RESUMO
ObjectiveDirected differentiation of human induced pluripotent stem cells （hiPSCs） into spinal cord γ-aminobutyric acid （GABA）-ergic progenitor cells were implanted into an decellularized optical nerve （DON） bioscaffold to construct a hiPSC-derived inhibitory neural network tissue with synaptic activities. This study aimed to provide a novel stem cell-based tissue engineering product for the study and the repair of central nervous system injury. MethodsThe combination of stepwise directional induction and tissue engineering technology was applied in this study. After hiPSCs were directionally induced into human neural progenitor cells （hNPCs） in vitro， they were seeded into a DON for three-dimensional culture， allowing further differentiation into inhibitory GABAergic neurons under the specific neuronal induction environment. Transmission electron microscopy and whole cell patch clamp technique were used to detect whether the hiPSCs differentiated neurons could form synapse-like structures and whether these neurons had spontaneous inhibitory postsynaptic currents， respectively， in order to validate that the hiPSC-derived neurons would form neural networks with synaptic transmission potentials from a structural and functional perspective. ResultsThe inhibitory neurons of GABAergic phenotype were successfully induced from hiPSCs in vitro， and maintained good viability after 28 days of culture. With the transmission electron microscopy， it was observed that many cell junctions were formed between hiPSC-derived neural cells in the three-dimensional materials， some of which presented a synapse- like structure， manifested as the slight thickness of cell membrane and a small number of vesicles within one side of the cell junctions， the typical structure of a presynatic component， and focal thickness of the membrane of the other side of the cell junctions， a typical structure of a postsynaptic component. According to whole-cell patch-clamp recording， the hiPSC-derived neurons had the capability to generate action potentials and spontaneous inhibitory postsynaptic currents were recorded in this biotissue. ConclusionsThe results of this study indicated that hiPSCs can be induced to differentiate into GABAergic progenitor cells in vitro and can successfully construct iPSC-derived inhibitory neural network tissue with synaptic transmission after implanted into a DON for three-dimensional culture. This study would provide a novel neural network tissue for future research and treatment of central nervous system injury by stem cell tissue engineering technology.
RESUMO
ObjectiveTo construct a neural network-like tissue with the potential of synaptic formation in vitro by seeding human induced pluripotent stem cell-derived neural precursor cells （hiPSC-NPCs） on decellularized optic nerve （DON）， so as to provide a promising approach for repair of nerve tissue injury. MethodsThrough directional induction and tissue engineering technology， human induced pluripotent stem cells （hiPSCs） and 3D DON scaffolds were combined to construct neural network-like tissues. Then the hiPSCs were directionally induced into human neural precursor cells （hNPCs） and neurons. Immunofluorescence staining was used to identify cell differentiation efficiency. 3D DON scaffolds were prepared. Morphology and cytocompatibility of scaffolds were identified by scanning electron microscopy and Tunnel staining. Induced hiPSC-NPCs were seeded on DON scaffolds. Immunofluorescence staining， scanning electron microscopy， transmission electron microscopy and patch clamp were used to observe the morphology and functional identification of constructed neural network tissues. Results①The results of immunofluorescence staining suggested that most of hiPSC-NPCs differentiated into neurons in vitro. We had successfully constructed a neural network dominated by neurons. ② The results of scanning electron microscopy and immunohistochemistry suggested that a neural network-like tissue with predominating excitatory neurons in vitro was successfully constructed. ③The results of immunohistochemical staining， transmission electron microscopy and patch clamp indicated that the neural network-like tissue had synaptic transmission function. ConclusionA neural network-like tissue mainly composed of excitatory neurons has been constructed by the combination of natural uniform-channel DON scaffold and hiPSC-NPCs， which has the function of synaptic transmission. This neural network plays a significant role in stem cell derived replacement therapy， and offers a promising prospect for repair of spinal cord injury （SCI） and other neural tissue injuries.
RESUMO
<p><b>OBJECTIVE</b>To discuss the correlation between the degree of cervical spinal cord compression and gender, age, height of intervertebral space and the abnormality curvature of cervical vertebrae. The multivariable linear regression was used to build the prediction model of cervical spinal cord compression.</p><p><b>METHODS</b>One hundred and twenty patients with cervical spondylosis were divided into 4 groups according to the degree of spinal cord compression. To measure the sagittal diameter of spinal cord (a) and medulla-pons junction (M) in MRI individually, and then calculate the ratio of a/M; according to the ratio to evaluate the degree of cervical spinal cord compression. In X-ray films to measure the height and angle of intervertebral space, curvature of cervical vertebrae (according to Borden's method and double line method of C2-7 Cobb angle to abtain data of curvature of cervical vertebrae). To performe linear regression and multiple linear regression to analyze the correlation above the data and build the prediction model of cervical spinal cord compression.</p><p><b>RESULTS</b>(1) With the exacerbation of cervical spinal cord compression, all measured data gradually decreased; there was significant difference in the anterior height and angle of intervertebral space, the ratio of a/M between any two groups (P < 0.05). (2) The ratio of a/M showed positive corrrelation with the anterior height of intervertebral space (R = 0.296, P < 0.001) and the curvature of cervical vertebrae (are to chord distance, R = 0.241, P < 0.001), but showed negative corrrelation with the proportion of cervical spinal cord compression (R = -0.821, P < 0.001); Borden's method are to chord distance) showed positive correlation with the double line method of C2-7 Cobb angle (R = 0.840, P < 0.001). The predictive equation of the ratio of a/M by multiple linear regression was obtained, and the F = 8.959, R = 0.434, P < 0.001, in which the age, the anterior height of intervertebral space and the curvature of cervical vertebrae were risk factors of cervical spinal cord compression.</p><p><b>CONCLUSION</b>The ratio of a/M can be a standard to evaluate degree of cervical spinal cord compression; the changes of the height of intervertebral disc, curvature of cervical vertebrae in X-ray films maybe have values to predict the degree of cervical spinal cord compression.</p>