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Bone fragility leading to increased risk of low energy (fragility) fractures that is independent of bone mineral density, occurring in individuals with detected and undetected hyperinsulinaemia.
Patients with type 2 diabetes mellitus (T2DM) and/or cardiovascular disease (CVD), conditions of hyperinsulinaemia, have lower levels of osteocalcin and bone remodelling, and increased rates of fragility fractures. Unlike osteoporosis with lower bone mineral density (BMD), T2DM bone fragility “hyperinsulinaemia-osteofragilitas” phenotype presents with normal to increased BMD. Hyperinsulinaemia and insulin resistance positively associate with increased BMD and fragility fractures.
Hyperinsulinaemia enforces glucose fuelling, which decreases NAD+-dependent antioxidant activity. This increases reactive oxygen species and mitochondrial fission, and decreases oxidative phosphorylation high-energy production capacity, required for osteoblasto/cytogenesis. Osteocytes directly mineralise and resorb bone, and inhibit mineralisation of their lacunocanalicular space via pyrophosphate. Hyperinsulinaemia decreases vitamin D availability via adipocyte sequestration, reducing dendrite connectivity, and compromising osteocyte viability. Decreased bone remodelling and micropetrosis ensues. Trapped/entombed magnesium within micropetrosis fossilisation spaces propagates magnesium deficiency (MgD), potentiating hyperinsulinaemia and decreases vitamin D transport. Vitamin D deficiency reduces osteocalcin synthesis and favours osteocyte apoptosis.
Carbohydrate restriction/fasting/ketosis increases beta-oxidation, ketolysis, NAD+-dependent antioxidant activity, osteocyte viability and osteocalcin, and decreases excess insulin exposure. Osteocalcin is required for hydroxyapatite alignment, conferring bone structural integrity, decreasing fracture risk and improving metabolic/endocrine homeodynamics. Patients presenting with fracture and normal BMD should be investigated for T2DM and hyperinsulinaemia.
1. Increased fragility fractures in T2DM patients with normal to higher bone mineral density: BMD does not predict bone fragility in this metabolic phenotype.
Increased fragility fractures are well documented in patients with type 2 diabetes mellitus (T2DM), a condition of chronic hyperinsulinaemia . Decreased skeletal bone mineral density (L-BMD) is the phenotype of “classical” osteoporosis . A higher BMD is considered to confer greater bone strength (fracture resistance). However, the T2DM bone fragility phenotype more often presents with normal to increased BMD (H-BMD) , and positively tracks with increased fracture risk .
In a cross-sectional non-intervention study of 146 Caucasian non-diabetic postmenopausal women with a mean age of 60 ± 2.7 years, HOMA-IR was found to be positively associated with volumetric bone mineral density (vBMD) . In addition, increased insulin resistance (IR) and treatment with exogenous insulin therapy is positively associated with higher BMD and increased fragility fractures. A study following 5994 T2DM males ≥ 65 years of age, found a higher non-vertebral fracture risk in those using insulin (HR 1.74, 95% CI 1.13, 2.69) who also had a higher BMD . Furthermore, in a population-based matched cohort study investigating primary care records of 2979 insulin users and 14,895 non-users, T2DM patients exposed to insulin therapy to manage glycaemia were found to have a 38% excess fracture risk . In both H-BMD-associated T2DM “osteofragilitas” (bone fragility) and L-BMD osteoporosis phenotypes, there is an increase in bone fragility and a loss in tensile and/or torsion strength, and bone ductility, resulting in higher rates of fractures.
Hyperinsulinaemia drives the pathogenesis of T2DM, which may precede hyperglycaemia by up to 24 years . Hyperinsulinaemia decreases osteoblastogenesis and propagates poorer-quality collagen production, a problem further compounded by hyperglycaemia increasing glycation damage on new or existing bone collagen. Hyperinsulinaemia drives chronic osteocyte distress via excess ceramide synthesis, which increases cellular reactive oxygen species (ROS) , leading to a unique type of mineralisation within their lacunae, coined by Bell, Kayser and Jones as “living fossilisation” . Concomitantly, hyperinsulinaemia decreases osteoclastogenesis, thus impeding the bone resorption needed to enable homeodynamic bone remodelling—a marker of good health  —and this results in a form of hyperinsulinaemia-hyperglycaemia pseudo-osteopetrosis. Combined, these hyperinsulinaemia-driven effects result in the increased bone mineral density seen in people with T2DM.
Osteocytes are the backbone of bone health, and consequently major players in whole-body metabolism . Osteocytes are able to directly mineralise and resorb bone and are central mediators in the regulatory control of osteoblasts and osteoclasts . Chronic hyperinsulinaemia diminishes the replenishment of osteocytes and drives the living fossilisation of the ones in existence, leading to the loss of the osteocyte’s dynamic orchestration of bone remodelling . We propose chronic hyperinsulinaemia provides a plausible explanation, a unifying theory of the mechanisms of action, for the increased BMD and bone fragility “osteofragilitas”, that leads to the increased fracture rates seen in the T2DM bone phenotype.
2. Osteocytes: Mediators of Bone Remodelling and Metabolic Heath
Within the adult skeleton, osteocytes comprise 90% to 95% of the total bone cells . Osteocytes embed within the bone after collagen formation by osteoblasts, of which some of these osteoblasts are fated to differentiate into the embedding osteocytes . The osteocytes, along with their neighbouring osteoblasts, continue to form mineralised bone onto the collagen scaffold, in the process forming the hydroxyapatite lacuna chamber around the embedding osteocytes . Continual morphological changes take place in the process of osteocytogenesis, leading to cells that bear little resemblance in structure to their predecessors, appearing visually more like neurons. Osteocytes have on average 50, and up to 100 dendrites, extending through highly connected intricate tunnels, called the canaliculi. The canaliculi are formed and maintained by the occupying osteocytes , and enable physical connection to other osteocytes and to the outside surface of bone, to osteoblasts, osteoclasts and the vasculature .
The lacunocanalicular system is a fluid-filled space, separated from the mineralised component of bone and maintained by the resident osteocytes . The osteocytes mediate bone remodelling, sense and respond to mechanical stress via their strategic distribution and network of a vast number of connected dendritic processes that enable intercellular communication. Their network detects fluid shear stress, enabling the translation of mechanical stimuli into biochemical messages. Osteocytes feed these signals forward via autocrine and paracrine mechanisms, to elicit homeostatic adaptive responses, which include: regulating bone remodelling in order to provide sensitive and continual changes to whole-body mineral needs, to effecting distant organ responses via endocrine signalling.
3. Dendritic Connectivity Is Essential for Function and Viability
Osteocytes connect to one another via their dendritic processes which form the osteocyte-lacunocanalicular network, and their health and viability are dictated by their dendritic connectivity . A compromise in osteocyte health diminishes their ability to actively inhibit mineralisation of their pericellular space , consequently reducing their connectivity. This further compromises the health of deeper osteocytes that become cut off from receiving nutrients which are no longer able to be delivered through fluid movement via the canalicular tunnels. Furthermore, osteocyte connectivity is required for the transduction of load-induced fluid flow that enables their mechanical sensory system to decrease apoptosis and increase osteocytogenesis . Maintenance of dendritic connectivity allows osteocyte-directed regulation of osteoblasts and osteoclasts, in addition to their own capacity to directly resorb bone .
It is highly likely that osteocytes rather than osteoclasts are responsible for bone resorption in the basal condition, as a function of whole-body mineral homeostasis, and that osteoclast bone resorption serves to function in the acute action/need/stress response . Patients with T2DM have decreased levels of carboxy-terminal collagen crosslinks (CTX) bone resorption marker, indicating a lower bone turnover . Osteocytes regulate both osteoblast and osteoclast differentiation and function, and are thus master regulators of dynamic bone remodelling, a function of healthy physiology (Figure 1) .
4. Hyperglycaemia Increases Advanced Glycation End-Product Formation in Bone Collagen
The anabolic hormone insulin is required at the basal level for healthy bone formation . However, chronic hyperinsulinaemia surpasses this threshold in a dose and duration manner. When coupled with hyperglycaemia, it results in the production of poorer-quality, more rigid collagen and glycation damage of existing collagen . Together, they drive increasing BMD by promoting skeletal mineral acquisition that is fragile in structure . Hyperglycaemia and hyperinsulinaemia increase advanced glycation end-product (AGE) formation. Increased glycation on fibrillar collagen negatively affects bone quality . In human tissue, the most abundant AGE is glucosepane, a lysine–arginine cross-linking, that forms the major AGE in bone type 1 collagen. Hyperglycaemia is one of the leading causes of AGE formation, affecting the structural and biochemical properties of protein binding sites, rendering them unrecognisable to other proteins and enzymes .
Hyperglycaemia-driven AGE formation of glucosepane in bone collagen causes a decrease in viscoelasticity and increases the production of a stiffer collagen, resulting in negative effects on the mechanical properties of load-bearing collagen in bone. This causes bone toughness to decrease, while a greater accumulation of AGE in bone results in increased fracture risk . Furthermore, an increase in stiffer/rigid collagen production occurs due to increased glycation effects on the vasculature, leading to increased hypoxia in the microenvironment . Hypoxia then compromises the osteoblasts’ capacity to generate sufficient ATP for collagen synthesis and for differentiation into osteocytes (Figure 1). Both of which are very energy intensive processes, requiring an efficient mitochondrial capacity to generate ATP via oxidative phosphorylation (OxPhos) . In addition, hyperinsulinaemia and hyperglycaemia inhibit beta-oxidation, whilst increasing mitochondrial (mt) reactive oxygen species (ROS) formation . This leads to increased H2O2 production, causing damage to intracellular protein synthesis machinery and consequently synthesis of poorer quality collagen. The typical methodology of assessing bone quality is via dual x-ray absorptiometry, however, this method is unable to detect the collagen aspect of bone quality . As a result, there is an increased frequency in missing early detection of hyperinsulinaemia-hyperglycaemia osteofragilitas fracture risk, which typically does not present with L-BMD. This suggests that BMD alone is a poor proxy/diagnostic marker for fracture risk in hyperinsulinaemic individuals.
Hyperinsulinaemia “enforces” cellular glucose substrate fuelling , and downregulates beta-oxidation by increasing intracellular ceramide production . Excess ceramide production compromises mtOxPhos capacity, by increasing dynamin-related protein 1 (Drp1) synthesis. Drp1 functions to increase mitochondrial fission, in addition to increasing the production of mtROS such as: superoxide (O2−), hydroxyl radical (−OH) and hydrogen peroxide (H2O2) . Concomitantly, ATP production from glucose oxidation reduces the intracellular pool of nicotinamide adenine dinucleotide (NAD+), consuming four NAD+ in the production of two acetyl moieties, in comparison to beta-oxidation, ketolysis or oxidation of acetoacetate, which consume two, one and zero respectively (Figure 2) . Thus, ATP synthesis that is increasingly reliant on glucose oxidation, negatively impacts the availability of NAD+.
Figure 2. Schematic representation of hyperinsulinaemia effects on cellular oxidative state, and bone homeodynamics. Acetoacetate (AcAc), adenosine triphosphate (ATP), advanced glycation end-products (AGE), beta-adrenergic receptor 2 (AdrB2), beta-hydroxybutyrate (BHB), heparan sulphate (HS), heparan sulphate proteoglycan (HSPG), hydrogen peroxide (H2O2), hyperinsulinaemia (HI), insulin receptor (InsR), isocitrate dehydrogenase 2 (Idh2), forkhead box O1 (FoxO1), glutathione oxidised form (GSSG), glutathione reduced form (GSH), lysine 68 (K68), magnesium (Mg), manganese superoxide dismutase 2 (MnSOD2), nicotinamide adenine dinucleotide (NAD+), nicotinamide adenine dinucleotide phosphate (NADP+), nuclear factor-kB (NF-kB), osteocalcin carboxylated (Gla-OCN or cOCN), osteocalcin un(der)carboxylated (Glu-OCN or ucOCN), osteoprotegerin (OPG), oxidative phosphorylation (OxPhos), receptor activator of nuclear factor-kB ligand (RANKL), reactive oxygen species (ROS), sirtuin 3 (SIRT3), superoxide (O2−), tumour necrosis factor α (TNFα), T cell immune regulator 1 (Tcirg1), tryptophan hydroxylase (Tph), vitamin D (D3), and vitamin K (K).
Sirtuin-3 (SIRT3) regulates the synthesis of endogenous antioxidant enzymes such as mitochondrial manganese superoxide dismutase (MnSOD2) and NADPH-dependent production of reduced glutathione (GSH) . An increased reliance on glucose fuelling and its effect on NAD+ availability, leads to a reduction in SIRT3 activity, since SIRT3 is NAD+ dependent. This in turn decreases signals for the transcription and synthesis of MnSOD2 and GSH . Furthermore, hyperinsulinaemia, diminishes the oxidative buffering capacity of the cellular redox antioxidant GSH . Beta-oxidation activity dramatically increases in osteoblasts as they mature. If beta-oxidation is diminished and/or inhibited, metabolic demand cannot be met, resulting in a decrease in precursor cellular differentiation capability, as osteoblastogenesis and osteocytogenesis are energy intensive processes . Bone neither stores nor synthesises any significant amount of fat. Thus, fatty acids delivered via chylomicron remnants (CR) and low-density lipoproteins (LDL) to bone are more likely to be used for ATP synthesis via beta-oxidation .
5. Hyperinsulinaemia Increases Osteocyte Mitochondrial Fission and Disassociation from the Endoplasmic Reticulum
6. MnSOD2 and SIRT3 Required for Osteoblastogenesis and Osteocytogenesis
7. Glucose Restriction Increases Glutathione Activity and Improved Cell Viability
8. Glucose Restriction Enables Osteocytogenesis; Hyperglycaemia Inhibits It
9. Pyrophosphate and Sclerostin
10. Osteocytes Produce Alkaline Phosphatase
Alkaline phosphatase (ALP) catalyses the hydrolysis of inorganic pyrophosphate (PPi) to phosphate (Pi). Phosphate forms part of calcium hydroxyapatite crystals, and an increase in phosphate promotes mineralisation . Nucleotide pyrophosphatase phosphodiesterase (NPP1) inhibits the action of ALP by increasing the concentration of calcification inhibitor pyrophosphate (ePPi) . Insulin and the fed-state reduces NPP1 gene (Enpp1) expression. Fasting increases the concentration of ePPi via NPP1, leading to inhibition of excessive mineralisation of the osteocyte lacunocanalicular space, which results in maintaining osteocyte viability, dendritic connectivity and consequent dynamic bone remodelling capacity. Insulin action leads to decreased ePPi concentration, subsequently decreasing the osteocytes ability to inhibit bone mineralisation, thus increased mineralisation of the osteocytes lacunocanalicular space occurs. The fasted state provokes the opposite effect, inhibition of mineralisation, and potentially enhances physiological levels of bone resorption via increased beta-hydroxybutyrate (BHB) .
If autumn were a time for humans to accumulate stored energy in preparation for a fasting period during winter, where foods available during autumn increase insulin secretion, activating energy storage mechanisms, logically mineral storage would also be required and may also be facilitated via insulin’s action on inhibiting NPP1 production of ePPi. This may provide an evolutionary explanation, where seasonal hyperinsulinaemia propagates increased osteocyte-lacunae mineralisation during autumn, as an adaptive survival mechanism. This would then be followed by a winter of fasting, which would subsequently lead to increased NPP1 activity, relinquishing back into the system the stored minerals. Hyperinsulinaemia T2DM could be described as a metabolic phenotype reflecting a constantly “fed-state”, the ever-lasting autumn. Thus, providing an explanation of the normal to increased BMD observed in people with T2DM.
12. Osteocalcin Endocrine Effects
13. Carboxylation of Osteocalcin
14. Osteocalcin and Insulin
Wei et al. investigated the potential role of OCN as a means to stimulate pancreatic beta-cell proliferation, given T1DM patients retain a small residual population of functional beta-cells , the logic of their hypothesis seems plausible. T2DM patients have low OCN levels too. However, this is with high insulin levels in non-insulin dependent diabetes mellites (NIDDM). The question remains then, would these patients benefit from exogenous OCN therapy? Or similar to conditions of T2DM, patients given exogenous insulin, serves only to mask the downstream problem (hyperglycaemia), while increasing hyperinsulinaemia and insulin resistance, driving the disease further . If OCN increases insulin secretion, then would we not see higher levels of OCN in hyperinsulinaemia pathologies such as T2DM, CVD and MetS? On the contrary, those with normal insulin levels have significantly higher OCN levels . This may indicate OCN levels and the carboxylation ratio, are firstly markers and then contributory makers of bone fracture resistance, and metabolic and endocrine health.
15. cOCN Levels Determine Hydroxyapatite Alignment Formation
Hyperinsulinaemia and hyperglycaemia propagation of impaired osteoblastogene-sis and osteocytogenesis results in decreased OCN production capacity, which impairs hydroxyapatite crystallographic orientation to collagen fibrils. Combined with poorer quality collagen synthesis and increased glycated collagen, the summative results may lead to compounding effects on bone fragility via compromised structural quality, that is independent of BMD. The sum of all of these dysregulated/impaired conditions, likely leads to the increased fracture rates seen in patients with the T2DM hyperinsulinaemia- osteofragilitas phenotype.
16. Osteocalcin Regulation of Ketosis
17. Vitamin D and Magnesium
Vitamin D is required for osteocyte viability and dendrite connectivity . Hyperinsulinaemia reduces vitamin D availability by increasing sequestration of the lipophilic hormone into adipocytes . Patients with pathologies of hyperinsulinaemia, including T2DM, CVD, obesity, MetS and metabolic cancers, are associated with having a lower vitamin D status . In addition, hyperinsulinaemia promotes magnesium deficiency (MgD) which decreases vitamin D transport in the blood . Chronic hyperinsulinaemia dysregulated vitamin D metabolism negatively affects osteocyte health and their subsequent ability to perform dynamic perilacunar remodelling .
Insulin is required for healthy bone mineralisation, as seen in insulin insufficient T1DM . Puberty and pregnancy are two stages of development where a natural state of hyperinsulinaemia occurs . Hyperinsulinaemia enables increased growth in bones during puberty, and mineral accretion during pregnancy, which may be in the form of micropetrosis (osteocyte-lacunae mineralisation), to ensure adequate provision of minerals for nursing offspring during lactation. These phases of life lend a physiological adaptive explanation as to why we would see an increase in BMD in T2DM hyperinsulinaemia . However, puberty and pregnancy are for a limited duration of time and come with either growth in bones (puberty), or lactation and subsequent resorption of bone minerals. Whereas T2DM hyperinsulinaemia may go undetected for many years, a pernicious chronic elevation, leading to excessive micropetrosis and living fossilisation. The resultant disconnecting of osteocytes from one another, impairs their ability to: sense and transmit information, modulate one another, and directly/indirectly modulate bone turnover . This hypothesis provides a plausible explanation as to why we would see normal to H-BMD in T2DM not conferring fracture resistance.
The article is from 10.3390/biomedicines9091165
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