1.Fibrin scaffold promoted the differentiation of neural stem cells to the neurons and inhibited the proliferation of astrocytes
Yanhong CUI ; Mubin WANG ; Jing CHEN ; Qian CHEN ; Xianglan SUN ; Aihua GONG ; Zhijian ZHANG ; Yongchang CHEN ; Ping JIANG
Acta Anatomica Sinica 2010;41(1):41-47
Objective To investigate the effects of the fibrin scaffold on the differentiation and the proliferation of neural stem cells and astrocytes. Methods Neural stem cells and the gliocytes derived from spinal cord were cultured in vitro respectively. The purified neural stem cells or gliocytes were seeded separately onto the fibrin scaffolds as experimental group and the glass slides modified with poly-L-lysine(PLL)as control group. At different time in culture the neural stem cells were immunofluorescence stained with antibodies against the marker of neurons I.e. Neurofilament(NF).The length of NF-positive neuritis was masured and the average value was calculated in the culture well (n=4). The gliocytes were immunofluorescence stained with antibodies against the marker of astrocytes I.e. Glial fibrillary acidic protein (GFAP ). The total number of the cells and the GFAP-positive cells were counted from 5 different fields of vision in the culture well (n=4), then the average ratio of GFAP-positive cells was calculated. The differentiation of neural stem cells, the extension of neurites and the proliferation of astrocytes on the fibrin scaffolds were compared with those on the slides. The protein of GFAP was detected by Western blotting to analyse the mature degree of astrocytes. All above experiments were repeated 3 times respectively. Results Immunofluorescence staining showed that the NF-positive neurites in the fibrin scaffold group were longer than those in the control group, whereas GFAP-positive cells were fewer than those in the control group. The expression of GFAP in the cells on the scaffold was lower than that in the control group.Conclusion The fibrin scaffold could promote differentiation of the neural stem cells to neurons and extension of the neurites. Meanwhile, the scaffold could inhibit proliferation and mature of the astrocytes.
2.Effect of tail vein transplantation of human amniotic mesenchymal stem cells with different transforming growth factor-β expressions on sciatic nerve function in xenografted mice
Mubin CHEN ; Qingqing LI ; Huihui CHAI ; Kuntai XIAO ; Xiaoguang HUANG ; Yuting XU ; Shan XUE ; Haitao SUN ; Yanwu GUO
Chinese Journal of Neuromedicine 2020;19(1):9-16
Objective To investigate the effect of tail vein transplantation of human amniotic mesenchymal stem cells (hAMSCs) with different transforming growth factor (TGF)-β expressions on recovery of sciatic nerve function in peripheral nerve xenotransplantation mice.Methods The hAMSCs were isolated from amnion membranes by healthy mother donors and identified by fluorescence activated cell sorter.The up-regulated and down-regulated TGF-β lentiviral plasmids were constructed and transfected into the purified hAMSCs;hAMSCs with stable up-regulated or down-regulated TGF-β expression were constructed.The sciatic nerves of C57BL/6 mice were isolated and cut out,and sciatic nerves of SD rats were isolated and transplanted into the sciatic nerve defected C57BL/6 mice to construct peripheral nerve xeno-transplanted mice models;these mice models were divided into 4 groups (n=10)according to random number table:control group,hAMSCs treatment group,high-expressed TGF-βhAMSCs treatment group,and low-expressed TGF-β hAMSCs treatment group;one d before modeling,phosphate buffer saline (PBS) or hAMSCs re-suspension were drawn with a syringe and slowly pushed into the tail veins of mice for transplantation treatment;14 d after treatment,DigGait analysis system was used to evaluate the recovery of sciatic nerve function in each group of mice.Result Fourteen d after treatment,the sciatic nerve function index (SFI) of the high-expressed TGF-β hAMSCs treatment group (-25.820±0.286) was significantly higher than that of the low-expressed TGF-β hAMSCs treatment group (-33.413±0.920) and hAMSCs treatment group (-30.755±0.421,P<0.05).Conclusion The tail vein transplantation of hAMSCs with TGF-β high expression can effectively improve the sciatic nerve function in peripheral nerve xenotransplantation mice,which may be a new breakthrough in the treatment of peripheral nerve defects.
3.Effect of human amniotic mesenchymal stem cell transplantation on motor behaviors in hSOD1-G93A mouse models of amyotrophic lateral sclerosis
Kuntai XIAO ; Xiaomei LIANG ; Ya GAO ; Mubin CHEN ; Duobin WU ; Xiaodan JIANG ; Yanwu GUO ; Haitao SUN
Chinese Journal of Neuromedicine 2017;16(7):649-656
Objective To explore the therapeutic effect of intravenous transplantation of human amniotic mesenchymal stem cells (hAMSCs) on protection of motor behaviors in hSOD1-G93A mouse models of amyotrophic lateral sclerosis (ALS).Methods Amnion membranes were obtained from placentas delivered by healthy mother donors.The hAMSCs were gradually isolated and purified from amnion membranes using tissue culture method.Immunophenotypes of the isolated hAMSCs were analyzed using fluorescence activated cell sorter (FACS).Transgenic mice harboring a high copy number of hSOD1-G93A (B6SJL-TgN [SOD1-G93A] 1Gur) transgene were used in this study.Hemizygous transgenic progenies were maintained by mating the transgenic males with F1 hybrid wild-type (WT)females.The progenies were genotyped by polymerase chain reaction (PCR) using genomic DNA isolated from mouse tail after birth.The study included hSOD1-G93A mice transplanted with hAMSCs,PBS-injected transgenic mice,and normal WT mice (n=12).The hAMSCs were administered intravenously in jugular vein of the mice under anesthesia.The cells (1 ×106) in 200 μL PBS were delivered over 10 min.Animals received cells or PBS at 12,14,and 16 weeks old,respectively.The disease onset and progression of ALS mice models were monitored using rotarod performance test,PaGE test,and CatWalk gait analysis since 8 weeks old every week.Results (1) The immunophenotype of the isolated hAMSCs was conformed using FACS.These cells were positive for CD29,CD44,CD73,CD90,and CD166,but negative for CD14,CD34,CD45,CD123,and human leukocyte (site) DR antigen.Interestingly,stage specific embryo surface antigen 4 and octuber binding transcription factor 4 were detected in hAMSCs.(2) ALS mice in the hAMSCs transplantation group had significantly improved motor functions than those in the PBS treatment group:motor performance on the rotarod test (from 14 to 18 weeks old),PaGE test (from 15 to 18 weeks old) and CatWalk gait analysis (from 15 to 19weeks old) in hAMSCs-injected ALS mice was significantly improved as compared with that in the PBS treatment group (P<0.05).Conclusions The multiple transplantation of hAMSCs by intravenous delivery can bring amelioration of the disease phenotype,as evidenced by improved motor function in hSOD1-G93A mouse models.The hAMSCs transplantation can be considered as a promising cellular treatment for ALS.