CircRNA_Maml2 promotes the proliferation and migration of intestinal epithelial cells after severe burns by regulating the miR-93-3p/FZD7/Wnt/β-catenin pathway

Abstract

Background: Circular RNA (circRNA) plays key regulatory roles in the development of many diseases. However the biological functions and potential molecular mechanisms of circRNA in the injury and repair of intestinal mucosa in mice after severe burns are yet to be elucidated. Methods: Cell counting kit-8 (CCK-8), 5-ethynyl-2′-deoxyuridine (EdU), wound healing and transwell assays were used to detect cell proliferation and migration ability. Real-time quantitative PCR was used to identify the expression of circRNA, microRNA and messenger RNA. Nuclear and cytoplasmic separation experiments were employed to perceive the location of circRNA_Maml2. Finally, in vitro and in vivo experiments were conducted to study the repairing effect of circRNA_Maml2 on the intestinal mucosa of mice after severe burns. Results: When compared with the control group, the expression of circRNA_Maml2 was significantly reduced in the severe burn group. Furthermore, overexpression of circRNA_Maml2 promoted the proliferation and migration of CT26.wt cells in vivo and the repair of damaged intestinal mucosa in vitro. CircRNA_Maml2 acted as a sponge adsorption molecule for miR-93-3p to enhance the expression of frizzled class receptor 7 and activate the downstream Wnt/β-catenin pathway, thereby promoting the repair of the intestinal mucosa. Conclusions: Our findings demonstrate that circRNA_Maml2 regulates the miR-93-3p/FZD7/Wnt/β-catenin pathway and promotes the repair of damaged intestinal mucosa. Hence, circRNA_Maml2 is a potential therapeutic target to promote intestinal mucosal repair.

Keywords

CircRNA_Maml2 / Severe burn / miR-93-3p / FZD7 / Wnt/β-catenin pathway / Proliferation / Migration

References

[1] Wilmore DW, Smith RJ, O'Dwyer ST, Jacobs DO, Ziegler TR, Wang XD. The gut: a central organ after surgical stress. Surgery. 1988; 104: 917-23.

[2] Porter C, Tompkins RG, Finnerty CC, Sidossis LS, Suman OE, Herndon DN. The metabolic stress response to burn trauma: current understanding and therapies. Lancet. 2016; 388: 1417-26.

[3] Zhuang M, Deng Y, Zhang W, Zhu B, Yan H, Lou J, et al. LncRNA Bmp1 promotes the healing of intestinal mucosal lesions via the miR-128-3p/PHF6/PI3K/AKT pathway. Cell Death Dis. 2021; 12: 595.

[4] Magnotti LJ, Deitch EA. Burns, bacterial translocation, gut barrier function, and failure. J Burn Care Rehabil. 2005; 26: 383-91.

[5] Li C, Li Y, Zhuang M, Zhu B, Zhang W, Yan H, et al. Long noncoding RNA H19 act as a competing endogenous RNA of let-7g to facilitate IEC-6 cell migration and proliferation via regulating EGF. J Cell Physiol. 2021; 236: 2881-92.

[6] Li X, Yang L, Chen LL. The biogenesis, functions, and challenges of circular RNAs. Mol Cell. 2018; 71: 428-42.

[7] Sanger HL, Klotz G, Riesner D, Gross HJ, Kleinschmidt AK. Viroids are single-stranded covalently closed circular RNA molecules existing as highly base-paired rod-like structures. Proc Natl Acad Sci U S A. 1976; 73: 3852-6.

[8] Cocquerelle C, Mascrez B, Hetuin D, Bailleul B. Mis-splicing yields circular RNA molecules. FASEB J. 1993; 7: 155-60.

[9] Arnaiz E, Sole C, Manterola L, Iparraguirre L, Otaegui D, Lawrie CH. CircRNAs and cancer: biomarkers and master regulators. Semin Cancer Biol. 2019; 58: 90-9.

[10] Li P, Chen S, Chen H, Mo X, Li T, Shao Y, et al. Using circular RNA as a novel type of biomarker in the screening of gastric cancer. Clin Chim Acta. 2015; 444: 132-6.

[11] Zhang W, Yan H, Deng Y, Lou J, Zhang P, Cui Q, et al. Expression profile and bioinformatics analysis of circular RNA in intestinal mucosal injury and repair after severe burns. Cell Biol Int. 2020; 44: 2570-87.

[12] Jinek M, Doudna JA. A three-dimensional view of the molecular machinery of RNA interference. Nature. 2009; 457: 405-12.

[13] Salmena L, Poliseno L, Tay Y, Kats L, Pandolfi PP. A ceRNA hypothesis: the Rosetta stone of a hidden RNA language? Cell. 2011; 146: 353-8.

[14] Wang Y, Juranek S, Li H, Sheng G, Tuschl T, Patel DJ. Structure of an argonaute silencing complex with a seed-containing guide DNA and target RNA duplex. Nature. 2008; 456: 921-6.

[15] Memczak S, Jens M, Elefsinioti A, Torti F, Krueger J, Rybak A, et al. Circular RNAs are a large class of animal RNAs with regulatory potency. Nature. 2013; 495: 333-8.

[16] Zhang DY, Qiu W, Jin P, Wang P, Sun Y. Role of autophagy and its molecular mechanisms in mice intestinal tract after severe burn. J Trauma Acute Care Surg. 2017; 83: 716-24.

[17] Ahmad A, Druzhyna N, Szabo C. Cystathionine-gamma-lyase deficient mice are protected against the development of multiorgan failure and exhibit reduced inflammatory response during burn. Burns. 2017; 43: 1021-33.

[18] Chen C, Wang P, Su Q, Wang S, Wang F. Myosin light chain kinase mediates intestinal barrier disruption following burn injury. PLoS One. 2012; 7: e34946.

[19] Wang L, Yang G, Zhu X, Wang Z, Wang H, Bai Y, et al. miR-93-3p inhibition suppresses clear cell renal cell carcinoma proliferation, metastasis and invasion. Oncotarget. 2017; 8: 82824-34.

[20] Feng Z, Chen R, Huang N, Luo C. Long non-coding RNA ASMTL-AS1 inhibits tumor growth and glycolysis by regulating the miR-93-3p/miR-660/FOXO1 axis in papillary thyroid carcinoma. Life Sci. 2020; 244: 117298.

[21] Li C, Wang F, Wei B, Wang L, Kong D. LncRNA AWPPH promotes osteosarcoma progression via activation of Wnt/beta-catenin pathway through modulating miR-93-3p/FZD7 axis. Biochem Biophys Res Commun. 2019; 514: 1017-22.

[22] MacDonald BT, Tamai K, He X. Wnt/beta-catenin signaling: components, mechanisms, and diseases. Dev Cell. 2009; 17: 9-26.

[23] Fernandez A, Huggins IJ, Perna L, Brafman D, Lu D, Yao S, et al. The WNT receptor FZD7 is required for maintenance of the pluripotent state in human embryonic stem cells. Proc Natl Acad Sci U S A. 2014; 111: 1409-14.

[24] He W, Wang Y, Wang P, Wang F. Intestinal barrier dysfunction in severe burn injury. Burns Trauma. 2019; 7: 24.

[25] Lachiewicz AM, Hauck CG, Weber DJ, Cairns BA, van Duin D. Bacterial infections after burn injuries: impact of multidrug resistance. Clin Infect Dis. 2017; 65: 2130-6.

[26] Udy AA, Roberts JA, Lipman J, Blot S. The effects of major burn related pathophysiological changes on the pharmacokinetics and pharmacodynamics of drug use: an appraisal utilizing antibiotics. Adv Drug Deliv Rev. 2018; 123: 65-74.

[27] Chen B, Huang S. Circular RNA: an emerging non-coding RNA as a regulator and biomarker in cancer. Cancer Lett. 2018; 418: 41-50.

[28] Qu S, Yang X, Li X, Wang J, Gao Y, Shang R, et al. Circular RNA: a new star of noncoding RNAs. Cancer Lett. 2015; 365: 141-8.

[29] Lasda E, Parker R. Circular RNAs: diversity of form and function. RNA. 2014; 20: 1829-42.

[30] Suzuki H, Tsukahara T. A view of pre-mRNA splicing from RNase R resistant RNAs. Int J Mol Sci. 2014; 15: 9331-42.

[31] Salzman J, Chen RE, Olsen MN, Wang PL, Brown PO. Cell-type specific features of circular RNA expression. PLoS Genet. 2013; 9: e1003777.

[32] Holdt LM, Kohlmaier A, Teupser D. Molecular roles and function of circular RNAs in eukaryotic cells. Cell Mol Life Sci. 2018; 75: 1071-98.

[33] Han D, Li J, Wang H, Su X, Hou J, Gu Y, et al. Circular RNA circMTO1 acts as the sponge of microRNA-9 to suppress hepatocellular carcinoma progression. Hepatology. 2017; 66: 1151-64.

[34] Zhang X, Wang S, Wang H, Cao J, Huang X, Chen Z, et al. Circular RNA circNRIP1 acts as a microRNA-149-5p sponge to promote gastric cancer progression via the AKT1/mTOR pathway. Mol Cancer. 2019; 18: 20.

[35] Wang K, Long B, Liu F, Wang JX, Liu CY, Zhao B, et al. A circular RNA protects the heart from pathological hypertrophy and heart failure by targeting miR-223. Eur Heart J. 2016; 37: 2602-11.

[36] Ng WL,Marinov GK,Chin YM,Lim YY,Ea CK. Transcriptomic analysis of the role of RasGEF1B circular RNA in the TLR4/LPS pathway. Sci Rep. 2017; 7: 12227.

[37] Qian P, Linbo L, Xiaomei Z, Hui P. Circ_0002770, acting as a competitive endogenous RNA, promotes proliferation and invasion by targeting miR-331-3p in melanoma. Cell Death Dis. 2020; 11: 264.

[38] Simonetti M, Agarwal N, Stosser S, Bali KK, Karaulanov E, Kamble R, et al. Wnt-Fzd signaling sensitizes peripheral sensory neurons via distinct noncanonical pathways. Neuron. 2014; 83: 104-21.

[39] Ueno K, Hiura M, Suehiro Y, Hazama S, Hirata H, Oka M, et al. Frizzled-7 as a potential therapeutic target in colorectal cancer. Neoplasia. 2008; 10: 697-705.