Programmed application of extracellular matrix promotes neural differentiation of mouse embryonic stem cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To study the role of extracellular matrix (ECM) in neural differentiation of mouse embryonic stem cells (ESCs).</p><p><b>METHODS</b>Mouse ESCs were incubated in the ESC conditioned medium, and the formation of embryonic bodies (EBs) were induced in bacteriological dishes using high-concentration all-trans retinoic acid (RA). The EBs were seeded on different matrixes (gelatin, fibronectin, and laminin/poly-L-ornithine) to test their impact on neural differentiation of the ESCs using immunofluorescence assay. The effect of laminin/poly-L-ornithine on the growth of neurites was evaluated with fluorescence microscopy.</p><p><b>RESULTS</b>High-concentration RA activated and accelerated the differentiation of ESCs toward nestin-positive neural progenitor cells. Fibronectin supplement in the matrix dose-dependently promoted ESC differentiation into neural progenitor cells, while laminin/poly-L-ornithine increased the growth of the neurites and induced the maturation of the differentiated neural cells.</p><p><b>CONCLUSION</b>ECM plays an important role in neural differentiation of mouse ESCs, and application of FN produces the most conspicuous effect during the differentiation of the ESCs into the neural progenitor cells;laminin/poly-L-ornithine is the most effective during their differentiation into neural cells.</p>

Transfection of embryonic stem cells with green fluorescent protein gene and their differentiation into neural cells / 中国地方病学杂志

Objective To establish embryonic stem cells (ESC) that can express green fluorescent protein (CFP) and differentiate them into neurons. It would provide tagging neurons for clinical transplantation to cure neural system diseases. Methods ESC (R1) was transfeeted with a plasmid containing the GFP by electroporation. A transgeuic cell line was obtained after selection with G418. The ESCs were characterized by AKP staining. Monolayer differentiation method was used to induce neural differentiation derived from GFP-ESC and immunofluorescence method was used to identify Tuj1 positive cells. Results There was no significant difference(X2=3.14,P0.05) in transfect rates between liposome and electroporation (65% vs 79%). The AKP staining of GFP-ESC was positive. GFP-ESC could be differentiated into neural cells. Conclusions These results show that ESC expressing GFP has been estabhshed, which can be differemiated into neurons.

Human neural stem cells promote corticospinal axons regeneration and synapse reformation in injured spinal cord of rats / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Axonal regeneration in lesioned mammalian central nervous system is abortive, and this causes permanent disabilities in individuals with spinal cord injuries. This paper studied the action of neural stem cell (NSC) in promoting corticospinal axons regeneration and synapse reformation in rats with injured spinal cord.</p><p><b>METHODS</b>NSCs were isolated from the cortical tissue of spontaneous aborted human fetuses in accordance with the ethical request. The cells were discarded from the NSC culture to acquire NSC-conditioned medium. Sixty adult Wistar rats were randomly divided into four groups (n = 15 in each): NSC graft, NSC medium, graft control and medium control groups. Microsurgical transection of the spinal cord was performed in all the rats at the T11. The NSC graft group received stereotaxic injections of NSCs suspension into both the spinal cord stumps immediately after transection; graft control group received DMEM injection. In NSC medium group, NSC-conditioned medium was administered into the spinal cord every week; NSC culture medium was administered to the medium control group. Hindlimb motor function was assessed using the BBB Locomotor Rating Scale. Regeneration of biotin dextran amine (BDA) labeled corticospinal tract was assessed. Differentiation of NSCs and the expression of synaptophysin at the distal end of the injured spinal cord were observed under a confocal microscope. Group comparisons of behavioral data were analyzed with ANOVA.</p><p><b>RESULTS</b>NSCs transplantation resulted in extensive growth of corticospinal axons and locomotor recovery in adult rats after complete spinal cord transection, the mean BBB scores reached 12.5 in NSC graft group and 2.5 in graft control group (P < 0.05). There was also significant difference in BBB score between the NSC medium (11.7) and medium control groups (3.7, P < 0.05). BDA traces regenerated fibers sprouted across the lesion site and entered the caudal part of the spinal cord. Synaptophysin expression colocalized with BDA positive axons and neurons distal to the injury site. Transplanted cells were found to migrate into the lesion, but not scatter along the route of axon grows. The cells differentiated into astrocytes or oligodendrocytes, but not into the neurons after transplantation. Furthermore, NSC medium administration did not limit the degree of axon sprouting and functional recovery of the injured rats compared to the NSC graft group.</p><p><b>CONCLUSIONS</b>Human embryonic neural stem cells can promote functional corticospinal axons regeneration and synapse reformation in the injured spinal cord of rats. The action is mainly through the nutritional effect of the stem cells on the spinal cord.</p>

Experimental study on islet cells in rats under condition of three-dimensional microgravity / 中华外科杂志

<p><b>OBJECTIVE</b>To study the survival rate and secretory function of islet cells in rats under condition of three-dimensional microgravity.</p><p><b>METHODS</b>Isolated islet cells were assigned to flask-cultured or bioreactor-cultured. Survival rate of islets cultured for days 3, 7, 14 in stationary flasks or microgravity bioreactors was measured by AO-PI double-staining. Cultured islets were identified by dithizone (DTZ) staining, and insulin contents of different culture liquids were measured by radioimmunoassay.</p><p><b>RESULTS</b>Pancreatic islets stained nacarat with DTZ were easily visualized. When islet cells were cultured for 7 days and 14 days, survival rate of bioreactor-cultured islets was (0.9000 +/- 0.0107)% and 0.8038% +/- 0.0092% and higher than flask-cultured islets (P < 0.01). Insulin level of bioreactor-cultured islets is (70.875 +/- 0.31) m micro /L on the cultured 7 days while flask-cultured islets is (41.246 +/- 0.35) m micro /L. There was statistically significant difference of insulin production between the two groups (P < 0.01). Bioreactor-cultured islet contents were higher than flask-cultured on the cultured 14, 21 and 30 days (P < 0.01).</p><p><b>CONCLUSIONS</b>Islet cells survival rate and secretory function revealed that bioreactor-cultured islets functioned better compared with flask-cultured islets.</p>