| Background:Oligodendrocyte precursors(OPCs)are precursors of oligodendrocytes.By differentiating into oligodendrocytes(OLs),OPCs play important role in developmental myelination in the central nervous system(CNS).White matter contains bundles of myelinated axons that connect the regions of gray matter allowing them to communicate with one and other.Changes in white matter that commonly occurs between perinatal period are myelination disturbances and demyelination.These changes will lead to various neurological disorders,including cognitive impairment,cerebral palsy and mental retardation.Perinatal period,between 25 weeks of gestation and 1-2 years after birth,is a stage of maturation of OPCs in the brain development.At this time,the developing CNS exhibits vulnerability to various insults such as hypoxic-ischemia,inflammation and congenital defects,causing the loss of OPCs,extensive myelination disturbances and demyelination,forming white matter lesions.Because endogenous OPCs are in a state of relative insufficiency,transplantation of exogenous OPCs may promote remyelination in the area of white matter lesions.Chapter 1:Evaluation of Myelin Reparation Effects of OPCs Transplantation on White Matter Injury in Premature Infants by Electron MicroscopyObjective:To investigate the effect of oligodendrocyte precursor cells(OPCs)transplantation on myelin reparation in premature infants with white matter injury(WMI).Methods:Thirty-two neonatal SD rats were randomly divided into 8 groups(n = 4):sham group,model control group,5d ventricle transplantation group,5d white matter transplantation group,9d ventricle transplantation group,9d white matter transplantation group,14d ventricle transplantation Group and 14d white matter transplantation group.In the transplantation group and the model control group,right common carotid artery were ligated and hypoxia for 80min at 3 days-old to establish a rat model of WMI in premature infants.At 5,9,and 14 days after modeling,3×105 OPCs were transplanted into the ventricle(ventricle transplants)or white matter(white matter transplants)via brain stereotaxic apparatus.At ages of 60 and 90 days,the myelin sheaths in the the corpus callosum regions were observed and evaluated by electron microscopy.Fifty myelin sheaths were randomly selected to calculate the g-ratio,in order to evaluate the thickness of myelin sheath.Results:At 60 days of age,the appearance of myelin in the sham group was normal,but in the model control group was loose,separated and disintegrated.In the 5d white matter and 5d ventricle transplantation group,myelin was thin and the lamellar structure remained relatively tense in shape.In the 9d white matter transplantation group,the myelin structure was loose.In the 9d ventricle,14d white matter and 14d ventricle transplantation group,separated and disintegrated appearance in the myelin were rare.At the 90th day of age,the appearance of myelin in the sham group was normal,but in the model control group was highly loose and disintegrated.In the 5d,9d and 14d transplantation group,whatever the ventricle or white matter group,the loose,separated and disintegrated appearance were relatively rare.Especially in the 14d ventricle transplantation group,separated and disintegrated appearance were highly rare and the lamellar structure was almost normal.The results of g-ratio showed that,there was no difference between the groups at 60 days of age.At 90 days of age,when comparing with the model control group,the 5d white matter transplantation group,the 5d ventricle transplantation group,the 9d white matter transplantation group showed significant drop in mean of g-ratio(P>0.05),suggesting the incease in thickness of myelin sheath,but when comparing the sham group,there was no significant difference between sham group and the above group(P>0.05);in the 5d ventricle transplantation group,9d ventricle transplantation group and 14d white matter transplantation group,there were significant difference between the results of 60 days and 90 days of age(P<0.05),the thickness of myelin sheath in the 60 days of age were increased;There were no significant difference between other groups(P>0.05).Myelin injury score results showed that compared with the model control group,in the 60-day-old rats,except for the 9-day white matter transplantation group,the rest of the transplantation group had less injury score(P<0.05).In the 90-day-old rats,except for 9d of the ventricle In the transplantation group,the rest of the transplantation group suffered less injury(P<0.05).The score of injury was significantly increased in all transplanted and model control groups compared with sham group(P<0.05).Conclusion:The results show that OPCs transplantation has a certain effects in myelin reparation in the premature infant with white matter injury.However,these effects are not obvious and need to be further confirmed in the next research.Chapter 2:Effects of OPCs Transplantation Combined with Miconazole on Myelin Reparation in Congenital Leukodystrophy Model MiceObjective:To investigate the effect of oligodendrocyte precursor cells transplantation combined with miconazole in the neonatal period on myelin reparation in mice with leukodystrophy.Methods:Using MBP knockout mice(shiverer mice)as a model of leukodystrophy,the newborn shi-/-mice were randomly assigned to negative control group and treatment group,each group of 5.In addition,5 newborn shi +/+ mice were selected as positive control group.In the treatment group,oligodendrocyte precursor cells(OPCs)were transplanted 24 hours after birth,and the control group was replaced by PBS.At the same time,in the treatment group miconazole was injected intraperitoneally 1-5 days after birth,and the control group was replaced by normal saline.The control group was replaced with normal saline.At the age of 8 weeks,Western blot were used to detect MBP protein expression and the myelin sheath structure of the corpus callosum was observed by transmission electron microscopy.Results:Compared with the negative control group,the MBP expression in the treatment group was significantly enhanced and the average fluorescence intensity was significantly increased(P<0.05),But when compared with the positive control group,the MBP expression in the treatment group was significantly weakened and the average fluorescence intensity significantly decreased(P<0.05).Under electron microscope,the myelin sheath structure in the corpus callosum region was significantly diverse among the groups.In the negative control group,the myelin sheath was rare and irregular,and the lamellar arrangement was loose and disorderly.In the positive control group,the myelin was significantly increased and thickened.With the normal morphology,the lamellar layer was closely arranged and the boundary was clear.In the treatment group,the number of myelin was significantly higher than that of the negative control group,and some myelin sheath was normal in shape,but most myelin was still slender.Conclusion:OPCs combined with miconazole in the neonatal period helps to improve demyelination in mice with leukodystrophy,but does not completely reverse the pathological status of injuryd myelin.Chapter 3:Cell Therapy for Cerebral Palsy:a Meta-analysis of Randomized Controlled TrialsObjective:To evaluate efficacy and safety of cell therapy in children with cerebral palsy.Methods:Searches were conducted by two independent researchers in April 2017 in foreign language database(including Embase,PubMed,and Cochrane Libraries)and Chinese database(including Chinese Biomedical Literature Database(CBM),ZhiWang and WanFang database).According to the inclusion and exclusion criteria,randomized controlled trials of cell therapy for cerebral palsy in children were included.The literature quality was assessed and the data was extracted.A meta-analysis was conducted using R 3.4.0 software.The primary outcomes were gross motor function measurement(GMFM)and the second outcomes were severe adverse events(SAEs).Effects were expressed as standardized mean difference(SMD)with 95%CI.Statistical analysis was performed at the same time forest map and plot map were made.Results:A total of 334 cases of five RCTs were included in the study.The GMFM score of the experimental group was higher than that of the control group(SMD =0.68,95%CI=0.44-0.91,P<0.0001).In the umbilical cord blood group,the score of the experimental group increased more(Q =5.53,P =0.02).The incidence of serious adverse events in the experimental group was approximately similar to that of the control group.Conclusion:Cell therapy for children with cerebral palsy has an effective short-term effect,but due to the limitations of the literature,more high-quality studies are needed to validate the results. |
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