Exosomal miR-1307 of osteosarcoma and the proliferation and apoptosis of osteosarcoma cells

doi:10.3969/j.issn.2095-4344.2185

Abstract

Abstract: BACKGROUND: Studies have shown that the occurrence of osteosarcoma is associated with abnormal expression of various microRNAs (miRNAs). Exosomes (Exos) containing miRNAs get much more attentions in intracellular communications.
OBJECTIVE: To investigate the effects of exosomal miR-1307 from osteosarcoma on proliferation and apoptosis of osteosarcoma cells and its mechanism.
METHODS: Human osteosarcoma cell lines (143B, MG63, U2OS, Saos-2 and SW1353) and human normal osteoblastic cell line (hFOB 1.19) were purchased. The expression level of miR-1307 in five kinds of osteosarcoma cell lines and normal osteoblastic cell line was detected by qRT-PCR. Finally, SW1353 cell line was selected for functional verification of osteosarcoma cells. Osteosarcoma cells and normal osteoblasts were cultured. Osteosarcoma exosomes and normal osteoblastic exosomes were extracted by differential centrifugation. Then miR-1307 inhibitor and miR-NC mimic were transfected into SW1353 osteosarcoma cells and normal osteoblasts, and then SW1353 osteosarcoma cell exosomes and normal osteoblast exosomes were extracted. The above exosomes (25 mg/L) were added to SW1353 osteosarcoma cells to intervene for 24 hours. Cell proliferation assay and flow cytometry were used to observe the proliferation and apoptosis of osteosarcoma cells. Targetscan, miRBase and miRDIP databases were used to predict the target genes of miR-1307. The activity of luciferin was assayed by luciferase reporter gene. mRNA and protein expression levels of AGAP1 were assayed by qRT-PCR and western blot assay in osteosarcoma cells.
RESULTS AND CONCLUSION: (1) Compared with hFOB 1.19-exosomes of osteoblasts, SW1353 osteosarcoma cell exosomes significantly promoted the proliferation and inhibited the apoptosis of osteosarcoma cells (P < 0.01). Compared with SW1353 osteosarcoma cell exosomes, the level of miR-1307 in SW1353 osteosarcoma cell exosomes was down-regulated; the proliferation of osteosarcoma cells was significantly decreased but the apoptosis rate was significantly increased (P < 0.01). (2) AGAP1 was verified as a direct target gene of miR-1307. Compared with miR-NC, miR-1307 significantly inhibited the mRNA and protein levels of AGAP1 (P < 0.01). Compared with miR-NC, miR-1307 significantly inhibited the luciferase activity of wild-type PGLO-AGAP1-WT 3'-UTR (P < 0.05). (3) The results show that exosomal miR-1307 of osteosarcoma promoted the proliferation and inhibited the apoptosis of osteosarcoma cells via targeting AGAP1.

Key words: bone, osteosarcoma, exosome, miR-1307, AGAP1, osteoblast, gene, experiment

CLC Number:

Han Fei, Pu Peidong, Ma Qingyuan, Zhu Zhoujun, Wang Mengyu Wang Chao, Shi Chong, Shi Chenhui, Wang Weishan. Exosomal miR-1307 of osteosarcoma and the proliferation and apoptosis of osteosarcoma cells[J]. Chinese Journal of Tissue Engineering Research, 2021, 25(13): 2036-2042.

Figures/Tables 7

References

[1] BROWN HK, TELLEZ-GABRIEL M, HEYMANN D. Cancer stem cells in osteosarcoma. Cancer Lett. 2017;386:189-195.
[2] MORENO F, CACCIAVILLANO W, CIPOLLA M, et al. Childhood osteosarcoma: Incidence and survival in Argentina. Report from the National Pediatric Cancer Registry, ROHA Network 2000-2013. Pediatr Blood Cancer. 2017; 64(10):e26533.
[3] BRICCOLI A, ROCCA M, SALONE M, et al. High grade osteosarcoma of the extremities metastatic to the lung: long-term results in 323 patients treated combining surgery and chemotherapy, 1985-2005. Surg Oncol. 2010;19(4): 193-199.
[4] MORTUS JR, ZHANG Y, HUGHES DP. Developmental pathways hijacked by osteosarcoma. Adv Exp Med Biol. 2014;804:93-118.
[5] POWERS M, ZHANG W, LOPEZ-TERRADA D, et al. The molecular pathology of sarcomas. Cancer Biomark. 2010;9(1-6):475-491.
[6] SHAO XJ, MIAO MH, XUE J, et al. The Down-Regulation of MicroRNA-497 Contributes to Cell Growth and Cisplatin Resistance Through PI3K/Akt Pathway in Osteosarcoma. Cell Physiol Biochem. 2015;36(5):2051-2062.
[7] SHEN K, MAO R, MA L, et al. Post-transcriptional regulation of the tumor suppressor miR-139-5p and a network of miR-139-5p-mediated mRNA interactions in colorectal cancer. FEBS J. 2014;281(16):3609-3624.
[8] XUE J, NIU J, WU J, et al. MicroRNAs in cancer therapeutic response: Friend and foe. World J Clin Oncol. 2014;5(4):730-743.
[9] MA C, ZHAN C, YUAN H, et al. MicroRNA-603 functions as an oncogene by suppressing BRCC2 protein translation in osteosarcoma. Oncol Rep. 2016;35(6): 3257-3264.
[10] LI BL, LU C, LU W, et al. miR-130b is an EMT-related microRNA that targets DICER1 for aggression in endometrial cancer. Med Oncol. 2013;30(1):484.
[11] QIU X, DOU Y. miR-1307 promotes the proliferation of prostate cancer by targeting FOXO3A. Biomed Pharmacother. 2017;88:430-435.
[12] HAN S, ZOU H, LEE JW, et al. miR-1307-3p Stimulates Breast Cancer Development and Progression by Targeting SMYD4. J Cancer. 2019;10(2): 441-448.
[13] SHIMOMURA A, SHIINO S, KAWAUCHI J, et al. Novel combination of serum microRNA for detecting breast cancer in the early stage. Cancer Sci. 2016;107(3): 326-334.
[14] CHEN WT, YANG YJ, ZHANG ZD, et al. MiR-1307 promotes ovarian cancer cell chemoresistance by targeting the ING5 expression. J Ovarian Res. 2017;10(1):1.
[15] CHEN S, WANG L, YAO B, et al. miR-1307-3p promotes tumor growth and metastasis of hepatocellular carcinoma by repressing DAB2 interacting protein. Biomed Pharmacother. 2019;117:109055.
[16] MINCIACCHI VR, FREEMAN MR, DI VIZIO D. Extracellular vesicles in cancer: exosomes, microvesicles and the emerging role of large oncosomes. Semin Cell Dev Biol. 2015;40:41-51.
[17] OHNO S, TAKANASHI M, SUDO K, et al. Systemically injected exosomes targeted to EGFR deliver antitumor microRNA to breast cancer cells. Mol Ther. 2013;21(1): 185-191.
[18] ALVAREZ-ERVITI L, SEOW Y, YIN H, et al. Delivery of siRNA to the mouse brain by systemic injection of targeted exosomes. Nat Biotechnol. 2011;29(4): 341-345.
[19] BRAICU C, TOMULEASA C, MONROIG P, et al. Exosomes as divine messengers: are they the Hermes of modern molecular oncology? Cell Death Differ. 2015; 22(1):34-45.
[20] RAIMONDI L, DE LUCA A, GALLO A, et al. Osteosarcoma cell-derived exosomes affect tumor microenvironment by specific packaging of microRNAs. Carcinogenesis. 2019:bgz130.
[21] WANG JW, WU XF, GU XJ, et al. Exosomal miR-1228 From Cancer-Associated Fibroblasts Promotes Cell Migration and Invasion of Osteosarcoma by Directly Targeting SCAI. Oncol Res. 2019;27(9):979-986.
[22] GONG L, BAO Q, HU C, et al. Exosomal miR-675 from metastatic osteosarcoma promotes cell migration and invasion by targeting CALN1. Biochem Biophys Res Commun. 2018;500(2):170-176.
[23] SHIMBO K, MIYAKI S, ISHITOBI H, et al. Exosome-formed synthetic microRNA-143 is transferred to osteosarcoma cells and inhibits their migration. Biochem Biophys Res Commun. 2014;445(2):381-387.
[24] OTTAVIANI G, JAFFE N. The epidemiology of osteosarcoma. Cancer Treat Res. 2009;152:3-13.
[25] TANG N, SONG WX, LUO J, et al. Osteosarcoma development and stem cell differentiation. Clin Orthop Relat Res. 2008;466(9):2114-2130.
[26] WANG B, QU XL, LIU J, et al. HOTAIR promotes osteosarcoma development by sponging miR-217 and targeting ZEB1. J Cell Physiol. 2019;234(5):6173-6181.
[27] ZHANG S, WANG Y, CHEN S, et al. Silencing of cytoskeleton-associated protein 2 represses cell proliferation and induces cell cycle arrest and cell apoptosis in osteosarcoma cells. Biomed Pharmacother. 2018;106:1396-1403.
[28] SANG W, ZHU L, MA J, et al. Lentivirus-Mediated Knockdown of CTHRC1 Inhibits Osteosarcoma Cell Proliferation and Migration. Cancer Biother Radiopharm. 2016;31(3):91-98.
[29] 崔国宁,刘喜平,虎峻瑞,等.不同来源外泌体与肿瘤发病相关性的研究与进展[J].中国组织工程研究,2020,24(13):2095-2101.
[30] LIN F, YIN HB, LI XY, et al. Bladder cancer cell‑secreted exosomal miR‑21 activates the PI3K/AKT pathway in macrophages to promote cancer progression. Int J Oncol. 2020;56(1):151-164.
[31] DUAN B, SHI S, YUE H, et al. Exosomal miR-17-5p promotes angiogenesis in nasopharyngeal carcinoma via targeting BAMBI. J Cancer. 2019;10(26):6681-6692.
[32] HE L, ZHU W, CHEN Q, et al. Ovarian cancer cell-secreted exosomal miR-205 promotes metastasis by inducing angiogenesis. Theranostics. 2019;9(26):8206-8220.
[33] SHANG D, XIE C, HU J, et al. Pancreatic cancer cell-derived exosomal microRNA-27a promotes angiogenesis of human microvascular endothelial cells in pancreatic cancer via BTG2. J Cell Mol Med. 2020;24(1):588-604.
[34] CHE X, JIAN F, CHEN C, et al. PCOS serum-derived exosomal miR-27a-5p stimulates endometrial cancer cells migration and invasion. J Mol Endocrinol. 2020;64(1):1-12.
[35] NIE Z, STANLEY KT, STAUFFER S, et al. AGAP1, an endosome-associated, phosphoinositide-dependent ADP-ribosylation factor GTPase-activating protein that affects actin cytoskeleton. J Biol Chem. 2002;277(50):48965-48975.
[36] MEURER S, PIOCH S, WAGNER K, et al. AGAP1, a novel binding partner of nitric oxide-sensitive guanylyl cyclase. J Biol Chem. 2004;279(47):49346-49354.
[37] BENDOR J, LIZARDI-ORTIZ JE, WESTPHALEN RI, et al. AGAP1/AP-3-dependent endocytic recycling of M5 muscarinic receptors promotes dopamine release. EMBO J. 2010;29(16):2813-2826.
[38] TSUTSUMI K, NAKAMURA Y, KITAGAWA Y, et al. AGAP1 regulates subcellular localization of FilGAP and control cancer cell invasion. Biochem Biophys Res Commun. 2020;522(3):676-683.