Castro Gustavo, Navajas Eduardo, Farah Michel Eid, Maia Mauricio, Rodrigues Eduardo Buchele
Vision Institute, Department of Ophthalmology, Federal University of São Paulo, 04021-001 São Paulo, SP, Brazil.
ISRN Ophthalmol. 2013 Feb 26;2013:752161. doi: 10.1155/2013/752161. eCollection 2013.
Purpose. The purpose of this work was to evaluate the retinal integration and differentiation of neurospheres formed by stem cells and mouse neural progenitor cells injected intravitreally in mice eyes with retinal injury. Methods. Eight male C57BL mice, 8 weeks old, were submitted to intraperitoneal injection of sodium iodate (2% NaIO3, 50 mg/kg). After 72 hours, 2 μ L of solution with mNPC were injected intravitreally (100.000 cells/ μ L). After 7 days, their eyes were dissected and cryoprotected in 30% sucrose in PB for at least 24 hours at 4°C. The material was analyzed by immunohistochemistry and the following primary antibodies evaluation. Results. The results showed that the grafted cells integrated and survived in the adult mice within the sinner retinal tissue for at least 7 days. Immunohistochemical analysis revealed mature neuronal pattern in some regions. The mNPC population in the transplants was tightly surrounded by neuroretinal cells, suggesting their active role in neuron survival. Notably, the appearance of GFP-positive mNPC was not the result of fusion between donor cells and endogenous neuroretinal cells. Conclusions. Migration, survival, and differentiation of mNPCs were observed after 7 days following a single application with neurosphere method. The results may be clinically relevant for future stem cell therapy to restore retinal degeneration.
目的。本研究旨在评估在视网膜损伤的小鼠眼玻璃体内注射干细胞和小鼠神经祖细胞形成的神经球的视网膜整合及分化情况。方法。8只8周龄雄性C57BL小鼠接受腹腔注射碘酸钠(2% NaIO₃,50 mg/kg)。72小时后,将2 μL含小鼠神经祖细胞(mNPC)的溶液(100,000个细胞/μL)玻璃体内注射。7天后,解剖其眼睛并在4℃下于含30%蔗糖的磷酸盐缓冲液(PB)中进行冷冻保护至少24小时。通过免疫组织化学和以下一抗评估对材料进行分析。结果。结果显示,移植的细胞在成年小鼠的视网膜内层组织中整合并存活至少7天。免疫组织化学分析在某些区域揭示了成熟的神经元模式。移植中的mNPC群体被神经视网膜细胞紧密包围,表明它们在神经元存活中发挥积极作用。值得注意的是,绿色荧光蛋白(GFP)阳性的mNPC的出现并非供体细胞与内源性神经视网膜细胞融合的结果。结论。采用神经球方法单次应用后7天观察到了mNPC的迁移、存活和分化。这些结果可能对未来恢复视网膜变性的干细胞治疗具有临床意义。