Neural stem cell(NSC)transplantation is a promising strategy for replacing lost neurons following spinal cord injury.However,the survival and differentiation of transplanted NSCs is limited,possibly owing to the neuro...Neural stem cell(NSC)transplantation is a promising strategy for replacing lost neurons following spinal cord injury.However,the survival and differentiation of transplanted NSCs is limited,possibly owing to the neurotoxic inflammatory microenvironment.Because of the important role of glucose metabolism in M1/M2 polarization of microglia/macrophages,we hypothesized that altering the phenotype of microglia/macrophages by regulating the activity of aldose reductase(AR),a key enzyme in the polyol pathway of glucose metabolism,would provide a more beneficial microenvironment for NSC survival and differentiation.Here,we reveal that inhibition of host AR promoted the polarization of microglia/macrophages toward the M2 phenotype in lesioned spinal cord injuries.M2 macrophages promoted the differentiation of NSCs into neurons in vitro.Transplantation of NSCs into injured spinal cords either deficient in AR or treated with the AR inhibitor sorbinil promoted the survival and neuronal differentiation of NSCs at the injured spinal cord site and contributed to locomotor functional recovery.Our findings suggest that inhibition of host AR activity is beneficial in enhancing the survival and neuronal differentiation of transplanted NSCs and shows potential as a treatment of spinal cord injury.展开更多
Objectives To establish a new transgenic mouse model for determining the function and role of human scavenger receptor A (SR-A) in atherosclerosis in vivo.Methods Human scavenger receptor minigene-driven mouse tie-1...Objectives To establish a new transgenic mouse model for determining the function and role of human scavenger receptor A (SR-A) in atherosclerosis in vivo.Methods Human scavenger receptor minigene-driven mouse tie-1 promoter was constructed and confirmed by endonuclease digestion and sequence analysis. Transgenic mice were generated via the microinjection method. PCR and Southern blot were used to screen the positive transgenic mice. RT-PCR and immunohistochemical analysis were used to detect the level and location of human SR-A Ⅰ expression in transgenic mice. The activity of human SR-A Ⅰ was determined by morphologic observation of aortic endothelial cells of transgenic mice under transmission electron microscopy.Results The electrophoresis assay showed the expected 4 fragments of 0.9 kb, 1.1 kb, 1.2 kb and 4.2 kb in the Sma Ⅰ digest and 2 fragments of 0.8 kb and 6.7 kb in Bgl Ⅱ digest of plasmids pTie-1/hSR-A. The fragment sequence of tie-1 promoter and human SR-A cDNA in plasmids pTie-1/hSR-A was correct and no ATG before the translation initiation sites of human SR-A was found by sequence analysis. 561 injected and surviving embryos with the purified human SR-A minigene were implanted into the oviducts of 19 ICR pseudopregnant mice. Among the 54 surviving pups from 13 foster mothers, 7 were identified by PCR and Southern blot analysis. The results of RT-PCR and imrnunohistochemical analysis showed human SR-A was specifically expressed on vascular endothelial cells of the aorta and renal artery, as well as hepatic sinusoidal endothelial cells in transgenic mice. Transmmion electron microscope (TEM) of aorta of transgenic mice showed that a large number of vesicles, multivesicle bodies and swollen mitochondria filled the plasma of endothelial cells.Conclusions A transgenic mouse model with overexpression of human SR-A in endothelial cells was successfully established. The transgene was integrated and transmitted into the chromosome of transgenic mice. Tie-1 promoter controlled the transgene to express in endothelial cells in mice. Pinocytic activity of aortic endothelial cells in transgenic mice was higher than that of C57BL/6J mice. Our studies will provide a new transcgenic model for investigation of atherosclerosis and functions of human SR-A.展开更多
Ischemia occurs in diabetic retinopathy with neuronal loss, edema, glial cell reactivity and oxidative stress. Epacs, consisting of Epac 1 and Epac2, are cAMP mediators playing important roles in maintenance of endoth...Ischemia occurs in diabetic retinopathy with neuronal loss, edema, glial cell reactivity and oxidative stress. Epacs, consisting of Epac 1 and Epac2, are cAMP mediators playing important roles in maintenance of endothelial barrier and neuronal functions To investigate the roles of Epacs in the pathogenesis of ischemic retinopathy, transient middle cerebral artery occlusion （tMCAO） was performed on Epacl-deficient （Epacl-/- ） mice, Epac2-deficient （Epac2-/-） mice, and their wild type counter-parts （Epacl＋/＋ and Epac2＋/＋）. Two-hour occlusion and 22-hour reperfusion were conducted to induce ischemia/reperfusion injury to the retina. After tMCAO, the contralateral retinae displayed similar morphology between different genotypes. Neu-ronal loss, retinal edema and increase in immunoreactivity for aquaporin 4 （AQP4）, glial fibrillary acidic protein （GFAP）, peroxiredoxin 6 （Prx6） were observed in ipsilateral retinae. Epac2 / ipsilateral retinae showed more neuronal loss in retinal ganglion cell layer, increased retinal thickness and stronger immunostaining of AQP4, GFAP, and Prx6 than those of Epac2＋/＋. However, Epacl-/- ipsilateral retinae displayed similar pathology as those in Epacl＋/＋ mice. Our observations suggest that Epac2-deficiency led to more severe ischemic retinopathy after retinal ischemia/reperfusion injury.展开更多
基金supported by the National Natural Science Foundation of China,Nos.81601056(to KZ),81901252(to QZ)Shaanxi Key Research and Development Program of China,No.2020SF-083(to KZ)+1 种基金Sanming Project of Medicine in Shenzhen of China,No.SZSM201911011(to SXW)the Key Laboratory of Spine and Spinal Cord Injury Repair and Regeneration(Tongji University,Ministry of Education)of China(to KZ).
文摘Neural stem cell(NSC)transplantation is a promising strategy for replacing lost neurons following spinal cord injury.However,the survival and differentiation of transplanted NSCs is limited,possibly owing to the neurotoxic inflammatory microenvironment.Because of the important role of glucose metabolism in M1/M2 polarization of microglia/macrophages,we hypothesized that altering the phenotype of microglia/macrophages by regulating the activity of aldose reductase(AR),a key enzyme in the polyol pathway of glucose metabolism,would provide a more beneficial microenvironment for NSC survival and differentiation.Here,we reveal that inhibition of host AR promoted the polarization of microglia/macrophages toward the M2 phenotype in lesioned spinal cord injuries.M2 macrophages promoted the differentiation of NSCs into neurons in vitro.Transplantation of NSCs into injured spinal cords either deficient in AR or treated with the AR inhibitor sorbinil promoted the survival and neuronal differentiation of NSCs at the injured spinal cord site and contributed to locomotor functional recovery.Our findings suggest that inhibition of host AR activity is beneficial in enhancing the survival and neuronal differentiation of transplanted NSCs and shows potential as a treatment of spinal cord injury.
基金ThisprojectwassupportedbytheNationalNaturalScienceFoundationofChina (No 3990 0 0 6 1)
文摘Objectives To establish a new transgenic mouse model for determining the function and role of human scavenger receptor A (SR-A) in atherosclerosis in vivo.Methods Human scavenger receptor minigene-driven mouse tie-1 promoter was constructed and confirmed by endonuclease digestion and sequence analysis. Transgenic mice were generated via the microinjection method. PCR and Southern blot were used to screen the positive transgenic mice. RT-PCR and immunohistochemical analysis were used to detect the level and location of human SR-A Ⅰ expression in transgenic mice. The activity of human SR-A Ⅰ was determined by morphologic observation of aortic endothelial cells of transgenic mice under transmission electron microscopy.Results The electrophoresis assay showed the expected 4 fragments of 0.9 kb, 1.1 kb, 1.2 kb and 4.2 kb in the Sma Ⅰ digest and 2 fragments of 0.8 kb and 6.7 kb in Bgl Ⅱ digest of plasmids pTie-1/hSR-A. The fragment sequence of tie-1 promoter and human SR-A cDNA in plasmids pTie-1/hSR-A was correct and no ATG before the translation initiation sites of human SR-A was found by sequence analysis. 561 injected and surviving embryos with the purified human SR-A minigene were implanted into the oviducts of 19 ICR pseudopregnant mice. Among the 54 surviving pups from 13 foster mothers, 7 were identified by PCR and Southern blot analysis. The results of RT-PCR and imrnunohistochemical analysis showed human SR-A was specifically expressed on vascular endothelial cells of the aorta and renal artery, as well as hepatic sinusoidal endothelial cells in transgenic mice. Transmmion electron microscope (TEM) of aorta of transgenic mice showed that a large number of vesicles, multivesicle bodies and swollen mitochondria filled the plasma of endothelial cells.Conclusions A transgenic mouse model with overexpression of human SR-A in endothelial cells was successfully established. The transgene was integrated and transmitted into the chromosome of transgenic mice. Tie-1 promoter controlled the transgene to express in endothelial cells in mice. Pinocytic activity of aortic endothelial cells in transgenic mice was higher than that of C57BL/6J mice. Our studies will provide a new transcgenic model for investigation of atherosclerosis and functions of human SR-A.
基金supported by the Research Grants Council of Hong Kong(RGC)HKU 764008M to Sookja Kim Chung
文摘Ischemia occurs in diabetic retinopathy with neuronal loss, edema, glial cell reactivity and oxidative stress. Epacs, consisting of Epac 1 and Epac2, are cAMP mediators playing important roles in maintenance of endothelial barrier and neuronal functions To investigate the roles of Epacs in the pathogenesis of ischemic retinopathy, transient middle cerebral artery occlusion （tMCAO） was performed on Epacl-deficient （Epacl-/- ） mice, Epac2-deficient （Epac2-/-） mice, and their wild type counter-parts （Epacl＋/＋ and Epac2＋/＋）. Two-hour occlusion and 22-hour reperfusion were conducted to induce ischemia/reperfusion injury to the retina. After tMCAO, the contralateral retinae displayed similar morphology between different genotypes. Neu-ronal loss, retinal edema and increase in immunoreactivity for aquaporin 4 （AQP4）, glial fibrillary acidic protein （GFAP）, peroxiredoxin 6 （Prx6） were observed in ipsilateral retinae. Epac2 / ipsilateral retinae showed more neuronal loss in retinal ganglion cell layer, increased retinal thickness and stronger immunostaining of AQP4, GFAP, and Prx6 than those of Epac2＋/＋. However, Epacl-/- ipsilateral retinae displayed similar pathology as those in Epacl＋/＋ mice. Our observations suggest that Epac2-deficiency led to more severe ischemic retinopathy after retinal ischemia/reperfusion injury.
