1.Effect of BMP9 over-expression on migration and invasion of human lung squamous-cell carcinoma NCI-H520 cells
Jing WANG ; Fang DENG ; Ya LI ; Mengtian FAN ; Yangliu GUO ; Qiong SHI
Chinese Journal of Pathophysiology 2015;(11):1921-1927
AIM:To investigate the effect of bone morphogenetic proteins 9 (BMP9) on the migration and in-vasion abilities of human lung squamous-cell carcinoma NCI-H520 cells and its mechanism.METHODS:The expression of BMP9 at mRNA and protein levels in the NCI-H520 cells and human bronchial epithelial ( HBE) cells was detected by RT-PCR and Western blot.The NCI-H520 cells were transfected with the recombinant adenovirus AdBMP9 and the expres-sion of BMP9 at mRNA and protein levels was validated by RT-PCR and Western blot.The migration and invasion abilities of the NCI-H520 cells were determined by wound-healing and Transwell assays.The mRNA and protein levels of the migra-tion-related factor matrix metalloproteinase 2 (MMP2) were detected by RT-PCR and Western blot.The level of phospho-rylated Smad1/5 (p-Smad1/5) was detected by Western blot.Meanwhile, NCI-H520 cells were treated with BMP specific antagonist AdNoggin and AdBMP9.The level of p-Smad1/5 and the cell migration ability were measured by Western blot, wound-healing and Transwell assays.RESULTS:The expression of BMP9 at mRNA and protein levels was lower in NCI-H520 cells than that in HBE cells.After AdBMP9 was stably transfected into the NCI-H520 cells, the expression of BMP9 at mRNA and protein levels was significantly up-regulated, cell migration and invasion abilities were significantly de-creased, and the mRNA and protein levels of MMP2 were decreased.Meanwhile, the level of p-Smad1/5 was increased. Noggin reversed BMP9-caused the increase in p-Smad1/5 and the decrease in cell migration ability.CONCLUSION:O-ver-expression of BMP9 inhibits the migration and invasion abilities of lung squamous-cell carcinoma NCI-H520 cells.The activation of BMP-Smad signaling pathway may be involved in this inhibitory process.
2.LncRNA HAGLR activates RUNX2 and inhibits NLRP3 inflammasome to promote tibial fracture healing
Wen Wang ; Xinyu Chen ; Ziyi Huang ; Yangliu Deng ; Hongwang Cui
Acta Universitatis Medicinalis Anhui 2023;58(5):830-837
Objective:
To study the effect of long non⁃coding RNA (LncRNA) HAGLR on the expression of NODlike receptor pyrin domain⁃associated protein 3 ( NLRP3 ) inflammasome and fracture healing in tibial fracture (TF) mice and to explore the mechanism .
Methods :
First , HAGLR in osteoblast MC3T3 ⁃E1 was silenced by in vitro . Cell viability was detected by CCK⁃8 assay , cell apoptosis was detected by TUNEL assay , and the expressions of bone alkaline phosphatase (BALP) and osteocalcin were detected by qPCR . Western blot assay was used to detect the expressions of RUNX2 , phosphorylated RUNX2 ( p ⁃RUNX2 ) , NLRP3 , cysteine aspartic protease 1 (Caspase1) , apoptosis⁃associated spot⁃like protein ( ASC) and interleukin⁃1β . TF mouse models were established by tibial fracture operation in mice . HAGLR was overexpressed in the model mice , and RUNX2 was silenced or an inflammatory body inhibitor MCC950 was added on the basis of overexpression of HAGLR . The expressions of HAGLR and RUNX2 were detected by qPCR , and the expressions of insulin⁃like growth factor (IGF⁃1) were detected by Western blot . microCT was used to measure the volume of mouse callus (MBV) and the total tibial wet weight .
Results :
The apoptosis rate of MC3T3 ⁃E1 cells increased and the expression levels of RUNX2 ,p⁃RUNX2 , BALP and osteocalcin decreased ( P < 0. 05 ) . The expressions of NLRP3 , Caspase1 , ASC and IL⁃1β increased ( P <0. 05) . Compared with healthy tissue , the expressions of HAGLR and RUNX2 in TF mice decreased . Overexpression of HAGLR promoted the expressions of HAGLR and RUNX2 in TF mice , and increased the expression of MBV and tibia wet weight and IGF⁃1 (P < 0. 05) . Silencing RUNX2 on the basis of overexpression of HAGLR resulted in decreased expression of MBV , tibial wet weight and IGF⁃1 in TF mice (P < 0. 05) . However , the addition of NLRP3 inflammasome inhibitor MCC950 on top of the overexpression of HAGLR resulted in increased expressions of MBV , full⁃length tibia wet weight and IGF⁃1 (P < 0. 05) .
Conclusion
LncRNA HAGLR promotes the healing of tibial fractures by activating RUNX2 and inhibiting NLRP3 inflammasome .