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Standardized fraction of Xylocarpus moluccensis inhibits inflammation by modulating MAPK-NFκB and ROS-HIF1α-PKM2 activation

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Abstract

Objective

Present study investigates the effect of Xylocarpus moluccensis (Lamk.) M. Roem fruit fraction (CDR) on endotoxemia and explores the underlying mechanisms.

Materials and methods

The effect of CDR (1–100 µg/ml) was assessed on cytokines, MAPKs, ROS, and metabolic reprogramming in LPS-induced cells (J774.2 and THP-1) by the conventional methodology of ELISA, PCR, and Western blotting. The effect of CDR (1–50 mg/kg, p.o.) was also evaluated in the mice model of endotoxemia and sepsis.

Results

CDR prevents LPS-induced cytokine production from murine and human whole blood and cell lines. CDR suppressed total cellular and mitochondrial superoxide generation and preserved mitochondrial function in LPS-stimulated phagocytes. Additionally, CDR abrogated LPS-induced MAPK’s phosphorylation and IκBα degradation in J774.2 cells. Moreover, CDR suppressed LPS-induced glycolytic flux as indicated from PKM2, HK-2, PDK-2, and HIF-1α expression in J774.2 cells. In vivo, CDR pre-treatment inhibited pro-inflammatory cytokines release, metabolic reprogramming from oxidative phosphorylation to glycolysis in both LPS-induced endotoxemia and cecal slurry-induced sepsis mice model.

Conclusion

Present study demonstrates the protective effect of CDR on LPS-induced inflammation and sepsis and identifies MAPK-NFκB and ROS-HIF1α-PKM2 as the putative target axis.

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Availability of data and materials

The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.

Abbreviations

LPS:

Lipopolysaccharide

TLRs:

Toll-like receptors

TNF-α:

Tumour necrosis factor alpha

IL-6:

Interleukin 6

IL-1β:

Interleukin 1 beta

NLRP3:

NLR family pyrin domain containing 3

IL-4:

Interleukin 4

IL-10:

Interleukin 10

MAPK:

Mitogen-activated protein kinase

ERK1/2:

Extracellular signal-regulated kinases 1/2

JNK1/2:

C-Jun N-terminal kinase 1/2

IκBα:

Inhibitor of kappa B

PKM2:

Pyruvate kinase muscle isozyme M2

HK-2:

Hexokinase-2

PDK-2:

Pyruvate dehydrogenase kinase 2

HIF-1α:

Hypoxia-inducible factor-1alpha

iNOS:

Inducible nitric oxide synthase

Arg-1:

Arginase-1

PAMP:

Pathogen-associated molecular pattern

ROS:

Reactive oxygen species

OXPHOS:

Oxidative phosphorylation

CNS:

Central nervous system

CDRI:

Central Drug Research Institute

RPMI:

Roswell Park Memorial Institute

DMEM:

Dulbecco's modified Eagle medium

PMs:

Peritoneal macrophages

i.p.:

Intraperitoneal

FBS:

Fetal bovine serum

PI:

Protease Inhibitor

DHE:

Dihydroethidium

MSR:

MitoSOX red

DMSO:

Dimethyl sulfoxide

ELISA:

Enzyme-linked immune sorbent assay

BCA:

Bicinchoninic acid assay

SDS-PAGE:

Sodium dodecyl sulfate polyacrylamide gel electrophoresis

PVDF:

Polyvinylidene difluoride

TBST:

Tris-buffered saline with Tween

IAEC:

Institutional Animal Ethics Committee

CMC:

Carboxy methyl cellulose

CS:

Cecal slurry

P.O.:

Peroral

TGA:

Thioglycollate

MMP:

Matrix metalloproteinases

RNS:

Reactive nitrogen species

ETC:

Electron transport chain

ATP:

Adenosine triphosphate

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Acknowledgements

Authors acknowledge the excellent technical help of Mr. C·P Pandey for the central instrument facility, Mr. A.L. Vishwakarma, and Mrs. M. Chaturvedi for the Flow Cytometry experiment from Sophisticated Analytical Instrument Facility; CSIR-CDRI, Lucknow. This paper bears CDRI communication number 10375.

Funding

Grant support to MKB from CSIR, CDRI, and DST (EMR/2016/005340) project is acknowledged. Award of senior research fellowships to HA (3/1/2/(7)/CVD/2018-NCD-II) and SSR (3/1/2/(1)/OBS/18/NCD-II) from ICMR is gratefully acknowledged.

Author information

Author notes

  1. Madhu Dikshit

    Present address: Translational Health Science and Technology Institute, Faridabad, Haryana, 121001, India

Authors and Affiliations

  1. Pharmacology Division, CSIR-Central Drug Research Institute, B.S. 10/1, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow, 226031, India

    Heena Agarwal, Santosh Reddy Sukka, Vishal Singh, Madhu Dikshit & Manoj Kumar Barthwal

  2. Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India

    Heena Agarwal, Santosh Reddy Sukka & Manoj Kumar Barthwal

Authors
  1. Heena Agarwal
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Contributions

HA performed the majority of experiments, generated figures, analysed the data, and wrote the manuscript draft. SSR involved in in vivo LPS-induced endotoxemia mice models, VS was involved initially in standardization of a few in vivo protocols and experiments, MD gave critical inputs related to in vitro, in vivo studies, and manuscript preparation, and MKB is responsible for the overall concept, project generation, and conduct and finalization of the manuscript.

Corresponding author

Correspondence to Manoj Kumar Barthwal.

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Conflict of interest

The authors declare no competing interests.

Ethics approval

All animal procedures were in accordance with the IAEC (approval number: IAEC/2013/53/Renew-04 (240/17)/Dated-07/03/2017 & IAEC/2016/152/Dated-04/11/2016), which follow the Committee for the Purpose of Control and Supervision of Experiments on Animals guidelines and conforming to the Indian National Science Academy international norms.

Consent to participate

Experiments using human blood were conducted out after getting necessary ethical permission from the Institute's human research ethics committee at King George's Medical University (KGMC), Lucknow, India.

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All the co-authors have seen and approved the content of the manuscript for publication.

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Agarwal, H., Sukka, S.R., Singh, V. et al. Standardized fraction of Xylocarpus moluccensis inhibits inflammation by modulating MAPK-NFκB and ROS-HIF1α-PKM2 activation. Inflamm. Res. 71, 423–437 (2022). https://doi.org/10.1007/s00011-022-01549-3

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  • DOI: https://doi.org/10.1007/s00011-022-01549-3

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