Asymmetric Organocatalysis Enables Rapid Assembly of Portimine Precursor Chains.

Sequential organocatalytic additions of 2-furanone and dihydroxyacetone derivatives to a crotonaldehyde lynchpin provide polyhydroxylated chains reminiscent of lactonized deoxo Kdn type sugars. Further homologation via Kulinkovich ring opening of the butyrolactone and acylation of the zinc homoenolate derived from the incipient cyclopropanol allows assembly of functionalized chain precursors to portimine. Our experiments probe the stability and reactivity of monosubstituted methylidene pyrrolines and generate advanced intermediates useful for exploring the biosynthesis and de novo synthesis of portimine.

A Mechanistic Exploration of Natural Organic Matter Aggregation and Surface Complexation in Smectite Mesopores.

Soil minerals and organic matter play critical roles in nutrient cycling and other life-essential biogeochemical processes, yet the structural and dynamical details of natural organic matter (NOM) film formation on smectites are not fully understood on the molecular scale. XRD of Suwannee River NOM-hectorite (a smectite clay) complexes shows that the humic and fulvic components of NOM bind predominantly at the external surfaces of packets of smectite platelets rather than in the interlayer slit pores, suggesting that the key behavior governing smectite-NOM interactions takes place in mesopores between smectite particles. New molecular dynamics modeling of a ∼110 ŠH2O-saturated smectite mesopore at near-neutral pH shows that model NOM molecules initially form small clusters of 2-3 NOM molecules near the center of the pore fluid. Formation of these clusters is driven by the hydrophobic mechanism, where aromatic/aliphatic regions associate with one another to minimize their interactions with H2O, and charge-balancing cations associated with the deprotonated carboxylate sites are located only at the outer surface of these clusters. Despite hydrophobicity driving the initial clustering, NOM clusters are formed more quickly when high-charge-density cations like Ca2+ are present vs low-charge-density cations like Cs+, as the former cations more effectively minimize the electrostatic repulsions between the negatively charged NOM molecules. Once the small hydrophobicity-driven NOM clusters form, the simulations show that Ca2+ promotes the aggregation of NOM clusters through tetradentate Ca2+ bridges involving carboxylate groups on two different NOM clusters. Importantly, our studies indicate that Ca2+ plays a crucial role in binding the NOM clusters to the smectite surface, which occurs through multiple quaternary complexes (Ob)-H2O-Ca2+-COO-NOM. In contrast, Cs+ never forms any coordination or acts like bridges between NOM molecules nor as ion bridges to the smectite surface. Additionally, we observe the formation of a metastable superaggregate involving all 16 NOM molecules several times in a Ca2+-bearing mesopore fluid. Superaggregates are never observed in the simulations involving Cs+. The modeling results are fully consistent with helium ion microscope images of NOM-hectorite complexes suggesting that NOM surface films develop when preformed NOM clusters interact with smectite surfaces. Overall, the binding of NOM clusters to the outer surfaces of smectite particles and the formation of large NOM aggregates at neutral pH occur through cation bridging, and cation bridging only occurs when high-charge-density cations like Ca2+ are present.

The osteogenic potential of the marine-derived multi-mineral formula aquamin is enhanced by the presence of vitamin D.

Bone degenerative diseases are on the increase globally and are often problematic to treat. This has led to a demand to identify supplements that aid bone growth and formation. Aquamin is a natural multi-mineral food supplement, derived from the red algae Lithothamnion species which contains calcium, magnesium and 72 other trace minerals. It has been previously reported to increase bone formation and mineralisation. This study aimed to investigate the 28 day in vitro osteogenic response of Aquamin supplemented with Vitamin D. The osteogenic potential of MC3T3-E1 osteoblast-like cells was analysed in standard osteogenic medium supplemented with Aquamin +/- Vitamin D3, and the controls consisted of osteogenic medium, +/- Vitamin D3. Proliferation of osteoblasts, metabolic activity and cell viability did not differ between Aquamin and the osteogenic control groups. Alkaline phosphatase (ALP) levels and mineralisation were increased by the supplementation of Aquamin, and the addition of Vitamin D3 increased mineralisation for all groups. The combination of Aquamin and Vitamin D3 yielded a significant increase in ALP and mineralisation over Aquamin alone and the standard osteogenic control +/- Vitamin D3. This study demonstrates that Aquamin aids osteogenesis, and that its osteogenic response can be enhanced by combining Aquamin with Vitamin D3.

Examination of osteoarthritis and subchondral bone alterations within the stifle joint of an ovariectomised ovine model.

The exact relationship between osteoporosis and osteoarthritis is still a matter for debate for many. The ovariectomised ewe is frequently used as a model for osteoporosis, resulting in significant alterations in bone morphometry and turnover in both trabecular and subchondral bone after 1 year. This study examines whether ovariectomy has any impact on development of osteoarthritis within the ovine stifle joint at the same time point. In addition, we investigate whether there are any significant correlations present between articular cartilage degeneration and alterations in microstructural parameters or turnover rates in the underlying bone. Twenty-two sheep were examined in this study; 10 of the sheep underwent ovariectomy and 12 were kept as controls. Five distinctive fluorochrome dyes were administered intravenously at 12-week intervals to both groups, to label sites of bone turnover. All animals were then sacrificed 12 months postoperatively. Although most specimens showed some evidence of osteoarthritis, no measurable difference between the two study groups was detected. Osteoarthritis was associated with a thinning of the subchondral plate, specifically the subchondral cortical bone; however, whereas previous studies have suggested a link between trabecular thinning and osteoarthritis, this was not confirmed. No correlation was found between osteoarthritis and bone turnover rates of either the subchondral trabecular bone or bone plate. In conclusion, despite the fact that ovariectomy results in marked morphological and structural changes in the ovine stifle joint at 1-year postoperatively, no evidence was found to suggest that it plays a direct role in the aetiology of osteoarthritis.

Subchondral osteopenia and accelerated bone remodelling post-ovariectomy - a possible mechanism for subchondral microfractures in the aetiology of spontaneous osteonecrosis of the knee?

Osteopenia and subchondral microfractures are implicated in the aetiology of spontaneous osteonecrosis of the knee (SPONK). The ovine tibia shows significant alterations of the trabecular architecture within the subchondral bone of the medial tibial plateau post-ovariectomy (OVX), including reduced trabecular bone volume fraction. We hypothesise that accelerated subchondral bone resorption may also play a role in increasing microfracture risk at this site. Twenty-two sheep were examined in this study; 10 of the sheep underwent OVX, while the remainder (n = 13) were kept as controls (CON). Five fluorochrome dyes were administered intravenously at 12-week intervals via the jugular vein to both groups, to label sites of bone turnover. These animals were then killed at 12 months post-operatively. Bone turnover was significantly increased in the OVX group in both trabecular bone (2.024 vs. 1.047 no. mm(-2) ; P = 0.05) and within the subchondral bone plate (4.68 vs. 0.69 no. mm(-2) ; P < 0.001). In addition to the classically described turnover visible along trabecular surfaces, we also found visual evidence of intra-trabecular osteonal remodelling. In conclusion, this study shows significant alterations in bone turnover in both trabecular bone and within the subchondral bone plate at 1 year post-OVX. Remodelling of trabecular bone was due to both classically described hemi-osteonal and intra-trabecular osteonal remodelling. The presence of both localised osteopenia and accelerated bone remodelling within the medial tibial plateau provide a possible mechanism for subchondral microfractures in the aetiology of SPONK. Further utilisation of the OVX ewe may be useful for further study in this field.

Estrogen plus estrogen receptor antagonists alter mineral production by osteoblasts in vitro.

In early postmenopausal women, estrogen withdrawal is associated with increased bone turnover leading to bone loss and increased risk of fracture. Recent studies have suggested that the remaining bone tissue is significantly stronger, stiffer and has an increased tissue-level mineral content. Such changes may occur to compensate for bone loss or as a direct result of estrogen deficiency. To date many details of the physiology of osteoblastic cells during estrogen deficiency are vague. In this study we test the hypothesis that osteoblastic matrix mineralisation is altered at the onset of estrogen deficiency. In vitro cell culture experiments were carried out up to 28 days to compare the mineral production of MC3T3-E1 osteoblastic cells subject to estrogen deficiency (fulvestrant), enhanced estrogen supplementation (17-ß-estradiol) or a combination of both. Mineralisation was detected using von Kossa staining and was quantified with alizarin red absorbance readings. The expression of osteocalcin and osteopontin proteins, markers of osteoblast differentiation and mineralisation, was monitored using immunohistochemistry. Our results demonstrate that estrogen enhancement improves matrix mineralisation by MC3T3 cells in vitro. Furthermore this study found a significant reduction in the level of mineralisation when cells were treated with a combination of estrogen and fulvestrant. In an estrogen deficient environment mineralisation by osteoblastic cells was not altered. These findings suggest that altered tissue mineralisation following estrogen deficiency is not a direct result of estrogen deficiency on osteoblasts. Rather, we propose that altered tissue mineralisation may be a compensatory mechanism by bone to counter bone loss and reduced strength.

Subchondral trabecular structural changes in the proximal tibia in an ovine model of increased bone turnover.

Ovariectomized (OVX) sheep are now considered to be useful models for a variety of metabolic bone disorders. The specific aim of this study was to determine the effects of ovariectomy on the structural parameters and material density of the subchondral bone of the ovine tibial plateau as measured by microcomputed tomography (MicroCT). Twenty-three sheep were examined in this study; 10 of the sheep underwent ovariectomy (OVX), and the remainder (n=13) were kept as controls (CON). These animals were then sacrificed at 12 months post-operatively. Three-dimensional analyses were performed of osteochondral samples (15 mm deep) which were obtained from the medial tibial plateau using MicroCT. Bone volume fraction of the subchondral trabecular bone was reduced in the ovariectomized sheep as compared to control animals (0.439 vs. 0.483, P=0.038). Trabeculae were also significantly thinner in the OVX group (0.220 vs. 0.252 mm, P=0.010), with reduced connectivity density (7.947 vs. 11.524 mm(-3) , P=0.014). There was a trend towards lower numbers of individual trabeculae present in the OVX group as compared to controls, but this did not reach significance (2.817 vs. 3.288 mm(-1) , P=0.1). There was also increased trabecular separation in the OVX group, which again fell short of significance (0.426 vs. 0.387 mm, P=0.251). There was no difference in hydroxyapatite concentration (HA) between the two groups (929 vs. 932 mgHA cm(-3) , P=0.687). In conclusion, significant alterations of the trabecular architecture under the tibial plateau were observed following 12 months of oestrogen-deficiency in this ovine model. Despite these marked morphological and structural density differences, the material densities were equal in the two groups.

The effects of estrogen deficiency and bisphosphonate treatment on tissue mineralisation and stiffness in an ovine model of osteoporosis.

While much research has been dedicated to understanding osteoporosis, the nature of mineral distribution and the mechanical property variation in diseased bone is poorly understood. The current study aimed to determine the effect of estrogen deficiency and bisphosphonate therapy on bone tissue properties using an ovine model of osteoporosis. Skeletally mature animals (4+ years) were divided into an ovariectomy group (ovx, n=20) and a non treatment control group (control, n=20). A zoledronic acid treated group was also included in which animals were estrogen deficient for 20 months prior to receiving treatment (Zol, n=4). Half of the control and ovx groups were euthanized 12 or 31 months post-operatively and all Zol animals were euthanised at 31 months. Individual trabeculae were removed from the proximal femur and were analysed at specific locations across the width of the trabeculae. The mineral content was measured using quantitative backscatter electron imaging and the modulus was measured using nanoindentation. The spatial distribution of tissue modulus and mineral content in bone from ovariectomised animals was similar to control. However, ovariectomy significantly reduced the overall mineral content and tissue modulus relative to the control group after 12 months. Interestingly, significant differences were not maintained 31 months post-OVX. Treatment with zoledronic acid increased the mineral content and tissue modulus relative to both the ovariectomised and control groups. Zoledronic acid was also found to alter the mineral and modulus gradients normally associated with healthy bone tissue. The current study provides evidence that both estrogen deficiency and zoledronic acid therapy significantly alter mineral content and the mechanical properties of trabecular tissue.

Inhibition of Listeria monocytogenes by acetate, benzoate and sorbate: weak acid tolerance is not influenced by the glutamate decarboxylase system.

AIMS: Weak acids are widely used by the food industry to prevent spoilage and to inhibit the growth of pathogenic micro-organisms. In this study the inhibitory effects of three commonly used weak acids, acetic acid, benzoic acid and sorbic acid, on the growth of Listeria monocytogenes were investigated. METHODS AND RESULTS: In a chemically defined medium at pH 6.4 benzoic acid had the greatest inhibitory effect (50% inhibition of growth at 4 mmol l(-1)), while acetate was the least inhibitory (50% inhibition of growth at 50 mmol l(-1)). Mutants lacking either sigma B (Delta sigB) or two of the glutamate decarboxylase systems (Delta gadAB) were used to investigate the contribution these systems make to weak acid tolerance in L. monocytogenes. CONCLUSIONS: The stress-inducible sigma factor sigma B (sigma(B)) was not required for protection against acetate and played only a minor role in tolerating benzoate and sorbate. The glutamate decarboxylase system, which plays an important role in tolerating inorganic acids, played no significant role in the ability of L. monocytogenes to tolerate these weak acids, and neither did the presence of glutamate in the growth medium. SIGNIFICANCE AND IMPACT OF THE STUDY: These results suggest that the effectiveness of weak acid preservatives in food will not be compromised by the presence of glutamate, at least under mildly acidic conditions.

Biomechanical properties across trabeculae from the proximal femur of normal and ovariectomised sheep.

The elastic behaviour of trabecular bone is a function not only of bone volume and architecture, but also of tissue material properties. Variation in tissue modulus can have a substantial effect on the biomechanical properties of trabecular bone. However, the nature of tissue property variation within a single trabecula is poorly understood. This study uses nanoindentation to determine the mechanical properties of bone tissue in individual trabeculae. Using an ovariectomised ovine model, the modulus and hardness distribution across trabeculae were measured. In both normal and ovariectomised bone, the modulus and hardness were found to increase towards the core of the trabeculae. Across the width of the trabeculae, the modulus was significantly less in the ovariectomised bone than in the control bone. However, in contrast to this hardness was found not to differ significantly between the two groups. This study provides valuable information on the variation of mechanical material properties in healthy and diseased trabecular bone tissue. The results of the current study will be useful in finite element modelling where more accurate values of trabecular bone modulus will enable the prediction of the macroscale behaviour of trabecular bone.

Biomechanical comparison of the pullout properties of external skeletal fixation pins in the tibiae of intact and ovariectomised ewes.

The pin-bone interface is the least stable component of the external skeletal fixator. Concerns exist regarding the ability to obtain adequate implant purchase in poor quality bone. Consequently, reduced bone quality has been viewed as a contra-indication for the use of external skeletal fixators. The aim of this study was to investigate the holding power of two different fixator pin designs in bone from entire and ovariectomised sheep. Thirty-two aged ewes were divided into two groups. Group 1 were controls, and Group 2 were ovariectomised (OVX). The ewes were sacrificed 12 months post-ovariectomy and five pairs of tibiae were harvested from each group. The holding power of cortical and cancellous fixator pins was assessed at five standardised locations on each tibia. An increase in mean cortical thickness was noted in the OVX group. The holding power of cancellous fixator pins was superior to that of cortical pins, irrespective of whether or not ovariectomy had been performed. Cancellous pins had an increased holding power in post ovariectomy bone compared to control bone. Cortical pin performance was not affected by ovariectomy. There was a lack of correlation between the incidence of insertional fractures of the far cortex and implant holding power. The results raise questions over the effectiveness of ovariectomy in establishing osteopaenic bone suitable for assessing implant performance, hence further investigations are warranted.

Breast cancer and problems with medical interactions: relationships with traumatic stress, emotional self-efficacy, and social support.

This investigation examined relationships between breast cancer patients' psychosocial characteristics (impact of the illness, traumatic stress symptoms, emotional self-efficacy, and social support) and problems they perceived in their medical interactions and their satisfaction with their physicians. Participants were 352 women enrolled in a multicenter trial of the effects of group therapy for women with recently diagnosed primary breast cancer. The findings reported here are from a cross-sectional analysis of baseline data gathered prior to randomization. Problems interacting with physicians and nurses were associated with greater levels of cancer-related traumatic stress (p < 0.01), less emotional self-efficacy for cancer (p < 0.05), less satisfaction with informational support from family, friends, and spouse, and a tendency to perceive those sources of support as more aversive (p < 0.05). Women who were less satisfied with emotional support from their family, friends and spouse were less likely to feel satisfied with their physicians (p < 0.05). These patient characteristics identify women with primary breast cancer who are likely to experience difficulty in their interactions with nurses and physicians and to be less satisfied with their physicians.