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Cellular and Molecular Life Sciences

, Volume 75, Issue 13, pp 2431–2446 | Cite as

An MD2-derived peptide promotes LPS aggregation, facilitates its internalization in THP-1 cells, and inhibits LPS-induced pro-inflammatory responses

  • Anshika Tandon
  • Munesh Kumar Harioudh
  • Nayab Ishrat
  • Amit Kumar Tripathi
  • Saurabh Srivastava
  • Jimut Kanti Ghosh
Original Article
  • 583 Downloads

Abstract

MD2, a 160-residue accessory glycoprotein, is responsible for the recognition and binding of Gram-negative bacterial membrane component, lipopolysaccharide (LPS). Internalization of pathogen inside the mononuclear phagocytes has also been attributed to MD2 which leads to the clearance of pathogens from the host. However, not much is known about the segments in MD2 that are responsible for LPS interaction or internalization of pathogen inside the defense cells. A 16-residue stretch (MD54) from MD2 protein has been identified that possesses a short heptad repeat sequence and four cationic residues enabling it to participate in both hydrophobic and electrostatic interactions with LPS. An MD54 analog of the same size was also designed in which a leucine residue at a heptadic position was replaced with an alanine residue. MD54 but not its analog, MMD54 induced aggregation of LPS and aided in its internalization within THP-1 monocytes. Furthermore, MD54 inhibited LPS-induced nuclear translocation of NF-κB in PMA-treated THP-1 and TLR4/MD2/CD14-transfected HEK-293T cells and the production of pro-inflammatory cytokines. In addition, in in vivo experiments, MD54 showed marked protection and survival of mice against LPS-induced inflammation and death. Overall, we have identified a short peptide with heptad repeat sequence from MD2 that can cause aggregation of LPS and abet in its internalization within THP-1 cells, resulting in attenuation of LPS-induced pro-inflammatory responses in vitro and in vivo.

Keywords

MD2 Lipopolysaccharide (LPS) aggregation Sepsis Attenuation of LPS-induced pro-inflammatory responses Heptad repeat Synthetic peptides 

Abbreviations

CD14

Cluster of differentiation 14

HEK-293T

Human embryonic kidney cells 293

LBP

Lipopolysaccharide-binding protein

LPS

Lipopolysaccharide

MD2

Lymphocyte antigen 96

NF-κB

Nuclear factor kappa-light-chain-enhancer of activated B cells

PAMPs

Pathogen-associated molecular pattern

PRRs

Pattern recognition receptor

TLR4

Toll-like receptor 4

PCNA

Proliferating cell nuclear antigen

ICU

Intensive Care Unit

AMP

Anti-microbial peptide

Notes

Acknowledgements

The CSIR-CDRI communication number for this article is 9609. This work was supported by a Council of Scientific and Industrial Research (CSIR) network project BioDiscovery (BSC0120). We are thankful to Dr. Douglous Golenbock (Medicine Department, University of Massachusetts Medical School, Worcester MA) and Dr. Anila Dwivedi (Endocrinology div. CSIR-CDRI, Lucknow.) for providing us with the plasmid constructs. The authors are extremely thankful to Prof. Surajit Bhattacharjya, School of Biological Sciences, Nanyang Technological University, Singapore for editing the revised version of the manuscript. The authors thankfully acknowledge the anonymous reviewers for their constructive criticism and comments in improving the quality of the manuscript. Garima Pant of Sophisticated Analytical Instrumentation Facility (SAIF) and Rima Roy from Molecular and Structural Biology Division, CSIR-CDRI are acknowledged for assistance in using the Confocal Microscope. A. L. Vishwakarma and R. K Purshottam from SAIF and CSIR-CDRI are acknowledged for assistance in using FACS and HPLC facility, respectively. We are thankful to Pharmaceutics Department for providing us the Zeta sizer instrument accessibility. We also thankfully acknowledge National Laboratory Animal Centre, CSIR-Central Drug Research Institute, Lucknow, India for providing us the animals.

Author contributions

JKG and AT conceived the idea and designed the experiments. AT did the major part of the experiments. MKH and NI helped in western and in vivo experiments. AKT and SS assisted us in the peptide synthesis and biophysical experiments, respectively. JKG and AT analyzed the results and wrote the manuscript. All the authors were consulted on the final version of the manuscript.

Compliance with ethical standards

Conflict of interest

The authors declare no competing financial interest.

Supplementary material

18_2017_2735_MOESM1_ESM.doc (1.1 mb)
Supplementary material 1 (DOC 1116 kb)

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Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  • Anshika Tandon
    • 1
  • Munesh Kumar Harioudh
    • 1
  • Nayab Ishrat
    • 1
  • Amit Kumar Tripathi
    • 1
  • Saurabh Srivastava
    • 1
  • Jimut Kanti Ghosh
    • 1
  1. 1.Molecular and Structural Biology DivisionCSIR-Central Drug Research InstituteLucknowIndia

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