Effects of icariin on proliferation and differentiation of MC3T3-E1 cells in an inflammatory environment

doi:10.12307/2024.455

Abstract

Abstract: BACKGROUND: Studies have shown that chronic apical periodontitis is one of the common inflammatory bone destruction diseases. Icariin can promote osteogenic differentiation, inhibit bone resorption, and may play a protective role in bone destruction caused by chronic apical periodontitis.
OBJECTIVE: To investigate the effect of icariin on the proliferation and differentiation of MC3T3-E1 cells in the inflammatory environment stimulated by lipopolysaccharides.
METHODS: Lipopolysaccharides were used to stimulate MC3T3-E1 cells to establish an inflammatory environment in vitro, and cell counting kit-8 was used to detect the best concentration and optimal action time of lipopolysaccharides. Cell counting kit-8 was used to detect the optimal concentration of icariin under the stimulation of lipopolysaccharides at a concentration of 1 μg/mL. Alkaline phosphatase detection, Real-time PCR and western blot assay were used to detect the effect of icariin on osteogenic differentiation of MC3T3-E1 cells in the inflammatory environment. Real-time PCR and western blot were used to detect the effects of icariin on the expression of interleukin-1β and interleukin-6 in MC3T3-E1 cells in the lipopolysaccharide-stimulated inflammatory environment.
RESULTS AND CONCLUSION: Cell counting kit-8 results showed that the optimal concentration of icariin was 0.1 μg/mL. In the inflammatory environment, icariin enhanced the expression of alkaline phosphatase and promoted osteoblast differentiation. Compared with the lipopolysaccharide group, the expression of osteogenesis-related factors alkaline phosphatase and Runx2 was increased in the lipopolysaccharide+icariin group. Compared with the lipopolysaccharide group, the expression levels of inflammation-related factors interleukin-1β and interleukin-6 decreased in the lipopolysaccharide+icariin group. To conclude, lipopolysaccharides weaken the osteogenic ability of MC3T3-E1 cells and aggravate the inflammatory response, but icariin has a protective effect on them.

Key words: MC3T3-E1 cell, icariin, chronic apical periodontitis, lipopolysaccharide, osteogenic differentiation

CLC Number:

Han Yue, Wang Yufei, Liu Wanqing, Dong Ming, Niu Weidong. Effects of icariin on proliferation and differentiation of MC3T3-E1 cells in an inflammatory environment[J]. Chinese Journal of Tissue Engineering Research, 2024, 28(23): 3709-3714.

Figures/Tables 5

References

[1] CAVALLA F, LETRA A, SILVA RM, et al. Determinants of Periodontal/Periapical Lesion Stability and Progression. J Dent Res. 2021;100(1):29-36.
[2] MÖLLER AJ, FABRICIUS L, DAHLÉN G, et al. Influence on periapical tissues of indigenous oral bacteria and necrotic pulp tissue in monkeys. Scand J Dent Res. 1981;89(6):475-484.
[3] DAHLÉN G. Microbiology and treatment of dental abscesses and periodontal-endodontic lesions. Periodontol 2000. 2002;28:206-239.
[4] LIN X, CHI D, MENG Q, et al. Single-Cell Sequencing Unveils the Heterogeneity of Nonimmune Cells in Chronic Apical Periodontitis. Front Cell Dev Biol. 2022;9:820274.
[5] GALLER KM, WEBER M, KORKMAZ Y, et al. Inflammatory Response Mechanisms of the Dentine-Pulp Complex and the Periapical Tissues. Int J Mol Sci. 2021;22(3):1480.
[6] LUO X, WAN Q, CHENG L, et al. Mechanisms of bone remodeling and therapeutic strategies in chronic apical periodontitis. Front Cell Infect Microbiol. 2022;12:908859.
[7] YU W, ZHONG L, YAO L, et al. Bone marrow adipogenic lineage precursors promote osteoclastogenesis in bone remodeling and pathologic bone loss. J Clin Invest. 2021;131(2):e140214.
[8] WEI W, LI J, LIU X, et al. Inhibition of RGS10 Aggravates Periapical Periodontitis via Upregulation of the NF-κB Pathway. J Endod. 2022; 48(10):1308-1318.
[9] HAJISHENGALLIS G. New developments in neutrophil biology and periodontitis. Periodontol 2000. 2020;82(1):78-92.
[10] TAN X, HUANG D, ZHOU W, et al. Dickkopf-1 may regulate bone coupling by attenuating wnt/β-catenin signaling in chronic apical periodontitis. Arch Oral Biol. 2018;86:94-100.
[11] XU R, GUO D, ZHOU X, et al. Disturbed bone remodelling activity varies in different stages of experimental, gradually progressive apical periodontitis in rats. Int J Oral Sci. 2019;11(3):27.
[12] WANG X, QUINN PJ. Lipopolysaccharide: Biosynthetic pathway and structure modification. Prog Lipid Res. 2010;49(2):97-107.
[13] WITTGOW WC JR, SABISTON CB JR. Microorganisms from pulpal chambers of intact teeth with necrotic pulps. J Endod. 1975;1(5):168-171.
[14] FELIPE PEREIRA R, WILLIAN LATTARI TESSARIN G, YAMAMOTO CHIBA F, et al. Apical periodontitis promotes insulin resistance and alters adaptive immunity markers in rats. Saudi Dent J. 2021;33(8):979-986.
[15] LOMBARDO BEDRAN TB, MARCANTONIO RA, SPIN NETO R, et al. Porphyromonas endodontalis in chronic periodontitis: a clinical and microbiological cross-sectional study. J Oral Microbiol. 2012;4. doi: 10.3402/jom.v4i0.10123.
[16] WANG R, YANG D, YU YQ, et al. Effect of inhibition of Porphyromonas endodontalis on osteoblast differentiation. Shanghai Kou Qiang Yi Xue. 2021;30(4):350-354.
[17] FU X, SUN X, ZHANG C, et al. Genkwanin Prevents Lipopolysaccharide-Induced Inflammatory Bone Destruction and Ovariectomy-Induced Bone Loss. Front Nutr. 2022;9:921037.
[18] YANG D, LI R, QIU LH, et al. Effects of lipopolysaccharides extracted from Porphyromonas endodontalis on the expression of IL-1beta mRNA and IL-6 mRNA in osteoblasts. Shanghai Kou Qiang Yi Xue. 2009;18(2):194-197.
[19] FANG C, HE M, LI D, et al. YTHDF2 mediates LPS-induced osteoclastogenesis and inflammatory response via the NF-κB and MAPK signaling pathways. Cell Signal. 2021;85:110060.
[20] HONG C, YANG L, ZHANG Y, et al. Epimedium brevicornum Maxim. Extract exhibits pigmentation by melanin biosynthesis and melanosome biogenesis/transfer. Front Pharmacol. 2022;13:963160.
[21] LUO Z, DONG J, WU J. Impact of Icariin and its derivatives on inflammatory diseases and relevant signaling pathways. Int Immunopharmacol. 2022;108:108861.
[22] MING LG, CHEN KM, XIAN CJ. Functions and action mechanisms of flavonoids genistein and icariin in regulating bone remodeling. J Cell Physiol. 2013;228(3):513-521.
[23] KARSENTY G, WAGNER EF. Reaching a genetic and molecular understanding of skeletal development. Dev Cell. 2002;2(4):389-406.
[24] TAKAYANAGI H. Osteoimmunology: shared mechanisms and crosstalk between the immune and bone systems. Nat Rev Immunol. 2007;7(4): 292-304.
[25] LI Y, LING J, JIANG Q. Inflammasomes in Alveolar Bone Loss. Front Immunol. 2021;12:691013.
[26] ALQURIA TA, ALFIRDOUS RA, GUPTA S, et al. Comparison of conventional and contemporary root canal disinfection protocols against bacteria, lipoteichoic acid (LTA), and lipopolysaccharide (LPS). Sci Rep. 2023;13(1):1206.
[27] SUDO H, KODAMA HA, AMAGAI Y, et al. In vitro differentiation and calcification in a new clonal osteogenic cell line derived from newborn mouse calvaria. J Cell Biol. 1983;96(1):191-198.
[28] TIAGO DM, CANCELA ML, LAIZÉ V. Proliferative and mineralogenic effects of insulin, IGF-1, and vanadate in fish osteoblast-like cells. J Bone Miner Metab. 2011;29(3):377-382.
[29] NIU T, ROSEN CJ. The insulin-like growth factor-I gene and osteoporosis: a critical appraisal. Gene. 2005;361:38-56.
[30] KITASE Y, PRIDEAUX M. Targeting osteocytes vs osteoblasts. Bone. 2023;170:116724.
[31] CAO FY, LONG Y, WANG SB, et al. Fluorescence light-up AIE probe for monitoring cellular alkaline phosphatase activity and detecting osteogenic differentiation. J Mater Chem B. 2016;4(26):4534-4541.
[32] VIMALRAJ S. Alkaline phosphatase: Structure, expression and its function in bone mineralization. Gene. 2020;754:144855.
[33] SILLER AF, WHYTE MP. Alkaline Phosphatase: Discovery and Naming of Our Favorite Enzyme. J Bone Miner Res. 2018;33(2):362-364.
[34] KOMORI T. Molecular Mechanism of Runx2-Dependent Bone Development. Mol Cells. 2020;43(2):168-175.
[35] INADA M, YASUI T, NOMURA S, et al. Maturational disturbance of chondrocytes in Cbfa1-deficient mice. Dev Dyn. 1999;214(4):279-290.
[36] KOMORI T, YAGI H, NOMURA S, et al. Targeted disruption of Cbfa1 results in a complete lack of bone formation owing to maturational arrest of osteoblasts. Cell. 1997;89(5):755-764.
[37] KOMORI T. Regulation of Proliferation, Differentiation and Functions of Osteoblasts by Runx2. Int J Mol Sci. 2019;20(7):1694.
[38] MALKOV MI, LEE CT, TAYLOR CT. Regulation of the Hypoxia-Inducible Factor (HIF) by Pro-Inflammatory Cytokines. Cells. 2021;10(9):2340.
[39] LOPEZ-CASTEJON G, BROUGH D. Understanding the mechanism of IL-1β secretion. Cytokine Growth Factor Rev. 2011;22(4):189-195.
[40] ZHANG H, LIU L, JIANG C, et al. MMP9 protects against LPS-induced inflammation in osteoblasts. Innate Immun. 2020;26(4):259-269.
[41] ZHOU M, GUO M, SHI X, et al. Synergistically Promoting Bone Regeneration by Icariin-Incorporated Porous Microcarriers and Decellularized Extracellular Matrix Derived From Bone Marrow Mesenchymal Stem Cells. Front Bioeng Biotechnol. 2022;10:824025.
[42] CAO H, KE Y, ZHANG Y, et al. Icariin stimulates MC3T3-E1 cell proliferation and differentiation through up-regulation of bone morphogenetic protein-2. Int J Mol Med. 2012;29(3):435-439.
[43] XIAO Q, CHEN A, GUO F. Effects of Icariin on expression of OPN mRNA and type I collagen in rat osteoblasts in vitro. J Huazhong Univ Sci Technolog Med Sci. 2005;25(6):690-692.
[44] SHAUKAT A, SHAUKAT I, RAJPUT SA, et al. Icariin Alleviates Escherichia coli Lipopolysaccharide-Mediated Endometritis in Mice by Inhibiting Inflammation and Oxidative Stress. Int J Mol Sci. 2022;23(18):10219.
[45] LI X, KHAN I, XIA W, et al. Icariin enhances youth-like features by attenuating the declined gut microbiota in the aged mice. Pharmacol Res. 2021;168:105587.
[46] LIU J, LI D, SUN X, et al. Icariine Restores LPS-Induced Bone Loss by Downregulating miR-34c Level. Inflammation. 2016;39(5):1764-1770.
[47] YU X, DONG M, WANG L, et al. Nanotherapy for bone repair: milk-derived small extracellular vesicles delivery of icariin. Drug Deliv. 2023;30(1):2169414.