PMID- 34270816
OWN - NLM
STAT- MEDLINE
DCOM- 20211229
LR  - 20220731
IS  - 1097-0231 (Electronic)
IS  - 0951-4198 (Print)
IS  - 0951-4198 (Linking)
VI  - 35
IP  - 20
DP  - 2021 Oct 30
TI  - Inflammation protein quantification by multiple reaction monitoring mass 
      spectrometry in lipopolysaccharide-stimulated THP-1 cells.
PG  - e9166
LID - 10.1002/rcm.9166 [doi]
LID - e9166
AB  - RATIONALE: Inflammation is a cascade of events mediated by a cytokine network 
      triggering the cellular response. In order to monitor the modulation of the 
      crucial inflammatory proteins, e.g., Tumour Necrosis Factor-alpha (TNF-alpha), 
      Interferon-gamma (INF-gamma), Interleukin-8 (IL-8) and Interleukin-10 (IL-10), upon 
      stimulation with endotoxins, differentiated and undifferentiated THP-1 cells were 
      treated with lipopolysaccharides (LPSs) from E. coli, key cell wall components of 
      Gram-negative bacteria. METHODS: The multiple reaction monitoring mass 
      spectrometry (MRM-MS) method was optimized by using the standard proteins to be 
      quantified, in order to construct external calibration curves and define the 
      analytical parameters. The developed method was used to quantify the 
      above-mentioned inflammatory proteins in THP-1 differentiated cells upon 
      stimulation with LPSs with high accuracy, sensitivity, and robustness. RESULTS: 
      The analysis of such proteins in MRM mode allowed the kinetics of stimulation 
      along the time up to 24 h to be followed and the MS results were found to be 
      comparable with those obtained by Western-blotting. A significant increase in 
      TNF-alpha release triggered a cascade mechanism leading to the production of INF-gamma 
      and IL-8. IL-10, instead, was found to be constant throughout the process. 
      CONCLUSIONS: The developed MRM-MS method allowed the quantification of TNF-alpha, 
      INF-gamma, IL-8 and IL-10 along a time-course from 2 to 24 h. Hence, a trace of the 
      kinetics of the inflammatory response in THP-1 cells upon stimulation with E. 
      coli LPSs was obtained. Finally, the extensibility of the developed MRM method to 
      serum samples and other matrices demonstrated the versatility of the approach and 
      the possibility to quantify multiple target proteins in different biological 
      samples by using a few microliters in a single analysis.
CI  - (c) 2021 The Authors. Rapid Communications in Mass Spectrometry published by John 
      Wiley & Sons Ltd.
FAU - Illiano, Anna
AU  - Illiano A
AUID- ORCID: 0000-0003-1491-8966
AD  - CEINGE Advanced Biotechnologies, University of Naples Federico II, Naples, Italy.
AD  - Department of Chemical Sciences, University of Naples Federico II, Naples, Italy.
AD  - Consorzio Interuniversitario Istituto Nazionale Biostrutture e Biosistemi, Rome, 
      Italy.
FAU - Pinto, Gabriella
AU  - Pinto G
AUID- ORCID: 0000-0001-9169-3452
AD  - Department of Chemical Sciences, University of Naples Federico II, Naples, Italy.
AD  - Consorzio Interuniversitario Istituto Nazionale Biostrutture e Biosistemi, Rome, 
      Italy.
FAU - Gaglione, Rosa
AU  - Gaglione R
AUID- ORCID: 0000-0003-2391-0237
AD  - Department of Chemical Sciences, University of Naples Federico II, Naples, Italy.
FAU - Arciello, Angela
AU  - Arciello A
AUID- ORCID: 0000-0001-8269-6459
AD  - Department of Chemical Sciences, University of Naples Federico II, Naples, Italy.
FAU - Amoresano, Angela
AU  - Amoresano A
AUID- ORCID: 0000-0002-8386-655X
AD  - Department of Chemical Sciences, University of Naples Federico II, Naples, Italy.
AD  - Consorzio Interuniversitario Istituto Nazionale Biostrutture e Biosistemi, Rome, 
      Italy.
LA  - eng
GR  - University of Naples Federico II/
PT  - Evaluation Study
PT  - Journal Article
PL  - England
TA  - Rapid Commun Mass Spectrom
JT  - Rapid communications in mass spectrometry : RCM
JID - 8802365
RN  - 0 (Interleukin-8)
RN  - 0 (Lipopolysaccharides)
RN  - 0 (Tumor Necrosis Factor-alpha)
RN  - 130068-27-8 (Interleukin-10)
RN  - 82115-62-6 (Interferon-gamma)
SB  - IM
MH  - Escherichia coli/immunology/physiology
MH  - Escherichia coli Infections/immunology/microbiology
MH  - Humans
MH  - Inflammation/*immunology/microbiology
MH  - Interferon-gamma/chemistry/immunology
MH  - Interleukin-10/chemistry/immunology
MH  - Interleukin-8/chemistry/immunology
MH  - Kinetics
MH  - Lipopolysaccharides/adverse effects/*immunology
MH  - Mass Spectrometry/*methods
MH  - Monocytes/*chemistry/*immunology
MH  - THP-1 Cells
MH  - Tumor Necrosis Factor-alpha/chemistry/immunology
PMC - PMC9285679
EDAT- 2021/07/17 06:00
MHDA- 2021/12/30 06:00
PMCR- 2022/07/15
CRDT- 2021/07/16 18:02
PHST- 2021/07/13 00:00 [revised]
PHST- 2021/04/09 00:00 [received]
PHST- 2021/07/13 00:00 [accepted]
PHST- 2021/07/17 06:00 [pubmed]
PHST- 2021/12/30 06:00 [medline]
PHST- 2021/07/16 18:02 [entrez]
PHST- 2022/07/15 00:00 [pmc-release]
AID - RCM9166 [pii]
AID - 10.1002/rcm.9166 [doi]
PST - ppublish
SO  - Rapid Commun Mass Spectrom. 2021 Oct 30;35(20):e9166. doi: 10.1002/rcm.9166.