Expression Profile of miR-199a and Its Role in the Regulation of Intestinal Inflammation
Abstract
:Simple Summary
Abstract
1. Introduction
2. Materials and Methods
2.1. Isolation of Jejunal Epithelial Cells
2.2. Genotype Identification
2.3. DSS Treatment
2.4. RNA Extraction and Real-Time PCR
2.5. Histopathological Evaluation
2.6. Intestinal Permeability Test
2.7. Immunofluorescence Staining
2.8. Serum Biochemical Analysis
2.9. ELISA Assay
2.10. Bioinformatics Analysis
2.11. Luciferase Reporter Assay
2.12. Statistical Analysis
3. Results
3.1. Expression Levels of miR-199a in Piglets during the Postweaning Period
3.2. Loss of miR-199a Increased Susceptibility to DSS-Induced Colitis
3.3. Deficiency of miR-199a Exacerbated Immune Response in Mice with DSS Challenge
3.4. Impaired Intestinal Barrier Function in miR-199a Knockout Mice following DSS Treatment
3.5. Identification of Key Pathways in Mice Lacking miR-199a
3.6. Validation of Ndrg1 as a Candidate Target of miR-199a-3p
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
- Liang, H.; Zhang, J.; Zen, K.; Zhang, C.-Y.; Chen, X. Nuclear microRNAs and their unconventional role in regulating non-coding RNAs. Protein Cell 2013, 4, 325–330. [Google Scholar] [CrossRef] [Green Version]
- Saliminejad, K.; Khorram Khorshid, H.R.; Soleymani Fard, S.; Ghaffari, S.H. An overview of microRNAs: Biology, functions, therapeutics, and analysis methods. J. Cell. Physiol. 2019, 234, 5451–5465. [Google Scholar] [CrossRef] [PubMed]
- Stefani, G.; Slack, F.J. Small non-coding RNAs in animal development. Nat. Rev. Mol. Cell Biol. 2008, 9, 219–230. [Google Scholar] [CrossRef] [PubMed]
- Lam, I.K.Y.; Chow, J.X.; Lau, C.S.; Chan, V.S.F. MicroRNA-mediated immune regulation in rheumatic diseases. Cancer Lett. 2018, 431, 201–212. [Google Scholar] [CrossRef] [PubMed]
- Takagi, T.; Naito, Y.; Mizushima, K.; Hirata, I.; Yagi, N.; Tomatsuri, N.; Ando, T.; Oyamada, Y.; Isozaki, Y.; Hongo, H.; et al. Increased expression of microRNA in the inflamed colonic mucosa of patients with active ulcerative colitis. J. Gastroenterol. Hepatol. 2010, 25 (Suppl. S1), S129–S133. [Google Scholar] [CrossRef]
- Singh, U.P.; Murphy, A.E.; Enos, R.T.; Shamran, H.A.; Singh, N.P.; Guan, H.; Hegde, V.L.; Fan, D.; Price, R.L.; Taub, D.D.; et al. miR-155 deficiency protects mice from experimental colitis by reducing T helper type 1/type 17 responses. Immunology 2014, 143, 478–489. [Google Scholar] [CrossRef]
- Tian, Y.; Xu, J.; Li, Y.; Zhao, R.; Du, S.; Lv, C.; Wu, W.; Liu, R.; Sheng, X.; Song, Y.; et al. MicroRNA-31 Reduces Inflammatory Signaling and Promotes Regeneration in Colon Epithelium, and Delivery of Mimics in Microspheres Reduces Colitis in Mice. Gastroenterology 2019, 156, 2281–2296.e6. [Google Scholar] [CrossRef]
- McDaneld, T.G. MicroRNA: Mechanism of gene regulation and application to livestock1. J. Anim. Sci. 2009, 87, E21–E28. [Google Scholar] [CrossRef] [Green Version]
- Liu, H.-C.; Hicks, J.A.; Trakooljul, N.; Zhao, S.-H. Current knowledge of microRNA characterization in agricultural animals. Anim. Genet. 2010, 41, 225–231. [Google Scholar] [CrossRef]
- Wu, F.; Guo, N.J.; Tian, H.; Marohn, M.; Gearhart, S.; Bayless, T.M.; Brant, S.R.; Kwon, J.H. Peripheral blood microRNAs distinguish active ulcerative colitis and Crohn’s disease. Inflamm. Bowel Dis. 2011, 17, 241–250. [Google Scholar] [CrossRef] [Green Version]
- Smith, F.; Clark, J.E.; Overman, B.L.; Tozel, C.C.; Huang, J.H.; Rivier, J.E.F.; Blisklager, A.T.; Moeser, A.J. Early weaning stress impairs development of mucosal barrier function in the porcine intestine. Am. J. Physiol. Liver Physiol. 2010, 298, G352–G363. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Moeser, A.J.; Pohl, C.S.; Rajput, M. Weaning stress and gastrointestinal barrier development: Implications for lifelong gut health in pigs. Anim. Nutr. 2017, 3, 313–321. [Google Scholar] [CrossRef]
- Xiong, X.; Tan, B.; Song, M.; Ji, P.; Kim, K.; Yin, Y.; Liu, Y. Nutritional Intervention for the Intestinal Development and Health of Weaned Pigs. Front. Vet. Sci. 2019, 6, 46. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Tao, X.; Xu, Z.; Men, X. Analysis of Serum microRNA Expression Profiles and Comparison with Small Intestinal microRNA Expression Profiles in Weaned Piglets. PLoS ONE 2016, 11, e0162776. [Google Scholar] [CrossRef] [Green Version]
- Tahamtan, A.; Teymoori-Rad, M.; Nakstad, B.; Salimi, V. Anti-Inflammatory MicroRNAs and Their Potential for Inflammatory Diseases Treatment. Front. Immunol. 2018, 9, 1377. [Google Scholar] [CrossRef] [Green Version]
- Miretti, S.; Lecchi, C.; Ceciliani, F.; Baratta, M. MicroRNAs as biomarkers for animal health and welfare in livestock. Front. Vet Sci. 2020, 7, 578193. [Google Scholar] [CrossRef]
- Tao, X.; Xu, Z. MicroRNA Transcriptome in Swine Small Intestine during Weaning Stress. PLoS ONE 2013, 8, e79343. [Google Scholar] [CrossRef] [Green Version]
- Jang, H.-J.; Lee, S.I. MicroRNA expression profiling during the suckling-to-weaning transition in pigs. J. Anim. Sci. Technol. 2021, 63, 854–863. [Google Scholar] [CrossRef]
- Paraskevi, A.; Theodoropoulos, G.; Papaconstantinou, I.; Mantzaris, G.; Nikiteas, N.; Gazouli, M. Circulating MicroRNA in inflammatory bowel disease. J. Crohn’s Colitis 2012, 6, 900–904. [Google Scholar] [CrossRef]
- Kaliyamoorthy, V.; Jacop, J.P.; Thirugnanasambantham, K.; Ibrahim, H.I.M.; Kandhasamy, S. The synergic impact of lignin and Lactobacillus plantarum on DSS-induced colitis model via regulating CD44 and miR 199a alliance. World J. Microbiol. Biotechnol. 2022, 38, 233. [Google Scholar] [CrossRef] [PubMed]
- Zou, L.; Xiong, X.; Yang, H.; Wang, K.; Zhou, J.; Lv, D.; Yin, Y. Identification of microRNA transcriptome reveals that miR-100 is involved in the renewal of porcine intestinal epithelial cells. Sci. China Life Sci. 2019, 62, 816–828. [Google Scholar] [CrossRef]
- Wang, L.X.; Zhu, F.; Li, J.Z.; Li, Y.L.; Ding, X.Q.; Yin, J.; Xiong, X.; Yang, H.S. Epidermal growth factor promotes intestinal secretory cell differentiation in weaning piglets via Wnt/β-catenin signalling. Animal 2020, 14, 790–798. [Google Scholar] [CrossRef]
- Xiong, X.; Yang, H.; Tan, B.; Yang, C.; Wu, M.; Liu, G.; Kim, S.W.; Li, T.; Li, L.; Wang, J.; et al. Differential expression of proteins involved in energy production along the crypt-villus axis in early-weaning pig small intestine. Am. J. Physiol. Liver Physiol. 2015, 309, G229–G237. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Wu, Z.; Pi, G.; Song, W.; Li, Y. Investigation of the Expression Pattern and Functional Role of miR-10b in Intestinal Inflammation. Animals 2023, 13, 1236. [Google Scholar] [CrossRef] [PubMed]
- Liao, S.; Tang, S.; Chang, M.; Qi, M.; Li, J.; Tan, B.; Gao, Q.; Zhang, S.; Li, X.; Yin, Y.; et al. Chloroquine Downregulation of Intestinal Autophagy to Alleviate Biological Stress in Early-Weaned Piglets. Animals 2020, 10, 290. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- de Groot, N.; Fariñas, F.; Cabrera-Gómez, C.G.; Pallares, F.J.; Ramis, G. Weaning causes a prolonged but transient change in immune gene expression in the intestine of piglets. J. Anim. Sci. 2021, 99, skab065. [Google Scholar] [CrossRef]
- Nejad, C.; Stunden, H.J.; Gantier, M.P. A guide to miRNAs in inflammation and innate immune responses. FEBS J. 2018, 285, 3695–3716. [Google Scholar] [CrossRef] [Green Version]
- Peng, J.; Jiang, J.; Wang, H.; Feng, X.; Dong, X. miR-199a-3p suppresses cervical epithelial cell inflammation by inhibiting the HMGB1/TLR4/NF-κB pathway in preterm birth. Mol. Med. Rep. 2020, 22, 926–938. [Google Scholar] [CrossRef]
- Yu, Y.; Zhou, H.; Xiong, Y.; Liu, J. Exosomal miR-199a-5p derived from endothelial cells attenuates apoptosis and inflammation in neural cells by inhibiting endoplasmic reticulum stress. Brain Res. 2020, 1726, 146515. [Google Scholar] [CrossRef]
- McKenna, L.B.; Schug, J.; Vourekas, A.; McKenna, J.B.; Bramswig, N.C.; Friedman, J.R.; Kaestner, K.H. MicroRNAs Control Intestinal Epithelial Differentiation, Architecture, and Barrier Function. Gastroenterology 2010, 139, 1654–1664.e1. [Google Scholar] [CrossRef] [Green Version]
- Kwon, J.; Lee, C.; Heo, S.; Kim, B.; Hyun, C.-K. DSS-induced colitis is associated with adipose tissue dysfunction and disrupted hepatic lipid metabolism leading to hepatosteatosis and dyslipidemia in mice. Sci. Rep. 2021, 11, 5283. [Google Scholar] [CrossRef] [PubMed]
- Li, Y.; Luan, Y.; Li, J.; Song, H.; Li, Y.; Qi, H.; Sun, B.; Zhang, P.; Wu, X.; Liu, X.; et al. Exosomal miR-199a-5p promotes hepatic lipid accumulation by modulating MST1 expression and fatty acid metabolism. Hepatol. Int. 2020, 14, 1057–1074. [Google Scholar] [CrossRef] [PubMed]
- Wang, H.; Li, W.; Xu, J.; Zhang, T.; Zuo, D.; Zhou, Z.; Lin, B.; Wang, G.; Wang, Z.; Sun, W.; et al. NDRG1 inhibition sensitizes osteosarcoma cells to combretastatin A-4 through targeting autophagy. Cell Death Dis. 2017, 8, 3048. [Google Scholar] [CrossRef] [PubMed]
- Shi, X.-H.; Larkin, J.C.; Chen, B.; Sadovsky, Y. The Expression and Localization of N-Myc Downstream-Regulated Gene 1 in Human Trophoblasts. PLoS ONE 2013, 8, e75473. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Li, X.L.; Zhou, C.Y.; Sun, Y.; Su, Z.Y.; Wang, X.; Jia, E.N.; Zhang, Q.; Jiang, X.F.; Qi, W.Q.; Xu, Y. Bioinformatic analysis of potential candidates for therapy of inflammatory bowel disease. Eur. Rev. Med. Pharmacol. Sci. 2015, 19, 4275–4284. [Google Scholar] [PubMed]
- Sevinsky, C.J.; Khan, F.; Kokabee, L.; Darehshouri, A.; Maddipati, K.R.; Conklin, D.S. NDRG1 regulates neutral lipid metabolism in breast cancer cells. Breast Cancer Res. 2018, 20, 55. [Google Scholar] [CrossRef]
Genes | Primers | Sequences (5′-3′) | Tm (°C) | GC (%) | Reference |
---|---|---|---|---|---|
TNF-α | Forward | CCCTCACACTCAGATCATCTTCT | 59 | 48 | NM_013693.3 |
Reverse | GCTACGACGTGGGCTACAG | 60 | 63 | ||
IL-1β | Forward | ACCTGTCCTGTGTAATGAAAGACG | 61 | 46 | NM_008361.4 |
Reverse | TGGGTATTGCTTGGGATCCA | 59 | 50 | ||
IL-6 | Forward | GCTTAATTACACATGTTCTCTGGGAAA | 60 | 37 | NM_031168.2 |
Reverse | CAAGTGCATCATCGTTGTTCATAC | 59 | 42 | ||
IL-23 | Forward | CACCTCCCTACTAGGACTCAGC | 61 | 59 | NM_031252.2 |
Reverse | TGGGCATCTGTTGGGTCT | 58 | 56 | ||
Claudin-1 | Forward | ACTCCTTGCTGAATCTGAACAGT | 60 | 43 | NM_016674.4 |
Reverse | GGACACAAAGATTGCGATCAG | 58 | 48 | ||
Occludin | Forward | ACTGGGTCAGGGAATATCCA | 57 | 50 | NM_008756.2 |
Reverse | TCAGCAGCAGCCATGTACTC | 60 | 55 | ||
Ndrg1 | Forward | CATTTTGCTGTCTGCCATG | 55 | 47 | NM_008681.2 |
Reverse | CCATGCCAATGACACTCTTG | 57 | 50 | ||
β-actin | Forward | GTGCTATGTTGCTCTAGACTTCG | 59 | 48 | NM_007393.5 |
Reverse | ATGCCACAGGATTCCATACC | 57 | 50 |
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content. |
© 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
Wu, Z.; Yan, Y.; Li, W.; Li, Y.; Yang, H. Expression Profile of miR-199a and Its Role in the Regulation of Intestinal Inflammation. Animals 2023, 13, 1979. https://doi.org/10.3390/ani13121979
Wu Z, Yan Y, Li W, Li Y, Yang H. Expression Profile of miR-199a and Its Role in the Regulation of Intestinal Inflammation. Animals. 2023; 13(12):1979. https://doi.org/10.3390/ani13121979
Chicago/Turabian StyleWu, Zijuan, Yanyun Yan, Wenli Li, Yali Li, and Huansheng Yang. 2023. "Expression Profile of miR-199a and Its Role in the Regulation of Intestinal Inflammation" Animals 13, no. 12: 1979. https://doi.org/10.3390/ani13121979
APA StyleWu, Z., Yan, Y., Li, W., Li, Y., & Yang, H. (2023). Expression Profile of miR-199a and Its Role in the Regulation of Intestinal Inflammation. Animals, 13(12), 1979. https://doi.org/10.3390/ani13121979