Glucose-dependent insulinotropic polypeptide (GIP) directly affects collagen fibril diameter and collagen cross-linking in osteoblast cultures.

Glucose-dependent insulinotropic polypeptide (GIP) is absolutely crucial in order to obtain optimal bone strength and collagen quality. However, as the GIPR is expressed in several tissues other than bone, it is difficult to ascertain whether the observed modifications of collagen maturity, reported in animal studies, were due to direct effects on osteoblasts or indirect through regulation of signals originating from other tissues. The aims of the present study were to investigate whether GIP can directly affect collagen biosynthesis and processing in osteoblast cultures and to decipher which molecular pathways were necessary for such effects. MC3T3-E1 cells were cultured in the presence of GIP ranged between 10 and 100pM. Collagen fibril diameter was investigated by electron microscopy whilst collagen maturity was determined by Fourier transform infra-red microspectroscopy (FTIRM). GIP treatment resulted in dose-dependent increases in lysyl oxidase activity and collagen maturity. Furthermore, GIP treatment shifted the collagen fiber diameter towards lower value but did not significantly affect collagen heterogeneity. GIP acted directly on osteoblasts by activating the adenylyl cyclase-cAMP pathway. This study provides evidences that GIP acts directly on osteoblasts and is capable of improving collagen maturity and fibril diameter

Optimal bone mechanical and material properties require a functional glucagon-like peptide-1 receptor.

Bone is permanently remodeled by a complex network of local, hormonal, and neuronal factors that affect osteoclast and osteoblast biology. Among these factors, a role for gastrointestinal hormones has been proposed based on the evidence that bone resorption dramatically falls after a meal. Glucagon-like peptide-1 (GLP1) is one of these gut hormones, and despite several reports suggesting an anabolic effect of GLP1, or its stable analogs, on bone mass, little is known about the effects of GLP1/GLP1 receptor on bone strength. In this study, we investigated by three-point bending, quantitative X-ray microradiography, microcomputed tomography, qBEI, and FTIRI bone strength and bone quality in male Glp1r knockout (Glp1r KO) mice when compared with control WT animals. Animals with a deletion of Glp1r presented with a significant reduction in ultimate load, yield load, stiffness, and total absorbed and post-yield energies when compared with WT animals. Furthermore, cortical thickness and bone outer diameter were significantly decreased in deficient animals. The mineral quantity and quality were not significantly different between Glp1r KO and WT animals. On the other hand, the maturity of the collagen matrix was significantly reduced in deficient animals and associated with lowered material properties. Taken together, these data support a positive effect of GLP1R on bone strength and quality

Efficacy of targeting bone-specific GIP receptor in ovariectomy-induced bone loss

Glucose-dependent insulinotropic polypeptide (GIP) has been recognized in the last decade as an important contributor of bone remodeling and is necessary for optimal bone quality. However, GIP receptors are expressed in several tissues in the body and little is known about the direct versus indirect effects of GIP on bone remodeling and quality. The aims of the present study were to validate two new GIP analogues, called [D-Ala2]-GIP-Tag and [D-Ala2]-GIP1-30, that specifically target either bone or whole body GIP receptors, respectively; and to ascertain the beneficial effects of GIP therapy on bone in a mouse model of ovariectomy-induced bone loss. Both GIP analogues exhibited similar binding capacities at the GIP receptor and intracellular responses as full-length GIP1-42. Furthermore, only [D-Ala2]-GIP-Tag, but not [D-Ala2]-GIP1-30, was undoubtedly found exclusively in the bone matrix and released at acidic pH. In ovariectomized animals, [D-Ala2]-GIP1-30 but not [D-Ala2]-GIP-Tag ameliorated bone stiffness at the same magnitude than alendronate treatment. Only [D-Ala2]-GIP1-30 treatment led to significant ameliorations in cortical microarchitecture. Although alendronate treatment increased the hardness of the bone matrix and the type B carbonate substitution in the hydroxyapatite crystals, none of the GIP analogues modified bone matrix composition. Interestingly, in ovariectomy-induced bone loss, [D-Ala²]-GIP-Tag failed to alter bone strength, microarchitecture and bone matrix composition. Overall, this study shows that the use of a GIP analogue that target whole body GIP receptors might be useful to improve bone strength in ovariectomized animals

Glucose-dependent insulinotropic polypeptide (GIP) receptor deletion leads to reduced bone strength and quality.

Bone is permanently remodeled by a complex network of local, hormonal and neuronal factors that affect osteoclast and osteoblast biology. In this context, a role for gastro-intestinal hormones has been proposed based on evidence that bone resorption dramatically falls after a meal. Glucose-dependent insulinotropic polypeptide (GIP) is one of the candidate hormones as its receptor, glucose-dependent insulinotropic polypeptide receptor (GIPR), is expressed in bone. In the present study we investigated bone strength and quality by three-point bending, quantitative x-ray microradiography, microCT, qBEI and FTIR in a GIPR knockout (GIPR KO) mouse model and compared with control wild-type (WT) animals. Animals with a deletion of the GIPR presented with a significant reduction in ultimate load (--11%), stiffness (-16%), total absorbed (-28%) and post-yield energies (-27%) as compared with WT animals. Furthermore, despite no change in bone outer diameter, the bone marrow diameter was significantly increased and as a result cortical thickness was significantly decreased by 20% in GIPR deficient animals. Bone resorption at the endosteal surface was significantly increased whilst bone formation was unchanged in GIPR deficient animals. Deficient animals also presented with a pronounced reduction in the degree of mineralization of bone matrix. Furthermore, the amount of mature cross-links of collagen matrix was significantly reduced in GIPR deficient animals and was associated with lowered intrinsic material properties. Taken together, these data support a positive effect of the GIPR on bone strength and quality

Alteration of the bone tissue material properties in type 1 diabetes mellitus: A Fourier transform infrared microspectroscopy study

Type 1 diabetes mellitus (T1DM) is a severe disorder characterized by hyperglycemia and hypoinsulinemia. A higher occurrence of bone fractures has been reported in T1DM, and although bone mineral density is reduced in this disorder, it is also thought that bone quality may be altered in this chronic pathology. Vibrational microscopies such as Fourier transform infrared microspectroscopy (FTIRM) represent an interesting approach to study bone quality as they allow investigation of the collagen and mineral compartment of the extracellular matrix in a specific bone location. However, as spectral feature arising from the mineral may overlap with those of the organic component, the demineralization of bone sections should be performed for a full investigation of the organic matrix. The aims of the present study were to (i) develop a new approach, based on the demineralization of thin bone tissue section to allow a better characterization of the bone organic component by FTIRM, (ii) to validate collagen glycation and collagen integrity in bone tissue and (iii) to better understand what alterations of tissue material properties in newly forming bone occur in T1DM. The streptozotocin-injected mouse (150mg/kg body weight, injected at 8weeks old) was used as T1DM model. Animals were randomly allocated to control (n=8) or diabetic (n=10) groups and were sacrificed 4weeks post-STZ injection. Bones were collected at necropsy, embedded in polymethylmethacrylate and sectioned prior to examination by FTIRM. FTIRM collagen parameters were collagen maturity (area ratio between 1660 and 1690cm(-1) subbands), collagen glycation (area ratio between the 1032cm(-1) subband and amide I) and collagen integrity (area ratio between the 1338cm(-1) subband and amide II). No significant differences in the mineral compartment of the bone matrix could be observed between controls and STZ-injected animals. On the other hand, as compared with controls, STZ-injected animals presented with significant higher value for collagen maturity (17%, p=0.0048) and collagen glycation (99%, p=0.0121), while collagen integrity was significantly lower by 170% (p=0.0121). This study demonstrated the profound effect of early T1DM on the organic compartment of the bone matrix in newly forming bone. Further studies in humans are required to ascertain whether T1DM also lead to similar effect on the quality of the bone matrix

Sitagliptin alters bone composition in high fat fed mice

Type 2 diabetes mellitus is recognized as a significant risk factor for fragility of bone. Among the newer anti-diabetic agents, dipeptidyl peptidase-4 inhibitors (DPP4i) have been reported to decrease the occurrence of bone fractures although the reason is unclear. The main aim of this study was to evaluate the impact of sitagliptin treatment on tissue bone strength and compositional parameters in the high-fat-fed mouse model. Male NIH swiss mice were allowed free access to high-fat diet for 150 days to induce chronic hyperglycemia and insulin resistance. Sitagliptin was administered once daily for 3 weeks. High-fat-fed mice administered with saline were used as controls. Bone strength was assessed at the organ and tissue level by three-point bending and nanoindentation, respectively. Bone microarchitecture was investigated by microcomputed tomography and bone composition was evaluated by Fourier transform infrared imaging and quantitative backscattered electron imaging. Administration of sitagliptin increased non-fasting insulin, improved glucose tolerance and increased insulin sensitivity. This was associated with clear ameliorations in bone strength at the organ and tissue level. No changes in trabecular or cortical microarchitectures were observed. On the other hand, higher values of Ca, Ca, collagen maturity, mineral/matrix ratio, mineral maturity and crystal size index were evidenced after sitagliptin treatment. Correlation analysis significantly linked the modifications of bone strength to changes in bone compositional parameters. These results bring new light on the mode of action of sitagliptin on bone physiology and demonstrate a benefit of DPP4i

Beneficial effects of a N-terminally modified GIP agonist on tissue-level bone material properties.

Bone remodeling is under complex regulation from nervous, hormonal and local signals, including gut hormones. Among the gut hormones, a role for the glucose-dependent insulinotropic polypeptide (GIP) has been suggested. However, the rapid degradation of GIP in the bloodstream by the ubiquitous enzyme dipeptidyl peptidase-4 (DPP-4) precludes therapeutic use. To circumvent this problem, a series of N-terminally modified GIP agonists have been developed, with N-AcGIP being the most promising. The aims of the present study were to investigate the effects of N-AcGIP on bone at the micro-level using trabecular and cortical microstructural morphology, and at the tissue-level in rats. Copenhagen rats were randomly assigned into control or N-AcGIP-treated groups and received daily injection for 4 weeks. Bone microstructural morphology was assessed by microCT and dynamic histomorphometry and tissue-level properties by nanoindentation, qBEI and infra-red microscopy. Four week treatment with N-AcGIP did not alter trabecular or cortical microstructural morphology. In addition, no significant modifications of mechanical response and properties at the tissue-level were observed in trabecular bone. However, significant augmentations in maximum load (12%), hardness (14%), indentation modulus (13%) and dissipated energy (16%) were demonstrated in cortical bone. These beneficial modifications of mechanical properties at the tissue-level were associated with increased mineralization (22%) and collagen maturity (13%) of the bone matrix. Taken together, the results support a beneficial role of GIP, and particularly stable analogs such as N-AcGIP, on tissue material properties of bone

Use of glucagon-like peptide-1 receptor agonists and bone fractures: a meta-analysis of randomized clinical trials.

BACKGROUND: Patients with type 2 diabetes mellitus (T2DM) are at a higher risk of bone fractures independent of the use of antidiabetic medications. Furthermore, antidiabetic medications could directly affect bone metabolism. Recently, the use of dipeptidyl peptidase-4 inhibitors has been associated with a lower rate of bone fracture. The aim of the present meta-analysis was to assess whether patients with T2DM treated with glucagon-like peptide-1 receptor agonists (GLP-1Ra) present a lower incidence of bone fracture compared with patients using other antidiabetic drugs. METHODS: A search on Medline, Embase, and http://www.clinicaltrials.gov, as well as a manual search for randomized clinical trials of T2DM treated with either a GLP-1Ra or another antidiabetic drug for a duration of ≥24 weeks was conducted by two authors (GM, AM) independently. RESULTS: Although 28 eligible studies were identified, only seven trials reported the occurrence of at least a bone fracture in one arm of the trial. The total number of fractures was 19 (13 and six with GLP-1Ra and comparator, respectively). The pooled Mantel-Haenszel odds ratio for GLP-1Ra was 0.75 (95% confidence interval 0.28-2.02, P = 0.569) in trials versus other antidiabetic agents. CONCLUSIONS: Although preliminary, our study highlighted that the use of GLP-1Ra does not modify the risk of bone fracture in T2DM compared with the use of other antidiabetic medications

Thiazolidinediones induce osteocyte apoptosis and increase sclerostin expression

Aims  Thiazolidinediones (TZDs) are associated with a higher risk of bone fracture in women compared with men. The aim of the present study was to investigate whether TZDs could influence osteocyte behaviour and contribute to the skeletal phenotype observed in TZD-treated patients.Methods  The murine MLO-Y4 cell line was used as a source of osteocytes. These cells were cultured for 24 h with 0, 10−8 m, 10−7 m, 10−6 m, 10−5 m or 10−4 m of pioglitazone, rosiglitazone or troglitazone in the presence or absence of 17β-oestradiol. The extent of osteocyte apoptosis was assessed, as was the expression of the bone formation inhibitor sclerostin and receptor activator for nuclear factor κB ligand (RANKL) also.Results  In the absence of 17β-oestradiol, pioglitazone, rosiglitazone and troglitazone induced osteocyte apoptosis dose-dependently even at the lowest concentration of 10−8 m. Furthermore, the expression of sclerostin but not RANKL was significantly increased in TZD-treated cultures compared with untreated cultures. The presence of 17β-oestradiol significantly reduced TZD-induced osteocyte apoptosis and also sclerostin up-regulation.Conclusions  These findings therefore raise the potential concern of using TZDs in post-menopausal women where the lack of oestrogen would not prevent osteocyte apoptosis and sclerostin up-regulation and may aggravate the reduction in bone mass in these patients

A new stable GIP-Oxyntomodulin hybrid peptide improved bone strength both at the organ and tissue levels in genetically-inherited type 2 diabetes mellitus

Obesity and type 2 diabetes mellitus (T2DM) progress worldwide with detrimental effects on several physiological systems including bone tissue mainly by affecting bone quality. Several gut hormones analogues have been proven potent in ameliorating bone quality. In the present study, we used the leptin receptor-deficient db/db mice as a model of obesity and severe T2DM to assess the extent of bone quality alterations at the organ and tissue levels. We also examined the beneficial effects of gut hormone therapy in this model by using a new triple agonist ([d-Ala(2)]GIP-Oxm) active at the GIP, GLP-1 and glucagon receptors. As expected, db/db mice presented with dramatic alterations of bone strength at the organ level associated with deterioration of trabecular and cortical microarchitectures and an augmentation in osteoclast numbers. At the tissue level, these animals presented also with alterations of bone strength (reduced hardness, indentation modulus and dissipated energy) with modifications of tissue mineral distribution, collagen glycation and collagen maturity. The use of [d-Ala(2)]GIP-Oxm considerably improved bone strength at the organ level with modest effects on trabecular microarchitecture. At the tissue level, [d-Ala(2)]GIP-Oxm ameliorated bone strength reductions with positive effects on collagen glycation and collagen maturity. This study provides support for including gut hormone analogues as possible new therapeutic strategies for improving bone quality in bone complications associated to T2DM