Effects of insulin therapy on porosity, non-enzymatic glycation and mechanical competence in the bone of rats with type 2 diabetes mellitus.

Type 2 diabetes mellitus increases skeletal fragility; however, the contributing mechanisms and optimal treatment strategies remain unclear. We studied the effects of diabetes and insulin therapy on non-enzymatic glycation (NEG), cortical porosity (Ct.Po) and biomechanics of the bone tissue in Zucker Diabetic Fatty (ZDF) rats. Eleven-week old ZDF diabetic and non-diabetic rats were given insulin to achieve glycaemic control or vehicle seven days per week over twelve weeks (insulin dose adapted individually 0.5 international units (IU) at week 1 to 13.0IU at week 12). The right femora were excised, micro-CT scanned, and tested in 3-point bending to measure biomechanics. NEG of the midshaft was determined from bulk fluorescence. Diabetes led to increased NEG (+50.1%, p=0.001) and Ct.Po (+22.9%, p=0.004), as well as to reduced mechanical competence (max. stress: -14.2%, p=0.041, toughness: -29.7%, p=0.016) in the bone tissue. NEG and Ct.Po both correlated positively to serum glucose (NEG: R(2)=0.41, p<0.001, Ct.Po: R(2)=0.34, p=0.003) and HbA1c (NEG: R(2)=0.42, p<0.001, Ct.Po: R(2)=0.28, p=0.008) levels, while NEG correlated negatively with bone biomechanics (elastic modulus: R(2)=0.21, p=0.023, yield stress: R(2)=0.17, p=0.047). Twelve weeks of insulin therapy had no significant effect on NEG or Ct.Po, and was unable to improve the mechanical competence of the bone tissue. A reduction of mechanical competence was observed in the bone tissue of the diabetic rats, which was explained in part by increased collagen NEG. Twelve weeks of insulin therapy did not alter NEG, Ct.Po or bone biomechanics. However, significant correlations between NEG and serum glucose and HbA1c were observed, both of which were reduced with insulin therapy. This suggests that a longer duration of insulin therapy may be required to reduce the NEG of the bone collagen and restore the mechanical competence of diabetic bone.

Effects of parathyroid hormone on cortical porosity, non-enzymatic glycation and bone tissue mechanics in rats with type 2 diabetes mellitus.

Type 2 diabetes mellitus increases skeletal fragility; however, the contributing mechanisms and the efficacy of bone-forming agents are unclear. We studied diabetes and parathyroid hormone (PTH) treatment effects on cortical porosity (Ct.Po), non-enzymatic glycation (NEG) and bone mechanics in Zucker diabetic fatty (ZDF) rats. Eleven-week old ZDF diabetic (DB) and non-diabetic (ND) rats were given 75µg/kg PTH (1-84) or vehicle 5days per week over 12weeks. The right femora and L4 vertebrae were excised, micro-CT scanned, and tested in 3-point bending and uniaxial compression, respectively. NEG of the samples was determined using fluorescence. Diabetes increased Ct.Po (vertebra (vert): +40.6%, femur (fem): +15.5% vs. ND group, p<0.05) but had no effect on NEG. PTH therapy reduced vertebral NEG in the ND animals only (-73% vs untreated group, p<0.05), and increased femoral NEG in the DB vs. ND groups (+63%, p<0.05). PTH therapy had no effect on Ct.Po. Diabetes negatively affected bone tissue mechanics where reductions in vertebral maximum strain (-22%) and toughness (-42%) were observed in the DB vs. ND group (p<0.05). PTH improved maximum strain in the vertebra of the ND animals (+21%, p<0.05) but did not have an effect in the DB group. PTH increased femoral maximum strain (+21%) and toughness (+28%) in ND and decreased femoral maximum stress (-13%) and toughness (-27%) in the DB animals (treated vs. untreated, p<0.05). Ct.Po correlated negatively with maximum stress (fem: R=-0.35, p<0.05, vert: R=-0.57, p<0.01), maximum strain (fem: R=-0.35, p<0.05, vert: R=-0.43, p<0.05) and toughness (fem: R=-0.34, p<0.05, vert: R=-0.55, p<0.01), and NEG correlated negatively with toughness at the femur (R=-0.34, p<0.05) and maximum strain at the vertebra (R=-0.49, p<0.05). Diabetes increased cortical porosity and reduced bone mechanics, which were not improved with PTH treatment. PTH therapy alone may worsen diabetic bone mechanics through formation of new bone with high AGEs cross-linking. Optimal treatment regimens must address both improvements of bone mass and glycemic control in order to successfully reduce diabetic bone fragility. This article is part of a Special Issue entitled "Bone and diabetes".

Bone defect regeneration and cortical bone parameters of type 2 diabetic rats are improved by insulin therapy.

Zucker Diabetic Fatty (ZDF) rats represent an established model of type 2 diabetes mellitus (T2DM) and display several features of human diabetic bone disease, including impaired osteoblast function, decreased bone strength, and delayed bone healing. Here, we determined whether glycemic control by insulin treatment prevents skeletal complications associated with diabetes. Subcritical femur defects were created in diabetic (fa/fa) and non-diabetic (+/+) ZDF rats. Diabetic rats were treated once daily with long-lasting insulin glargin for 12weeks for glycemic control. Insulin treatment successfully maintained serum levels of glycated hemoglobin, while untreated diabetic rats showed a 2-fold increase. Trabecular and cortical bone mass measured by µCT were decreased in diabetic rats. Insulin treatment increased bone mass of the cortical, but not of the trabecular bone compartment. Dynamic histomorphometry revealed a lower bone formation rate at the trabecular and periosteal cortical bone in diabetic animals and decreased serum procollagen type 1 N-terminal propeptide (P1NP, -49%) levels. Insulin treatment partially improved these parameters. In T2DM, serum levels of tartrate-resistant acid phosphatase (TRAP, +32%) and C-terminal telopeptide (CTX, +49%) were increased. Insulin treatment further elevated TRAP levels, but did not affect CTX levels. While diabetes impaired bone defect healing, glycemic control with insulin fully reversed these negative effects. In conclusion, insulin treatment reversed the adverse effects of T2DM on bone defect regeneration in rats mainly by improving osteoblast function and bone formation. This article is part of a Special Issue entitled Bone and diabetes.