Register      Login
Reproduction, Fertility and Development Reproduction, Fertility and Development Society
Vertebrate reproductive science and technology
RESEARCH ARTICLE

TallyHO obese female mice experience poor reproductive outcomes and abnormal blastocyst metabolism that is reversed by metformin

Erica D. Louden A , Kerri M. Luzzo A , Patricia T. Jimenez A , Tiffany Chi A , Maggie Chi A and Kelle H. Moley A B
+ Author Affiliations
- Author Affiliations

A Department of Obstetrics and Gynecology, Washington University School of Medicine, 425 S. Euclid Ave, Campus Box 8064, St Louis, MO 63110, USA.

B Corresponding author. Email: moleyk@wustl.edu

Reproduction, Fertility and Development 27(1) 31-39 https://doi.org/10.1071/RD14339
Published: 4 December 2014

Abstract

Obese women experience worse reproductive outcomes than normal weight women, specifically infertility, pregnancy loss, fetal malformations and developmental delay of offspring. The aim of the present study was to use a genetic mouse model of obesity to recapitulate the human reproductive phenotype and further examine potential mechanisms and therapies. New inbred, polygenic Type 2 diabetic TallyHO mice and age-matched control C57BL/6 mice were superovulated to obtain morula or blastocyst stage embryos that were cultured in human tubal fluid (HTF) medium. Deoxyglucose uptake was determined for individual insulin-stimulated blastocysts. Apoptosis was detected by confocal microscopy using the terminal deoxyribonucleotidyl transferase-mediated dUTP–digoxigenin nick end-labelling (TUNEL) assay and Topro-3 nuclear dye. Embryos were scored for TUNEL-positive as a percentage of total nuclei. AMP-activated protein kinase (AMPK) activation, tumour necrosis factor (TNF)-α expression and adiponectin expression were analysed by western immunoblot and confocal immunofluorescent microscopy. Lipid accumulation was assayed by BODIPY. Comparisons were made between TallyHO morulae cultured to blastocyst embryos in either HTF medium or HTF medium with 25 μg mL–1 metformin. TallyHO mice developed whole body abnormal insulin tolerance, had decreased litter sizes and increased non-esterified fatty acid levels. Blastocysts from TallyHO mice exhibited increased apoptosis, decreased insulin sensitivity and decreased AMPK. A possible cause for the insulin resistance and abnormal AMPK phosphorylation was the increased TNF-α expression and lipid accumulation, as detected by BODIPY, in TallyHO blastocysts and decreased adiponectin. Culturing TallyHO morulae with the AMPK activator metformin led to a reversal of all the abnormal findings, including increased AMPK phosphorylation, improved insulin-stimulated glucose uptake and normalisation of lipid accumulation. Women with obesity and insulin resistance experience poor pregnancy outcomes. Previously we have shown in mouse models of insulin resistance that AMPK activity is decreased and that activators of AMPK reverse poor embryo outcomes. Here, we show for the first time using a genetically altered obese model, not a diet-induced model, that metformin reverses many of the adverse effects of obesity at the level of the blastocyst. Expanding on this we determine that activation of AMPK via metformin reduces lipid droplet accumulation, presumably by eliminating the inhibitory effects of TNF-α, resulting in normalisation of fatty acid oxidation and HADH2 (hydroxyacyl-CoA dehydrogenase/3-ketoacyl-CoA thiolase/enoyl-CoA hydratase (trifunctional protein), alpha subunit) activity. Metformin exposure in vitro was able to partially reverse these effects, at the level of the blastocyst, and may thus be effective in preventing the adverse effects of obesity on pregnancy and reproductive outcomes.

Additional keywords: insulin sensitivity, lipid accumulation, mitochondrial metabolism, obesity, preimplantation embryos.


References

Allemand, M. C., Irving, B. A., Asmann, Y. W., Klaus, K. A., Tatpati, L., Coddington, C. C., and Nair, K. S. (2009). Effect of testosterone on insulin stimulated IRS1 Ser phosphorylation in primary rat myotubes: a potential model for PCOS-related insulin resistance. PLoS ONE 4, e4274.
Effect of testosterone on insulin stimulated IRS1 Ser phosphorylation in primary rat myotubes: a potential model for PCOS-related insulin resistance.Crossref | GoogleScholarGoogle Scholar | 19169352PubMed |

Antuna-Puente, B., Feve, B., Fellahi, S., and Bastard, J. P. (2008). Adipokines: the missing link between insulin resistance and obesity. Diabetes Metab. 34, 2–11.
Adipokines: the missing link between insulin resistance and obesity.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1cXkslOksr8%3D&md5=50fabb6e83b3d02c8f83ed3a8aace8cdCAS | 18093861PubMed |

Balkwill, F. (2009). Tumour necrosis factor and cancer. Nat. Rev. Cancer 9, 361–371.
Tumour necrosis factor and cancer.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXjvF2gtLk%3D&md5=653ff15846f65ac5ccca5ca493d51692CAS | 19343034PubMed |

Barb, D., Williams, C. J., Neuwirth, A. K., and Mantzoros, C. S. (2007). Adiponectin in relation to malignancies: a review of existing basic research and clinical evidence. Am. J. Clin. Nutr. 86, s858–s866.
| 18265479PubMed |

Blackmore, H. L., and Ozanne, S. E (2013). Maternal diet-induced obesity and offspring cardiovascular health. J. Dev. Orig. Health Dis. 4, 338–347.
Maternal diet-induced obesity and offspring cardiovascular health.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DC%2BC2cfntlCitg%3D%3D&md5=5bf76fc0c7298c10292753b69fd2f2deCAS | 24970727PubMed |

Chi, M. M., Pingsterhaus, J., Carayannopoulos, M., and Moley, K. H. (2000). Decreased glucose transporter expression triggers BAX-dependent apoptosis in the murine blastocyst. J. Biol. Chem. 275, 40 252–40 257.
Decreased glucose transporter expression triggers BAX-dependent apoptosis in the murine blastocyst.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3MXhsFelsw%3D%3D&md5=07eb7d1327c114bc4586e95756a97463CAS |

Chi, M. M., Hoehn, A., and Moley, K. H. (2002). Metabolic changes in the glucose-induced apoptotic blastocyst suggest alterations in mitochondrial physiology. Am. J. Physiol. Endocrinol. Metab. 283, E226–E232.
| 1:CAS:528:DC%2BD38XmslSgs70%3D&md5=41dfedafabe19cc9cdfefed5843654c8CAS | 12110526PubMed |

Claret, M., Smith, M. A., Batterham, R. L., Selman, C., Choudhury, A. I., Fryer, L. G., Clements, M., Al-Qassab, H., Heffron, H., Xu, A. W., Speakman, J. R., Barsh, G. S., Viollet, B., Vaulont, S., Ashford, M. L., Carling, D., and Withers, D. J. (2007). AMPK is essential for energy homeostasis regulation and glucose sensing by POMC and AgRP neurons. J. Clin. Invest. 117, 2325–2336.
AMPK is essential for energy homeostasis regulation and glucose sensing by POMC and AgRP neurons.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2sXoslOnsLo%3D&md5=0f5fd564981045f552cdcb1333f679a0CAS | 17671657PubMed |

Cocksedge, K. A., Li, T. C., Saravelos, S. H., and Metwally, M. (2008). A reappraisal of the role of polycystic ovary syndrome in recurrent miscarriage. Reprod. Biomed. Online 17, 151–160.
A reappraisal of the role of polycystic ovary syndrome in recurrent miscarriage.Crossref | GoogleScholarGoogle Scholar | 18616903PubMed |

Dale, P. O., Tanbo, T., Haug, E., and Abyholm, T. (1998). The impact of insulin resistance on the outcome of ovulation induction with low-dose follicle stimulating hormone in women with polycystic ovary syndrome. Hum. Reprod. 13, 567–570.
The impact of insulin resistance on the outcome of ovulation induction with low-dose follicle stimulating hormone in women with polycystic ovary syndrome.Crossref | GoogleScholarGoogle Scholar | 1:STN:280:DyaK1c3jt1Whtg%3D%3D&md5=1d05833403f98c7eb23d920bd21461a3CAS | 9572412PubMed |

Dzamko, N. L., and Steinberg, G. R. (2009). AMPK-dependent hormonal regulation of whole-body energy metabolism. Acta Physiol. (Oxf.) 196, 115–127.
AMPK-dependent hormonal regulation of whole-body energy metabolism.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXlt1KrtLo%3D&md5=b581d650483061ce828e0952b4315cf5CAS | 19245657PubMed |

Eng, G. S., Sheridan, R. A., Wyman, A., Chi, M. M., Bibee, K. P., Jungheim, E. S., and Moley, K. H. (2007). AMP kinase activation increases glucose uptake, decreases apoptosis, and improves pregnancy outcome in embryos exposed to high IGF-I concentrations. Diabetes 56, 2228–2234.
AMP kinase activation increases glucose uptake, decreases apoptosis, and improves pregnancy outcome in embryos exposed to high IGF-I concentrations.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2sXhtV2gs7bI&md5=c9962d52dc86a5275e4680fc81db6a04CAS | 17575082PubMed |

Fain, J. N., Buehrer, B., Tichansky, D. S., and Madan, A. K. (2008). Regulation of adiponectin release and demonstration of adiponectin mRNA as well as release by the non-fat cells of human omental adipose tissue. Int. J. Obes. (Lond.) 32, 429–435.
Regulation of adiponectin release and demonstration of adiponectin mRNA as well as release by the non-fat cells of human omental adipose tissue.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1cXjsVSjsLc%3D&md5=8266fe0efa8332b7b945c9af0e7f4159CAS |

Flegal, K. M., Carroll, M. D., Kit, B. K., and Ogden, C. L. (2012). Prevalence of obesity and trends in the distribution of body mass index among US adults, 1999–2010. JAMA 307, 491–497.
Prevalence of obesity and trends in the distribution of body mass index among US adults, 1999–2010.Crossref | GoogleScholarGoogle Scholar | 22253363PubMed |

Hammond, R. A. (2009). Complex systems modeling for obesity research. Prev. Chronic Dis. 6, A97.
| 19527598PubMed |

Igosheva, N., Abramov, A. Y., Poston, L., Eckert, J. J., Fleming, T. P., Duchen, M. R., and McConnell, J. (2010). Maternal diet-induced obesity alters mitochondrial activity and redox status in mouse oocytes and zygotes. PLoS ONE 5, e10074.
Maternal diet-induced obesity alters mitochondrial activity and redox status in mouse oocytes and zygotes.Crossref | GoogleScholarGoogle Scholar | 20404917PubMed |

Isakson, P., Hammarstedt, A., Gustafson, B., and Smith, U. (2009). Impaired preadipocyte differentiation in human abdominal obesity: role of Wnt, tumor necrosis factor-alpha, and inflammation. Diabetes 58, 1550–1557.
Impaired preadipocyte differentiation in human abdominal obesity: role of Wnt, tumor necrosis factor-alpha, and inflammation.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXos1Cquro%3D&md5=4d9e0e5bbf4731a1c5e93c217b098792CAS | 19351711PubMed |

Jimenez, P. T., Frolova, A. I., Chi, M. M., Grindler, N. M., Willcockson, A. R., Reynolds, K. A., Zhao, Q., and Moley, K. H. (2013). DHEA-mediated inhibition of the pentose phosphate pathway alters oocyte lipid metabolism in mice. Endocrinology 154, 4835–4844.
DHEA-mediated inhibition of the pentose phosphate pathway alters oocyte lipid metabolism in mice.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3sXhvV2itL7E&md5=a9fb8b574e7eaf6af3f5d69c366ad366CAS | 24036000PubMed |

Jungheim, E. S., and Moley, K. H. (2010). Current knowledge of obesity’s effects in the pre- and periconceptional periods and avenues for future research. Am. J. Obstet. Gynecol. 203, 525–530.
Current knowledge of obesity’s effects in the pre- and periconceptional periods and avenues for future research.Crossref | GoogleScholarGoogle Scholar | 20739012PubMed |

Jungheim, E. S., Schoeller, E. L., Marquard, K. L., Louden, E. D., Schaffer, J. E., and Moley, K. H. (2010). Diet-induced obesity model: abnormal oocytes and persistent growth abnormalities in the offspring. Endocrinology 151, 4039–4046.
Diet-induced obesity model: abnormal oocytes and persistent growth abnormalities in the offspring.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXht1Oju7rN&md5=2a66eae9ecb5a3078757a6b784deea0cCAS | 20573727PubMed |

Jungheim, E. S., Louden, E. D., Chi, M. M., Frolova, A. I., Riley, J. K., and Moley, K. H. (2011). Preimplantation exposure of mouse embryos to palmitic acid results in fetal growth restriction followed by catch-up growth in the offspring. Biol. Reprod. 85, 678–683.
Preimplantation exposure of mouse embryos to palmitic acid results in fetal growth restriction followed by catch-up growth in the offspring.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3MXht12ht7zE&md5=7a6bcf18ed14ec50a617db9709291711CAS | 21653893PubMed |

Kim, J. H., Sen, S., Avery, C. S., Simpson, E., Chandler, P., Nishina, P. M., Churchill, G. A., and Naggert, J. K. (2001). Genetic analysis of a new mouse model for non-insulin-dependent diabetes. Genomics 74, 273–286.
Genetic analysis of a new mouse model for non-insulin-dependent diabetes.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3MXksVChsLc%3D&md5=1154f80795bb54f15f5ddd89a75483e2CAS | 11414755PubMed |

Kim, J. H., Stewart, T. P., Zhang, W., Kim, H. Y., Nishina, P. M., and Naggert, J. K. (2005). Type 2 diabetes mouse model TallyHO carries an obesity gene on chromosome 6 that exaggerates dietary obesity. Physiol. Genomics 22, 171–181.
Type 2 diabetes mouse model TallyHO carries an obesity gene on chromosome 6 that exaggerates dietary obesity.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2MXpvFOltrc%3D&md5=31ce49c57ef74faab6f89eb2dbadd53cCAS | 15870394PubMed |

Kim, S. T., Marquard, K., Stephens, S., Louden, E., Allsworth, J., and Moley, K. H. (2011). Adiponectin and adiponectin receptors in the mouse preimplantation embryo and uterus. Hum. Reprod. 26, 82–95.
Adiponectin and adiponectin receptors in the mouse preimplantation embryo and uterus.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXhs1Wqu7%2FN&md5=8d0d515471c2f088ea27c2faaa0aae55CAS | 21106494PubMed |

Li, L., Yang, G., Shi, S., Yang, M., Liu, H., and Boden, G. (2009). The adipose triglyceride lipase, adiponectin and visfatin are downregulated by tumor necrosis factor-alpha (TNF-alpha) in vivo. Cytokine 45, 12–19.
The adipose triglyceride lipase, adiponectin and visfatin are downregulated by tumor necrosis factor-alpha (TNF-alpha) in vivo.Crossref | GoogleScholarGoogle Scholar | 19026557PubMed |

Li, H., Chen, X., Guan, L., Qi, Q., Shu, G., Jiang, Q., Yuan, L., Xi, Q., and Zhang, Y. (2013). MiRNA-181a regulates adipogenesis by targeting tumor necrosis factor-alpha (TNF-alpha) in the porcine model. PLoS ONE 8, e71568.
MiRNA-181a regulates adipogenesis by targeting tumor necrosis factor-alpha (TNF-alpha) in the porcine model.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3sXhsFOrt77P&md5=ab8406eec0a16a2b198fcdbdb9689efdCAS | 24098322PubMed |

Louden, E., Chi, M. M., and Moley, K. H. (2008). Crosstalk between the AMP-activated kinase and insulin signaling pathways rescues murine blastocyst cells from insulin resistance. Reproduction 136, 335–344.
Crosstalk between the AMP-activated kinase and insulin signaling pathways rescues murine blastocyst cells from insulin resistance.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1cXht1GmtL3N&md5=eed5814f0bc6ed41c171c2fc68ce8a7cCAS | 18577554PubMed |

Luzzo, K. M., Wang, Q., Purcell, S. H., Chi, M., Jimenez, P. T., Grindler, N., Schedl, T., and Moley, K. H. (2012). High fat diet induced developmental defects in the mouse: oocyte meiotic aneuploidy and fetal growth retardation/brain defects. PLoS ONE 7, e49217.
High fat diet induced developmental defects in the mouse: oocyte meiotic aneuploidy and fetal growth retardation/brain defects.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC38XhslKjtrvN&md5=77377d5e28d49e82d44d033465038ccfCAS | 23152876PubMed |

Ogden, C. L., Carroll, M. D., Kit, B. K., and Flegal, K. M. (2012). ‘Prevalence of Obesity in the United States, 2009–2010.’ NCHS Data Brief, volume 82. pp. 1–8.

Passonneau, J. V., and Lowry, O. H. (1993) ‘Enzymatic Analysis: A Practical Guide.’ (Humana Press: New York.)

Riley, J. K., Carayannopoulos, M. O., Wyman, A. H., Chi, M., and Moley, K. H. (2006). Phosphatidylinositol 3-kinase activity is critical for glucose metabolism and embryo survival in murine blastocysts. J. Biol. Chem. 281, 6010–6019.
Phosphatidylinositol 3-kinase activity is critical for glucose metabolism and embryo survival in murine blastocysts.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD28XhvVeju7k%3D&md5=e3e5069ec7ac2e82d21557c4a252ffb4CAS | 16272157PubMed |

Sam, S., and Mazzone, T. (2014). Adipose tissue changes in obesity and the impact on metabolic function. Transl. Res , .
Adipose tissue changes in obesity and the impact on metabolic function.Crossref | GoogleScholarGoogle Scholar | 24929206PubMed |

Shibata, R., Sato, K., Pimentel, D. R., Takemura, Y., Kihara, S., Ohashi, K., Funahashi, T., Ouchi, N., and Walsh, K. (2005). Adiponectin protects against myocardial ischemia–reperfusion injury through AMPK- and COX-2-dependent mechanisms. Nat. Med. 11, 1096–1103.
Adiponectin protects against myocardial ischemia–reperfusion injury through AMPK- and COX-2-dependent mechanisms.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD2MXhtVOiurfK&md5=e720fdcdf9c9f1e9b50ad080975cd082CAS | 16155579PubMed |

Steinberger, J., Daniels, S. R., American Heart Association Atherosclerosis, Hypertension, and Obesity in the Young Committee (Council on Cardiovascular Disease in the Young) American Heart Association Diabetes Committee (Council on Nutrition, Physical Activity, and Metabolism) (2003). Obesity, insulin resistance, diabetes, and cardiovascular risk in children: an American Heart Association scientific statement from the Atherosclerosis, Hypertension, and Obesity in the Young Committee (Council on Cardiovascular Disease in the Young) and the Diabetes Committee (Council on Nutrition, Physical Activity, and Metabolism). Circulation 107, 1448–1453.
Obesity, insulin resistance, diabetes, and cardiovascular risk in children: an American Heart Association scientific statement from the Atherosclerosis, Hypertension, and Obesity in the Young Committee (Council on Cardiovascular Disease in the Young) and the Diabetes Committee (Council on Nutrition, Physical Activity, and Metabolism).Crossref | GoogleScholarGoogle Scholar | 12642369PubMed |

Street, M. E., Volta, C., Ziveri, M. A., Viani, I., and Bernasconi, S. (2009). Markers of insulin sensitivity in placentas and cord serum of intrauterine growth-restricted newborns. Clin. Endocrinol. (Oxf.) 71, 394–399.
Markers of insulin sensitivity in placentas and cord serum of intrauterine growth-restricted newborns.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXhtFagtL%2FE&md5=5f166acc385572182b3433782f86aab8CAS | 19226262PubMed |

Sweet, I. R., Gilbert, M., Maloney, E., Hockenbery, D. M., Schwartz, M. W., and Kim, F. (2009). Endothelial inflammation induced by excess glucose is associated with cytosolic glucose 6-phosphate but not increased mitochondrial respiration. Diabetologia 52, 921–931.
Endothelial inflammation induced by excess glucose is associated with cytosolic glucose 6-phosphate but not increased mitochondrial respiration.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXktVOrs7Y%3D&md5=fdbcd911a5b6f2ba7910a6ae26e9256bCAS | 19219423PubMed |

Tschritter, O., Fritsche, A., Thamer, C., Haap, M., Shirkavand, F., Rahe, S., Staiger, H., Maerker, E., Haring, H., and Stumvoll, M. (2003). Plasma adiponectin concentrations predict insulin sensitivity of both glucose and lipid metabolism. Diabetes 52, 239–243.
Plasma adiponectin concentrations predict insulin sensitivity of both glucose and lipid metabolism.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD3sXhtFagu70%3D&md5=0ba7091723450a8954c8649f3fbb87c8CAS | 12540592PubMed |

Waller, D. K., Shaw, G. M., Rasmussen, S. A., Hobbs, C. A., Canfield, M. A., Siega-Riz, A. M., Gallaway, M. S., Correa, A., National Birth Defects Prevention Study (2007). Prepregnancy obesity as a risk factor for structural birth defects. Arch. Pediatr. Adolesc. Med. 161, 745–750.
Prepregnancy obesity as a risk factor for structural birth defects.Crossref | GoogleScholarGoogle Scholar | 17679655PubMed |

Wu, L. L., Dunning, K. R., Yang, X., Russell, D. L., Lane, M., Norman, R. J., and Robker, R. L. (2010). High-fat diet causes lipotoxicity responses in cumulus–oocyte complexes and decreased fertilization rates. Endocrinology 151, 5438–5445.
High-fat diet causes lipotoxicity responses in cumulus–oocyte complexes and decreased fertilization rates.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BC3cXhsVKks7bL&md5=34ca33a770155dc3a74cdf7f220e3190CAS | 20861227PubMed |

Xu, X. J., Valentine, R. J., and Ruderman, N. B. (2014). AMP-activated protein kinase (AMPK): does this master regulator of cellular energy state distinguish insulin sensitive from insulin resistant obesity? Curr. Obes. Rep. 3, 248–255.
AMP-activated protein kinase (AMPK): does this master regulator of cellular energy state distinguish insulin sensitive from insulin resistant obesity?Crossref | GoogleScholarGoogle Scholar | 24891985PubMed |

Yamauchi, T., Kamon, J., Minokoshi, Y., Ito, Y., Waki, H., Uchida, S., Yamashita, S., Noda, M., Kita, S., Ueki, K., Eto, K., Akanuma, Y., Froguel, P., Foufelle, F., Ferre, P., Carling, D., Kimura, S., Nagai, R., Kahn, B. B., and Kadowaki, T. (2002). Adiponectin stimulates glucose utilization and fatty-acid oxidation by activating AMP-activated protein kinase. Nat. Med. 8, 1288–1295.
Adiponectin stimulates glucose utilization and fatty-acid oxidation by activating AMP-activated protein kinase.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD38XotlWlsrg%3D&md5=846215a1cb613a3b6d9898932a398623CAS | 12368907PubMed |

Yoo, H. J., and Choi, K. M. (2014). Adipokines as a novel link between obesity and atherosclerosis. World J. Diabetes 5, 357–363.
Adipokines as a novel link between obesity and atherosclerosis.Crossref | GoogleScholarGoogle Scholar | 24936256PubMed |

Zhang, S., and Kim, K. H. (1995). TNF-alpha inhibits glucose-induced insulin secretion in a pancreatic beta-cell line (INS-1). FEBS Lett. 377, 237–239.
TNF-alpha inhibits glucose-induced insulin secretion in a pancreatic beta-cell line (INS-1).Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DyaK28Xnt1er&md5=5093ea2f9ba0628892a8523200c42efdCAS | 8543058PubMed |

Zhang, Z., Zhao, M., Li, Q., Zhao, H., Wang, J., and Li, Y. (2009). Acetyl-l-carnitine inhibits TNF-alpha-induced insulin resistance via AMPK pathway in rat skeletal muscle cells. FEBS Lett. 583, 470–474.
Acetyl-l-carnitine inhibits TNF-alpha-induced insulin resistance via AMPK pathway in rat skeletal muscle cells.Crossref | GoogleScholarGoogle Scholar | 1:CAS:528:DC%2BD1MXotlKrsg%3D%3D&md5=dc87e713fead30349e621cda1b543d14CAS | 19121314PubMed |