The Herbal Medicine KBH-1 Inhibits Fat Accumulation in 3T3-L1 Adipocytes and Reduces High Fat Diet-Induced Obesity through Regulation of the AMPK Pathway

The aim of this study was to investigate whether a novel formulation of an herbal extract, KBH-1, has an inhibitory effect on obesity. To determine its anti-obesity effects and its underlying mechanism, we performed anti-obesity-related experiments in vitro and in vivo. 3T3-L1 preadipocytes were analyzed for lipid accumulation as well as the protein and gene expression of molecular targets involved in fatty acid synthesis. To determine whether KBH-1 oral administration results in a reduction in high-fat diet (HFD)-induced obesity, we examined five groups (n = 9) of C57BL/6 mice as follows: 10% kcal fat diet-fed mice (ND), 60% kcal fat diet-fed mice (HFD), HFD-fed mice treated with orlistat (tetrahydrolipstatin, marketed under the trade name Xenical), HFD-fed mice treated with 150 mg/kg KBH-1 (KBH-1 150) and HFD-fed mice treated with 300 mg/kg KBH-1 (KBH-1 300). During adipogenesis of 3T3-L1 cells in vitro, KBH-1 significantly reduced lipid accumulation and down-regulated the expression of master adipogenic transcription factors, including CCAAT/enhancer binding protein (C/EBP) β, C/EBP α and peroxisome proliferation-activity receptor (PPAR) γ, which led to the suppression of the expression of several adipocyte-specific genes and proteins. KBH-1 also markedly phosphorylated the adenosine monophosphate-activated protein kinase (AMPK) and acetyl-CoA carboxylase (ACC). In addition, KBH-1-induced the inhibition effect on lipid accumulation and AMPK-mediated signal activation were decreased by blocking AMPK phosphorylation using AMPK siRNA. Furthermore, daily oral administration of KBH-1 resulted in dose-dependent decreases in body weight, fat pad mass and fat tissue size without systemic toxicity. These results suggest that KBH-1 inhibits lipid accumulation by down-regulating the major transcription factors of the adipogenesis pathway by regulating the AMPK pathway in 3T3-L1 adipocytes and in mice with HFD-induced obesity. These results implicate KBH-1, a safe herbal extract, as a potential anti-obesity therapeutic agent.

本研究旨在探究新型草本提取物配方KBH-1是否对肥胖具有抑制作用。为明确其抗肥胖功效与潜在作用机制，我们开展了体外与体内抗肥胖相关实验。我们对3T3-L1前脂肪细胞（3T3-L1 preadipocytes）的脂质积累情况，以及参与脂肪酸合成的分子靶点的蛋白与基因表达水平进行了分析。为验证KBH-1口服给药是否可减轻高脂饮食（high-fat diet, HFD）诱导的肥胖，我们将C57BL/6小鼠分为5组（每组n=9），具体分组如下：10%热量脂肪饮食喂养小鼠（正常饮食组，ND）、60%热量脂肪饮食喂养小鼠（高脂饮食组，HFD）、给予奥利司他（tetrahydrolipstatin，商品名赛尼可（Xenical））的高脂饮食喂养小鼠、给予150 mg/kg KBH-1的高脂饮食喂养小鼠（KBH-1 150组），以及给予300 mg/kg KBH-1的高脂饮食喂养小鼠（KBH-1 300组）。在体外3T3-L1细胞的成脂分化过程中，KBH-1可显著减少脂质积累，并下调成脂关键转录因子的表达，包括CCAAT/增强子结合蛋白β（CCAAT/enhancer binding protein β, C/EBPβ）、C/EBPα以及过氧化物酶体增殖活性受体γ（peroxisome proliferation-activity receptor γ, PPARγ），进而抑制多种脂肪细胞特异性基因与蛋白的表达。KBH-1还可显著磷酸化腺苷单磷酸活化蛋白激酶（adenosine monophosphate-activated protein kinase, AMPK）与乙酰辅酶A羧化酶（acetyl-CoA carboxylase, ACC）。此外，通过AMPK小干扰RNA（AMPK siRNA）阻断AMPK磷酸化后，KBH-1诱导的脂质积累抑制作用与AMPK介导的信号激活均被削弱。进一步研究发现，每日口服KBH-1可呈剂量依赖性降低小鼠体重、脂肪垫重量与脂肪组织体积，且未引发全身性毒性反应。上述结果表明，在3T3-L1脂肪细胞与高脂饮食诱导肥胖的小鼠模型中，KBH-1可通过调控AMPK通路，下调成脂通路的关键转录因子，从而抑制脂质积累。本研究结果提示，安全的草本提取物KBH-1有望成为潜在的抗肥胖治疗药物。