Necroptosis Mediates Muscle Protein Degradation in a Cachexia Model of Weanling Pig with Lipopolysaccharide Challenge
Abstract
:1. Introduction
2. Results
2.1. LPS Challenge Dynamically Changes Physiological and Biochemical Indexes for Diagnosis of Cachexia in Weanling Pigs
2.2. LPS Challenge Dynamically Induces Inflammatory Response and Activates Muscle Protein Degradation Signaling Pathway in Weanling Pigs
2.3. LPS Challenge Dynamically Induces Muscle Necroptosis in Weanling Pigs
2.4. Nec-1 Inhibits Muscle Necroptosis Induced by LPS in Weanling Pigs
2.5. Inhibition of Necroptosis by Nec-1 Relieves Muscle Structure Injury, Inflammation, and Protein Degradation Induced by LPS in Weanling Pigs
3. Discussion
4. Materials and Methods
4.1. Animals and Experimental Design
4.2. Blood and Muscle Sample Collection
4.3. Plasma Hormone Measurement
4.4. Muscle Morphologic Analysis
4.5. Muscle Ultrastructural Observation
4.6. Muscle Protein and DNA Concentration Determination
4.7. mRNA Abundance Analysis
4.8. Protein Expression Analysis
4.9. Statistical Analysis
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Gene | Primer Sequence | Amplification Length | SERIAL NUMBER |
---|---|---|---|
TNF-α | F: TCCAATGGCAGAGTGGGTATG | 67 | NM_214022.1 |
R: AGCTGGTTGTCTTTCAGCTTCAC | |||
IL-1β | F: GCTAACTACGGTGACAACAATAATG | 186 | NM_214055.1 |
R: CTTCTCCACTGCCACGATGA | |||
IL-6 | F: AAGGTGATGCCACCTCAGAC | 151 | JQ839263.1 |
R: TCTGCCAGTACCTCCTTGCT | |||
Akt | F: GAAGAAGGAGGTCATCGT | 178 | NM_001159776.1 |
R: GGACAGGTGGAAGAAGAG | |||
FOXO-1 | F: TTCACCAGGCACCATCAT | 236 | NM_214014.2 |
R: GAGGAGAGTCGGAAGTAAGT | |||
MuFR1 | F: ATGGAGAACCTGGAGAAGCA | 219 | FJ905227.1 |
R: ACGGTCCATGATCACCTCAT | |||
MAFbx | F: TCACAGCTCACATCCCTGAG | 167 | NM_001044588.1 |
R: GACTTGCCGACTCTCTGGAC | |||
RIP1 | F: ACATCCTGTACGGCAACTCT | 175 | XM_005665538.2 |
R: CGGGTCCAGGTGTTTATCC | |||
RIP3 | F: CTTGTTGTCTGTCCGTGAGC | 238 | XM_001927424.3 |
R: GAGGAGGTTGGGCTGTTGA | |||
MLKL | F: TCTCGCTGCTGCTTCA | 105 | XM_013998184.1 |
R: CTCGCTTGTCTTCCTCTG | |||
GAPDH | F: CGTCCCTGAGACACGATGGT | 194 | AF017079.1 |
R: GCCTTGACTGTGCCGTGGAAT |
Antibodies (Host) | Catalog Number | Manufacturers | Dilution Ratio |
---|---|---|---|
RIP1 (rabbit) | LS-B8214 | LifeSpan BioSciences (Lynnwood, WA, USA) | 1:1000 |
p-RIP1 (rabbit) | 91702S | Cell Signaling Technology (Danvers, MA, USA) | 1:1000 |
RIP3 (rabbit) | SC-135170 | Santa Cruz Biotechnology (Dallas, TX, USA) | 1:1000 |
p-RIP1 (rabbit) | 91702S | Cell Signaling Technology | 1:1000 |
MLKL (rabbit) | 37705S | Cell Signaling Technology | 1:1000 |
p-MLKL (rabbit) | 62233S | Cell Signaling Technology | 1:1000 |
Akt (rabbit) | 9272S | Cell Signaling Technology | 1:1000 |
p-Akt (rabbit) | 9271S | Cell Signaling Technology | 1:1000 |
FOXO1 (rabbit) | 9454S | Cell Signaling Technology | 1:1000 |
p-FOXO1 (rabbit) | 9464S | Cell Signaling Technology | 1:1000 |
GAPDH (mouse) | FD0063 | Hangzhou Fude Biological Technology (Hangzhou, China) | 1:5000 |
IgG-HRP (goat anti-rabbit) | ANT020 | Wuhan Antejie Biotechnology(Wuhan, China) | 1:5000 |
IgG-HRP (goat anti-mouse) | ANT019 | Wuhan Antejie Biotechnology | 1:5000 |
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Guo, J.; Qin, X.; Wang, Y.; Li, X.; Wang, X.; Zhu, H.; Chen, S.; Zhao, J.; Xiao, K.; Liu, Y. Necroptosis Mediates Muscle Protein Degradation in a Cachexia Model of Weanling Pig with Lipopolysaccharide Challenge. Int. J. Mol. Sci. 2023, 24, 10923. https://doi.org/10.3390/ijms241310923
Guo J, Qin X, Wang Y, Li X, Wang X, Zhu H, Chen S, Zhao J, Xiao K, Liu Y. Necroptosis Mediates Muscle Protein Degradation in a Cachexia Model of Weanling Pig with Lipopolysaccharide Challenge. International Journal of Molecular Sciences. 2023; 24(13):10923. https://doi.org/10.3390/ijms241310923
Chicago/Turabian StyleGuo, Junjie, Xu Qin, Yang Wang, Xiangen Li, Xiuying Wang, Huiling Zhu, Shaokui Chen, Jiangchao Zhao, Kan Xiao, and Yulan Liu. 2023. "Necroptosis Mediates Muscle Protein Degradation in a Cachexia Model of Weanling Pig with Lipopolysaccharide Challenge" International Journal of Molecular Sciences 24, no. 13: 10923. https://doi.org/10.3390/ijms241310923