Non-alcoholic fatty liver disease (NAFLD) is associated with abnormal bone metabolism, potentially mediated by elevated levels of proinflammatory cytokines such as tumor necrosis factor alpha (TNF-ɑ) and interleukin 6 (IL-6). This study aims to investigate the direct regulatory effects of liver tissues on osteoblast and osteoclast functions in vitro, focusing on the liver-bone axis in NAFLD. Twelve-week-old C57BL/6 mice were fed either a control diet or a high-fat diet (HFD) for 12 weeks. Bone structural parameters were assessed using microCT. Primary hepatocyte cultures were established from control and HFD-fed C57BL/6 mice, as well as IL-6-/- and TNF-α-/- mice. The supernatants from these hepatocyte cultures were used to induce differentiation in bone marrow cell-derived osteoblasts and osteoclasts in vitro. Results showed that mice on a HFD exhibited increased lipid infiltration in liver and bone marrow tissues, alongside reduced bone mass. Moreover, the supernatants from hepatocyte cultures from mice on a HFD displayed elevated TNF-α and IL-6 levels. These supernatants, particularly those derived from HFD-fed and IL-6-/- mice, significantly enhanced osteoclast differentiation in vitro. In contrast, supernatants from TNF-α-/- mice did not significantly affect osteoblast or osteoclast differentiation in vitro. In conclusions, this current study suggested that fatty liver tissues may negatively impact bone metabolism. Additionally, knockout of TNF-α and IL-6 genes revealed distinct influence on osteoblast and osteoclast functions, highlighting the complex interplay between live pathology and bone health.© 2024 The Author(s).