Quantitative proteomics analysis using 2D-PAGE to investigate the effects of cigarette smoke and aerosol of a prototypic modified risk tobacco product on the lung proteome in C57BL/6 mice.
Smoking is associated with several serious diseases, such as lung cancer and chronic obstructive pulmonary disease (COPD). Within our systems toxicology framework, we are assessing whether potential modified risk tobacco products (MRTP) can reduce smoking-related health risks compared to conventional cigarettes. In this article, we evaluated to what extent 2D-PAGE/MALDI MS/MS (2D-PAGE) can complement the iTRAQ LC-MS/MS results from a previously reported mouse inhalation study, in which we assessed a prototypic MRTP (pMRTP). Selected differentially expressed proteins identified by both LC-MS/MS and 2D-PAGE approaches were further verified using reverse-phase protein microarrays. LC-MS/MS captured the effects of cigarette smoke (CS) on the lung proteome more comprehensively than 2D-PAGE. However, an integrated analysis of both proteomics data sets showed that 2D-PAGE data complement the LC-MS/MS results by supporting the overall trend of lower effects of pMRTP aerosol than CS on the lung proteome. Biological effects of CS exposure supported by both methods included increases in immune-related, surfactant metabolism, proteasome, and actin cytoskeleton protein clusters. Overall, while 2D-PAGE has its value, especially as a complementary method for the analysis of effects on intact proteins, LC-MS/MS approaches will likely be the method of choice for proteome analysis in systems toxicology investigations.



Quantitative proteomics is anticipated to play a growing role within systems toxicology assessment frameworks in the future. To further understand how different proteomics technologies can contribute to toxicity assessment, we conducted a quantitative proteomics analysis using 2D-PAGE and isobaric tag-based LC-MS/MS approaches and compared the results produced from the 2 approaches. Using a prototypic modified risk tobacco product (pMRTP) as our test item, we show compared with cigarette smoke, how 2D-PAGE results can complement and support LC-MS/MS data, demonstrating the much lower effects of pMRTP aerosol than cigarette smoke on the mouse lung proteome. The combined analysis of 2D-PAGE and LC-MS/MS data identified an effect of cigarette smoke on the proteasome and actin cytoskeleton in the lung.
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Ashraf
A
Elamin
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Bjoern
B
Titz
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Sophie
S
Dijon
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Celine
C
Merg
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland; Bayer Technology Services GmbH, 51368 Leverkusen, Germany.
Marcel
M
Geertz
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Thomas
T
Schneider
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Florian
F
Martin
Biology Consultant, Max-Baermann-Str. 21, 51429, Bergisch Gladbach, Germany.
Walter K
WK
Schlage
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Stefan
S
Frentzel
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Fabio
F
Talamo
Philip Morris International Research Laboratories (part of Philip Morris International group of companies), 50 Science Park Road, 117406, Singapore.
Blaine
B
Phillips
Philip Morris International Research Laboratories (part of Philip Morris International group of companies), 50 Science Park Road, 117406, Singapore.
Emilija
E
Veljkovic
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Nikolai V
NV
Ivanov
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Patrick
P
Vanscheeuwijck
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland.
Manuel C
MC
Peitsch
Philip Morris Research and Development, Philip Morris Products SA (part of Philip Morris International group of companies), Quai Jeanrenaud 5, 2000 Neuchâtel, Switzerland. Electronic address: Julia.hoeng@pmi.com.
Julia
J
Hoeng
D000199:Actins / Q000187:drug effects; D000336:Aerosols / Q000009:adverse effects*; D000818:Animals; D002853:Chromatography, Liquid; D003599:Cytoskeleton / Q000187:drug effects; D015180:Electrophoresis, Gel, Two-Dimensional; D019570:Inhalation Exposure / Q000009:adverse effects; D008168:Lung / Q000737:chemistry* / Q000473:pathology; D051379:Mice; D008810:Mice, Inbred C57BL; D046988:Proteasome Endopeptidase Complex / Q000187:drug effects; D020543:Proteome / Q000032:analysis / Q000187:drug effects*; D040901:Proteomics / Q000379:methods*; D012906:Smoke / Q000009:adverse effects*; D053719:Tandem Mass Spectrometry; D062789:Tobacco Products
D000199:Actins; D000336:Aerosols; D020543:Proteome; D012906:Smoke; D046988:Proteasome Endopeptidase Complex
2D-PAGE; Data integration; Inhalation toxicology; Modified risk tobacco product; Quantitative proteomics; RPPA; Systems toxicology