Han Qin1, Jun Cai2. 1. Department of Stomatology, The Lianyungang Affiliated Hospital of Xuzhou Medical University, #182 Tongguan Road, Liangyungang 222002, Jiangsu Province, China. Electronic address: qinhan2005@163.com. 2. Department of Anesthesia, The Maternal and Child Health Care Hospital of Lianyungang City, Lianyungang, #10 Cangwu Road, 222002, Jiangsu Province, China.
Abstract
OBJECTIVE: This study aimed to investigate the effect of Runx2 silencing on autophagy and RANKL expression in mouse osteoblasts, and provide an experimental basis to assess obstacles in dental eruption. METHODS: In accordance with previously reported methods, LVpFU-GW-016PSC60109-1 virus was used to transfect mouse osteoblasts (MOI = 40). Target gene expression was assessed via cytometer, and the effect of silencing Runx2 was assessed via a two-step quantitative real-time polymerase chain reaction (qRT-PCR)-based method. Western blotting was performed to assess LC3, Beclin-1 and RANKL expression. RESULTS: As confirmed via qRT-PCR analysis, Runx2 was efficiently silenced in the experimental group (>90% efficiency). Western blotting revealed that LC3 and RANKL proteins were significantly down -regulated in the experimental group (group KD), their expression levels being particularly lower than those in the control group (group NC). However, Beclin-1 protein expression was not significantly different from that of the control. CONCLUSION: Upon Runx2 silencing, autophagy-related proteins and RANKL were repressed in osteoblasts, thereby potentially causing the tooth eruption disorder.
OBJECTIVE: This study aimed to investigate the effect of Runx2 silencing on autophagy and RANKL expression in mouse osteoblasts, and provide an experimental basis to assess obstacles in dental eruption. METHODS: In accordance with previously reported methods, LVpFU-GW-016PSC60109-1 virus was used to transfect mouse osteoblasts (MOI = 40). Target gene expression was assessed via cytometer, and the effect of silencing Runx2 was assessed via a two-step quantitative real-time polymerase chain reaction (qRT-PCR)-based method. Western blotting was performed to assess LC3, Beclin-1 and RANKL expression. RESULTS: As confirmed via qRT-PCR analysis, Runx2 was efficiently silenced in the experimental group (>90% efficiency). Western blotting revealed that LC3 and RANKL proteins were significantly down -regulated in the experimental group (group KD), their expression levels being particularly lower than those in the control group (group NC). However, Beclin-1 protein expression was not significantly different from that of the control. CONCLUSION: Upon Runx2 silencing, autophagy-related proteins and RANKL were repressed in osteoblasts, thereby potentially causing the tooth eruption disorder.
Authors: Azadeh Montaseri; Claudia Giampietri; Michela Rossi; Anna Riccioli; Andrea Del Fattore; Antonio Filippini Journal: Biomolecules Date: 2020-09-30