| Stroke,a common serious CNS disease with high disability and high mortality,is caused by sunden defect of cerebral blood flow and seriously affects the quality of patients' life.Systematic researches of pathological mechanism of stroke,particularly ischemic stroke,are urgently required to promote the clinical investigation to improve prevention and treatment of the disease.Microglia is one kind of the most important neural cells and widely distributed in the central nervous system(CNS).as homogenous partner of macrophage,it serves as the main immune cell in CNS,which plays an extremely crucial role in the maintenance of CNS homeostasis as well as the repairment.The functional behaviors of activated microglia by stroke,such as proliferation,migration,ingestion,synthesis and secretion of inflammation and inflammatory factors,are directly related to the progress and prognosis of stroke and the factors affecting the functional status of microglia will impact the pathological development and the prognosis of the disease.Aquaporins(AQPs)are membrane channel protein,which are widely located on the plasma membrane of prokaryotic and eukaryotic cells,and specifically mediate water and some small molecues transporting across membrane.The accumulated evidences showed that AQPs-mediated water transporting were involved in many important life events,such as cell migration,differentiation,material transport and signal transduction,which widely participate in a variety of physiological and pathological processes.As a member of the aquaporin family,AQP3,one of the aquaglyceroporins,can efficiently transport some small size molecules such as glycerol besides water.Our previous results have showed that AQP3 was expressed in macrophage and deletion of AQP3 significantly decreases the capability of macrophage to ingest and phagocytose foreign matters,consequently,greatly reducing animal resistance to infection.Mechanism study showed that,compared to that of the wildtype counterpart,the migration and quick deformation of AQP3-/-macroghage were undermined and the intracellular ATP concentration reduced,which meant the impairment of ingesting and phagocytosing of AQP3-/-macrophage may be attributed to disability of morphology alteration and energy metabolism resulting from AQP3 gene knock-out.So far,only very limited literatures reported the expression of AQP3 in brain tissues,mainly in Cerebro-Spinal-fluid-secreting related Choroid plexus.As for the functions of AQP3 in CNS,there was a complete blank.Based on the facts: the expression of AQP3 in primary microglia cells was detected in our previous studies;the similarity between microglia and macrophage in origin and functions;the crucial protective role of microglia in stroke,wehypothesized that AQP3 gene deletion may play a vital role in the development and prognosis of stroke.In this study,using AQP3 gene knockout mice,we roughly investigated the functions and underlying mechanisms of AQP3 in ischemic stroke.Firstly,by RT-PCR,Western Blot methods,we confirmed the expression of AQP3 in mouse brain tissue.Then,we first found the expression at high level of AQP3 in primary microglia through RT-PCR,Western blot and immunofluorescence analysis.We have established AQP3+/+ and AQP3-/-ischemic stroke mouse models separately according to the classical MACO protocol and evaluated neurological deficit by angle holding test and histopathological structure analysis by TTC dye.The results showed that AQP3-knock-out ischemic stroke mice attained lower neurological scores and have larger infarct volume in brain tissue(holding angle test analysis: 2.0±0.67 VS.2.6±0.65(AQP3-/-VS.AQP3+/+);TTC staining: 41% VS.31%(AQP3-/-VS.AQP3+/+)).The mechanism analysis indicated a similarity in proliferation rate of both genotypes(AQP3+/+and AQP3-/-)of microglia,but a significant difference in migration : the movement rate of AQP3-/-microglia retarded much behind wt microglia under each condition(wound healing test: 0.83±0.025 vs.0.46± 0.031 and transwell assay: 19.3 ± 1.70 vs.12.3 ± 1.34).Also,Intracellular content of ATP in AQP3-/-microglia was much lower than wt microglia(120.1±10.29 vs.229.8±11.68).The evidences in our study strongly indicated a possibility that AQP3 gene knock-out could undermine the ability of microglia quickly moving to lesions area and interfering with normal energy supply,therefore aggravating impairment and malignant outcome of ischemic stroke.More systematical researches are still needed.Our study assured the involvement of protective roles of aquaglyceroporin AQP3 in pathological process of ischemic stroke and roughly revealed the possible mechanism.The results of this study is helpful for fully understanding the pathological mechanism of ischemic stroke,paving a new way in clinical trial experiments of this disease and also giving new insightful suggestions to reveal pathological mechanism of other neural degenerative diseases related to dysfunction of microglia,such as Parkinson's disease,Multiple Sclerosis,Alzheimer disease. |
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