Kinase inhibit region of SOCS3 attenuates IL6-induced proliferation and astrocytic differentiation of neural stem cells via cross talk between signaling pathways.

AIMS: Efficiency of neural stem cells (NSCs) therapy for brain injury is restricted by astrogliosis around the damaged region, in which JAK2/STAT3 signaling plays a key role. The SOCS3 that can directly inhibit JAK/STAT3 pathway. Here, we investigated the effects of a fusion peptide that combined kinase inhibitory region (KIR) of SOCS3 and virus trans-activator of transcription (TAT) on biological behavior of cultured NSCs under inflammatory conditions. METHODS: NSCs were isolated from embryonic brain of SD rats, TAT-KIR was synthesized, and penetration rate was evaluated by flow cytometry (FACS). CCK8, immunostaining, and FACS were used to detected of TAT-KIR on the proliferation of NSCs. The expressions of GFAP and ß tubulin III positive cells induced by IL6 with/without TAT-KIR were examined by immunostaining and Western blotting to observe the NSCs differentiation, and the effect of TAT-KIR on signaling cross talk was observed by Western blotting. RESULTS: Penetration rate of TAT-KIR into primary cultured NSCs was up to 94%. TAT-KIR did not affect the growth and viability of NSCs. It significantly reduced the NSCs proliferation that enhanced by IL-6 stimulation via blocking the cell cycle progression from the G0/G1 to S phase. In addition, TAT-KIR attenuated astrocytic differentiation and kept high level of neuronal differentiation derived from IL-6-induced NSCs. The fate of NSCs differentiation under inflammatory conditions was affected by TAT-KIR, which was associated with synchronous inhibition of STAT3 and AKT, while promoting JNK expression. CONCLUSION: TAT-KIR mimetic of SOCS3 could be a promising approach for brain repair via regulating the biological behaviors of exogenous NSCs.

[DNA aptamer selection in vitro for determining ketamine by FluMag-SELEX].

OBJECTIVE: To select specific DNA aptamer for determining ketamine by FluMag-SELEX. METHODS: Based on magnetic beads with tosyl surface modification as solid carrier and ketamine as target, a random ssDNA library with total length of 78 bp in vitro was compounded. After 13 rounds screening, DNA cloning and sequencing were done. Primary and secondary, structures were analyzed. The affinity, specificity and Kd values of selected aptamer were measured by monitoring the fluorescence intensity. RESULTS: Two ssDNA aptamers (Apt#4 and Apt#8) were successfully selected with high and specific abilities to bind ketamine as target with Kd value of 0.59 and 0.66 µmol/L. The prediction of secondary structure was main stem-loop and G-tetramer. The stem was the basis of stability of aptamer's structure. And loop and G-tetramer was the key of specific binding of ketamine. CONCLUSION: FluMag-SELEX can greatly improve the selection efficiency of the aptamer, obtain the ketamine-binding DNA aptamer, and develop a new method for rapid detection of ketamine.