Untargeted Metabolomic Characteristics of Skeletal Muscle Dysfunction in Rabbits Induced by a High Fat Diet
Abstract
:Simple Summary
Abstract
1. Introduction
2. Materials and Methods
2.1. Ethics Statement
2.2. Animals and Feeding Strategy
2.3. Skeletal Muscle Tissue Collection and Preparation
2.4. Metabolomic Profiling
2.5. Metabolite Recognition and Data Preprocessing
2.6. Statistical Analysis
3. Results
3.1. Effect of HFD on Skeletal Tissue Metabolomics
3.2. Effects of HFD on the Metabolic Pathways of Endogenous Substances in Skeletal Muscle
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
- Lieb, W.; Sullivan, L.M.; Harris, T.B.; Roubenoff, R.; Benjamin, E.J.; Levy, D.; Fox, C.S.; Wang, T.J.; Wilson, P.W.; Kannel, W.B.; et al. Plasma leptin levels and incidence of heart failure, cardiovascular disease, and total mortality in elderly individuals. Diabetes Care 2008, 32, 612–616. [Google Scholar] [CrossRef] [Green Version]
- Rasool, S.; Geetha, T.; Broderick, T.L.; Babu, J.R. High fat with high sucrose diet leads to obesity and induces myodegeneration. Front. Physiol. 2018, 9, 1054. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Hruby, A.; Hu, F.B. The epidemiology of obesity: A big picture. Pharmacoeconomics 2015, 33, 673–689. [Google Scholar] [CrossRef]
- Bittencourt, A.; Brum, P.O.; Ribeiro, C.T.; Gasparotto, J.; Bortolin, R.C.; De Vargas, A.R.; Heimfarth, L.; De Almeida, R.F.; Moreira, J.C.F.; De Oliveira, J.; et al. High fat diet-induced obesity causes a reduction in brain tyrosine hydroxylase levels and non-motor features in rats through metabolic dysfunction, neuroinflammation and oxidative stress. Nutr. Neurosci. 2020, 1–15. [Google Scholar] [CrossRef]
- Merz, K.E.; Thurmond, D.C. Role of skeletal muscle in insulin resistance and glucose uptake. Compr. Physiol. 2020, 10, 785–809. [Google Scholar] [CrossRef]
- DeFronzo, R.A.; Tripathy, D. Skeletal muscle insulin resistance is the primary defect in type 2 diabetes. Diabetes Care 2009, 32, S157–S163. [Google Scholar] [CrossRef] [Green Version]
- Ferrannini, E.; Simonson, D.C.; Katz, L.D.; Reichard, G.; Bevilacqua, S.; Barrett, E.J.; Olsson, M.; DeFronzo, R.A. The disposal of an oral glucose load in patients with non-insulin-dependent diabetes. Metabolism 1988, 37, 79–85. [Google Scholar] [CrossRef]
- Thiebaud, D.; Jacot, E.; Defronzo, R.A.; Maeder, E.; Jequier, E.; Felber, J.P. The effect of graded doses of insulin on total glucose uptake, glucose oxidation, and glucose storage in man. Diabetes 1982, 31, 957–963. [Google Scholar] [CrossRef] [PubMed]
- DeFronzo, R.A.; Jacot, E.; Jequier, E.; Maeder, E.; Wahren, J.; Felber, J.P. The effect of insulin on the disposal of intravenous glucose: Results from Indirect calorimetry and hepatic and femoral venous catheterization. Diabetes 1981, 30, 1000–1007. [Google Scholar] [CrossRef] [PubMed]
- Donkers, S.J.; Nickel, D.; Paul, L.; Wiegers, S.R.; Knox, K.B. Adherence to physiotherapy-guided web-based exercise for persons with moderate-to-severe multiple sclerosis: A randomized controlled pilot study. Int. J. MS Care 2020, 22, 208–214. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Lee, K.; Jin, H.; Chei, S.; Oh, H.-J.; Lee, J.-Y.; Lee, B.-Y. Effect of dietary silk peptide on obesity, hyperglycemia, and skeletal muscle regeneration in high-fat diet-fed mice. Cells 2020, 9, 377. [Google Scholar] [CrossRef] [Green Version]
- Want, E.J.; Masson, P.; Michopoulos, F.; Wilson, I.D.; Theodoridis, G.; Plumb, R.S.; Shockcor, J.; Loftus, N.; Holmes, E.; Nicholson, J. Global metabolic profiling of animal and human tissues via UPLC-MS. Nat. Protoc. 2012, 8, 17–32. [Google Scholar] [CrossRef] [PubMed]
- Shao, J.; Wang, J.; Li, Y.; Elzo, M.A.; Tang, T.; Lai, T.; Ma, Y.; Gan, M.; Wang, L.; Jia, X.; et al. Growth, behavioural, serum biochemical and morphological changes in female rabbits fed high-fat diet. J. Anim. Physiol. Anim. Nutr. 2021, 105, 345–353. [Google Scholar] [CrossRef] [PubMed]
- Manickam, R.; Duszka, K.; Wahli, W. PPARs and microbiota in skeletal muscle health and wasting. Int. J. Mol. Sci. 2020, 21, 8056. [Google Scholar] [CrossRef]
- Borradaile, N.M.; Han, X.; Harp, J.D.; Gale, S.E.; Ory, D.S.; Schaffer, J.E. Disruption of endoplasmic reticulum structure and integrity in lipotoxic cell death. J. Lipid Res. 2006, 47, 2726–2737. [Google Scholar] [CrossRef] [Green Version]
- Wei, Y.; Wang, D.; Topczewski, F.; Pagliassotti, M.J. Saturated fatty acids induce endoplasmic reticulum stress and apoptosis independently of ceramide in liver cells. Am. J. Physiol. Metab. 2006, 291, E275–E281. [Google Scholar] [CrossRef]
- D’Souza, K.; Paramel, G.V.; Kienesberger, P.C. Lysophosphatidic acid signaling in obesity and insulin resistance. Nutrition 2018, 10, 399. [Google Scholar] [CrossRef] [Green Version]
- Okudaira, S.; Yukiura, H.; Aoki, J. Biological roles of lysophosphatidic acid signaling through its production by autotaxin. Biochimie 2010, 92, 698–706. [Google Scholar] [CrossRef]
- Dusaulcy, R.; Rancoule, C.; Grès, S.; Wanecq, E.; Colom, A.; Guigné, C.; van Meeteren, L.A.; Moolenaar, W.H.; Valet, P.; Saulnier-Blache, J.S. Adipose-specific disruption of autotaxin enhances nutritional fattening and reduces plasma lysophosphatidic acid. J. Lipid Res. 2011, 52, 1247–1255. [Google Scholar] [CrossRef] [Green Version]
- Rancoule, C.; Attane, C.; Greß, S.; Fournel, A.; Dusaulcy, R.; Bertrand, C.; Vinel, C.; Treguer, K.; Prentki, M.; Valet, P.; et al. Lysophosphatidic acid impairs glucose homeostasis and inhibits insulin secretion in high-fat diet obese mice. Diabetologia 2013, 56, 1394–1402. [Google Scholar] [CrossRef] [Green Version]
- Kim, T.; He, L.; Johnson, M.S.; Li, Y.; Zeng, L.; Ding, Y.; Long, Q.; Moore, J.F.; Sharer, J.D.; Nagy, T.R.; et al. Carnitine palmitoyltransferase 1b deficiency protects mice from diet-induced insulin resistance. J. Diabetes Metab. 2014, 5, 361. [Google Scholar] [CrossRef] [Green Version]
- Muoio, D.M. Metabolic inflexibility: When mitochondrial indecision leads to metabolic gridlock. Cell 2014, 159, 1253–1262. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Bruls, Y.M.; de Ligt, M.; Lindeboom, L.; Phielix, E.; Havekes, B.; Schaart, G.; Kornips, E.; Wildberger, J.E.; Hesselink, M.K.; Muoio, D.; et al. Carnitine supplementation improves metabolic flexibility and skeletal muscle acetylcarnitine formation in volunteers with impaired glucose tolerance: A randomised controlled trial. EBioMedicine 2019, 49, 318–330. [Google Scholar] [CrossRef] [Green Version]
- Randle, P.; Garland, P.; Hales, C.; Newsholme, E. The glucose fatty-acid cycle its role in insulin sensitivity and the metabolic disturbances of diabetes mellitus. Lancet 1963, 281, 785–789. [Google Scholar] [CrossRef]
- Stephens, F.B.; Constantin-Teodosiu, D.; Greenhaff, P.L. New insights concerning the role of carnitine in the regulation of fuel metabolism in skeletal muscle. J. Physiol. 2007, 581, 431–444. [Google Scholar] [CrossRef] [PubMed]
- Campbell, C.D.; Ganesh, J.; Ficicioglu, C. Two newborns with nutritional vitamin B12 deficiency: Challenges in newborn screening for vitamin B12 deficiency. Haematology 2005, 90, ECR45. [Google Scholar]
- Stephens, F.B.; Chee, C.; Wall, B.T.; Murton, A.; Shannon, C.E.; van Loon, L.J.; Tsintzas, K. Lipid-induced insulin resistance is associated with an impaired skeletal muscle protein synthetic response to amino acid ingestion in healthy young men. Diabetes 2014, 64, 1615–1620. [Google Scholar] [CrossRef] [Green Version]
- Bender, A.D. The metabolism of “surplus” amino acids. Br. J. Nutr. 2012, 108, S113–S121. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Schutz, Y. Protein turnover, ureagenesis and gluconeogenesis. Int. J. Vitam. Nutr. Res. 2011, 81, 101–107. [Google Scholar] [CrossRef] [PubMed]
- Hameed, A.; Mojsak, P.; Buczynska, A.; Suleria, H.A.R.; Kretowski, A.; Ciborowski, M. Altered metabolome of lipids and amino acids species: A source of early signature biomarkers of T2DM. J. Clin. Med. 2020, 9, 2257. [Google Scholar] [CrossRef]
- Settembre, C.; Ballabio, A. Lysosome: Regulator of lipid degradation pathways. Trends Cell Biol. 2014, 24, 743–750. [Google Scholar] [CrossRef] [Green Version]
- Newgard, C.B.; An, J.; Bain, J.R.; Muehlbauer, M.J.; Stevens, R.D.; Lien, L.F.; Haqq, A.M.; Shah, S.H.; Arlotto, M.; Slentz, C.A.; et al. A branched-chain amino acid-related metabolic signature that differentiates obese and lean humans and contributes to insulin resistance. Cell Metab. 2009, 9, 311–326. [Google Scholar]
- Wang-Sattler, R.; Yu, Z.; Herder, C.; Messias, A.C.; Floegel, A.; He, Y.; Heim, K.; Campillos, M.; Holzapfel, C.; Thorand, B.; et al. Novel biomarkers for pre-diabetes identified by metabolomics. Mol. Syst. Biol. 2012, 8, 615. [Google Scholar] [CrossRef] [PubMed]
- Floegel, A.; Stefan, N.; Yu, Z.; Mühlenbruch, K.; Drogan, D.; Joost, H.-G.; Fritsche, A.; Häring, H.-U.; De Angelis, M.H.; Peters, A.; et al. Identification of serum metabolites associated with risk of type 2 diabetes using a targeted metabolomic approach. Diabetes 2012, 62, 639–648. [Google Scholar] [CrossRef] [Green Version]
- Derave, W.; Everaert, I.; Beeckman, S.; Baguet, A. Muscle carnosine metabolism and beta-alanine supplementation in relation to exercise and training. Sports Med. 2010, 40, 247–263. [Google Scholar] [CrossRef] [Green Version]
- Dahl, T.A.; Midden, W.R.; Hartman, P.E. Some prevalent biomolecules as defenses against singlet oxygen damage. Photochem. Photobiol. 1988, 47, 357–362. [Google Scholar] [CrossRef]
- Kopple, J.D.; Swendseid, M.E. Effect of histidine intake of plasma and urine histidine levels, nitrogen balance and N tau-methylhistidine excretion in normal and chronically uremic men. J. Nutr. 1981, 111, 931–942. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Chan, W.K.M.; Decker, E.; Chow, C.K.; Boissonneault, G.A. Effect of dietary carnosine on plasma and tissue antioxidant concentrations and on lipid oxidation in rat skeletal muscle. Lipids 1994, 29, 461–466. [Google Scholar] [CrossRef]
- HoleČek, M.; Vodeničarovová, V. Effects of histidine supplementation on amino acid metabolism in rats. Physiol. Res. 2020, 69, 99–111. [Google Scholar] [CrossRef]
- Quirici, V.; Botero-Delgadillo, E.; González-Gómez, P.L.; Espíndola-Hernández, P.; Zambrano, B.; Cuevas, E.; Wingfield, J.C.; Vásquez, R.A. On the relationship between baseline corticosterone levels and annual survival of the thorn-tailed rayadito. Gen. Comp. Endocrinol. 2021, 300, 113635. [Google Scholar] [CrossRef] [PubMed]
- Sharma, A.K.; Shi, X.; Isales, C.M.; McGee-Lawrence, M.E. Endogenous glucocorticoid signaling in the regulation of bone and marrow adiposity: Lessons from metabolism and cross talk in other tissues. Curr. Osteoporos. Rep. 2019, 17, 438–445. [Google Scholar] [CrossRef]
- Uehara, M.; Yamazaki, H.; Yoshikawa, N.; Kuribara-Souta, A.; Tanaka, H. Correlation among body composition and metabolic regulation in a male mouse model of Cushing’s syndrome. Endocr. J. 2020, 67, 21–30. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Li, M.; Hu, X.; Xu, Y.; Hu, X.; Zhang, C.; Pang, S. A Possible mechanism of metformin in improving insulin resistance in diabetic rat models. Int. J. Endocrinol. 2019, 2019, 3248527. [Google Scholar] [CrossRef] [Green Version]
- Bonaldo, P.; Sandri, M. Cellular and molecular mechanisms of muscle atrophy. Dis. Model. Mech. 2013, 6, 25–39. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Adhikary, S.; Kothari, P.; Choudhary, D.; Tripathi, A.K.; Trivedi, R. Glucocorticoid aggravates bone micro-architecture deterioration and skeletal muscle atrophy in mice fed on high-fat diet. Steroids 2019, 149, 108416. [Google Scholar] [CrossRef]
- D’Souza, A.M.; Beaudry, J.L.; Szigiato, A.-A.; Trumble, S.J.; Snook, L.A.; Bonen, A.; Giacca, A.; Riddell, M.C. Consumption of a high-fat diet rapidly exacerbates the development of fatty liver disease that occurs with chronically elevated glucocorticoids. Am. J. Physiol. Liver Physiol. 2012, 302, G850–G863. [Google Scholar] [CrossRef] [Green Version]
- Appiakannan, H.S.; Kestyus, D.R.; Weber, E.T. Effects of high fat diet and chronic circadian challenge on glucocorticoid regulation in C57BL/6J mice. Physiol. Behav. 2019, 204, 100–105. [Google Scholar] [CrossRef] [PubMed]
Ingredient | SND | HFD | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Proportion (%) | DE (MJ/kg) | CP (%) | EE (%) | CF (%) | Ca (%) | P (%) | Proportion (%) | DE (MJ/kg) | CP (%) | EE (%) | CF (%) | Ca (%) | P (%) | |
Straw powder | 26 | 0.855 | 1.248 | 0.364 | 7.748 | 0.073 | 0.021 | 23.5 | 0.773 | 1.128 | 0.329 | 7.000 | 0.066 | 0.019 |
Maize | 18 | 2.889 | 1.602 | 0.648 | 0.576 | 0.005 | 0.070 | 16 | 2.568 | 1.424 | 0.576 | 0.512 | 0.004 | 0.062 |
Barley | 20 | 2.808 | 2.040 | 0.34 | 0.860 | 0.020 | 0.093 | 18 | 2.527 | 1.836 | 0.306 | 0.774 | 0.018 | 0.084 |
Bran | 15 | 1.631 | 2.310 | 2.475 | 0.765 | 0.050 | 0.072 | 13.5 | 1.468 | 2.079 | 2.228 | 0.689 | 0.045 | 0.065 |
Bean cake | 16 | 2.166 | 6.768 | 0.304 | 0.576 | 0.045 | 0.091 | 14.5 | 1.963 | 6.134 | 0.276 | 0.522 | 0.041 | 0.082 |
Fish meal | 3.5 | 0.552 | 2.047 | 0.196 | 0.137 | 0.104 | 3.15 | 0.497 | 1.842 | 0.176 | 0.123 | 0.094 | ||
Lard | 10 | 3.683 | 9.8 | |||||||||||
Stone powder | 1.0 | 0.350 | 0.9 | 0.315 | ||||||||||
Salt | 0.5 | 0.45 | ||||||||||||
Total | 100 | 10.91 | 16.015 | 4.327 | 10.525 | 0.68 | 0.431 | 100 | 13.479 | 14.443 | 13.691 | 9.497 | 0.621 | 0.406 |
Name | Formula | RT [min] | m/z | p-Value | VIP | Trend |
---|---|---|---|---|---|---|
PC (14:1e/3:0) | C25 H50 N O7 P | 14.594 | 508.33981 | 0.00060252 | 1.96834418 | ↓ |
ACar 10:2 | C17 H30 N O4 | 10.771 | 312.21707 | 0.001022871 | 2.78164921 | ↑ |
Hexanoylcarnitine | C13 H25 N O4 | 8.781 | 260.18542 | 0.001221548 | 2.134885168 | ↑ |
Phenylacetylglycine | C10 H11 N O3 | 8.22 | 194.08138 | 0.002987006 | 1.562157041 | ↓ |
LPC 15:1 | C23 H46 N O7 P | 14.323 | 480.30856 | 0.004218472 | 1.679235509 | ↓ |
Palmitoylcarnitine | C23 H45 N O4 | 13.56 | 400.34161 | 0.004662458 | 2.728685353 | ↑ |
ACar 22:6 | C29 H46 N O4 | 13.404 | 472.34155 | 0.005910097 | 2.009955994 | ↑ |
5-chloro-2,8-dimethyl-4-[(3-nitro-2-pyridyl)oxy]quinoline | C16 H12 Cl N3 O3 | 1.602 | 330.05746 | 0.006079831 | 1.752888258 | ↓ |
ACar 18:1 | C25 H48 N O4 | 13.664 | 426.35709 | 0.006620219 | 2.680982815 | ↑ |
ACar 7:0 | C14 H28 N O4 | 9.819 | 274.20126 | 0.007692279 | 2.101077514 | ↑ |
ACar 17:2 | C24 H44 N O4 | 13.239 | 410.32602 | 0.010393589 | 2.48157315 | ↑ |
ACar 18:2 | C25 H46 N O4 | 13.426 | 424.34134 | 0.010520878 | 2.366450915 | ↑ |
Ala-Gln | C8 H15 N3 O4 | 1.305 | 218.11295 | 0.011721656 | 2.174643548 | ↑ |
ACar 16:1 | C23 H44 N O4 | 13.29 | 398.32599 | 0.011757863 | 2.207576851 | ↑ |
N-Acetyl-L-carnosine | C11 H16 N4 O4 | 1.381 | 269.12411 | 0.012378517 | 1.528953387 | ↑ |
LPC 22:6 | C30 H50 N O7 P | 14.45 | 568.33807 | 0.016506546 | 1.321877932 | ↑ |
PC (14:1e/2:0) | C24 H48 N O7 P | 14.352 | 494.32413 | 0.016550657 | 1.102232178 | ↓ |
ACar 20:5 | C27 H44 N O4 | 13.206 | 446.32581 | 0.017118755 | 2.11188684 | ↑ |
Muscone | C16 H30 O | 14.308 | 239.23694 | 0.018553432 | 1.998565512 | ↓ |
ACar 20:4 | C27 H46 N O4 | 13.413 | 448.3414 | 0.019178682 | 2.014369263 | ↑ |
Propionylcarnitine | C10 H19 N O4 | 2.882 | 218.13879 | 0.020501301 | 1.659387379 | ↓ |
ACar 17:1 | C24 H46 N O4 | 13.479 | 412.34155 | 0.024652794 | 2.259139096 | ↑ |
ACar 20:3 | C27 H48 N O4 | 13.578 | 450.35712 | 0.027362927 | 2.267210155 | ↑ |
LPC 15:0 | C23 H48 N O7 P | 14.46 | 482.32388 | 0.027825481 | 1.548906089 | ↓ |
ACar 15:1 | C22 H42 N O4 | 13.095 | 384.3103 | 0.029353037 | 2.072617982 | ↑ |
LPE 17:0 | C22 H46 N O7 P | 15.085 | 468.30875 | 0.030453786 | 1.236219 | ↓ |
Corticosterone | C21 H30 O4 | 11.761 | 347.22134 | 0.032938755 | 2.163429757 | ↓ |
ACar 15:0 | C22 H44 N O4 | 13.365 | 386.3259 | 0.032970783 | 2.228659726 | ↑ |
1-Palmitoyl-Sn-Glycero-3-Phosphocholine | C24 H50 N O7 P | 13.491 | 496.33948 | 0.033163212 | 1.248603083 | ↓ |
ACar 18:3 | C25 H44 N O4 | 13.203 | 422.32593 | 0.034156704 | 2.196673054 | ↑ |
ACar 12:1 | C19 H36 N O4 | 12.16 | 342.26349 | 0.034297235 | 2.186103895 | ↑ |
Tetrahydrocorticosterone | C21 H34 O4 | 12.513 | 368.27911 | 0.035427687 | 2.2470103 | ↑ |
Prolylleucine | C11 H20 N2 O3 | 2.098 | 229.15422 | 0.035890864 | 1.53343442 | ↓ |
DL-Carnitine | C7 H15 N O3 | 1.31 | 162.11217 | 0.036302614 | 2.164262278 | ↓ |
L-Histidine | C6 H9 N3 O2 | 1.619 | 156.07664 | 0.036883235 | 2.296408916 | ↑ |
Carnosine | C9 H14 N4 O3 | 1.602 | 227.11374 | 0.038157621 | 1.374481323 | ↑ |
Lysops 22:5 | C28 H46 N O9 P | 14.594 | 572.29755 | 0.043926394 | 1.145501212 | ↑ |
Betaine | C5 H11 N O2 | 1.32 | 118.0863 | 0.046878628 | 1.626178977 | ↓ |
ACar 18:0 | C25 H50 N O4 | 13.889 | 428.37296 | 0.049277304 | 1.980841845 | ↑ |
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. |
© 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
Share and Cite
Fan, H.; Li, Y.; Wang, J.; Shao, J.; Tang, T.; Elzo, M.A.; Wang, L.; Lai, T.; Ma, Y.; Gan, M.; et al. Untargeted Metabolomic Characteristics of Skeletal Muscle Dysfunction in Rabbits Induced by a High Fat Diet. Animals 2021, 11, 1722. https://doi.org/10.3390/ani11061722
Fan H, Li Y, Wang J, Shao J, Tang T, Elzo MA, Wang L, Lai T, Ma Y, Gan M, et al. Untargeted Metabolomic Characteristics of Skeletal Muscle Dysfunction in Rabbits Induced by a High Fat Diet. Animals. 2021; 11(6):1722. https://doi.org/10.3390/ani11061722
Chicago/Turabian StyleFan, Huimei, Yanhong Li, Jie Wang, Jiahao Shao, Tao Tang, Mauricio A. Elzo, Li Wang, Tianfu Lai, Yuan Ma, Mingchuan Gan, and et al. 2021. "Untargeted Metabolomic Characteristics of Skeletal Muscle Dysfunction in Rabbits Induced by a High Fat Diet" Animals 11, no. 6: 1722. https://doi.org/10.3390/ani11061722
APA StyleFan, H., Li, Y., Wang, J., Shao, J., Tang, T., Elzo, M. A., Wang, L., Lai, T., Ma, Y., Gan, M., Jia, X., & Lai, S. (2021). Untargeted Metabolomic Characteristics of Skeletal Muscle Dysfunction in Rabbits Induced by a High Fat Diet. Animals, 11(6), 1722. https://doi.org/10.3390/ani11061722