX mice was up-regulated,but not statistically significant.Additionally,compared with the WT-EX group,the expression of CAMR1 protein in the brain tissue of TG-EX mice showed a highly significant up-regulation(P<0.001).The expression of CAMR2 protein was up-regulated in the brain tissue of TG-EX mice,but there was no significant difference in the expression of CAMR1 protein in the brain tissue of TG-EX mice.Compared with the WT-S group,the expression of PPTAA and PPTAB protein in the brain tissue of the TG-S group was significantly up-regulated.In the case of PPTAC protein,the expression in the brain tissue of the TG-S group was down-regulated.As expected,the expression of PPTAB and PPTAC protein was down-regulated in TG-EX mice,and the difference was highly significant(P<0.001),while the expression of PPTAA protein in TG-EX mice was up-regulated(P<0.001).Compared with TG-S group,the expression of PPTAB protein and PPTAC protein in TG-EX mice were down-regulated(P<0.001),whereas the expression of PPTAA protein was up-regulated in TG-EX mice.In the brain tissue of the TG-S group,the expression of PPTBA and PPTBB proteins was significantly higher than that in the WT-S group.Also,there was a very significant difference in both PPTBA and PPTBB proteins.Compared with WT-EX group,there was a down-regulation in the expression of both PPTBA and PPTBB proteins in TG-EX mice.The expression of PPTBA and PPTBB protein in TG-EX mice showed a significantly up-regulated trend compared with that in TG-S group.Conclusion:FNC_TG transgenic mice have arrhythmia,right ventricular block and other symptoms due to abnormal expression of calcium-associated protein.Exhaustive PPCS intervention has aggravated arrhythmia symptoms in transgenic mice,resulting in cardiac quenching.Therefore,after exhaustive PPCS intervention,there was a massive hemorrhage in brain tissue,resulting in sudden cerebral death.Also,a large amount of amyloid deposition in the cerebral vascular wall was observed.Calcium related proteins are involved in the regulation of cardiovascular pathology and vascular development in transgenic mice.The abnormal expression of calcium related protein leads to cerebrovascular aneurysms in the transgenic mice.The results show that screening for cardiovascular and cerebrovascular diseases among people with genetic defects require moderate intensity exercise intervention. |
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