Tumour necrosis factor alpha induces neuroinflammation and insulin resistance in immortalized hypothalamic neurons through independent pathways

Abstract

The links between obesity, inflammation and insulin resistance, all key characteristics of Type 2 diabetes mellitus (T2DM), are yet to be delineated in the brain. One of the key neuroinflammatory proteins detected in the hypothalamus with over‐nutrition is tumour necrosis factor alpha (TNFα). Using immortalized embryonic rat and mouse hypothalamic cell lines (rHypoE‐ 7 and mHypoE‐46) that express orexigenic neuropeptide Y (NPY) and agouti‐related peptide (AgRP), we studied changes in insulin signalling and inflammatory gene marker mRNA expression after TNFα exposure. A qPCR array of 84 inflammatory markers (cytokines, chemokines, and receptors) demonstrated an increase in the expression of multiple genes encoding inflammatory markers upon exposure to 100 ng/mL TNFα for 4 h. Furthermore, neurons pre‐exposed to TNFα (50 ng/mL) for 6 or 16 h exhibited a significant reduction in pAkt compared to control after insulin treatment, indicating the attenuation of insulin signalling. The mRNA expression of insulin signalling related genes were also decreased with exposure to TNFα. TNFα significantly increased mRNA expression of IκBα, Tnfrsf1a, and IL6 at 4 and 24 h, activating a pro‐inflammatory state. An inhibitor study using an IKK‐β inhibitor, PS1145, demonstrated that TNFα‐induced neuroinflammatory marker expression occurs through the IKK‐β/NF‐κB pathway, whereas oleate, a monounsaturated fatty acid, had no effect on inflammatory markers. To test the efficacy of anti‐inflammatory treatment to reverse insulin resistance, neurons were treated with TNFα and PS1145, which did not significantly restore the TNFα‐induced changes in cellular insulin sensitivity, indicating an alternative pathway may be involved. In conclusion, exposure to the inflammatory cytokine TNFα causes cellular insulin resistance and inflammation marker expression in the rHypoE‐7 and mHypoE‐46 neurons, consistent with effects seen with TNFα in peripheral tissues. It also mimics insulin‐ and palmitate‐induced insulin resistance in hypothalamic neurons. This study provides further evidence that altered central energy metabolism may be caused by obesity‐induced cytokine expression.

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