目的 本研究通过动态监测生酮饮食（ketogenic diet，KD）干预C57/BL6J雌性小鼠骨微结构参数演变规律，首次系统评估KD疗法的时间依赖性骨代谢毒性效应，揭示KD干预时长与骨代谢紊乱的剂量效应关系，精准定位骨量丢失关键时间窗，为临床KD疗程安全周期划定提供重要实验依据。 方法 将45只C57/BL6J雌性小鼠分为9组：正常饮食4周(Con+1)、8周(Con+2)、12周（Con+3），KD4周(KD+1)、8周(KD+2)、12周（KD+3），双侧卵巢切除4周(OVX+1)、8周(OVX+2)、12周（OVX+3）组；并监测体重、血酮和血糖；进行Micro-CT扫描、HE染色观察各个时间点股骨骨微结构及形态变化。 结果 Con+1、KD+1及OVX+1骨微结构无显著差异。KD+2的TMD比Con+2明显减少。KD+3的TMD、BV/TV、Tb.N、Conn.D、Ct.Th、Barea及BA/TA比Con+3明显降低，Tb.Sp显著升高。KD组BV/TV在三个时间点分别为5.8%，5.0%，3.6%。OVX在三个时间点BV/TV分别为4.4%，2.9%，3.0%。HE结果进一步验证Micro-CT结果。 结论 4周以内短期的KD疗法对小鼠骨微结构影响较小，可作为较安全的干预周期；KD诱导小鼠OP模型需要12周，可作为KD诱导小鼠骨质疏松病理机制和防治研究的重点阶段，KD具有 “时间依赖性骨损害”，该发现对推动KD疗法的精准安全应用具有重要转化价值。
Abstract
Objective To investigate the changes in bone microstructure of female C57BL/6J mice after 4-, 8-, and 12-week ketogenic diet (KD) interventions, aiming to determine the optimal time window for establishing a KD-induced osteoporosis (OP) model. The findings will provide theoretical insights for preventing the adverse effects of KD therapy on bone microstructure, thereby promoting the clinical translation and broader application of KD therapeutic strategies. Methods Experimental animals were divided into groups with normal diet for 4 weeks (Con+1), 8 weeks (Con+2), and 12 weeks (Con+3); KD for 4 weeks (KD+1), 8 weeks (KD+2), and 12 weeks (KD+3); and bilateral ovariectomy for 4 weeks (OVX+1), 8 weeks (OVX+2), and 12 weeks (OVX+3). Body weight, blood ketones, and blood glucose were monitored. Micro-CT scanning and HE staining were performed to observe changes in femoral bone microstructure and morphology at each time point. Results There were no significant differences in bone microstructure between Con+1, KD+1, and OVX+1. The trabecular mineral density (TMD) of KD+2 was significantly reduced compared to Con+2. The TMD, bone volume fraction (BV/TV), trabecular number (Tb.N), connectivity density (Conn.D), cortical thickness (Ct.Th), bone area (Barea), and bone area/total area (BA/TA) of KD+3 were significantly lower than those of Con+3, while trabecular separation (Tb.Sp) was significantly increased. The BV/TV of the KD group at the three time points was 5.8%, 5.0%, and 3.6%, respectively. The BV/TV of the OVX group at the three time points was 4.4%, 2.9%, and 3.0%, respectively. The HE results confirmed the Micro-CT findings. Conclusions Short-term ketogenic diet (KD) interventions within 4 weeks exhibited minimal impact on bone microstructure in mice, suggesting a relatively safe therapeutic window. In contrast, a 12-week KD regimen successfully induced an osteoporosis (OP) model in mice, serving as a critical phase for investigating the pathological mechanisms of KD-induced OP and exploring preventive/therapeutic strategies. KD has "time-dependent bone damage", and this discovery has significant translational value for promoting the precise and safe application of KD therapy.
关键词
生酮饮食 /
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骨质疏松 /
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模型 /
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骨微结构
Key words
Ketogenic diet /
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Osteoporosis /
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Model /
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Bone microarchitecture
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