The Role And Mechanism Of Calcium Response Factor (CaRf) In Microglia Mediated Chronic Hypoperfusion White Matter Injury

Vascular cognitive impairment(VCI)has become the disease with the second highest incidence of dementia after Alzheimer disease(AD).With the registration of China's seventh census in December 2020,the total population has been close to zero growth.At the same time,due to the improvement of medical conditions and living standards,the current long-term trend of population aging will be irreversible,and actively responding to the problem of population aging has become a national strategy.Since VCI includes the entire process from mild cognitive impairment to dementia,understanding the pathogenesis of the disease and early intervention are effective means to prevent the progression of VCI.Insufficiency of cerebral perfusion is an important factor leading to VCI and dementia,which has a significant correlation with the severity of dementia,and leads to white matter hypertension(WMH)in the T2 sequence of magnetic resonance imaging(MRI).The existence and severity of these white matter changes,especially in the frontal lobe,are important determinants of cognitive function and dementia.However,the pathological mechanism of white matter damage caused by this hypoperfusion has not been fully elucidated,and there are no effective drugs to treat this process effectively.Therefore,in-depth study of the pathogenesis of this pathological process and finding an effective target for the treatment of white matter damage caused by chronic hypoperfusion will provide new hope for the treatment of VCI and severe dementia.Microglia are the innate immune cells of the central nervous system.Once the central nervous system is damaged,microglia are rapidly activated and participate in neuroinflammatory reactions together with immune cells such as lymphocytes and natural killer cells.In the animal model of chronic cerebral hypoperfusion,microglia activate and proliferate in the early stage of ischemia throughout the entire pathological process of chronic hypoperfusion.By removing microglia or inducing microglia to transform to an anti-inflammatory phenotype,it can promote the remyelination of the corpus callosum,increase the integrity of the white matter,and improve cognitive function,which indicates the regulation of the microglia phenotype Transformation can be used as an effective method to treat low-perfusion white matter injury.However,the specific reasons for the phenotypic changes of microglia are still not very clear.It is necessary to understand and find the key targets that can regulate the directional transformation of microglia.Calcium response factor(Ca RF)is a calcium-responsive protein with transcriptional function.It acts as a transcription factor in neurons,which is highly expressed in the brain,and is conserved in humans and animals.It has a differential expression trend during the phenotypic transformation of microglia.It can have a high expression pattern in M2 anti-inflammatory microglia activated by IL-4,and has the potential to regulate the phenotypic transformation of microglia.So,whether Ca RF can mediate chronic hypoperfusion white matter damage through microglia,there is no relevant research yet.This study uses a variety of experimental methods to verify the role of Ca RF in chronic hypoperfusion white matter injury models and WMH patients by regulating the phenotype of microglia to affect white matter injury and cognitive function.It make a solid foundation for the later study of the signal pathway of microglia phenotype,and also find a new path for the treatment of clinical VCI lesions or dementia.The main results of the research are as follows:1.The establishment of bilateral common carotid artery stenosis(BCAS)model and its damage to cognitive function and brain white matterA).The BCAS model has a good postoperative survival rate during inhalation anesthesia surgery;B).The anterior circulation cerebral blood flow(CBF)of immediately(57.73%±20.06)and 30 days(82.05%±7.9)after the operation were significantly lower than the preoperative CBF,showing chronic hypoperfusion;C).BCAS mice showed decreased learning ability and spatial memory impairment in behavioral experiments;D).BCAS mouse corpus callosum and striatum showed varying degrees of demyelination changes,and the expression of myelin basic protein(MBP)and myelin associated glycoprotein(MAG)decreased with time.2.The expression mode of Ca RF in the BCAS modelA).The expression of Ca RF decreased in microglia after BCAS.B).Microglia showed a significant pro-inflammatory phenotype on the 7th day,an anti-inflammatory phenotype on the 14th day,and a simple proliferation state on the 30th day of BCAS,with no significant changes in the phenotype.3.Conditional knockout of Ca RF in microglia aggravates the cognitive dysfunction and white matter demyelination of BCASA).Conditionally knocking out Ca RF in CX₃CR1 cells can cause memory disorder,loss of corpus callosum myelin sheath and reduction of MBP/MAG protein,and this damage is more significant in BCAS.B).Myelin ultrastructure is damaged in chronic hypoperfusion.C).Ca RF has a significant regulatory effect in the process of microglia activation and pro-inflammatory phenotypic transformation.4.Ca RF regulates microglia to protect oligodendrocytes and neuronsA).Ca RF regulates microglia to promote the differentiation and maturation of oligodendrocyte precursor cells(OPCs)and reduce the apoptosis of OPCs.B).The lack of Ca RF in microglia results in the reduction of synapses in the medial prefrontal cortex(m PFC),and the damage of synapses is further increased in BCAS5.Drug screening on microglia with Ca RF as the target and the protective effect of degarelix on BCASA).Using FDA-approved 1959 compounds to screen microglia,it was found that degarelix can increase the expression of Ca RF in microglia at both the RNA level and the protein level.B).Degarelix can improve the cognitive function of mice in BCAS,and the0.2mg dose group shows a better therapeutic effect in the process of learning and working memory.C).Degarelix can reduce the demyelination caused by chronic hypoperfusion injury,and protect the integrity of neuronal synapse protein and the number of dendritic spines.6.The diagnostic value of Ca RF in patients with chronic hypoperfusion white matter hyperintensityA).Compared with the control group,the proportion of age,hypertension,and MOCA scores are risk factors for WMH.B).The level of Ca RF in peripheral blood mononuclear macrophages in WMH patients was significantly down-regulated,and it was significantly negatively correlated with the expression of TNF-?and IL-1?,while it was significantly positively correlated with ARG-1.B).There is a significant negative correlation between the change of Ca RF expression and the degree of WMH,and Ca RF has the efficacy and advantages of alone for predicting and diagnosing.Conclusions:1.Ca RF participates in the process of chronic hypoperfusion white matter injury through microglia.The absence of Ca RF will promote the transformation of microglia into an inflammatory phenotype,resulting in demyelination of the corpus callosum and cognitive dysfunction.2.Ca RF has the ability to regulate microglia to promote the differentiation and maturation of OPCs and reduce the apoptosis of OPCs.The lack of Ca RF in microglia leads to the decrease of m PFC synapses,and the damage of synapses is further increased in the environment of BCAS.3.Degarelix can increase the expression of Ca RF in microglia,protect the corpus callosum from demyelination damage and the integrity of dendritic spines,and improve the cognitive function of BCAS mice.4.The expression of Ca RF in patients with chronic hypoperfusion and WMH was decreased,and Ca RF has the efficacy and advantages for predicting and diagnosing.