Authors: Leng Yunxia, Zhang Meng, Wu Min, Ren Guoliang, Gao Zongyin
To investigate the regulation effects of Ngn2 gene transfection on retinal neuron differnetion in three-dimentional optic vesicle (OV) of mice.
OV was cultured in vitro using mouse induced pluripotent stem cells (iPSC) under specific conditions.During OV culture, it was transfected multiple times by lentivirus-mediated Ngn2 gene and then it was induced after maturation.The cells were specificly differentiated toward retinal nerve cells in OV.Using the green fluorescent protein (EGFP) gene as control, the differentiation of retinal nerve cells in OV was detected by immunohistochemistry.Reverse transcription PCR and Western blot were used to quantitatively detect the expressions of retinal neuron-specific proteins Pax6, Islet1 and Brn3b.
The mouse iPS-derived OV was successfully cultured.The number of neural cells in the OV transfected with the Ngn2 gene was increased by the lentiviral-mediated lentivirus.The expressions of PAX6, Islet1 and Brn3b in the Ngn2 transfection group were significantly higher at the gene and protein levels than those in the control group, with significant differences between the two groups (P<0.05).
The Ngn2 gene can effectively increase the number of retinal neuron differentiation in OV and make in vitro cultured OV more mature and form a more perfect retinal cell neural circuit.