Immune modulating effects of additional supplementation of estradiol combined with testosterone in murine testosterone-deficient NAFLD model.

Nonalcoholic fatty liver disease (NAFLD) is associated with testosterone deficiency. However, NAFLD patients generally do not respond to treatment with testosterone alone. We investigated the innate immune mechanisms underlying the effects of treatment with testosterone alone, estrogen alone, or combined testosterone and estrogen on high-fat diet (HFD)-induced NAFLD due to testosterone deficiency. Orchiectomized (OCX) male Rag2-/- mice were used as a model of testosterone deficiency. To assess NAFLD severity, NAFLD activity score (NAS) is adopted. Moreover, immunological change was analyzed by multicolor flow cytometry. Treatment with both testosterone and estrogen significantly decreased body weight to that of the sham mice/normal diet (ND). NAS and liver fibrosis in OCX-HFD mice were significantly deteriorated, and treatment with testosterone and estrogen improved same as sham-ND mice. HFD increased the ratio of both type 2 and 3 innate lymphoid cells (ILC2s and ILC3s) to CD45-positive cells in the liver. Treatment with testosterone alone decreased the ratio of ILC2 to CD45 but not the ILC3-to-CD45 ratio. Addition of estrogen to the treatment reduced the ratios of ILC2-to-CD45 and ILC3-to-CD45 to the same level observed in sham-HFD mice. Moreover, OCX-HFD mice had a decreased proportion of M2 macrophages compared with sham-ND mice. Treatment with testosterone alone did not restore the proportion of M2 macrophages; however, combination treatment with both estrogen and testosterone increased that to the same level as that in sham-HFD mice. Treatment with both testosterone and estrogen improves liver fibrosis and decreases ILC3 and increases M2 macrophage abundance in the liver.NEW & NOTEWORTHY The progression of nonalcoholic fatty liver disease (NAFLD) is associated with testosterone deficiency. NAFLD patients generally do not respond to treatment with testosterone alone. In animal studies, treatment with testosterone and estrogen reduced the ratios of ILC2:CD45 and ILC3:CD45 and increased M2 macrophages in liver. Our study suggests, based on our immunological data, that a combination of estrogen and testosterone may be clinically relevant for the treatment of NAFLD in patients with male menopause.