Ketogenic Diet Might Regulate Autophagy of BMSCs via mTOR Signaling Contributing to Osteoporosis in Mice.

Rapamycin (Rapa), an mTOR inhibitor, enhances autophagy to alleviate bone loss in senile and estrogen-deficient osteoporosis by restoring the biological properties of bone marrow stromal cells (BMSCs). However, Rapa against bone loss induced by ketogenic diet (KD) and the underlying mechanism remain elusive. This study aims to evaluate Rapa's influence on KD-induced osteoporosis and clarify its underlying mechanisms. Microarchitectures of bone, serum concentrations of bone alkaline phosphatase (ALP) and tartrate-resistant acid phosphatase (TRAP), along with osteocalcin (OCN), microtubule-associated protein light chain 3 (LC3), TRAP, and Sca-1 immunohistochemistry staining were evaluated in Sham, the OVX (ovariectomy), the OVX+ Rapa, the KD, and the KD+ Rapa groups. Expression of mTOR and the autophagy-related proteins, including LC3-II/LC3-I, Beclin1, ATG7, and P62 were also analyzed. Both KD and OVX induced significant bone loss in the cancellous bone of the distal femur and the L5 vertebra. Furthermore, KD effectively decreased serum ALP and increased TRAP in mice. In addition, KD and OVX downregulated OCN, ALP, RUNX2, the autophagy-related proteins, upregulated TRAP, PPARγ, p-mTOR, and P62, and decreased LC3 and Sca-1 expressions in mice. On the other hand, Rapa reduced significant bone loss in cancellous bone induced by KD or OVX. Additionally, Rapa effectively increased serum ALP and decreased TRAP in KD mice and decreased serum TRAP in OVX mice. It upregulated OCN, ALP, RUNX2, LC3-II/LC3-I, Beclin1, and ATG7 and downregulated TRAP, PPARγ, p-mTOR, and P62 in the KD and OVX cohorts. The findings demonstrated that autophagy downregulation might contribute to KD-induced osteoporosis, which may be part of the pathogenic mechanism responsible for bone loss during KD.