Bone marrow mesenchymal stem cells versus bone marrow mononuclear cells for treating cerebral palsy / 中国组织工程研究

BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) and bone marrow mononuclear cells(BMMNCs) have been both used to treat spastic cerebral palsy. However, the differences in their therapeutic effects remain unknown. OBJECTIVE: To compare the therapeutic effects of BMMSCs and BMMNCs in cerebral palsy children as well as on fine motor function. METHODS: 105 children with spastic cerebral palsy were enrolled and randomly assigned to three groups: BMMSCs group, BMMNCs group and control group. Patients in the two transplantation groups received four intrathecal cell injections, and those in the control group received Bobath therapy, twice a day, for consecutive 3 weeks. The Gross Motor Function Measure (GMFM) and Fine Motor Function Measure (FMFM) were used to evaluate the therapeutic efficacy at 3, 6 and 12 months after transplantation. RESULTS AND CONCLUSION: At 3 months after cell transplantation, scores in A dimension of GMFM and in A, C dimensions of FMFM in BMMSC group were all superior to those of BMMNC group and control group (P < 0.05). At 6 months after cell transplantation, scores in A, B dimensions of GMFM and in A, B, C, D and E dimensions of FMFM in BMMSC group were better than those of BMMNC group and control group (P< 0.05), and total scores of GMFM and FMFM were also better in the BMMSC group (P < 0.05). At 12 months after cell transplantation, scores in A, B and C dimensions of GMFM and A, B, C, D and E dimensions of FMFM scores in BMMSC group were all superior to those of BMMNC group and control group (P < 0.05) as well as the total GMFM and FMFM scores. There were six cases of low intracranial pressure headache in BMMNC group and six cases of low-grade fever in BMMSC group. In summary, both BMMSCs transplantation and BMMNCs transplantation are safe, effective and feasible for the treatment of spastic cerebral palsy in children, and moreover, BMMSCs transplantation is a better method than BMMNCs transplantation to improve gross and fine motor functions of spastic cerebral palsy children.

Ferumoxide labeled Flk1+ CD31- CD34- human bone marrow mesenchymal stem cells and its in vivo tracing in the brains of Macaca Fascicularis / 中国医学科学院学报

<p><b>OBJECTIVE</b>To explore the method for labeling Flk1+ CD31- CD34- human bone marrow mesenchymal stem cells (hBMSCs) with ferumoxide-PLL and evaluate the feasibility of its tracing after transplantation into the brains of Macaca Fascicularis.</p><p><b>METHODS</b>The hBMSCs were incubated with ferumoxide-PLL. Trypan blue staining, Prussian blue staining, and transmission electron microscope were performed to show intracellular iron, marking efficiency, and the vigor of the labeled cells. After the hBMSCs were transplanted into the brains of cynomolgus monkeys by stereotaxis, magnetic resonance imaging (MRI) was performed to trace the cells in vivo. Cell survival and differentiation were studied with immunohistochemistry, Prussian blue staining, and HE staining.</p><p><b>RESULTS</b>The marking efficiency of the ferumoxide-PLL was 96%. Iron particles were found intracytoplasmic of the hBMSCs by Prussian blue staining and transmission electron microscopy. The relaxation rates of labeled cells in MRI were 4.4 and 4.2 times higher than those of the unlabeled cells. Hypointensity area was found by MRI three weeks after transplantation. Many hBMSCs and new vessels were found in the transplantation zone by pathological and immunofluorescence methods.</p><p><b>CONCLUSIONS</b>Ferumoxide-PLL can effectively label hBMSCs and thus increase its contrast in MRI results. The cells can survive in the brains of cynomolgus monkeys. The labeled hBMSCs can be traced in vivo by MRI.</p>

Induction of functional recovery by co-transplantation of neural stem cells and Schwann cells in a rat spinal cord contusion injury model / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To study the transplantation efficacy of neural stem cells (NSCs) and Schwann cells (SC) in a rat model of spinal cord contusion injury.</p><p><b>METHODS</b>Multipotent neural stem cells (NSCs) and Schwann cells were harvested from the spinal cords of embryonic rats at 16 days post coitus and sciatic nerves of newborn rats, respectively. The differential characteristics of NSCs in vitro induced by either serum-based culture or co-culture with SC were analyzed by immunofluorescence. NSCs and SCs were co-transplanted into adult rats having undergone spinal cord contusion at T9 level. The animals were weekly monitored using the Basso-Beattie-Bresnahan locomotor rating system to evaluate functional recovery from contusion-induced spinal cord injury. Migration and differentiation of transplanted NSCs were studied in tissue sections using immunohistochemical staining.</p><p><b>RESULTS</b>Embryonic spinal cord-derived NSCs differentiated into a large number of oligodendrocytes in serum-based culture upon the withdrawal of mitogens. In cocultures with SCs, NSCs differentiated into neuron more readily. Rats with spinal cord contusion injury which had undergone transplantation of NSCs and SCs into the intraspinal cavity demonstrated a moderate improvement in motor functions.</p><p><b>CONCLUSIONS</b>SC may contribute to neuronal differentiation of NSCs in vitro and in vivo. Transplantation of NSCs and SCs into the affected area may be a feasible approach to promoting motor recovery in patients after spinal cord injury.</p>

Induction of Endogenous Neural Stem Cells with Hyperbaric Oxygen after Half Cut-off of Spinal Cord in Rats / 中国康复理论与实践

@#ObjectiveTo investigate the effect of hyperbaric oxygen (HBO) on proliferation and differentiation of endogenous neural stem cells after acute spinal cord half cut-off in rats. MethodsThe differences of proliferation and differentiation of endogenous neural stem cells between injured group and intervention group were compared. ResultsThere were remarkable differences between injured group and intervention group. ConclusionHBO can promote the proliferation and differentiation of the neural stem cells in rats after spine cord injury.

A Model of Focal Cortical Infarctionin Rat:Mini mally Invasive Craniotomy / 中国康复理论与实践

@#ObjectiveTo develop a stable model of focal cerebral infarction in rat to study the curative effect of neural stem cells transplantation.MethodsThirty-seven rats were selected which were divided into two groups in random, experimental group and control group. The focal infarction model was developed by the ligation of the left middle cerebral artery followed by the ligation of the ipsilateral common carotid artery and the temporary clip occlusion of the contralateral common carotid artery for 1.5 h. The operation adopted minimally invasive craniotomy though temporal bone. The model was evaluated by examining the neurologic deficits, ink perfusion, TTC staining and Magnetic Resonance imaging.ResultsAll the rats were in good condition after the operation, the mortality rate was 6.25% after 4 weeks. Ink perfusion and TTC staining confirmed that the ischemia was confined to the cortex. The areas of infarction measured 83.52 mm3 by Magnetic Resonance imaging after 4 weeks.ConclusionA stable focal cerebral infarction model can be achieved by minimally invasive craniotomy. It is superior for its homogeneity of infarction volume and site, and its low mortality. It can be used for the study of transplantation of neural stem cells.

Differentiation of rat neural stem cells and its relationship with environment / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To explore the differentiation fates of rat neural stem cells (NSCs) in different environmental conditions.</p><p><b>METHODS</b>NSCs derived from 16-day-old rat embryo were proliferated in vitro and implanted into the brain of rats with intra-cerebral hemorrhage. At the same time some NSCs were co-cultured in vitro with Schwann cells derived from newborn rats. MAP-2, GFAP and GalC (which are the specific markers of neural cells, astrocytes and oligodendrocytes respectively), BrdU and beta-tubulin were detected by immunohistochemical and immunofluorescent methods.</p><p><b>RESULTS</b>BrdU positive cells that were implanted into the brain distributed around the hemorrhagic area. The majority of them were GFAP positive astrocytes while a few of them were beta-tubulin positive neural cells or GalC positive oligodendrocytes. After being co-cultured with Schwann cells in vitro, NSCs are predominately shown beta-tubulin and MAP-2 positive, and only a minority of them were GFAP or GalC positive.</p><p><b>CONCLUSIONS</b>The hemorrhagic environment in vivo induces NSCs to differentiate mainly into astrocytes while co-culture with Schwann cells in vitro induce the majority of NSCs to differentiate into neural cells.</p>

Experimental study of rat cerebral hemorrhage model via autologous blood injection / 中国康复理论与实践

@#ObjectiveTo observe the effect of rat cerebral hemorrhage model via autologous blood injection.MethodsBlood extracted from rats' cardiac ventricles was injected into the right caudate nucleus via stereotactic method. Pathological examination,ultrastructural visualization and motor functional test were applied to validate the feasibility of this method.ResultsRats showed paralysis of left extremities rapidly after surgery. Pathological examination confirmed the formation of hematoma in caudate nucleus. The deviation of hematoma volume and location among individuals was significantly lower than that of rats made by collagenase injection. Transmission electron microscopy showed wide mylin degeneration in right caudate nucleus. Motor functional test revealed the dysfunction of left extremities with marked lower deviation among individuals compared with that of collagenase injection (P<0.05).ConclusionAutologous blood injection can result in significantly lower deviation of hematoma volume and location as well as dysfunctional degree compared with that of collagenase injection.

Effect of combined therapy of mild hypothermia and hibernation on severe brain injury / 中国康复理论与实践

@#ObjectiveTo investigate the efficacy of combined therapy of mild hypothermia and hibernation to treat severe brain injury. Methods24 patients with severe brain injury were randomly divided into combined therapy group and normothermia group. Glasgow Coma Scale scores of all the patients were in the range of 3 to 8. No later than 10 hours after their injury, hypothermia patients were given half dosage of No.1 hibernation cocktail and had been cooled by cooling blankets to 32℃-34℃ (rectal temperature) for 5 days, then to 35℃ for 24 hours, and slowly increased to their normal level. 3 days and 7 days after their admission, intracranial pressure,creatine phosphate kinase,partial pressure of arterial O2 and CO2, platelet and Na+,K+ were measured.7 days after their admission, Glasgow Outcome Scale scores of each patient and mortality of each group were measured. ResultsThe mortality of combined therapy group(25.0%) was significantly lower than that of normothermia group (66.6%,P<0.05). The decreased values of intracranial pressure, creatine phosphate kinase and platelet number of combined therapy group were all significantly higher than that of normothermia group respectively (P<0.05). There were no significant difference in mean artery pressure, blood electrolyte, and partial pressure of arterial O2 and CO2 between these two groups(P>0.05). ConclusionThe combined therapy of mild hypothermia and hibernation can effectively reduce the mortality of patients with severe brain injury as it is much easier, less invasive and with less complications.

Research on repairing facial nerve injury of rabbits by neural stem cells and autologous fasia / 中国康复理论与实践

@#ObjectiveTo observe the effect on repairing facial nerve injury of rabbits by neural stem cells and autologous fasia. Methods22 rabbits with transected facial nerve were divided into 2 groups randomly, control group (8 rabbits,15 sides totally), which transected facial nerve were wrapped by autologous fasia, and treament group (14 rabbits, 20 sides totally), which were wrapped by neural stem cells and autologous fasia. Six weeks after transplantation, neuro-electrophysiological test, immunohistochemical examination were done. The number and thickness of myelin in the re-connected area of transected facial nerve were observed. ResultsThe transplanted animals recovered much better than that in control group (P<0.05). Immunohistochemical examination showed a great deal of BrdU positive cells around the re-connected area of transected facial nerve. Immunohistochemical staining also found plenty of regenerative myelins in this area in the treatment group. While in control group, there were no BrdU positive cells and only a few of regenerative myelins in the same area. ConclusionTransplantation of neural stem cells combined with autologous fasia might become the new method to treat facial nerve injury.

Ultrastructure of neural stem cells of Wistar rats cultured in vitro / 中国康复理论与实践

@#ObjectiveTo detect the ultrastructure of neural stem cells (NSCs) cultured in vitro .MethodsNSCs separated from the cortex of 17—19 days Wistar rat fetus were cultured and induced to differentiate in vitro. Electron microscopes were used to visualize the ultrastructure of these cells before and after differentiation.ResultsNSCs had the similar cellular size, morphology and intracellular structures pre-differentiation. Cells were able to proliferate via mitosis. The nucleus/cytoplasm ratio was very high. The nucleus was poly-morphological. Cells had very little cytoplasm and no mature organelles. After differentiation, several processes protruded out from cellular surface. Cells became flat shape, the volume of cytoplasm increased dramatically and various kinds of mature organelles appeared in the cytoplasm. Cells differentiated into two kinds of cells,neural cells and glial cells,with quite different morphology and intracellular structure. ConclusionNSC is one kind of original cells which can be induced to differentiate into mature neural cells and glial cells.

Effect of rat Schwann cell secretion on proliferation and differentiation of human neural stem cells / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To investigate the effect of rat Schwann cell secretion on the proliferation and differentiation of human embryonic neural stem cells (NSCs).</p><p><b>METHODS</b>The samples were divided into three groups. In Group One, NSCs were cultured in DMED/F12 in which Schwann cells had grown for one day. In Group Two, NSCs and Schwann cells were co-cultured. In Group Three, NSCs were cultured in DMEM/F12. The morphology of NSCs was checked and beta-tubulin, GalC, hoechst 33342 and GFAP labellings were detected.</p><p><b>RESULTS</b>In Group One, all neural spheres were attached to the bottom and differentiated. The majority of them were beta-tubulin positive while a few of cells were GFAP or GalC positive. In Group Two, neural spheres remained undifferentiated and their proliferation was inhibited in places where Schwann cells were robust. In places where there were few Schwann cells, NSCs performed in a similar manner as in Group One. In Group Three, the cell growth state deteriorated day after day. On the 7th day, most NSCs died.</p><p><b>CONCLUSION</b>The secretion of rat Schwann cells has a growth supportive and differentiation-inducing effect on human NSCs.</p>

Schwann cells transplantation promoted and the repair of brain stem injury in rats / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To explore the possibility of Schwann cells transplantation to promote the repair of injured brain stem reticular structure in rats.</p><p><b>METHODS</b>Schwann cells originated from sciatic nerves of 1 to 2-day-old rats were expanded and labelled by BrdU in vitro, transplanted into rat brain stem reticular structure that was pre-injured by electric needle stimulus. Immunohistochemistry and myelin-staining were used to investigate the expression of BrdU, GAP-43 and new myelination respectively.</p><p><b>RESULTS</b>BrdU positive cells could be identified for up to 8 months and their number increased by about 23%, which mainly migrated toward injured ipsilateral cortex. The GAP-43 expression reached its peak in 1 month after transplantation and was significantly higher than that in the control group. New myelination could be seen in destructed brain stem areas.</p><p><b>CONCLUSION</b>The transplantation of Schwann cells can promote the restoration of injured brain stem reticular structure.</p>