The current studies present evidence for an importance of efficient hepatic insulin clearance in the maintenance of bone homeostasis. In the L-SACC1 murine model we have shown that impaired insulin clearance in the liver, which results in high levels of circulating insulin and insulin resistance in peripheral tissues, leads to decreased bone turnover. It affects bone remodeling process by decreasing both, bone formation and bone resorption. It also changes myelopoietic cell commitment toward the osteoclast and B-cell lineages. We have demonstrated in both, in vivo
and in vitro
systems that high levels of insulin affect recruitment and differentiation of osteoclasts by impairing the RANKL signaling pathway. In osteoclasts, expression of the RANK receptor is under the control of the c-fos transcription factor (1
), and insulin modulates the expression of both genes. Using PBMC derived from normoinsulinemic animals, we showed that exogenously added insulin affects osteoclast differentiation and the expression of RANK and c-fos in a manner identical to L-SACC1 animals. These findings provide evidence that insulin negatively regulates the phenotype of cells of the osteoclast lineage, and suggest that systemic hyperinsulinemia has a permanent effect on osteoclast progenitors.
The effect of insulin on cells of mesenchymal lineage is less evident. Differentiation potential of MSCs derived from L-SACC1 animals toward osteoblast lineage and the expression of osteoblast gene markers were not different from MSCs derived from WT animals, with exception to IGF-1 which expression was decreased. In contrast to studies of other’s (4
), insulin neither increase alkaline phosphatase activity nor cell proliferation when added to the PBMC culture. However, serum parameters of bone formation in L-SACC1 were significantly different from that of WT animals. These data suggest that systemic changes, which result from CEACAM1 mutation in the liver, affect osteoblast function in the bone in an indirect manner, which does not involve intrinsic changes in MSC differentiation potential. In contrast to cells of hematopoietic lineage, in vitro
effect of insulin on MSC osteoblast phenotype does not recapitulate attenuated bone formation in L-SACC1 animals, suggesting that CEACAM1 mutation in the liver affects marrow mesenchymal cells by different mechanism than cells of hematopoietic lineage.
We have reported previously that PPARγ2, an adipocyte-specific transcription factor, suppresses the expression of IGF-1 in bone, especially its longer isoform which contains exon 6 (13
). Here, we showed that L-SACC1 mice are characterized by increased expression of PPARγ2 in marrow MSC. Thus, PPARγ2 may be responsible for decreased IGF-1 expression. Interestingly, although we did not observe an increase in CFU-AD formation in ex vivo
conditions, L-SACC1 animals possess higher number of adipocytes in the bone marrow. The discrepancy between ex vivo
and in vivo
observation indicate that in in vivo
conditions, an additional factor, perhaps insulin, which possess proadipocytic activity (27
), enhanced adipocytic diiferentiation of MSC. Taken together, these results suggest that prolonged exposure to high levels of insulin induces the adipocyte phenotype in MSC and decreases the expression of IGF-1, a positive regulators of osteoblast function (19
), which may contribute to the decreased bone formation rate observed in L-SACC1 animals.
Several studies point toward a decreased bone turnover rate in T2DM (2
). An analysis of serum bone turnover parameters in T2DM patients and in diabetic Pima Indians showed reduced bone formation, as assessed by low levels of alkaline phosphatase, osteocalcin and IGF-1 (2
). Another study based on longitudinal observations of bone status showed attenuation of the rate of bone loss with aging and relatively higher bone mass in T2DM patients (9
). The higher bone mass in L-SACC1 animals is entirely consistent with low bone turnover, which results from attenuated bone resorption and decreased bone formation.
A paradox between normal or higher BMD and increased fracture risk in T2DM suggests altered quality in diabetic bone. In addition to hyperinsulinemia leading to low bone turnover, hyperglycemia may account for changes in bone biomaterial quality by modification of collagen fibers (18
). Highly reactive glucose metabolites (AGEs), of which circulating levels are increased in diabetic hyperglycemia, are implicated in forming cross-links between collagen fibers, which affect bone biomechanical properties by increasing its stiffness and fragility (22
Using the L-SACC1 model of hyperinsulinemia caused by liver-specific impairment in insulin clearance, we have demonstrated that high insulin levels affect bone resorption and impair bone formation, leading to a decreased bone turnover rate. Because altered insulin action in L-SACC1 mice is primarily hepatic, our studies suggest that insulin metabolism in the liver, which regulates overall insulin action in peripheral tissues (muscle and adipose tissue) (16
), regulates also the maintenance of bone mass. The similarity of the phenotype of L-SACC1 mice with hyperinsulinemia in humans emphasizes the relevance of the current studies in advancing our understanding of the altered bone homeostasis in diabetes.