Molecular Biology Reports

, Volume 46, Issue 2, pp 2447–2459 | Cite as

Exploration of immunomodulatory and protective effect of Withania somnifera on trace metal oxide (zinc oxide nanoparticles) induced toxicity in Balb/c mice

  • Jitendra KumarEmail author
  • Murli Dhar Mitra
  • Ahmad Hussain
  • Gautam Kaul
Original Article


The current study was undertaken to investigate the immunomodulatory and protective effects of Withania somnifera (WS) extract and Withaferin A (WA) supplementation on zinc oxide nanoparticles mediated toxicity in Balb/c mice. The animals were exposed to ZnO NPs along with WS and WA for 28 days and various parameters like body weight, organ coefficient, cytotoxicity, nitric oxide (NO), total serum protein, phagocytosis, and the gene expression levels of TLR6 and ARG genes were determined. In vivo study showed that, dose-dependent reduction in phagocytosis, an increase in the levels of NO production along with up-regulation of TLR6, arginase gene was significant (P < 0.05) when ZnO NPs were given. However toxicity of ZnO NP was reduced in presence of WS and WA with decreased TLR6 over expression and restoration of phagocytic activities. Our results provided a valuable insight into the underlying mechanism for the protective effects of WS.

Graphical Abstract

Mechanism of toxicity induced by Zinc oxide nanoparticles ZnO NPs and immunomodulatory protective effects of Withania somnifera extract (WS) and Withaferin A (WA), in Balb/c mice modal of peritoneal macrophages. Red arrows: effect of ZnO NPs independently leads to ROS production which attenuated the phagocytosis of yeast by macrophages through, up-regulation of TLR6 and down-regulation of arginase gene expression. Green arrows: co-treatment, Impact of Withania somnifera extract with zinc oxide nanoparticles (WS + ZnO NPs), Withaferin A along with zinc oxide nanoparticles (WA + ZnO NPs)-enhance phagocytic activity by counteracting mechanism of ZnO NPs toxicity. Black arrows: increasing or decreasing effects. Per oral (P.O).


Balb/c mice Immunomodulatory Oxidative stress Phagocytosis Withania somnifera Zinc oxide nanoparticles 





Dimethyl sulfoxide


Dulbecco’s Modified Eagle’s Medium Hams F-12


Fetal calf serum


Instability (∆HME+), EC50 dose of nanoparticle that gives half-maximal response


Reactive oxygen species

WA + ZnO NP5

Withaferin A and zinc oxide nanoparticle 5 mg/kg body weight

WA + ZnO NP50

Withaferin A and zinc oxide nanoparticle 50 mg/kg body weight


Withania somnifera extract

WS + ZnO NP5

Withania somnifera extract and zinc oxide nanoparticle 5 mg/kg body weight

WS + ZnO NP50

Withania somnifera extract and zinc oxide nanoparticle 50 mg/kg body weight


Zinc oxide nanoparticle

ZnO NPs5

Zinc oxide nanoparticle 5 mg/kg body weight

ZnO NPs50

Zinc oxide nanoparticle 50 mg/kg body weight



The authors wish to thank the Director, Indian Council of Agriculture Research-National Dairy Research institute, Karnal (Haryana)-132001, India for the research grants. Also thanks to Dr. Sadeesh E. M. and Dr. Chander Datt, M.D. Mitra, Dr. Abhishek, Sandeep, Taruna, Vikash, Kavita Rani, Kush Pandey and Dr. Hanuman, for unconditional support and help during research.

Author contributions

Conceived and designed the experiments: JK, performed the experiments; JK, AH and MDM, analyzed the data: JK, AH, and wrote the paper: JK, MDM.


This work was financially supported by the Director, Indian Council of Agriculture Research (ICAR)-National Dairy Research institute, Karnal-132001, Haryana, India for the fellowship provided to Jitendra Kumar for research.

Compliance with ethical standards

Conflict of interest

The authors declare no competing financial interest in our study.


All experimental animals were managed according to standard guidelines as approved by the Institutional Animal Ethics Committee (IACE). The approval for this study and experimentation was also taken from Institutional Animal Ethics Committee (Reg. No. 1705/GO/AC/13/CPCSEA), Livestock Research Center, National Dairy Research Institute (ICAR-NDRI), Karnal-132001, Haryana, India.


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Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  1. 1.Animal Biochemistry DivisionICAR-National Dairy Research InstituteKarnalIndia
  2. 2.Department of ChemistryIndian Institute of Technology (IIT), Banaras Hindu University (BHU)VaranasiIndia

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