Effects of let-7a on proliferation, osteogenic differentiation and apoptosis of human dental pulp stem cells

doi:10.19439/j.sjos.2023.05.004

Abstract

Abstract: PURPOSE: To study the effect and possible mechanism of let-7a on proliferation, differentiation and apoptosis of human dental pulp stem cell (hDPSCs). METHODS: The cells were divided into four groups: overexpression control (let-7a control/let-7a agomir control), overexpression let-7a (let-7a mimics/let-7a agomir), knockdown let-7a control (let-7a inhibitor control) and knockdown let-7a (let-7a inhibitor). Cell counting kit-8 assay(CCK-8) was used to detect the proliferation of cells at 24 hours, 48 hours and 72 hours after transfection. Calcified nodules were detected by Alizarin red staining. The protein expression of alkaline phosphatase (ALP), osteopontin (OPN), 4E-binding protein 1 (4EBP1), p-4EBP1, mammalian target of rapamycin (mTOR) and p-mTOR were detected by Western blot. Annexin V-APC/7-AAD cell apoptosis detection kit was used to detect the level of apoptosis after transfection. Statistical analysis was performed using GraphPad Prism 5.0 software. RESULTS: Let-7a inhibited proliferation of hDPSCs and promoted odontoblast differentiation and apoptosis. Let-7a down-regulated the expression of 4EBP1, p-4EBP1, mTOR and p-mTOR. CONCLUSIONS: Let-7a may inhibit proliferation of hDPSCs and promote their differentiation and apoptosis by inhibiting mTOR-4EBP1 molecular pathway.

Key words: Let-7a, Human dental pulp stem cells, Proliferation, Differentiation, Apoptosis

CLC Number:

R781.3

LIU Yan, SHAN Dan-ni, SUN Jing, ZOU Yu-xi, YUAN Chang-yong. Effects of let-7a on proliferation, osteogenic differentiation and apoptosis of human dental pulp stem cells[J]. Shanghai Journal of Stomatology, 2023, 32(5): 468-474.

References

[1] Gronthos S, Mankani M, Brahim J, et al.Postnatal human dental pulp stem cells (DPSCs) in vitro and in vivo[J]. Proc Natl Acad Sci USA, 2000, 97(25): 13625-13630.
[2] Moussa DG, Aparicio C.Present and future of tissue engineering scaffolds for dentin-pulp complex regeneration[J]. J Tissue Eng Regen Med, 2019, 13(1): 58-75.
[3] Chang K, Chen RS, Chang FH, et al.Promoting dentinogenesis of DPSCs through inhibiting microRNA-218 by using magnetic nanocarrier delivery[J]. J Formos Med Assoc, 2019, 118(6): 1005-1013.
[4] Huang X, Xu S, Gao J, et al.miRNA expression profiling identifies DSPP regulators in cultured dental pulp cells[J]. Int J Mol Med, 2011, 28(4): 659-667.
[5] Liu F, Wang X, Yang Y, et al.The suppressive effects of miR-508-5p on the odontogenic differentiation of human dental pulp stem cells by targeting glycoprotein non-metastatic melanomal protein B[J]. Stem Cell Res Ther, 2019, 10(1): 35-45.
[6] Yang ZY, Wang Y, Liu Q, et al.microRNA cluster MC-let-7a-1~let-7d promotes autophagy and apoptosis of glioma cells by down-regulating STAT3[J]. CNS Neurosci Ther, 2020, 26(3): 319-331.
[7] Ma W, Dou Q, Ha X.Let-7a-5p inhibits BMSCs osteogenesis in postmenopausal osteoporosis mice[J]. Biochem Biophys Res Commun, 2019, 510(1): 53-58.
[8] Xie Z, Shen Z, Zhan P, et al.Functional dental pulp regeneration: basic research and clinical translation[J]. Int J Mol Sci, 2021, 22(16): 8991-9018.
[9] Nixdorf DR, Moana-filho EJ, Law AS, et al. Frequency of nonodontogenic pain after endodontic therapy: a systematic review and meta-analysis[J]. J Endod, 2010, 36(9): 1494-1498.
[10] Moussa DG, Aparicio C.Present and future of tissue engineering scaffolds for dentin-pulp complex regeneration[J]. J Tissue Eng Regen Med, 2019, 13(1): 58-75.
[11] 王付燕, 杜立群. 牙髓干细胞在组织工程及再生医学中的应用研究进展[J]. 解剖学杂志, 2020, 43(3): 231-235.
[12] Piva E, Tarlé SA, Nör JE, et al.Dental pulp tissue regeneration using dental pulp stem cells isolated and expanded in human serum[J]. J Endod, 2017, 43(4): 568-574.
[13] Nakashima M, Iohara K.Regeneration of dental pulp by stem cells[J]. Adv Dent Res, 2011, 23(3): 313-319.
[14] Nakashima M, Iohara K, Murakami M, et al.Pulp regeneration by transplantation of dental pulp stem cells in pulpitis: a pilot clinical study[J]. Stem Cell Res Ther, 2017, 8(1): 61-74.
[15] Wang BG, Jiang LY, Xu Q. A comprehensive evaluation for polymorphisms in let-7 family in cancer risk and prognosis: a system review and meta-analysis[J]. Biosci Rep, 2018, 38(4): BSR20180273.
[16] Liu B, Liu Y, Zhou M, et al.Comprehensive ceRNA network analysis and experimental studies identify an IGF2-AS/miR-150/IGF2 regulatory axis in colorectal cancer[J]. Pathol Res Pract, 2020, 216(10): 153104.
[17] Wang T, Zhu H, Yang S, et al.Let-7a-5p may participate in the pathogenesis of diabetic nephropathy through targeting HMGA2[J]. Mol Med Rep, 2019, 19(5): 4229-4237.
[18] 王丽, 张静, 张洪宇, 等. Let-7a调控自身免疫性甲状腺炎发生发展及其机制研究[J]. 中西医结合心血管病电子杂志, 2020, 8(22): 182-182.
[19] Johnson CD, Esquela-kerscher A, Stefani G, et al. The let-7 microRNA represses cell proliferation pathways in human cells[J]. Cancer Res, 2007, 67(16): 7713-7722.
[20] Chen Y, Qiao L, Zhang Z, et al.Let-7a inhibits proliferation and promotes apoptosis of human asthmatic airway smooth muscle cells[J]. Exp Ther Med, 2019, 17(5): 3327-3334.
[21] Sui C, Zhang L, Hu Y.MicroRNA-let-7a inhibition inhibits LPS-induced inflammatory injury of chondrocytes by targeting IL6R[J]. Mol Med Rep, 2019, 20(3): 2633-2640.
[22] Ranjbarnejad F, Nadri S, Biglari A, et al.Effect of let-7a overexpression on the differentiation of conjunctiva mesenchymal stem cells into photoreceptor-like cells[J]. Iran J Basic Med Sci, 2019, 22(8): 878-883.
[23] Ma W, Dou Q, Ha X.Let-7a-5p inhibits BMSCs osteogenesis in postmenopausal osteoporosis mice[J]. Biochem Biophys Res Commun, 2019, 510(1): 53-58.
[24] 杨鹏宇, 张建生. mTOR抑制剂治疗胶质瘤的研究进展[J]. 国际神经病学神经外科学杂志, 2009, 36(6): 548-551.