BMP2 Modified by the m6A Demethylation Enzyme ALKBH5 in the Ossification of the Ligamentum Flavum Through the AKT Signaling Pathway

Abstract

Ossification of the ligamentum flavum (OLF) is characterized by a process of ectopic bone formation in the ligamentum flavum. The definitive pathophysiology of OLF still remains unclear, but the epigenetic m 6 A modification plays an important role in OLF. In addition, no studies have reported the function of ALKBH5 in OLF development. In this study, we investigated the function of the m 6 A demethylation enzyme ALKBH5 in OLF. To evaluate the function of ALKBH5, OLF tissues and normal ligamentum flavum tissues were collected. In vitro methods, including HE, IHC and western blotting assays, were used to evaluate the association of ALKBH5 with OLF. In addition, we verified the effects of ALKBH5 on osteogenesis using alizarin red and ALP staining. MeRIP q-PCR was performed to investigate the methylation level of BMP2. Moreover, the mechanism of ALKBH5-mediated regulation of the ossification of the ligamentum flavum cells through the AKT signaling pathway was also verified. The present study showed that the expression of ALKBH5 increased in OLF tissues. The overexpression of ALKBH5 increased the expression of osteogenic genes and promoted the ossification of ligamentum flavum cells. Furthermore, BMP2 was significantly enriched in the ligamentum flavum cells of the anti-m 6 A group compared with those of the IgG group. The overexpression of ALKBH5 led to the activation of p-AKT, and BMP2 was regulated by ALKBH5 through the AKT signaling pathway. ALKBH5 promoted the osteogenesis of the ligamentum flavum cells through BMP2 demethylation and AKT activation. ALKBH5 was shown to be an important demethylation enzyme in OLF development.

The biological function of m6A demethylase ALKBH5 and its role in human disease

“…ALKBH5 was increased in ossification of the ligamentum flavum (OLF) tissues, and promoted the ossification of the ligamentum flavum cells through BMP2 demethylation and activating AKT signaling pathway [14]. The METTL3-mediated m6A methylation was found to be dynamically reversed by the demethylase ALKBH5, which acted to reverse the suppression of METTL3 in osteogenic progression, partly though regulating expression of MYD88 and facilitating the activation of NF-κB which was a well-known repressor of osteogenesis [28].…”

“…For example, ALKBH5 acted to influence spermatogenesis and trophoblast invasion, thus affecting human reproductive system diseases [23,29]. In addition, ALKBH5 also played an important role in the ossification of the ligamentum flavum cells and osteogenic progression via some famous signaling pathways, such as AKT and NF-κB signaling pathway [14,28].…”

“…Human AlkB homolog H5 (ALKBH5), was a famous m6A demethylase (also called m6A eraser), which had spawned significant biological and pharmacological interest among researchers [ 10 ]. Recently, the effects of ALKBH5 on many biological processes had been demonstrated, including proliferation [ 11 ], invasion [ 12 ], metastasis [ 13 ], ossification [ 14 ] and so on. In addition, ALKBH5 was also involved in kinds of cancers or non-cancers, such as glioblastoma [ 15 ], pancreatic cancer [ 16 ], colon cancer [ 17 ], breast cancer [ 18 ], gastric cancer [ 19 ], lung cancer [ 20 ], ovarian cancer [ 21 ], diabetes [ 22 ] and reproductive system diseases [ 23 ] (Figs.…”

Screening and Identifying m6A Regulators in Pancreatic Cancer Based on The Cancer Genome Atlas Database

“…Additionally, signaling associated with multiple cancers is dysregulated in PAAD development. We found that in patients with PAAD, a high ALKBH5 mRNA expression level was associated with the activation of AKT signaling pathways, which participate in important cellular pathological processes in PAAD development [27], suggesting that the mRNAs of molecules in the AKT pathway may be the m6a modi cation target mediated by ALKBH5 [28]. Recently, a study has shown that ALKBH5 functions as an anti-tumor protein in pancreatic cancer progression [29]; in this paper, upregulated ALKBH5 sensitized pancreatic cancer to gemcitabine-chemotherapy, and knockdown of ALKBH5 decreased pancreatic cancer cell invasion, migration, proliferation, metastasis, and tumorigenesis.…”

Screening and Identifying m6A Regulators as an Independent Prognostic Biomarker in Pancreatic Cancer Based on The Cancer Genome Atlas Database

“…Additionally, signaling associated with multiple cancers is dysregulated in PAAD development. We found that in patients with PAAD, a high ALKBH5 mRNA expression level was associated with the activation of AKT signaling pathways, which participate in important cellular pathological processes in PAAD development [ 27 ], suggesting that the mRNAs of molecules in the AKT pathway may be the m6a modification target mediated by ALKBH5 [ 28 ]. Recently, a study has shown that ALKBH5 functions as an antitumor protein in pancreatic cancer progression [ 29 ]; in this paper, upregulated ALKBH5 sensitized pancreatic cancer to gemcitabine chemotherapy, and knockdown of ALKBH5 decreased pancreatic cancer cell invasion, migration, proliferation, metastasis, and tumorigenesis [ 29 ].…”