Abstract
Heat stress affects milk yield and quality in lactating dairy cows in summer. Bovine mammary epithelial cells (bMECs) play a key role in milk secretion, and microRNAs (miRNAs) regulate numerous functions of bMEC. Previous reports have verified that miR-216b regulated cell apoptosis through repressing target genes in several cancer cells. So, our purpose was to explore the potential involvement of miR-216b in heat stress-induced cell apoptosis in bMECs. Firstly, the heat stress model was constructed and we found that apoptotic rates of bMECs significantly increased under heat stress. The expression of miR-216b, Bax mRNA, and caspase-3 mRNA was upregulated. However, Bcl-2 mRNA level was detected to differentially downregulated. Overexpression of miR-216b remarkably downregulated the expression of caspase-3 and Bax mRNA and protein, and the mRNA and protein level of Bcl-2 was increased. Inhibition of miR-216b increased the activity of caspase-3 and Bax, and the level of Bcl-2 was inhibited. Moreover, Fas was identified as a target gene of miR-216b through bioinformatic analysis and dual-luciferase reporter assay. Fas activity was significantly inhibited and enhanced respectively after transfecting miRNA mimics and inhibitor. Finally, inhibition of Fas via the small interfering RNA (siRNA) also inhibited cell apoptosis induced by heat stress. Taken together, our results indicated that miR-216b exerted as an anti-apoptotic effect under heat stress in bMECs by targeting Fas.
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References
Bartel DP (2004) MicroRNAs: genomics, biogenesis, mechanism, and function. Cell 116:281–297. https://doi.org/10.1016/S0092-8674(04)00045-5
Boise LH, González-García M, Postema CE, Ding L, Lindsten T, Turka LA, Mao X, Nuñez G, Thompson CB (1993) bcl-x, a bcl-2-related gene that functions as a dominant regulator of apoptotic cell death. Cell 74:597–608. https://doi.org/10.1016/0092-8674(93)90508-N
Chen H, Wang J, Hu B, Wu X, Chen Y, Li R, Yuan W (2015) MiR-34a promotes Fas-mediated cartilage endplate chondrocyte apoptosis by targeting Bcl-2. Mol Cell Biochem 406:21–30. https://doi.org/10.1007/s11010-015-2420-4
Clemons NJ, Buzzard K, Steel R, Anderson RL (2005) Hsp72 inhibits Fas-mediated apoptosis upstream of the mitochondria in type II cells. J Biol Chem 280:9005–9012. https://doi.org/10.1074/jbc.M414165200
Degenhardt K, Chen G, Lindsten T, White E (2002) BAX and BAK mediate p53-independent suppression of tumorigenesis. Cancer Cell 2:193–203. https://doi.org/10.1016/S1535-6108(02)00126-5
Deng M, Tang H, Zhou Y, Zhou M, Xiong W, Zheng Y, Ye Q, Zeng X, Liao Q, Guo X (2011) miR-216b suppresses tumor growth and invasion by targeting KRAS in nasopharyngeal carcinoma. J Cell Sci 124:2997–3005. https://doi.org/10.1242/jcs.085050
Elmore S (2007) Apoptosis: a review of programmed cell death. Toxicol Pathol 35:495–516. https://doi.org/10.1080/01926230701320337
Gu ZT, Li L, WU F, Zhao P, Yang H, Liu YS, Geng Y, Zhao M, Su L (2015) Heat stress induced apoptosis is triggered by transcription-independent p53, Ca2+ dyshomeostasis and the subsequent Bax mitochondrial translocation. Sci Rep 5:11497. https://doi.org/10.1038/srep11497
Guo X, Chi S, Cong X, Li H, Jiang Z, Cao R, Tian W (2015) Baicalin protects sertoli cells from heat stress-induced apoptosis via activation of the Fas/FasL pathway and Hsp72 expression. Reprod Toxicol 57:196–203. https://doi.org/10.1016/j.reprotox.2015.06.049
Han N, Shi L, Guo Q, Sun W, Yu Y, Yang L, Zhang X, Zhang M, Han N, Shi L (2017) HAT1 induces lung cancer cell apoptosis via up regulating Fas. Oncotarget 8:89970–89977. https://doi.org/10.18632/oncotarget.21205
Hsu YL, Yu HS, Lin HC, Wu KY, Yang RC, Kuo PL (2011) Heat shock induces apoptosis through reactive oxygen species involving mitochondrial and death receptor pathways in corneal cells. Exp Eye Res 93:405–412. https://doi.org/10.1016/j.exer.2011.06.005
Jana S, Sengupta S, Biswas S, Chatterjee A, Roy H, Bhattacharyya A (2017) miR-216b suppresses breast cancer growth and metastasis by targeting SDCBP. Biochem Biophys Res Commun 482:126–133. https://doi.org/10.1016/j.bbrc.2016.10.003
Ji YB, Chen N, Zhu HW, Ling N, Li WL, Song DX, Gao SY, Zhang WC, Ma NN (2014) Alkaloids from beach spider lily (Hymenocallis littoralis) induce apoptosis of HepG-2 cells by the fas-signaling pathway. Asian Pac J Cancer Prev APJCP 15:9319–9325. https://doi.org/10.7314/APJCP.2014.15.21.9319
Kadzere CT, Murphy MR, Silanikove N, Maltz E (2002) Heat stress in lactating dairy cows: a review. Livest Prod Sci 77:59–91. https://doi.org/10.1016/S0301-6226(01)00330-X
Khate K, Yadav BR (2010) Incidence of mastitis in Sahiwal cattle and Murrah buffaloes of a closed organized herd. Indian J Anim Sci 80:467–469
Li Z, Meng J, Xu TJ, Qin XY, Zhou XD (2013) Sodium selenite induces apoptosis in colon cancer cells via Bax-dependent mitochondrial pathway. Eur Rev Med Pharmacol Sci 17:2166–2171
Liossis SN, Ding XZ, Kiang JG, Tsokos GC (1997) Overexpression of the heat shock protein 70 enhances the TCR/CD3- and Fas/Apo-1/CD95-mediated apoptotic cell death in Jurkat T cells. J Immunol 158:5668–5675
Liu F, Zhou S, Deng Y, Zhang Z, Zhang E, Wu Z, Huang Z, Chen X (2015) MiR-216b is involved in pathogenesis and progression of hepatocellular carcinoma through HBx-miR-216b-IGF2BP2 signaling pathway. Cell Death Dis 6:e1670. https://doi.org/10.1038/cddis.2015.46
Liu HX, Liu S, Qu W, Yang HY, Wen X, Chen T, Hou LF, Ping J (2017) α7 nAChR mediated Fas demethylation contributes to prenatal nicotine exposure-induced programmed thymocyte apoptosis in mice. Oncotarget 8:93741–93756. https://doi.org/10.18632/oncotarget.21526
Maglara AA, Vasilaki A, Jackson MJ, Mcardle A (2003) Damage to developing mouse skeletal muscle myotubes in culture: protective effect of heat shock proteins. J Physiol 548:837–846. https://doi.org/10.1113/jphysiol.2002.034520
Mendell JT, Olson EN (2012) MicroRNAs in stress signaling and human disease. Cell 148:1172–1187. https://doi.org/10.1016/j.cell.2012.02.005
Miura M, Sasagawa I, Suzuki Y, Nakada T, Fujii J (2002) Apoptosis and expression of apoptosis-related genes in the mouse testis following heat exposure. Fertil Steril 77:787–793. https://doi.org/10.1016/s0015-0282(01)03255-1
Mo JS, Alam KJ, Kang IH, Park WC, Seo GS, Choi SC, Kim HS, Moon HB, Yun KJ, Chae SC (2015) MicroRNA 196B regulates FAS-mediated apoptosis in colorectal cancer cells. Oncotarget 6:2843–2855. https://doi.org/10.18632/oncotarget.3066
Moulin M, Arrigo AP (2006) Long lasting heat shock stimulation of TRAIL-induced apoptosis in transformed T lymphocytes. Exp Cell Res 312:1765–1784. https://doi.org/10.1016/j.yexcr.2006.02.008
Salakou S, Kardamakis D, Tsamandas AC, Zolota V, Apostolakis E, Tzelepi V, Papathanasopoulos P, Bonikos DS, Papapetropoulos T, Petsas T (2007) Increased Bax/Bcl-2 ratio up-regulates caspase-3 and increases apoptosis in the thymus of patients with myasthenia gravis. Vivo 21:123
Sonkoly E, Ståhle M, Pivarcsi A (2008) MicroRNAs and immunity: novel players in the regulation of normal immune function and inflammation. Semin Cancer Biol 18:131–140. https://doi.org/10.1016/j.semcancer.2008.01.005
Tran SE, Meinander A, Holmström TH, Riveromüller A, Heiskanen KM, Linnau EK, Courtney MJ, Mosser DD, Sistonen L, Eriksson JE (2003) Heat stress downregulates FLIP and sensitizes cells to Fas receptor-mediated apoptosis. Cell Death Differ 10:1137–1147. https://doi.org/10.1038/sj.cdd.4401278
Trauth BC, Klas C, Matzku S, Möller P, Falk W, Debatin KM, Krammer PH (1989) Monoclonal antibody-mediated tumor regression by induction of apoptosis. Science 245:301
Vaux DL, Korsmeyer SJ (1999) Cell death in development. Cell 96:245–254
Wang X, Yuan B, Dong W, Yang B, Yang Y, Lin X, Gu G (2015) Humid heat exposure induced oxidative stress and apoptosis in cardiomyocytes through the angiotensin II signaling pathway. Heart Vessel 30:396–405. https://doi.org/10.1007/s00380-014-0523-6
Xu X, Fu Y, Tong J, Fan S, Xu K, Sun H, Liang Y, Yan C, Yuan Z, Ge Y (2014) MicroRNA-216b/Beclin 1 axis regulates autophagy and apoptosis in human Tenon’s capsule fibroblasts upon hydroxycamptothecin exposure. Exp Eye Res 123:43–55. https://doi.org/10.1016/j.exer.2014.03.008
Zhang R, Wang L, He L, Yang B, Yao C, Du P, Xu Q, Cheng W, Hua ZC (2014) Fas-associated protein with death domain regulates notch signaling during muscle regeneration. Cells Tissues Organs 200:253–264. https://doi.org/10.1159/000437258
Zhang JF, Shi LL, Zhang L, Zhao ZH, Liang F, Xu X, Zhao LY, Yang PB, Zhang JS, Tian YF (2016) MicroRNA-25 negatively regulates cerebral ischemia/reperfusion injury-induced cell apoptosis through Fas/FasL pathway. J Mol Neurosci 58:507–516. https://doi.org/10.1007/s12031-016-0712-0
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This research was supported by the Research on the Germplasm Resources of Main Livestock and Poultry and Public Service in Sichuan (No. 2016NYZ0050).
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SJL and MCC contributed to concept design. YSH and THZ cultured the cells and constructed the heat stress models; MCC, THZ, and HBH performed RNA and protein isolation and Western blot; WDX, YS, and BWL performed data analysis; MCC, JW, and SYC performed manuscript preparing. All authors read and approved the final manuscript.
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Cai, M., Hu, Y., Zheng, T. et al. MicroRNA-216b inhibits heat stress-induced cell apoptosis by targeting Fas in bovine mammary epithelial cells. Cell Stress and Chaperones 23, 921–931 (2018). https://doi.org/10.1007/s12192-018-0899-9
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DOI: https://doi.org/10.1007/s12192-018-0899-9