Bis-picolinamide Ruthenium(III) Dihalide Complexes: Dichloride-to-Diiodide Exchange Generates Single trans Isomers with High Potency and Cancer Cell Selectivity.

A library of new bis-picolinamide ruthenium(III) dihalide complexes of the type [RuX2 L2 ] (X=Cl or I, L=picolinamide) have been synthesised and characterised. The complexes exhibit different picolinamide ligand binding modes, whereby one ligand is bound (N,N) and the other bound (N,O). Structural studies revealed a mixture of cis and trans isomers for the [RuCl2 L2 ] complexes but upon a halide exchange reaction to yield [RuI2 L2 ], only single trans isomers were detected. High cytotoxic activity against human cancer cell lines was observed, with the potencies of some complexes similar to or better than cisplatin. The conversion to [RuI2 L2 ] substantially increased the activity towards cancer cell lines by more than twelvefold. The [RuI2 L2 ] complexes displayed potent activity against the A2780cis (cisplatin-resistant human ovarian cancer) cell line, with a more than fourfold higher potency than cisplatin. Equitoxic activity was observed against normoxic and hypoxic cancer cells, which indicates the potential to eradicate both the hypoxic and aerobic fractions of solid tumours with similar efficiency. The activity of selected complexes against non-cancer ARPE-19 cells was also tested. The [RuI2 L2 ] complexes were found to be more potent than the [RuCl2 L2 ] analogues and also more selective towards cancer cells with a selectivity factor in excess of sevenfold.

Ascorbic acid alters cell fate commitment of human neural progenitors in a WNT/ß-catenin/ROS signaling dependent manner.

BACKGROUND: Improving the neuronal yield from in vitro cultivated neural progenitor cells (NPCs) is an essential challenge in transplantation therapy in neurological disorders. In this regard, Ascorbic acid (AA) is widely used to expand neurogenesis from NPCs in cultures although the mechanisms of its action remain unclear. Neurogenesis from NPCs is regulated by the redox-sensitive WNT/ß-catenin signaling pathway. We therefore aimed to investigate how AA interacts with this pathway and potentiates neurogenesis. METHODS: Effects of 200 µM AA were compared with the pro-neurogenic reagent and WNT/ß-catenin signaling agonist lithium chloride (LiCl), and molecules with antioxidant activities i.e. N-acetyl-L-cysteine (NAC) and ruthenium red (RuR), in differentiating neural progenitor ReNcell VM cells. Cells were supplemented with reagents for two periods of treatment: a full period encompassing the whole differentiation process versus an early short period that is restricted to the cell fate commitment stage. Intracellular redox balance and reactive oxygen species (ROS) metabolism were examined by flow cytometry using redox and ROS sensors. Confocal microscopy was performed to assess cell viability, neuronal yield, and levels of two proteins: Nucleoredoxin (NXN) and the WNT/ß-catenin signaling component Dishevelled 2 (DVL2). TUBB3 and MYC gene responses were evaluated by quantitative real-time PCR. DVL2-NXN complex dissociation was measured by fluorescence resonance energy transfer (FRET). RESULTS: In contrast to NAC which predictably exhibited an antioxidant effect, AA treatment enhanced ROS metabolism with no cytotoxic induction. Both drugs altered ROS levels only at the early stage of the differentiation as no changes were held beyond the neuronal fate commitment stage. FRET studies showed that AA treatment accelerated the redox-dependent release of the initial pool of DVL2 from its sequestration by NXN, while RuR treatment hampered the dissociation of the two proteins. Accordingly, AA increased WNT/ß-catenin signaling output i.e. MYC mRNA level, whereas RuR attenuated it. Moreover, AA improved neurogenesis as much as LiCl as both TUBB3-positive cell yield and TUBB3 mRNA level increased, while NAC or RuR attenuated neurogenesis. Markedly, the neurogenesis outputs between the short and the full treatment with either NAC or AA were found unchanged, supporting our model that neuronal yield is altered by events taking place at the early phase of differentiation. CONCLUSIONS: Our findings demonstrate that AA treatment elevates ROS metabolism in a non-lethal manner prior to the NPCs commitment to their neuronal fate. Such effect stimulates the redox-sensitive DVL2 activation and WNT/ß-catenin signaling response that would enhance the ensuing neuronal cell differentiation.

Activation and deactivation of a robust immobilized Cp*Ir-transfer hydrogenation catalyst: a multielement in situ X-ray absorption spectroscopy study.

A highly robust immobilized [Cp*IrCl2]2 precatalyst on Wang resin for transfer hydrogenation, which can be recycled up to 30 times, was studied using a novel combination of X-ray absorption spectroscopy (XAS) at Ir L3-edge, Cl K-edge, and K K-edge. These culminate in in situ XAS experiments that link structural changes of the Ir complex with its catalytic activity and its deactivation. Mercury poisoning and "hot filtration" experiments ruled out leached Ir as the active catalyst. Spectroscopic evidence indicates the exchange of one chloride ligand with an alkoxide to generate the active precatalyst. The exchange of the second chloride ligand, however, leads to a potassium alkoxide-iridate species as the deactivated form of this immobilized catalyst. These findings could be widely applicable to the many homogeneous transfer hydrogenation catalysts with Cp*IrCl substructure.

Physical activity: benefit or weakness in metabolic adaptations in a mouse model of chronic food restriction?

In restrictive-type anorexia nervosa (AN) patients, physical activity is usually associated with food restriction, but its physiological consequences remain poorly characterized. In female mice, we evaluated the impact of voluntary physical activity with/without chronic food restriction on metabolic and endocrine parameters that might contribute to AN. In this protocol, FRW mice (i.e., food restriction with running wheel) reached a crucial point of body weight loss (especially fat mass) faster than FR mice (i.e., food restriction only). However, in contrast to FR mice, their body weight stabilized, demonstrating a protective effect of a moderate, regular physical activity. Exercise delayed meal initiation and duration. FRW mice displayed food anticipatory activity compared with FR mice, which was strongly diminished with the prolongation of the protocol. The long-term nature of the protocol enabled assessment of bone parameters similar to those observed in AN patients. Both restricted groups adapted their energy metabolism differentially in the short and long term, with less fat oxidation in FRW mice and a preferential use of glucose to compensate for the chronic energy imbalance. Finally, like restrictive AN patients, FRW mice exhibited low leptin levels, high plasma concentrations of corticosterone and ghrelin, and a disruption of the estrous cycle. In conclusion, our model suggests that physical activity has beneficial effects on the adaptation to the severe condition of food restriction despite the absence of any protective effect on lean and bone mass.

Rhodium, iridium, and ruthenium half-sandwich picolinamide complexes as anticancer agents.

Novel rhodium, iridium, and ruthenium half-sandwich complexes containing (N,N)-bound picolinamide ligands have been prepared for use as anticancer agents. The complexes show promising cytotoxicities, with the presence, position, and number of halides having a significant effect on the corresponding IC50 values. One ruthenium complex was found to be more cytotoxic than cisplatin on HT-29 and MCF-7 cells after 5 days and 1 h, respectively, and it remains active with MCF-7 cells even under hypoxic conditions, making it a promising candidate for in vivo studies.

Residues required for activity in Escherichia coli o-succinylbenzoate synthase (OSBS) are not conserved in all OSBS enzymes.

Understanding how enzyme specificity evolves will provide guiding principles for protein engineering and function prediction. The o-succinylbenzoate synthase (OSBS) family is an excellent model system for elucidating these principles because it has many highly divergent amino acid sequences that are <20% identical, and some members have evolved a second function. The OSBS family belongs to the enolase superfamily, members of which use a set of conserved residues to catalyze a wide variety of reactions. These residues are the only conserved residues in the OSBS family, so they are not sufficient to determine reaction specificity. Some enzymes in the OSBS family catalyze another reaction, N-succinylamino acid racemization (NSAR). NSARs cannot be segregated into a separate family because their sequences are highly similar to those of known OSBSs, and many of them have both OSBS and NSAR activities. To determine how such divergent enzymes can catalyze the same reaction and how NSAR activity evolved, we divided the OSBS family into subfamilies and compared the divergence of their active site residues. Correlating sequence conservation with the effects of mutations in Escherichia coli OSBS identified two nonconserved residues (R159 and G288) at which mutations decrease efficiency ≥200-fold. These residues are not conserved in the subfamily that includes NSAR enzymes. The OSBS/NSAR subfamily binds the substrate in a different orientation, eliminating selective pressure to retain arginine and glycine at these positions. This supports the hypothesis that specificity-determining residues have diverged in the OSBS family and provides insight into the sequence changes required for the evolution of NSAR activity.

Gastric Schwannoma: A Case Report and Literature Review.

Gastric schwannomas (GS) are very rare spindle cell, submucosal mesenchymal tumors that arise from Schwann cells of nerve plexuses in the stomach wall. They are usually benign but can become malignant and metastasize to other organs. Surgical resection with biopsy is the gold standard for diagnosis and management of GS. In this article, we present a 68-year-old female patient who presented with abdominal pain, nausea, vomiting, and belching for a couple of months. Upon further evaluation, she was found to have a 4.2 cm gastric mass, which was consistent with gastric schwannoma through biopsy and immunohistochemistry. The patient underwent complete surgical resection of the tumor without any complications. In this article, we will discuss the literature about GS including its clinical presentation, diagnosis, and management options.

Dieulafoy Lesion of the Colon: A Rare Finding During Colonoscopy.

Dieulafoy lesions are common dilated submucosal vessels that can present with gastrointestinal (GI) bleeding. These lesions are usually found in the stomach or esophagus and colonic Dieulafoy lesions are very rare. Clinical presentation can vary from mild non-threatening GI bleeding to massive and recurrent hemorrhage. Here, we discuss a case of a 71-year-old female patient who presented with a bright red bleed per rectum. Colonoscopy was performed, which revealed a bleeding Dieulafoy lesion in the descending colon with clotted blood in the transverse and descending colon. In this article, we will also review the literature related to the epidemiology, clinical presentation, diagnosis, and management of Dieulafoy lesions.

The extracellular matrix of human bone marrow adipocytes and glucose concentration differentially alter mineralization quality without impairing osteoblastogenesis.

Bone marrow adipocytes (BMAds) accrue in various states of osteoporosis and interfere with bone remodeling through the secretion of various factors. However, involvement of the extracellular matrix (ECM) produced by BMAds in the impairment of bone marrow mesenchymal stromal cell (BM-MSC) osteoblastogenesis has received little attention. In type 2 diabetes (T2D), skeletal fragility is associated with several changes in bone quality that are incompletely understood, and BMAd quantity increases in relationship to poor glycemic control. Considering their altered phenotype in this pathophysiological context, we aimed to determine the contribution of the ECM of mature BMAds to osteoblastogenesis and mineralization quality in the context of chronic hyperglycemia. Human BM-MSCs were differentiated for 21 days in adipogenic medium containing either a normoglycemic (LG, 5.5 mM) or a high glucose concentration (HG, 25 mM). The ECM laid down by BMAds were devitalized through cell removal to examine their impact on the proliferation and differentiation of BM-MSCs toward osteoblastogenesis in LG and HG conditions. Compared to control plates, both adipocyte ECMs promoted cell adhesion and proliferation. As shown by the unmodified RUNX2 and osteocalcin mRNA levels, BM-MSC commitment in osteoblastogenesis was hampered by neither the hyperglycemic condition nor the adipocyte matrices. However, adipocyte ECMs or HG condition altered the mineralization phase with perturbed expression levels of type 1 collagen, MGP and osteopontin. Despite higher ALP activity, mineralization levels per cell were decreased for osteoblasts grown on adipocyte ECMs compared to controls. Raman spectrometry revealed that culturing on adipocyte matrices specifically prevents type-B carbonate substitution and favors collagen crosslinking, in contrast to exposure to HG concentration alone. Moreover, the mineral to organic ratio was disrupted according to the presence of adipocyte ECM and the glucose concentration used for adipocyte or osteoblast culture. HG concentration and adipocyte ECM lead to different defects in mineralization quality, recapitulating contradictory changes reported in T2D osteoporosis. Our study shows that ECMs from BMAds do not impair osteoblastogenesis but alter both the quantity and quality of mineralization partly in a glucose concentration-dependent manner. This finding sheds light on the involvement of BMAds, which should be considered in the compromised bone quality of T2D and osteoporosis patients more generally.

Incentives and penalties tied to sales volume in contracts between beverage companies and public universities in the United States.

OBJECTIVE: To assess whether and how beverage companies incentivize universities to maximize sugar-sweetened beverage (SSB) sales through pouring rights contracts. METHODS: Cross-sectional study of contracts between beverage companies and public U.S. universities with 20,000 or more students active in 2018 or 2019. We requested contracts from 143 universities. The primary measures were presence of financial incentives and penalties tied to sales volume. RESULTS: 124 universities (87%) provided 131 unique contracts (64 Coca-Cola, 67 Pepsi). 125 contracts (95%) included at least one provision tying payments to sales volume. The most common incentive type was commissions, found in 104 contracts (79%). Nineteen contracts (15%) provided higher commissions or rebates for carbonated soft drinks compared to bottled water. CONCLUSIONS: Most contracts between universities and beverage companies incentivized universities to market and sell bottled beverages, particularly SSBs. Given the health risks associated with consumption of SSBs, universities should consider their role in promoting them.

Pouring rights contracts between universities and beverage companies: Provisions related to scientific research.

Pouring rights contracts between universities and beverage companies are common and grant companies the exclusive right to serve, sell, and market specific beverages on campuses. In exchange, universities receive financial payments and other incentives. At the same time, beverage industry-sponsored research at universities has increased. Pouring rights contracts may include provisions that allocate funds for or place limitations on scientific research. In this cross-sectional study, we assessed whether pouring rights contracts contained provisions that allocated funds for or placed limitations on scientific research. From 2019 to 2020, we obtained contracts through requests under public records laws from US universities (public, 4-year, ≥ 20,000 students) with contracts active 2018-2019. Of the 143 requests, 6 did not have contracts and 9 declined to provide contracts. Our final sample included 131 contracts from 124 universities in 38 states. Thirty contracts (22.9%) referenced research (18 Coke; 12 Pepsi). Three contracts (2.3%) included provisions that made direct grants or gifts of research funding, 3 (2.3%) permitted the university to acknowledge funding from competitors, and 26 (19.8%) allowed for research using beverages from competing companies. Given increases in industry-sponsored research, the absence of provisions that made direct grants or gifts of research funding suggests that sponsorship of research is occurring through other mechanisms. Additionally, universities must be able to acknowledge funding and conduct research on any beverage and should not need permission via contract provisions to do so. Future studies should consider practical implications of these provisions in pouring rights contracts and assess whether they facilitate or hinder research.

Are Students Paid to Market Sugar-Sweetened Beverages to Peers? A Review of University Pouring Rights Contracts.

Background: Many university students regularly consume sugar-sweetened beverages (SSBs), which are associated with obesity and related chronic diseases. Moreover, students are strongly influenced by both their peers and product marketing. Our exploratory study examined pouring rights contracts between universities and beverage companies, focusing on provisions establishing campus/brand ambassador positions and marketing/merchandising manager positions whose jobs are to market SSBs on campus. Methods: For this cross-sectional study conducted in late 2020, two independent coders reviewed 131 pouring rights contracts between Coca-Cola or Pepsi and 124 unique public universities with 20,000 or more students enrolled. Contracts were active in 2018 or 2019. Results: Twenty-six contracts (20%) contained provisions specifically establishing either campus/brand ambassador positions (n = 16), marketing/merchandising manager positions (n = 7), both (n = 1), or unclear language related to these positions (n = 2). Thirteen contracts (10%) required that the position be filled by a current student. The objectives for both types of positions included increasing revenue and driving beverage sales. When stated in the contracts (n = 5), the payments allocated for these positions ranged between $5,000 and $10,000 per year. Conclusions: Given the association between SSBs and obesity and other related health outcomes, combined with the influence that peers and product marketing may have on adolescents' and young adults' attitudes toward consumption of these beverages, universities should be more transparent when these provisions are included in their pouring rights contracts and should carefully consider whether it is appropriate for these contracts to include funding for students to market SSBs.

Marketing of sugar-sweetened beverages to youth through U.S. university pouring rights contracts.

Child-targeted marketing can influence children's food preferences and childhood consumption of sugar-sweetened beverages (SSBs) is associated with negative health outcomes in both childhood and adulthood. This study explores how beverage companies are using pouring rights contracts (PRCs) with U.S. public universities to market SSBs to youth under 18 years of age. We obtained 139 PRCs (64 Coca-Cola, 67 Pepsi, 8 Gatorade) from 132 universities between June 2019 and August 2020. Each contract was coded by two reviewers who extracted quotes relevant to youth-targeted marketing activities. Twenty-two contracts in our sample (16%) contained a total of 25 provisions related to youth-targeted marketing. Nearly all youth-targeted marketing provisions (n = 24 of 25) were tied to university athletics. Most provisions (n = 19) described the marketing of specific beverages or involved the use of brand names that are also beverages (e.g., "Gatorade," "Coca-Cola"). Fifteen contracts included advertising or support for youth summer camps; five contracts allowed the beverage company to sponsor free experiences for children at university athletic events; three contracts allowed advertising at high school athletic events hosted at university facilities; and two contracts established programs for "underprivileged" or "disadvantaged" youth. Five contracts acknowledged that their provisions may be affected by laws or self-regulatory policies that limit advertising to children. Beverage companies should reconsider marketing to youth through PRCs, universities should carefully consider PRCs with youth-targeted provisions, and the government should further regulate and prevent youth-targeted marketing.

Guidelines for Biobanking of Bone Marrow Adipose Tissue and Related Cell Types: Report of the Biobanking Working Group of the International Bone Marrow Adiposity Society.

Over the last two decades, increased interest of scientists to study bone marrow adiposity (BMA) in relation to bone and adipose tissue physiology has expanded the number of publications using different sources of bone marrow adipose tissue (BMAT). However, each source of BMAT has its limitations in the number of downstream analyses for which it can be used. Based on this increased scientific demand, the International Bone Marrow Adiposity Society (BMAS) established a Biobanking Working Group to identify the challenges of biobanking for human BMA-related samples and to develop guidelines to advance establishment of biobanks for BMA research. BMA is a young, growing field with increased interest among many diverse scientific communities. These bring new perspectives and important biological questions on how to improve and build an international community with biobank databases that can be used and shared all over the world. However, to create internationally accessible biobanks, several practical and legislative issues must be addressed to create a general ethical protocol used in all institutes, to allow for exchange of biological material internationally. In this position paper, the BMAS Biobanking Working Group describes similarities and differences of patient information (PIF) and consent forms from different institutes and addresses a possibility to create uniform documents for BMA biobanking purposes. Further, based on discussion among Working Group members, we report an overview of the current isolation protocols for human bone marrow adipocytes (BMAds) and bone marrow stromal cells (BMSCs, formerly mesenchymal), highlighting the specific points crucial for effective isolation. Although we remain far from a unified BMAd isolation protocol and PIF, we have summarized all of these important aspects, which are needed to build a BMA biobank. In conclusion, we believe that harmonizing isolation protocols and PIF globally will help to build international collaborations and improve the quality and interpretation of BMA research outcomes.

Collagen glycation alters neovascularization in vitro and in vivo.

Microvascular network formation is required for the success of many therapies in regenerative medicine. The process of vessel assembly is fundamentally altered, however, in many people within the potential patient population, including the elderly and people with diabetes. Significant research has been performed to determine how cellular dysfunction contributes to this inadequate neovascularization, but alterations in the extracellular matrix (ECM) may also influence this process. Glycation of ECM proteins, specifically type I collagen, increases as people age and is accelerated due to uncontrolled diabetes. This glycation results in increased ECM stiffness and resistance to degradation. The goal of this research is to determine whether collagen glycation consistent with changes in aged (defined as people older than 80 years old) and diabetic individuals influences neovascularization. Collagen gels that were incubated in glucose-6-phopshate (G6P) for varying times exhibited cross-linking (26.2+/-8.1% and 31.3+/-5.6% for incubation in 375 mM G6P for 5 and 8 days, respectively), autofluorescence, and advanced glycation end product levels (666+/-481 and 2122+/-501 pmol/mg protein for 5 and 8 days of 375 mM G6P, respectively) consistent with aged and diabetic populations. Three-dimensional culture models showed that sprouting angiogenesis was delayed in collagen gels with high levels of glycation. When implanted in vivo, glycated gels were degraded (44.4+/-4.2% and 49.5+/-11.7% nondegraded gel remaining for gels incubated for 5 and 8 days in 375 mM G6P, respectively) and vascularized (75.5+/-32.0 and 73.7+/-23.6 vessels/mm(2)) more slowly than controls (22.3+/-9.9% gel remaining and 133.3+/-31.0 vessels/mm(2)). These results suggest that glycation of collagen can alter neovascularization and may contribute to alterations in vessel assembly observed as people age and due to diabetes.

Hormone-sensitive lipase (HSL) is also a retinyl ester hydrolase: evidence from mice lacking HSL.

Here, we investigated the importance of hormone-sensitive lipase (HSL) as a retinyl ester hydrolase (REH). REH activity was measured in vitro using recombinant HSL and retinyl palmitate. The expression of retinoic acid (RA)-regulated genes and retinoid metabolites were measured in high-fat diet fed HSL-null mice using real-time quantitative PCR and triple-stage liquid chromatography/tandem mass spectrometry, respectively. Age- and gender-matched wild-type littermates were used as controls. The REH activity of rat HSL was found to be higher than that against the hitherto best known HSL substrate, i.e., diacylglycerols. REH activity in white adipose tissue (WAT) of HSL-null mice was completely blunted and accompanied by increased levels of retinyl esters and decreased levels of retinol, retinaldehyde and all-trans RA. Accordingly, genes known to be positively regulated by RA were down-regulated in HSL-null mice, including pRb and RIP140, key factors promoting differentiation into the white over the brown adipocyte lineage. Dietary RA supplementation partly restored WAT mass and the expression of RA-regulated genes in WAT of HSL-null mice. These findings demonstrate the importance of HSL as an REH of adipose tissue and suggest that HSL via this action provides RA and other retinoids for signaling events that are crucial for adipocyte differentiation and lineage commitment.

Fibroblast growth factor-1 (FGF-1) loaded microbeads enhance local capillary neovascularization.

BACKGROUND: Growth of new blood vessels (neovascularization) occurs naturally in the body, but the slow rate of the process may not be sufficient for survival of engineered tissues and transplanted cells, such as pancreatic islets. For transplanted islets, it is crucial that the transplantation site has sufficient vasculature to support the needs of the islets. Therefore, the specific aim of this research was quantify the effect of FGF-1 incorporation into alginate microbeads on neovascularization of such capsules in an in vivo rat transplant model. MATERIALS AND METHODS: Microbeads loaded with FGF-1 or control beads (beads without FGF-1) were implanted in the rat omental pouch model. Animals were sacrificed 7 d post-implantation. RESULTS: Microbeads loaded with FGF-1 stimulated a significant increase in vascular density compared with control rats implanted with control beads. CONCLUSIONS: These results indicate that alginate microbeads loaded with FGF-1 enhance local neovascularization around implanted microbeads. These data provide a compelling impetus for experimental pursuit of FGF-loaded alginate microcapsules for vascularization of transplanted islets.

Sustained delivery of FGF-1 increases vascular density in comparison to bolus administration.

The use of growth factors for the therapeutic stimulation of neovascularization in regenerative medicine has been extensively investigated, but the inability to control their temporal delivery may limit clinical success. A strategy that delivers continuous therapeutic concentrations of growth factors may increase the protein's efficacy. The present study investigates the ability of sustained delivery of fibroblast growth factor-1 (FGF-1), to induce neovascularization in vivo. Alginate microbeads were synthesized to release active FGF-1 for three weeks. Microbeads loaded with FGF-1 (total amount 150 ng) were implanted into a surgically created omentum pouch in rats and were compared to control empty microbead implants and a single bolus injection of 150 ng of FGF-1 with empty microbead implant. Animals were sacrificed at either 3 or 6 weeks post implantation and omenta were analyzed for vascular density and mural cell interactions. Vascular area for bolus FGF-1 and FGF-1 loaded microbeads was higher than control at 3 weeks. At 6 weeks, vascular density in the group with FGF-1 loaded microbeads was significantly higher than the group with bolus administration of FGF-1, primarily due to an increase in the number of vessels less than 20 microm in diameter. Vascular density in omenta of the group receiving the bolus FGF-1 returned to control levels by 6 weeks. Staining for smooth muscle actin showed that 50% of vessels had associated mural cells. There was a trend of increased mural cell staining at 6 weeks for the FGF-1 loaded beads compared to bolus FGF-1 and control levels. Results in these studies suggest that sustained release of FGF-1 increases the duration of the vascular response in contrast to a bolus injection of FGF-1.

High Glucose Level Impairs Human Mature Bone Marrow Adipocyte Function Through Increased ROS Production.

Bone marrow adipocytes (BMAds) accumulate in aging, menopause, and metabolic diseases such as Type 2 diabetes. These osteoporotic conditions are associated with oxidative stress and hyperglycemia which are both considered as critical factors underlying bone fragility. Glucose excess and reactive oxygen species (ROS) are known to favor adipogenesis over osteoblastogenesis. In this study, we investigated whether high glucose exposure could determine dysfunction of mature BMAds, specifically through ROS production. The effects of low (LG, 5 mM) or high glucose (HG, 25 mM) concentrations were examined using human bone mesenchymal stromal cells (hBMSCs) in the time course of differentiation, and, up to 21 days once adipocytes were mature. HG did not alter the adipocyte differentiation process of hBMSCs. Yet, after 21 days under HG exposure, PPARG, CEBPA, and adiponectin mRNA expressions were decreased. These alterations were also observed following adipogenic inducer withdrawal as well as in adipocytes fully differentiated in LG then cultured in HG for the last 11 days. Without inducers, HG condition also led to decreased leptin mRNA level. Importantly, intracellular and extracellular ROS concentrations measured using Amplex Red were significantly raised by 50% under HG exposure. This rise was observed once adipocytes ended differentiation and was reproduced within the different cell culture settings without any cytotoxicity. Among genes involved in ROS metabolism, the mRNA level of the H2O2 generating enzyme NOX4 was found upregulated in the presence of HG. Following cell separation, mature BMAds were shown to overproduce ROS and to display the gene alterations in contrast to non-lipid-laden cells. Finally, a non-lethal treatment with a pro-oxidant agent under LG condition reduces the mRNA levels of PPARG, adiponectin, and leptin as the HG condition does in the absence of inducers, and amplifies the effect of glucose excess on gene expression. HG concentration drives mature BMAds toward altered expression of the main adipokines and transcriptional factors. These perturbations are associated with a rise in ROS generation likely mediated through enhanced expression of NOX4. Mature BMAds are thus responsive to changes in glucose and ROS concentrations, which is relevant regarding with their phenotype and function in age- or metabolic disease-related osteoporosis.

Bone disorders associated with diabetes mellitus and its treatments.

Both type 1 and type 2 diabetes mellitus are associated with bone disorders, albeit via different mechanisms. Early studies in patients with type 1 diabetes suggested a 10-fold increase in the hip fracture risk compared to non-diabetic controls. Meta-analyses published more recently indicate a somewhat smaller risk increase, with odds ratios of 6 to 7. Diminished bone mineral density is among the contributors to the increased fracture risk. Both types of diabetes are associated with decreased bone strength related to low bone turnover. The multiple and interconnected pathophysiological mechanisms underlying the bone disorders seen in type 1 diabetes include insulin deficiency, accumulation of advanced glycation end products, bone microarchitecture alterations, changes in bone marrow fat content, low-grade inflammation, and osteocyte dysfunction. The bone alterations are less severe in type 2 diabetes. Odds ratios for hip fractures have ranged across studies from 1.2 to 1.7, and bone mineral density is higher than in non-diabetic controls. The odds ratio is about 1.2 for all bone fragility fractures combined. The pathophysiological mechanisms are complex, particularly as obesity is very common in patients with type 2 diabetes and is itself associated with an increased risk of fractures at specific sites (humerus, tibia, and ankle). The main mechanisms underlying the bone fragility are an increase in the risk of falls, sarcopenia, disorders of carbohydrate metabolism, vitamin D deficiency, and alterations in cortical bone microarchitecture and bone matrix. The medications used to treat both types of diabetes do not seem to play a major role. Nevertheless, thiazolidinediones and, to a lesser extent, sodium-glucose cotransporter inhibitors may have adverse effects on bone, whereas metformin may have beneficial effects. For the most part, the standard management of bone fragility applies to patients with diabetes. However, emphasis should be placed on preventing falls, which are particularly common in this population. Finally, there is some evidence to suggest that anti-fracture treatments are similarly effective in patients with and without diabetes.