Type 2 diabetic mice demonstrate slender long bones with increased fragility secondary to increased osteoclastogenesis
Introduction
Diabetes is an emerging epidemic that has consequences for musculo-skeletal growth and development, and bone fracture risk [1], [2], [3], [4], [5], [6], [7], [8]. Yet, Type 2 diabetics (non-insulin-dependent, T2D) often demonstrate increased bone mineral density (BMD) [9], [10], [11], [12], [13]. Furthermore, the use of anti-diabetic oral thiazolidinediones (PPARγ agonists) in T2D has recently been shown to increase fracture risk in the absence of a reduction in BMD [14]. These apparent paradoxes suggest that the increased bone fragility in T2D may not be discerned from bone mass, but may depend on aspects of bone structure or tissue quality that are not revealed from BMD. A number of factors have been postulated to be etiologically associated with poor bone accrual in diabetes, including an insulin-related direct reduction in bone formation and reduced force generating capacity of muscle.
Studies of bone turnover in animal models are most accurately assessed by histomorphometry. Several models have demonstrated diabetes-induced alterations in bone and could aid in elucidating the cellular and biomechanical mechanisms that may lead to increased risk of fragility fractures [15], [16], [17], [18], [19], [20]. Models for Type 1 diabetes (T1D) are most prevalent and have demonstrated decreased breaking strength of femoral shafts when compared to controls [21], [22]. However, many of these are drug-induced models confounded by side-effects that can result in significant health issues. For example, the pancreatic β-cell toxins streptozotocin or alloxan have been injected into both rats and mice to induce an uncontrolled diabetic state [23], [24]. Growth trajectories are immediately altered such that diabetic animals fail to grow (i.e., the rates of long bone growth and body mass gains are significantly stunted) [25], [26]. However, stunted longitudinal growth is not normally associated with diabetes. Most recently, using a model of streptozotocin-induced insulinopenia, Hamada et al. confirmed this affect on growth as well as a reduction in trabecular bone volume with reduction in both bone formation and bone resorption in young animals, changes that were reversed by insulin replacement therapy [25]. The main conclusion of that study was that in T1D, insulinopenia is the primary etiology for osteopenia, though secondary mechanisms, such as oxidative stress, may also play a role [25]. The cellular mechanisms behind these differences require further examination.
Models of T2D have thus far been less biologically insightful because many induce obesity (another state of insulin resistance) or are limited to rats [27], [28], [29], [30]. There is limited evidence for increased fracture risk due to greater levels of obesity and obesity may even protect a subject from fracture, despite being associated with lower physical activity [3]. Obesity increases the load-bearing forces during locomotion and, absent a bone formation defect, elicits structural compensation, i.e., more weight requires a larger frame [31]. Obesity may also enhance endogenous production of estrogen, a hormone important for bone cell regulation in both sexes [32]. Non-obese T2D occurs spontaneously in rats such as the Torii, which was outbred from Sprague–Dawley in 1997, and also exhibits stunted longitudinal bone growth; however, the disease has unknown etiology and is associated with insulinopenia [28].
We have generated a non-obese transgenic mouse model of T2D (MKR mice) by blocking both the insulin and IGF-signaling pathways specifically in skeletal muscle. Blocking of both receptors in muscle abrogates downstream signaling, particularly the AKT pathway, which regulates glucose uptake into muscle. MKR mice are born with naturally occurring hyperinsulinemia and insulin resistance in muscle. Early in life (∼ 2 weeks) the MKR mice develop dyslipidemia, and the mice eventually develop frank diabetes with hyperglycemia (∼ 7–8 weeks) [33], [34]. This scenario of disease development is similar to the human disease, where insulin resistance and hyperinsulinemia, with dyslipidemia eventually progress to diabetes. The MKR mouse model is useful since no exogenous agents are required to induce disease, thus removing potential confounding aspects. In addition, MKR mice are non-obese, excluding another confounding variable. In this study we employed the MKR mouse model to examine the effect of progression of T2D on skeletal integrity. Skeletal characterization was performed at the pre-diabetic (3 weeks of age), early diabetic (8 weeks) and established diabetic stages (16 weeks). Here we demonstrate that early onset of insulin resistance does not affect longitudinal growth but alters transverse expansion of skeletal structures, which worsens with the progression to the adult diabetic state.
Section snippets
Materials and methods
The generation of the MKR Type 2 diabetic mouse has been described elsewhere [33]. MKR male mice, on a FVB/N background, were bred to homozygosity, and were compared to wild-type FVB/N controls (WT). MKR mice demonstrate severe insulin resistance starting at birth, hyperinsulinemia and hyperlipidemia at 3–4 weeks of age but with normoglycemia [33]. At 6–8 weeks of age MKR mice develop diabetes with blood glucose levels of 250–400 mg/dl versus 130–160 mg/dl for WT. The mice were kept on a 12-h
Diabetes phenotype of MKR mice
Male MKR and FVB/N control mice (WT) were weaned at 3 weeks of age and following genotyping as previously described were monitored weekly for blood glucose levels [33]. All mice remained normoglycemic from 3 weeks to 5 weeks of age. By 7 weeks of age, fed blood glucose levels in MKR mice rose to diabetic levels (between 250 and 400 mg/dl), whereas WT remained normoglycemia (between 130 and 160 mg/dl). The hyperglycemia of the MKR mice remained at 16 weeks of age. Hyperlipidemia and
Discussion
Fragility fractures are associated with both Types 1 and 2 diabetes despite the fact that bone mass (BMD) losses do not occur in Type 2 diabetes (T2D) [9], [10], [11], [12], [13]. The reductions in BMD with Type 1 diabetes (T1D) occur early in the disease process and are associated with increased bone fragility and fractures later in life. While the reductions in BMD in T1D have been well established, the cellular cause(s) have not been defined completely, with both increased bone turnover and
Conflict of interest statement
The authors have nothing to disclose.
Acknowledgments
We are indebted to Ken Inagaki, Damien Laudier, Valerie Williams, David Berman, and Bin Hu for their help on this project. Financial support received from funding agencies in the United States including the Mount Sinai School of Medicine (Steckler Fund Grant to DL), NIAMS (NIH Grants AR41210 to MBS, AR44927 to KJJ and AR054919 and AR055141 to SY), National Space Biomedical Research Institute (NASA Grant NCC 9-58 to MBS) and the Charles H. Revson Foundation (Fellowship to JCF). The statements
References (62)
- et al.
Skeletal involution by age-associated oxidative stress and its acceleration by loss of sex steroids
J. Biol. Chem.
(2007) - et al.
Transgenic mice with osteoblast-targeted insulin-like growth factor-I show increased bone remodeling
Bone
(2006) - et al.
The role of insulin in chondrogenesis
Mol. Cell. Endocrinol.
(2006) - et al.
Histomorphometric analysis of diabetic osteopenia in streptozotocin-induced diabetic mice: a possible role of oxidative stress
Bone
(2007) - et al.
The biomechanical integrity of bone in experimental diabetes
Diabetes Res. Clin. Pract.
(2001) - et al.
Osteoporosis in the Cohen diabetic rat: correlation between histomorphometric changes in bone and microangiopathy
Lab. Invest.
(2002) - et al.
Bone formation in spontaneously diabetic Torii-newly established model of non-obese Type 2 diabetes rats
Bone
(2008) - et al.
Loading induces site-specific increases in mineral content assessed by microcomputed tomography of the mouse tibia
Bone
(2005) - et al.
TDAG51 mediates the effects of insulin-like growth factor I (IGF-I) on cell survival
J. Biol. Chem.
(2004) - et al.
Stress fracture in military recruits: gender differences in muscle and bone susceptibility factors
Bone
(2000)
Glucose-induced inhibition of in vitro bone mineralization
Bone
Oxidative stress inhibits osteoblastic differentiation of bone cells by ERK and NF-kappaB
Biochem. Biophys. Res. Commun.
Oxidative stress modulates osteoblastic differentiation of vascular and bone cells
Free Radic. Biol. Med.
Osteoclastic function is accelerated in male patients with Type 2 diabetes mellitus: the preventive role of osteoclastogenesis inhibitory factor/osteoprotegerin (OCIF/OPG) on the decrease of bone mineral density
Diabetes Res. Clin. Pract.
Osteoporosis in men
Endocrinol. Metab. Clin. North. Am.
Projection of diabetes burden through 2050: impact of changing demography and disease prevalence in the U.S
Diabetes Care
Diabetes mellitus and the incidence of hip fracture: results from the Nord-Trondelag Health Survey
Diabetologia
Decreased lumbar spine bone mass and low bone turnover in children and adolescents with insulin dependent diabetes mellitus followed longitudinally
J. Pediatr. Endocrinol. Metab.
Osteoporosis in patients with diabetes mellitus
J. Bone Miner. Res.
Osteoporosis and diabetes mellitus
Rev. Endocr. Metab. Disord.
Osteopenia in insulin-dependent diabetes mellitus; prevalence and aspects of pathophysiology
J. Endocrinol. Invest.
Fracture risk in Type 2 diabetes: update of a population-based study
J. Bone Miner. Res.
Type 1 and Type 2 diabetes and incident hip fractures in postmenopausal women
Diabetes Care
Proximal femur density in Type 1 and 2 diabetic patients
Diabet Metab.
Bone mineral density and fracture risk in Type-2 diabetes mellitus: the Rotterdam Study
Osteoporos. Int.
The risk of hip fractures in older individuals with diabetes: a population-based study
Diabetes Care
Diabetes and bone loss at the hip in older black and white adults
J. Bone Miner. Res.
Increased mortality in patients with a hip fracture-effect of pre-morbid conditions and post-fracture complications
Osteoporos. Int.
Glycemic durability of rosiglitazone, metformin, or glyburide monotherapy
N. Engl. J. Med.
Generation of a new congenic mouse strain to test the relationships among serum insulin-like growth factor I, bone mineral density, and skeletal morphology in vivo
J. Bone Miner. Res.
Intensive insulin therapy and bone mineral density in Type 1 diabetes mellitus: a prospective study
Osteoporos. Int.
Cited by (68)
Induction and rescue of skeletal fragility in a high-fat diet mouse model of type 2 diabetes: An in vivo and in vitro approach
2022, BoneCitation Excerpt :Bones from the HFD and LFD group were harvested for characterization. In line with previous studies [59–65], our bone microarchitecture findings showed that diabetes caused an increase in the degree of mineralization, as well as alteration of its microstructure. Furthermore, similar to ours, a previous study [56] also reports trabecular structure to be affected.
Characteristics of bone strength and metabolism in type 2 diabetic model Tsumura, Suzuki, Obese Diabetes mice
2018, Bone ReportsCitation Excerpt :A previous study on osteoclasts derived from db/db T2DM mice reported that the differentiation of osteoclasts was enhanced by hyperglycemia (Catalfamo et al., 2013). However, these results were not consistent with those of previous studies on bone metabolism in T2DM animal models although this may be due to the fact that these studies on bone metabolism were conducted in mice with low BMD (Devlin et al., 2014; Fu et al., 2015; Fujii et al., 2008; Hamann et al., 2011; Kawashima et al., 2009; Omi et al., 1998 Turner et al., 2013; Zhang et al., 2009;). We suggest that the increased functional ability of osteoclasts in pre-diabetic conditions in TSOD mice may involve hyperinsulinemia and that the function of osteoclasts is maintained by chronic hyperglycemia in established diabetic conditions.
LOX-related collagen crosslink changes act as an initiator of bone fragility in a ZDF rats model
2018, Biochemical and Biophysical Research Communications
- 1
Both of these authors contributed equally to this work.