| BackgroundWith the continuous development of society, more developed of traffic facilities, and increased motor vehicles, construction sites, factories, traffic accidents and injuries increase sharply. Among the injuries, trauma wounded increased greatly, and traumatic brain injury (TBI) is more than others, which is the chief factor of death cause. In the surviving patients, the mutilation rate is also higher, which do not only bring distress to patients, but also is a burden of family, even society. Although the first aid, image technology, emergency treatment and rehabilitation of TBI has made significant progress, the goals of treatment is only preserving residual neurological function and reducing the death rate of disability as far as possible, and not treating the occurred nerve damage effectively. The hottest and most difficult point of current research is how to promote the recovery of neurological function promoted by nerve regeneration after brain injury. After TBI, the main problems of recovery we faced are primary and secondary neurons missing and there are many factors, mainly myelin associated axon growth inhibition factor (Nogo-A, MAG,OMgp), inflammatory reaction, changes of extracellular matrix (ECM), formation of glial scar, that are not advantage to axon regeneration existed in the local microenvironment of damaged parts. Therefore, one of the strategies treating brain injury is to add the missing neurons and to improve local microenvironment of the damage. Although a large number of experiments have showed that neural stem cells (neural stem cells, NSCs) are both in adult and juvenile animal central nervous system (central nerve system, CNS). NSCs is a kind of cell mass that can self-renew, differentiate into neurons and glial cells, proliferate, migrate and differentiate under certain conditions and play a role in participating in repairing the defective neural functions. However, self-regeneration ability is limited in this kind of cells after central nervous system injury in adult, and can't finish the nerve regeneration function, especially in large damaged areas of central nervous tissues, which can't be finished only on its own neural stem cells to recovery. With recent research advances in the stem cells, there have been achieved exciting results in lots of studies through autologous or allogeneic transplantation of different stem cells (including different sources of adult stem and embryonic stem, etc). So stem cell transplantation for treatment of central nervous system impairment after traumatic brain injury brought us new hope. Bone marrow stromal cells are rich sources and easily obtained, researches showed that adult bone marrow stromal cells in vitro can be quickly and efficiently induced to differentiate into adult bone marrow-derived neural stem cells (human adult bone marrow-derived neural stem cells, MDNSCs), and MDNSCs have similar biological characteristics with their own brain-derived neural stem cells. If they are applied to autotransplantation, it can avoid potential immune rejection and ethical controversy. Besides seed cells, we also should improve the micro-environment of local damage, which is suitable for nerve tissue reparation and transplantation into the exogenous cells replacing of the dysfunction or death cells, and form functional integration to the host cells, which improve the dysfunction of nerve functions. The development of tissue engineering scaffolds brings hopes of solving the problem, which is similar to structure and function of the extracellular matrix, provides seed cell adhesion stent and plays an important role in place of the extracellular matrix. The main neural tissue engineering scaffolds include natural biological materials, synthetic materials, chemical modification of materials, composite materials, bio-derived materials and nano materials. On account of these factors, our research used bone marrow-derived neural stem cells as seed cell with Matrigel as scaffold (BD Matrigel) transplantation to observe the brain injury repair.The enrichment of the source of seed cells and development of tissue engineering scaffold material promotes the development therapy of stem cell transplantation. However, there are many problems to solve before the application of stem cell therapy, one of which is how to monitor and evaluate the situation of migration, differentiation and function of stem cells in vivo by non-invasive method. The mechanism of interaction and function contaction of transplanted cells and host cells is unclear, which is the problem needed to solve before the application to clinical with stem cells transplantation therapy. Meanwhile, an effective non-invasive method of assessing the transplantation therapies is equally important for our clinical strategy and evaluation of prognosis. In many available methods, one of the most popular methods is neuroimaging for intravital tracing, including magnetic resonance imaging, nuclear medicine techniques, fluorescence and bioluminescence technology. By long-term longitudinal evaluation, not need to put to animals to death, intravital tracing identifies the migration of transplanted cells and transplanted cells' mechanism of action preferably, which can not only promote the development of stem cell therapy, but also have more practical value for future clinical application.Now, the home and abroad researches are mostly tracking of transplanted stem cells'survive and migration in vivo in structure, such as SPIO (Superparamagnetic iron oxide, SPIO), etc. However, these methods combination of imaging can't trace their function inside the host nerve tissue, so we need to assess and trace the functional integration of transplanted stem cells and host tissue in vivo. Functional imaging studies in the past used FDG PET and BOLD MRI method can only provide indirect regional brain tissue activity information-the local sugar brain metabolic changes in neural activity, particularly at the cellular level, which is still not reflect the function of stem cell transplantation activity directly. Thus we took into account the application of MEMRI technique assessment of functional recovery after neural stem cell transplantation.Manganese ion imaging technique is a functional magnetic resonance imaging technique. In 1997, Lin YJ and Koretsky AP firstly used this technology to image brain functional electrical stimulation of forelimbs of rat, and explained the principle of MEMRI, that is voltage-dependent calcium ion channels open when electrical activity in nerve cells occurs, and manganese ions as a calcium analogue, which can through opening voltage-dependent calcium ion channels into the cell, combined with manganese ion is a paramagnetic substance, which can play a role in marking electrical activity of nerve cells, in place where the role of these activated regions obtained magnetic resonance enhancement, which based on through its open voltage-dependent calcium ion channels into the nerve cells, and in the local administration can be passed along the nerve tract anterograde characteristics, besides, when the nerve activity takes place manganese ions quickly into the activated neurons, but its removal in the cell has a longer period, manganese ions can increase in the aggregate when activate the neurons continued to, this manganese ion imaging-term cumulative effects will contribute to capture to the more subtle, more transient electrical activity of neurons from millions of the transplanted cells, their past applications include neurological imaging, nerve tract imaging and brain imaging studies to build the near future will also be applied to imaging nervous system diseases.In view of rat visual system function and anatomical certainty as well as domestic and foreign rare the application of manganese ion imaging techniques in rat visual cortex, this experiment chosen visual cortex as research object, try application dynamic MEMRI method tracer rat visual cortex function activities in order to supple methodological support for nervous system study.Part I:Transplantation of BMSC-D-NSCs with Matrigel for treatment of brain injury in ratsObjective:Currently, in the repair of central nervous function defect, especially the study of brain injury, less study on the stem cell transplantation with tissue engineering repair brain damage, yet rarely seen with bone marrow-derived neural stem cells and Matrigel for treatment of brain injury research. This study focuses on research effect of neural stem cells derived from bone marrow stromal cells with Matrigel transplantation on brain injury in the repair of neurological function.Methods:Isolation and identification of bone mesenchymal stem cells, with bFGF, EGF, and N2 induce bone marrow-derived to neural stem cells, and then labeled with fluorescent dye DiI before the transplant.32 Wistar rats were randomly divided into four groups:normal group, saline group, simply BMSC-D-NSCs transplantation group, BMSC-D-NSCs with Matrigel co-transplantation group. Use the modified free-fall impact brain injury model, after damaged 7 days transplant into damage area, respectively,1,7,14,21,28 days after cell transplantation in animal behavior score, after 4 weeks transplantation animals were sacrificed, calculated cavity volume caused by damage, and analysis the transplanted cells survival and migration, All statistical analysis used SPSS 13.0 software, compared the volume of different groups after treatment, use One-Way analysis of variance; different groups at five time points of behavioral score using repeated measures analysis of variance, P<0.05 was considered significant.Results:Defects volume of BMSC-D-NSCs transplantation group and BMSC-D-NSCs with Matrigel co-transplantation group were significantly reduced compare with saline group (P=0.000), and the BMSC-D-NSCs with Matrigel co-transplantation group had a more significant improvement than the BMSC-D-NSCs transplantation group (P=0.000). In mNSS, compare with BMSC-D-NSCs and saline group BMSC-D-NSCs with Matrigel co-transplantation group functional improvement is obvious, the difference was significant (P<0.05).Conclusion:BMSC-D-NSCs with Matrigel co-transplantation group can better promote the repair of brain damage than the BMSC-D-NSCs group.Part II:Dynamic manganese-enhanced functional MRI on rat visual cortex.Objective:Although our preliminary study showed that, BMSC-D-NSCs with matrigel co-transplantation can better promote the recovery of neural function than BMSC-D-NSCs alone, but its function in vivo situation is still not clear, our group proposed use dynamic MEMRI functional imaging to study the BMSC-D-NSCs in vivo function after transplantation, we carried out some research on the imaging study, this part of the paper, we use dynamic MEMRI technology imaging rat visual cortex in order to provides information on the methodology for nervous system function reseach.Method:Six adult male Wistar rats were chosen and the process was divided into 4 continuous phases. No agent was injected into the rats in the first phase (5min). Opening the BBB with mannitol and injecting manganese chloride were performed by right internal carotid artery (ICA) in the second phase (10min). In the third phase (15min), manganese chloride was administrated by ICA and vision stimulation was performed before the imaging process. And the mixed liquor of manganese chloride and glutamate was injected in the forth phase (5min).Result:No specific enhanced region was found in rat brain in the first and second phases. The right visual cortex was enhanced specifically on T1WI in the third. And many brain regions of right hemisphere, the sites that agent was injected, were obviously enhanced in the forth phase. ROI analysis showed that the signal intensity in the third phrase (1.897±0.172) was significantly stronger as compared with that in the second phrase (1.549±0.163) (P<0.05).Conclusion:The dynamic mangamese-enhanced functional magnetic resonance imaging can analyze the function activities of the vision cortex in rats and pvovide a new method for researching the function of nervous system. |
PDF Full Text Request