AIM： To explore the effects of ultrasound exposure combined with microbubble contrast agent （SonoVue） on the permeability of the cellular membrane and on the expression of plasrnid DNA encoding enhanced green fluor...AIM： To explore the effects of ultrasound exposure combined with microbubble contrast agent （SonoVue） on the permeability of the cellular membrane and on the expression of plasrnid DNA encoding enhanced green fluorescent protein （pEGFP） transfer into human umbilical vein endothelial cells （HUVECs）. METHODS： HUVECs with fluorescein isothiocyanatedextran （FD500） and HUVECs with pEGFP were exposed to continuous wave （1.9 MHz, 80.0 mW/cm^2） for 5 min, with or without a SonoVue. The percentage of FD500 taken by the HUVECs and the transient expression rate of pEGFP in the HUVECs were examined by fluorescence microscopy and flow cytornetry, respectively. RESULTS： The percentage of FDS00-positive HUVECs in the group of ultrasound exposure combined with SonoVue was significantly higher than that of the group of ultrasound exposure alone （24.0%± 5.5% vs 66.6% ± 4.1%, P 〈 0.001）. Compared with the group of ultrasound exposure alone, the transfection expression rate of pEGFP in HUVECs was markedly increased with the addition of SonoVue （16.1% ± 1.9% vs 1.5% ± 0.2%, P 〈 0.001）. No statistical significant difference was observed in the HUVECs survival rates between the ultrasound group with and without the addition of SonoVue （94.1% ± 2.3% vs 91.1% ± 4.1% ）. CONCLUSION： The cell membrane permeability of HUVECs and the transfection efficiency of pEGFP into HUVECs exposed to ultrasound are significantly increased after addition of an ultrasound contrast agent without obvious damage to the survival of HUVECs. This non- invasive gene transfer method may be a useful tool for clinical gene therapy of hepatic tumors.展开更多
Voltage-dependent anion channel （VDAC）I is the main channel of the mitochondrial outer membrane （MOM） and it has been proposed to be part of the permeability transition pore （PTP）, a putative multiprotein comple...Voltage-dependent anion channel （VDAC）I is the main channel of the mitochondrial outer membrane （MOM） and it has been proposed to be part of the permeability transition pore （PTP）, a putative multiprotein complex candidate agent of the mitochondrial permeability transition （MPT）. Working at the single live cell level, we found that overexpression of VDAC1 triggers MPT at the mitochondrial inner membrane （MIM）. Conversely, silencing VDAC1 ex- pression results in the inhibition of MPT caused by selenite-induced oxidative stress. This MOM-MIM crosstalk was modulated by Cyclosporin A and mitochondrial Cyclophilin D, but not by Bcl-2 and BcI-XL, indicative of PTP operation. VDAC1-dependent MPT engages a positive feedback loop involving reactive oxygen species and p38-MAPK, and secondarily triggers a canonical apoptotic response including Bax activation, cytochrome e release and caspase 3 activation. Our data thus support a model of the PTP complex involving VDAC1 at the MOM, and indicate that VDACl-dependent MPT is an upstream mechanism playing a causal role in oxidative stress-induced apoptosis.展开更多
Objective： To culture olfactory ensheathing cells （OECs） of rats in vitro and to investigate its morphology, mitosis and immunocytochemistry, and to explore if the OECs could be a new donation for transplantation. ...Objective： To culture olfactory ensheathing cells （OECs） of rats in vitro and to investigate its morphology, mitosis and immunocytochemistry, and to explore if the OECs could be a new donation for transplantation. Methods： OECs were harvested from olfactory mucosa of Sprague Dawleys rats based on the differing rates of attachment of the various cell types, followed by glial fibrillary acidic protein （GFAP）, nerve growth factor （ NGF）, anti-low affinity receptor for NGF （ NGFRp75 ）, brain-derived neurotrophic factor （BDNF） , neurotrophino3 （ NTo3 ） and S-100 immunocytochemistry. The morphological changes and mitosis were observed under a phase contrast microscope at different culture time. Results - Three morphologically distinct types of cells, bipolar, multipolar and flat morphology were present in the primary culture of adult rat olfactory mucosa. Mitosis was characterized by a retraction of all processes, forming a sphere that divided into spherical daughter cells, the daughter cells sent out their processes. The OECs were immunoreactive for GFAP, NGFRp75, S-100, NGF, BDNF and NT-3. Conclusions： The OECs from nasal olfactory mucosa cultivated in the medium with fetal bovine serum could survive, divide, differentiate, and express the neurotrophin. It may become an accessible source for autologous grafting in spinal cord injury.展开更多
基金Supported by grants from the Nationl Natural Scientific Foundation of China, No.30300082, 30470467, and Scientific Foundation Committee of Guangdong Province, China, No.04009360
文摘AIM： To explore the effects of ultrasound exposure combined with microbubble contrast agent （SonoVue） on the permeability of the cellular membrane and on the expression of plasrnid DNA encoding enhanced green fluorescent protein （pEGFP） transfer into human umbilical vein endothelial cells （HUVECs）. METHODS： HUVECs with fluorescein isothiocyanatedextran （FD500） and HUVECs with pEGFP were exposed to continuous wave （1.9 MHz, 80.0 mW/cm^2） for 5 min, with or without a SonoVue. The percentage of FD500 taken by the HUVECs and the transient expression rate of pEGFP in the HUVECs were examined by fluorescence microscopy and flow cytornetry, respectively. RESULTS： The percentage of FDS00-positive HUVECs in the group of ultrasound exposure combined with SonoVue was significantly higher than that of the group of ultrasound exposure alone （24.0%± 5.5% vs 66.6% ± 4.1%, P 〈 0.001）. Compared with the group of ultrasound exposure alone, the transfection expression rate of pEGFP in HUVECs was markedly increased with the addition of SonoVue （16.1% ± 1.9% vs 1.5% ± 0.2%, P 〈 0.001）. No statistical significant difference was observed in the HUVECs survival rates between the ultrasound group with and without the addition of SonoVue （94.1% ± 2.3% vs 91.1% ± 4.1% ）. CONCLUSION： The cell membrane permeability of HUVECs and the transfection efficiency of pEGFP into HUVECs exposed to ultrasound are significantly increased after addition of an ultrasound contrast agent without obvious damage to the survival of HUVECs. This non- invasive gene transfer method may be a useful tool for clinical gene therapy of hepatic tumors.
文摘Voltage-dependent anion channel （VDAC）I is the main channel of the mitochondrial outer membrane （MOM） and it has been proposed to be part of the permeability transition pore （PTP）, a putative multiprotein complex candidate agent of the mitochondrial permeability transition （MPT）. Working at the single live cell level, we found that overexpression of VDAC1 triggers MPT at the mitochondrial inner membrane （MIM）. Conversely, silencing VDAC1 ex- pression results in the inhibition of MPT caused by selenite-induced oxidative stress. This MOM-MIM crosstalk was modulated by Cyclosporin A and mitochondrial Cyclophilin D, but not by Bcl-2 and BcI-XL, indicative of PTP operation. VDAC1-dependent MPT engages a positive feedback loop involving reactive oxygen species and p38-MAPK, and secondarily triggers a canonical apoptotic response including Bax activation, cytochrome e release and caspase 3 activation. Our data thus support a model of the PTP complex involving VDAC1 at the MOM, and indicate that VDACl-dependent MPT is an upstream mechanism playing a causal role in oxidative stress-induced apoptosis.
文摘Objective： To culture olfactory ensheathing cells （OECs） of rats in vitro and to investigate its morphology, mitosis and immunocytochemistry, and to explore if the OECs could be a new donation for transplantation. Methods： OECs were harvested from olfactory mucosa of Sprague Dawleys rats based on the differing rates of attachment of the various cell types, followed by glial fibrillary acidic protein （GFAP）, nerve growth factor （ NGF）, anti-low affinity receptor for NGF （ NGFRp75 ）, brain-derived neurotrophic factor （BDNF） , neurotrophino3 （ NTo3 ） and S-100 immunocytochemistry. The morphological changes and mitosis were observed under a phase contrast microscope at different culture time. Results - Three morphologically distinct types of cells, bipolar, multipolar and flat morphology were present in the primary culture of adult rat olfactory mucosa. Mitosis was characterized by a retraction of all processes, forming a sphere that divided into spherical daughter cells, the daughter cells sent out their processes. The OECs were immunoreactive for GFAP, NGFRp75, S-100, NGF, BDNF and NT-3. Conclusions： The OECs from nasal olfactory mucosa cultivated in the medium with fetal bovine serum could survive, divide, differentiate, and express the neurotrophin. It may become an accessible source for autologous grafting in spinal cord injury.
