Skip to main content
SpringerLink
Log in

Silver Nanoparticles Induced Oxidative Stress and Mitochondrial Injuries Mediated Autophagy in HC11 Cells Through Akt/AMPK/mTOR Pathway

  • Published:
Biological Trace Element Research Aims and scope Submit manuscript

Abstract

Silver nanoparticles (AgNPs) are widely used in industrial products, and they have good antibacterial properties, with potential for prevention and treatment of cow mastitis. However, concerns exist about the cytotoxicity of AgNPs. Thus, we have studied the role of autophagy in AgNP-induced cytotoxicity in mouse HC11 mammary epithelium cells. We found that AgNPs injured HC11 cells, with release of lactate dehydrogenase (LDH). AgNPs also induced autophagy in HC11 cells, which was associated with oxidative stress, as indicated by increased reactive oxygen species (ROS) and increased expression of hemoxygenase-1(HO-1) and Nrf2. Mitochondria were altered by AgNPs: mitochondrial membrane potential (MMP) was decreased and the expression of PINK1 and Parkin was increased. AgNPs also increased the expression of p-AMPK and decreased the expression of p-Akt and p-mTOR. The addition of 3-methyl adenine inhibited autophagy and enhanced the cytotoxicity of AgNPs, indicating that autophagy is protective against AgNP-induced cell death. In summary, AgNPs induced protective autophagy in HC11 cells via the Akt/AMPK/mTOR pathway, associated with cellular oxidative stress and mitochondrial alterations. Our research confirms that AgNPs may damage the breast tissue in clinical applications and should be used with caution. Further research is necessary to clarify whether the damage caused by AgNPs will affect the lactation function of the mammary glands and possible residues in milk.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Data Availability

All data generated or analyzed during this study are included in this published article.

Abbreviations

LDH:

lactate dehydrogenase

LC3:

microtubule-associated protein 1-light chain 3

Beclin:

Beclin 1, autophagy related

P62:

sequestosome 1

ROS:

reactive oxygen stress

HO-1:

heme oxygenase1

Nrf2:

nuclear factor erythroid-2p45-related factor2

Keap1:

Kelch-like ECH-associated protein 1

Bach 1:

BTB and CNC homology 1

NAC:

N-acetyl-l-cysteine

MMP:

mitochondrial membrane potential

JC-1:

5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolylcarbocyanine iodide

PINK1:

Putative kinase 1

Akt:

RAC-alpha serine/threonine-protein kinase

AMPK:

5′-AMP-activated protein kinase catalytic subunit

mTOR:

rapamycin target protein

References

  1. Derrien TL, Hamada S, Zhou M, Smilgies D-M, Luo D (2020) Three-dimensional nanoparticle assemblies with tunable plasmonics via a layer-by-layer process. Nano Today 30:100823. https://doi.org/10.1016/j.nantod.2019.100823

    Article  CAS  Google Scholar 

  2. Zhao Y, Li L, Zhang PF, Shen W, Liu J, Yang FF, Liu HB, Hao ZH (2015) Differential regulation of gene and protein expression by zinc oxide nanoparticles in Hen’s ovarian granulosa cells: specific roles of nanoparticles. PLoS One 10(10):e0140499. https://doi.org/10.1371/journal.pone.0140499

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Shati AA, Elsaid FG (2020) Biosynthesized silver nanoparticles and their genotoxicity. J Biochem Mol Toxicol 34(1):e22418. https://doi.org/10.1002/jbt.22418

    Article  CAS  PubMed  Google Scholar 

  4. Li L, Li L, Zhou X, Yu Y, Li Z, Zuo D, Wu Y (2019) Silver nanoparticles induce protective autophagy via Ca(2+)/CaMKKbeta/AMPK/mTOR pathway in SH-SY5Y cells and rat brains. Nanotoxicology 13(3):369–391. https://doi.org/10.1080/17435390.2018.1550226

    Article  CAS  PubMed  Google Scholar 

  5. Huo L, Chen R, Zhao L, Shi X, Bai R, Long D, Chen F, Zhao Y, Chang YZ, Chen C (2015) Silver nanoparticles activate endoplasmic reticulum stress signaling pathway in cell and mouse models: the role in toxicity evaluation. Biomaterials 61:307–315. https://doi.org/10.1016/j.biomaterials.2015.05.029

    Article  CAS  PubMed  Google Scholar 

  6. Simard JC, Vallieres F, de Liz R, Lavastre V, Girard D (2015) Silver nanoparticles induce degradation of the endoplasmic reticulum stress sensor activating transcription factor-6 leading to activation of the NLRP-3 inflammasome. J Biol Chem 290(9):5926–5939. https://doi.org/10.1074/jbc.M114.610899

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Zhang XF, Choi YJ, Han JW, Kim E, Park JH, Gurunathan S, Kim JH (2015) Differential nanoreprotoxicity of silver nanoparticles in male somatic cells and spermatogonial stem cells. Int J Nanomedicine 10:1335–1357. https://doi.org/10.2147/IJN.S76062

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Li R, Luo X, Zhu Y, Zhao L, Li L, Peng Q, Ma M, Gao Y (2017) ATM signals to AMPK to promote autophagy and positively regulate DNA damage in response to cadmium-induced ROS in mouse spermatocytes. Environ Pollut 231:1560–1568. https://doi.org/10.1016/j.envpol.2017.09.044

    Article  CAS  PubMed  Google Scholar 

  9. Zhou W, Miao Y, Zhang Y, Liu L, Lin J, Yang JY, Xie Y, Wen L (2013) Induction of cyto-protective autophagy by paramontroseite VO2nanocrystals. Nanotechnology 24(16):165102

    Article  Google Scholar 

  10. Li R, Ji Z, Qin H, Kang X, Sun B, Wang M, Chang CH, Wang X, Zhang H, Zou H, Nel AE, Xia T (2014) Interference in autophagosome fusion by rare earth nanoparticles disrupts autophagic flux and regulation of an interleukin-1beta producing inflammasome. ACS Nano 8(10):10280–10292. https://doi.org/10.1021/nn505002w

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Ziglari T, Anderson DS, Holian A (2020) Determination of the relative contribution of the non-dissolved fraction of ZnO NP on membrane permeability and cytotoxicity. Inhal Toxicol 32(2):86–95. https://doi.org/10.1080/08958378.2020.1743394

    Article  CAS  PubMed  Google Scholar 

  12. Ji Wang YY, Lu K, Yang M, Li Y, Zhou X, Sun Z (2018) Silica nanoparticles induce autophagy dysfunction via lysosomal impairment in hepatocytes. Nanomedicine 14(5):1837. https://doi.org/10.1016/j.nano.2017.11.264

    Article  Google Scholar 

  13. Xuemei Liu BT, Jiang X, Xu G, Bai L, Zhang L, Pan M, Qin X, Chen C, Zou Z (2019) Lysosomal dysfunction is associated with persistent lung injury in dams caused by pregnancy exposure to carbon black nanoparticles. Life Sci 233:116741. https://doi.org/10.1016/j.lfs.2019.116741

    Article  CAS  PubMed  Google Scholar 

  14. Miyayama T, Fujiki K, Matsuoka M, Miyayama T, Fujiki K, Matsuoka M, Miyayama T, Fujiki K, Matsuoka M (2017) Silver nanoparticles induce lysosomal-autophagic defects and decreased expression of transcription factor EB in A549 human lung adenocarcinoma cells. Toxicol in Vitro 46:148–154

    Article  Google Scholar 

  15. Farah MA, Ali MA, Chen S-M, Li Y, Al-Hemaid FM, Abou-Tarboush FM, Al-Anazi KM, Lee J (2016) Silver nanoparticles synthesized from Adenium obesum leaf extract induced DNA damage, apoptosis and autophagy via generation of reactive oxygen species. Colloids Surf B: Biointerfaces 141:158–169

    Article  Google Scholar 

  16. Xin L, Wang J, Fan G, Che B, Cheng K, Dong G (2016) Comparative oxidative stress elicited by nanosilver in stable HSPA1A promoter-driven luciferase reporter HepG2 and A549 cells. Toxicol Res (Camb) 5:5–1305. https://doi.org/10.1039/c6tx00195e

    Article  Google Scholar 

  17. Lee YH, Cheng FY, Chiu HW, Tsai JC, Fang CY, Chen CW, Wang YJ (2014) Cytotoxicity, oxidative stress, apoptosis and the autophagic effects of silver nanoparticles in mouse embryonic fibroblasts. Biomaterials 35(16):4706–4715. https://doi.org/10.1016/j.biomaterials.2014.02.021

    Article  CAS  PubMed  Google Scholar 

  18. Akter M, Sikder MT, Rahman MM, Ullah A, Hossain KFB, Banik S, Hosokawa T, Saito T, Kurasaki M (2018) A systematic review on silver nanoparticles-induced cytotoxicity: physicochemical properties and perspectives. J Adv Res 9(C):1–16. https://doi.org/10.1016/j.jare.2017.10.008

    Article  CAS  PubMed  Google Scholar 

  19. Maurer LL, Meyer JN (2016) A systematic review of evidence for silver nanoparticle-induced mitochondrial toxicity. Environmental Science Nano 3(2):311–322. https://doi.org/10.1039/c5en00187k

    Article  CAS  Google Scholar 

  20. Leermakers PA, Remels AHV, Zonneveld MI, Rouschop KMA, Gosker HR (2020) Iron deficiency-induced loss of skeletal muscle mitochondrial proteins and respiratory capacity; the role of mitophagy and secretion of mitochondria-containing vesicles. FASEB J 34(5):1–15. https://doi.org/10.1096/fj.201901815R

    Article  CAS  Google Scholar 

  21. Francesco Napoletano OB, Vandenabeele P, Mollereau B, Fanto M (2019) Intersections between regulated cell death and autophagy. Trends Cell Biol 29(4):323–338. https://doi.org/10.1016/j.tcb.2018.12.007

    Article  CAS  PubMed  Google Scholar 

  22. Zhang X, Yin H, Li Z, Zhang T, Yang Z (2016) Nano-TiO2 induces autophagy to protect against cell death through antioxidative mechanism in podocytes. Cell Biol Toxicol 32(6):513–527

    Article  CAS  Google Scholar 

  23. Towers CG, Thorburn A (2016) Therapeutic targeting of autophagy. Ebiomedicine 14:15–23

    Article  Google Scholar 

  24. Hu J, Wu H, Wang D, Yang Z, Zhuang L, Yang N, Dong J (2018) Weicao capsule ameliorates renal injury through increasing autophagy and NLRP3 degradation in UAN rats. Int J Biochem Cell Biol 96:1–8. https://doi.org/10.1016/j.biocel.2018.01.001

    Article  CAS  PubMed  Google Scholar 

  25. Singh R, Cuervo AM (2011) Autophagy in the cellular energetic balance. Cell Metab 13(5):495–504

    Article  CAS  Google Scholar 

  26. Martínez-García GG, Mario G (2020) Autophagy role in environmental pollutants exposure. Prog Mol Biol Transl Sci. https://doi.org/10.1016/bs.pmbts.2020.02.003

  27. Yik-Lam Cho HWST, Saquib Q, Ren Y, Ahmad J, Wahab R, He W, Bay B-H, Shen H-M (2020) Dual role of oxidative stress-JNK activation in autophagy and apoptosis induced by nickel oxide nanoparticles in human cancer cells. Free Radic Biol Med 153:173–186. https://doi.org/10.1016/j.freeradbiomed.2020.03.027

    Article  CAS  PubMed  Google Scholar 

  28. José Antonio Pérez-Arizti JLV-G, Juárez REG, del Pilar Ramos-Godinez M, Colín-Val Z, López-Marure R (2020) Titanium dioxide nanoparticles promote oxidative stress, autophagy and reduce NLRP3 in primary rat astrocytes. Chem Biol Interact 317:108966. https://doi.org/10.1016/j.cbi.2020.108966

    Article  CAS  PubMed  Google Scholar 

  29. Tanida I (2011) Autophagosome formation and molecular mechanism of autophagy. Antioxid Redox Signal 14(11):2201–2214

    Article  CAS  Google Scholar 

  30. Meng Y-C, Lou X-L, Yang L-Y, Li D, Hou Y-Q (2020) Role of the autophagy-related marker LC3 expression in hepatocellular carcinoma: a meta-analysis. J Cancer Res Clin Oncol 146(suppl 2):1103–1113. https://doi.org/10.1007/s00432-020-03174-1

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Liang C, Feng Z, Manthari RK, Wang C, Zhang J (2020) Arsenic induces dysfunctional autophagy via dual regulation of mTOR pathway and Beclin1-Vps34/PI3K complex in MLTC-1 cells. J Hazard Mater 391:122227. https://doi.org/10.1016/j.jhazmat.2020.122227

    Article  CAS  PubMed  Google Scholar 

  32. Xu YWL, Bai R, Zhang T, Chen C (2015) Silver nanoparticles impede phorbol myristate acetate-induced monocyte–macrophage differentiation and autophagy. Nanoscale 7(38):16100–16109

    Article  CAS  Google Scholar 

  33. Ahmed AD, Mohammed H, Soo L, Kyeongseok K, Subbroto S, Gwang-Mo Y, Hye C, Ssang-Goo C (2017) The role of reactive oxygen species (ROS) in the biological activities of metallic nanoparticles. Int J Mol Sci 18(1):120

    Article  Google Scholar 

  34. Szwed M, Sonstevold T, Overbye A, Engedal N, Grallert B, Morch Y, Sulheim E, Iversen TG, Skotland T, Sandvig K, Torgersen ML (2019) Small variations in nanoparticle structure dictate differential cellular stress responses and mode of cell death. Nanotoxicology 13(6):761–782. https://doi.org/10.1080/17435390.2019.1576238

    Article  CAS  PubMed  Google Scholar 

  35. Bellezza I, Giambanco I, Minelli A, Donato R (2018) Nrf2-Keap1 signaling in oxidative and reductive stress. Biochim Biophys Acta, Mol Cell Res 1865(5):721–733

    Article  CAS  Google Scholar 

  36. Teodoro JOS, Rolo AP, Palmeira CM (2013) The NAD ratio redox paradox: why does too much reductive power cause oxidative stress? Toxicol Mech Methods 23(5):297–302

    Article  CAS  Google Scholar 

  37. Teodoro J, Soeiro P, Carlos M, Silva R, Duarte F (2016) Low-dose, subchronic exposure to silver nanoparticles causes mitochondrial alterations in Sprague-Dawley rats. Nanomedicine 11(11):1359–1375

    Article  CAS  Google Scholar 

  38. Lee TY, Liu MS, Huang LJ, Lue SI, Yang RC (2013) Bioenergetic failure correlates with autophagy and apoptosis in rat liver following silver nanoparticle intraperitoneal administration. Part Fibre Toxicol 10(1):40

    Article  Google Scholar 

  39. Ma W, Jing L, Valladares A, Mehta SL, Wang Z, Li PA, Bang JJ (2015) Silver nanoparticle exposure induced mitochondrial stress, caspase-3 activation and cell death: amelioration by sodium selenite. Int J Biol Sci 11(8):860–867. https://doi.org/10.7150/ijbs.12059

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Xin L, Wang J, Fan G, Che B, Wu Y, Guo S, Tong J (2016) Oxidative stress and mitochondrial injury-mediated cytotoxicity induced by silver nanoparticles in human A549 and HepG2 cells. Environ Toxicol 31(12):1691–1699. https://doi.org/10.1002/tox.22171

    Article  CAS  PubMed  Google Scholar 

  41. Wei L, Wang J, Chen A, Liu J, Feng X, Shao L (2017) Involvement of PINK1/parkin-mediated mitophagy in ZnO nanoparticle-induced toxicity in BV-2 cells. Int J Nanomedicine 12:1891–1903. https://doi.org/10.2147/IJN.S129375

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Shan S, Shen Z, Zhang C, Kou R, Xie K, Song F (2019) Mitophagy protects against acetaminophen-induced acute liver injury in mice through inhibiting NLRP3 inflammasome activation. Biochem Pharmacol 169:113643. https://doi.org/10.1016/j.bcp.2019.113643

    Article  CAS  PubMed  Google Scholar 

  43. Ivankovic D, Chau K-Y, Schapira AHV, Gegg ME (2016) Mitochondrial and lysosomal biogenesis are activated following PINK1/parkin-mediated mitophagy. J Neurochem 136(2):388–402

    Article  CAS  Google Scholar 

  44. Al-Bari MAA, Xu P (2020) Molecular regulation of autophagy machinery by mTOR-dependent and -independent pathways. Ann N Y Acad Sci 1467:3–20. https://doi.org/10.1111/nyas.14305

    Article  CAS  PubMed  Google Scholar 

  45. Chang HJ, Ro SH, Jing C, Otto NM, Kim DH (2010) mTOR regulation of autophagy. FEBS Lett 584:1287–1295

    Article  Google Scholar 

  46. Yu Y, Hou L, Song H, Xu P, Sun Y, Wu K (2017) Akt/AMPK/mTOR pathway was involved in the autophagy induced by vitamin E succinate in human gastric cancer SGC-7901 cells. Mol Cell Biochem 424(1–2):173–183. https://doi.org/10.1007/s11010-016-2853-4

    Article  CAS  PubMed  Google Scholar 

  47. Liu Y, Yu H, Zhang X, Wang Y, Zhang LW (2018) The protective role of autophagy in nephrotoxicity induced by bismuth nanoparticles through AMPK/mTOR pathway. Nanotoxicology 12(20):1–16

    Google Scholar 

  48. Kim SH, Kim G, Han DH, Lee M, Kim I, Kim B, Kim KH, Song YM, Yoo JE, Wang HJ, Bae SH, Lee YH, Lee BW, Kang ES, Cha BS, Lee MS (2017) Ezetimibe ameliorates steatohepatitis via AMP activated protein kinase-TFEB-mediated activation of autophagy and NLRP3 inflammasome inhibition. Autophagy 13(10):1767–1781. https://doi.org/10.1080/15548627.2017.1356977

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Pan H, Zhong XP, Lee S (2016) Sustained activation of mTORC1 in macrophages increases AMPKα-dependent autophagy to maintain cellular homeostasis. BMC Biochem 17(1):1–12. https://doi.org/10.1186/s12858-016-0069-6

    Article  CAS  Google Scholar 

  50. Chen Y, Wang M, Zhang T, Du E, Liu Y, Qi S, Xu Y, Zhang Z (2018) Autophagic effects and mechanisms of silver nanoparticles in renal cells under low dose exposure. Ecotoxicol Environ Saf 166:71–77. https://doi.org/10.1016/j.ecoenv.2018.09.070

    Article  CAS  PubMed  Google Scholar 

  51. Jing Pang FL, Xu F, Yang H, Han L, Fan Y, Nie H, Wang Z, Wang F, Zhang Y (2018) Influences of different dietary energy level on sheep testicular development associated with AMPK/ULK1/autophagy pathway. Theriogenology 108:362–370. https://doi.org/10.1016/j.theriogenology.2017.12.017

    Article  CAS  PubMed  Google Scholar 

  52. Liu HL, Zhang YL, Yang N, Zhang YX, Jiang C-Y (2011) A functionalized single-walled carbon nanotube-induced autophagic cell death in human lung cells through Akt–TSC2-mTOR signaling. Cell Death Dis 2(5):e159

    Article  Google Scholar 

  53. Shi M, Cheng L, Zhang Z, Liu Z, Mao X (2015) Ferroferric oxide nanoparticles induce prosurvival autophagy in human blood cells by modulating the Beclin 1/Bcl-2/VPS34 complex. Int J Nanomedicine 10:207–216

    CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We thank the College of Veterinary Medicine, Sichuan Agricultural University, Chengdu 611130, China, for kindly providing facilities.

Author information

Authors and Affiliations

  1. College of Veterinary Medicine, Sichuan Agricultural University, 211 Huimin Road, Chengdu, 611130, Sichuan, China

    Jin Hou, Ling Zhao, Huaqiao Tang, Gang Ye, Fei Shi, Min Kang, Helin Chen & Yinglun Li

  2. College of Science, Sichuan Agricultural University, 211 Huimin Road, Chengdu, 611130, Sichuan, China

    Xiaoli He

Authors
  1. Jin Hou
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ling Zhao
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Huaqiao Tang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Xiaoli He
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Gang Ye
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Fei Shi
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Min Kang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Helin Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Yinglun Li
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Conceptualization, Ling Zhao; data curation, Jin Hou; investigation, Min Kang; methodology, Jin Hou and Huaqiao Tang; project administration, Yinglun Li; resources, Gang Ye and Fei Shi; supervision, Jin Hou, Ling Zhao, Huaqiao Tang, Xiaoli He, Helin Chen, and Yinglun Li; visualization, Ling Zhao; writing—original draft, Jin Hou; and writing—review and editing, Jin Hou and Huaqiao Tang.

Corresponding author

Correspondence to Yinglun Li.

Ethics declarations

Conflict of Interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Hou, J., Zhao, L., Tang, H. et al. Silver Nanoparticles Induced Oxidative Stress and Mitochondrial Injuries Mediated Autophagy in HC11 Cells Through Akt/AMPK/mTOR Pathway. Biol Trace Elem Res 199, 1062–1073 (2021). https://doi.org/10.1007/s12011-020-02212-w

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12011-020-02212-w

Keywords

Search

Navigation